-------
1 trends, Balogh et al. (2010) observed that the mean Pb concentration peaked in the 1970s
2 then declined, with levels from the 1990s below 1930s levels.
3 Data from select regions of the U.S. illustrate that Pb concentrations in surface waters and
4 sediment are likely to be higher in urbanized areas compared with rural locations. Figure
5 3-29 illustrates such variability within a single watershed for the Apalachicola,
6 Chattahoochee, and Flint River Basin, which runs south from north of the greater Atlanta,
7 GA metropolitan area and drains into the Gulf of Mexico at the Apalachicola Bay in the
8 Florida panhandle. Sediment concentrations peaked near the Atlanta area and diminished
9 as distance from the Apalachicola Bay decreased. This observation suggests that rural
10 areas have lower Pb sediment levels compared with urban areas. Consistent with the
11 WACAP trends shown in Figure 3-28. the data also illustrated that Pb concentrations in
12 sediment have declined in the U.S. since 1975 (Figure 3-30). Note that Figure 3-30 does
13 not include data near Atlanta, so the urban peak cannot be seen here as in Figure 3-29.
November 2012 3-122 Draft - Do Not Cite or Quote
-------
100
90
80
•3 70
j? 60
1
* 50
I 40
£ 30
20
10
0
0
100 200 300 400 500
River km above Apalachteola Bay, FL
Downstream
600
700
soo
—*—Pb in streambed-sediment and reservoir-core samples
X Pb background in streambed-sediment and baseline reservoir-core samples
Note: The background refers to concentrations from undeveloped geographic regions and baseline samples are obtained from the
bottom of the sediment core to minimize anthropogenic effects on the sample. Pb concentrations reported on a dry basis.
The lakes and reservoirs along the Apalachicola, Chattahoochee, and Flint River Basin (ACF) feed from north of the Atlanta, GA
metropolitan area into the Gulf of Mexico at Apalachicola Bay in the Florida panhandle.
Source: Reprinted with permission of the American Chemical Society, Callenderand Rice (2000).
Figure 3-29 Sediment core data (1992-1994) for the lakes and reservoirs along
the Apalachicola, Chattahoochee, and Flint River Basin (ACF).
November 2012
3-123
Draft - Do Not Cite or Quote
-------
160
140
120
I
"w 10°
¥
£ so
I
3 60
fi
40
20
—•—1975-1980
-••--1980-1985
-A-1985-1990
—X- 1990-1995
Downstream
100 200 300 400 500
River km above Apalachicola Bay, FL
600
700
800
Note: The background refers to concentrations from undeveloped geographic regions and baseline samples are obtained from the
bottom of the sediment core to minimize anthropogenic effects on the sample. Pb concentrations reported on a dry basis. Sediment
samples were not obtained for various time periods in Atlanta, so the graph does not indicate a lack of elevated sediment Pb in
Atlanta.
Lakes and reservoirs along the Apalachicola, Chattahoochee, and Flint River Basin (ACF) feed from north of the Atlanta, GA
metropolitan area into the Gulf of Mexico at Apalachicola Bay in the Florida panhandle.
Source: Reprinted with permission of the American Chemical Society, Callenderand Rice (2000).
Figure 3-30 Sediment core data (1975-1995) for the lakes and reservoirs along
the Apalachicola, Chattahoochee, and Flint River Basin (ACF).
1 Many recent studies have illustrated the effects of natural disasters on Pb concentrations
2 in surface water and sediment in the wake of Hurricane Katrina, which made landfall on
3 August 29, 2005 in New Orleans, LA, and Hurricane Rita, which made landfall west of
4 New Orleans on September 23, 2005. Pardue et al. (2005) sampled floodwaters on
5 September 3 and September 7, 2005 following the hurricanes and observed that elevated
6 concentrations of Pb along with other trace elements and contaminants were not irregular
7 for stormwater but were important because human exposure to the stormwater was more
8 substantial for Hurricane Katrina than for a typical storm. Floodwater samples obtained
9 throughout the city on September 18, 2005 and analyzed for Pb by Presley et al. (2006)
10 were below the limit of detection (0.04 (ig/mL). Likewise, Hou et al. (2006) measured
11 trace metal concentration in the water column of Lake Pontchartrain and at various
12 locations within New Orleans during the period September 19 through October 9, 2005
13 and found that almost all Pb concentrations were below the limit of detection
November 2012
3-124
Draft - Do Not Cite or Quote
-------
1 (0.0020 mg/kg). However, several studies noted no appreciable increase in Pb
2 concentration within Lake Pontchartrain soils and sediments (Abel et al.. 2010; Abel et
3 al.. 2007; Schwab et al.. 2007; Cobb et al.. 2006; Presley et al.. 2006). Shi et al. (2010)
4 analyzed Lake Pontchartrain sediment samples using a factored approach and found that
5 most Pb was sequestered in carbonate-rich, iron oxide-rich, and magnesium oxide-rich
6 sediments in which it can be more readily mobilized and potentially more bioaccessible.
7 Zahran et al. (2010) and Presley et al. (2010) noted that soil Pb samples obtained outside
8 schools also tended to decrease in the wake of Hurricanes Katrina and Rita, with some
9 sites observing substantial increases and others noting dramatic reductions. These studies
10 suggest that floodwaters can change the spatial distribution of Pb in soil and sediments to
11 result in increased or reduced concentrations.
3.6.3 Rain
12 There are currently no routine measurements of Pb in precipitation in the U.S. Recent
13 results from locations outside the U.S. were consistent with decreasing rain water
14 concentrations described in the 2006 Pb AQCD, reflecting the elimination of Pb from
15 on-road gasoline in most countries. From the 2006 Pb AQCD (U.S. EPA. 2006b). volume
16 weighted Pb concentrations in precipitation collected in 1993-94 from Lake Superior,
17 Lake Michigan and Lake Erie ranged from -0.7 to ~1.1 (ig/L (Sweet etal.. 1998). These
18 values fit well with the temporal trend reported in Watmough and Dillon (2007). who
19 calculated annual volume-weighted Pb concentrations to be 2.12, 1.17 and 0.58 (ig/L for
20 1989-1990, 1990-1991 and 2002-2003, respectively, in precipitation from a central
21 Ontario, Canada, forested watershed. A similar value of 0.41 (ig/L for 2002-03 for Plastic
22 Lake, Ontario, was reported in Landre et al. (2009). For the nearby Kawagama Lake,
23 Shotyk and Krachler (2010) gave Pb concentrations in unfiltered rainwater collected in
24 2008. For August and September 2008, the values were 0.45 and 0.22 (ig/L, respectively,
25 and so there had been little discernible change over the post-2000 period. In support, Pb
26 concentrations in snow pit samples collected in 2005 and 2009 collected 45 km northeast
27 of Kawagama Lake had not changed to any noticeable extent(0.13, 0.17, and 0.28 (ig/L
28 in 2005; 0.15 and 0.26 (ig/L in 2009) (Shotvk and Krachler. 2010).
29 There have also been a few recently published, long-term European studies of Pb
30 concentration in precipitation including Berg et al. (2008) and Farmer et al. (2010). Berg
31 et al. (2008) compared the trends in Pb concentration in precipitation at Norwegian
32 background sites in relation to the decreasing European emissions of Pb over the period
33 1980-2005. The Birkenes site at the southern tip of Norway is most affected by long-
34 range transport of Pb from mainland Europe but there had been a 97% reduction in the
35 concentration of Pb in precipitation over the 26-year time period. This was similar to the
November 2012 3-125 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
reductions of 95% and 92% found for the more northerly sites, Karvatn and
Jergul/Karasjok, respectively (Figure 3-31). A decline of-95% in Pb concentrations in
moss (often used as a biomonitor of Pb pollution) from the southernmost part of Norway,
collected every 5 years over the period 1977-2005, agreed well with the Birkenes
precipitation results (Berg et al., 2008). The reductions in Pb concentration in both
precipitation and moss appear to agree well with the reductions in emissions in Europe
(-85%) and Norway (-99%). Similar to the situation in the U.S., the greatest reductions
occurred prior to the late 1990s, and relatively minor reductions have occurred thereafter;
see Figure 3-31.
G)
-0
Q-
Birkenes
Karvatn
Jergul/Karasjok
1980
1985
1990
1995
2000
2005
Source: Reprinted with permission of Pergamon Press, Berg et al. (2008)
Figure 3-31 Trends in Pb concentration in precipitation from various sites in
Norway over the period 1980-2005.
10 Farmer et al. (2010) showed the trends in concentration of Pb in precipitation collected in
11 a remote part of northeastern Scotland over the period 1989-2007. The 2.6- and 3.0-fold
12 decline in mean concentration from 4.92 (ig/L (1989-1991) to 1.88 (ig/L (1999) and then
13 to 0.63 (ig/L (2006-2007) is qualitatively but not quantitatively in line with the sixfold
14 decline in annual total U.K. emissions of Pb to the atmosphere over each of these time
15 periods. After leaded on-road gasoline was banned in the U.K. in 2000, the ratio of
16 rainwater Pb concentrations to Pb emissions (metric tons) appears to have stabilized to a
17 near-constant value of 0.009 (ig/L per metric ton. The concentrations in precipitation
18 reported in these studies are all at the lower end of the range reported in the
November 2012
3-126
Draft - Do Not Cite or Quote
-------
1 2006 Pb AQCD (U.S. EPA. 2006b). and similar to concentrations reported for those
2 studies conducted after the removal of Pb from on-road gasoline. Overall, recent studies
3 of wet deposition tended to confirm the conclusions of the 2006 Pb AQCD (U.S. EPA,
4 2006b) that wet deposition fluxes have greatly decreased since the removal of Pb from
5 on-road gasoline.
3.6.4 Snowpack
6 The location of Pb deposition impacts its further environmental transport. For example,
7 Pb deposited to some types of soil may be relatively immobile, while Pb deposited to
8 snow is likely to undergo further transport more easily when snow melts. Deposition to
9 snow was investigated in several studies. Measurements of Pb in snowmelt during the
10 WACAP study, showed that median Pb concentration ranged form 20-60 ng/L, with 95th
11 percentile values ranging from 30-130 ng/L; see Figure 3-32 (NPS. 2011). Measurements
12 in WACAP of Hg and particulate carbon deposition onto snow were thought to reflect
13 coal combustion, and Pb was not significantly correlated with Hg in terms of either
14 concentration or of calculated enrichment factors normalized to Al concentrations.
15 Shotyk and Krachler (2009) reported considerably higher concentrations at two North
16 American sites, Johnson and Parnell, in Ontario, Canada. Mean Pb concentration for
17 contemporary snow was 672 (Johnson, n = 6; Parnell, n = 3) ng/L. Shotyk et al. (2010)
18 presented additional values for Pb in contemporary snow samples in Simcoe County,
19 Ontario, and these were higher than for ground and surface waters. Luther Bog and Sifton
20 Bog snow had mean Pb concentrations of 747 and 798 ng/L, respectively. The relatively
21 high concentrations in snow were attributed to contamination with predominantly
22 anthropogenic Pb, although it was noted that the extent of contamination was
23 considerably lower than in past decades.
24 Seasonal patterns of heavy metal deposition to snow on Lambert Glacier basin, east
25 Antarctica, were determined by Hur et al. (2007). The snow pit samples covered the
26 period from austral spring 1998 to summer 2002 and Pb concentrations ranged from
27 1.29-9.6 pg/g with a mean value of 4.0 pg/g. This was similar to a mean value of 4.7 pg/g
28 (1965-1986) obtained by Planchon et al. (2003) for Coats Land, northwest Antarctica.
29 Estimated contributions to the Pb in Lambert Glacier basin snow were ~1 % from rock
30 and soil dust (based on Al concentrations) and -4.6% from volcanoes (based on the
31 concentrations of nss-sulfate). There was almost negligible contribution from seaspray
32 (based on Na concentrations), and so it was suggested that a substantial part of the
33 measured Pb concentration must originate from anthropogenic sources, ffighest Pb
34 concentrations were generally observed in spring/summer with an occasional peak in
35 winter. This contrasts with data for the Antarctic Peninsula, where highest concentrations
November 2012 3-127 Draft - Do Not Cite or Quote
-------
1 occurred during autumn/winter, and again with Coats Land, where high concentrations
2 were observed throughout the winter. These differences were attributed to spatial changes
3 in input mechanism of Pb aerosols arriving at different sites over Antarctica, which could
4 be due to their different source areas and transport pathways. Hur et al. (2007). however,
5 suggested that the good correlation between Pb and crustal metals in snow samples shows
6 that Pb pollutants and crustal PM are transported and deposited in Lambert Glacier basin
7 snow in a similar manner.
o
"
o
r-j
d
O
r-j
o
"(5
1=
111
o
O
o
d
CO
o
d
P
d
g
d
I I I I I I I I I
DENA NOAT MORA NOCA OLYM SEKI GAAR ROMO GLAC
Source: WACAP Database (NFS. 2011)
(DENA = Denali, GAAR = Gates of the Arctic, GLAC = Glacier, MORA = Mount Ranier, NOCA = North Cascades, NOAT = Noatak,
OLYM = Olympic, ROMO = Rocky Mountain, SEKI = Sequoia and Kings Canyon)
Figure 3-32 Box plots illustrating Pb concentration in snow melt at nine
National Parks and Preserves.
Lee et al. (2008b) collected 42 snow samples during the period autumn 2004-summer
2005 from a 2.1-meter snow pit at a high-altitude site on the northeast slope of Mount
November 2012
3-128
Draft - Do Not Cite or Quote
-------
1 Everest, Himalayas. Pb concentrations ranged from 5-530 pg/g with a mean value of
2 77 pg/g. The mean value is clearly higher than the Hur et al. (2007) value for Antarctica
3 but is substantially lower than a mean concentration of 573 pg/g for snow from Mont
4 Blanc, France [ 1990-1991; Lee et al. (2008b)]. The mean Pb concentration for Mount
5 Everest snow was lower during the monsoon (28 pg/g) compared with the non-monsoon
6 periods (137 pg/g). From calculated enrichment factors (Pb/Alsnow:Pb/Alcmst),
7 anthropogenic inputs of Pb were partly important but soil and rock dust also contributed.
8 The low Pb concentrations during monsoon periods are thought to be attributable to low
9 levels of atmospheric loadings of crustal dusts. Lee et al. (2008b) noted that their
10 conclusions differ from those in Kang et al. (2007). who stated that anthropogenic
11 contributions of Pb to Mount Everest snow were negligible because the Everest
12 concentrations were similar to those in Antarctica. Kang et al. (2007) did not take account
13 of the difference in accumulation rates at the two sites and had also used Pb
14 concentrations for Antarctic snow from a study by Ikegawa et al. (1999). Lee et al.
15 (2008b) suggested that these Pb concentrations were much higher than expected and that
16 their snow samples may have suffered from contamination during sampling and analysis.
3.6.5 Natural Waters
17 Monitoring data for streams, rivers, and lakes are summarized in periodic national
18 assessments of surface waters that are carried out periodically by EPA, and they include
19 measurement of major biological and chemical stressors. Human exposure to Pb in
20 drinking water is described in Section 4.1.3.3. Pb concentrations in natural waters also
21 may reflect deposition of Pb even in remote locations. WACAP data at five National
22 Parks and Preserves show median Pb concentrations in surface waters to range from 6 to
23 75 ng/L (NPS. 2011); see Figure 3-33. Four sites (Denali, Mt. Ranier, Glacier, and
24 Olympic National Parks) were in the lower range of 6 to 20 ng/L. One site (Noatak)
25 reported a single value of 75 ng/L. With the exception of the Noatak site, the WACAP
26 values were in line with measurements by Shotyk and Krachler (2007) of Pb
27 concentrations in six artesian flows in Simcoe County, near Elmvale, Ontario, Canada.
28 The values ranged from 0.9 to 18 ng/L with a median (n = 18) of 5.1 ng/L. These are
29 comparable with reports of a range of 0.3-8 ng/L for Lake Superior water samples (Field
30 and Sherrell. 2003). Shotyk and Krachler (2007) also commented that such low
31 concentrations for ground and surface waters are not significantly different from those
32 (5.1 ± 1.4 ng/L) reported for Arctic ice from Devon Island, Canada, dating from
33 4,000-6,000 years ago. In a separate study, Shotyk and Krachler (2009) reported
34 concentrations of Pb in groundwater (from two locations, Johnson and Parnell), surface
35 water (Kawagama Lake [Ontario, Canada]) and contemporary snow (Johnson and
November 2012 3-129 Draft - Do Not Cite or Quote
-------
1 Parnell, as described in Section 3.6.4). The lowest mean dissolved Pb concentrations
2 were found for groundwater: 5.9 (Johnson, n = 11) and 3.4 (Parnell, n = 12) ng/L. For
3 lake water the mean Pb concentration was 57 (Kawagama Lake, n = 12) ng/L. The
4 extremely low concentrations of Pb in the groundwaters were attributed to natural
5 removal processes. Specifically, at the sampling location in Canada, there is an
6 abundance of clay minerals with high surface area and high cation exchange capacity and
7 these, combined with the elevated pH values (pH=8.0) resulting from flow through a
8 terrain rich in limestone and dolostone, provide optimal circumstances for the removal of
9 trace elements such as Pb from groundwater. Although such removal mechanisms have
10 not been demonstrated, the vast difference between Pb concentration in snow and that in
11 the groundwaters, indicate that the removal process is very effective. Shotyk and
12 Krachler (2010) speculate that even at these very low Pb concentrations, much if not
13 most of the Pb is likely to be colloidal, as suggested by the 2006 Pb AQCD (U.S. EPA.
14 2006b). Finally, Shotyk et al. (2010) suggest that the pristine groundwaters from Simcoe
15 County, Canada, provide a useful reference level against which other water samples can
16 be compared.
November 2012 3-130 Draft - Do Not Cite or Quote
-------
1=
o
o
O
i--
CD
to
CD
CD
CD
Q
CD
CO
CD
CD
CD
CD
CD
CD
DENA
MORA
GU\C
OLYM
NO AT
Source: WACAP Database (NFS. 2011)
Note: (DENA = Denali, GLAC = Glacier, MORA = Mount Ranier, NOAT = Noatak, OLYM = Olympic)
Figure 3-33 Boxplots of Pb concentration in surface waters measured at five
National Parks and Preserves.
i
2
o
J
4
5
6
7
8
9
10
11
Although Pb concentrations in Kawagama Lake (Ontario, Canada) water were
approaching "natural values," the 206Pb/207Pb ratios for the samples that had the lowest
dissolved Pb concentrations of 10, 10 and 6 ng/L were 1.16, 1.15 and 1.16, respectively.
These values are inconsistent with those expected for natural Pb (the clay fraction from
the lake sediments dating from the pre-industrial period had values of 1.19-1.21) and it
was concluded that most of the dissolved Pb in the lake water was of industrial origin.
Shotyk and Krachler (2010) found that the full range of isotope ratios for Kawagama
Lake water samples (Ontario, Canada) was 1.09 to 1.15; this was not only much lower
than the stream water values entering the lake but also lower than the values attributed to
leaded on-road gasoline in Canada (-1.15). The streamwater ratio values were ~1.16 to
1.17, while those for rainwater were as low as 1.09; in good agreement with the lower
November 2012
3-131
Draft - Do Not Cite or Quote
-------
1 lake water values. This means that there must be an additional atmospheric source of Pb,
2 which has a lower 206Pb/207Pb ratio than leaded on-road gasoline. Supporting evidence
3 came from contemporary samples such as near surface peat, rainwater and snow, all of
4 which confirmed a trend away from natural Pb (1.191 to 1.201) to lower 206Pb/207Pb
5 ratios. The local smelting activities (Sudbury) were unlikely to be the source of
6 anthropogenic Pb as Sudbury-derived emissions exhibit a typical 206Pb/207Pb ratio of
7 -1.15, similar to leaded on-road gasoline. Instead, it was suggested that long-range
8 transport of Pb from the smelter at Rouyn-Noranda (known as the "Capital of Metal,"
9 NW Quebec) may still be impacting on Kawagama Lake but no Pb isotope data was
10 quoted to support this supposition. Several studies, summarized in Mager (2012).
11 reported Pb concentrations in matched reference and mining-disturbed streams in
12 Missouri and the western U.S. They are summarized in Table 3-11.
13 The range of Pb levels in various saltwater environments are available from several
14 studies although the values are not specific to the U.S. A range of 0.005-0.4 (ig Pb/L for
15 seawater was reported by Leland and Kuwabara (1985) to reflect localized anthropogenic
16 inputs in marine environments based on references from prior to 1980 and 0.01 to
17 27 (ig Pb/L by Sadiq (1992). In general, Pb in seawater is higher in coastal areas and
18 estuaries since these locations are closer to sources of Pb contamination and loading from
19 terrestrial systems (U.S. EPA. 2008b).
November 2012 3-132 Draft - Do Not Cite or Quote
-------
Table 3-11 Pb concentrations from stream food-webs; in mining-disturbed areas
of Missouri and the western U.S.
Area
Total Pb in water (H9/L)
Dissolved Pb (\igl\-)
Animas River, CO (Besser et al., 2001):
Reference Streams
Mining-disturbed areas
Boulder River, MT (Faraa et al., 2007):
Reference Streams
Mining-disturbed areas
<1.8
0.9-8.6
0.4 (colloidal)
0.1-44
<0.2
<0. 1-6.9
0.3-0.4
0.1-2
Coeur d'Alene River, ID (Clark, 2003; Farag et al., 1998):
Reference Streams
Mining-disturbed areas
New Lead Belt, MO (Besser et al., 2007; Brumbaugh et al.
Reference Streams
Mining-disturbed areas
2-20
6-2,000
, 2007):
NR
NR
0.01-2
2-50
O.01-1.6
0.02-1 .7
Adapted with permission of Elsevier: Table 4-4 in Mager (2012)
3.6.6 Vegetation
1 The 2006 Pb AQCD (U.S. EPA. 2006b) presented data on Pb in vegetation. The main
2 conclusions were that Pb uptake was strongly affected by pH, and acidic soils are most
3 likely to have Pb in solution for absorption by plants. Additionally, the 2006 Pb AQCD
4 (U.S. EPA. 2006b) states that most Pb stored within vegetation is stored in roots rather
5 than fruits or shoots. Recent measurements from the WACAP study (NPS. 2011) have
6 shown some Pb storage in lichens. Median Pb concentrations ranged from 0.3 mg/kg in
7 Noatak National Park (Alaska) to 5 mg/kg in Glacier National Park (Montana), with
8 substantial variation in the Glacier and Olympic National Park (Washington State)
9 samples; Figure 3-34. Landers et al. (2008) state that lichen Pb concentrations have
10 decreased substantially from the 1980s and that metal concentrations were within
11 background levels for these remote Western sites.
November 2012
3-133
Draft - Do Not Cite or Quote
-------
o
O
NOAT DENA GAAR MORA OLYM SEKI GLAC
Source: WACAP Database (NFS. 2011)
Note: (DENA = Denali, GAAR = Gates of the Arctic, GLAC = Glacier, MORA = Mount Ranier, NOAT = Noatak, OLYM = Olympic,
SEKI = Sequoia and Kings Canyon)
Figure 3-34 Boxplots of Pb concentration in lichen measured at seven
National Parks and Preserves.
i
2
o
J
4
5
6
7
8
9
10
11
Mosses can be used effectively for monitoring trends in Pb deposition as demonstrated in
many studies (Harmens et al., 2010; Harmens et al., 2008). For example, Harmens et al.
(2008) showed that a 52% decrease in deposited Pb concentrations corresponded to a
57% decrease in Pb concentrations in moss. Farmer et al. (2010) showed that there was
good agreement between the 206Pb/207Pb ratio for precipitation and mosses collected in
northeast Scotland. A study in the Vosges Mountains (France) also found a ratio value of
1.158 for a moss sample and a surface soil litter value of 1.167 and concluded that 1.158
to 1.167 represented the current atmospheric baseline (Geagea et al.. 2008). For rural
northeast Scotland, a combination of sources is giving rise to a 206Pb/207Pb ratio of-1.15
in recent precipitation and mosses (Farmer et al.. 2010). Clearly, sources with a lower
ratio than coal (-1.20) must be contributing substantially to the overall emissions. Pb
November 2012
3-134
Draft - Do Not Cite or Quote
-------
1 from waste incineration has been implicated as a possible current source (cf. typical
2 206Pb/207Pb ratios for Pb from European incineration plants are -1.14 to 1.15 [de la Cruz
3 et al. (2009) and references therein].
4 Pb has been measured on vegetation near roads in recent years. Hasselbach et al. (2005)
5 measured Pb and other metals in mosses to assess deposition of metals along a haul road
6 leading from a port to the Red Dog Zn-Pb mine in Northwest Alaska. They observed that
7 moss concentrations of Pb decreased with increasing distance from the road, while
8 subsurface soils (average depth = 62 meters) did not vary with distance from the road.
9 The strong moss Pb gradient and constant subsurface soil Pb concentrations imply that Pb
10 concentrations in mosses were primarily attributed to deposition and did not have
11 appreciable contributions from soil. Throughout the study area, median moss Pb
12 concentration was 16.2 mg/kg (dry basis), with a range of 1.1-912.5 mg/kg.
13 Concentrations along the port road also diminished with increasing distance from the
14 port, where ore loading operations take place. Hasselbach et al. (2005) attributed the
15 concentrations to ore dust generated during loading operations at the port and mine along
16 with fugitive dust escaping during truck transport. Maher et al. (2008) measured average
17 Pb loading onto tree leaves near highways to be 29 (ig/m2 (max: 81 (ig/m2) at elevations
18 ranging from 0.30 to 2.1 meters.
19 Trends in Pb concentration among flora have decreased in recent years. For example,
20 Franzaring et al. (2010) evaluated data from a 20-year biological monitoring study of Pb
21 concentration in permanent forest and grassland plots in Baden-Wurttemberg, southwest
22 Germany. Grassland and tree foliage samples were collected from 1985-2006. The
23 samples were not washed and so atmospheric deposition rather than uptake from the soil
24 probably predominates. For all foliage (beech and spruce), Pb concentrations have shown
25 large reductions over time, particularly in the early 1990s. The Pb concentrations in the
26 grassland vegetation also decreased from the late 1980s to the early 1990s but the trend
27 thereafter was found to be statistically non-significant. The reduction corresponded to the
28 phase-out of leaded on-road gasoline in Germany. Similarly, Aznar et al. (2008a)
29 observed that the decline in Pb concentrations in the outer level of tree rings
30 corresponded with the decline in Cu smelter emissions in Gaspe Peninsula in Canada;
31 Figure 3-35. Both Pb concentrations and Pb isotope ratios declined with distance from the
32 smelter (Aznar et al.. 2008b; Aznar etal., 2008a).
November 2012 3-135 Draft - Do Not Cite or Quote
-------
Trends
1
0.8
0.6-
0.4
0.2
0
Regional
pollution
800 •
700 -
600 •
500 •
400 •
300 •
200 -
100 •
Humus
•
ii (11)
.
• (41)
i (29)
o (21)
1950 1960 1970 1980 1990 2000
Tree rings
2002 1950 1960 1970 1980 1990 2000
Sapwood-heartwood
boundary
Source: Reprinted with permission of Elsevier Publishing, Aznar et al. (2008a)
Notes: Humus Pb concentration reported in units of mg/kg dry basis, and tree ring Pb concentration reported in units of ug/kg dry
basis.
Figure 3-35 Trends in regional pollution near a copper (Cu) smelter in Canada
and Pb concentrations at the boundary of heartwood trees within
roughly 75 km of the smelter.
3.6.7 Aquatic Bivalves
1 Data from invertebrate waterborne populations can serve as in indicator of Pb
2 contamination because animals such as mussels and oysters take in contaminants during
3 filter feeding. Kimbrough et al. (2008) surveyed Pb concentrations in mussels, zebra
O O V / J '
4 mussels, and oysters along the coastlines of the continental U.S. In general, they observed
5 the highest concentrations of Pb in the vicinity of urban and industrial areas. Company et
6 al. (2008) measured Pb concentrations and Pb isotope ratios in bivalves along the
7 Guadiana River separating Spain and Portugal. Analysis of Pb isotope ratio data
8 suggested that high Pb concentrations were related to historical mining activities in the
November 2012
3-136
Draft - Do Not Cite or Quote
-------
1 region. Elevated Pb concentrations were also observed by Company et al. (2008) in the
2 vicinity of more populated areas. Couture et al. (2010) report data from a survey of the
3 isotopic ratios of Pb mMytilus edulis blue mussel, collected off the coast of France from
4 1985-2005. The results indicated that the likely source of Pb in mussel tissue is from
5 resuspension of contaminated sediments enriched with Pb runoff from wastewater
6 treatment plants, municipal waste incinerators, smelters and refineries rather than from
7 atmospheric deposition (Couture et al.. 2010).
3.6.8 Vertebrate Populations
8 Pb concentrations in fish fillet and liver were measured through the WACAP study in
9 eight National Parks and Preserves (NPS. 2011). For fish fillet, Pb concentrations ranged
10 from 0.0033-0.30 mg/kg dry basis, with a median of 0.016 mg/kg dry basis. Liver stores
11 were several times higher, with Pb concentrations ranging from 0.011-0.97 mg/kg dry
12 basis and a median of 0.096 mg/kg dry basis. Pb concentrations in moose meat and liver
13 were also measured at the Denali National Park and Preserve (Alaska) as part of WACAP
14 (NPS. 2011). Moose meat Pb concentrations ranged from 0.021-0.23 mg/kg dry basis
15 with a median of 0.037 mg/kg dry basis. Pb concentrations in moose liver ranged from
16 0.025-0.11 mg/kg dry basis with a median of 0.053 mg/kg dry basis. Boxplots of
17 measured Pb concentrations in fish fillet and liver are shown in Figure 3-36. and boxplots
18 of measured Pb concentrations for moose meat and liver are shown in Figure 3-37. For
19 fish and meat tissues, median and maximum Pb concentrations were substantially lower
20 than values reported in the 2006 Pb AQCD (U.S. EPA. 2006b). Similarly, in a study of
21 Pb levels in moose teeth from Isle Royale, MI, (Vucetich et al.. 2009) median and mean
22 Pb levels underwent a statistically significant decrease from the period 1952-1982 to
23 1983-2002 in both calves and adult moose. For 1952-1982, Pb concentrations were
24 relatively constant, and a linear decline (R2 = 0.86) was observed for 1983-2002. These
25 findings suggest an overall decline but still some Pb accumulation in fish and moose in
26 these remote locations occurring recently.
November 2012 3-137 Draft - Do Not Cite or Quote
-------
1.0 -
o.s -
0.6 -
o
"(5
c
o
O
0.2 -
o.o -
^J -L Q
1 1 1 1 1 1 1 1 1 1 1 1 1
O 01
CC -i-i
ill ,11 :=
i
"2
UJ
tn
= .s;=i = .s= S = .s= i^Sil
L^ _l U- Ll Ll- -I U_ I ^- -I Ll- Lj i7 ' I
(DENA = Denali, GAAR = Gates of the Arctic, GLAC = Glacier, MORA = Mount Ranier, NOAT = Noatak, OLYM = Olympic,
ROMO = Rocky Mountain, SEKI = Sequoia and Kings Canyon)
Note: Tissue concentration reported on a dry basis.
Source: WACAP Database (NFS. 2011)
Figure 3-36 Boxplots of Pb concentration in fish fillet and fish liver, measured
at eight National Parks and/or Preserves.
November 2012
3-138
Draft - Do Not Cite or Quote
-------
CD
CD
o
"oo
O
O
m
z;
i.U
01
CD
CD
Liver
Meat
Note: Tissue concentration reported on a dry basis.
Source: WACAP Database (NFS. 2011)
Figure 3-37 Boxplots of Pb concentration in moose meat and moose liver
measured at Denali National Park and Preserve.
November 2012
3-139
Draft - Do Not Cite or Quote
-------
3.7 Summary and Conclusions
3.7.1 Sources of Atmospheric Pb
1 The 2006 Pb AQCD (U.S. EPA. 2006b) documented the decline in ambient air Pb
2 emissions following the ban on alkyl-Pb additives for on-road gasoline. Pb emissions
3 declined by 98% from 1970 to 1995 and then by an additional 76% from 1995 to 2008, at
4 which time national Pb emissions were 964 tons/year. As was the case for the 2008
5 NAAQS review, piston-engine aircraft emissions currently comprise the largest share
6 (57%) of total atmospheric Pb emissions nationally (U.S. EPA. 201 la). Other sources of
7 ambient air Pb, in approximate order of importance with regard to national totals, include
8 metal working and mining, fuel combustion, other industrial sources, roadway related
9 sources, and historic Pb. Although piston-engine aircraft collectively comprise the largest
10 emissions source, the highest emitting individual industrial sites produce more ambient
11 air Pb emissions than individual airports.
3.7.2 Fate and Transport of Pb
12 The atmosphere is the main environmental transport pathway for Pb, and on a global
13 scale atmospheric Pb is primarily associated with fine PM. Pb in fine PM is transported
14 long distances and found in remote areas. Atmospheric Pb deposition peaked in the
15 1970s, followed by a decline. Both wet and dry deposition are important removal
16 mechanisms for atmospheric Pb. Wet deposition is more important for fine Pb, and Pb
17 associated with coarse PM is usually removed by dry deposition. Local deposition fluxes
18 are much higher near industrial sources, and a substantial amount of emitted Pb is
19 deposited near sources, leading to increased soil Pb concentrations. Deposition does not
20 cause an ultimate sink for Pb because particles are potentially resuspended and
21 redeposited many times before reaching a site where further transport is unlikely,
22 especially for dry deposition.
23 In water, Pb is transported as free ions, soluble chelates, or on surfaces of iron and
24 organic rich colloids. In most surface waters, atmospheric deposition is the largest source
25 of Pb, but urban runoff and industrial discharge are also considerable. A substantial
26 portion of Pb in runoff ultimately originates from atmospheric deposition, but substantial
27 amounts of Pb from vehicle wear and building materials can also be transported by runoff
28 waters without becoming airborne. Often the majority of Pb transport by runoff occurs at
29 the beginning of a rainfall event. Pb is rapidly dispersed in water, and highest
30 concentrations of Pb are observed near sources where Pb is deposited.
November 2012 3-140 Draft - Do Not Cite or Quote
-------
1 Transport in surface waters is largely controlled by exchange with sediments. The cycling
2 of Pb between water and sediments is governed by chemical, biological, and mechanical
3 processes, which are affected by many factors. Organic matter in sediments has a high
4 capacity for accumulating trace elements like Pb. Binding of anoxic sediments to sulfides
5 is a particularly important process that affects Pb bioavailability. Pb is relatively stable in
6 sediments, with long residence times and limited mobility. However, Pb-containing
7 sediment particles can be remobilized into the water column. Resuspended Pb is largely
8 associated with OM or Fe and Mn particles. This resuspension of contaminated sediments
9 strongly influences the lifetime of Pb in water bodies and can be a more important Pb
10 source to the water column than atmospheric deposition. Resuspension of sediments
11 largely occurs during discrete events related to storms.
12 A complex variety of factors influence Pb retention in soil, including hydraulic
13 conductivity, solid composition, OM content, clay mineral content, microbial activity,
14 plant root channels, animal holes, geochemical reactions, colloid amounts, colloidal
15 surface charge, and pH. Leaf litter can be an important temporary sink for metals from
16 the soil around and below leaves, and decomposition of leaf litter can reintroduce
17 substantial amounts of Pb into soil "hot spots," where re-adsorption of Pb is favored. A
18 small fraction of Pb in soil is present as the free Pb2+ ion. The fraction of Pb in this form
19 is strongly dependent on soil pH.
20 In summary, environmental distribution of Pb occurs mainly through the atmosphere,
21 from where it is deposited into surface waters and soil. Pb associated with coarse PM
22 deposits to a great extent near sources, while fine Pb-PM can be transported long
23 distances. Surface waters act as an important reservoir, with half-lives of Pb in the water
24 column largely controlled by rates of deposition to and resuspension from bottom
25 sediments. Pb retention in soil depends on Pb speciation and a variety of factors intrinsic
26 to the soil.
3.7.3 Ambient Pb Monitoring
27 Since the publication of the 2006 Pb AQCD for Pb (U.S. EPA. 2006b) there has been
28 little progress in the state of the science regarding monitoring technology and monitor
29 siting criteria for representation of population exposures to airborne Pb and Pb of
30 atmospheric origin. Our understanding of sampling errors in the existing FRM, of
31 possible alternatives to existing Pb-TSP sampling technology, and of particle size ranges
32 of Pb particles occurring in different types of locations have changed little in that time. In
33 addition to monitors used historically for sampling Pb-PM, several single stage and
34 multi-stage impactors and inlets used for sampling PM are also potential options for
November 2012 3-141 Draft - Do Not Cite or Quote
-------
1 monitoring Pb particles smaller than 15 pirn. Ambient air Pb deposits onto soil or dust. As
2 described in Section 4.1, the size distribution of dust and soil Pb particles is larger than
3 the size distribution of ambient air Pb particles. The existing samplers reasonably capture
4 the airborne fraction of ambient Pb that is available for human exposure.
5 The current Pb monitoring network design requirements include two types of monitoring
6 sites: source-oriented and non-source-oriented. Source-oriented monitoring sites are
7 required near sources of air Pb emissions which are expected to or have been shown to
8 contribute to ambient air Pb concentrations in excess of the NAAQS. Non-source-
9 oriented monitoring of Pb-TSP or Pb-PMi0 is also required at NCore sites in CBSAs with
10 a population of at least 500,000.
11 In addition to Pb-TSP monitoring for the purposes of judging attainment with the
12 NAAQS, Pb is also routinely measured in smaller PM fractions in the CSN, IMPROVE,
13 and the NATTS networks. While monitoring in multiple networks provides extensive
14 geographic coverage, measurements between networks are not directly comparable in all
15 cases because different PM size ranges are sampled in different networks. Depending on
16 monitoring network, Pb is monitored in TSP, PM10, or PM2 5 using high-volume or
17 low-volume samplers.
3.7.4 Ambient Air Pb Concentrations
18 Ambient air Pb concentrations have declined drastically over the period 1980-2010. The
19 median annual maximum 3-month average concentration of Pb-TSP has dropped by 97%
20 from 0.87 ug/m3 in 1980 to 0.03 ug/m3 in 2010. The decline can be attributed to the
21 phase-out of Pb antiknock agents in on-road fuel and reductions in industrial use and
22 processing of Pb, as described in Section 3.2.1. The mean of maximum 3-month average
23 concentrations for source-oriented monitors was skewed toward the 75th percentile of the
24 data distribution and exceeded the level of the NAAQS, indicating that highest ambient
25 air Pb concentrations occur near a subset of source-oriented monitors. Studies in the peer-
26 reviewed literature have shown slightly elevated Pb concentrations downwind of
27 industrial sources and airports.
28 Spatial variability was observed in ratios and correlations of Pb within different size
29 fractions. Urban or suburban land types did not appear to affect sampled size
30 distributions. Studies in the peer-reviewed literature suggest that proximity to industrial
31 sources or some roadways can affect the Pb-PM size distribution. Pb concentrations
32 exhibit varying degrees of association with other criteria pollutant concentrations.
33 Overall, non-source Pb-TSP was moderately associated with CO, PM2 5, and PMi0, which
34 may indicate some role of traffic in Pb exposure. Among trace metals speciated from
November 2012 3-142 Draft - Do Not Cite or Quote
-------
1 PM2 5, Pb was not associated with most pollutants; Pb did associate with Zn, although
2 that association was low-to-moderate, suggesting mobile source emissions contributing to
3 the Pb. EC, Cu, OC, and Br concentrations also exhibited low-to-moderate associations
4 with Pb concentrations. Such correlations may suggest some common sources affecting
5 the pollutants. Finally, the evidence on natural background Pb suggests a plausible
6 background airborne Pb range of 0.02 to 1 ng/m3.
3.7.5 Ambient Pb Concentrations in Non-Air Media and Biota
7 Atmospheric deposition has led to measurable Pb concentrations observed in rain,
8 snowpack, soil, surface waters, sediments, agricultural plants, livestock, and wildlife
9 across the world, with highest concentrations near Pb sources, such as metal smelters.
10 Since the phase-out of Pb from on-road gasoline, concentrations in these media have
11 decreased to varying degrees. In rain, snowpack, and surface waters, Pb concentrations
12 have decreased considerably. Declining Pb concentrations in tree foliage, trunk sections,
13 and grasses have also been observed. In contrast, Pb is retained in soils and sediments,
14 where it provides a historical record of deposition and associated ambient concentrations.
15 In remote lakes, sediment profiles indicate higher Pb concentrations in near surface
16 sediment as compared to pre-industrial era sediment from greater depth and indicate peak
17 concentrations between 1960 and 1980, when leaded on-road gasoline was at peak use.
18 Concentrations of Pb in moss, lichens, peat, and aquatic bivalves have been used to
19 understand spatial and temporal distribution patterns of air Pb concentrations. Ingestion
20 and water intake are the major routes of Pb exposure for aquatic organisms, and food,
21 drinking water, and inhalation are major routes of exposure for livestock and terrestrial
22 wildlife. Overall, Pb concentrations have decreased substantially in media through which
23 Pb is rapidly transported, such as air and water. Substantial Pb remains in soil and
24 sediment sinks. In areas less affected by major local sources, the highest concentrations
25 are below the surface layers and reflect the previous use of Pb in on-road gasoline and
26 emissions reductions from other sources.
November 2012 3-143 Draft - Do Not Cite or Quote
-------
3.8 Chapter 3 Appendix (Supplemental Material)
3.8.1
Variability across the U.S.
Table 3-12 Distribution of 1-month average Pb-TSP concentrations (ug/m3) nationwide, source-oriented
monitors, 2008-2010.
State/ County
Year Season County State name
Site N: mo
ID means
N
sites Mean
Min
1
5
10
25
50
75
90
95
99
max
Nationwide statistics
2008-2010
2008
2009
2010
Winter
Spring
Summer
Fall
2,318
548
629
1141
554
579
601
584
0.202
0.318
0.212
0.141
0.202
0.239
0.186
0.184
0.000
0.004
0.002
0.000
0.000
0.000
0.001
0.000
0.003
0.004
0.004
0.002
0.002
0.003
0.003
0.004
0.006
0.013
0.008
0.005
0.006
0.007
0.006
0.007
0.010
0.024
0.013
0.008
0.008
0.012
0.010
0.011
0.029
0.050
0.038
0.018
0.026
0.034
0.030
0.026
0.063
0.110
0.084
0.045
0.055
0.070
0.066
0.064
0.217
0.348
0.256
0.136
0.184
0.272
0.212
0.206
0.578
0.841
0.611
0.408
0.502
0.738
0.559
0.505
0.856
1.240
0.856
0.625
0.883
0.977
0.755
0.758
1.576
2.557
1.357
1.233
2.438
1.905
1.233
1.178
4.440
4.440
2.438
1.828
3.103
3.123
4.440
4.225
November 2012
3-144
Draft - Do Not Cite or Quote
-------
Year Season
State/
County
State
County Site
name ID
N: mo
means
N
sites
Mean
Min
1
5
10 25
50
75 90
95
99 max
Nationwide statistics, pooled by site
2008-2010
2008
2009
2010
Winter
Spring
Summer
Fall
Statistics for individual counties
(2008-2010)
111
47
54
101
108
110
111
110
0.161
0.323
0.214
0.140
0.156
0.185
0.148
0.152
0.002
0.007
0.007
0.002
0.000
0.002
0.002
0.002
0.003
0.007
0.007
0.003
0.003
0.002
0.003
0.004
0.008
0.022
0.013
0.005
0.006
0.010
0.006
0.009
0.013 0.031
0.028 0.055
0.018 0.043
0.013 0.030
0.009 0.021
0.015 0.027
0.012 0.025
0.013 0.034
0.056
0.148
0.090
0.052
0.048
0.057
0.050
0.062
0.177 0.441
0.419 0.890
0.343 0.669
0.165 0.392
0.160 0.475
0.210 0.568
0.153 0.430
0.168 0.421
0.687
1.205
0.849
0.586
0.879
0.921
0.696
0.616
0.997 1 .275
1 .540 1 .540
0.921 0.921
0.888 1.185
1.130 1.488
1.189 1.548
0.882 1 .031
1.081 1.189
01109
06037
12057
13015
13215
17031
17115
17119
17143
17195
17201
18035
18089
18097
18127
19155
AL
CA
FL
GA
GA
IL
IL
IL
IL
IL
IL
IN
IN
IN
IN
IA
Pike
Los Angeles
Hillsborough
Bartow
Muscogee
Cook
Macon
Madison
Peoria
Whiteside
Winnebago
Delaware
Lake
Marion
Porter
Pottawattamie
32
131
81
12
12
11
12
36
24
12
11
59
57
70
12
12
1
4
3
1
1
1
1
1
2
1
1
2
3
2
1
1
0.5252
0.2380
0.1755
0.0128
0.0361
0.1515
0.0800
0.1367
0.0119
0.0194
0.0339
0.2746
0.0309
0.0195
0.0125
0.1536
0.054
0.018
0.007
0.007
0.004
0.028
0.018
0.018
0.010
0.010
0.010
0.034
0.004
0.003
0.004
0.025
0.054
0.019
0.007
0.007
0.004
0.028
0.018
0.018
0.010
0.010
0.010
0.034
0.004
0.003
0.004
0.025
0.083
0.026
0.017
0.007
0.004
0.028
0.018
0.022
0.010
0.010
0.010
0.040
0.007
0.005
0.004
0.025
0.164 0.252
0.034 0.047
0.020 0.053
0.008 0.008
0.010 0.013
0.028 0.050
0.025 0.035
0.024 0.037
0.010 0.010
0.012 0.012
0.014 0.020
0.049 0.080
0.008 0.012
0.005 0.008
0.005 0.007
0.026 0.063
0.402
0.085
0.104
0.014
0.027
0.074
0.074
0.068
0.010
0.015
0.024
0.128
0.020
0.012
0.009
0.164
0.798 1 .053
0.246 0.602
0.187 0.530
0.016 0.017
0.043 0.058
0.196 0.304
0.118 0.144
0.175 0.304
0.012 0.016
0.024 0.036
0.032 0.050
0.241 0.427
0.035 0.052
0.025 0.046
0.021 0.024
0.257 0.276
1.117
0.905
0.567
0.019
0.140
0.580
0.168
0.363
0.023
0.040
0.118
1.011
0.079
0.050
0.026
0.282
1.277 1.277
2.501 2.880
1 .007 1 .007
0.019 0.019
0.140 0.140
0.580 0.580
0.168 0.168
0.836 0.836
0.024 0.024
0.040 0.040
0.118 0.118
4.440 4.440
0.298 0.298
0.125 0.125
0.026 0.026
0.282 0.282
November 2012
3-145
Draft - Do Not Cite or Quote
-------
State/
Year Season County
20169
21019
21151
26067
27003
27037
27145
29093
29099
29179
31053
31127
36071
39035
39051
39091
39101
39151
39155
40121
41071
42003
42007
42011
42045
42055
42063
State
KS
KY
KY
Ml
MN
MN
MN
MO
MO
MO
NE
NE
NY
OH
OH
OH
OH
OH
OH
OK
OR
PA
PA
PA
PA
PA
PA
County
name
Saline
Boyd
Madison
Ionia
Anoka
Dakota
Stearns
Iron
Jefferson
Reynolds
Dodge
Nemaha
Orange
Cuyahoga
Fulton
Logan
Marion
Stark
Trumbull
Pittsburg
Yamhill
Allegheny
Beaver
Berks
Delaware
Franklin
Indiana
Site N: mo
ID means
11
7
12
12
12
36
12
171
453
48
9
8
105
72
34
102
10
11
8
11
12
24
54
117
12
11
12
N
sites
1
1
1
1
1
1
1
7
19
4
1
1
3
3
1
4
1
1
1
1
1
2
3
6
1
1
1
Mean
0.2020
0.0042
0.0255
0.1781
0.0157
0.1966
0.0028
0.3388
0.4795
0.0428
0.0515
0.0476
0.0281
0.0941
0.1462
0.0480
0.0358
0.0175
0.0075
0.0023
0.0157
0.0369
0.1130
0.0989
0.0452
0.0449
0.0454
Min
0.043
0.002
0.004
0.016
0.003
0.037
0.000
0.007
0.011
0.007
0.005
0.008
0.001
0.004
0.009
0.003
0.025
0.008
0.004
0.002
0.006
0.006
0.042
0.034
0.043
0.042
0.042
1
0.043
0.002
0.004
0.016
0.003
0.037
0.000
0.008
0.015
0.007
0.005
0.008
0.001
0.004
0.009
0.003
0.025
0.008
0.004
0.002
0.006
0.006
0.042
0.035
0.043
0.042
0.042
5
0.043
0.002
0.004
0.016
0.003
0.048
0.000
0.014
0.033
0.008
0.005
0.008
0.003
0.007
0.009
0.004
0.025
0.008
0.004
0.002
0.006
0.006
0.044
0.038
0.043
0.042
0.042
10
0.044
0.002
0.008
0.023
0.005
0.058
0.000
0.018
0.048
0.011
0.005
0.008
0.004
0.008
0.026
0.005
0.026
0.009
0.004
0.002
0.007
0.006
0.047
0.042
0.043
0.043
0.043
25
0.083
0.004
0.013
0.054
0.007
0.084
0.000
0.033
0.141
0.017
0.021
0.010
0.006
0.014
0.057
0.020
0.027
0.010
0.005
0.002
0.008
0.010
0.068
0.048
0.043
0.043
0.043
50
0.133
0.004
0.017
0.169
0.011
0.137
0.003
0.093
0.336
0.027
0.031
0.024
0.018
0.038
0.091
0.042
0.033
0.018
0.007
0.002
0.016
0.017
0.096
0.066
0.045
0.045
0.044
75
0.320
0.004
0.022
0.279
0.021
0.259
0.005
0.518
0.659
0.060
0.053
0.049
0.044
0.121
0.170
0.070
0.041
0.024
0.008
0.003
0.020
0.040
0.128
0.119
0.047
0.047
0.046
90
0.457
0.007
0.032
0.361
0.022
0.424
0.006
0.850
1.118
0.087
0.149
0.206
0.063
0.210
0.420
0.090
0.054
0.025
0.017
0.003
0.025
0.121
0.198
0.200
0.048
0.047
0.047
95
0.488
0.007
0.121
0.414
0.054
0.572
0.008
1.110
1.451
0.099
0.149
0.206
0.081
0.400
0.490
0.100
0.066
0.028
0.017
0.003
0.037
0.144
0.272
0.295
0.048
0.047
0.058
99
0.488
0.007
0.121
0.414
0.054
0.738
0.008
2.557
2.220
0.268
0.149
0.206
0.101
0.719
0.510
0.120
0.066
0.028
0.017
0.003
0.037
0.149
0.286
0.347
0.048
0.047
0.058
max
0.488
0.007
0.121
0.414
0.054
0.738
0.008
4.225
3.123
0.268
0.149
0.206
0.134
0.719
0.510
0.170
0.066
0.028
0.017
0.003
0.037
0.149
0.286
0.348
0.048
0.047
0.058
November 2012
3-146
Draft - Do Not Cite or Quote
-------
State/
County
Year Season County State name
Site
ID
N: mo
means
N
sites
Mean Min 1
10
25
50
75
90
95
99
max
42073
PA
Lawrence
0.0438 0.042 0.042 0.042 0.042 0.043 0.044 0.045 0.046 0.046 0.046 0.046
42079
PA
Luzerne
10
0.0953 0.043 0.043 0.043 0.044 0.045 0.071 0.102 0.215 0.268 0.268 0.268
42129
PA
Westmoreland
12
0.0439 0.041 0.041 0.041 0.041 0.043 0.044 0.045 0.046 0.047 0.047 0.047
47093
TN
Knox
48
0.0165 0.002 0.002 0.005 0.006 0.008 0.012 0.019 0.032 0.038 0.063 0.063
47163
TN
Sullivan
120
0.0534 0.021 0.023 0.030 0.032 0.037 0.045 0.059 0.083 0.124 0.145 0.156
48085
TX
Collin
108
0.3062 0.007 0.028 0.040 0.052 0.104 0.189 0.438 0.717 0.904 1.178 1.564
48375
TX
Potter
0.0044 0.004 0.004 0.004 0.004 0.004 0.004 0.005 0.006 0.006 0.006 0.006
51770
VA
Roanoke City
12
0.0412 0.005 0.005 0.005 0.008 0.010 0.015 0.035 0.054 0.272 0.272 0.272
55117
Wl
Sheboygan
12
0.0802 0.001 0.001 0.001 0.003 0.007 0.054 0.136 0.182 0.279 0.279 0.279
72013
PR
Arecibo
(Puerto Rico)
12
0.1774 0.038 0.038 0.038 0.064 0.102 0.178 0.264 0.290 0.310 0.310 0.310
Statistics for individual sites where overall average monthly mean > national 90th percentile (2008-2010)
11090003
32
0.525 0.054 0.054 0.083 0.164 0.252 0.402 0.798 1.053 1.117 1.277 1.277
060371405
36
0.671 0.100 0.100 0.188 0.235 0.285 0.359 0.771 2.086 2.501 2.880 2.880
290930016
36
0.670 0.166 0.166 0.186 0.219 0.330 0.466 0.726 0.974 2.435 4.225 4.225
290930021
36
0.681 0.082 0.082 0.084 0.095 0.194 0.650 0.879 1.437 2.438 2.557 2.557
290990004
36
0.997 0.256 0.256 0.307 0.408 0.598 0.918 1.236 1.690 1.905 2.416 2.416
290990015
21
1.275 0.340 0.340 0.421 0.646 0.756 1.118 1.349 2.440 3.103 3.123 3.123
290990020a
31
0.687 0.191 0.191 0.195 0.297 0.368 0.620 0.808 1.111 1.280 2.220 2.220
290990021a
21
0.719 0.084 0.084 0.141 0.359 0.572 0.666 0.876 1.164 1.168 1.553 1.553
290990022a
31
0.441
0.140 0.140 0.171 0.208 0.303 0.409 0.599 0.683 0.754 0.861 0.861
290999001"
24
0.850 0.186 0.186 0.208 0.319 0.449 0.845 1.071 1.382 1.558 1.623 1.623
290999005a
24
0.986 0.155 0.155 0.250 0.330 0.558 0.864 1.487 1.802 1.828 1.985 1.985
480850009a
36
0.601
0.137 0.137 0.138 0.185 0.420 0.579 0.757 1.101 1.178 1.564 1.564
"Sites listed in the bottom six rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-147
Draft - Do Not Cite or Quote
-------
Table 3-13 Distribution of 1 -month average Pb-TSP concentrations
monitors, 2008-2010.
Year Season ^^ State C°^ S,'Je me™
N sites Mean
»
Min
1
(ug/m3) nationwide,
5
10
25
50
non-source-oriented
75
90
95
99
max
Nationwide statistics
2008-2010 2290
2008 685
2009 768
2010 837
Winter 556
Spring 574
Summer 584
Fall 576
0.0120
0.0126
0.0114
0.0120
0.0109
0.0122
0.0119
0.0129
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.002
0.002
0.002
0.000
0.001
0.002
0.002
0.002
0.004
0.005
0.004
0.004
0.004
0.004
0.005
0.005
0.010
0.010
0.010
0.009
0.008
0.009
0.010
0.010
0.015
0.015
0.014
0.016
0.013
0.015
0.016
0.016
0.026
0.029
0.023
0.026
0.022
0.028
0.026
0.026
0.040
0.040
0.040
0.036
0.038
0.040
0.040
0.040
0.052
0.052
0.048
0.054
0.056
0.052
0.050
0.053
0.136
0.066
0.128
0.136
0.087
0.128
0.057
0.136
Nationwide statistics, pooled by site
2008-2010
2008
2009
2010
Winter
Spring
Summer
Fall
88 0.0120
59 0.0125
66 0.0116
73 0.0119
88 0.0115
86 0.0119
88 0.0117
88 0.0130
0.000
0.001
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.001
0.000
0.000
0.000
0.000
0.000
0.000
0.001
0.002
0.001
0.001
0.001
0.001
0.000
0.001
0.002
0.003
0.002
0.001
0.001
0.002
0.001
0.003
0.005
0.006
0.004
0.005
0.004
0.004
0.005
0.005
0.011
0.010
0.010
0.010
0.009
0.009
0.010
0.011
0.016
0.016
0.014
0.018
0.016
0.016
0.016
0.017
0.024
0.024
0.024
0.023
0.025
0.027
0.026
0.028
0.033
0.043
0.032
0.028
0.038
0.032
0.034
0.031
0.046
0.051
0.050
0.046
0.048
0.059
0.043
0.054
0.046
0.051
0.050
0.046
0.048
0.059
0.043
0.054
Statistics for individual counties (2008-2010)
04013 AZ Maricopa 6
06025 CA Imperial 33
06037 CA Los Angeles 224
06065 CA Riverside 72
1 0.0218
1 0.0162
8 0.0098
2 0.0077
0.009
0.004
0.000
0.000
0.009
0.004
0.000
0.000
0.009
0.006
0.000
0.003
0.009
0.009
0.002
0.004
0.014
0.011
0.006
0.006
0.021
0.015
0.010
0.008
0.028
0.019
0.012
0.010
0.038
0.025
0.017
0.010
0.038
0.032
0.020
0.012
0.038
0.035
0.038
0.014
0.038
0.035
0.044
0.014
November 2012
3-148
Draft - Do Not Cite or Quote
-------
State/
Year Season
County
06071
08005
08031
13089
17031
17117
17119
17143
17163
18089
18097
18163
25025
26081
26163
27017
27037
27053
27075
27123
27137
27163
29097
29187
29189
36047
State
CA
CO
CO
GA
IL
IL
IL
IL
IL
IN
IN
IN
MA
Ml
Ml
MN
MN
MN
MN
MN
MN
MN
MO
MO
MO
NY
County
name
San
Bernardino
Arapahoe
Denver
DeKalb
Cook
Macoupin
Madison
Peoria
Saint Clair
Lake
Marion
Vanderburgh
Suffolk
Kent
Wayne
Carlton
Dakota
Hennepin
Lake
Ramsey
Saint Louis
Washington
Jasper
Saint Francois
Saint Louis
Kings
Site N: mo
ID means
71
9
12
10
288
24
36
36
36
36
35
33
31
12
36
12
118
126
10
71
72
72
12
24
33
24
N sites
2
1
1
1
8
1
1
1
1
1
1
2
2
1
2
1
5
4
1
3
2
3
1
2
1
1
Mean
0.0091
0.0120
0.0056
0.0033
0.0195
0.0101
0.0188
0.0105
0.0206
0.0150
0.0058
0.0045
0.0087
0.0053
0.0112
0.0000
0.0035
0.0032
0.0000
0.0062
0.0015
0.0016
0.0125
0.0327
0.0230
0.0131
Min
0.001
0.004
0.003
0.002
0.010
0.010
0.010
0.010
0.010
0.005
0.002
0.001
0.004
0.003
0.003
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.007
0.008
0.005
0.010
1
0.001
0.004
0.003
0.002
0.010
0.010
0.010
0.010
0.010
0.005
0.002
0.001
0.004
0.003
0.003
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.007
0.008
0.005
0.010
5
0.003
0.004
0.003
0.002
0.010
0.010
0.010
0.010
0.010
0.005
0.002
0.001
0.004
0.003
0.003
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.007
0.009
0.005
0.010
10
0.004
0.004
0.004
0.002
0.010
0.010
0.010
0.010
0.012
0.005
0.003
0.002
0.005
0.003
0.004
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.007
0.009
0.005
0.010
25
0.007
0.007
0.005
0.003
0.012
0.010
0.012
0.010
0.014
0.008
0.004
0.003
0.007
0.005
0.005
0.000
0.000
0.000
0.000
0.002
0.000
0.000
0.009
0.018
0.006
0.011
50
0.010
0.012
0.005
0.003
0.016
0.010
0.016
0.010
0.018
0.014
0.005
0.004
0.008
0.005
0.009
0.000
0.002
0.002
0.000
0.004
0.000
0.000
0.012
0.032
0.008
0.012
75
0.012
0.016
0.006
0.004
0.025
0.010
0.020
0.010
0.026
0.019
0.008
0.005
0.010
0.006
0.015
0.000
0.005
0.005
0.000
0.008
0.002
0.003
0.017
0.039
0.050
0.014
90
0.014
0.018
0.008
0.005
0.034
0.010
0.032
0.013
0.032
0.030
0.010
0.006
0.013
0.008
0.021
0.000
0.008
0.006
0.000
0.013
0.004
0.004
0.018
0.054
0.050
0.018
95
0.014
0.018
0.008
0.006
0.040
0.010
0.053
0.013
0.038
0.033
0.012
0.010
0.016
0.008
0.023
0.000
0.010
0.008
0.000
0.020
0.006
0.005
0.019
0.080
0.050
0.020
99
0.022
0.018
0.008
0.006
0.060
0.012
0.066
0.014
0.054
0.049
0.013
0.010
0.020
0.008
0.032
0.000
0.017
0.010
0.000
0.028
0.010
0.006
0.019
0.089
0.066
0.020
max
0.022
0.018
0.008
0.006
0.070
0.012
0.066
0.014
0.054
0.049
0.013
0.010
0.020
0.008
0.032
0.000
0.036
0.044
0.000
0.028
0.010
0.006
0.019
0.089
0.066
0.020
November 2012
3-149
Draft - Do Not Cite or Quote
-------
State/
Year Season
County
39017
39029
39035
39049
39143
39167
40115
42003
42021
42045
42101
42129
48061
48141
48201
48479
49035
51087
State
OH
OH
OH
OH
OH
OH
OK
PA
PA
PA
PA
PA
TX
TX
TX
TX
UT
VA
County
name
Butler
Columbiana
Cuyahoga
Franklin
Sandusky
Washington
Ottawa
Allegheny
Cambria
Delaware
Philadelphia
Westmoreland
Cameron
El Paso
Harris
Webb
Salt Lake
Henrico
Site N: mo
ID means
34
107
107
36
12
54
16
36
23
20
24
24
35
68
32
29
12
7
N sites
1
3
3
1
1
2
2
1
1
1
1
1
1
3
1
1
1
1
Mean
0.0055
0.0155
0.0143
0.0092
0.0048
0.0048
0.0124
0.0105
0.0463
0.0432
0.0210
0.0419
0.0041
0.0206
0.0053
0.0134
0.0173
0.0066
Min
0.002
0.004
0.004
0.004
0.003
0.002
0.003
0.000
0.040
0.040
0.011
0.037
0.002
0.014
0.003
0.004
0.003
0.003
1
0.002
0.004
0.004
0.004
0.003
0.002
0.003
0.000
0.040
0.040
0.011
0.037
0.002
0.014
0.003
0.004
0.003
0.003
5
0.003
0.006
0.006
0.005
0.003
0.002
0.003
0.000
0.040
0.040
0.011
0.040
0.003
0.014
0.003
0.005
0.003
0.003
10
0.004
0.007
0.007
0.005
0.003
0.003
0.005
0.000
0.040
0.040
0.012
0.040
0.003
0.014
0.004
0.006
0.006
0.003
25
0.004
0.008
0.009
0.007
0.004
0.003
0.006
0.004
0.040
0.040
0.014
0.040
0.003
0.015
0.004
0.008
0.009
0.003
50
0.005
0.011
0.012
0.009
0.005
0.005
0.013
0.009
0.040
0.043
0.020
0.040
0.004
0.017
0.005
0.011
0.011
0.004
75
0.007
0.018
0.017
0.011
0.006
0.006
0.017
0.015
0.044
0.046
0.027
0.042
0.005
0.019
0.006
0.018
0.024
0.005
90
0.008
0.027
0.024
0.013
0.006
0.007
0.021
0.019
0.054
0.047
0.033
0.050
0.006
0.029
0.007
0.026
0.040
0.024
95
0.009
0.034
0.030
0.014
0.007
0.008
0.025
0.024
0.058
0.048
0.033
0.050
0.007
0.056
0.008
0.028
0.043
0.024
99
0.009
0.065
0.041
0.016
0.007
0.010
0.025
0.053
0.128
0.048
0.039
0.053
0.009
0.087
0.010
0.035
0.043
0.024
max
0.009
0.136
0.041
0.016
0.007
0.010
0.025
0.053
0.128
0.048
0.039
0.053
0.009
0.087
0.010
0.035
0.043
0.024
November 2012
3-150
Draft - Do Not Cite or Quote
-------
"*
10
25
50
75
90
95
99 max
Statistics for individual sites where overall average monthly mean > national 90th percentile (2008-2010)
170310022
170310026
170316003
291870006"
291870007"
42021 0808a
420450002"
421290007"
481410002"
36
36
36
12
12
23
20
24
23
0.0330
0.0282
0.0249
0.0383
0.0271
0.0463
0.0432
0.0419
0.0236
0.012
0.014
0.012
0.009
0.008
0.040
0.040
0.037
0.016
0.012
0.014
0.012
0.009
0.008
0.040
0.040
0.037
0.016
0.014
0.014
0.014
0.009
0.008
0.040
0.040
0.040
0.016
0.016
0.018
0.018
0.015
0.009
0.040
0.040
0.040
0.016
0.020
0.020
0.020
0.024
0.013
0.040
0.040
0.040
0.017
0.033
0.028
0.026
0.035
0.026
0.040
0.043
0.040
0.018
0.040
0.034
0.031
0.042
0.035
0.044
0.046
0.042
0.021
0.056
0.044
0.033
0.080
0.052
0.054
0.047
0.050
0.033
0.062
0.048
0.038
0.089
0.054
0.058
0.048
0.050
0.056
0.070
0.052
0.040
0.089
0.054
0.128
0.048
0.053
0.087
0.070
0.052
0.040
0.089
0.054
0.128
0.048
0.053
0.087
'Sites listed in the bottom six rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-151
Draft - Do Not Cite or Quote
-------
Table 3-14 Distribution of 3-month moving average Pb-TSP concentrations (ug/m3) nationwide, source-oriented
monitors, 2008-2010.
Year
Nationwide
j O*«*A/ ! ! LI
Season i State JCountyname Site ID I ° N sites Mean Min 1 5 10 25 50 75 90 95 99 max
jCounty | jmeans
statistics3
2008-2010
2008
2,112
0.2134
0.000
0.004
0.010
0.014
0.035
0.079
0.250
0.600
0.881
1.555
2.889
III I J537 I I 0.3225 I 0.005 I 0.006 I 0.016 I 0.028 I 0.056 I 0.129 I 0.385 I 0.900 I 1.197 I 2.452 I 2.889
2009
2010
Winter
jSpring
Nationwide
Summer
Fall
statistics, pooled b
2008-2010 |
2008
2009
2010
ysite
600
975
443
535
572
562
106
47
54
96
0.2177
0.1507
0.2366
0.2376
0.2022
0.1835
0.1671
0.3309
0.2203
0.1415
0.004
0.000
0.003
0.000
0.002
0.002
0.002
0.007
0.007
0.002
0.005
0.002
0.004
0.004
0.003
0.004
0.003
0.007
0.007
0.002
0.011
0.008
0.011
0.011
0.009
0.009
0.012
0.024
0.013
0.008
0.016
0.012
0.014
0.014
0.015
0.013
0.015
0.029
0.019
0.013
0.040
0.024
0.040
0.035
0.034
0.033
0.030
0.056
0.042
0.027
0.090
0.052
0.083
0.078
0.077
0.078
0.059
0.154
0.086
0.053
0.292
0.173
0.272
0.323
0.240
0.220
0.173
0.461
0.311
0.163
0.622
0.436
0.647
0.642
0.580
0.521
0.577
0.814
0.632
0.407
0.799
0.694
0.963
0.999
0.869
0.714
0.717
1.284
0.840
0.619
1.217
1.055
2.070
2.017
1.261
1.186
1.009
1.639
0.886
1.110
2.070
1.375
2.621
2.889
2.163
2.456
1.316
1.639
0.886
1.110
iWinter
M04
0.1700 i 0.003 i 0.004 i 0.011 i 0.013 i 0.025 i 0.055 i 0.171 i 0.522 i 0.827 i 1.097 i 1.324
Statistics fc
Spring
Summer
Fall | | |
>r individual counties (2008-2010)
1 01 109 JAL I Pike
1 06037 |CA | Los Angeles
12057
FL
Hillsbo rough
|13015 |GA |Bartow
J13215 JGA jMuscogee
25
131
79
11
12
101
106
105
1
4
3
1
1
0.1904
0.1597
0.1538
0.5771
0.2521
0.1940
0.0125
0.0367
0.001
0.002
0.002
0.223
0.023
0.011
0.009
0.014
0.002
0.004
0.004
0.223
0.023
0.011
0.009
0.014
0.013
0.010
0.009
0.247
0.036
0.015
0.009
0.014
0.016
0.015
0.012
0.256
0.041
0.037
0.009
0.020
0.028
0.028
0.032
0.302
0.055
0.063
0.011
0.022
0.060
0.058
0.066
0.574
0.078
0.110
0.013
0.031
0.186
0.174
0.170
0.719
0.237
0.249
0.014
0.052
0.502
0.520
0.462
1.088
0.543
0.423
0.015
0.066
0.874
0.788
0.630
1.178
0.832
0.582
0.016
0.070
1.231
0.989
0.960
1.210
2.452
1.770
0.016
0.070
1.740
1.104
1.161
1.210
2.489
1.770
0.016
0.070
November 2012
3-152
Draft - Do Not Cite or Quote
-------
Year
i State/
Season i
| County
1 17031
J17115
17119
J17143
J17195
State
IL
IL
IL
IL
IL
County name
Cook
Macon
Madison
Peoria
Whiteside
Site ID !me^n°s
i9
J10
36
J20
ho
Hs«es
1
1
1
2
1
Mean
0.1364
0.0806
0.1346
0.0121
0.0191
Min
0.068
0.048
0.027
0.010
0.012
1
0.068
0.048
0.027
0.010
0.012
5
0.068
0.048
0.035
0.010
0.012
10
0.068
0.052
0.036
0.010
0.014
25
0.109
0.067
0.063
0.011
0.016
50
0.135
0.080
0.113
0.012
0.019
75
0.150
0.088
0.207
0.014
0.022
90
0.241
0.117
0.283
0.015
0.025
95
0.241
0.123
0.341
0.016
0.025
99
0.241
0.123
0.416
0.016
0.025
max
0.241
0.123
0.416
0.016
0.025
17201
18035
18089
18097
18127
19155
20169
21151
26067
27003
27037
27145
29093
IL
IN
IN
IN
IN
IA
KS
KY
Ml
MN
MN
MN
MO
Winnebago
Delaware
Lake
Marion
Porter
Pottawattamie
Saline
Madison
Ionia
Anoka
Dakota
Stearns
Iron
9
57
46
66
10
12
9
10
10
10
36
10
158
1
2
2
2
1
1
1
1
1
1
1
1
6
0.0356
0.2866
0.0305
0.0198
0.0131
0.1581
0.2286
0.0212
0.1980
0.0161
0.2026
0.0032
0.3465
0.019
0.053
0.007
0.005
0.007
0.034
0.096
0.013
0.106
0.006
0.068
0.000
0.010
0.019
0.053
0.007
0.005
0.007
0.034
0.096
0.013
0.106
0.006
0.068
0.000
0.011
0.019
0.059
0.011
0.006
0.007
0.034
0.096
0.013
0.106
0.006
0.072
0.000
0.019
0.019
0.073
0.012
0.007
0.007
0.067
0.096
0.014
0.110
0.008
0.088
0.001
0.022
0.021
0.090
0.016
0.011
0.007
0.113
0.107
0.015
0.128
0.010
0.104
0.002
0.033
0.027
0.159
0.027
0.014
0.013
0.153
0.231
0.017
0.212
0.013
0.216
0.004
0.142
0.057
0.246
0.036
0.025
0.017
0.220
0.324
0.024
0.259
0.022
0.248
0.004
0.549
0.063
0.495
0.040
0.036
0.020
0.246
0.421
0.037
0.273
0.029
0.357
0.005
0.901
0.063
1.867
0.057
0.043
0.022
0.263
0.421
0.049
0.284
0.031
0.415
0.005
1.167
0.063
2.163
0.129
0.079
0.022
0.263
0.421
0.049
0.284
0.031
0.429
0.005
2.076
0.063
2.163
0.129
0.079
0.022
0.263
0.421
0.049
0.284
0.031
0.429
0.005
2.456
29099
29179
31053
31127
MO
MO
NE
NE
Jefferson
Reynolds
Dodge
Nemaha
423
40
7
6
19
4
1
1
0.4925
0.0397
0.0474
0.0447
0.023
0.012
0.019
0.019
0.033
0.012
0.019
0.019
0.050
0.014
0.019
0.019
0.071
0.015
0.019
0.019
0.187
0.017
0.020
0.024
0.385
0.031
0.060
0.032
0.723
0.057
0.067
0.075
0.989
0.087
0.072
0.087
1.186
0.089
0.072
0.087
2.017
0.100
0.072
0.087
2.889
0.100
0.072
0.087
36071
39035
NY
OH
Orange
Cuyahoga
99
70
3
3
0.0271
0.0905
0.003
0.006
0.003
0.006
0.004
0.010
0.005
0.011
0.007
0.021
0.027
0.050
0.037
0.122
0.068
0.221
0.075
0.287
0.086
0.531
0.086
0.531
139051
iOH i Fulton
i30
0.1609 i 0.025 i 0.025 i 0.027 i 0.046 i 0.054 i 0.092 i 0.254 i 0.354 i 0.453 i 0.567 i 0.567
39091
39101
OH
OH
Logan
Marion
100
8
4
1
0.0499
0.0379
0.004
0.032
0.004
0.032
0.004
0.032
0.006
0.032
0.033
0.034
0.047
0.037
0.072
0.042
0.090
0.047
0.095
0.047
0.100
0.047
0.100
0.047
November 2012
3-153
Draft - Do Not Cite or Quote
-------
Year
i State/
Season i
| County
1 391 51
| 391 55
40121
J41071
I 42003
State
OH
OH
OK
OR
PA
County name
Stark
Trumbull
Pittsburg
Yamhill
Allegheny
Site ID !me^n°s
i9
J6
9
|10
J20
Hs«es
1
1
1
1
2
Mean
0.0180
0.0080
0.0021
0.0166
0.0414
Min
0.015
0.005
0.002
0.009
0.009
1
0.015
0.005
0.002
0.009
0.009
5
0.015
0.005
0.002
0.009
0.011
10
0.015
0.005
0.002
0.011
0.012
25
0.016
0.006
0.002
0.013
0.017
50
0.018
0.008
0.002
0.016
0.030
75
0.019
0.010
0.002
0.019
0.054
90
0.023
0.011
0.003
0.026
0.099
95
0.023
0.011
0.003
0.027
0.120
99
0.023
0.011
0.003
0.027
0.138
max
0.023
0.011
0.003
0.027
0.138
42007
42011
42045
42055
42063
42079
42129
47093
47163
48085
51770
55117
72013
PA
PA
PA
PA
PA
PA
PA
TN
TN
TX
VA
Wl
PR
Beaver
Berks
Delaware
Franklin
Indiana
Luzerne
Westmoreland
Knox
Sullivan
Collin
Roanoke City
Sheboygan
Arecibo
(Puerto Rico)
41
105
10
7
10
6
10
44
118
108
10
10
10
3
6
1
1
1
1
1
2
4
3
1
1
1
0.1160
0.0995
0.0447
0.0447
0.0447
0.1078
0.0434
0.0165
0.0554
0.3101
0.0466
0.0897
0.1725
0.043
0.038
0.043
0.043
0.043
0.084
0.041
0.007
0.030
0.048
0.013
0.012
0.059
0.043
0.039
0.043
0.043
0.043
0.084
0.041
0.007
0.030
0.051
0.013
0.012
0.059
0.052
0.041
0.043
0.043
0.043
0.084
0.041
0.009
0.033
0.070
0.013
0.012
0.059
0.056
0.045
0.043
0.043
0.043
0.084
0.042
0.009
0.035
0.085
0.016
0.034
0.068
0.083
0.051
0.043
0.043
0.043
0.085
0.042
0.012
0.039
0.120
0.019
0.058
0.129
0.114
0.078
0.045
0.045
0.044
0.103
0.044
0.016
0.045
0.217
0.026
0.076
0.194
0.159
0.145
0.046
0.046
0.046
0.135
0.044
0.020
0.060
0.469
0.097
0.126
0.213
0.170
0.183
0.047
0.046
0.049
0.137
0.046
0.023
0.100
0.682
0.108
0.164
0.241
0.187
0.197
0.047
0.046
0.049
0.137
0.046
0.027
0.125
0.753
0.109
0.170
0.245
0.206
0.242
0.047
0.046
0.049
0.137
0.046
0.035
0.134
1.189
0.109
0.170
0.245
0.206
0.251
0.047
0.046
0.049
0.137
0.046
0.035
0.168
1.262
0.109
0.170
0.245
November 2012
3-154
Draft - Do Not Cite or Quote
-------
Year
! State/
Season i
| County
State
County name
ojtpin |N:mo
Slte ID imeans
N sites
Mean
Min
1
5
10
25
50
75
90
95
99
max
Statistics for individual sites where overall average monthly mean > national 90th percentile (2008-2010)
011090003
25
0.5771 0.223 0.223 0.247 0.256 0.302 0.574 0.719 1.088 1.178 1.210 1.210
060371405!
36
i 0.7174 i 0.188 i 0.188 i 0.234 i 0.237 i 0.309 i 0.476 i 0.791 i 2.178 i 2.452 i 2.489 i 2.489
290930016
290930021
290990004
290990015"
290990020"
290990021"
290999001"
290999005"
480850009"
36
36
36
21
29
21
22
22
36
0.6682
0.6950
1 .0090
1.3162
0.6680
0.7317
0.8413
0.9875
0.6068
0.207
0.173
0.640
0.612
0.452
0.429
0.587
0.612
0.196
0.207
0.173
0.640
0.612
0.452
0.429
0.587
0.612
0.196
0.258
0.192
0.655
0.632
0.471
0.435
0.592
0.630
0.268
0.313
0.218
0.699
0.743
0.482
0.507
0.600
0.644
0.335
0.418
0.346
0.775
0.921
0.555
0.547
0.699
0.783
0.469
0.543
0.689
0.913
1.074
0.651
0.685
0.845
0.995
0.585
0.634
0.954
1.081
1.258
0.754
0.900
0.963
1.220
0.704
1.167
1.214
1.555
2.621
0.891
0.999
1.061
1.271
0.965
2.076
1.275
2.011
2.634
0.943
1.013
1.100
1.278
1.189
2.456
1.937
2.017
2.889
0.989
1.141
1.204
1.375
1.262
2.456
1.937
2.017
2.889
0.989
1.141
1.204
1.375
1.262
aThe 3-month averages presented here were created using a simplified approach of the procedures detailed in 40 CFR part 50 appendix R and as such cannot be directly compared to the Pb NAAQS for
determination of compliance with the Pb NAAQS.
"Sites listed in the bottom six rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-155
Draft - Do Not Cite or Quote
-------
Table 3-15 Distribution of 3-month moving average Pb-TSP concentrations (ug/m3) nationwide, non-source-
oriented monitors, 2008-2010.
Year
JO*«*»I I I
Season! 1ST JCountyname Site ID N: mo N Mean Min 1 5 10 25 50 75 90 95 99 max
County | means sites
Nationwide statistics
2008-2010
2008
2,164
0.0120
0.000
0.000
0.001
0.002
0.005
0.010
0.015
0.025
0.037
0.048
0.073
III I I 663 I I 0.0130 I 0.000 I 0.000 I 0.001 I 0.002 I 0.005 I 0.011 I 0.016 I 0.027 I 0.040 I 0.050 I 0.055
2009
2010
Winter
Spring
Summer
Fall
Nationwide statistics, p
2008-2010
2008
2009
2010
>oolec
i by site
727
774
494
548
565
557
86
59
65
71
0.0114
0.0118
0.0113
0.0119
0.0121
0.0126
0.0120
0.0127
0.0117
0.0118
0.000
0.000
0.000
0.000
0.000
0.000
0.000
0.001
0.001
0.000
0.000
0.000
0.001
0.000
0.000
0.000
0.000
0.001
0.001
0.000
0.001
0.001
0.001
0.001
0.001
0.001
0.001
0.002
0.001
0.001
0.002
0.001
0.002
0.002
0.002
0.002
0.002
0.003
0.003
0.001
0.004
0.005
0.005
0.005
0.005
0.005
0.005
0.005
0.004
0.005
0.009
0.010
0.009
0.009
0.010
0.011
0.010
0.011
0.010
0.010
0.014
0.016
0.014
0.015
0.016
0.017
0.016
0.016
0.014
0.017
0.024
0.025
0.023
0.025
0.026
0.027
0.024
0.024
0.026
0.022
0.038
0.035
0.037
0.036
0.037
0.037
0.034
0.043
0.031
0.028
0.043
0.047
0.050
0.050
0.046
0.048
0.046
0.050
0.049
0.045
0.073
0.057
0.055
0.073
0.053
0.057
0.046
0.050
0.049
0.045
iWinter
84 i 0.0118 i 0.000 i 0.000 i 0.001 i 0.002 i 0.005 i 0.010 i 0.015 i 0.025 i 0.036 i 0.048 i 0.048
Statist
Spring
Summer
Fall | |
ics for individual counties (2008-2010
06025JCA jlmperial
06037 1 CA | Los Angeles
06065
CA
Riverside
06071 |CA | San Bernardino
08005JCO jArapahoe
)
31
218
72
69
7
83
86
86
1
8
2
2
1
0.0118
0.0118
0.0126
0.0165
0.0100
0.0078
0.0091
0.0126
0.000
0.000
0.000
0.007
0.000
0.002
0.003
0.011
0.000
0.000
0.000
0.007
0.000
0.002
0.003
0.011
0.001
0.001
0.001
0.008
0.002
0.004
0.005
0.011
0.002
0.002
0.002
0.011
0.004
0.005
0.006
0.011
0.004
0.005
0.005
0.013
0.006
0.007
0.007
0.011
0.010
0.009
0.011
0.017
0.009
0.008
0.009
0.013
0.015
0.016
0.016
0.021
0.013
0.010
0.011
0.014
0.025
0.023
0.026
0.023
0.016
0.011
0.013
0.014
0.034
0.037
0.030
0.023
0.020
0.011
0.014
0.014
0.059
0.043
0.046
0.023
0.028
0.011
0.017
0.014
0.059
0.043
0.046
0.023
0.035
0.011
0.017
0.014
November 2012
3-156
Draft - Do Not Cite or Quote
-------
Year
istate/
Season i
| County
08031
13089
17031
17117
17119
ST
CO
GA
IL
IL
IL
County name
Denver
DeKalb
Cook
Macoupin
Madison
Site ID
N:mo
means
10
8
287
24
36
N
sites
1
1
8
1
1
Mean
0.0054
0.0035
0.0196
0.0101
0.0188
Min
0.004
0.003
0.010
0.010
0.010
1
0.004
0.003
0.010
0.010
0.010
5
0.004
0.003
0.010
0.010
0.010
10
0.004
0.003
0.010
0.010
0.011
25
0.005
0.003
0.012
0.010
0.014
50
0.006
0.004
0.017
0.010
0.016
75
0.006
0.004
0.025
0.010
0.022
90
0.006
0.004
0.033
0.011
0.036
95
0.006
0.004
0.038
0.011
0.036
99
0.006
0.004
0.047
0.011
0.039
max
0.006
0.004
0.051
0.011
0.039
17143
17163
18089
18097
18163
25025
26081
26163
27017
27037
27053
27075
27123
27137
27163
29097
29187
IL
IL
IN
IN
IN
MA
Ml
Ml
MN
MN
MN
MN
MN
MN
MN
MO
MO
Peoria
Saint Clair
Lake
Marion
Vanderburgh
Suffolk
Kent
Wayne
Carlton
Dakota
Hennepin
Lake
Ramsey
Saint Louis
Washington
Jasper
Saint Francois
36
36
36
33
31
24
10
32
10
112
124
8
65
70
70
10
21
1
1
1
1
2
2
1
2
1
5
4
1
3
2
3
1
2
0.0105
0.0204
0.0149
0.0056
0.0047
0.0093
0.0055
0.0119
0.0000
0.0036
0.0033
0.0000
0.0061
0.0016
0.0017
0.0135
0.0337
0.010
0.012
0.007
0.003
0.002
0.005
0.004
0.004
0.000
0.000
0.000
0.000
0.001
0.000
0.000
0.009
0.011
0.010
0.012
0.007
0.003
0.002
0.005
0.004
0.004
0.000
0.000
0.001
0.000
0.001
0.000
0.000
0.009
0.011
0.010
0.012
0.007
0.003
0.003
0.006
0.004
0.004
0.000
0.001
0.001
0.000
0.001
0.000
0.000
0.009
0.012
0.010
0.014
0.007
0.003
0.003
0.006
0.005
0.005
0.000
0.001
0.001
0.000
0.001
0.000
0.000
0.011
0.012
0.010
0.016
0.010
0.004
0.004
0.008
0.005
0.005
0.000
0.001
0.002
0.000
0.002
0.001
0.001
0.012
0.027
0.010
0.020
0.014
0.005
0.005
0.009
0.006
0.012
0.000
0.003
0.003
0.000
0.005
0.001
0.001
0.014
0.035
0.011
0.024
0.018
0.007
0.005
0.011
0.006
0.017
0.000
0.005
0.004
0.000
0.008
0.002
0.003
0.015
0.042
0.012
0.029
0.024
0.009
0.006
0.013
0.006
0.021
0.000
0.007
0.006
0.000
0.014
0.004
0.004
0.016
0.048
0.012
0.033
0.032
0.010
0.007
0.015
0.006
0.023
0.000
0.012
0.006
0.000
0.016
0.004
0.004
0.017
0.053
0.013
0.036
0.037
0.011
0.007
0.016
0.006
0.024
0.000
0.013
0.015
0.000
0.017
0.005
0.005
0.017
0.054
0.013
0.036
0.037
0.011
0.007
0.016
0.006
0.024
0.000
0.015
0.016
0.000
0.017
0.005
0.005
0.017
0.054
29189
36047
MO
NY
Saint Louis
Kings
33
24
1
1
0.0243
0.0131
0.005
0.011
0.005
0.011
0.005
0.011
0.006
0.011
0.007
0.012
0.008
0.013
0.050
0.014
0.050
0.016
0.050
0.018
0.055
0.019
0.055
0.019
39017iOH iButler
30
0.0055 i 0.003 i 0.003 i 0.004 i 0.004 i 0.005 i 0.006 i 0.006 i 0.007 i 0.007 i 0.008 i 0.008
39029
39035
OH
OH
Columbiana
Cuyahoga
105
105
3
3
0.0148
0.0144
0.005
0.005
0.005
0.006
0.007
0.006
0.008
0.008
0.010
0.010
0.013
0.013
0.017
0.018
0.021
0.023
0.028
0.027
0.054
0.033
0.057
0.035
November 2012
3-157
Draft - Do Not Cite or Quote
-------
Year
istate/
Season i
| County
39049
39143
39167
40115
42003
ST
OH
OH
OH
OK
PA
County name
Franklin
Sandusky
Washington
Ottawa
Allegheny
Site ID
N:mo
means
36
10
48
12
36
N
sites
1
1
2
2
1
Mean
0.0092
0.0052
0.0047
0.0128
0.0101
Min
0.005
0.004
0.002
0.005
0.000
1
0.005
0.004
0.002
0.005
0.000
5
0.005
0.004
0.002
0.005
0.000
10
0.005
0.004
0.003
0.006
0.000
25
0.008
0.005
0.004
0.010
0.007
50
0.010
0.005
0.004
0.014
0.012
75
0.011
0.006
0.006
0.016
0.014
90
0.011
0.006
0.007
0.018
0.016
95
0.012
0.006
0.007
0.019
0.018
99
0.012
0.006
0.008
0.019
0.025
max
0.012
0.006
0.008
0.019
0.025
42021
42045
42101
42129
48061
48141
48201
48479
49035
PA
PA
PA
PA
TX
TX
TX
TX
UT
Cambria
Delaware
Philadelphia
Westmoreland
Cameron
El Paso
Harris
Webb
Salt Lake
23
14
22
24
33
56
30
23
10
1
1
1
1
1
3
1
1
1
0.0459
0.0427
0.0214
0.0417
0.0042
0.0212
0.0051
0.0121
0.0145
0.040
0.040
0.013
0.037
0.002
0.014
0.004
0.006
0.007
0.040
0.040
0.013
0.037
0.002
0.014
0.004
0.006
0.007
0.040
0.040
0.014
0.040
0.003
0.014
0.004
0.007
0.007
0.040
0.040
0.014
0.040
0.003
0.015
0.004
0.007
0.007
0.040
0.040
0.018
0.040
0.004
0.016
0.005
0.008
0.008
0.041
0.042
0.022
0.041
0.004
0.018
0.005
0.010
0.011
0.046
0.045
0.025
0.043
0.005
0.023
0.006
0.016
0.016
0.069
0.046
0.029
0.046
0.005
0.038
0.006
0.021
0.032
0.070
0.047
0.029
0.047
0.006
0.040
0.007
0.022
0.036
0.073
0.047
0.030
0.048
0.006
0.040
0.007
0.026
0.036
0.073
0.047
0.030
0.048
0.006
0.040
0.007
0.026
0.036
Statistics for individual sites where overall average monthly mean > national 90th percentile (2008-2010)
170310022
170310026
170316003
291870006"
291870007"
291892003"
420210808"
420450002"
421290007"
36
36
36
10
11
33
23
14
24
0.0335
0.0281
0.0245
0.0412
0.0268
0.0243
0.0459
0.0427
0.0417
0.016
0.018
0.015
0.017
0.011
0.005
0.040
0.040
0.037
0.016
0.018
0.015
0.017
0.011
0.005
0.040
0.040
0.037
0.018
0.019
0.015
0.017
0.011
0.005
0.040
0.040
0.040
0.026
0.022
0.017
0.026
0.012
0.006
0.040
0.040
0.040
0.028
0.023
0.020
0.035
0.012
0.007
0.040
0.040
0.040
0.032
0.026
0.025
0.043
0.028
0.008
0.041
0.042
0.041
0.038
0.032
0.028
0.048
0.035
0.050
0.046
0.045
0.043
0.047
0.038
0.031
0.054
0.036
0.050
0.069
0.046
0.046
0.048
0.043
0.035
0.054
0.041
0.050
0.070
0.047
0.047
0.051
0.046
0.036
0.054
0.041
0.055
0.073
0.047
0.048
0.051
0.046
0.036
0.054
0.041
0.055
0.073
0.047
0.048
aThe 3-month averages presented here were created using a simplified approach of the procedures detailed in 40 CFR part 50 appendix R and as such cannot be directly compared to the Pb NAAQS for
determination of compliance with the Pb NAAQS.
"Sites listed in the bottom six rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-158
Draft - Do Not Cite or Quote
-------
Table 3-16 Distribution of annual 1-month site maxima TSP Pb concentrations (ug/m3) nationwide, source-
oriented monitors, 2008-2010.
Year
jSitelD-year
N (sites)
Mean
Min
10
25
50
75
90
95
99
max
Nationwide statistics
2008-2010
2008
2009
111
47
54
0.5003
0.8138
0.4486
0.003
0.012
0.016
0.006
0.012
0.016
0.016
0.052
0.022
0.032
0.057
0.050
0.066
0.096
0.090
0.156
0.320
0.170
0.575
0.850
0.618
1.530
2.557
1.280
2.416
3.123
1.623
4.225
4.440
2.438
4.440
4.440
2.438
2010
101
0.3105
0.003
0.006
0.008
0.024
0.054
0.142
0.347
0.854
1.117
1.576
1.828
Annual site max 1 -month means >= national 90th percentile (2008-20010)
060371405-2008
180350009-2008
290930016-2008
290930021-2008
290930021-2009
290990004-2008
290990004-2009
290990004-2010
2.8800
4.4400
4.2252
2.5566
2.4380
2.4156
1.5599
1.5762
1 29099001 1-2008 1.5295
1 29099001 5a-2008 3.1228
290990020a-2008
29099002 1a-2008
2.2204
1 .5528
i 290999001 a-2009 1.6228
1 290999001 a-2010
290999005a-2009
290999005a-2010
480850009a-2008
1 .5576
1 .9850
1 .8278
1 .5640
aSites listed in the bottom eight rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-159
Draft - Do Not Cite or Quote
-------
Table 3-17 Distribution of annual 1-month site maxima TSP Pb concentrations (ug/m3) nationwide, non-source-
oriented monitors, 2008-2010.
Year
jSitelD-year
N (sites)
Mean
Min
10
25
50
75
90
95
99
max
Nationwide statistics
2008-2010
2008
2009
88
59
66
0.0284
0.0232
0.0210
0.000
0.004
0.003
0.000
0.004
0.003
0.004
0.005
0.005
0.006
0.006
0.006
0.010
0.010
0.008
0.020
0.016
0.014
0.041
0.033
0.026
0.057
0.053
0.040
0.070
0.058
0.056
0.136
0.066
0.128
0.136
0.066
0.128
2010
73
0.0233
0.000
0.000
0.002
0.004
0.008
0.015
0.029
0.049
0.065
0.136
0.136
Annual site max 1-month means >= national 90th percentile (2008-2010)
170310022-2009
170310022-2010
0.0700
0.0620
1171193007-2008
0.0660
291870006a-2010
291892003a-2008
39029001 9a-2010
390290022a-2010
42021 0808a-2008
42021 0808a-2009
481410002a-2010
481410033a-2009
0.0894
0.0660
0.1360
0.0652
0.0583
0.1280
0.0870
0.0570
aSites listed in the bottom eight rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-160
Draft - Do Not Cite or Quote
-------
Table 3-18 Distribution of annual 3-month site maxima Pb-TSP concentrations (ug/m3) nationwide, source-
oriented monitors, 2008-2010.
Year
jSitelD-year
N (sites)
Mean
Min
10
25
50
75
90
95
99
max
Nationwide statistics3
2008-2010
2007
2008
2009
Annual site max
3-month means >= nation
011090003-2008
060371405-2008
120571066-2008
106
47
54
96
al 90th percent
0.3605
0.5831
0.3611
0.2112
ile (2008-2C
1.2100
2.4890
1 .7700
0.003
0.009
0.012
0.003
110)
0.005
0.009
0.012
0.003
0.016
0.038
0.017
0.011
0.023
0.043
0.035
0.021
0.047
0.085
0.060
0.046
0.109
0.242
0.121
0.091
0.378
0.815
0.467
0.262
1.204
2.017
1.079
0.630
1.937
2.456
1.258
0.865
2.489
2.889
2.070
1.375
2.889
2.889
2.070
1.375
180350009-2008
29093001 6b-2008
29093001 6b-2009
29093002 1b-2009
2.1630
2.4560
2.0700
1 .9370
290990004b-2008
29099001 5b-2008
290999001 b-2009
290999005b-2009
2.0170
2.8890
1 .2040
1 .2580
J290999005b-2010
|480850009b-2008
1 .3750
1 .2620
"The 3-month averages presented here were created using a simplified approach of the procedures detailed in 40 CFR part 50 appendix R and as such cannot be directly compared to the Pb NAAQS for
determination of compliance with the Pb NAAQS.
bSites listed in the bottom nine rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-161
Draft - Do Not Cite or Quote
-------
Table 3-19 Distribution of annual 3-month site maxima Pb-TSP concentrations (ug/m3) nationwide, non-source-
oriented monitors, 2008-2010.
Year
jSitelD-year
| N (sites) | Mean j Min j 1
10
25
50
75
90
95
99
max
Nationwide statistics3
2008-2010
86
0.0198 0.000 0.000 0.002 0.004 0.007 0.015 0.028 0.044 0.051 0.073 0.073
2008
2009
2010
59
65
71
0.0176
0.0162
0.0171
0.002
0.002
0.000
0.002
0.002
0.000
0.004
0.003
0.001
0.005
0.004
0.002
0.007
0.006
0.006
0.014
0.013
0.013
0.024
0.021
0.024
0.039
0.038
0.037
0.048
0.041
0.047
0.055
0.073
0.057
0.055
0.073
0.057
Annual site max 3-month means >= national 90th percentile (2008-2010)
170310022-2008
170310022-2009
0.0480
0.0470
170310022-2010
170310026b-2008
291870006b-2010
291892003b-2008
39029001 9b-2010
390290022b-2010
420210808b-2008
0.0510
0.0460
0.0540
0.0550
0.0570
0.0440
0.0490
420210808b-2009
420450002b-2010
421290007b-2008
0.0730
0.0470
0.0480
"The 3-month averages presented here were created using a simplified approach of the procedures detailed in 40 CFR part 50 appendix R and as such cannot be directly compared to the Pb
NAAQS for determination of compliance with the Pb NAAQS.
b Sites listed in the bottom nine rows of this table fall in the upper 90th percentile of the data pooled by site.
November 2012
3-162
Draft - Do Not Cite or Quote
-------
3
4
5
Concentrations of Pb Measured using PM10 Monitors (for Concentrations
and Trends)
Figure 3-38 displays maximum 3-month averages for Pb-PMi0 concentrations for 36
counties in which measurements were obtained. Among the 36 counties in which PM10
monitoring was conducted, only one county, Gila County, AZ, reported concentrations
above 0.076 ug/m3. Three other counties reported concentrations greater than
0.016 ug/m3: Wayne County, MI, Boyd County, KY, and the county of St. Louis City,
MO.
2007-2009 Pb-PM10 County Maximum 3-Month Mean
Concentration:
* >= 0.076 ng/m3 (1 county)
* 0.016 -0.075 n£/m (3 counties)
• 0.006 - 0.015 ug/m- (17 counties)
<= .005 ug/m- (15 counties)
_1 no data
Figure 3-38 Highest county-level Pb-PMi0 concentrations (ug/m ), maximum
3-month average, 2007-2009.
November 2012
3-163
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
Concentrations of Pb Measured using PM2.s Monitors (for Speciation
Concentrations and Trends)
Figure 3-39 displays maximum 3-month average county-level data for Pb in PM2 5
concentrations for 323 counties in which PM2 5 measurements were obtained for
speciation in the CSN and IMPROVE networks. The data presented here are not
compared to the NAAQS because PM2 5 monitors are not deployed for the purpose of
evaluating compliance for the NAAQS. Among the 323 counties in which PM25
monitoring was conducted, only eleven counties reported concentrations greater than
0.016 ug/m3: Jefferson, AL, San Bernardino, CA, Imperial, CA, Wayne, MI, Jefferson,
MO, Erie, NY, Lorain, OH, Allegheny, PA, Berks, PA, Davidson, TN, and El Paso, TX.
2007-2009 Pb-PM2 5 County Maximum 3-Month Mean
.v -? -V " __--
V * -- - -^~~ M
• 0.016 -0.075 pg/m-ill counties)
• 0.006 -0.015 yg/m!(71counties)
<= .005 H£/m: (241 counties)
Figure 3-39 Highest county-level Pb-PM2.s concentrations (ug/m ), maximum
3-month average, 2007-2009.
November 2012
3-164
Draft - Do Not Cite or Quote
-------
3.8.2 Intra-urban Variability
1 Intra-urban variability in Pb concentrations reported to AQS was described in detail for
2 Los Angeles County, CA (Los Angeles), Hillsborough and Pinellas Counties, FL
3 (Tampa), Cook County, IL (Chicago), Jefferson County, MO (Herculaneum), Cuyahoga
4 County, OH (Cleveland), and Sullivan County, TN (Bristol) were selected for this
5 assessment to illustrate the variability in Pb concentrations measured across different
6 metropolitan regions with varying Pb source characteristics. Four of the counties
7 encompass large cities (Los Angeles, Tampa, Chicago, and Cleveland). All six counties
8 contain source-oriented monitors. Maps and wind roses (graphs representing wind
9 direction and wind speed at a location) are presented in this Chapter 3 Appendix for each
10 of the six urban areas. Additionally, annual and seasonal box plots of the Pb
11 concentration distributions and intra-monitor correlation tables are presented to illustrate
12 the level of variability throughout each urban area.
13 Maps of six areas (Los Angeles County, CA; Hillsborough/Pinellas Counties, FL; Cook
14 County, IL; Jefferson County, MO; Cuyahoga County, OH; and Sullivan County, TN) are
15 shown to illustrate the location of all Pb monitors meeting the inclusion criteria. Wind
16 roses for each season are also provided to help put the source concentration data in
17 context. Letters on the maps identify the individual monitor locations and correspond
18 with the letters provided in the accompanying concentration box plots and pair-wise
19 monitor comparison tables. The box plots for each monitor include the annual and
20 seasonal concentration median and interquartile range with whiskers extending from the
21 5th to the 95th percentile. Data from 2008-2010 were used to generate the box plots,
22 which are stratified by season as follows: 1 = winter (December-February), 2 = spring
23 (March-May), 3 = summer (June-August), and 4 = fall (September-November). The
24 comparison tables include the Pearson correlation coefficient (R), Spearman rank-ordered
25 correlation coefficient (p), the 90th percentile of the absolute difference in concentrations
26 (P90) in (ig/m3, the coefficient of divergence (COD) and the straight-line distance
27 between monitor pairs (d) in km. The COD provides an indication of the variability
28 across the monitoring sites within each county and is defined as follows:
CODJk =
p /_, \Xt] + Xtk
'-1
Equation 3A-1
29 where Xtj and Xik represent the observed hourly concentrations for time period / at sites j
30 and k, and/? is the number of paired hourly observations. A COD of 0 indicates there are
November 2012 3-165 Draft - Do Not Cite or Quote
-------
1 no differences between concentrations at paired sites (spatial homogeneity), while a COD
1 approaching 1 indicates extreme spatial heterogeneity.
3 In certain cases, the information contained in these figures and tables should be used with
4 some caution since many of the reported concentrations forthe years 2008-2010 are near
5 or below the analysis method's stated method detection limit (MDL). The MDL is
6 generally taken as 0.01 because it is the upper value of the range of MDLs reported for
7 atomic absorption (AA) and Emissions Spectra ICAP methods, which were the two
8 methods reported in the AQS to have been used for analysis of FRM samples (Rice.
9 2007). Generally, data are reported to the hundredth place, so this assumption is
10 reasonable. The approximate percentage of data below the MDL (to the nearest 5%) is
11 provided for each site along with box plots of seasonal Pb concentration at monitors
12 within each urban area studied.
13 Figure 3-40 illustrates Pb monitor locations within Los Angeles County, CA. Ten
14 monitors are located within Los Angeles County, five of which were source-oriented and
15 the other five were non-source-oriented monitors. Monitor A was located immediately
16 downwind of the Quemetco battery recycling facility in the City of Industry, CA. This
17 source was estimated to produce 0.32 tons of Pb/yr (U.S. EPA. 2008c). Monitor C was
18 sited in a street canyon just upwind of the Exide Pb recycling facility, which was
19 estimated to produce 2.0 tons of Pb/yr (U.S. EPA. 2008c). Monitor D was situated
20 slightly northwest of the same Pb recycling facility. It is still in relatively close proximity
21 but not downwind on most occasions. Monitor B was located 12 km downwind of the
22 Exide facility. Monitor E was located nearby the Trojan Battery recycling facility, which
23 emitted 0.79 tons Pb/yr (U.S. EPA. 2008c). Location of the non-source-oriented monitors
24 varied. Monitor F was positioned on a rooftop 60 meters away from a 4-lane arterial road
25 and 100 meters from of a railroad. Monitor G was located on a rooftop approximately
26 20 meters from an 8-lane arterial road, and monitor H was positioned at the curbside of a
27 four-lane road roughly 650 meters north of that road's junction with Interstate 1-405.
28 Monitor I was sited in a parking lot roughly 80 meters from a four-lane road, and monitor
29 J was located approximately 130 meters south of a 4-lane highway. Figure 3-41 displays
30 seasonal wind roses for Los Angeles County. In spring, summer, and fall, the
31 predominant winds come from the west-southwest. During winter, wind direction varies
32 with a portion from the west-southwest and the remainder from the east. The highest
33 winds during winter come more frequently from the west-southwest.
34 The maps shown in Figure 3-40 for source-oriented monitors A-E illustrate the different
35 conditions captured by the monitors; this informs analysis of the seasonal and year-round
36 concentrations reported in Figure 3-42. The average annual concentration at monitor A
37 was 0.074 ug/m3. The 95th percentile exceeded the level of the NAAQS in the spring
November 2012 3-166 Draft - Do Not Cite or Quote
-------
1 (0.16 ug/m3) and summer (0.18 ug/m3). Monitor C reported the highest concentrations in
2 Los Angeles County, with a year-round mean of 0.68 ug/m3. Given the position of this
3 monitor with respect to the Exide facility, there is the potential for recirculation of
4 fugitive Pb emissions in the air sampled by that monitor. The average annual Pb
5 concentration at monitor D was 0.12 ug/m3, and the 75th percentile of year-round data
6 exceeded the level of the NAAQS; in spring, the 70th percentile exceeded 0.15 ug/m3.
7 Monitor B reported the lowest values among the source-oriented monitors with an
8 average annual concentration of 0.013 ug/m3. Note that 75% of reported values were
9 below the MDL for this site, and no data from this site exceeded the level of the NAAQS.
10 The annual average concentration at monitor E was 0.068 ug/m3, and the 95th percentile
11 of concentration was 0.17 ug/m3.
12 The non-source-oriented monitors located at sites F-J all recorded low concentrations,
13 with average values ranging from 0.004 to 0.018 ug/m3 (Figure 3-42). The highest
14 average year-round concentrations were recorded at site F. The 95th percentiles at these
15 sites ranged from 0.01 to 0.04 ug/m3. There is much less certainty in the data recorded at
16 the non-source-oriented sites, because 45-95% of the data from these monitors were
17 below the MDL. Additionally, only one of the non-source-oriented monitors (monitor H)
18 was positioned at roadside, and none of the non-source-oriented monitors were located at
19 the side of a major highway.
20 Intersampler correlations (Table 3-20). illustrate that Pb has high intra-urban spatial
21 variability. For the source-oriented monitors, the highest correlation (R = 0.59, p = 0.57)
22 occurred for monitors C and D, which covered the same site. Because monitor D was
23 slightly farther from the Exide source and slightly upstream of the predominant wind
24 direction, the signal it received from the source site was correspondingly lower. Hence,
25 the correlation between these sites was moderate despite their relatively close proximity.
26 In general, low or even negative correlations were observed between the source-oriented
27 and non-source-oriented monitors. The exception to this was the Spearman-ranked
28 correlation between source-oriented monitor B and non-source-oriented monitor F, with
29 p = 0.74. Pearson correlation was much lower for this pair (R = 0.33). Monitors B and F
30 are roughly 16 km apart, whereas monitor B is only 12 km from monitors D and C, 8 km
31 from monitor E, and 6 km from monitor A. It is possible that monitors B and F both
32 captured a source that was either longer in range or more ubiquitous and so would have
33 been obscured by the stronger source signals at sites A, C, D, and E. Comparisons
34 between the non-source-oriented monitors revealed moderate correlation between sites
35 (G to J [R = 0.29 to 0.71, p = 0.37 to 0.65]). Sites G, H, I and J are all located in the
36 southwestern quadrant of Los Angeles. It is possible that they are also exposed to a
37 ubiquitous source that produces a common signal at these four sites.
November 2012 3-167 Draft - Do Not Cite or Quote
-------
Legend
0 TSP Source Monitors
• TSP Non-source Monitors
• City-based Population Center
• County-based Population Center
Inters) ate s
Mitjof Highways
Bodies o( Waler
Urban Areas
V.CA
0 10 20 40 Kilometers
Note: Monitor locations are denoted by green markers, and source locations are denoted by red markers. Top: view of all Pb FRM
monitors in Los Angeles County. Bottom left: Close up of the industrial site near monitors C and D. Bottom right: Close up of the
populated area captured by monitor F.
Figure 3-40 Pb TSP monitor and source locations within Los Angeles County,
CA (06-037), 2007-2009.
November 2012
3-168
Draft - Do Not Cite or Quote
-------
I
i
Note: Clockwise from top left: January, April, July, and October. Note that the wind percentages vary from month to month.
Source: NRCS (2011).
Figure 3-41 Wind roses for Los Angeles County, CA, from meteorological data
at the Los Angeles International Airport, 1961-1990.
November 2012
3-169
Draft - Do Not Cite or Quote
-------
Site
SITE ID
MEAN
SD
OBS
% BELOW
MDL
Source
orientation
A
06-037-
1404
0.074
0.040
66
0
Source
B
06-037-
1602
0.013
0.017
112
75
Source
C
06-037-
1405
0.68
1.0
617
0
Source
D
06-037-
1406
0.12
0.092
242
0
Source
E
06-037-
1403
0.068
0.052
128
0
Source
F
06-037-
1103
0.018
0.011
121
45
Non-
source
G
06-037-
1301
0.015
0.012
108
65
Non-
source
H
06-037-
4002
0.0083
0.0068
120
85
Non-
source
1
06-037-
4004
0.0087
0.0069
117
85
Non-
source
J
06-037-
5005
0.0040
0.0064
109
95
Non-
source
E
15
01
C.
o
J.U -
2.9-
2.8 -
2.7
2.6-
2.5
2.4-
2.3 -
2.2-
2.1 -
2.0-
1.9-
1.7-
1.6
1.4-
1.3 -
1.1 -
1.0-
0.9-
0.8 -
0.7
0.6-
0.5 -
0.4-
0.3 -
0.2-
0.1 -
0.0 -
A
*lt*+
B
= , 4
C
D
m
E
iii = i
F
G
*i
H
I
J
Y1
234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234
season
Figure 3-42 Box plots of annual and seasonal 24-h Pb TSP concentrations
(ug/m3) from source-oriented and non-source-oriented monitors
within Los Angeles County, CA (06-037), 2007-2009.
November 2012
3-170
Draft - Do Not Cite or Quote
-------
Table 3-20 Comparisons between Pb TSP concentrations from source-oriented and non-source-oriented
monitors within Los Angeles County, CA (06-037), 2007-2009.
A Source R
P
P90
COD
B Source R
P
P90
COD
C Source R
P
P90
COD
D Source R
P
P90
COD
E Source R
P
P90
COD
ABC
Source Source Source
1.00 -0.04 0.14
1.00 0.16 0.10
0.00 0.08 0.49
0.00 0.63 0.64
1.00 0.06
1.00 0.05
0.00 3.59
0.00 0.96
1.00
1.00
0.00
0.00
D
Source
0.10
0.08
0.10
0.31
0.17
0.05
0.25
0.84
0.59
0.57
1.76
0.68
1.00
1.00
0.00
0.00
E
Source
0.17
0.27
0.10
0.34
-0.06
0.07
0.10
0.71
0.08
0.03
2.14
0.77
0.18
0.12
0.17
0.42
1.00
1.00
0.00
0.00
F
Non-
Source
0.03
-0.15
0.08
0.57
0.33
0.74
0.02
0.46
0.12
-0.08
3.59
0.95
0.33
0.17
0.24
0.78
0.05
0.13
0.10
0.61
G
Non-
Source
0.00
0.00
0.06
0.57
0.29
0.12
0.02
0.48
0.24
0.26
4.22
0.96
0.09
0.11
0.25
0.80
0.07
0.06
0.10
0.64
H
Non-
Source
-0.08
0.14
0.08
0.79
0.40
0.28
0.01
0.61
0.28
0.28
3.59
0.98
0.32
0.24
0.25
0.89
0.00
0.24
0.11
0.78
1
Non-
Source
-0.07
-0.02
0.08
0.77
0.22
0.11
0.02
0.60
0.18
0.20
3.59
0.98
0.20
0.21
0.25
0.89
0.09
0.07
0.11
0.79
J
Non-
Source
-0.27
-0.09
0.08
0.85
0.20
0.10
0.02
0.81
0.08
0.13
3.92
0.99
0.03
0.07
0.25
0.95
-0.07
0.18
0.11
0.90
November 2012
3-171
Draft - Do Not Cite or Quote
-------
F Non-Source R
P
P90
COD
G Non-Source R
P
P90
COD
H Non-Source R
P
P90
COD
1 Non-Source R
P
P90
COD
J Non-Source R
P
P90
COD
ABCDEFGH
Non- Non- Non-
Source Source Source Source Source
Source Source Source
1.00 0.10 0.43
1.00 0.02 0.19
0.00 0.02 0.02
0.00 0.39 0.61
1.00 0.71
1 .00 0.65
0.00 0.01
0.00 0.54
1.00
1.00
0.00
0.00
1
Non-
Source
0.34
0.09
0.02
0.58
0.55
0.39
0.02
0.61
0.60
0.51
0.01
0.55
1.00
1.00
0.00
0.00
J
Non-
Source
0.21
0.09
0.02
0.82
0.54
0.38
0.02
0.85
0.51
0.40
0.01
0.77
0.29
0.37
0.01
0.78
1.00
1.00
0.00
0.00
Each comparison contains (in order): Pearson rank-order correlation (R), Spearman rank-order correlation (p), the difference between the 90th and 10th percentile data (P90), and the
coefficient of divergence (COD).
November 2012 3-172 Draft - Do Not Cite or Quote
-------
1 Figure 3-43 illustrates Pb monitor locations within Hillsborough and Pinellas Counties in
2 FL, which comprise the greater Tampa-St. Petersburg metropolitan area. Two source-
3 oriented monitors (A and B) were located within Hillsborough County, and one non-
4 source-oriented monitor (C) was located in Pinellas County. Monitor A was located
5 360 meters north-northeast of the EnviroFocus Technologies battery recycling facility,
6 which produced 1.3 tons/year (U.S. EPA. 2008d). and monitor B was located 320 meters
7 southwest of the same facility. Monitor C was located next to a two-lane road in Pinellas
8 Park, FL.
9 Figure 3-44 displays seasonal wind roses for the Tampa-St. Petersburg metropolitan area.
10 These wind roses suggest shifting wind directions throughout the winter, spring, and
11 summer. During the winter, the highest winds came from the north and northeast with
12 little influence from the west and southwest. During spring and summer, easterly and
13 westerly winds were evident from the wind rose, with winds from the west being slightly
14 higher in wind speed. During autumn, winds came predominantly from the northeast with
15 little signal from the west or south.
16 Seasonal and year-round concentrations are reported for Hillsborough and Pinellas
17 Counties in Figure 3-45. The average annual concentration at monitor A was 0.15 ug/m3,
18 and the 95th percentile was 0.70 ug/m3. During winter, the 60th percentile of the data met
19 the level of the NAAQS. At this site, the highest concentrations occurred during summer,
20 which corresponded to the time when westerly winds were stronger. Concentration data
21 at monitor B were much higher, with an annual average of 0.45 ug/m3 and a 95th
22 percentile of 1.9 ug/m3. Annually, the 55th percentile exceeded the level of the NAAQS,
23 and in autumn the 45th percentile exceeded the NAAQS. The highest concentrations
24 occurred in autumn, coinciding with the time when winds blew from the northeast, when
25 monitor B was most often downwind of the battery recycling facility. The non-source-
26 oriented monitor C always reported concentrations of 0.0 ug/m3. This is likely related to
27 its location next to a quiet road in a small city.
28 Intersampler correlations, shown in Table 3-21. illustrate that Pb has high intra-urban
29 spatial variability. The source-oriented monitors were anticorrelated (R = -0.09,
30 p = -0.08). This was likely related to the fact that they were designated to monitor the
31 same source and were downwind of the source at different times.
November 2012 3-173 Draft - Do Not Cite or Quote
-------
Top: view of all Pb FRM monitors in Hillsborough and Pinellas Counties.
Bottom: Close up of industrial site around monitors A and B.
Figure 3-43 Pb TSP monitor locations within Hillsborough and Pinellas
Counties, FL (12-057 and 12-103), 2007-2009.
November 2012
3-174
Draft - Do Not Cite or Quote
-------
I
•
I
I
Note: Clockwise from top left: January, April, July, and October. Note that wind percentages vary from month to month.
Source: NRCS (2011).
Figure 3-44 Wind roses for Hillsborough/Pinellas Counties, FL, obtained from
meteorological data at Tampa International Airport, 1961-1990.
November 2012
3-175
Draft - Do Not Cite or Quote
-------
Site
SITE ID
12-057-1073
12-057-1066
12-103-3005
MEAN
0.15
0.45
0.00
SD
0.27
1.08
0.00
DBS
154
155
58
, BELOW MDL
20
95
Source orientation
E
1
c
O
c
(U
u
c
O
u
Source
Source
Non-source
3.0-
2.9 -
2.8 -
2.7-
2.6 -
2.5 -
2.4-
2.3 -
2.2 -
2.1 -
2.0 -
1.9 -
1.8 -
1.7 -
1.6 -
1.5 -
1.4 -
1.3 -
1.2 -
1.1 -
1.0 -
0.9 -
0.8 -
0.7-
0.6 -
0.5 -
0.4-
0.3 -
0.2 -
0.1 -
o.o -
/
B
C
u
Y1234 Y1234 Y1234
season
Figure 3-45 Box plots of annual and seasonal 24-h Pb TSP concentrations
(ug/m3) from source-oriented and non-source-oriented monitors
within Hillsborough and Pinellas Counties, FL (12-057 and
12-103), 2007-2009.
November 2012
3-176
Draft - Do Not Cite or Quote
-------
Table 3-21 Correlations between Pb TSP concentrations from source-oriented
and non-source-oriented monitors within Hillsborough and Pinellas
Counties, FL (12-057 and 12-103), 2007-2009.
A Source R
P
P90
COD
g Source R
P
P90
COD
Q Non-source R
P
P90
COD
A B
Source Source
1 .00 -0.09
1 .00 -0.08
0.00 1.20
0.00 0.71
1.00
1.00
0.00
0.00
C
Non-source
0.50
1.00
2.20
1.00
1.00
1.00
0.00
0.00
Each comparison contains (in order): Pearson rank-order correlation (R), Spearman rank-order correlation (p), the difference
between the 90th and 10th percentile data (P90), and the coefficient of divergence (COD).
1
2
o
5
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
Figure 3-46 illustrates Pb monitor locations within Cook County, IL. Eight monitors were
located within Cook County, four of which were designated by the Illinois Environmental
Protection Agency (IEPA) in data reported to the AQS as source-oriented and the other
four were non-source-oriented monitors. Monitor A was situated within 10 km of 6
sources ranging in emissions from 0.14 to 1.08 tons/year (U.S. EPA. 2008a). Monitor A
was also sited in the median of Interstate I-90/I-94. Monitor B was located on the
northern roadside of Interstate 1-290, 5 meters from the closest lane of traffic and was
within 10 km of 2 Pb sources (0.41 and 1.08 tons/year) (U.S. EPA. 2008a). Monitor C
was also located within 10 km of 6 sources in Cook County and Lake County, IN; the
largest of those sources was 2.99 tons/year and was located 8 km southeast of monitor C
(U.S. EPA. 2008a). Monitor C was placed on the roof of a high school. Monitor D was
located roughly 60 meters west of Interstate 1-294 and adjacent to O'Hare International
Airport. Monitor E was located on the rooftop of a building rented for government offices
in Alsip, IL, a suburb south of Chicago. This location was roughly 1 km north of
Interstate 1-294 but not located on an arterial road; it was 9 km southeast of a
0.56 tons/year source (U.S. EPA. 2008a). Monitor F was sited in the parking lot of a
water pumping station, 100 meters north of Interstate 1-90 and 300 meters northwest of
the junction between Interstates 1-90 and 1-94. This site was 2 km north-northwest of a
0.10 tons/year source (U.S. EPA. 2008a). Monitor G was situated atop an elementary
school in a residential neighborhood on the south side of Chicago, roughly 100 meters
November 2012
3-177
Draft - Do Not Cite or Quote
-------
1 south of a rail line and over 300 meters west of the closest arterial road. Although not
2 designated as a source monitor, monitor G was located 2 km southwest of facilities
3 emitting 0.30 and 0.41 tons/year (U.S. EPA. 2008a). Monitor H was sited on the grounds
4 of the Northbrook Water Plant. Interstate 1-94 curves around this site and was
5 approximately 700 meters from the monitor to the east and around to the north. Figure
6 3-47 displays seasonal wind roses for Cook County. Wind patterns were quite variable
7 during each season for this area. During the winter, winds mostly came from the west,
8 with smaller contributions from the northwest, southwest, and south. In spring,
9 measurable winds were omni-directional, with the highest winds coming from the south
10 and northeast. Winds originated predominantly from the southwest and south during the
11 summer, with measurable contributions from the northeast as well. In autumn, wind flow
12 was predominantly from the south, but smaller contributions also came from the
13 southwest, west, and northwest.
14 Figure 3-48 presents seasonal box plots of Pb concentration at the eight monitors located
15 within Cook County. The maximum 95th percentile concentration on this plot was
16 0.14 ug/m3, so the scale of this box plot makes the variability in these data appear wider
17 than the data presented for Los Angeles County and Hillsborough/Pinellas Counties.
18 Monitor C was in closest proximity to the industrial steel facilities located in Lake
19 County, IN. The average of concentrations measured at monitor C was 0.031 ug/m3, with
20 a median of 0.02 ug/m3 and a maximum concentration of 0.31 ug/m3. In winter, the 95th
21 percentile of data was 0.14 ug/m3. The higher values could potentially be attributed to
22 transport of emissions; winds blow from the southeast roughly 10-15% of the time
23 throughout the year. No other monitors in Cook County reported values above the level
24 oftheNAAQS.
25 Three "near-road" monitors, A, B, and D can be compared with the other monitors to
26 consider the possibility of roadside resuspension of Pb dust from contemporaneous
27 sources, as discussed in Section 3.2.2.6. It would be expected that resuspension would
28 diminish with distance from the road. The 2 roadside monitors, A and B, reported
29 average concentrations of 0.030 ug/m3 and 0.024 ug/m3, respectively. The median
30 concentrations for monitors A and B were 0.02 ug/m3. Fifteen percent of data were below
31 the MDL for monitor A, and 25% were below the MDL for monitor B. Note that data
32 obtained from monitor A may reflect industrial emissions as well. Monitor D was located
33 roughly 60 meters from the closest interstate and 570 meters from the closest runway at
34 O'Hare International Airport. The average concentration at this site was 0.012 ug/m3, and
35 85% of data were below the MDL. Non-source monitors, E, F, G, and H had average
36 concentrations of 0.011-0.017 ug/m3. It is possible that the difference between Pb
37 concentrations at monitors A and B and Pb concentrations at the other monitors was
November 2012 3-178 Draft - Do Not Cite or Quote
-------
1 related to proximity to the roadway, although this cannot be stated with certainty without
2 source apportionment data to confirm or refute the influence of industrial plumes from
3 Lake County, IN or local sources at each of the monitors.
4 Comparison among the monitor data demonstrates a high degree of spatial variability
5 (Table 3-22). None of the source-oriented monitors were well correlated with each other.
6 The highest correlation between source-oriented monitors occurred for monitors (A and
7 B [R = 0.32, p = 0.26]). This might have reflected more substantial differences related to
8 the additional influence of industrial sources nearby monitor A. Monitors (C and D) were
9 uncorrelated with each other and with monitors (A and B), likely because their exposure
10 to sources was substantially different. The source-oriented and non-source-oriented
11 monitors were generally not well correlated. The highest Spearman correlation occurred
12 between monitors D and H (p = 0.53), but Pearson correlation was much lower for this
13 pair (R = 0.19). Both were located on the north side of Cook County, but monitor H was
14 roughly 20 km northeast of monitor D. Winds blew from the southwest roughly 20-30%
15 of the time throughout the year and from the northeast 20-25% of the time between the
16 months of March and July, so the correlation may have been related to a common signal
17 transported across both sites. Monitors B and F (R = 0.52, p = 0.46) were also moderately
18 correlated. Monitor F is roughly 12 km northeast of monitor B, so the same common
19 wind influence for monitors D and H may have also caused the moderate correlation
20 between monitors (B and F). Monitor F was also moderately correlated with the other 3
21 non-source monitors (R = 0.42 to 0.54, p = 0.36 to 0.45), and the correlation between
22 monitors (E and G) was moderate (R = 0.65, p = 0.40). The data from monitor H did not
23 correlate well with those from monitors E and G. The non-source monitors were oriented
24 from north to south over a distance of roughly 50 km in the following order: monitor H,
25 monitor F, monitor G, and monitor E. The correlation pattern may have been related to
26 distance between samplers. Monitor H was located in the suburb of Northbrook, monitors
27 F and G were sited within the Chicago city limits, and monitor E was situated in a town
28 near the south side of Chicago. Differences among land use may have been related to the
29 lack of correlation of the monitor H data with those from monitors E and G. It is likely
30 that data from monitor F was at times belter correlated with monitors E and G and at
31 other times with monitor H, since it had moderate correlation with all three other
32 non-source monitors.
November 2012 3-179 Draft - Do Not Cite or Quote
-------
Legend
O TSP Source Monitors
• TSP Non-source Monitors
• City-based Population Cenlei
• County-based Population Cenlei
— I nlor states
Major Highways
Bodies of Water
Urban Aieas
Cook County. IL
"-«**HE
0 S 10
Top: view of all Pb FRM monitors in Cook County.
Bottom left: Close up of the high traffic site around monitor A.
Bottom right: Close up of O'Hare International Airport adjacent to monitor D.
Figure 3-46 Pb TSP Monitor locations within Cook County, IL (17-031),
2007-2009.
November 2012
3-180
Draft - Do Not Cite or Quote
-------
I
I
I
I
Note: Clockwise from the top left: January, April, July, and October. Note that the wind percentages vary from month to month.
Source: NRCS (2011)
Figure 3-47 Wind roses for Cook County, IL, obtained from meteorological
data at O'Hare International Airport, 1961-1990.
November 2012
3-181
Draft - Do Not Cite or Quote
-------
Site
A
B
C
D
E F
G H
SITE ID 17-031-0026 17-031-6003 17-031-0022 17-031-3103 17-031-0001 17-031-0052 17-031-3301 17-031-4201
MEAN
SD
DBS
% BELOW
MDL
Source
orientation
0.15 -
0.14 -
0.13 -
0.12 -
0.11 -
0.10 -
- 0.09 -
E
^ 0.08 -
° 0.07 -
4-1
c 0.06 -
01
o
§ 0.05 -
0.04 -
0.03 -
0.02 -
0.01 -
0.00 -
0.030
0.020
179
15
0.024
0.013
175
25
Source
Source
0.031
0.036
177
25
0.012
0.0062
168
85
Source Source
0.013 0.017
0.0078 0.0098
177 175
75 55
Non-source Non-
source
|
I
A
I
I
B
1
f
||
11
1
1
C
i
|
II
1
It
|
1
||
'I
D
LI
E
.nil
F
III
0.017 0.011
0.0097 0.0031
171 168
50 95
Non- Non-source
source
G
I
III!
H
.i.l.
Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234
season
Figure 3-48 Box plots of annual and seasonal 24-h Pb TSP concentrations
(ug/m3) from source-oriented and non-source-oriented monitors
within Cook County, IL (17-031), 2007-2009.
November 2012
3-182
Draft - Do Not Cite or Quote
-------
Table 3-22 Correlations between Pb TSP concentrations from source-oriented
and non-source-oriented monitors within Cook County, IL (17-031),
2007-2009.
A
B
C
D
E
F
G
H
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Non-Source R
P
P90
COD
Non-Source R
P
P90
COD
Non-Source R
P
P90
COD
Non-Source R
P
P90
COD
A B C D
Source Source Source Source
1.00 0.32 0.00 0.05
1.00 0.26 -0.01 0.08
0.00 0.03 0.06 0.04
0.00 0.29 0.38 0.43
1.00 0.14 0.07
1.00 0.05 0.10
0.00 0.04 0.03
0.00 0.33 0.36
1.00 0.01
1.00 0.04
0.00 0.05
0.00 0.40
1.00
1.00
0.00
0.00
Each comparison contains (in order): Pearson rank-order correlation (R), Spearman
90th and 10th percentile data (P90), and the coefficient of divergence (COD).
Figure 3-49 illustrates Pb monitor locations
E
Non-
Source
0.17
0.06
0.04
0.41
0.54
0.32
0.03
0.34
0.24
0.16
0.05
0.39
0.18
0.21
0.01
0.19
1.00
1.00
0.00
0.00
F
Non-
Source
0.39
0.32
0.03
0.36
0.52
0.46
0.02
0.29
0.05
0.10
0.04
0.35
0.12
0.37
0.01
0.24
0.42
0.36
0.02
0.24
1.00
1.00
0.00
0.00
G
Non-
Source
0.34
0.18
0.03
0.36
0.60
0.35
0.02
0.30
0.19
0.17
0.05
0.35
0.08
0.07
0.02
0.28
0.65
0.40
0.01
0.24
0.54
0.41
0.01
0.24
1.00
1.00
0.00
0.00
H
Non-
Source
0.06
0.06
0.04
0.45
0.06
-0.01
0.03
0.40
-0.04
0.06
0.05
0.42
0.19
0.53
0.01
0.15
-0.01
0.07
0.01
0.20
0.42
0.45
0.02
0.26
0.01
0.05
0.02
0.27
1.00
1.00
0.00
0.00
rank-order correlation (p), the difference between the
with Jefferson County, MO. Ten source-
oriented monitors surrounded the Doe Run primary Pb smelter in Herculaneum, MO on
the west and northwestern sides. The largest distance between these monitors was
November 2012
3-183
Draft - Do Not Cite or Quote
-------
1 approximately 1.5 km. Monitor E located on the Doe Run facility roughly 20 meters west
2 of the nearest building. Monitors A, B, C, D, F, G, and H were all located approximately
3 200 meters west of the facility. Monitors D, E, and H were situated alongside service
4 roads to the facility. Monitor I was sited 100 meters north of the smelter, and monitor J
5 was located approximately 600 meters northwest of the facility. The Doe Run smelter
6 was the only active primary smelter in the U.S. at the time of this review, and the facility
7 was estimated to have emitted 41.1 tons Pb/yr (U.S. EPA. 2008f). Figure 3-50 displays
8 seasonal wind roses for Jefferson County. During winter, predominant winds originated
9 from the northwest, with a smaller fraction of calmer winds originating in the south-
10 southeast. During the spring, the south-southeasterly winds became more prevalent with a
11 measurable fraction of stronger winds still originating in the north-northwest. In the
12 summer, winds were omni-directional and generally calmer. A slightly larger percentage
13 came from the south compared with other wind directions. Autumn winds were most
14 predominantly south-southeastern, with a smaller fraction from the west and northwest.
15 Figure 3-51 illustrates the seasonal distribution of concentrations at monitors A-J in
16 Jefferson County. The annual average concentrations ranged from 0.18 to 1.36 ug/m3
17 across the monitors. The maximum concentration was measured at monitor C to be
18 21.6 ug/m3, which was 144 times higher than the level of the standard. For this monitor,
19 the 25th percentile of the data was at the level of the standard. In general, median and
20 75th percentile concentrations were highest during the springtime and second highest
21 during the fall. These seasons coincide with periods when the southeastern winds were
22 stronger and more prevalent. Because the Doe Run facility had two 30-meter stacks
23 (Bennett, 2007). it is possible that the Pb measured at the closer monitors were due to
24 either fugitive emissions from the plant; or, if vechiles and ground equipment were
25 operated nearby, the previously-deposited emissions from the plant were resuspended.
26 Spatial variability among the monitors is lower than at many sites, because the monitors
27 are relatively close together and are located on one side of the same source (Table 3-23).
28 Correlations range substantially (R = -0.03 to 0.96, p = -0.04 to 0.96). High correlations
29 (R > 0.75, p > 0.75) occurred for monitors (A and C), (A and D), (C and D), (D and F),
30 (E and F), (G and H), and (I and J). Monitors (A and C), (A and D), (C and D), (D and F),
31 (E and F), and (G and H) are all within 250 meters of each other. For the highest
32 correlation (R = 0.96, p = 0.96, [for monitors E and F]), monitor F is 250 meters directly
33 east of monitor E. Low correlation (R < 0.25, p < 0.25) generally occurred when monitors
34 B, I, and J were compared with monitors A, C, D, E, F, G, and H. Monitors B, I, and J
35 were on the outskirts of the measurement area and so were likely oriented such that the
36 southeasterly winds did not carry pollutants to these sites concurrently with the signal
37 recorded by the other monitors.
November 2012 3-184 Draft - Do Not Cite or Quote
-------
1-MIJfll.l
» T&PSoulcs Mwllor*
• c-ty wsea Population C»nl«r
• Coui%-tkl«*d Populalion C»nw
Note: All monitors surround the Doe Run industrial facility. Top: Map view of all monitors in Jefferson County. Bottom: Satellite view
of the monitors and the Doe Run facility.
Figure 3-49 Pb TSP Monitor locations within Jefferson County, MO (29-099),
2007-2009.
November 2012
3-185
Draft - Do Not Cite or Quote
-------
i
I
i
•
Note: Clockwise from top left: January, April, July, and October. Note wind percentages vary from month to month.
Source: NRCS (2011)
Figure 3-50 Wind roses for Jefferson County, MO, obtained from
meteorological data at St. Louis/Lambert International Airport,
1961-1990.
November 2012
3-186
Draft - Do Not Cite or Quote
-------
Site
SITE ID
MEAN
SD
OBS
% BELOW
MDL
Source
orientation
7.0-
A
29-099-00
22
0.43
0.54
622
0
Source
B
C
29-099-002 29-099-00
4 15
0.36
0.49
209
5
Source
1.36
1.97
1E3
0
Source
D E F G
29-099-002 29-099-00 29-099-00 29-099-002
3 04 20 1
0.39 1.12 0.69 0.75
0.54 1.67 1.01 1.25
632 1E3 575 953
0505
Source Source Source Source
I I I
H
29-099-00
05
0.29
0.59
351
25
Source
1
29-099-001
1
0.34
0.85
366
5
Source
J
29-099-00
13
0.18
0.33
177
15
Source
7.0 -
6.5-
6.0 -
5.5 -
5.0 -
4.5 -
4.0 -
3.5 -
3.0
2.5 -
2.0-
1.5-
1.0-
0.5 -
A
I.
II
!
i
B
ll
III
C
D
E
I
F
|
!
G
H
,
ll
I
!
I
,
[I
ii
j
i.i.i
Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234 Y1234
season
Figure 3-51 Box plots of annual and seasonal 24-h Pb TSP concentrations
(ug/m3) from source-oriented and non-source-oriented monitors
within Jefferson County, MO (29-099), 2007-2009.
November 2012
3-187
Draft - Do Not Cite or Quote
-------
Table 3-23 Correlations between Pb TSP concentrations from source-oriented
and non-source-oriented monitors within Jefferson County, MO
(29-099), 2007-2009.
A
B
C
D
E
F
G
H
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
Source R
P
P90
COD
A B C D
Source Source Source Source
1.00 0.66 0.80 0.84
1.00 0.59 0.80 0.83
0.00 0.71 1.55 0.42
0.00 0.46 0.48 0.30
1 .00 0.54 0.40
1 .00 0.53 0.43
0.00 1 .86 0.87
0.00 0.58 0.51
1 .00 0.86
1 .00 0.86
0.00 1.56
0.00 0.50
1.00
1.00
0.00
0.00
E
Source
0.60
0.57
1.93
0.55
0.15
0.10
2.77
0.69
0.56
0.59
2.26
0.50
0.70
0.71
1.83
0.50
1.00
1.00
0.00
0.00
F
Source
0.65
0.64
1.14
0.45
0.15
0.14
1.96
0.62
0.72
0.72
1.26
0.46
0.80
0.80
1.02
0.36
0.96
0.96
0.86
0.35
1.00
1.00
0.00
0.00
G
Source
0.33
0.33
1.41
0.57
0.08
0.07
2.08
0.68
0.28
0.26
2.94
0.60
0.41
0.41
1.38
0.53
0.57
0.54
2.16
0.49
0.56
0.56
1.13
0.47
1.00
1.00
0.00
0.00
H
Source
0.32
0.35
0.74
0.64
0.16
0.22
0.94
0.68
0.32
0.27
2.65
0.74
0.48
0.56
0.76
0.61
0.53
0.46
2.50
0.66
0.56
0.54
1.51
0.63
0.85
0.87
1.53
0.61
1.00
1.00
0.00
0.00
1
Source
0.07
0.07
0.92
0.67
0.11
0.10
1.04
0.65
-0.03
-0.04
3.18
0.73
0.17
0.14
0.88
0.63
0.09
0.06
3.09
0.70
0.12
0.10
1.74
0.65
0.36
0.28
2.10
0.63
0.24
0.20
0.89
0.67
J
Source
0.05
0.05
0.78
0.69
0.01
0.09
0.91
0.65
-0.03
0.04
2.60
0.73
0.10
0.18
0.70
0.66
0.14
0.16
2.57
0.72
0.20
0.19
1.40
0.70
0.34
0.38
2.08
0.66
0.33
0.30
0.56
0.65
November 2012
3-188
Draft - Do Not Cite or Quote
-------
1 Source R
P
P90
COD
J Source R
P
P90
COD
ABODE FGH 1
1.00
1.00
0.00
0.00
J
0.87
0.79
0.62
0.48
1.00
1.00
0.00
0.00
Each comparison contains (in order): Pearson rank-order correlation (R), Spearman rank-order correlation (p), the difference
between the 90th and 10th percentile data (P90), and the coefficient of divergence (COD).
1 Figure 3-52 illustrates Pb monitor locations in Cuyahoga County, OH. Five monitors are
2 located within Cuyahoga County, three of which were designated by the Ohio EPA
3 (OEPA) as source-oriented and the other two were non-source-oriented monitors.
4 Monitors A, B, and C were all located within 1-10 km of six 0.1 tons/year source
5 facilities and one 0.2 tons/year source (U.S. EPA. 2008g). Additionally, monitor B was
6 located 30 meters north of the Ferro Corporation headquarters. This facility was stated in
7 the 2005 NEI to have no emissions, but it was thought by the OEPA to be the source of
8 exceedances at this monitor (U.S. EPA. 2008g). Monitor A was sited roughly 300 meters
9 south of the Ferro Corporation facility. Monitor C was located 2.2 km west-northwest of
10 the 0.5 tons/year Victory White Metal Co. facility. Monitor C was also roughly 250
11 meters southeast of Interstate 1-490. Monitors D and E were designated as non-source-
12 oriented monitors, although monitor D was just 600 meters further from the Victory
13 White Metal facility than was monitor C. Monitor D was sited on a residential street
14 located 50 meters north of Interstate 1-490. Monitor E was located on the rooftop of a
15 building within 20 meters of a four-lane arterial road. Figure 3-53 displays seasonal wind
16 roses for Cuyahoga County. During winter, summer, and autumn, the predominant winds
17 were from the southwest, with stronger winds recorded during the winter. In the spring,
18 the strongest winds still emanated from the south-southwest, but measurable winds were
19 also scattered from the northeast to the northwest.
20 Figure 3-54 illustrates the seasonal distribution of Pb concentration data at the five
21 monitoring sites. The influence of southern winds, along with close proximity to a
22 potentially-emitting facility, could have caused the elevated concentrations observed at
23 monitor B (average: 0.10 ug/m3). The 80th percentile of data was at the level of the
24 NAAQS at this monitor, and during autumn the 60th percentile of data met the level of
25 the NAAQS. The maximum concentration during fall and for the monitor year-round was
26 0.22 ug/m3. Concentration data from all other monitors were below the level of the
27 NAAQS. For monitor A, the average concentration was 0.025 ug/m3, and the median
November 2012 3-189 Draft - Do Not Cite or Quote
-------
1 reached 0.04 ug/m3 during the summer. Maximum concentration at this monitor was
2 0.07 ug/m3. Concentrations at monitor C averaged 0.017 ug/m3, and those at monitors D
3 and E averaged 0.014 ug/m3 and 0.013 ug/m3, respectively. Maximum concentrations
4 reached 0.04 ug/m3 at all three monitors.
5 The level of spatial variability is illustrated by the intersampler correlations presented in
6 Table 3-24. Monitors A and B appear to be anticorrelated (R = -0.06, p = -0.13). If the
7 Ferro site was the dominant source in this area, then the anticorrelation was likely caused
8 by the positioning of monitors A and B on opposite sides of that facility. At any given
9 time, potential emissions from the Ferro plant may have affected monitors A and B at
10 distinct times. Monitors C, D, and E correlated moderately to well with each other
11 (R = 0.37 to 0.74, p = 0.67 to 0.77). Given that all 3 monitors are separated by roughly
12 2.8 km, it is possible that the relatively high correlations related to common sources, as
13 suggested in the previous paragraph. Little correlation was observed between the source-
14 oriented and non-source-oriented monitors.
November 2012 3-190 Draft - Do Not Cite or Quote
-------
Legend
TSP Source Monrtots
TSP Non-source Monitors
City-based Population Center
County-based Population Center
- Interslates
- Majw Highways
Bodies of Wfater
Urban Areas
Cuyahoga County, OH
Note: Top: view of all Pb FRM monitors in Cuyahoga County. Bottom left: Close up of industrial site around monitors A and B.
Bottom right: Close up of monitor D north of Interstate 1-490.
Figure 3-52 Pb TSP Monitor locations within Cuyahoga County, OH (39-035),
2007-2009.
November 2012
3-191
Draft - Do Not Cite or Quote
-------
I
I
I
I
Note: Clockwise from top left: Jan, April, July, and October. Note wind percentages vary from month to month.
Source: NRCS (2011)
Figure 3-53 Wind roses for Cuyahoga County, OH, obtained from
meteorological data at Cleveland/Hopkins International Airport,
1961-90.
November 2012
3-192
Draft - Do Not Cite or Quote
-------
Site
SITE ID
MEAN
SD
DBS
% BELOW MDL
Source orientation
0.25 -
0.24 -
0.23 -
0.22 -
0.21 ;
0.20 -
0.19 -
0.18 -
0.17 -
0.16 -
"E 0.15 -
"M 0.14 -
~ 0.13 -
•B 0.12 :
(D
£ 0.11 -
g 0.10 -
I °-09:
0.08 -
0.07 -
0.06 -
0.05 -
0.04 -
0.03 -
0.02 -
0.01 -
o.oo -
A
B
39-035-0050 39-035-0049
0.025
0.018
36
20
Source
0.10
0.060
36
0
Source
A
|
i
I
B
C
D E
39-035-0061 39-035-0038 39-035-0042
0.017
0.010
36
30
Source
0.014 0.013
0.0072 0.0076
35 36
45 45
Non-source Non-source
C
^
I
D
i(
i,
E
Lit
Figure 3-54
Y1234 Y1234 Y1234 Y1234 Y1234
season
Box plots of annual and seasonal 24-h Pb TSP concentrations
(ug/m3) from source-oriented and non-source-oriented monitors
within Cuyahoga County, OH (39-035), 2007-2009.
November 2012
3-193
Draft - Do Not Cite or Quote
-------
1
2
o
J
4
5
6
7
8
9
10
11
Table 3-24
A Source
B Source
C Source
D Non-Source
E Non-Source
Each comparison
between the 90th
Correlations between Pb TSP concentrations from source-oriented
and non-source-oriented monitors within Cuyahoga County, OH
(39-035), 2007-2009.
A
Source
R 1.00
p 1.00
P90 0.00
COD 0.00
R
P
P90
COD
R
P
P90
COD
R
P
P90
COD
R
P
P90
COD
B C
Source Source
-0.06 0.21
-0.13 0.24
0.18 0.05
0.64 0.33
1.00 0.26
1.00 0.31
0.00 0.18
0.00 0.69
1.00
1.00
0.00
0.00
D
Non-Source
0.17
0.19
0.04
0.35
0.43
0.24
0.19
0.71
0.74
0.77
0.01
0.17
1.00
1.00
0.00
0.00
contains (in order): Pearson rank-order correlation (R), Spearman rank-order correlation (p),
and 10th percentile data (P90), and the coefficient of divergence (COD).
Figure 3-55 illustrates Pb monitor locations within Sullivan County, TN
E
Non-Source
0.24
0.21
0.05
0.37
0.11
0.34
0.19
0.73
0.51
0.67
0.01
0.18
0.37
0.67
0.01
0.17
1.00
1.00
0.00
0.00
the difference
. Three source-
oriented monitors were situated around an Exide Pb recycling facility emitting
0.78 tons/year (U.S. EPA. 200810. Monitors A and C are positioned along the facility's
service road and are approximately 100 meters and 200 meters away from the facility,
respectively. Monitor A is directly next to the road, and monitor C is roughly 15 meters
from the road. Monitor B is located in the facility's parking lot roughly 50 meters from
the closest building. The facility and all three monitors are approximately 1.5 km
northwest of the Bristol Motor Speedway and Dragway racetracks, which hosts a variety
of auto races each year, including NASCAR, KART, and drag racing. Although the
NASCAR circuit no longer uses tetraethyl Pb as an anti-knock agent in its fuel, some of
the smaller racing circuits continue to do so. However, the speedway is rarely upwind of
November 2012
3-194
Draft - Do Not Cite or Quote
-------
1 the monitoring sites and so likely had minimal influence on the reported concentrations.
2 Figure 3-56 displays seasonal wind roses for Sullivan County. During winter and spring,
3 the predominant winds come from the southwest and west. In the summer, the percentage
4 of wind coming from the west and southwest is roughly equal to that for wind coming
5 from the east and northeast, although the easterly winds are calmer. During autumn,
6 winds come predominantly from the northeast and east, although these winds tend to be
7 calmer than those originating from the southwest and west.
8 The data presented in Figure 3-57 illustrates that concentrations above the level of the
9 NAAQS occurred frequently at the monitors. The average concentrations at monitors A,
10 B, and C were 0.11 ug/m3, 0.051 ug/m3, and 0.059 ug/m3, respectively. Median
11 concentrations were 0.08 ug/m3, 0.03 ug/m3, and 0.04 ug/m3, respectively. The 75th
12 percentile of year-round data at monitor A was at the level of the NAAQS, while the 95th
13 percentile of data were below the NAAQS level for monitors B and C. The maxima at
14 each monitor were 0.76 ug/m3, 0.26 ug/m3, and 0.43 ug/m3 for monitors A, B, and C. The
15 concentrations measured at monitor A tended to be higher because the predominant and
16 stronger winds came from the southwest, so in many cases monitor A was upwind of the
17 facility. It is possible that Pb that had either deposited or was stored in waste piles
18 became readily resuspended by traffic-related turbulence and was measured at monitor A
19 since that monitor was closest to the road. The slightly higher concentrations at monitor
20 A compared with those from monitor C are consistent with the southwestern winds.
21 Not surprisingly, the correlations of monitor A with monitors B and C (R = 0.06 to 0.14,
22 p = -0.04 to 0.13) were quite low (Table 3-25). The correlation between monitors B and
23 C was moderate (R = 0.31, p = 0.45). It makes sense that the correlation for these
24 monitors would be somewhat higher because they are both oriented to the east of the Pb
25 recycling facility, although monitor C is to the northeast and monitor B to the east-
26 southeast.
November 2012 3-195 Draft - Do Not Cite or Quote
-------
Legend
• TSPSource Monitors
* City-based Population Canter
• County-based Population Center
Interstate*
Major Highways
Bodies of Water
Urban Areas
Sullivan County, TN
0 S 10 20 Kilometers
Note: Top: Map, bottom: Satellite image. Monitors A, B, and C surround the Exide Pb recycling facility. Just to the southeast is the
Bristol motor speedway.
Figure 3-55 Pb TSP Monitor locations within Sullivan County, TN (47-163),
2007-2009.
November 2012
3-196
Draft - Do Not Cite or Quote
-------
i
•
S:
Source: NRCS (2011)
Note: Clockwise from top left: January, April, July, and October. Note that the wind percentages vary from month to month.
Figure 3-56 Wind roses for Sullivan County, TN, obtained from meteorological
data at Bristol/Tri City Airport, 1961-90.
November 2012
3-197
Draft - Do Not Cite or Quote
-------
Site
SITE ID
MEAN
SD
OBS
% BELOW MDL
Source orientation
0.44 -
0.42 -
0.40 -
0.38 -
0.36 -
0.34 -
0.32 -
0.30 -
0.28 -
mE 0.26 -
"M 0.24 -
"c 0.22 -
0
'•£ °-20 "
c 0.18 -
0)
c 0.16 -
0
u o.H -
0.12 -
0.10 -
0.08 -
0.06 -
0.04 -
0.02 -
o.oo -
A
47-163-3001
0.11
0.11
334
0
Source
A
B
47-163-3002
0.051
0.036
362
0
Source
B
c
47-1 63-3003
0.059
0.047
345
0
Source
C
Figure 3-57
Y1234 Y1234 Y1234
season
Box plots of annual and seasonal 24-h Pb TSP concentrations
(ug/m3) from source-oriented monitors within Sullivan County, TN
(47-163), 2007-2009.
November 2012
3-198
Draft - Do Not Cite or Quote
-------
Table 3-25 Correlations between Pb TSP concentrations from source-oriented
monitors within Sullivan County, TN (47-163), 2007-2009.
A Source R
P
P90
COD
B Source R
P
P90
COD
C Source R
P
P90
COD
A B
Source Source
1.00 0.06
1 .00 -0.04
0.00 0.21
0.00 0.47
1.00
1.00
0.00
0.00
C
Source
0.14
0.13
0.19
0.43
0.31
0.45
0.06
0.23
1.00
1.00
0.00
0.00
Each comparison contains (in order): Pearson rank-order correlation (R), Spearman rank-order correlation (p), the difference
between the 90th and 10th percentile data (P90), and the coefficient of divergence (COD).
3.8.3 Seasonal Variation in Pb Concentrations
1 Monthly average Pb concentrations averaged over multiple sites and over 3 years from
2 2008-2010 are shown for Pb-TSP from source-oriented monitors (Figure 3-58). Pb-TSP
3 from non-source-oriented monitors (Figure 3-59). Pb-PMi0 (Figure 3-60). and Pb-PM2 5
4 (Figure 3-61). For source-oriented Pb-TSP (Figure 3-58). monthly average concentrations
5 were determined from between 146 and 154 samples in each month. For non-source-
6 oriented TSP (Figure 3-59). monthly average concentrations were determined from
7 between 141 and 151 samples in each month. A winter minimum was observed with
8 December, January, and February exhibiting the three lowest monthly averages. In both
9 cases, there is little seasonal variation. Minor variations in monthly averages are probably
10 driven by exceptional events. Monthly median concentrations are very similar for all
11 months.
November 2012
3-199
Draft - Do Not Cite or Quote
-------
CO
2
-------
0.05
0.04-
0.03 -
0 0.02
0.01
0.00-
A
1
Ja
K
r
n
j|
F(
I
;b
M
ar
A
nr
M
ay
Ji
n
vlontr
J
j|
A
jg
S<
5P
O
ct
N
DV
^
D
K
3C
Note: Box and whisker plots are used for each month, with the box comprising the interquartile range of the data and the whiskers
comprising the range within the 5th to 95th percentiles. The median is noted by the red line, and the blue star denotes the mean.
Figure 3-59 Monthly non-source-oriented Pb-TSP average (ug/m3) over 12
months of the year, 2008-2010.
0.028 -
0.026 -
0.024 •
0.022 -
0.020 -
CO
j= 0.018-
D)
-3 0.016-
•B 0.014-
•E 0.012-
0)
^ 0.010-
o
0 0.008 -
0.006 •
0.004 -
0.002 -
0.000 -
T
-L
I
Jan
I
i
Feb
_3
r
=>=
1
Mar
3
f
->-
I
Apr
3
)c
-L-
1
May
^
Jun
— . —
r^
{-,
-L-
i
Jul
3
C
-L-
i
Aug
3
C
-1-
I
Sep
•}
c
1
I
Oct
__^_
3
f
J-
I
Nov
)
I
-L-
Dec
Month
Note: Box and whisker plots are used for each month, with the box comprising the interquartile range of data and the whiskers
comprising the range from 5th to 95th percentiles. The median is noted by the red line, and the blue star denotes the mean.
Figure 3-60 Monthly Pb-PMi0 average (ug/m3) over 12 months of the year,
2007-2009.
November 2012
3-201
Draft - Do Not Cite or Quote
-------
0.010-
0.009 -
0.008 -
ff 0.007-
E
g 0.006 -
•2 0.005 -
1 0.004 •
|
° 0.003 -
0.002 -
0.001 •
0.000 -
—
_l_
i
Jan
*
i
i
Feb
*
_L
Mar
3
C
J_
Apr
3
C
J_
i
May
-j
L.
Jr-
_l_
i
Jun
3
K
_l_
i
Jul
__^
3
C
_l_
i
Aug
p
t
_L
i
Sep
J_
i
Oct
J
C
i
Nov
3
C
_L
Dec
Month
Note: Box and whisker plots are used for each month, with the box comprising the interquartile range of the data and whiskers
comprising the range from 5th to 95th percentiles. The median is noted by the red line, and the blue star denotes the mean.
Figure 3-61 Monthly Pb-PM2 5 average (ug/m3) over 12 months of the year,
2007-2009.
i
2
3
4
5
For both Pb-PMi0 (Figure 3-60) and Pb-PM2 5, (Figure 3-61) there is also little seasonal
variation, with minor fluctuations in monthly averages probably driven by exceptional
events, and similar monthly median concentrations for all months. Pb-PMi0 monthly
average concentrations were determined from between 100 and 109 samples and
Pb-PM2.5 from between 866 and 1,034 samples each month.
3.8.4 Size Distribution of Pb-bearing PM
6 Table 3-26 presents data for co-located Pb-TSP, Pb-PMi0, and/or Pb-PM2 5 monitors.
7 Table 3-27 contains metadata for studies in Section 3.5.3 involving size distribution data,
8 and Table 3-28 contains the size distribution data for those studies. At times, the data
9 were extracted from figures in the original references.
November 2012
3-202
Draft - Do Not Cite or Quote
-------
Table 3-26 Correlations and average of the concentration ratios for co-located monitors, TSP versus PMio, TSP
versus PM2.s, and PM™ versus PM2.s.
Site ID*
060190008
060190008
060190008
060190008
060250004
060250005
060250005
060251003
060290004
060290004
060290014
060290014
060290014
060290014
060292004
060310003
060310004
060370002
060374002
CBSA
Fresno, CA
Fresno, CA
Fresno, CA
Fresno, CA
El Centra, CA
El Centra, CA
El Centra, CA
El Centra, CA
Bakersfield, CA
Bakersfield, CA
Bakersfield, CA
Bakersfield, CA
Bakersfield, CA
Bakersfield, CA
Bakersfield, CA
Hanford-Corcoran, CA
Hanford-Corcoran, CA
Los Angeles-Long Beach-
Santa Ana, CA
Los Angeles-Long Beach-
Santa Ana, CA
Land Type
Suburban
Suburban
Suburban
Suburban
Suburban
Suburban
Suburban
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Suburban
Unknown
Suburban
Suburban
Suburban
Avg
Years Corr Ratio
PMio: TSP
1995-2001 0.93 0.82
1995-2001 0.93 0.83
1996-2001 0.79 0.98
1996-2001 0.92 0.89
1995-2000 0.94 0.75
1995-2000 0.92 0.78
1995-2000 0.47 0.80
1995-2000 0.43 0.81
1995-2000 0.76 0.38
Years Corr
Avg
Ratio
PM2.5: TSP
1992-2001 0.82
1992-2001 0.80
0.59
0.56
1996-2001 0.77
1996-2001 0.91
0.73
0.64
1992-1994 0.90
1992-1994 0.94
1994-2000 0.96
1994-2000 0.92
1995-2000 0.28
1995-2000 0.27
0.55
0.43
0.51
0.53
0.60
0.62
1992-2000 0.62
0.30
Years
Corr
Avg
Ratio
PM2.s: PMio
1995-2001
1995-2001
1995-2001
1995-2001
1995-1996
1996-2001
0.99
0.96
0.99
0.98
0.96
0.99
0.77
0.82
0.79
0.77
0.80
0.71
1995-1995
0.81
0.72
1995-2000
1995-2000
1995-2000
1995-2000
1995-2000
1995-1998
1996-2000
1995-2000
1995-2000
0.96
0.91
0.84
0.98
0.74
0.97
0.95
0.89
0.91
0.71
0.77
0.80
0.72
0.74
0.83
0.77
0.59
0.62
November 2012
3-203
Draft - Do Not Cite or Quote
-------
060374002
060390001
060631008
060658001
060658001
060670010
060710014
060771002
060771002
060850004
060850004
060850004
060990002
060990002
060990005
060990005
061072002
170310022
170310052
171190010
Los Angeles-Long Beach-
Santa Ana, CA
Madera-Chowchilla, CA
NONE (Plumas Co., CA)
Riverside-San Bernardino-
Ontario, CA
Riverside-San Bernardino-
Ontario, CA
Sacramento-Arden-Arcade-
-Roseville, CA
Riverside-San Bernardino-
Ontario, CA
Stockton, CA
Stockton, CA
San Jose-Sunnyvale-
Santa Clara, CA
San Jose-Sunnyvale-
Santa Clara, CA
San Jose-Sunnyvale-
Santa Clara, CA
Modesto, CA
Modesto, CA
Modesto, CA
Modesto, CA
Visalia-Porterville, CA
Chicago-Nape rville-Joliet,
IL-IN-WI
Chicago-Nape rville-Joliet,
IL-IN-WI
St. Louis, MO-IL
Suburban
Urban and Center City
Unknown
Suburban
Suburban
Urban and Center City
Suburban
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Suburban
Suburban
Urban and Center City
Avg
Years Corr Ratio
1995-2000 0.87 0.72
1995-1996 0.13 0.39
1995-1997 0.93 0.72
1995-2000 0.70 0.84
1995-2000 0.91 0.74
1995-2000 0.87 0.63
1995-1998 0.96 0.79
1992 0.81 0.94
1992 0.84 0.86
1992 0.40 0.96
Avg
Years Corr Ratio
1992-2000 0.43 0.44
1992-1996 0.31 0.33
1992-1997 0.86 0.46
1992-2000 0.59 0.56
1992-2000 0.76 0.48
1994-1997 0.11 0.36
1992-1993 0.42 0.37
1992-2000 0.54 0.39
1992-1998 0.24 0.61
Years
Avg
Corr Ratio
1995-1996
1997-1999
1995-1997
0.98 0.90
0.95 0.72
0.94 0.67
1995-2001
1996-2000
1995-2000
0.99 0.75
0.78 0.73
0.94 0.71
1995-2000
0.95 0.69
1995-1998
1997-1998
1998-2000
1998-2000
1995-2000
0.64 0.80
0.50 0.73
0.99 0.71
0.97 0.71
0.99 0.70
November 2012
3-204
Draft - Do Not Cite or Quote
-------
171191007
171630010
180890023
201730007
201730008
201730009
201731012
201770007
202090015
202090020
270530053
300490719
300490719
450430001
450790014
450791003
450791003
St. Louis, MO-IL
St. Louis, MO-IL
Chicago-Nape rville-Joliet,
IL-IN-WI
Wichita, KS
Wichita, KS
Wichita, KS
Wichita, KS
Topeka, KS
Kansas City, MO-KS
Kansas City, MO-KS
Minneapolis-St. Paul-
Bloomington, MN-WI
Helena, MT
Helena, MT
Georgetown, SC
Columbia, SC
Columbia, SC
Columbia, SC
Urban and Center City
Suburban
Urban and Center City
Suburban
Suburban
Suburban
Suburban
Urban and Center City
Urban and Center City
Urban and Center City
Urban and Center City
Suburban
Suburban
Urban and Center City
Suburban
Urban and Center City
Urban and Center City
Years Corr
1992 0.92
1992 0.96
2007 - 2008 0.73
1990-1997 0.18
1990-1997 0.34
1990-1997 0.54
1990-1997 0.85
1990-1997 0.56
1990-1997 0.75
1990-1997 0.99
1996-2001 0.54
1990-1991 0.81
1990 0.79
1990-1991 0.71
1990 0.82
1990 0.90
1990 0.90
Avg
Ratio
0.91
0.91
0.86
1.28
1.12
1.05
0.89
0.99
0.80
0.81
0.57
0.48
0.49
0.60
0.89
0.94
0.83
Avg
Years Corr Ratio
Avg
Years Corr Ratio
*Note: For comparability, comparisons are limited to monitors where all samples were above the MDL, at least 30 co-located samples were obtained, and both monitors reported data at standard
temperature and pressure.
November 2012
3-205
Draft - Do Not Cite or Quote
-------
Table 3-27 Metadata for studies of Pb-PM size distribution.
Reference
Location
Nearest source
Proximity to source
Sampling dates
Jersey City, NJ- an
urban/industrial area
New Brunswick, NJ- a suburban
area
Jersey City- near Manhattan, NJ Turnpike,
Hudson River- high gas/oil consumption for
industry/domestic heating, heavy gasoline &
diesel powered vehicles and ship traffic from
harbor.
New Brunswick- near NJ parkway and Garden
State Parkway
Close
ASD measurements:
June 10-20,2002
Bein et al. (2006)
Pittsburg, PA
Article does not describe sources; other articles
also use data from Pittsburgh Air Quality Study
(PAQS) and may have more info on sources
July 2001-September
2002
Pekey et al. (2010)
Kocaeli, Turkey
Kocaeli is a very industrialized and urbanized
region in Turkey; sources include a large
refinery, a petrochemical complex, a hazardous
waste incinerator and industry operations for
textile, machine, mine, metal, food, automotive,
paper, chemistry, wood, petroleum, tanning,
and coal sectors, plus heavy traffic
Singh etal. (2002)
Downey, CA- a city in Los Angeles
County along "Alameda corridor"
joining coastal area to downtown
LA
Riverside, CA- an inland county
east of LA
Downey- a "source" site affected by fresh PM
emissions from nearby oil refineries, industry,
and heavy diesel emissions
Riverside- a "receptor" site affected by aged
PM emissions including high vehicle emissions
in LA
Downey: 10 km downwind of
refineries; 2-4 km from Interstates
1-710 and I-605
Riverside- 70 km east of
downtown LA
Downey: September
2000-January 2001
Riverside: February
2001-June 2001
Dall'Osto et al. (2008)
U.K. national air quality monitoring
station in Port Talbot, U.K.
One of the U.K.'s largest integrated steelworks;
near major roadways; (Table 1 of this study
describes the plants, operations, emission
types, and emission components for steelwork
sources)
Monitoring site next to steelwork April 24-May 5, 2006
Weitkamp et al. (2005)
Coking facility near Pittsburgh, PA
Large coking facility that converts 6 million tons
of coal to 4 million tons of metallurgic coke
every year
Sampling site downwind, directly
across river from coke
facility(~400m)
August 22-September 5,
2002
Pekey et al. (2010)
Indoor/outdoor sample points for
15 homes in Kocaeli, Turkey
Kocaeli is a very industrialized and urbanized
region in Turkey; sources include a large
refinery, a petrochemical complex, a hazardous
waste incinerator and industry operations for
textile, machine, mine, metal, food, automotive,
paper, chemistry, wood, petroleum, tanning,
and coal sectors, plus heavy traffic
15 Kocaeli homes chosen as
representative sample; 10 close to
high traffic roads, 5 near
low/moderate traffic roads
Summer: May 31-June
29, 2006
Winter: December 16-
January 20, 2007
November 2012
3-206
Draft - Do Not Cite or Quote
-------
Reference
Location
Nearest source
Proximity to source
Sampling dates
Near Interstate 1-405 between
Sunset Blvd and Wilshire Blvd,
Los Angeles, CA
Heavy traffic on Interstate 1-405 Freeway
UP: 150m upwind of Interstate
I-405 (background)
DW1: 10m downwind
DW2: 150m downwind
DW3: 450m downwind
April 13-May 1, 2004
Song and Gao (2011)
Carlstadt, NJ
Heavy traffic on NJ Turnpike
~5m from roadside of the highway
Winter: December 2007-
February 2008
Summer: July 2008
Zereini et al. (2005)
3 Sites in Frankfurt, Germany
with different traffic densities
Vehicle emissions
Site 1: next to main street with
32500 cars/day
Site 2: next to side street with
<1000 cars/day
Site 3: large garden 8km NWof
city
August 2001 to July 2002
Lough et al. (2005)
Two road traffic tunnels in
Milwaukee, Wl
Traffic emissions
5 m upwind from entrance (inside
tunnel); 15 m upwind from tunnel
exit
Summer: July 31-August
28, 2000
Winter: December 13-
January 17, 2001
Haysetal. (2011)
20m downwind of Interstate I-440
highway in Raleigh, NC
Traffic emissions from highway
20 m downwind
July 26-31 and August
3-10,2006
Roadside site and site inside
highway tunnel in Taipei, Taiwan
Vehicle emissions
Roadside: sidewalk 4m from road
Tunnel: relay station in tunnel
1.4km from outlet; 2 m from traffic
lane
January to December
2008
Birmili et al. (2006)
Remote background: Mace Head
atmospheric research station in
Connemara, Ireland
Urban background: University of
Birmingham campus, U.K.
Roadside: A38 Bristol Road,
Birmingham, U.K.
Road Tunnel: Queensway
underpass in Birmingham, U.K.
Traffic emissions
Remote background:
Urban background: at least 100m
from road traffic
Roadside: 4 m from traffic
Road Tunnel: 30 m from tunnel
exit
Remote background:
August 8-28, 2002
Urban background:
April 23-October 7, 2002
Roadside: July 8-12,
2002
Road Tunnel: July 2,
2002
Bruggemann et al. (2009)
Harrison et al. (2003)
Roadside in
Dresden, Germany
Roadside in
Birmingham, U.K.
Traffic emissions from busy street (traffic
density-55,000 per day; 8% trucks), tramline,
railway station
Traffic emissions from A38
Next to road, near tramline
crossing, 200 m to railway station
9m from road
September 2003-August
2004
October 26, 2000 to
January 17, 2001
November 2012
3-207
Draft - Do Not Cite or Quote
-------
Reference
Location
Nearest source
Proximity to source
Sampling dates
Suburb of Kanazawa, Japan- a
western coastal city; the largest in
Hokuriku region of Japan
Emissions from road traffic, nearby incinerators
and electricity generation plants, and sea salt
Next to road; ~5km from
incinerators and electricity
generation plants; situated on
west coast
May-June 2003
Martuzevicius et al. (2004)
9 Locations in Cincinnati, Ohio
metropolitan area
Vehicle emissions from Cincinnati highway
network; emission from industry (233 facilities
within municipal area limits)
11 Sites- varying distance to linear
and point sources; distance to
major highways ranges from
21 Om to 4,590m
December 2001-
November2002
Moreno et al. (2008)
3 Sampling sites in Mexico City
(Mexico) Metropolitan Area:
1 Site in the industrial center (TO),
1 Site NE outside city limits (T1),
and
1 Rural site north of city (T2)
Urban pollution sources- traffic emissions,
industry
3 Sites with varying relation to
"Mexico urban plume" by
distance, wind direction
March 2006
Goforth and Christoforou
(2006)
Lake Hartwell, GA
(rural southeast U.S.)
February-March 2003
Makkonen et al. (2010)
Virolahti EMEP station, Finland
European Route E18 (3,000 vehicles/day)
5 km
August 2007
Wojas and Almquist (2007)
Oxford, OH and other towns in
Greater Cincinnati region
Transportation, manufacturing processes, and
coal-fired power plants
-80 km to northwest Cincinnati
January to December
2005
Lin et al. (2005)
Roadside in city in southern
Taiwan
Traffic emissions (avg traffic load = 72,000
vehicles/day), Pingtung Industrial Park
(146 factories), Nearby crematory
10 m from road, 2 km from
industrial park (146 factories,
i.e., electron apparatus, metal,
and food manufacturing), 1 km
from crematory
February to April 2004
Csavina et al. (2011)
Main sampling site in Winkelman,
Arizona at Hayden High School
Nearby an active mining and smelting site in
Hayden, Arizona
2km from mine tailings pile; 1km
from smelting operations, main
smoke stack, and slag pile
December 2008-
November2009
November 2012
3-208
Draft - Do Not Cite or Quote
-------
Reference Location Nearest source Proximity to source Sampling dates
Fang and Huang (2011) 3 Sites in central Taiwan: Emissions from vehicle traffic and industry Hung-kuang (HK): in residential November 2010-
A school area 2 km from maJ°r expressway December 2010
(Hung-kuang), Gaomei in Taichung (GM): 300
A wetland hectare wetland with coal
(Gaomei in Taichung), combustion-based Taichung
Thermal Power Plant (located
An industrial site a, the coast of the west side of
(Quan-xing) the sampNng site)
Quan-xing (QX): town with lots of
industry including metal
manufacturing, textiles, petroleum
and coal products
7 Sites around Los Angeles, CA, Not stated - August, 2002 - June,
including 6 urban watershed sites 2003
and one non-urban coastal
watershed.
November 2012 3-209 Draft - Do Not Cite or Quote
-------
Table 3-28 Size distribution data for various studies described in Table 3-27.
Reference Location Size Bin
Yi et al. (2QQ6) Jersey City, NJ 0.18-0.32
0.32-0.56
0.56-1
1-1.8
1 .8-3.2
3.2-5.6
5.6-10
10-14.4
14.4-19.9
19.9-26.1
26.1-36.1
36.1-100
New Brunswick, NJ 0.18-0.32
0.32-0.56
0.56-1
1-1.8
1 .8-3.2
3.2-5.6
5.6-10
10-14.4
14.4-19.9
19.9-26.1
26.1-36.1
36.1-100
Bein et al. (2006) Pittsburgh, PA 1 -1 .8
1 .8-3.2
3.2-5.6
5.6-10
Concentration
0.001054
0.000668
0.000952
0.000852
0.000609
0.001229
0.001591
0.000948
0.000171
0.000693
0.000333
0.000447
0.001146
0.00078
0.001733
0.001083
0.000373
0.000446
0.000347
0.000182
2.02E-09
1 .35E-05
1 .62E-05
0.000152
0.096608
0.314846
0.187393
0.239094
November 2012
3-210
Draft - Do Not Cite or Quote
-------
Reference Location Size Bin
Singh et al. (2002) Downey, CA <0.1 |jm
0.1-0.35
0.35-1.0
1 .0-2.5
2.5-10
Riverside, CA <0.1 |jm
0.1-0.35
0.35-1.0
1 .0-2.5
2.5-10
Dall'Osto et al. (2008) Port Talbot, U.K. 0.1-0.196
0.196-0.356
0.356-0.57
0.57-1
1-1.8
1.8-3.1
3.1-6.2
6.2-9.9
9.9-18
Sabin et al. (2006b) Los Angeles, CA, 10 m downwind of road <6
6-11
11-20
20-29
>29
Concentration
0.00133
0.00419
0.00334
0.00189
0.00175
0.0004
0.00089
0.0018
0.001
0.00297
0.00021 1
0.001871
0.005424
0.004935
0.010229
0.002216
0.001847
0.000807
0.000141
0.007953
0.004172
0.00013
0.000522
0.004563
November 2012
3-211
Draft - Do Not Cite or Quote
-------
Reference Location Size Bin
Zereini et al. (2005) Frankfurt, Germany <0.43
Main street 0.43-0.63
0.63-1.1
1.1-2.1
2.1-3.3
3.3-4.7
4.7-5.8
5.8-9.0
>9.0
Side street <0.43
0.43-0.63
0.63-1.1
1.1-2.1
2.1-3.3
3.3-4.7
4.7-5.8
5.8-9.0
>9.0
Rural background <0.43
0.43-0.63
0.63-1.1
1.1-2.1
2.1-3.3
3.3-4.7
4.7-5.8
5.8-9.0
>9.0
Concentration
0.005904
0.005332
0.004285
0.002857
0.002666
0.002857
0.001809
0.002476
0.004285
0.00332
0.002818
0.002239
0.001544
0.000849
0.000772
0.000386
0.00054
0.000733
0.003312
0.002442
0.002208
0.001405
0.000602
0.000502
0.000201
0.000201
0.000502
November 2012
3-212
Draft - Do Not Cite or Quote
-------
Reference Location Size Bin
Hays etal. (2011) Raleigh, NC 0.03-0.06
0.06-0.108
0.108-0.17
0.17-0.26
0.26-0.4
0.4-0.65
0.65-1
1-1.6
1 .6-2.5
2.5-4.4
4.4-6.8
6.8-10
10-18
Chen et al. (201 Ob) Taipei, Taiwan tunnel <0.1
0.1-2.5
2.5-10
Concentration
0.000186
0.000395
0.000732
0.001486
0.003593
0.007315
0.00423
0.002719
0.002701
0.003346
0.00123
0.000934
0.001265
0.018409
0.019773
0.030682
<0.1 0.00125
0.1-2.5
2.5-10
Bruggemann et al. (2009) Dresden, Germany 0.05-0.14
Summer 0.14-0.42
0.42-1.2
1 .2-3.5
3.5-10
Winter 0.05-0.14
0.14-0.42
0.42-1.2
1 .2-3.5
3.5-10
0.020625
0.024375
0.001078
0.002874
0.004671
0.001617
0.000539
0.002335
0.00521
0.013293
0.003054
0.000539
November 2012
3-213
Draft - Do Not Cite or Quote
-------
Reference Location Size Bin
Harrison et al. (2003) Birmingham, U.K. <0.2
0.2-1
1-2
2-10
Concentration
0.00685
0.014923
0.002446
0.00318
>10 0.000489
Wang et al. (2006d) Kanazawa, Japan 0.1-0.43
0.43-0.65
0.65-1.1
1.1-2.1
2.1-3.3
3.3-4.7
4.7-7
7-11
11-18
Martuzevicius et al. (2004) Cincinnati, OH ~0.1-0.18
Cycle VIII 0.18-0.32
0.32-0.56
0.56-1
1-1.8
1 .8-3.2
3.2-5.6
5.6-10
Cycle IX -0.1 -0.1 8
0.18-0.32
0.32-0.56
0.56-1
1-1.8
1 .8-3.2
3.2-5.6
5.6-10
0.000792
0.000748
0.00118
0.00103
0.000393
0.000678
0.000375
0.000229
0.000125
0.000758
0.002045
0.003258
0.00447
0.005758
0.00697
0.008333
0.009545
0.000455
0.001591
0.002879
0.004091
0.005379
0.006667
0.007879
0.009091
November 2012
3-214
Draft - Do Not Cite or Quote
-------
Reference
Location
Size Bin
Concentration
Lim et al. (2006)
'Summer and Fall data not provided
because there is more uncertainty in
the sites for those datasets.
Los Angeles, CA
Los Angeles River Watershed #1
Winter
Spring
Los Angeles River Watershed #2
Winter
Spring
Los Angeles River Watershed #3
Winter
Spring
Santa Ana River Watershed
Winter
6-11
11-20
20-29
>29
6-11
11-20
20-29
>29
6-11
11-20
20-29
>29
6-11
11-20
20-29
>29
6-11
11-20
20-29
>29
6-11
11-20
20-29
>29
6-11
11-20
20-29
>29
1.315
0.743
0.821
2.302
1.212
1.485
3.025
1.251
0.547
1.212
1.042
1.212
0.312
0.782
1.116
1.055
0.547
1.016
2.299
1.055
0.508
1.524
0.097
0.235
0.195
November 2012
3-215
Draft - Do Not Cite or Quote
-------
Reference Location Size Bin
Spring 6-11
11-20
20-29
>29
Ballona Creek Watershed 6-1 1
Winter 11-20
20-29
>29
Spring 6-11
11-20
20-29
>29
Dominguez Creek Watershed 6-1 1
Winter 11-20
20-29
>29
Spring 6-11
11-20
20-29
>29
Malibu Creek (non-urban) 6-1 1
Winter 11-20
Concentration
0.185
0.235
0.039
0.156
1.263
1.016
0.313
0.664
5.064
1.29
0.312
2.58
2.315
1.368
0.547
0.625
0.683
0.469
0.078
0.508
0.201
0.391
20-29
>29
Spring 6-11
11-20
0.039
0.211
0.156
20-29
>29
0.117
November 2012
3-216
Draft - Do Not Cite or Quote
-------
3.8.5
Pb Concentration in a Multipollutant Context
CO
PM10
N02
PM2.5
SO2
03
0
O 00
0
O 0® C
OCO ODGDD
> OOOarOTDOOO 0 0
ooo
fjTlTi OCX IHQQDKS
O OCOBOOQD
CO 0 OOO «
3SDDO O OO O
-1
-0.5
0.5
Note: Correlations were calculated from available data when data were above MDL and there were at least 30 data pairs available
for comparison.
Correlations for individual sites are shown with black open circles, while median correlations are illustrated with a red square.
Figure 3-62 Spearman correlations of monitored non-source Pb-TSP
concentration with daily averages of copollutant concentrations,
2008-2010.
November 2012
3-217
Draft - Do Not Cite or Quote
-------
Source, SO2
Non-Source, SO2
Source, PM2s
Non-Source, PM25
Source, PMio
Non-Source, PMio
Source, O3
Non-Source, O3
Source, N02
Non-Source, NO2
Source, CO
Non-Source, CO
Source, SO2
Non-Source, SO2
Source, PM2 5
Non-Source, PM25
Source, PMio
Non-Source, PM10
Source, O3
Non-Source, O3
Source, NO2
Non-Source, NO2
Source, CO
Non-Source, CO
US Summer
o oo
OOO GOKHDGD QDO O O O
O O
OOO OCD O O OBDODDODO
O O O O
O OOGD CHD GOOD OOO QtJOO O dD CH8DCXBD 00 O (
OO
O O OODOOOOIBEMDCDO C5DCHWZID <3D
O O
O O O OOCDDSD O O dHOSDCJ) OO (3D
1.0
-0.5 0.0 0.5
Spearman Correlation Coefficient
1.0
Note: Top panel: Summer; Bottom panel: Fall.
Figure 3-63
Seasonal correlations of monitored Pb-TSP concentration with
copollutant concentrations, 2007-2008.
November 2012
3-218
Draft - Do Not Cite or Quote
-------
Non-Source, SO2 ' "
Source, PM2 5
Non-Source, PM2s
Non-Source, PM,0
Non-Source, O3
Non-Source, N02 • -
Non-Source, CO . -
US Winter
o
0 0
000 0
0
o o o o o
O 000 O (
0
0 0
0 0 00 0
-1.0
-0.5 0.0 0.5
Spearman Correlation Coefficient
1.0
Non-Source, S02 -
Source, PM2.s -
Non-Source, PM25 -
Non-Source, PMio
Non-Source, 03 .
Non-Source, NO2 .
Non-Source, CO .
US Spring
o o
QD O
-1.0
Note: Top panel: Winter; Bottom panel: Spring.
oo o
o oo
QD
-0.5 0.0 0.5
Spearman Correlation Coefficient
1.0
Figure 3-64 Seasonal correlations of monitored Pb-TSP concentration with
copollutant concentrations, 2009.
November 2012
3-219
Draft - Do Not Cite or Quote
-------
Non-Source, SO2
Source. PM25 "
Non-Source, PM2 5 -
Non-Source, PM,0 -
Non-Source, O3 -
Non-Source, NO2
Non-Source, CO -
US Summer
ooo o
o
O 00 O
o o o
O ODD O
O O
OO O O
-1.0
-0.5 0.0 0.5
Spearman Correlation Coefficient
1.0
Non-Source, SO2
Source, PM2s
Non-Source, PM2 5
Non-Source, PM,0
Non-Source, Os
Non-Source, NO2
Non-Source. CO
US Fall
O 00
00 O O
000
O 00 O O
o o
-1.0
Note: Top panel: Summer; Bottom panel: Fall.
-0.5 0.0 0.5
Spearman Correlation Coefficient
1.0
Figure 3-65 Seasonal correlations of monitored Pb-TSP concentration with
copollutant concentrations, 2009.
November 2012
3-220
Draft - Do Not Cite or Quote
-------
Top panel: Winter; Bottom panel: Spring.
Note: "nvol" = non-volatile, "vol" = volatile, and organic carbon (OC) samples were blank-adjusted.
Figure 3-66 Seasonal correlations of monitored Pb-PM2.s concentration with
copollutant concentrations, 2007-2009.
November 2012
3-221
Draft - Do Not Cite or Quote
-------
Zn —
Br -
Cu -
K -
s -
$04-2 —
Ca -
luh -
Cmstal —
Fe —
Se -
N03- -
11 -
EC -
^ -
Si —
Mg -
K+ —
V —
NH4t -
Ha -
Hg
Cd —
Na+2 -
vol N03- -
Ni —
fs —
nvol H03- —
0 | 1 |
o TO o i H |
0, 1 ,
0 | 1 |
0 0 1 1 I
0 00 1 < I
0 0 1 1 1
o o cm 1 | |-
0 | ( , |_
0 0 1 H 1 H-
0 1 1 1 1
0 01 1 • h
0 0 1 1 | |
1 1 1 1
1 _J 1 J
OO O 1 1 1 1 1
0 | 1 , |
1 1 1 1
(. 1
-] 1
( ,
1
]::::::::::
1 o
1 0
1
1 0
10 0
100 00
o o
-HO 0
Zn —
K —
Cu -
Br -
oc -
Fe -
vol H03- -
Cmstal —
Ca -
K+ -
NH4+ -
Si -
Cr -
V -
Mg
Na+2 -
Ni -
Hg -
Cl —
Cd -
cool 1 | h 1
0 , H , | ,
O O OTC O 1 ^ 1 1 1
i-CD
o 1 0 0
o mi- n__J__^ — r H
1 • — i H
000 1 1 1 1 1 OJ 0
1 | | | 1010
Top panel: Summer; Bottom panel: Fall.
Note: "nvol" = non-volatile, "vol" = volatile, and organic carbon (OC) samples were blank-adjusted.
Figure 3-67 Seasonal correlations of monitored Pb-PM2.s concentration with
copollutant concentrations, 2007-2009.
November 2012
3-222
Draft - Do Not Cite or Quote
-------
Table 3-29 Copollutant exposures for various trace metal studies.
Adgate et al. (2007)
Location
PM2.5
Pb
S
Ca
Al
Na
Fe
Mg
K
Ti
Zn
Cu
Ni
Mn
Sb
Cd
V
La
Cs
Th
Sc
Ag
Co
Cr
Si
Cl
Se
Rb
Sr
As
l-R(med)a'b
Personal
(median)0
Minnesota, U.S.
1.5
272.1
85.0
23.3
20.6
43.1
16.3
38.4
0.8
6.5
1.-0.15
2.4
0.21
0.12
0.12
0.05
0.00
0.00
0.00
0.00
0.07
0.02
1.2
3.2
351.6
174.1
58.6
31.9
78.6
27.5
47.5
1.4
9.6
4.9
1.8
2.3
0.30
0.14
0.16
0.11
0.00
0.00
0.01
0.08
0.07
2.6
Riedikeretal. (2003)
Vehicle
(range)0
Roadside
(range)0
New Jersey, U.S.
24,000
2-3
905-1 ,592
31-44
307-332
6-75
9-10
5-10
18-32
0
3-4
4-6
1
2
198-464
7-32
1
1
5-28
1
31 ,579
4-6
1,416-2,231
18-40
82-163
23-57
6-10
14-17
8-16
0
3
4-7
1
1
338-672
3-9
1-2
1
1
1
Pekey et al.
(2010) Molnar et al. (2007)
I -near industry I-R l-School l-Pre-School
(range)3 (median)a'b (median)3 (median)3
Kocaeli, Turkey Stockholm, Sweden
24,400-29,800
34-85 2.8 2.5 1.7
435-489 330 290 220
309-452 70 110 58
53-60
44-58 57 100 71
160-215 120 96 67
29-39 8.0 13 8.7
51-88 14 17 11
21-58 9.3 1.7 2.1
2-3 0.99 1.0 0.72
28-32 2.2 2.5 2.1
3-5 2.5 2.7 1.8
3-8 <1.1 1.3 1.1
387-401
1-2
Mo
Br
al: Indoor; Units
bR: Residential;
°Units: ng/m3
: ng/m3
Units: ng/m3
2.1 1.3 1.3
November 2012
3-223
Draft - Do Not Cite or Quote
-------
References for Chapter 3
Abel. MT; Cobb. GP; Presley. SM; Ray. GL; Rainwater. TR; Austin. GP; Cox. SB; Anderson. TA;
Leftwich. BD; Kendall. RJ: Suedel. BC. (2010). Lead distributions and risks in New Orleans
following hurricanes Katrina and Rita. Environ Toxicol Chem 29: 1429-1437.
http://dx.doi.org/10.1002/Etc.205
Abel. MT; Presley. SM; Rainwater. TR; Austin. GP: Cox. SB; Mcdaniel. LN; Marsland. EJ; Leftwich.
BD; Anderson. TA; Kendall. RJ: Cobb. GP. (2007). Spatial and temporal evaluation of metal
concentrations in soils and sediments from New Orleans, Louisiana, USA, following hurricanes
Katrina and Rita. Environ Toxicol Chem 26: 2108-2114. http://dx.doi.Org/10.1897/06-595R.l
Adgate. JL; Mongin. SJ: Pratt. GC: Zhang. J; Field. MP; Ramachandran. G: Sexton. K. (2007).
Relationships between personal, indoor, and outdoor exposures to trace elements in PM2.5. Sci Total
Environ 386: 21-32. http://dx.doi.Org/10.1016/i.scitotenv.2007.07.007
Aelion. CM; Davis. HT; Mcdermott. S: Lawson. AB. (2009). Soil metal concentrations and toxicity:
Associations with distances to industrial facilities and implications for human health. Sci Total
Environ 407: 2216-2223. http://dx.doi.Org/10.1016/i.scitotenv.2008.ll.033
Al-Malack. MH. (2001). Migration of lead from unplasticized polyvinyl chloride pipes. J Hazard Mater
82: 263-274. http://dx.doi.org/10.1016/S0304-3894(00)00366-6
Al-Masri. MS: Al-Kharfan. K; Al-Shamali. K. (2006). Speciation of Pb, Cu and Zn determined by
sequential extraction for identification of air pollution sources in Syria. Atmos Environ 40: 753-761.
http://dx.doi.0rg/10.1016/i.atmosenv.2005.10.008
Allott. RW: Hewitt. CN; Kelly. MR. (1989). The environmental half-lives and mean residence times of
contaminants in dust for an urban environment: Barrow-in-Furness. Sci Total Environ 93: 403-410.
http://dx.doi.org/10.1016/0048-9697(90)90131-0
Alpert. DJ: Hopke. PK. (1981). A determination of the sources of airborne particles collected during the
regional air pollution study. Atmos Environ 15: 675-687.
http://dx.doi.org/10.1016/0004-6981(81)90272-9
ALRA. Airborne Lead Reduction Act of 1984. S. 2609. (1984).
http://heinonline.org/HOL/Page?collection=congrec&handle=hein.cbhear/cbhearings2029&tvpe=Ima
ge&id=l
Amato. F; Pandolfi. M; Viana. M; Querol. X; Alastuev. A; Moreno. T. (2009). Spatial and chemical
patterns of PM10 in road dust deposited in urban environment. Atmos Environ 43: 1650-1659.
http://dx.doi.0rg/10.1016/i.atmosenv.2008.12.009
Annibaldi. A; Truzzi. C: Illuminati. S: Bassotti. E; Scarponi. G. (2007). Determination of water-soluble
and insoluble (dilute-HCl-extractable) fractions of Cd, Pb and Cu in Antarctic aerosol by square wave
anodic stripping voltammetry: Distribution and summer seasonal evolution at Terra Nova Bay
(Victoria Land). Anal Bioanal Chem 387: 977-998. http://dx.doi.org/10.1007/s00216-006-0994-0
Anttila. P; Makkonen. U; Hellen. H; Kyllonen. K; Leppanen. S: Saari. H; Hakola. H. (2008). Impact of
the open biomass fires in spring and summer of 2006 on the chemical composition of background air
in south-eastern Finland. Atmos Environ 42: 6472-6486.
http://dx.doi.0rg/10.1016/i.atmosenv.2008.04.020
Ariola. V: D'Alessandro. A; Lucarelli. F; Marcazzan. G: Mazzei. F; Nava. S: Garcia-Orellana. I; Prati. P;
Valli. G: Vecchi. R; Zucchiatti. A. (2006). Elemental characterization of PM10, PM2.5 and PM1 in the
town of Genoa (Italy). Chemosphere 62: 226-232.
http://dx.doi.0rg/10.1016/i.chemosphere.2005.05.004
November 2012 3-224 Draft - Do Not Cite or Quote
-------
ASTM (American Society for Testing and Materials). (2007). Standard specification for aviation
gasolines. (ASTM D910-06). West Conshohocken, PA: American Society for Testing and Material.
http://www.astm.org/Standards/D910.htm
Atkinson. CA; Jollev. DF; Simpson. SL. (2007). Effect of overlying water pH, dissolved oxygen, salinity
and sediment disturbances on metal release and sequestration from metal contaminated marine
sediments. Chemosphere 69: 1428-1437. http://dx.doi.Org/10.1016/i.chemosphere.2007.04.068
Aucott. M; Caldarelli. A. (2012). Quantity of Lead Released to the Environment in New Jersey in the
Form of Motor Vehicle Wheel Weights. Water Air Soil Pollut 223: 1743 -175 2.
httrj://dx.doi.org/10.1007/sll270-011-0979-2
Aznar. JC: Richer-Lafleche. M; Begin. C: Rodrigue. R. (2008a). Spatiotemporal reconstruction of lead
contamination using tree rings and organic soil layers. Sci Total Environ 407: 233-241.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.09.044
Aznar. JC: Richer-Lafleche. M; Cluis. D. (2008b). Metal contamination in the lichen Alectoria
sarmentosa near the copper smelter of Murdochville, Quebec. Environ Pollut 156: 1.
http://dx.doi.0rg/10.1016/i.envpol.2007.12.037
Bacardit. M; Camarero. L. (2010a). Atmospherically deposited major and trace elements in the winter
snowpack along a gradient of altitude in the Central Pyrenees: the seasonal record of long-range fluxes
over SWEurope. Atmos Environ 44: 582-595. http://dx.doi.Org/10.1016/i.atmosenv.2009.06.022
Bacardit. M; Camarero. L. (201 Ob). Modelling Pb, Zn and As transfer from terrestrial to aquatic
ecosystems during the ice-free season in three Pyrenean catchments. Sci Total Environ 408:
5854-5861. http://dx.doi.Org/10.1016/i.scitotenv.2010.07.088
Bacon. SN; Mcdonald. EV; Amit. R; Enzel. Y; Crouvi. O. (2011). Total suspended paniculate matter
emissions at high friction velocities from desert landforms. J Geophys Res 116: F03019.
http://dx.doi.org/10.1029/2011JF001965
Balogh. SJ: Triplett. LD; Engstrom. PR; Nollet. YH. (2010). Historical trace metal loading to a large river
recorded in the sediments of Lake St. Croix, USA. J Paleolimnol 44: 517-530.
http://dx.doi.org/10.1007/sl0933-010-9434-4
Barrett. J: Taylor. K; Hudson-Edwards. K; Charnock. J. (2010). Solid-phase speciation of Pb in urban
road dust sediment: A XANES and EXAFS study. Environ Sci Technol 44: 2940-2946.
http://dx.doi.org/10.1021/es903737k
Batonneau. Y; Bremard. C: Gengembre. L; Laureyns. J: Le Maguer. A; Le Maguer. D; Perdrix. E;
Sobanska. S. (2004). Speciation of PM10 sources of airborne nonferrous metals within the 3-km zone
of lead/zinc smelters. Environ Sci Technol 38: 5281-5289. http://dx.doi.org/10.1021/es0497173
Bein. KJ; Zhao. Y; Peknev. NJ; Davidson. CI; Johnston. MV; Wexler. AS. (2006). Identification of
sources of atmospheric PM at the Pittsburgh supersite, part II: Quantitative comparisons of single
particle, particle number, and particle mass measurements. Atmos Environ 40: S424-S444.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.01.039
Bein. KJ: Zhao. YJ; Johnston. MV: Wexler. AS. (2007). Identification of sources of atmospheric PM at
the Pittsburgh supersite, part III: Source characterization. Atmos Environ 41: 3974-3992.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.01.039
Bench. G: Grant. PG: Ueda. D: Cliff. SS: Perry. KD; Cahill. TA. (2002). The use of STIM and PESA to
measure profiles of aerosol mass and hydrogen content, respectively, across Mylar rotating drums
impactor samples. Aerosol Sci Technol 36: 642-651. http://dx.doi.org/10.1080/02786820252883874
Bennett. JD. (2007). Memorandum regarding Doe Run-Herculaneum State Implementation Plan (SIP)
dispersion modeling review. Jefferson City, MO: Missouri Department of Natural Resources.
http://www.dnr.mo.gov/env/apcp/docs/drhsipmodmemol.pdf
November 2012 3-225 Draft - Do Not Cite or Quote
-------
Berg. T; Aas. W: Pacytia. J: Uggerud HT; Vadset M. (2008). Atmospheric trace metal concentrations at
Norwegian background sites during 25 years and its relation to European emissions. Atmos Environ
42: 7494-7501. http://dx.doi.Org/10.1016/i.atmosenv.2008.05.020
Berthelot. Y; Valton. E; Auroy. A; Trottier. B; Robidoux. PY. (2008). Integration of lexicological and
chemical tools to assess the bioavailability of metals and energetic compounds in contaminated soils.
Chemosphere 74: 166-177. http://dx.doi.Org/10.1016/i.chemosphere.2008.07.056
Besser. JM; Brumbaugh. WG: May. TW: Church. SE; Kimball. BA. (2001). Bioavailability of metals in
stream food webs and hazards to brook trout (Salvelinus fontinalis) in the upper Animas River
watershed, Colorado. Arch Environ Contam Toxicol 40: 48-59.
http://dx.doi.org/10.1007/s002440010147
Besser. JM; Brumbaugh. WG: May. TW: Schmitt CJ. (2007). Biomonitoring of lead, zinc, and cadmium
in streams draining lead-mining and non-mining areas, Southeast Missouri, USA. Environ Monit
Assess 129: 227-241. http://dx.doi.org/10.1007/sl0661-006-9356-9
Biasioli. M; Barberis. R; Ajmone-Marsan. F. (2006). The influence of a large city on some soil properties
and metals content. Sci Total Environ 356: 154-164. http://dx.doi.Org/10.1016/i.scitotenv.2005.04.033
Biggins. PDE: Harrison. RM. (1979). Atmospheric chemistry of automotive lead. Environ Sci Technol 13:
558-565. http://dx.doi.org/10.1021/es60153a017
Biggins. PDE: Harrison. RM. (1980). Chemical speciation of lead compounds in street dusts. Environ Sci
Technol 14: 336-339. http://dx.doi.org/10.1021/es60163a005
Bindler. R; Renberg. I; Klaminder. J. (2008). Bridging the gap between ancient metal pollution and
contemporary biogeochemistry. J Paleolimnol 40: 755-770.
http://dx.doi.org/10.1007/sl0933-008-9208-4
Birch. G: McCreadv. S. (2009). Catchment condition as a major control on the quality of receiving basin
sediments (Sydney Harbour, Australia). Sci Total Environ 407: 2820-2835.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.12.051
Birch. G: O'Hea. L. (2007). The chemistry of suspended paniculate material in a highly contaminated
embayment of Port Jackson (Australia) under quiescent, high-wind and heavy-rainfall conditions.
Environ Geol 53: 501-516. http://dx.doi.org/10.1007/s00254-007-0662-5
Birmili. W: Allen. AG: Barv. F; Harrison. RM. (2006). Trace metal concentrations and water solubility in
size-fractionated atmospheric particles and influence of road traffic. Environ Sci Technol 40:
1144-1153. http://dx.doi.org/10.1021/es0486925
Bonnard. R: Mckone. TE. (2009). Integration of the predictions of two models with dose measurements in
a case study of children exposed to the emissions from a lead smelter. Hum Ecol Risk Assess 15:
1203-1226. http://dx.doi.org/10.1080/10807030903304849
Boonfueng. T; Axe. L; Xu. Y; Tyson. TA. (2006). Nickel and lead sequestration in manganese oxide-
coated montmorillonite. J Colloid Interface Sci 303: 87-98.
http://dx.doi.0rg/10.1016/i.jcis.2006.07.056
Boutron. CF; Goerlach. U; Candelone. JP; Bolshov. MA; Delmas. RJ. (1991). Decrease in anthropogenic
lead, cadmium and zinc in Greenland snows since the late 1960s. Nature 353: 153-156.
http://dx.doi.org/10.1038/353153aO
Bouvet. M; Francois. D; Schwartz. C. (2007). Road soil retention Pb leached from MSWI bottom ash.
Waste Manag 27: 840-849. http://dx.doi.Org/10.1016/j.wasman.2006.06.003
Britter. RE; Hanna. SR. (2003). Flow and dispersion in urban areas. Annu Rev Fluid Mech 35: 469-496.
http://dx.doi.org/10.1146/annurev.fluid.35.101101.161147
Brown. RW: Gonzales. C: Hooper. MJ; Bavat. AC: Fornerette. AM; McBride. TJ; Longoria. T; Mielke.
HW. (2008). Soil lead (Pb) in residential transects through Lubbock, Texas: A preliminary assessment.
Environ Geochem Health 30: 541-547. http://dx.doi.org/10.1007/sl0653-008-9180-y
November 2012 3-226 Draft - Do Not Cite or Quote
-------
Bruggemann. E; Gerwig. H; Gnauk. T; Muller. K; Herrmann. H. (2009). Influence of seasons, air mass
origin and day of the week on size-segregated chemical composition of aerosol particles at a kerbside.
Atmos Environ 43: 2456-2463. http://dx.doi.Org/10.1016/i.atmosenv.2009.01.054
Brumbaugh. WG: May. TW: Besser. JM; Allert. AL; Schmitt. CJ. (2007). Assessment of elemental
concentrations in streams of the new lead belt in southeastern Missouri, 2002-05.
(USGSSIR20075057). Reston, VA: U.S. Geological Survey, http://pubs.usgs.gov/sir/2007/5057
Bukowiecki. N; Lienemann. P; Hill. M; Furger. M; Richard. A; Amato. F; Prevot. ASH; Baltensperger. U;
Buchmann. B; Gehrig. R. (2010). PM10 emission factors for non-exhaust particles generated by road
traffic in an urban street canyon and along a freeway in Switzerland. Atmos Environ 44: 2330-2340.
http://dx.doi.0rg/10.1016/i.atmosenv.2010.03.039
Buonanno. G: Fuoco. FC: Stabile. L. (2011). Influential parameters on particle exposure of pedestrians in
urban microenvironments. Atmos Environ 45: 1434-1443.
http://dx.doi.0rg/10.1016/i.atmosenv.2010.12.015
Bur. T; Probst. JL; N'Guessan. M; Probst. A. (2009). Distribution and origin of lead in stream sediments
from small agricultural catchments draining Miocene molassic deposits (SW France). Appl Geochem
24: 1324-1338. http://dx.doi.Org/10.1016/i.apgeochem.2009.04.004
Cahill. CF. (2003). Asian aerosol transport to Alaska during ACE-Asia. J Geophys Res 108: 8664.
http://dx.doi.org/10.1029/2002JD003271
Cahill. TA. (1981). Innovative aerosol sampling devices based upon PIXE capabilities. Nuclear
Instruments and Methods 181: 473-480. http://dx.doi.org/10.1016/0029-554X(81)90652-2
Cahill. TA; Feenev. PJ: Eldred. RA. (1987). Size-time composition profile of aerosols using the drum
sampler. Nucl lustrum Methods Phys Res B 22: 344-348.
http://dx.doi.org/10.1016/0168-583X(87)90355-7
Callender. E; Rice. KG. (2000). The urban environmental gradient: Anthropogenic influences on the
spatial and temporal distributions of lead and zinc in sediments. Environ Sci Technol 34: 232-238.
http://dx.doi.org/10.1021/es990380s
Canepari. S: Astolfi. ML; Moretti. S: Curini. R. (2010). Comparison of extracting solutions for elemental
fractionation in airborne particulate matter. Talanta 82: 834-844.
http://dx.doi.0rg/10.1016/i.talanta.2010.05.068
Canepari. S: Cardarelli. E; Perrino. C: Catrambone. M; Pietrodangelo. A; Strincone. M. (2006). Two-
stage chemical fractionation method for the analysis of elements and non-volatile inorganic ions in
PM10 samples: Application to ambient samples collected in Rome (Italy). Atmos Environ 40:
7908-7923. http://dx.doi.Org/10.1016/i.atmosenv.2006.07.005
Canepari. S: Perrino. C: Olivieri. F; Astolfi. ML. (2008). Characterisation of the traffic sources of PM
through size-segregated sampling, sequential leaching and ICP analysis. Atmos Environ 42:
8161-8175. http://dx.doi.0rg/10.1016/i.atmosenv.2008.07.052
Canovas. CR; Olias. M; Nieto. JM; Galvan. L. (2010). Wash-out processes of evaporitic sulfate salts in
the Tinto river: Hydrogeochemical evolution and environmental impact. Appl Geochem 25: 288-301.
http://dx.doi.0rg/10.1016/i.apgeochem.2009.ll.014
Cantwell. MG: Burgess. RM; King. JW. (2008). Resuspension of contaminated field and formulated
reference sediments Part I: Evaluation of metal release under controlled laboratory conditions.
Chemosphere73: 1824-1831. http://dx.doi.Org/10.1016/i.chemosphere.2008.08.007
Caravanos. J: Weiss. AL; Blaise. MJ; Jaeger. RJ. (2006a). A survey of spatially distributed exterior dust
lead loadings in New York City. Environ Res 100: 165-172.
http://dx.doi.0rg/10.1016/i.envres.2005.05.001
November 2012 3-227 Draft - Do Not Cite or Quote
-------
Carr. E; Lee. M; Marin. K; Holder. C: Hover. M; Pedde. M; Cook. R; Touma. J. (2011). Development and
evaluation of an air quality modeling approach to assess near-field impacts of lead emissions from
piston-engine aircraft operating on leaded aviation gasoline. Atmos Environ 45: 5795-5804.
http://dx.doi.0rg/10.1016/i.atmosenv.2011.07.017
Chalmers. AT; Van Metre. PC: Callender. E. (2007). The chemical response of particle-associated
contaminants in aquatic sediments to urbanization in New England, USA. J Contam Hydrol 91: 4-25.
http://dx.doi.0rg/10.1016/i.jconhvd.2006.08.007
Chan. YC: Cohen. DP; Hawas. O: Stelcer. E; Simpson. R; Denison. L; Wong. N; Hodge. M; Comino. E;
Carswell. S. (2008). Apportionment of sources of fine and coarse particles in four major Australian
cities by positive matrix factorisation. Atmos Environ 42: 374-389.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.09.030
Chen. FL; Vanderpool. R; Williams. R: Dimmick. F; Grover. BD; Long. R; Murdoch. R. (2011 a). Field
evaluation of portable and central site PM samplers emphasizing additive and differential mass
concentration estimates. Atmos Environ 45: 4522-4527.
http://dx.doi.0rg/10.1016/i.atmosenv.2011.02.006
Chen. S: Tsai. C: Huang. C: Chen. H; Chen. S: Lin. C: Tsai. J: Chou. C. (2010b). Chemical mass closure
and chemical characteristics of ambient ultrafine particles and other PM fractions. Aerosol Sci Technol
44: 713-723.
Cheng. H; Hu. Y. (2010). Lead (Pb) isotopic fingerprinting and its applications in lead pollution studies in
China: A review [Review]. Environ Pollut 158: 1134-1146.
http://dx.doi.0rg/10.1016/i.envpol.2009.12.028
Chester. R; Lin. FJ; Murphy. KJT. (1989). A three stage sequential leaching scheme for the
characterisation of the sources and environmental mobility of trace metals in the marine aerosol.
Environ Technol 10: 887 - 900. http://dx.doi.org/10.1080/09593338909384810
Cho. SH; Richmond-Bryant. J: Thornburg. J: Portzer. J: Vanderpool. R; Cavender. K; Rice. J. (2011). A
literature review of concentrations and size distributions of ambient airborne Pb-containing particulate
matter. Atmos Environ 45: 5005-5015. http://dx.doi.Org/10.1016/j.atmosenv.2011.05.009
Choel M; Deboudt. K; Flament. P; Lecornet. G: Perdrix. E; Sobanska. S. (2006). Fast evolution of
tropospheric Pb- and Zn-rich particles in the vicinity of a lead smelter. Atmos Environ 40: 4439-4449.
http://dx.doi.0rg/10.1016/i.atmosenv.2006.04.027
Chon. HS: Ohandja. DG: Voulvoulis. N. (2010). Implementation of E.U. water framework directive:
Source assessment of metallic substances at catchment levels. J Environ Monit 12: 36-47.
http://dx.doi.org/10.1039/b907851g
Chow. TJ; Johnstone. MS. (1965). Lead isotopes in gasoline and aerosols of Los Angeles Basin,
California. Science 147: 502-503. http://dx.doi.org/10.1126/science.147.3657.502
Cizmecioglu. SC: Muezzinoglu. A. (2008). Solubility of deposited airborne heavy metals. Atmos Res 89:
396-404. http://dx.doi.0rg/10.1016/i.atmosres.2008.03.012
Clark. GM. (2003). Occurrence and transport of cadmium, lead, and zinc in the spokane river basin, Idaho
and Washington, water years 1999-2001. (Water-Resources Investigations Report 024183). Boise, ID:
U.S. Geological Survey. http://id.water.usgs.gov/PDF/wri024183/clarkfinal2.pdf
Clark. HF; Brabander. DJ; Erdil. RM. (2006). Sources, sinks, and exposure pathways of lead in urban
garden soil. J Environ Qual 35: 2066-2074. http://dx.doi.org/10.2134/ieq2005.0464
Cloquet. C: Carignan. J: Libourel. G. (2006). Isotopic composition of Zn and Pb atmospheric depositions
in an urban/periurban area of northeastern France. Environ Sci Technol 40: 6594-6600.
http://dx.doi.org/10.1021/es0609654
CNL (Crocker Nuclear Laboratory). (2001). Version II IMPROVE sampler: Operating procedures manual
for use in the IMPROVE monitoring network. (TI 201 A). Davis, CA.
http://vista.cira.colostate.edu/improve/publications/SOPs/ucdavis sops/ti201a v2.pdf
November 2012 3-228 Draft - Do Not Cite or Quote
-------
Cobb. GP: Abel MT; Rainwater. TR; Austin. GP; Cox. SB; Kendall. RJ: Marsland. EJ; Anderson. TA;
Leftwich. BD; Zak. JC: Presley. SM. (2006). Metal distributions in New Orleans following hurricanes
Katrina and Rita: A continuation study. Environ Sci Technol 40: 4571-4577.
http://dx.doi.org/10.1021/es060041g
Cohen. DP; Crawford. J: Stelcer. E; Bac. VT. (2010). Characterisation and source apportionment of fine
paniculate sources at Hanoi from 2001 to 2008. Atmos Environ 44: 320-328.
http://dx.doi.0rg/10.1016/i.atmosenv.2009.10.037
Company. R; Serafim. A; Lopes. B; Cravo. A; Shepherd. TJ; Pearson. G: Bebianno. MJ. (2008). Using
biochemical and isotope geochemistry to understand the environmental and public health implications
of lead pollution in the lower Guadiana River, Iberia: A freshwater bivalve study. Sci Total Environ
405: 109-119. http://dx.doi.0rg/10.1016/j.scitotenv.2008.07.016
Cong. Z; Kang. S: Liu. X; Wang. G. (2007). Elemental composition of aerosol in the Nam Co region,
Tibetan Plateau, during summer monsoon season. Atmos Environ 41: 1180-1187.
http://dx.doi.0rg/10.1016/i.atmosenv.2006.09.046
Conor Pacific Environmental Technologies Inc. (2000). Airborne particulate matter, lead and manganese
at Buttonville airport. (041-6710). Toronto, Ontario: Environment Canada.
Courtin-Nomade. A; Soubrand-Colin. M; Marcus. MA; Fakra. SC. (2008). Evidence for the incorporation
of lead into barite from waste rock pile materials. Environ Sci Technol 42: 2867-2872.
http://dx.doi.org/10.1021/es702822k
Couture. RM; J-F. C: Auger. D; Claisse. D; Gobeil. C: Cossa. D. (2010). Seasonal and decadal variations
in lead sources to eastern North Atlantic mussels. Environ Sci Technol 44: 1211-1216.
http://dx.doi.org/10.1021/es902352z
Coynel. A; Schafer. J; Blanc. G: Bossy. C. (2007). Scenario of particulate trace metal and metalloid
transport during a major flood event inferred from transient geochemical signals. Appl Geochem 22:
821-836. http://dx.doi.0rg/10.1016/i.apgeochem.2006.10.004
Csavina. JA; Landazuri. A; Wonaschutz. A; Rine. K; Rheinheimer. P; Barbaris. B; Conant. W: Saez. AE;
Betterton. EA. (2011). Metal and metalloid contaminants in atmospheric aerosols from mining
operations. Water Air Soil Pollut 221: 145-157. http://dx.doi.org/10.1007/sll270-011-0777-x
Curtis. CJ: Evans. CD; Helliwell. RC: Monteith. DT. (2005). Nitrate leaching as a confounding factor in
chemical recovery from acidification in U.K. upland waters. Environ Pollut 137: 73-82.
http://dx.doi.0rg/10.1016/i.envpol.2004.12.032
Dahl. O; Poykio. R; Nurmesniemi. H. (2008). Concentrations of heavy metals in fly ash from a coal-fired
power plant with respect to the new Finnish limit values. Journal of Material Cycles and Waste
Management 10: 87-92. http://dx.doi.org/10.1007/sl0163-007-0189-6
DaH'Osto. M; Booth. MJ: Smith. W: Fisher. R; Harrison. RM. (2008). A study of the size distributions and
the chemical characterization of airborne particles in the vicinity of a large integrated steelworks.
Aerosol Sci Technol 42: 981-991. http://dx.doi.org/10.1080/02786820802339587
Das. SK; Routh. J: Roychoudhury. AN; Klump. JV. (2008). Major and trace element geochemistry in
Zeekoevlei, South Africa: A lacustrine record of present and past processes. Appl Geochem 23:
2496-2511. http://dx.doi.0rg/10.1016/i.apgeochem.2008.02.011
Dawson. JJC: Tetzlaff. D: Carey. AM; Raab. A; Soulsbv. C: Killham. K; Meharg. AA. (2010).
Characterizing Pb mobilization from upland soils to streams using (206)Pb/(207)Pb isotopic ratios.
Environ Sci Technol 44: 243-249. http://dx.doi.org/10.1021/es902664d
de la Cruz. MT; Laborda. F; Callen. MS: Lopez. JM; Mastral. AM. (2009). Study of Pb sources by Pb
isotope ratios in the airborne PM10 of Zaragoza, Spain. J Environ Monit 11: 2052-2057.
http://dx.doi.org/10.1039/b912274e
November 2012 3-229 Draft - Do Not Cite or Quote
-------
Deng. X; Jennings. AA. (2006). Evaluating an eletrokinetically driven extraction method for measuring
heavy metal soil contamination. J Environ Eng 132: 527-537.
http://dx.doi.org/10.1061/(ASCE)0733-9372(2006)132:4(527)
Dermont. G: Bergeron. M; Richer-Lafleche. M; Mercier. G. (2010). Remediation of metal-contaminated
urban soil using flotation technique. Sci Total Environ 408: 1199-1211.
http://dx.doi.0rg/10.1016/i.scitotenv.2009.ll.036
Desta. MB; Bruen. M; Higgins. N; Johnston. P. (2007). Highway runoff quality in Ireland. J Environ
Monit 9: 366-371. http://dx.doi.org/10.1039/b702327h
Diaz-Somoano. M; Kylander. ME; Lopez-Anton. MA; Suarez-Ruiz. I; Martinez-Tarazona. MR; Ferrat.
M; Kober. B; Weiss. DJ. (2009). Stable lead isotope compositions in selected coals from around the
world and implications for present day aerosol source tracing. Environ Sci Technol 43: 1078-1085.
http://dx.doi.org/10.1021/es801818r
Porn. CR: Pierce. JO. II; Philips. PE; Chase. GR. (1976). Airborne Pb, Cd, Zn, and Cu concentrations by
particle size near a Pb smelter. Atmos Environ 10: 443-446.
http://dx.doi.org/10.1016/0004-6981(76)90024-X
Dos Santos. M; Gomez. D; Dawidowski. L; Gautier. E; Smichowski. P. (2009). Determination of water-
soluble and insoluble compounds in size classified airborne paniculate matter. Microchem J 91:
133-139. http://dx.doi.0rg/10.1016/i.microc.2008.09.001
DRRC (Doe Run Company). (2010). DRRC multi-media Consent Decree. Herculaneum, MO: Doe Run
Resources Corporation.
Dunlap. CE; Alpers. CN; Bouse. R; Taylor. HE; Unruh. DM; Flegal. AR. (2008). The persistence of lead
from past gasoline emissions and mining drainage in a large riparian system: Evidence from lead
isotopes in the Sacramento River, California. Geochim Cosmo Act 72: 5935-5948.
http://dx.doi.org/10.1016/i.gca.20Q8.10.006
Duzgoren-Aydin. NS. (2007). Sources and characteristics of lead pollution in the urban environment of
Guangzhou. Sci Total Environ 385: 182-195. http://dx.doi.Org/10.1016/i.scitotenv.2007.06.047
Duzgoren-Aydin. NS: Weiss. AL. (2008). Use and abuse of Pb-isotope fingerprinting technique and GIS
mapping data to assess lead in environmental studies. Environ Geochem Health 30: 577-588.
http://dx.doi.org/10.1007/sl0653-008-9179-4
Edwards. RD; Lam. NL; Zhang. L; Johnson. MA; Kleinman. MT. (2009). Nitrogen dioxide and ozone as
factors in the availability of lead from lead-based paints. Environ Sci Technol 43: 8516-8521.
http://dx.doi.org/10.1021/es901077m
EIA (Energy Information Administration). (2012). Electric power monthly. Table 1.2. Net generation by
energy source: Electric utilities, 2002-June 2012.
http://www.eia.gov/electricitv/monthlv/epm table grapher.cfm?t=epmt 1 2
Erel. Y. (1998). Mechanisms and velocities of anthropogenic Pb migration in Mediterranean soils.
Environ Res 78: 112-117. http://dx.doi.org/10.1006/enrs.1997.3811
Erel. Y; Davan. U; Rabi. R; Rudich. Y; Stein. M. (2006). Trans boundary transport of pollutants by
atmospheric mineral dust. Environ Sci Technol 40: 2996-3005. http://dx.doi.org/10.1021/es0515021
ERG (Eastern Research Group Inc.). (2011). Documentation for aircraft component of the national
emissions inventory methodology. (0245.03.402.001). Morrisville, NC: Eastern Research Group.
Evans. GC: Norton. SA; Fernandez. IJ; Kahl. JS: Hanson. D. (2005). Changes in concentrations of major
elements and trace metals in northeastern US-Canadian sub-alpine forest floors. Water Air Soil Pollut
163: 245-267.
Ewen. C: Anagnostopoulou. MA; Ward. NL (2009). Monitoring of heavy metal levels in roadside dusts of
Thessaloniki, Greece in relation to motor vehicle traffic density and flow. Environ Monit Assess 157:
483-498. http://dx.doi.org/10.1007/sl0661-008-0550-9
November 2012 3-230 Draft - Do Not Cite or Quote
-------
Ewing. SA; Christensen. JN; Brown. ST; Vancuren. RA; Cliff. SS: Depaolo. DJ. (2010). Pb isotopes as an
indicator of the Asian contribution to paniculate air pollution in urban California. Environ Sci Technol
44: 8911-8916. http://dx.doi.org/10.1021/esl01450t
FAA (U.S. Federal Aviation Administration). (2011). Emissions and Dispersion Modeling System
(EDMS) (Version 5.1.3) [Computer Program]. Washington, D.C. Retrieved from
http://www.faa.gov/about/office org/headquarters offices/apl/research/models/edms model/
Falgayrac. G: Sobanska. S: Laureyns. J: Bremard. C. (2006). Heterogeneous chemistry between PbSO4
and calcite microparticles using Raman microimaging. Spectrochim Acta A Mol Biomol Spectrosc 64:
1095-1101. http://dx.doi.0rg/10.1016/j.saa.2005.ll.032
Falta. T; Limbeck. A; Koellensperger. G: Hann. S. (2008). Bioaccessibility of selected trace metals in
urban PM2.5 and PM10 samples: A model study. Anal Bioanal Chem 390: 1149-1157.
http://dx.doi.org/10.1007/s00216-007-1762-5
Fang. GC: Huang. CS. (2011). Atmospheric Particulate and Metallic Elements (Zn, Ni, Cu, Cd and Pb)
Size Distribution at Three Characteristic Sampling Sites. Environ Forensics 12: 191-199.
http://dx.doi.org/10.1080/15275922.2011.595052
Fang. TH; Li. JY; Feng. HM; Chen. HY. (2009). Distribution and contamination of trace metals in surface
sediments of the East China Sea. Mar Environ Res 68: 178-187.
http://dx.doi.0rg/10.1016/i.marenvres.2009.06.005
Farag. AM; Nimick. DA; Kimball. BA; Church. SE; Harper. DP; Brumbaugh. WG. (2007).
Concentrations of metals in water, sediment, biofilm, benthic macroinvertebrates, and fish in the
Boulder River watershed, Montana, and the role of colloids in metal uptake. Arch Environ Contam
Toxicol 52: 397-409. http://dx.doi.org/10.1007/s00244-005-0021-z
Farag. AM; Woodward. DF; Goldstein. JN; Brumbaugh. W: Meyer. JS. (1998). Concentrations of metals
associated with mining waste in sediments, biofilm, benthic macroinvertebrates, and fish from the
Coeur d'Alene River Basin, Idaho. Arch Environ Contam Toxicol 34: 119-127.
http://dx.doi.org/10.1007/s002449900295
Farfel. MR; Orlova. AO; Lees. PSJ: Rohde. C: Ashley. PJ: Chisolm. JJ. Jr. (2003). A study of urban
housing demolitions as sources of lead in ambient dust: Demolition practices and exterior dust fall.
Environ Health Perspect 111: 1228-1234.
Farfel. MR; Orlova. AO: Lees. PSJ: Rohde. C: Ashley. PJ: Chisolm. JJ. Jr. (2005). A study of urban
housing demolition as a source of lead in ambient dust on sidewalks, streets, and alleys. Environ Res
99: 204-213. http://dx.doi.Org/10.1016/i.envres.2004.10.005
Farmer. JG: Eades. LJ; Graham. MC: Cloy. JM; Bacon. JR. (2010). A comparison of the isotopic
composition of lead in rainwater, surface vegetation and tree bark at the long-term monitoring site,
Glensaugh, Scotland, in 2007. Sci Total Environ 408: 3704-3710.
http://dx.doi.0rg/10.1016/i.scitotenv.2010.03.050
Farmer. JG: Eades. LJ: MacKenzie. AB; Kirika. A; Bailey-Watts. TE. (1996). Stable lead isotope record
of lead pollution in Loch Lomond sediments since 1630 AD. Environ Sci Technol 30: 3080-3083.
http://dx.doi.org/10.1021/es960162o
Farmer. JG: Graham. MC: Bacon. JR; Dunn. SM; Vinogradoff. SI. (2005). Isotopic characterisation of the
historical lead deposition record at Glensau, an organic-rich, upland catchment in rural NE Scotland.
Sci Total Environ 346: 121-137. http://dx.doi.Org/10.1016/j.scitotenv.2004.ll.020
Fauser. P. (1999) Particulate air pollution with emphasis on traffic generated aerosols. (Doctoral
Dissertation). Technical University of Denmark (DTU), Kgs. Lyngby, Denmark.
Fernandez. C: Labanowski. J; Cambier. P; Jongmans. AG: Van Port. F. (2007). Fate of airborne metal
pollution in soils as related to agricultural management. 1. Zn and Pb distributions in soil profiles. Eur
J Soil Sci 58: 547-559. http://dx.doi.Org/10.llll/i.1365-2389.2006.00827.x
November 2012 3-231 Draft - Do Not Cite or Quote
-------
Fernandez Espinosa. AJ; Ternero Rodriguez. M; Barragan De La Rosa. FJ; Jimenez Sanchez. JC. (2002).
A chemical speciation of trace metals for fine urban particles. Atmos Environ 36: 773-780.
http://dx.doi.org/10.1016/81352-2310(01)00534-9
Fernandez Espinosa. AJ: Ternero Rodriguez. M; Fernandez Alvarez. F. (2004). Source characterisation of
fine urban particles by multivariate analysis of trace metals speciation. Atmos Environ 38: 873-886.
http://dx.doi.0rg/10.1016/i.atmosenv.2003.10.046
Fernando. HJS. (2010). Fluid dynamics of urban atmospheres in complex terrain. Annu Rev Fluid Mech
42: 365-389. http://dx.doi.org/10.1146/annurev-fluid-121108-145459
Field. PM; Sherrell. RM. (2003). Direct determination of ultra-trace levels of metals in fresh water using
desolvating micronebulization and HR-ICP-MS: Application to Lake Superior waters. J Anal At
Spectrom 18: 254-259. http://dx.doi.org/10.1039/b210628k
Filippelli. GM; Laidlaw. MAS. (2010). The elephant in the playground: Confronting lead-contaminated
soils as an important source of lead burdens to urban populations. Perspect Biol Med 53: 31-45.
http://dx.doi.0rg/10.1353/pbm.0.0136
Filippelli. GM; Laidlaw. MAS: Latimer. JC: Raftis. R. (2005). Urban lead poisoning and medical
geology: An unfinished story. Geological Society of America Today 15: 4-11.
http://dx.doi.org/10.1130/1052-5173(2005)015<4:ULPAMG>2.0.CO:2
Finlavson-Pitts. BJ; Pitts. JN. Jr. (2000). Chemistry of the upper and lower atmosphere: theory,
experiments and applications. San Diego, CA: Academic Press.
Flint. KR; Davis. AP (2007). Pollutant mass flushing characterization of highway storm water runoff from
an ultra-urban area. J Environ Eng 133: 616-626.
http://dx.doi.org/10.1061/(ASCE)0733-9372(2007)133:6(616)
Fowler. D; McDonald. AG: Crosslev. A; Nemitz. E; Leaver. D; Cape. JN; Smith. RI; Anderson. D;
Rowland. P; Ainsworth. G: Lawlor. AJ: Guvatt H; Harmens. H. (2006). U.K. heavy metal monitoring
network. (AS 06/07). Edinburgh, U.K.: NERC/Centre for Ecology and Hydrology.
http://nora.nerc.ac.uk/3323
Fowler. D; Smith. RI: Muller. JB; Havman. G: Vincent. KJ. (2005). Changes in the atmospheric
deposition of acidifying compounds in the U.K. between 1986 and 2001. Environ Pollut 137: 15-25.
http://dx.doi.0rg/10.1016/i.envpol.2004.12.028
Franzaring. J: Holz. I; Zipperle. J: Fangmeier. A. (2010). Twenty years of biological monitoring of
element concentrations in permanent forest and grassland plots in Baden-Wurttemberg (SW
Germany). Environ Sci Pollut Res Int 17: 4-12. http://dx.doi.org/10.1007/sll356-009-0181-x
Fraser. JL; Lum. KR. (1983). Availability of elements of environmental importance in incinerated sludge
ash. Environ Sci Technol 17: 52-54. http://dx.doi.org/10.1021/es00107a013
Freme. F. (2004). U.S. coal supply and demand: 2004 Review. Washington, DC: U.S. Energy Information
Administration. http://tonto.eia.doe.gov/FTPROOT/features/feature04.pdf
Fujiwara. F; Dos Santos. M; Marrero. J: Polla. G: Gomez. D; Dawidowskia. L; Smichowski. P. (2006).
Fractionation of eleven elements by chemical bonding from airborne paniculate matter collected in an
industrial city in Argentina. J Environ Monit 8: 913-922. http://dx.doi.org/10.1039/b604307k
Funasaka. K; Tojo. T; Katahira. K; Shinya. M; Miyazaki. T; Kamiura. T; Yamamoto. O: Moriwaki. H;
Tanida. H; Takaoka. M. (2008). Detection of Pb-LIII edge XANES spectra of urban atmospheric
particles combined with simple acid extraction. Sci Total Environ 403: 230-234.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.05.020
Gallon. C: Tessier. A; Gobeil. C. (2006). Historical perspective of industrial lead emissions to the
atmosphere from a Canadian smelter. Environ Sci Technol 40: 741-747.
http://dx.doi.org/10.1021/es051326g
November 2012 3-232 Draft - Do Not Cite or Quote
-------
Gao. Y; Lesven. L; Gillan. D; Sabbe. K; Billon. G: De Galan. S: Elskens. M; Baevens. W: Leermakers.
M. (2009). Geochemical behavior of trace elements in sub-tidal marine sediments of the Belgian
coast. Mar Chem 117: 88-96. http://dx.doi.Org/10.1016/j.marchem.2009.05.002
Garland. JA; Nicholson. KW. (1991). A review of methods for sampling large airborne particles and
associated radioactivity [Review]. J Aerosol Sci 22: 479-499.
http://dx.doi.org/10.1016/0021-8502(91)90006-4
Garrido. F; Serrano. S: Campbell. CG: Barrios. L; Garcia-Gonzalez. AT. (2008). Evidence of physical and
chemical nonequilibrium in lead and cadmium transport and sorption in acid soils. Soil Sci Soc Am J
72: 1434-1444. http://dx.doi.org/10.2136/sssaj2007.0411
Geagea. ML; Stille. P; Gauthier-Lafave. F; Perrone. T; Aubert. D. (2008). Baseline determination of the
atmospheric Pb, Sr and Nd isotopic compositions in the Rhine valley, Vosges mountains (France) and
the Central Swiss Alps. Appl Geochem 23: 1703-1714.
http://dx.doi.0rg/10.1016/i.apgeochem.2008.02.004
Gidnev. J: Twigg. M; Kittelson. D. (2010). Effect of organometallic fuel additives on nanoparticle
emissions from a gasoline passenger car. Environ Sci Technol 44: 2562-2569.
http://dx.doi.org/10.1021/es901868c
Giere. R; Blackford. M; Smith. K. (2006). TEM study of PM2.5 emitted from coal and tire combustion in
a thermal power station. Environ Sci Technol 40: 6235-6240. http://dx.doi.org/10.1021/es060423m
Gillette. DA; Blifford. IH. Jr; Fryrear. DW. (1974). The influence of wind velocity on the size
distributions of aerosols generated by the wind erosion of soils. J Geophys Res 79: 4068-4075.
http://dx.doi.org/10.1029/JC079i027p04068
Goforth. MR; Christoforou. CS. (2006). Particle size distribution and atmospheric metals measurements
in a rural area in the South Eastern USA. Sci Total Environ 356: 217-227.
http://dx.doi.0rg/10.1016/i.scitotenv.2005.03.017
Goval. A; Small. MJ; Von Stackelberg. K; Burmistrov. D. (2005). Estimation of fugitive lead emission
rates from secondary lead facilities using hierarchical Bayesian models. Environ Sci Technol 39:
4929-4937. http://dx.doi.org/10.1021/es035465e
Graham. MC: Vinogradoff. SI; Chipchase. AJ; Dunn. SM; Bacon. JR; Farmer. JG. (2006). Using size
fractionation and Pb isotopes to study Pb transport in the waters of an organic-rich upland catchment.
Environ Sci Technol 40: 1250-1256. http://dx.doi.org/10.1021/es0517670
Granev. JR: Landis. MS: Norris. GA. (2004). Concentrations and solubility of metals from indoor and
personal exposure PM2.5 samples. Atmos Environ 38: 237-247.
http://dx.doi.0rg/10.1016/i.atmosenv.2003.09.052
Griffith. DA. (2002). The geographic distribution of soil lead concentration: Description and concerns.
URISA Journal 14: 5-16.
Griffith. DA; Johnson. PL; Hunt. A. (2009). The geographic distribution of metals in urban soils: The
case of Syracuse, NY. GeoJournal 74: 275-291. http://dx.doi.org/10.1007/sl0708-008-9233-x
Gulson. B; Korsch. M; Dickson. B; Cohen. D; Mizon. K; Davis. JM. (2007). Comparison of lead isotopes
with source apportionment models, including SOM, for air particulates. Sci Total Environ 381:
169-179. http://dx.doi.0rg/10.1016/i.scitotenv.2007.03.018
Guo. T; Sansalone. J: Piro. P. (2006a). The role of in situ unit operation/process infiltration treatment on
partitioning and speciation of rainfall-runoff. Water Sci Technol 54: 255-261.
http://dx.doi.org/10.2166/wst.2006.589
Guo. XY: Zhang. SZ: Shan. XO: Luo. L: Pei. ZG: Zhu. YG: Liu. T: Xie. YN: Gault. A. (2006b).
Characterization of Pb, Cu, and Cd adsorption on paniculate organic matter in soil. Environ Toxicol
Chem 25: 2366-2373. http://dx.doi.Org/10.1897/05-636R.l
November 2012 3-233 Draft - Do Not Cite or Quote
-------
Gutierrez-Castillo. ME; Olivos-Ortiz. M; De Vizcava-Ruiz. A; Cebrian. ME. (2005). Chemical
characterization of extractable water soluble matter associated with PM10 from Mexico City during
2000. Chemosphere 61: 701-710. http://dx.doi.Org/10.1016/i.chemosphere.2005.03.063
H. K; M. K; W. S. (2010). Atmospheric Pb and Ti accumulation rates from sphagnum moss: Dependence
upon plant productivity. Environ Sci Technol 44: 5509-5515. http://dx.doi.org/10.1021/esl00366d
Halm. I; Brixev. LA; Wiener. RW: Henkle. SW: Baldauf. R. (2009). Characterization of traffic-related PM
concentration distribution and fluctuation patterns in near-highway urban residential street canyons. J
Environ Monit 11: 2136-2145. http://dx.doi.org/10.1039/b907130j
Hallberg. M; Renman. G: Lundbom. T. (2007). Seasonal variations of ten metals in highway runoff and
their partition between dissolved and paniculate matter. Water Air Soil Pollut 181: 183-191.
http://dx.doi.org/10.1007/sll270-006-9289-5
Harmens. H; Norris. DA; Koerber. GR; Buse. A; Steinnes. E; Ruhling. A. (2008). Temporal trends
(1990-2000) in the concentration of cadmium, lead and mercury in mosses across Europe. Environ
Pollut 151: 368-376. http://dx.doi.Org/10.1016/i.envpol.2007.06.043
Harmens. H; Norris. DA; Steinnes. E; Kubin. E; Piispanen. J; Alber. R; Aleksiavenak. Y; Blum. O;
Coskun. M; Dam. M; De Temmerman. L; Fernandez. JA; Frolova. M; Frontasveva. M; Gonzalez-
Miqueo. L; Grodzinska. K; Jeran. Z; Korzekwa. S: Krmar. M; Kvietkus. K; Leblond. S: Liiv. S:
Magnusson. SH; Mankovska. B; Pesch. R; Ruhling. A; Santamaria. JM; Schroder. W: Spiric. Z;
Suchara. I; Thoni. L; Urumov. V; Yurukova. L; Zechmeister. HG. (2010). Mosses as biomonitors of
atmospheric heavy metal deposition: Spatial patterns and temporal trends in Europe. Environ Pollut
158: 3144-3156. http://dx.doi.Org/10.1016/i.envpol.2010.06.039
Harris. AR; Davidson. CI. (2005). The role of resuspended soil in lead flows in the California South Coast
Air Basin. Environ Sci Technol 39: 7410-7415. http://dx.doi.org/10.1021/es050642s
Harris. AR; Davidson. CI. (2008). Particle resuspension in turbulent flow: A stochastic model for
individual soil grains. Aerosol Sci Technol 42: 613-628.
http://dx.doi.org/10.1080/02786820802227337
Harrison. RM; Tilling. R; Callen Romero. MS: Harrad. S: Jarvis. K. (2003). A study of trace metals and
polycyclic aromatic hydrocarbons in the roadside environment. Atmos Environ 37: 2391-2402.
http://dx.doi.org/10.1016/81352-2310(03)00122-5
Harrison. RM; Yin. J. (2010). Chemical speciation of PM2.5 particles at urban background and rural sites
in the U.K. atmosphere. J Environ Monit 12: 1404-1414. http://dx.doi.org/10.1039/c000329h
Hartnett. M; Berry. A. (2010). Transport of lead in the Mersey Estuary: The development of a novel
approach to deriving partition coefficients. Adv Eng Software 41: 84-91.
http://dx.doi.0rg/10.1016/i.advengsoft.2008.ll.007
Hasselbach. L; Ver Hoef. JM; Ford. J: Neitlich. P; Crecelius. E; Berryman. S: Wolk. B; Bohle. T. (2005).
Spatial patterns of cadmium and lead deposition on and adjacent to National Park Service lands in the
vicinity of Red Dog Mine, Alaska. Sci Total Environ 348: 211-230.
http://dx.doi.0rg/10.1016/i.scitotenv.2004.12.084
Hassellov. M; von der Kammer. F. (2008). Iron oxides as geochemical nanovectors for metal transport in
soil-river systems. Elements 4: 401-406. http://dx.doi.Org/10.2113/gselements.4.6.401
Hays. MD; Cho. SH; Baldauf. R: JJ. S: Shafer. M. (2011). Particle size distributions of metal and non-
metal elements in an urban near-highway environment. Atmos Environ 45: 925-934.
http://dx.doi.0rg/10.1016/i.atmosenv.2010.ll.010
Heal. MR; Hibbs. LR; Agius. RM: Beverland. IJ. (2005). Interpretation of variations in fine, coarse and
black smoke paniculate matter concentrations in a northern European city. Atmos Environ 39:
3711-3718. http://dx.doi.0rg/10.1016/i.atmosenv.2005.03.007
November 2012 3-234 Draft - Do Not Cite or Quote
-------
Helmreich. B; Hilliges. R; Schriewer. A; Horn. H. (2010). Runoff pollutants of a highly trafficked urban
road: Correlation analysis and seasonal influences. Chemosphere 80: 991-997.
http://dx.doi.0rg/10.1016/i.chemosphere.2010.05.037
Hinds. WC. (1999). Aerosol technology: Properties, behavior, and measurement of airborne particles (2nd
ed.). New York: John Wiley & Sons.
Hirner. AV. (2006). Speciation of alkylated metals and metalloids in the environment. Anal Bioanal Chem
385: 555-567. http://dx.doi.org/10.1007/s00216-006-0368-7
Hitzenberger. R; Berner. A; Galambos. Z; Maenhaut. W: Cafmever. J; Schwarz. J; Muller. K; Spindler. G:
Wieprecht. W: Acker. K; Hillamo. R; Makela. T. (2004). Intercomparison of methods to measure the
mass concentration of the atmospheric aerosol during INTERCOMP2000 - Influence of
instrumentation and size cuts. Atmos Environ 38: 6467-6476.
http://dx.doi.0rg/10.1016/i.atmosenv.2004.08.025
Hjortenkrans. DST; Bergback. BG: Haggerud. AV. (2007). Metal emissions from brake linings and tires:
Case studies of Stockholm, Sweden 1995/1998 and 2005. Environ Sci Technol 41: 5224-5230.
http://dx.doi.org/10.1021/es070198o
Holsen. TM; Noll. KB; Fang. GC: Lee. WJ: Lin. JM; Keeler. GJ. (1993). Dry deposition and particle size
distributions measured during the Lake Michigan Urban Air Toxics Study. Environ Sci Technol 27:
1327-1333. http://dx.doi.org/10.1021/es00044a006
Hong. S: Candelone. JP; Patterson. CC: Boutron. CF. (1994). Greenland ice evidence of hemispheric lead
pollution two millennia ago by Greek and Roman civilizations. Science 265: 1841-1843.
http://dx.doi.org/10.1126/science.265.5180.1841
Horowitz. AJ; Elrick. KA; Smith. JJ. (2008). Monitoring urban impacts on suspended sediment, trace
element, and nutrient fluxes within the City of Atlanta, Georgia, USA: Program design,
methodological considerations, and initial results. Hydrolog Process 22: 1473-1496.
http://dx.doi.org/10.1002/hyp.6699
Horowitz. AJ; Stephens. VC. (2008). The effects of land use on fluvial sediment chemistry for the
conterminous U.S. Results from the first cycle of the NAWQA Program: Trace and major elements,
phosphorus, carbon, and sulfur. Sci Total Environ 400: 290-314.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.04.027
Hossain. MA; Furumai. H; Nakajima. F; Aryal. RK. (2007). Heavy metals speciation in soakaways
sediment and evaluation of metal retention properties of surrounding soil. Water Sci Technol 56:
81-89. http://dx.doi.org/10.2166/wst.2007.746
Hou. A; Laws. E; Gambrell. R: Tan. M; Delaune. R: Li. Y; Roberts. H. (2006). Pathogen indicator
microbes and heavy metals in Lake Pontchartrain following Hurricane Katrina. Environ Sci Technol
40: 5904-5910. http://dx.doi.org/10.1021/es060946u
Hsu. SC: Chen Liu. S: Jeng. WL; Chou. CC. KG: Hsu. RT; Huang. YT; Chen. YY. (2006). Lead isotope
ratios in ambient aerosols from Taipei, Taiwan: Identifying long-range transport of airborne Pb from
the Yangtze Delta. Atmos Environ 40: 5393-5404. http://dx.doi.Org/10.1016/j.atmosenv.2006.05.003
Hsu. SC: Liu. SC: Huang. YT: Chou. CCK: Lung. SCC: Liu. TH: Tu. JY: Tsai. FJ. (2009c). Long-range
southeastward transport of Asian biosmoke pollution: Signature detected by aerosol potassium in
Northern Taiwan. J Geophys Res 114: D14301. http://dx.doi.org/10.1029/2009jd011725
Hu. S: Herner. JD; Shafer. M; Robertson. W: Schauer. JJ: Dwver. H; Collins. J: Huai. T; Ayala. A.
(2009a). Metals emitted from heavy-duty diesel vehicles equipped with advanced PM and NOx
emission controls. Atmos Environ 43: 2950-2959. http://dx.doi.Org/10.1016/i.atmosenv.2009.02.052
Huang. S: Conte. MH. (2009). Source/process apportionment of major and trace elements in sinking
particles in the Sargasso sea. Geochim Cosmo Act 73: 65-90.
http://dx.doi.0rg/10.1016/i.gca.2008.08.023
November 2012 3-235 Draft - Do Not Cite or Quote
-------
Hur. SD: Cunde. X; Hong. S: Barbante. C: Gabrielli. P; Lee. K; Boutron. CF; Ming. Y. (2007). Seasonal
patterns of heavy metal deposition to the snow on Lambert Glacier basin, East Antarctica. Atmos
Environ 41: 8567-8578. http://dx.doi.Org/10.1016/i.atmosenv.2007.07.012
Huston. R; Chan. YC: Gardner. T; Shaw. G: Chapman. H. (2009). Characterisation of atmospheric
deposition as a source of contaminants in urban rainwater tanks. Water Res 43: 1630-1640.
http://dx.doi.0rg/10.1016/i.watres.2008.12.045
IAEA (International Atomic Energy Agency). (1993). The "GENT" Stacked Filter Unit (SFU) sampler for
the collection of atmospheric aerosols in two size fractions: Description and instructions for
installation and use. Vienna, VA.
Ikegawa. M; Kimura. M; Honda. K; Akabane. I; Makita. K; Motovama. H; Fujii. Y; Itokawa. Y. (1999).
Geographical variations of major and trace elements in East Antarctica. Atmos Environ 33:
1457-1467. http://dx.doi.org/10.1016/S1352-2310(98)00243-X
Illinois Environmental Protection Agency. (2002). Chicago OHare Airport air toxic monitoring program.
Springfield, IL. http://www.epa.state.il.us/air/ohare/
Ishizaka. T; Tohno. S: Ma. CJ: Morikawa. A; Takaoka. M; Nishiyama. F; Yamamoto. K. (2009).
Reactivity between PbS04 and CaCO3 particles relevant to the modification of mineral particles and
chemical forms of Pb in particles sampled at two remote sites during an Asian dust event. Atmos
Environ 43: 2550-2560. http://dx.doi.Org/10.1016/i.atmosenv.2009.02.041
Jennings. AA; Ma. J. (2007). Variation in North American regulatory guidance for heavy metal surface
soil contamination at commercial and industrial sites. Journal of Environmental Engineering and
Science 6: 587-609. http://dx.doi.org/10.1139/s07-010
Jensen. MB; Holm. PE; Laursen. J; Hansen. HCB. (2006). Contaminant aspects of blackish surface
deposits on highway roadsides. Water Air Soil Pollut 175: 305-321.
http://dx.doi.org/10.1007/sll270-006-9140-z
Jerez. SB; Zhang. Y; Mcclure. JW: Jacobson. L; Heber. A; Hoff S: Koziel. J: Beaslev. D. (2006).
Comparison of measured total suspended paniculate matter concentrations using tapered element
oscillating microbalance and a total suspended paniculate sampler. J Air Waste Manag Assoc 56:
261-270.
Jimenez. S: Perez. M; Ballester. J. (2008). Vaporization of trace elements and their emission with
submicrometer aerosols in biomass combustion. Energy Fuels 22: 2270-2277.
http://dx.doi.org/10.1021/ef800111u
Jitaru. P; Infante. HG: Adams. FC. (2004). Simultaneous multi-elemental speciation analysis of
organometallic compounds by solid-phase microextraction and multicapillary gas chromatography
hyphenated to inductively coupled plasma-time-of-flight-mass spectrometry. J Anal At Spectrom 19:
867-875. http://dx.doi.org/10.1039/B404106B
Johnson. AH; Richter. SL. (2010). Organic-horizon lead, copper, and zinc contents of Mid-Atlantic forest
soils, 1978-2004. Soil Sci Soc Am J 74: 1001-1009. http://dx.doi.org/10.2136/sssai2008.0337
Johnson. KS: de Foy. B; Zuberi. B; Molina. IT; Molina. MJ; Xie. Y; Laskin. A; Shutthanandan. V
(2006). Aerosol composition and source apportionment in the Mexico City Metropolitan Area with
PIXE/PESA/STIM and multivariate analysis. Atmos Chem Phys 6: 4591-4600.
http://dx.doi.org/10.5194/acp-6-4591-2006
Johnson. KS: Laskin. A; Jimenez. JL; Shutthanandan. V; Molina. IT; Salcedo. D; Dzepina. K; Molina.
MJ. (2008). Comparative analysis of urban atmospheric aerosol by particle-induced X-ray emission
(PIXE), proton elastic scattering analysis (PESA), and aerosol mass spectrometry (AMS). Environ Sci
Technol 42: 6619-6624. http://dx.doi.org/10.1021/es800393e
Joshi. UM; Viiavaraghavan. K; Balasubramanian. R. (2009). Elemental composition of urban street dusts
and their dissolution characteristics in various aqueous media. Chemosphere 77: 526-533.
http://dx.doi.0rg/10.1016/i.chemosphere.2009.07.043
November 2012 3-236 Draft - Do Not Cite or Quote
-------
Kang. S: Zhang. Q; Kaspari. S: Qin. D; Cong. Z; Ren. J; Mavewski. PA. (2007). Spatial and seasonal
variations of elemental composition in Mt. Everest (Qomolangma) snow/firn. Atmos Environ 41:
72087218. http://dx.doi.0rg/10.1016/i.atmosenv.2007.05.024
Kaste. J: Friedland. A; Sturup. S. (2003). Using stable and radioactive isotopes to trace atmospherically
deposited Pb in montane forest soils. Environ Sci Technol 37: 3560-3567.
http://dx.doi.org/10.1021/es026372k
Kaste. JM; Bostick. BC: Friedland. AJ; Schroth. AW: Siccama. TG. (2006). Fate and speciation of
gasoline-derived lead in organic horizons of the northeastern USA. Soil Sci Soc Am J 70: 1688-1698.
http://dx.doi.org/10.2136/sssai2005.0321
Kastner-Klein. P; Fedorovich. E; Ketzel. M; R. B; Britter. R. (2003). The modelling of turbulence from
traffic in urban dispersion models part II: Evaluation against laboratory and full-scale concentration
measurements in street canyons. Environ Fluid Mech 3: 145-172.
http://dx.doi.Org/10.1023/A:1022049224166
Kastner-Klein. P; Fedorovich. E; Rotach. MW. (2001). A wind tunnel study of organised and turbulent air
motions in urban street canyons. J Wind Eng Ind Aerod 89: 849-861.
http://dx.doi.org/10.1016/S0167-6105(01)00074-5
Kastner-Klein. P; Rotach. MW. (2004). Mean flow and turbulence characteristics in an urban roughness
sublayer. Boundary Layer Meteorol 111: 55-84.
http://dx.doi.Org/10.1023/B:BOUN.0000010994.32240.bl
Kayhanian. M; Suverkropp. C: Ruby. A; Tsav. K. (2007). Characterization and prediction of highway
runoff constituent event mean concentration. J Environ Manage 85: 279-295.
http://dx.doi.0rg/10.1016/i.jenvman.2006.09.024
Kenny. L; Beaumont. G: Gudmundsson. A; Thorpe. A; Koch. W (2005). Aspiration and sampling
efficiencies of the TSP and louvered paniculate matter inlets. J Environ Monit 7: 481-487.
http://dx.doi.org/10.1039/b419001g
Keskinen. J: Pietarinen. K; Lehtimaki. M. (1992). Electrical low pressure impactor. J Aerosol Sci 23:
353-360. http://dx.doi.org/10.1016/0021-8502(92)90004-F
Kidwell. CB; Ondov. JM. (2001). Development and evaluation of a prototype system for collecting sub-
hourly ambient aerosol for chemical analysis. Aerosol Sci Technol 35: 596-601.
http://dx.doi.org/10.1080/02786820118049
Kidwell. CB; Ondov. JM. (2004). Elemental analysis of sub-hourly ambient aerosol collections. Aerosol
Sci Technol 38: 205-218. http://dx.doi.org/10.1080/02786820490261726
Kimbrough. KL; Lauenstein. GG: Christensen. JD; Apeti. DA. (2008). An assessment of two decades of
contaminant monitoring in the nation's coastal zone. Silver Spring, MD: National Centers for Coastal
Ocean Science, http://aquaticcommons.org/2232/
Klaminder. J: Bindler. R; Emteryd. O: Applebv. P: Grip. H. (2006a). Estimating the mean residence time
of lead in the organic horizon of boreal forest soils using 210-lead, stable lead and a soil
chronosequence. Biogeochemistry 78: 31-49. http://dx.doi.org/10.1007/sl0533-005-2230-y
Klaminder. J: Bindler. R; Emteryd. O: Renberg. I. (2005). Uptake and recycling of lead by boreal forest
plants: Quantitative estimates from a site in northern Sweden. Geochim Cosmo Act 69: 2485-2496.
http://dx.doi.0rg/10.1016/i.gca.2004.ll.013
Klaminder. J: Bindler. R; Laudon. H; Bishop. K; Emteryd. O: Renberg. I. (2006b). Flux rates of
atmospheric lead pollution within soils of a small catchment in northern Sweden and their
implications for future stream water quality. Environ Sci Technol 40: 4639-4645.
http://dx.doi.org/10.1021/es0520666
Klaminder. J: Bindler. R; Renberg. I. (2008a). The biogeochemistry of atmospherically derived Pb in the
boreal forest of Sweden. Appl Geochem 23: 2922-2931.
http://dx.doi.0rg/10.1016/i.apgeochem.2008.04.007
November 2012 3-237 Draft - Do Not Cite or Quote
-------
Klaminder. J; Bindler. R; Rydberg. J; Renberg. I. (2008b). Is there a chronological record of atmospheric
mercury and lead deposition preserved in the mor layer (O-horizon) of boreal forest soils? Geochim
Cosmo Act 72: 703-712. http://dx.doi.Org/10.1016/i.gca.2007.10.030
Klaminder. J: Hammarlund. D; Kokfelt. U: Vonk. JE; Bigler. C. (2010). Lead contamination of subarctic
lakes and its response to reduced atmospheric fallout: Can the recovery process be counteracted by the
ongoing climate change? Environ Sci Technol 44: 2335-2340. http://dx.doi.org/10.1021/es903025z
Knowlton. SW: Moran. SB. (2010). Stable Pb isotope ratios in aerosols, precipitation, and size-
fractionated paniculate matter in the Gulf of Maine, Scotian Shelf, and Labrador Sea. Mar Pollut Bull
60: 984-989. http://dx.doi.Org/10.1016/i.marpolbul.2010.02.005
Kumar. P; Hopke. PK; Raja. S: Casuccio. G: Lersch. TL; West. RR. (2012). Characterization and
heterogeneity of coarse particles across an urban area. Atmos Environ 46: 449-459.
Kylander. ME; Cortizas. AM; Rauch. S: Weiss. DJ. (2008). Lead penetration and leaching in a complex
temperate soil profile. Environ Sci Technol 42: 3177-3184. http://dx.doi.org/10.1021/es702358e
Kyotani. T; Iwatsuki. M. (2002). Characterization of soluble and insoluble components in PM25 and
PM10 fractions of airborne particulate matter in Kofu city, Japan. Atmos Environ 36: 639-649.
http://dx.doi.org/10.1016/81352-2310(01)00494-0
Laeudu. URK: Raia. S: Hopke. PK: Chalupa. DC: Utell. MJ: Casuccio. G: Lersch. TL: West. RR. (2011).
Heterogeneity of Coarse Particles in an Urban Area. Environ Sci Technol 45: 3288-3296.
http://dx.doi.org/10.1021/esl03831w
Lai. X; Norisuve. K; Mikata. M; Minami. T; Bowie. AR; Sohrin. Y. (2008b). Spatial and temporal
distribution of Fe, Ni, Cu and Pb along 140 degrees E in the Southern Ocean during austral summer
2001/02. Mar Chem 111: 171-183. http://dx.doi.Org/10.1016/i.marchem.2008.05.001
Laidlaw. MAS: Filippelli. GM. (2008). Resuspension of urban soils as a persistent source of lead
poisoning in children: A review and new directions. Appl Geochem 23: 2021-2039.
http://dx.doi.0rg/10.1016/i.apgeochem.2008.05.009
Laidlaw. MAS: Zahran. S: Mielke. HW: Taylor. MP; Filippelli. GM. (2012). Re-suspension of lead
contaminated urban soil as a dominant source of atmospheric lead in Birmingham, Chicago, Detroit
and Pittsburgh, USA. Atmos Environ 49: 302-310. http://dx.doi.Org/10.1016/i.atmosenv.2011.ll.030
Landers. DH; Simonich. SL; Jaffe. DA; Geiser. LH; Campbell DH; Schwindt. AR; Schreck. CB; Kent.
ML; Hafner. WD; Taylor. HE; Hageman. KJ; Usenko. S: Ackerman. LK; Schrlau. JE; Rose. NL; Blett.
TF; Erwav. MM. (2008). The fate, transport, and ecological impacts of airborne contaminants in
western national parks (USA). (EPA/600/R-07/138). Corvallis, Oregon: U.S. Environmental
Protection Agency, NHEERL, Western Ecology Division.
http://www.nature.nps.gov/air/studies/air toxics/WACAPreport.cfm
Landers. DH: Simonich. SM; Jaffe. D; Geiser. L; Campbell. DH: Schwindt. A; Schreck. C: Kent. M;
Hafner. W: Taylor. HE; Hageman. K; Usenko. S: Ackerman. L; Schrlau. J: Rose. N; Blett. T; Erwav.
MM. (2010). The Western Airborne Contaminant Assessment Project (WACAP): An interdisciplinary
evaluation of the impacts of airborne contaminants in western U.S. National Parks. Environ Sci
Technol 44: 855-859. http://dx.doi.org/10.1021/es901866e
Landre. AL; Watmough. SA; Dillon. PJ. (2009). The effects of dissolved organic carbon, acidity and
seasonality on metal geochemistry within a forested catchment on the Precambrian Shield, central
Ontario, Canada. Biogeochemistry 93: 271-289. http://dx.doi.org/10.1007/sl0533-009-9305-0
Landre. AL; Watmough. SA; Dillon. PJ. (2010). Metal pools, fluxes, and budgets in an acidified forested
catchment on the Precambrian Shield, Central Ontario, Canada. Water Air Soil Pollut 209: 209-228.
htrp://dx.doi.org/10.1007/sll270-009-0193-7
Lankev. RL; Davidson. CI; McMichael. FC. (1998). Mass balance for lead in the California south coast
air basin: an update. Environ Res 78: 86-93.
November 2012 3-238 Draft - Do Not Cite or Quote
-------
Lasheen. MR; Sharabv. CM; El-Kholy. NG: Elsherif. IY; El-Wakeel. ST. (2008). Factors influencing lead
and iron release from some Egyptian drinking water pipes. J Hazard Mater 160: 675-680.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.03.040
Leal-Granadillo. IA; Alonso. JIG: Sanz-Medel. A. (2000). Determination of the speciation of organolead
compounds in airborne paniculate matter by gas chromatography-inductively coupled plasma mass
spectrometry. Anal Chim Acta 423: 21-29. http://dx.doi.org/10.1016/S0003-2670(00)01032-l
Lee. G: Faure. G: Bigham. JM; Williams. DJ. (2008a). Metal release from bottom sediments of Ocoee
Lake No. 3, a primary catchment area for the Ducktown Mining District. J Environ Qua! 37: 344-352.
http://dx.doi.org/10.2134/iea2007.0223
Lee. K; Hur. SD; Hou. S: Hong. S: Qin. X; Ren. J: Liu. Y; Rosman. KJR; Barbante. C: Boutron. CF.
(2008b). Atmospheric pollution for trace elements in the remote high-altitude atmosphere in central
Asia as recorded in snow from Mt. Qomolangma (Everest) of the Himalayas. Sci Total Environ 404:
171-181. http://dx.doi.0rg/10.1016/i.scitotenv.2008.06.022
Lee. RE. Jr. (1972). Size distribution of suspended particulates in air. Res/Dev 23: 18-21.
Lee. RE. Jr; Patterson. RK; Wagman. J. (1968). Particle-size distribution of metal components in urban
air. Environ Sci Technol 2: 288-290.
Leith. D; Sommerlatt. D; Boundv. MG. (2007). Passive sampler for PM10-2.5 aerosol. J Air Waste Manag
Assoc 57: 332-336. http://dx.doi.org/10.1080/10473289.2007.10465336
Leland. HV; Kuwabara. JS. (1985). Trace metals. In GM Rand; SR Petrocelli (Eds.), Fundamentals of
aquatic toxicology: Methods and applications (pp. 374-415). Washington, DC: Hemisphere
Publishing.
Levin. R; Brown. MJ; Kashtock. ME; Jacobs. DE; Whelan. EA; Rodman. J: Schock. MR; Padilla. A;
Sinks. T. (2008). Lead exposures in US children, 2008: Implications for prevention. Environ Health
Perspect 116: 1285-1293. http://dx.doi.org/10.1289/ehp.11241
Lewis. J: Sjostrom. J: Skyllberg. U; Hagglund. L. (2010). Distribution, chemical speciation, and mobility
of lead and antimony originating from small arms ammunition in a coarse-grained unsaturated surface
sand. J Environ Qual 39: 863-870. http://dx.doi.org/10.2134/ieq2009.0211
Li. C: Nguyen. Q; Ryan. PH; Lemasters. GK; Spitz. H; Lobaugh. M; Glover. S: Grinshpun. SA. (2009a).
School bus pollution and changes in the air quality at schools: A case study. J Environ Monit 11:
1037-1042. http://dx.doi.org/10.1039/b819458k
Li. L; Pala. F; Jiang. M; Krahforst. C: Wallace. GT. (2010a). Three-dimensional modeling of Cu and Pb
distributions in Boston Harbor, Massachusetts and Cape Cod Bays. Estuar Coast Shelf Sci 88:
450-463. http://dx.doi.0rg/10.1016/i.ecss.2010.05.003
Lim. JH; Sabin. LD; Schiff. KG: Stolzenbach. KD. (2006). Concentration, size distribution, and dry
deposition rate of particle-associated metals in the Los Angeles region. Atmos Environ 40: 7810-7823.
http://dx.doi.0rg/10.1016/i.atmosenv.2006.07.025
Lin. CC: Chen. SJ: Huang. KL; Hwang. WI; Chang-Chien. GP; Lin. WY. (2005). Characteristics of
metals in nano/ultrafine/fme/coarse particles collected beside a heavily trafficked road. Environ Sci
Technol 39: 8113-8122.
Liu. W: Hopke. PK; van Curen. RA. (2003). Origins of fine aerosol mass in the western United States
using positive matrix factorization. J Geophys Res 108: 4716.
http://dx.doi.org/10.1029/2003JD003678
Lough. GC: Schauer. JJ; Park. JS: Shafer. MM; Deminter. JT; Weinstein. JP (2005). Emissions of metals
associated with motor vehicle roadways. Environ Sci Technol 39: 826-836.
Mager. EM. (2012). Lead. In Homeostasis and toxicology of non-essential metals. U.K.: Academic Press.
http://www.elsevierdirect.com/ISBN/9780123786340/Fish-Phvsiologv-Homeostasis-and-Toxicologv-
of-NonEssential-Metals
November 2012 3-239 Draft - Do Not Cite or Quote
-------
Magill. N: Sansalone. J. (2010). Distribution of particulate-bound metals for source area snow in the Lake
Tahoe watershed. J Environ Eng 136: 185-193.
http://dx.doi.org/10.1061/(ASCE)EE. 1943-7870.0000146
Maher. BA; Moore. C: Matzka. J. (2008). Spatial variation in vehicle-derived metal pollution identified
by magnetic and elemental analysis of roadside tree leaves. Atmos Environ 42: 364-373.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.09.013
Mahler. BJ; Van Metre. PC: Callender. E. (2006). Trends in metals in urban and reference lake sediments
across the United States, 1970 to 2001. Environ Toxicol Chem 25: 1698-1709.
Makkonen. U; Hellen. H; Anttila. P; Perm. M. (2010). Size distribution and chemical composition of
airborne particles in south-eastern Finland during different seasons and wildfire episodes in 2006. Sci
Total Environ 408: 644-651. http://dx.doi.Org/10.1016/i.scitotenv.2009.10.050
Marple. VA; Rubow. KL; Behm. SM. (1991). A microorifice uniform deposit impactor (MOUDI):
Description, calibration, and use. Aerosol Sci Technol 14: 434-446.
http://dx.doi.org/10.1080/02786829108959504
Martuzevicius. D; Grinshpun. SA; Reponen. T; Gorny. RL; Shukla. R; Lockev. J: Hu. S: McDonald. R;
Biswas. P; Kliucininkas. L; LeMasters. G. (2004). Spatial and temporal variations of PM2.5
concentration and composition throughout an urban area with high freeway density: The Greater
Cincinnati study. Atmos Environ 38: 1091-1105. http://dx.doi.Org/10.1016/j.atmosenv.2003.ll.015
Marx. SK; Kamber. BS: McGowan. HA. (2008). Scavenging of atmospheric trace metal pollutants by
mineral dusts: Inter-regional transport of Australian trace metal pollution to New Zealand. Atmos
Environ 42: 2460-2478. http://dx.doi.Org/10.1016/i.atmosenv.2007.12.014
McConnell. JR; Edwards. R. (2008). Coal burning leaves toxic heavy metal legacy in the Arctic. PNAS
105: 12140-12144. http://dx.doi.org/10.1073/pnas.08035641Q5
McGee. CJ: Fernandez. IJ; Norton. SA; Stubbs. CS. (2007). Cd, Ni, Pb, and Zn concentrations in forest
vegetation and soils in Maine. Water Air Soil Pollut 180: 141-153.
http://dx.doi.org/10.1007/sll270-006-9257-0
McKenzie. ER; Money. JE; Green. PG: Young. TM. (2009). Metals associated with stormwater-relevant
brake and tire samples. Sci Total Environ 407: 5855-5860.
http://dx.doi.0rg/10.1016/i.scitotenv.2009.07.018
McKenzie. ER; Wong. CM; Green. PG: Kayhanian. M; Young. TM. (2008). Size dependent elemental
composition of road-associated particles. Sci Total Environ 398: 145-153.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.02.052
Mercer. TT; Tillery. MI; Newton. GJ. (1970). A multi-stage, low flow rate cascade impactor. J Aerosol Sci
1: 9-15. http://dx.doi.org/10.1016/0021-8502(70)90024-8
MetOne (Met One Instruments Inc.). (2009). SASS/SUPER SASS speciation samplers: Specifications.
Available online at http://www.metone.com/documents/SASS0301Particulate.pdf
Mielke. HW: Gonzales. C. (2008). Mercury (Hg) and lead (Pb) in interior and exterior New Orleans
house paint films. Chemosphere 72: 882-885. http://dx.doi.Org/10.1016/i.chemosphere.2008.03.061
Mielke. HW: Gonzales. C: Powell. E; Mielke. PW. Jr. (2008). Urban soil-lead (Pb) footprint:
Retrospective comparison of public and private properties in New Orleans. Environ Geochem Health
30: 231-242. http://dx.doi.org/10.1007/sl0653-007-9111-3
Mielke. HW: Gonzales. CR; Powell. E; Jartun. M; Mielke. PW. Jr. (2007a). Nonlinear association
between soil lead and blood lead of children in metropolitan New Orleans, Louisiana: 2000-2005. Sci
Total Environ 388: 43-53. http://dx.doi.Org/10.1016/j.scitotenv.2007.08.012
Mielke. HW: Laidlaw. MA; Gonzales. C. (2010). Lead (Pb) legacy from vehicle traffic in eight California
urbanized areas: Continuing influence of lead dust on children's health [Review]. Sci Total Environ
408: 3965-3975. http://dx.doi.Org/10.1016/i.scitotenv.2010.05.017
November 2012 3-240 Draft - Do Not Cite or Quote
-------
Mielke. HW: Laidlaw. MA; Gonzales. CR. (2011c). Estimation of leaded (Pb) gasoline's continuing
material and health impacts on 90 US urbanized areas [Review]. Environ Int 37: 248-257.
http://dx.doi.0rg/10.1016/i.envint.2010.08.006
Mielke. HW: Powell. ET; Gonzales. CR: Mielke. PW. Jr. (2007b). Potential lead on play surfaces:
Evaluation of the "PLOPS" sampler as a new tool for primary lead prevention. Environ Res 103:
154-159. http://dx.doi.0rg/10.1016/i.envres.2006.08.007
Moffet. RC: de Foy. B; Molina. LT; Molina. MJ; Prather. KA. (2008a). Measurement of ambient aerosols
in northern Mexico City by single particle mass spectrometry. Atmos Chem Phys 8: 4499-4516.
http://dx.doi.org/10.5194/acp-8-4499-2008
Moffet. RC: Desvaterik. Y; Hopkins. RJ: Tivanski. AV; Gilles. MK; Wang. Y; Shutthanandan. V; Molina.
LT; Abraham. RG: Johnson. KS: Mugica. V: Molina. MJ: Laskin. A; Prather. KA. (2008b).
Characterization of aerosols containing Zn, Pb, and Cl from an industrial region of Mexico City.
Environ Sci Technol 42: 7091-7097. http://dx.doi.org/10.1021/es7030483
Molnar. P; Bellander. T; Sallsten. G: Boman. J. (2007). Indoor and outdoor concentrations of PM2.5 trace
elements at homes, preschools and schools in Stockholm, Sweden. J Environ Monit 9: 348-357.
http://dx.doi.org/10.1039/b616858b
Moreno. T; Querol. X; Pev. J: Minguillon. MC: Perez. N; Alastuev. A; Bernabe. RM; Blanco. S:
Cardenas. B; Eichinger. W: Salcido. A; Gibbons. W. (2008). Spatial and temporal variations in
inhalable CuZnPb aerosols within the Mexico City pollution plume. J Environ Monit 10: 370-378.
http://dx.doi.org/10.1039/b716507b
Murakami. M; Nakaiima. F; Furumai. H; Tomiyasu. B; Owari. M. (2007). Identification of particles
containing chromium and lead in road dust and soakaway sediment by electron probe microanalyser.
Chemosphere 67: 2000-2010. http://dx.doi.Org/10.1016/i.chemosphere.2006.ll.044
Murphy. DM; Capps. SL; Daniel. JS: Frost. GJ: White. WH. (2008). Weekly patterns of aerosol in the
United States. Atmos Chem Phys 8: 2729-2739. http://dx.doi.org/10.5194/acp-8-2729-2008
Murphy. DM; Hudson. PK; Cziczo. DJ; Gallavardin. S: Froyd. KD; Johnston. MV; Middlebrook. AM;
Reinard. MS: Thomson. DS: Thornberrv. T; Wexler. AS. (2007). Distribution of lead in single
atmospheric particles. Atmos Chem Phys 7: 3195-3210. http://dx.doi.org/10.5194/acp-7-3195-2007
N'guessan. YM; Probst. JL; Bur. T; Probst. A. (2009). Trace elements in stream bed sediments from
agricultural catchments (Gascogne region, S-W France): Where do they come from? Sci Total Environ
407: 2939-2952. http://dx.doi.Org/10.1016/i.scitotenv.2008.12.047
NAS (National Academy of Sciences). (1980). Lead in the human environment. Washington, DC:
National Academy Press.
Nicholson. KW (1988). Areview of particle resuspension [Review]. Atmos Environ 22: 2639-2651.
http://dx.doi.org/10.1016/0004-6981(88)90433-7
Nicholson. KW: Branson. JW. (1990). Factors affecting resuspension by road traffic. Sci Total Environ
93: 349-358. http://dx.doi.org/10.1016/0048-9697(90)90126-F
Nishiguchi. K; Utani. K; Fujimori. E. (2008). Real-time multielement monitoring of airborne particulate
matter using ICP-MS instrument equipped with gas converter apparatus. J Anal At Spectrom 23:
1125-1129. http://dx.doi.org/10.1039/b802302f
Noble. SR: Horstwood. MSA; Daw. P; Pashlev. V: Spiro. B; Smith. S. (2008). Evolving Pb isotope
signatures of London airborne particulate matter (PM10): Constraints from on-filter and solution-
mode MC-ICP-MS. J Environ Monit 10: 830-836. http://dx.doi.org/10.1039/b80215la
Noll. KE. (1970). A rotary inertia! impactor for sampling giant particles in the atmosphere. Atmos
Environ 4: 9-19. http://dx.doi.org/10.1016/0004-6981(70)90050-8
Norton. SA. (2007). Atmospheric metal pollutants: Archives, methods, and history. Water Air Soil Pollut
7: 93-98. http://dx.doi.org/10.1007/sll267-006-9089-3
November 2012 3-241 Draft - Do Not Cite or Quote
-------
Novak. M; Erel. Y; Zemanova. L; Bottrell. SH; Adamova. M. (2008). A comparison of lead pollution
record in sphagnum peat with known historical Pb emission rates in the British Isles and the Czech
Republic. Atmos Environ 42: 8997-9006. http://dx.doi.Org/10.1016/i.atmosenv.2008.09.031
NFS (U.S. National Park Service). (2011). WACAP database.
http://www.nature.nps.gov/air/studies/air toxics/WACAPreport.cfm
NRCS (Natural Resources Conservation Service). (2011). USDANational Water and Climate Center.
Available online at http://www.wcc.nrcs.usda.gov/ftpref/downloads/climate/windrose/ (accessed
January 11,2011).
Nriagu. JO. (1989). A global assessment of natural sources of atmospheric trace metals. Nature 338:
47-49. http://dx.doi.org/10.1038/338047aO
Obernberger. I; Brunner. T; Barnthaler. G. (2006). Chemical properties of solid biofuels—significance and
impact. Biomass and Bioenergy 30: 973-982. http://dx.doi.Org/10.1016/j.biombioe.2006.06.011
Oberts. GL. (2000). Influence of snowmelt dynamics on stormwater runoff quality. Watershed Protection
Techniques 1: 55-61.
Odigie. KO; Flegal. AR. (2011). Pyrogenic remobilization of historic industrial lead depositions. Environ
Sci Technol 45: 6290-6295. http://dx.doi.org/10.1021/es200944w
Ogulei. D; Hopke. PK; Zhou. L; Pancras. JP; Nair. N; Ondov. JM. (2006). Source apportionment of
Baltimore aerosol from combined size distribution and chemical composition data. Atmos Environ 40:
396-410. http://dx.doi.0rg/10.1016/i.atmosenv.2005.ll.075
Ondov. JM; Buckley. TJ; Hopke. PK; Ogulei. D; Parlange. MB; Rogge. WF; Squibb. KS: Johnston. MV;
Wexler. AS. (2006). Baltimore Supersite: Highly time- and size-resolved concentrations of urban
PM2.5 and its constituents for resolution of sources and immune responses. Atmos Environ 40:
224-237. http://dx.doi.0rg/10.1016/i.atmosenv.2005.ll.072
Osterberg. E; Mavewski. P; Kreutz. K; Fisher. D; Handlev. M; Sneed. S: Zdanowicz. C: Zheng. J;
Demuth. M; Waskiewicz. M; Bourgeois. J. (2008). Ice core record of rising lead pollution in the North
Pacific atmosphere. Geophys Res Lett 35: L05810. http://dx.doi.org/10.1029/2007gl032680
Pacyna. EG: Pacyna. JM: Fudala. J: Strzelecka-Jastrzab. E; Hlawiczka. S: Panasiuk. D; Nitter. S: Pregger.
T; Pfeiffer. H; Friedrich. R. (2007). Current and future emissions of selected heavy metals to the
atmosphere from anthropogenic sources in Europe. Atmos Environ 41: 8557-8566.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.07.040
Pacyna. JM: Pacyna. EG. (2001). An assessment of global and regional emissions of trace metals to the
atmosphere from anthropogenic sources worldwide. Environ Rev 9: 269-298.
http://dx.doi.org/10.1139/a01-012
Pancras. JP: Ondov. JM: Poor. N; Landis. MS: Stevens. RK. (2006). Identification of sources and
estimation of emission profiles from highly time-resolved pollutant measurements in Tampa, FL.
Atmos Environ 40: 467-481. http://dx.doi.Org/10.1016/i.atmosenv.2005.12.036
Pancras. JP: Ondov. JM: Zeisler. R. (2005). Multi-element electrothermal AAS determination of 11
marker elements in fine ambient aerosol slurry samples collected with SEAS-II. Anal Chim Acta 538:
303-312. http://dx.doi.0rg/10.1016/i.aca.2005.01.062
Paode. RD: Sofuoglu. SC: Sivadechathep. J: Noll. KB; Holsen. TM; Keeler. GJ. (1998). Dry deposition
fluxes and mass size distributions of Pb, Cu, and Zn measured in southern Lake Michigan during
AEOLOS. Environ Sci Technol 32: 1629-1635. http://dx.doi.org/10.1021/es970892b
Pardue. JH; Moe. WM; Mcinnis. D: Thibodeaux. LJ; Valsaraj. KT; Maciasz. E; van Heerden. I; Korevec.
N; Yuan. QZ. (2005). Chemical and microbiological parameters in New Orleans floodwater following
Hurricane Katrina. Environ Sci Technol 39: 8591-8599. http://dx.doi.org/10.1021/es0518631
Park. JM; Rock. JC: Wang. L; Seo. YC: Bhatnagar. A; Kim. S. (2009a). Performance evaluation of six
different aerosol samplers in aparticulate matter generation chamber. Atmos Environ 43: 280-289.
http://dx.doi.0rg/10.1016/i.atmosenv.2008.09.028
November 2012 3-242 Draft - Do Not Cite or Quote
-------
Patra. A; Colvile. R; Arnold. S: Bowen. E; Shallcross. D; Martin. D; Price. C: Tate. J: ApSimon. H;
Robins. A. (2008). On street observations of paniculate matter movement and dispersion due to traffic
on an urban road. Atmos Environ 42: 3911-3926. http://dx.doi.Org/10.1016/j.atmosenv.2006.10.070
Pedrot. M; Dia. A; Davranche. M; Bouhnik-Le Coz. M; Henin. O; Gruau. G. (2008). Insights into colloid-
mediated trace element release at the soil/water interface. J Colloid Interface Sci 325: 187-197.
http://dx.doi.0rg/10.1016/i.icis.2008.05.019
Pekev. B; Bozkurt. ZB; Pekev. H; Dogan. G: Zararsiz. A; Efe. N; Tuncel. G. (2010). Indoor/outdoor
concentrations and elemental composition of PM10/PM2.5 in urban/industrial areas of Kocaeli City,
Turkey. Indoor Air 20: 112-125. http://dx.doi.Org/10.llll/i.1600-0668.2009.00628.x
Peknev. NJ; Davidson. CI; Bein. KJ; Wexler. AS: Johnston. MV. (2006). Identification of sources of
atmospheric PM at the Pittsburgh supersite, part I: Single particle analysis and filter-based positive
matrix factorization. Atmos Environ 40: 411-423. http://dx.doi.Org/10.1016/i.atmosenv.2005.12.072
Perdrial. N; Liewig. N; Delphin. JE; Elsass. F. (2008). TEM evidence for intracellular accumulation of
lead by bacteria in subsurface environments. Chem Geol 253: 196-204.
http://dx.doi.0rg/10.1016/i.chemgeo.2008.05.008
Perrino. C: Canepari. S: Pappalardo. S: Marconi. E. (2010). Time-resolved measurements of water-
soluble ions and elements in atmospheric paniculate matter for the characterization of local and long-
range transport events. Chemosphere 80: 1291-1300.
http://dx.doi.0rg/10.1016/i.chemosphere.2010.06.050
Pingitore. NE. Jr; Clague. JW: Amava. MA; Maciejewska. B; Reynoso. JJ. (2009). Urban airborne lead:
X-ray absorption spectroscopy establishes soil as dominant source. PLoS ONE 4: e5019.
http://dx.doi.org/10.1371/iournal.pone.0005019
Planchon. FAM; van de Velde. K; Rosman. KJR; Wolff. EW: Ferrari. CP; Boutron. CF. (2003). One
hundred fifty-year record of lead isotopes in Antarctic snow from Coats Land. Geochim Cosmo Act
67: 693-708. http://dx.doi.org/10.1016/S0016-7037(02)01136-5
Point. D: Monperrus. M; Tessier. E; Amouroux. D; Chauvaud. L; Thouzeau. G: Jean. F; Amice. E; Grail.
J; Leynaert. A; Clavier. J: Donard. OFX. (2007). Biological control of trace metal and organometal
benthic fluxes in a eutrophic lagoon (Thau Lagoon, Mediterranean Sea, France). Estuar Coast Shelf
Sci 72: 457-471. http://dx.doi.Org/10.1016/j.ecss.2006.ll.013
Pokrovsky. OS: Schott. J. (2002). Iron colloids/organic matter associated transport of major and trace
elements in small boreal rivers and their estuaries (NW Russia). Chem Geol 190: 141-179.
http://dx.doi.org/10.1016/S0009-2541(02)00115-8
Pokrovsky. OS: Schott. J: Dupre. B. (2006). Trace element fractionation and transport in boreal rivers and
soil porewaters of permafrost-dominated basaltic terrain in Central Siberia. Geochim Cosmo Act 70:
3239-3260. http://dx.doi.Org/10.1016/i.gca.2006.04.008
Pokrovsky. OS: Viers. J: Shirokova. LS: Shevchenko. VP; Filipov. AS: Dupre. B. (2010). Dissolved,
suspended, and colloidal fluxes of organic carbon, major and trace elements in the Severnaya Dvina
River and its tributary. Chem Geol 273: 136-149. http://dx.doi.Org/10.1016/j.chemgeo.2010.02.018
Polissar. AV; Hopke. PK; Poirot. RL. (2001). Atmospheric aerosol over Vermont: chemical composition
and sources. Environ Sci Technol 35: 4604-4621.
Polissar. AV: Paatero. P; Hopke. PK; Malm. WC: Sisler. JF. (1998). Atmospheric aerosol over Alaska: 2.
Elemental composition and sources. J Geophys Res 103: 19045-19057.
Poperechna. N; Heumann. KG. (2005). Species-specific GC/ICP-IDMS for trimethyllead determinations
in biological and environmental samples. Anal Chem 77: 511-516.
http://dx.doi.org/10.1021/ac048757m
Popp. M; Hann. S: Koellensperger. G. (2010). Environmental application of elemental speciation analysis
based on liquid or gas chromatography hyphenated to inductively coupled plasma mass spectrometry—
a review. Anal Chim Acta 668: 114-129. http://dx.doi.Org/10.1016/i.aca.2010.04.036
November 2012 3-243 Draft - Do Not Cite or Quote
-------
Potts. PJ: Webb. PC. (1992). X-ray fluorescence spectrometry. J Geochem Explor 44: 251-296.
http://dx.doi.org/10.1016/0375-6742(92)90052-A
Poykio. R; Nurmesniemi. H; Dahl. O. (2007). Concentrations of nutrients and heavy metals in cyclone fly
ash from the grate-fired boiler at a small municipal district heating plant (6 MW). J Resid Sci Tech 4:
127-136.
Prather. KA; Nordmever. T; Salt. K. (1994). Real-time characterization of individual aerosol particles
using time-of-flight mass spectrometry. Anal Chem 66: 1403-1407.
http://dx.doi.org/10.1021/ac00081a007
Pratt. C: Lottermoser. BG. (2007). Mobilisation of traffic-derived trace metals from road corridors into
coastal stream and estuarine sediments, Cairns, northern Australia. Environ Geol 52: 437-448.
http://dx.doi.org/10.1007/s00254-006-0471-2
Preciado. HF; Li. LY. (2006). Evaluation of metal loadings and bioavailability in air, water and soil along
two highways of British Columbia, Canada. Water Air Soil Pollut 172: 81-108.
http://dx.doi.org/10.1007/sll270-005-9063-0
Presley. SM; Abel. MT; Austin. GP; Rainwater. TR; Brown. RW: Mcdaniel. LN; Marsland. EJ;
Fornerette. AM; Dillard. ML; Rigdon. RW: Kendall. RJ: Cobb. GP. (2010). Metal concentrations in
schoolyard soils from New Orleans, Louisiana before and after hurricanes Katrina and Rita.
Chemosphere 80: 67-73. http://dx.doi.Org/10.1016/i.chemosphere.2010.03.031
Presley. SM; Rainwater. TR; Austin. GP: Platt. SG: Zak. JC: Cobb. GP: Marsland. EJ: Tian. K; Zhang. B;
Anderson. TA; Cox. SB; Abel. MT; Leftwich. BD; Huddleston. JR; Jeter. RM; Kendall. RJ. (2006).
Assessment of pathogens and toxicants in New Orleans, LA following hurricane Katrina. Environ Sci
Technol 40: 468-474. http://dx.doi.org/10.1021/es052219p
Prospero. JM. (1999). Assessing the impact of advected African dust on air quality and health in the
United States. Hum Ecol Risk Assess 5: 471-479.
Pruvot C: Douav. F; Herve. F; Waterlot. C. (2006). Heavy metals in soil, crops and grass as a source of
human exposure in the former mining areas. Journal of Soils and Sediments 6: 215-220.
http://dx.doi.org/10.1065/iss2006.10.186
Pyrzynska. K. (1996). Organolead speciation in environmental samples: A review [Review]. Mikrochim
Acta 122: 279-293. http://dx.doi.org/10.1007/BF01245788
Qi. J: Feng. L; Li. X; Zhang. M. (2006). An X-ray photoelectron spectroscopy study of elements on the
surface of aerosol particles. J Aerosol Sci 37: 218-227.
http://dx.doi.0rg/10.1016/i.jaerosci.2005.04.005
Quevauviller. P. (2000). Accuracy and traceability in environmental monitoringpitfalls in methylmercury
determinations as a case study. J Environ Monit 2: 292-299. http://dx.doi.org/10.1039/b001852j
Quinton. JN; Catt. JA. (2007). Enrichment of heavy metals in sediment resulting from soil erosion on
agricultural fields. Environ Sci Technol 41: 3495-3500. http://dx.doi.org/10.1021/es062147h
Qureshi. S: Dutkiewicz. VA; Khan. AR; Swami. K; Yang. KX; Husain. L; Schwab. JJ: Demerjian. KL.
(2006). Elemental composition of PM2.5 aerosols in Queens, New York: Solubility and temporal
trends. Atmos Environ 40: S238-S251. http://dx.doi.Org/10.1016/i.atmosenv.2005.12.070
Raabe. OG: Braaten. DA; Axelbaum. RL; Teague. SV: Cahill. TA. (1988). Calibration studies of the drum
impactor. JAerosol Sci 19: 183-195. http://dx.doi.org/10.1016/0021-8502(88)90222-4
Rabinowitz. MB. (1986). Stable isotopes of ambient environmental lead in Boston, Massachusetts.
(Technical Report 1-28). Washington, DC: U.S. Environmental Protection Agency.
Radakovitch. O; Roussiez. V; Ollivier. P; Ludwig. W: Grenz. C: Probst. JL. (2008). Input of particulate
heavy metals from rivers and associated sedimentary deposits on the Gulf of Lion continental shelf.
Estuar Coast Shelf Sci 77: 285-295. http://dx.doi.Org/10.1016/i.ecss.2007.09.028
November 2012 3-244 Draft - Do Not Cite or Quote
-------
Rauch. JN; Pacyna. JM. (2009). Earth's global Ag, Al, Cr, Cu, Fe, Ni, Pb, and Zn cycles. Global
Biogeochem Cycles 23: GB2001. http://dx.doi.org/10.1029/2008gb003376
Reinard. MS: Adoua. K; Martini. JM; Johnston. MV. (2007). Source characterization and identification by
real-time single particle mass spectrometry. Atmos Environ 41: 9397-9409.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.09.001
Reuer. MK; Weiss. DJ. (2002). Anthropogenic lead dynamics in the terrestrial and marine environment.
Proc Biol Sci 360: 2889-2904.
Rice. J. (2007). Memorandum: Summary of method detection limits for ambient lead methods. Research
Triangle Park, NC: United States Environmental Protection Agency.
Richter. P; Grino. P; Ahumada. I; Giordano. A. (2007). Total element concentration and chemical
fractionation in airborne particulate matter from Santiago, Chile. Atmos Environ 41: 6729-6738.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.04.053
Riediker. M; Williams. R; Devlin. R; Griggs. T; Bromberg. P. (2003). Exposure to particulate matter,
volatile organic compounds, and other air pollutants inside patrol cars. Environ Sci Technol 37:
2084-2093. http://dx.doi.org/10.1021/es026264y
Rodes. CE; Evans. EG. (1985). Preliminary assessment of 10 "mu"m particulate sampling at eight
locations in the United States. Atmos Environ 19: 293-303.
http://dx.doi.org/10.1016/0004-6981(85)90097-6
Roonev. CP; McLaren. RG: Condron. LM. (2007). Control of lead solubility in soil contaminated with
lead shot: Effect of soil pH. Environ Pollut 149: 149-157.
http://dx.doi.0rg/10.1016/i.envpol.2007.01.009
Root. RA. (2000). Lead loading of urban streets by motor vehicle wheel weights. Environ Health Perspect
108: 937-940. http://dx.doi.org/10.1289/ehp.00108937
Ross. JM; Sherrell. PJVI. (1999). The role of colloids in tracemetal transport and adsorption behavior in
New Jersey. Limnol Oceanogr44: 1019-1034.
Rothwell. JJ: Evans. MG: Allott. TEH. (2007a). Lead contamination of fluvial sediments in an eroding
blanket peat catchment. Appl Geochem 22: 446-459.
http://dx.doi.0rg/10.1016/i.apgeochem.2006.ll.002
Rothwell. JJ: Evans. MG: Allott. TEH. (2008). In-stream processing of sediment-associated metals in
peatland fluvial systems. Water Air Soil Pollut 187: 53-64.
http://dx.doi.org/10.1007/sll270-007-9496-8
Rothwell. JJ: Evans. MG: Daniels. SM; Allott. TEH. (2007b). Baseflow and stormflow metal
concentrations in streams draining contaminated peat moorlands in the Peak District National Park
(U.K.). JHydrol341: 90-104. http://dx.doi.Org/10.1016/i.ihvdrol.2007.05.004
Rothwell. JJ: Lindsay. JB; Evans. MG: Allott. TEH. (2010). Modelling suspended sediment lead
concentrations in contaminated peatland catchments using digital terrain analysis. Ecol Eng 36:
623-630. http://dx.doi.0rg/10.1016/i.ecoleng.2008.10.010
Roulier. JL; Belaud. S: Coquery. M. (2010). Comparison of dynamic mobilization of Co, Cd and Pb in
sediments using DGT and metal mobility assessed by sequential extraction. Chemosphere 79:
839-843. http://dx.doi.0rg/10.1016/i.chemosphere.2010.02.056
Roulier. S: Robinson. B; Kuster. E; Schulin. R. (2008b). Analysing the preferential transport of lead in a
vegetated roadside soil using lysimeter experiments and a dual-porosity model. Eur J Soil Sci 59:
61-70. http://dx.doi.0rg/10.llll/i.1365-2389.2007.00954.x
Sabin. LD; Lim. JH; Stolzenbach. KD; Schiff. KG. (2006a). Atmospheric dry deposition of trace metals in
the coastal region of Los Angeles, California, USA. Environ Toxicol Chem 25: 2334-2341.
http://dx.doi.Org/10.1897/05-300R.l
November 2012 3-245 Draft - Do Not Cite or Quote
-------
Sabin. LD; Lim. JH; Venezia. MT; Winer. AM; Schiff. KG: Stolzenbach. KD. (2006b). Dry deposition
and resuspension of particle-associated metals near a freeway in Los Angeles. Atmos Environ 40:
7528-7538. http://dx.doi.Org/10.1016/i.atmosenv.2006.07.004
Sabin. LD; Schiff. KG. (2008). Dry atmospheric deposition rates of metals along a coastal transect in
southern California. Atmos Environ 42: 6606-6613. http://dx.doi.Org/10.1016/i.atmosenv.2008.04.042
Sadiq. M. (1992). Lead in marine environments. In Toxic metal chemistry in marine environments. New
York, NY: Marcel Dekker, Inc.
Sakata. M; Asakura. K. (2008). Evaluating relative contribution of atmospheric mercury species to
mercury dry deposition in Japan. Water Air Soil Pollut 193: 51-63.
http://dx.doi.org/10.1007/sll270-008-9667-2
Salcedo. D; Onasch. TB; Aiken. AC: Williams. LR; de Foy. B; Cubison. MJ; Worsnop. PR; Molina. LT;
Jimenez. JL. (2010). Determination of particulate lead using aerosol mass spectrometry:
MILAGRO/MCMA-2006 observations. Atmos Chem Phys 10: 5371-5389.
http://dx.doi.org/10.5194/acp-10-5371-2010
Sammut. ML; Noack. Y; Rose. J: Hazemann. JL; Proux. O; Depoux. M; Ziebel. A; Fiani. E. (2010).
Speciation of Cd and Pb in dust emitted from sinter plant. Chemosphere 78: 445-450.
http://dx.doi.0rg/10.1016/i.chemosphere.2009.10.039
Sanchez-Ccovllo. OR; Ynoue. RY; Martins. LD: Astolfo. R; Miranda. RM; Freitas. ED; Borges. AS:
Fornaro. A; Freitas. H; Moreira. A; Andrade. MF. (2009). Vehicular particulate matter emissions in
road tunnels in Sao Paulo, Brazil. Environ Monit Assess 149: 241-249.
http://dx.doi.org/10.1007/sl0661-008-0198-5
Sansalone. J: Ying. G: Lin. H. (2010). Distribution of metals for particulate matter transported in source
area rainfall-runoff. J Environ Eng 136: 172-184.
http://dx.doi.org/10.1061/(ASCE)EE. 1943-7870.0000139
Sansalone. JJ: Buchberger. SG. (1996). Characterization of metals and solids in urban highway winter
snow and spring rainfall-runoff. Trans Res Rec 1523: 147-159. http://dx.doi.org/10.3141/1523-18
Sato. K; Tamura. T; Furuta. N. (2008). Partitioning between soluble and insoluble fractions of major and
trace elements in size-classified airborne particulate matter collected in Tokyo. J Environ Monit 10:
211-218. http://dx.doi.org/10.1039/B709937A
Schauer. JJ: Lough. GC: Shafer. MM; Christensen. WF; Arndt. MF; DeMinter. JT; Park. JS. (2006).
Characterization of metals emitted from motor vehicles. (133). Boston, MA: Health Effects Institute.
http://pubs.healtheffects.org/view.php?id=150
Scheetz. CD; Rimstidt. JD. (2009). Dissolution, transport, and fate of lead on a shooting range in the
Jefferson National Forest near Blacksburg, VA, USA. Environ Geol 58: 655-665.
http://dx.doi.org/10.1007/s00254-008-1540-5
Scheid. S: Gunthardt-Goerg. MS: Schulin. R; Nowack. B. (2009). Accumulation and solubility of metals
during leaf litter decomposition in non-polluted and polluted soil. Eur J Soil Sci 60: 613-621.
http://dx.doi.0rg/10.llll/i.1365-2389.2009.01153.x
Schleicher. N; Norra. S: Chai. F; Chen. Y; Wang. S: Stuben. D. (2010). Anthropogenic versus geogenic
contribution to total suspended atmospheric particulate matter and its variations during a two-year
sampling period in Beijing, China. J Environ Monit 12: 434-441. http://dx.doi.org/10.1039/b914739j
Schroth. AW: Bostick. BC: Kaste. JM; Friedland. AJ. (2008). Lead sequestration and species
redistribution during soil organic matter decomposition. Environ Sci Technol 42: 3627-3633.
http://dx.doi.org/10.1021/es703002b
Schulz-Zunkel. C: Krueger. F. (2009). Trace metal dynamics in floodplain soils of the river Elbe: a review
[Review]. J Environ Qual 38: 1349-1362. http://dx.doi.org/10.2134/iea2008.0299
November 2012 3-246 Draft - Do Not Cite or Quote
-------
Schuwirth. N; Voegelin. A; Kretzschmar. R; Hofmann. T. (2007). Vertical distribution and speciation of
trace metals in weathering flotation residues of a zinc/lead sulfide mine. J Environ Qua! 36: 61-69.
http://dx.doi.org/10.2134/ieq2006.0148
Schwab. AP; Zhu. DS: Banks. MK. (2008). Influence of organic acids on the transport of heavy metals in
soil. Chemosphere 72: 986-994. http://dx.doi.Org/10.1016/i.chemosphere.2008.02.047
Schwab. KJ; Gibson. KE; Williams. PL; Kulbicki. KM; Lo. CP; Mihalic. JN; Brevsse. PN; Curriero. FC:
Geyh. AS. (2007). Microbial and chemical assessment of regions within New Orleans, LA impacted
by hurricane Katrina. Environ Sci Technol 41: 2401-2406. http://dx.doi.org/10.1021/es062916x
Sedefian. L; Rao. ST; Czapski. U. (1981). Effects of traffic-generated turbulence on near-field dispersion.
Atmos Environ 15: 527-536. http://dx.doi.org/10.1016/0004-6981(81)90182-7
Sengor. SS: Spvcher. NF; Ginn. TR; Sani. RK; Peyton. B. (2007). Biogeochemical reactive-diffusive
transport of heavy metals in Lake Coeur d'Alene sediments. Appl Geochem 22: 2569-2594.
http://dx.doi.0rg/10.1016/i.apgeochem.2007.06.011
Shacklette. HT; Boerngen. JG. (1984). Element concentrations in soils and other surficial materials of the
conterminous United States. (1270). Washington D.C.: Government Printing Office.
http://pubs.usgs.gov/pp/1270/
Shi. H; Witt. E. Ill; Shu. S: Su. T; Wang. J; Adams. C. (2010). Toxic trace element assessment for
soils/sediments deposited during hurricanes Katrina and Rita from southern Louisiana, USA: A
sequential extraction analysis. Environ Toxicol Chem 29: 1419-1428.
http://dx.doi.org/10.1002/etc.218
Shotbolt. LA; Rothwell. JJ: Lawlor. AJ. (2008). Amass balance approach to quantifying Pb storage and
fluxes in an upland catchment of the Peak District, north-central England. Earth Surface Processes and
Landforms33: 1721-1741. http://dx.doi.org/10.1002/esp.1644
Shotyk. W: Krachler. M. (2007). Lead in bottled waters: Contamination from glass and comparison with
pristine groundwater. Environ Sci Technol 41: 3508-3513. http://dx.doi.org/10.1021/es062964h
Shotyk. W: Krachler. M. (2009). Determination of trace element concentrations in natural freshwaters:
How low is "low," and how low do we need to go? J Environ Monit 11: 1747-1753.
http://dx.doi.org/10.1039/b917090c
Shotvk. W: Krachler. M. (2010). The isotopic evolution of atmospheric Pb in central Ontario since AD
1800, and its impacts on the soils, waters, and sediments of a forested watershed, Kawagama Lake.
Geochim Cosmo Act 74: 1963-1981. http://dx.doi.Org/10.1016/j.gca.2010.01.009
Shotvk. W: Krachler. M; Aeschbach-Hertig. W: Hillier. S: Zheng. JC. (2010). Trace elements in recent
groundwater of an artesian flow system and comparison with snow: Enrichments, depletions, and
chemical evolution of the water. J Environ Monit 12: 208-217. http://dx.doi.org/10.1039/B909723F
Shotvk. W: Le Roux. G. (2005). Biogeochemistry and cycling of lead [Review]. In A Sigel; H Sigel;
RKO Sigel (Eds.), Biogeochemical cycles of elements (pp. 239-275). Boca Raton, FL: Taylor &
Francis. http://dx.doi.org/10.1201/9780824751999.chlO
Shotvk. W: Weiss. D; Heisterkamp. M; Cheburkin. AK; Applebv. PG: Adams. FC. (2002). New peat bog
record of atmospheric lead pollution in Switzerland: Pb concentrations, enrichment factors, isotopic
composition, and organolead species. Environ Sci Technol 36: 3893-3900.
http://dx.doi.org/10.1021/es010196i
Sillanpaa. M; Saarikoski. S: Hillamo. R; Pennanen. A; Makkonen. U; Spolnik. Z; Van Grieken. R;
Koskentalo. TA; Salonen. RO. (2005). Chemical composition, mass size distribution and source
analysis of long-range transported wildfire smokes in Helsinki. Sci Total Environ 350: 119-135.
http://dx.doi.0rg/10.1016/i.scitotenv.2005.01.024
Silva. PJ: Prather. KA. (1997). On-line characterization of individual particles from automobile
emissions. Environ Sci Technol 31: 3074-3080. http://dx.doi.org/10.1021/es961063d
November 2012 3-247 Draft - Do Not Cite or Quote
-------
Singh. M; Jaques. PA; Sioutas. C. (2002). Size distribution and diurnal characteristics of particle-bound
metals in source and receptor sites of the Los Angeles Basin. Atmos Environ 36: 1675-1689.
http://dx.doi.org/10.1016/81352-2310(02)00166-8
Smichowski. P; Polla. G: Gomez. D. (2005). Metal fractionation of atmospheric aerosols via sequential
chemical extraction: a review [Review]. Anal Bioanal Chem 381: 302-316.
http://dx.doi.org/10.1007/s00216-004-2849-x
Smichowski. P; Polla. G: Gomez. D; Espinosa. AJF; Lopez. AC. (2008). A three-step metal fractionation
scheme for fly ashes collected in an Argentine thermal power plant. Fuel 87: 1249-1258.
http://dx.doi.0rg/10.1016/i.fuel.2007.07.011
Smith. GR. (2001). Minerals yearbook 1999: lead. Washington, DC: U.S. Department of the Interior, U.S.
Geological Survey.
Snyder. DC: Schauer. JJ; Gross. DS: Turner. JR. (2009). Estimating the contribution of point sources to
atmospheric metals using single-particle mass spectrometry. Atmos Environ 43: 4033-4042.
http://dx.doi.0rg/10.1016/i.atmosenv.2009.05.011
Sobanska. S: Ricq. N; Laboudigue. A; Guillermo. R; Bremard. C: Laureyns. J: Merlin. JC: Wignacourt
JP (1999). Microchemical investigations of dust emitted by a lead smelter. Environ Sci Technol 33:
1334-1339. http://dx.doi.org/10.1021/es9805270
Solomon. P; Derrick. M; Movers. J: Hyde. P. (1982). Performance Comparison of Three Samplers of
Suspended Airborne Paniculate Matter. J Air Pollut Control Assoc 32: 373-375.
http://dx.doi.org/10.1080/00022470.1982.10465411
Song. F; Gao. Y. (2011). Size distributions of trace elements associated with ambient particular matter in
the affinity of a major highway in the New JerseyNew York metropolitan area. Atmos Environ 45:
6714-6723. http://dx.doi.Org/10.1016/i.atmosenv.2011.08.031
Song. XH; Polissar. AV; Hopke. PK. (2001). Sources of fine particle composition in the northeastern US.
Atmos Environ 35: 5277-5286.
Soto-Jimenez. MF; Flegal. AR. (2009). Origin of lead in the Gulf of California Ecoregion using stable
isotope analysis. J Geochem Explor 101: 209-217. http://dx.doi.Org/10.1016/i.gexplo.2008.07.003
Soto-Jimenez. MF; Paez-Osuna. F. (2010). A first approach to study the mobility and behavior of lead in
hypersaline salt marsh sediments: Diffusive and advective fluxes, geochemical partitioning and Pb
isotopes. J Geochem Explor 104: 87-96. http://dx.doi.Org/10.1016/i.gexplo.2009.12.006
Soulhac. L; Garbero. V; Salizzoni. P; Mejean. P; Perkins. RJ. (2009). Flow and dispersion in street
intersections. Atmos Environ 43: 2981-2996. http://dx.doi.Org/10.1016/j.atmosenv.2009.02.061
Spalinger. SM; von Braun. MC: Petrosvan. V; von Lindern. IH. (2007). Northern Idaho house dust and
soil lead levels compared to the Bunker Hill superfund site. Environ Monit Assess 130: 57-72.
http://dx.doi.org/10.1007/sl0661-006-9450-z
Steinnes. E; Sjobakk. TE; Donisa. C: Brannvall. ML. (2005). Quantification of pollutant lead in forest
soils. Soil Sci Soc Am J 69: 1399-1404. http://dx.doi.org/10.2136/sssaj2004.0095
Stolpe. B; Hassellov. M. (2007). Changes in size distribution of fresh water nanoscale colloidal matter
and associated elements on mixing with seawater. Geochim Cosmo Act 71: 3292-3301.
http://dx.doi.0rg/10.1016/i.gca.2007.04.025
Sturm. A; Crowe. SA; Fowle. DA. (2008). Trace lead impacts biomineralization pathways during
bacterial iron reduction. Chem Geol 249: 282-293. http://dx.doi.Org/10.1016/i.chemgeo.2008.01.017
Swarbrick. JC: Skyllberg. U; Karlsson. T; Glatzel. P. (2009). High energy resolution X-ray absorption
spectroscopy of environmentally relevant lead(II) compounds. Inorg Chem 48: 10748-10756.
http://dx.doi.org/10.1021/ic9015299
Sweet. CW: Gatz. DF. (1998). Summary and analysis of available PM25 measurements in Illinois. Atmos
Environ 32: 1129-1133.
November 2012 3-248 Draft - Do Not Cite or Quote
-------
Sweet. CW: Weiss. A; Vermette. SJ. (1998). Atmospheric deposition of trace metals at three sites near the
Great Lakes. Water Air Soil Pollut 103: 423-439. http://dx.doi.Org/10.1023/A:1004905832617
Taillefert. M; Lienemann. CP; Gaillard. JF; Ferret. D. (2000). Speciation, reactivity, and cycling of Fe and
Pb in a meromictic lake. Geochim Cosmo Act 64: 169-183.
http://dx.doi.org/10.1016/S0016-7037(99)00285-9
Takamatsu. T; Watanabe. M; Koshikawa. MK; Murata. T; Yamamura. S: Havashi. S. (2010). Pollution of
montane soil with Cu, Zn, As, Sb, Pb, and nitrate in Kanto, Japan. Sci Total Environ 408: 1932-1942.
http://dx.doi.0rg/10.1016/i.scitotenv.2010.01.016
Tan. MG: Zhang. GL; Li. XL; Zhang. YX; Yue. WS: Chen. JM; Wang. YS: Li. AG: Li. Y; Zhang. YM;
Shan. ZC. (2006). Comprehensive study of lead pollution in Shanghai by multiple techniques. Anal
Chem 78: 8044-8050. http://dx.doi.org/10.1021/ac061365q
Taylor. MP; Mackav. AK; Hudson-Edwards. KA; Holz. E. (2010). Soil Cd, Cu, Pb and Zn contaminants
around Mount Isa city, Queensland, Australia: Potential sources and risks to human health. Appl
Geochem 25: 841-855. http://dx.doi.Org/10.1016/j.apgeochem.2010.03.003
Tessier. A; Campbell. PGC: Bisson. M. (1979). Sequential extraction procedure for the speciation of
paniculate trace-metals. Anal Chem 51: 844-851. http://dx.doi.org/10.1021/ac50043a017
Theodosi. C: Markaki. Z; Tselepides. A; Mihalopoulos. N. (2010). The significance of atmospheric inputs
of soluble and paniculate major and trace metals to the eastern Mediterranean seawater. Mar Chem
120: 154-163. http://dx.doi.0rg/10.1016/i.marchem.2010.02.003
Thurston. GD; Ito. K; Lall. R. (2011). A source apportionment of U.S. fine paniculate matter air pollution.
Atmos Environ 45: 3924-3936. http://dx.doi.Org/10.1016/i.atmosenv.2011.04.070
Tipping. E; Lawlor. AJ; Lofts. S: Shotbolt. L. (2006). Simulating the long-term chemistry of an upland
U.K. catchment: Heavy metals. Environ Pollut 141: 139-150.
http://dx.doi.0rg/10.1016/i.envpol.2005.08.019
Tuccillo. ME. (2006). Size fractionation of metals in runoff from residential and highway storm sewers.
Sci Total Environ 355: 288-300. http://dx.doi.Org/10.1016/j.scitotenv.2005.03.003
U.S. EPA (U.S. Environmental Protection Agency). (1977). Air quality criteria for lead [EPA Report].
(EPA-600/8-77-017). Washington, D.C.
http://nepis.epa.gov/Exe/ZvPURL.cgi?Dockev=20013GWR.txt
U.S. EPA (U.S. Environmental Protection Agency). (1986a). Air quality criteria for lead [EPA Report].
(EPA/600/8-83/028aF-dF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
U.S. EPA (U.S. Environmental Protection Agency). (1986b). Air quality criteria for lead: Volume I of IV
[EPA Report]. (EPA-600/8-83/028aF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
U.S. EPA (U.S. Environmental Protection Agency). (1999). Particulate matter (PM2.5) speciation
guidance: Final draft (Edition 1) [EPA Report]. Research Triangle Park, NC.
http://www.epa.gov/ttn/amtic/files/ambient/pm25/spec/specpln2.pdf
U.S. EPA (U.S. Environmental Protection Agency). (2003b). Guidance for developing ecological soil
screening levels (Eco-SSLs): Review of background concentration for metals - Attachment 1-4 [EPA
Report]. (OSWER Directive 92857-55). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2004). Air quality criteria for paniculate matter [EPA
Report]. (EPA/600/P-99/002aF-bF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=87903
U.S. EPA (U.S. Environmental Protection Agency). (2006a). 2002 National Emissions Inventory data and
documentation [EPA Report]. Research Triangle Park, NC.
http://www.epa.gov/ttnchiel/net/2002inventory.html
November 2012 3-249 Draft - Do Not Cite or Quote
-------
U.S. EPA (U.S. Environmental Protection Agency). (2006b). Air quality criteria for lead: Volume I of II
[EPA Report]. (EPA/600/R-05/144aF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/CFM/recordisplav.cfm?deid=158823
U.S. EPA (U.S. Environmental Protection Agency). (2007d). Guidance for developing ecological soil
screening levels [EPA Report]. (OSWER Directive 92857-55). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2007h). Review of the national ambient air quality
standards for lead: Policy assessment of scientific and technical Information: OAQPS staff paper [EPA
Report]. (EPA-452/R-07-013). Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2008a). 2005 National Emissions Inventory data and
documentation. Available online at http://www.epa.gov/ttn/chief/net/2005inventory.html (accessed
November 1, 2010).
U.S. EPA (U.S. Environmental Protection Agency). (2008b). Ambient aquatic life water quality: Lead
(draft) [EPA Report]. (EPA Contract No. 68-C-04-006). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2008c). California area designations for the 2008
lead National Ambient Air Quality Standards [EPA Report]. Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2008d). Florida area designations for the 2008 lead
National Ambient Air Quality Standards [EPA Report]. Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2008f). Missouri area designations for the 2008 lead
National Ambient Air Quality Standards [EPA Report]. Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2008g). Ohio area designations for the 2008 lead
National Ambient Air Quality Standards [EPA Report]. Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2008h). Tennessee area designations for the 2008
lead National Ambient Air Quality Standards [EPA Report]. Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2009a). Integrated science assessment for paniculate
matter [EPA Report]. (EPA/600/R-08/139F). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=216546
U.S. EPA (U.S. Environmental Protection Agency). (2010). Integrated science assessment for carbon
monoxide [EPA Report]. (EPA/600/R-09/019F). Research Triangle Park, NC.
http ://cfpub. epa. gov/ncea/cfm/recordisplav. cfm?deid=218686
U.S. EPA (U.S. Environmental Protection Agency). (2011a). 2008 National Emissions Inventory data and
documentation. Available online at http://www.epa.gov/ttn/chief/net/2008inventory.html (accessed
February 17,2011).
U.S. EPA (U.S. Environmental Protection Agency). (2011b). Emission factor supporting documentation
for the final mercury and air toxics standards [EPA Report]. (EPA-454/R-11-012). Research Triangle
Park, NC. http://www.epa.gov/ttn/atw/utilitv/mats efs casestudies currentbaseei.pdf
U.S. EPA (U.S. Environmental Protection Agency). (20111). List of designated reference and equivalent
methods [EPA Report]. Research Triangle Park, NC.
http://www.epa.gov/ttn/amtic/Files/ambient/criteria/reference-equivalent-methods-list.pdf
UC (University of California). (1995). IMPROVE data guide. Davis, CA: University of California Davis.
USGS (U.S. Geological Survey). (1996). Mineral commodities summary: Lead. In Mineral commodity
summaries 1996 (pp. 92-93). U.S. Department of the Interior.
http://minerals.usgs.gov/minerals/pubs/commoditv/lead/lead mcs96.pdf
USGS (U.S. Geological Survey). (2005). Historical statistics for mineral and material commodities in the
United States. Available online at http://minerals.usgs.gov/ds/2005/140/ (accessed May 10, 2010).
November 2012 3-250 Draft - Do Not Cite or Quote
-------
USGS (U.S. Geological Survey). (2006). Mineral commodities summary: Lead. In Mineral commodity
summaries 2006 (pp. 96-97). U.S. Department of the Interior.
http://minerals.usgs.gov/minerals/pubs/commoditv/lead/lead mcs06.pdf
USGS (U.S. Geological Survey). (2012). 2010 minerals yearbook: Lead [advance release]. Washington,
DC: U.S. Department of the Interior.
Uzu. G: Sobanska. S: Aliouane. Y; Pradere. P; Dumat. C. (2009). Study of lead phytoavailability for
atmospheric industrial micronic and sub-micronic particles in relation with lead speciation. Environ
Pollut 157: 1178-1185. http://dx.doi.Org/10.1016/i.envpol.2008.09.053
van Herwijnen. R; Hutchings. TR; Al-Tabbaa. A; Moffat AJ; Johns. ML; Ouki. SK. (2007). Remediation
of metal contaminated soil with mineral-amended composts. Environ Pollut 150: 347-354.
http://dx.doi.0rg/10.1016/i.envpol.2007.01.023
van Lith. SC: Alonso-Ramirez. V; Jensen. PA; Frandsen. FJ; Glarborg. P. (2006). Release to the gas phase
of inorganic elements during wood combustion: Part 1: Development and evaluation of quantification
methods. Energy Fuels 20: 964-978. http://dx.doi.org/10.1021/ef050131r
van Lith. SC: Jensen. PA; Frandsen. FJ: Glarborg. P. (2008). Release to the gas phase of inorganic
elements during wood combustion: Part 2. Influence of fuel composition. Energy Fuels 22:
1598-1609. http://dx.doi.org/10.1021/ef060613i
Van Metre. PC: Mahler. BJ; Wilson. JT; Callender. E. (2006). Major and Trace Elements in 35 Lake and
Reservoir Sediment Cores From Across the United States, 19942001. (Data Series 166). Reston, VA:
U.S. Geological Survey. http://www.ntis.gov/search/product.aspx?ABBR=PB2006113440
Vermette. SJ: Irvine. KN; Drake. JJ. (1991). Temporal variability of the elemental composition in urban
street dust. Environ Monit Assess 18: 69-77. http://dx.doi.org/10.1007/BF00394478
Verstraete. S: Van Meirvenne. M. (2008). A multi-stage sampling strategy for the delineation of soil
pollution in a contaminated brownfield. Environ Pollut 154: 184-191.
http://dx.doi.0rg/10.1016/i.envpol.2007.10.014
Vogl. J: Heumann. KG. (1997). Determination of heavy metal complexes with humic substances by
HPLC/ICP-MS coupling using on-line isotope dilution technique. Fresenius' Journal of Analytical
Chemistry 359: 438-441. http://dx.doi.org/10.1007/s002160050606
Voutsa. D; Samara. C. (2002). Labile and bioaccessible fractions of heavy metals in the airborne
paniculate matter from urban and industrial areas. Atmos Environ 36: 3583-3590.
http://dx.doi.org/10.1016/81352-2310(02)00282-0
Vucetich. JA; Outridge. PM; Peterson. RO; Eide. R; Isrenn. R. (2009). Mercury, lead and lead isotope
ratios in the teeth of moose (Alces alces) from Isle Royale, US Upper Midwest, from 1952 to 2002. J
Environ Monit 11: 1352-1359. http://dx.doi.org/10.1039/b903417j
Wagner. J: Leith. D. (2001). Field tests of a passive aerosol sampler. J Aerosol Sci 32: 33-48.
http://dx.doi.org/10.1016/S0021-8502(00)00055-0
Waheed. A; Zhang. YX; Bao. LM; Cao. QC: Zhang. GL; Li. Y; Li. XL. (2010). Study of seasonal
variation and source characteristic of PM10 of Shanghai urban atmosphere using PIXE. Journal of
Radioanal Chem 283: 427-432. http://dx.doi.org/10.1007/sl0967-009-0390-2
Wahlin. P; Berkowicz. R; Palmgren. F. (2006). Characterisation of traffic-generated paniculate matter in
Copenhagen. Atmos Environ 40: 2151-2159. http://dx.doi.Org/10.1016/i.atmosenv.2005.ll.049
Walder. AJ: Freedman. PA. (1992). Isotopic ratio measurement using a double focusing magnetic sector
mass analyser with an inductively coupled plasma as an ion source. J Anal At Spectrom 7: 571-575.
http://dx.doi.org/10.1039/JA9920700571
Wang. G: Huang. L; Gao. S: Gao. S: Wang. L. (2002). Characterization of water-soluble species of PM10
and PM2.5 aerosols in urban area in Nanjing, China. Atmos Environ 36: 1299-1307.
http://dx.doi.org/10.1016/81352-2310(01)00550-7
November 2012 3-251 Draft - Do Not Cite or Quote
-------
Wang. LJ; Wanjura. JD; Parnell. CB; Lacev. RE; Shaw. BW. (2005b). Performance characteristics of a
low-volume PM10 sampler. Trans ASAE 48: 739-748.
Wang. W: Liu. XD; Zhao. LW: Quo. DF; Tian. XD; Adams. F. (2006c). Effectiveness of leaded petrol
phase-out in Tianjin, China based on the aerosol lead concentration and isotope abundance ratio. Sci
Total Environ 364: 175-187. http://dx.doi.Org/10.1016/i.scitotenv.2005.07.002
Wang. XL; Sato. T; Xing. BS. (2006d). Size distribution and anthropogenic sources apportionment of
airborne trace metals in Kanazawa, Japan. Chemosphere 65: 2440-2448.
http://dx.doi.0rg/10.1016/i.chemosphere.2006.04.050
Wanjura. JD: Parnell. CB. Jr; Shaw. BW: Lacev. RE. (2005). Design and evaluation of a low-volume total
suspended paniculate sampler. Trans ASAE 48: 1547-1552.
Warnken. KW: Santschi. PH. (2009). Delivery of trace metals (Al, Fe, Mn, V, Co, Ni, Cu, Cd, Ag, Pb)
from the Trinity River Watershed towards the ocean. Estuaries Coasts 32: 158-172.
http://dx.doi.org/10.1007/sl2237-008-9088-x
Watmough. SA; Dillon. PJ. (2007). Lead biogeochemistry in a central Ontario forested watershed.
Biogeochemistry 84: 143-159. http://dx.doi.org/10.1007/sl0533-007-9110-6
Watmough. SA; Hutchinson. TC: Dillon. PJ. (2004). Lead dynamics in the forest floor and mineral soil in
south-central Ontari. Biogeochemistry 71: 43-68. http://dx.doi.org/10.1007/sl0533-004-7661-3
Watson. JG: Chow. JC: Shah. JJ; Pace. TG. (1983). The effect of sampling inlets on the PM-10 and
PM-15 to TSP concentration ratios. J Air Waste Manag Assoc 33: 114-119.
Wedding. JB; McFarland. AR; Cermak. JE. (1977). Large particle collection characteristics of ambient
aerosol samplers. Environ Sci Technol 11: 387-390. http://dx.doi.org/10.1021/es60127a005
Wedepohl. KH. (1978). Handbook of geochemistry. In KH Wedepohl (Ed.). New York, NY: Springer.
Weiss. AL; Caravanos. J: Blaise. MJ; Jaeger. RJ. (2006). Distribution of lead in urban roadway grit and its
association with elevated steel structures. Chemosphere 65: 1762-1771.
http://dx.doi.0rg/10.1016/i.chemosphere.2006.04.079
Weitkamp. EA; Lipsky. EM; Pancras. PJ: Ondov. JM; Polidori. A; Turpin. BJ; Robinson. AL. (2005). Fine
particle emission profile for a large coke production facility based on highly time-resolved fence line
measurements. Atmos Environ 39: 6719-6733. http://dx.doi.Org/10.1016/j.atmosenv.2005.06.028
Westerlund. C: Viklander. M. (2006). Particles and associated metals in road runoff during snowmelt and
rainfall. Sci Total Environ 362: 143-156. http://dx.doi.Org/10.1016/i.scitotenv.2005.06.031
Widory. D. (2006). Lead isotopes decipher multiple origins within single PM10 samples in the
atmosphere of Paris. Isotopes Environ Health Stud 42: 97-105.
http://dx.doi.org/10.1080/10256010500502736
Wiersma. GB; Elvir. JA; Eckhoff. JD. (2007). Forest vegetation monitoring and foliar chemistry of red
spruce and red maple at Acadia National Park in Maine. Environ Monit Assess 126: 27-37.
http://dx.doi.org/10.1007/sl0661-006-9329-z
Williams. AGB; Scheckel. KG: Tolaymat. T; Impellitteri. CA. (2006). Mineralogy and characterization of
arsenic, iron, and lead in a mine waste-derived fertilizer. Environ Sci Technol 40: 4874-4879.
http://dx.doi.org/10.1021/es060853c
Witt. M; Baker. AR; Jickells. TD. (2006). Atmospheric trace metals over the Atlantic and South Indian
Oceans: Investigation of metal concentrations and lead isotope ratios in coastal and remote marine
aerosols. Atmos Environ 40: 5435-5451. http://dx.doi.Org/10.1016/i.atmosenv.2006.04.041
Wojas. B; Almquist. C. (2007). Mass concentrations and metals speciation of PM2.5, PM10, and total
suspended solids in Oxford, Ohio and comparison with those from metropolitan sites in the Greater
Cincinnati region. Atmos Environ 41: 9064-9078. http://dx.doi.Org/10.1016/i.atmosenv.2007.08.010
November 2012 3-252 Draft - Do Not Cite or Quote
-------
Wong. CSC: Li. XD; Thornton. I. (2006). Urban environmental geochemistry of trace metals [Review].
Environ Pollut 142: 1-16. http://dx.doi.Org/10.1016/i.envpol.2005.09.004
Wu. J: Edwards. R; He. XE; Liu. Z; Kleinman. M. (2010). Spatial analysis of bioavailable soil lead
concentrations in Los Angeles, California. Environ Res 110: 309-317.
http://dx.doi.0rg/10.1016/i.envres.2010.02.004
Wu. SC: Luo. YM; Cheung. KG: Wong. MH. (2006). Influence of bacteria on Pb and Zn speciation,
mobility and bioavailability in soil: A laboratory study. Environ Pollut 144: 765-773.
http://dx.doi.0rg/10.1016/i.envpol.2006.02.022
Xie. L; Giammar. DE. (2007). Equilibrium solubility and dissolution rate of the lead phosphate
chloropyromorphite. Environ Sci Technol 41: 8050-8055. http://dx.doi.org/10.1021/es071517e
Xie. RK; Seip. HM; Liu. L; Zhang. DS. (2009). Characterization of individual airborne particles in
Taiyuan City, China. Air Qual Atmos Health 2: 123-131. http://dx.doi.org/10.1007/sll869-009-0039-x
Yadav. S: Raiamani. V. (2006). Air quality and trace metal chemistry of different size fractions of aerosols
in N-NW India - implications for source diversity. Atmos Environ 40: 698-712.
http://dx.doi.0rg/10.1016/i.atmosenv.2005.10.005
Yang. HH; Chiang. CF; Lee. WJ: Hwang. KP; Wu. EMY. (1999). Size distribution and dry deposition of
road dust PAHs. Environ Int 25: 585-597. http://dx.doi.org/10.1016/S0160-4120(99)00036-7
Yesilonis. ID; Pouvat. RV: Neerchal. NK. (2008). Spatial distribution of metals in soils in Baltimore,
Maryland: Role of native parent material, proximity to major roads, housing age and screening
guidelines. Environ Pollut 156: 723-731. http://dx.doi.Org/10.1016/i.envpol.2008.06.010
Yi. SM; Totten. LA; Thota. S: Yan. S: Offenberg. JH; Eisenreich. SJ: Granev. J: Holsen. TM. (2006).
Atmospheric dry deposition of trace elements measured around the urban and industrially impacted
NY-NJ harbor. Atmos Environ 40: 6626-6637. http://dx.doi.Org/10.1016/i.atmosenv.2006.05.062
Ying. G: Sansalone. J. (2008). Granulometric relationships for urban source area runoff as a function of
hydrologic event classification and sedimentation. Water Air Soil Pollut 193: 229-246.
http://dx.doi.org/10.1007/sll270-008-9685-0
Yu. CH; Fan. ZH; Meng. Q; Zhu. X; Korn. L; Bonanno. LJ. (2011). Spatial/temporal variations of
elemental carbon, organic carbon, and trace elements in PM10 and the impact of land-use patterns on
community air pollution in Paterson, NJ. J Air Waste Manag Assoc 61: 673-688.
Zahran. S: Mielke. HW: Gonzales. CR; Powell. ET; Weiler. S. (2010). New Orleans before and after
hurricanes Katrina/Rita: A quasi-experiment of the association between soil lead and children's blood
lead. Environ Sci Technol 44: 4433-4440. http://dx.doi.org/10.1021/esl00572s
Zak. K; Rohovec. J: Navratil. T. (2009). Fluxes of heavy metals from a highly polluted watershed during
flood events: A case study of the Litavka River, Czech Republic. Water Air Soil Pollut 203: 343-358.
htrp://dx.doi.org/10.1007/sll270-009-0017-9
Zereini. F; Alt. F; Messerschmidt. J: Wiseman. C: Feldmann. I; Von Bohlen. A; Muller. J: Liebl. K;
Puttmann. W (2005). Concentration and distribution of heavy metals in urban airborne paniculate
matter in Frankfurt am Main, Germany. Environ Sci Technol 39: 2983-2989.
http://dx.doi.org/10.1021/es040040t
Zhang. LF; Peng. SQ: Wang. S. (2009a). Decreased aortic contractile reaction to 5-hydroxytryptamine in
rats with long-term hypertension induced by lead (Pb2+) exposure. Toxicol Lett 186: 78-83.
http://dx.doi.0rg/10.1016/i.toxlet.2009.01.004
Zheng. RQ; Li. CY. (2009). Effect of lead on survival, locomotion and sperm morphology of Asian
earthworm, Pheretima guillelmi. J Environ Sci 21: 691-695.
http://dx.doi.org/10.1016/sl001-0742(08)62325-6
Zota. AR; Willis. R; Jim. R; Norris. GA; Shine. JP; Duvall. RM; Schaider. LA; Spengler. JD. (2009).
Impact of mine waste on airborne respirable particulates in northeastern Oklahoma, United States. J
Air Waste Manag Assoc 59: 1347-1357. http://dx.doi.org/10.3155/1047-3289.59.ll.1347
November 2012 3-253 Draft - Do Not Cite or Quote
-------
Zuna. M; Mihaljevic. M; Sebek. O; Ettler. V; Handlev. M; Navratil. T; Golias. V. (2011). Recent lead
deposition trends in the Czech Republic as recorded by peat bogs and tree rings. Atmos Environ 45:
4950-4958. http://dx.doi.Org/10.1016/i.atmosenv.2011.06.007
November 2012 3-254 Draft - Do Not Cite or Quote
-------
4 EXPOSURE, TOXICOKINETICS, AND
BIOMARKERS
4.1 Exposure Assessment
1 The purpose of this section is to present recent studies that provide insight about human
2 exposure to Pb through various pathways. Pb is considered to be a multimedia
3 contaminant with multiple pathways of exposure. The relative importance of various
4 media in affecting Pb exposure changes with source strength and location, location and
5 time activity of the exposed individuals, behavior of the exposed individuals, and risk
6 factors such as age and socioeconomic factors (risk factors are discussed in detail in
7 Chapter_5). Blood Pb and bone Pb biomarkers (discussed in Sections 4.3. 4.4. and 4.5).
8 are often used to indicate composite Pb exposure resulting from multiple media and
9 pathways of exposure.
10 The recent information provided here builds upon the conclusions of the 2006 Pb AQCD
11 (U.S. EPA. 2006b). which found that air Pb concentrations and blood Pb levels have
12 decreased substantially following the restrictions on Pb in on-road vehicle gasoline, Pb in
13 household paints, the use of Pb solder, and reductions in industrial Pb emissions that have
14 occurred since the late 1970s. Nevertheless, detectable quantities of Pb have still been
15 observed to be bioaccessible in various media types. It was reported in the
16 2006 Pb AQCD (U.S. EPA. 2006b) that airborne maximum quarterly Pb concentrations
17 in the U.S. were in the range of 0.03-0.05 ug/m3 for non-source-oriented monitors for the
18 years 2000-2004 and were 0.10-0.22 ug/m3 for source-oriented monitors during that time
19 period, while blood Pb levels reached a median of 1.70 ug/dL among children (1-5 years
20 of age) in 2001-2002. It was also observed that Pb exposures were associated with nearby
21 industrial Pb sources, presence of Pb-based paint, and Pb deposited onto food in several
22 of the studies described in the 2006 Pb AQCD.
4.1.1 Pathways for Pb Exposure
23 Pathways of Pb exposure are difficult to disentangle because Pb has multiple sources in
24 the environment and passes through various environmental media. These issues are
25 described in detail in Sections 3.2 and 3.3. Air-related pathways of Pb exposure are the
26 focus of this ISA. Pb can be emitted to air, soil, or water and then cycle through any or all
27 of these media. In addition to primary emission of particle-bound or gaseous Pb to the
28 atmosphere, Pb can be resuspended to the air from soil or dust. Additionally, Pb-bearing
29 PM can be deposited from the air to soil or water through wet and dry deposition. Air-
November 2012 4-1 Draft - Do Not Cite or Quote
-------
1 related Pb exposures also include inhalation and ingestion of Pb-contaminated food,
2 water or other materials following atmospheric deposition of Pb; these exposures include
3 dust and soil via hand-to-mouth contact. In general, air-related pathways include those
4 pathways where Pb passes through ambient air on its path from a source to human
5 exposure. Some non-air-related exposures of Pb include ingestion of indoor Pb paint, Pb
6 in diet as a result of inadvertent additions during food processing, and Pb in drinking
7 water attributable to Pb in distribution systems, as well as other generally less prevalent
8 pathways.
9 The complicated nature of Pb exposure is illustrated in Figure 4-1. in which the Venn
10 diagram depicts how Pb can cycle through multiple environmental media prior to human
11 exposure. The "air/soil/water" arrows illustrate Pb exposures to plants, animals, and/or
12 humans via contact with Pb-containing media. The exposures are air-related if Pb passed
13 through the air compartment. When animals consume plant material or water exposed to
14 Pb that has at some point passed through the air compartment, and when human diet
15 includes animals, plants or drinking water exposed to Pb that has passed through the air
16 compartment, these are also considered air-related Pb exposures. As a result of the
17 multitude of possible air-related exposure scenarios and the related difficulty of
18 constructing Pb exposure histories, most studies of Pb exposure through air, water, and
19 soil can be informative to this review. Figure 4-1 also illustrates other exposures, such as
20 occupational exposures, contact with consumer goods in which Pb has been used, or
21 ingestion of Pb in drinking water conveyed through Pb pipes. Most Pb biomarker studies
22 do not indicate speciation or isotopic signature, and so exposures that are not related to
23 Pb in ambient air are also reviewed in this section because they can contribute to Pb body
24 burden. Many of the studies presented in the subsequent material focus on observations
25 of Pb exposure via one medium: air, water, soil and dust, diet, or occupation.
November 2012 4-2 Draft - Do Not Cite or Quote
-------
Newly Emitted Pb
Historically Emitted Pb
OOTDOORSOIL
NDDUST
NATURAL WATERS
AND SEDIMENTS
Non-air Pb
eleases
•— --.
AIR
SOIL
WATER
1 — - -
**• —
PLANT
EXPOSURE
-+
^^.
AIR
SOIL
WATER
--. — — -
**• — — -— ,
r ANIMAL
t EXPOSURE]]"
AIR
SOIL
WATER
•——_—--
.-•— •
HUMAN
^EXPOSURE
( COSMETICS )
OYS et
Note: The Venn diagram is used to illustrate the passage of Pb through multiple environmental media compartments through which
exposure can occur.
Figure 4-1 Conceptual model of multimedia Pb exposure.
1 The relative importance of different sources or pathways of potential exposure to Pb in
2 the environment is often difficult to discern. Individual factors such as home
3 environment, location, and risk factors (described in more detail in Chapter_5) may
4 influence exposures. The National Human Exposure Assessment Survey (NHEXAS)
5 study sampled Pb, as well as other pollutants and VOCs, in multiple exposure media from
6 subjects across six states in EPA Region 5 (Illinois, Indiana, Michigan, Minnesota, Ohio,
7 and Wisconsin) (Clayton et al.. 1999) as well as in Arizona (O'Rourke et al.. 1999) and
8 Maryland (Egeghy et al., 2005). Results from NHEXAS indicate that personal exposure
9 concentrations of Pb are higher than indoor or outdoor concentrations of Pb, perhaps
10 suggesting a personal cloud effect; see Table 4-1. Pb levels in windowsill dust were
11 higher than Pb levels in surface dust collected from other surfaces. Clayton et al. (1999)
12 suggested that higher windowsill levels could be attributed to the presence of Pb-based
13 paint and/or to accumulation of infiltrated outdoor Pb-bearing PM. Pb levels in food were
14 higher than in beverages, and Pb levels in standing tap water (also referred to as "first
15 flush" or "first draw") were higher than Pb levels obtained after allowing water to run for
16 three minutes to flush out pipes. Layton and Beamer (2009) estimated that 34-66% of Pb
17 in floor dust was tracked in from outdoors and originated as ambient air Pb, based on
November 2012
4-3
Draft - Do Not Cite or Quote
-------
1992 levels in Sacramento; in 1992, phase-out of Pb usage in gasoline was near complete,
but industrial emissions were still higher than current levels; see Section 3.2.
Table 4-1 Estimates of Pb measurements for EPA Region 5 from the
NHEXAS study.
Medium3
Personal air
(ng/m3)d
Indoor air
(ng/m3)d
Outdoor air
(ng/m3)d
Surface dust
(ng/cm2)
Surface dust
(mg/kg)
Window sill dust
(ng/cm2)
Window sill dust
(mg/kg)
Standing tap water
(ug/L)
Flushed tap water
(ug/L)
Solid food
(ug/kg)
Beverages
(ug/kg)
Food + Beverages
(ug/kg)
Food intake
(ug/day)
Beverage intake
(ug/day)
Food + Beverage intake
(ug/day)
Blood
(ug/dL)
N
167
213
87
245
244
239
239
444
443
159
160
156
159
160
156
165
Percentage above
LODb (CLs)c
81.6
(71 .3; 92.0)
49.8
(37.2; 62.3)
73.8
(56.3; 91 .3)
92.1
(87.4; 96.8)
92.1
(87.4; 96.8)
95.8
(92.5; 99.0)
95.8
(92.5; 99.0)
98.8
(97.6; 100.0)
78.7
(70.7; 86.7)
100.0
(100.0; 100.0)
91.5
(85.2; 97.8)
100.0
(100.0; 100.0)
100.0
(100.0; 100.0)
91.5
(85.2; 97.8)
100.0
(100.0; 100.0)
94.2
(88.2; 100.0)
Mean (CLs)c
26.83
(17.60; 36.06)
14.37
(8.76; 19.98)
11.32
(8.16; 14.47)
514.43
(-336.6; 1365.5)
463.09
(188.1 5; 738.04)
1,822.6
(481.49; 3,163.6)
954.07
(506.70; 1 ,401 .4)
3.92
(3.06; 4.79)
0.84
(0.60; 1.07)
10.47
(6.87; 14.07)
1.42
(1.13; 1.72)
4.48
(2.94; 6.02)
7.96
(4.25; 1 1 .68)
2.15
(1.66; 2.64)
10.20
(6.52; 13.89)
2.18
(1.78; 2.58)
50th (CLs)c
13.01
(11.13; 18.13)
6.61
(4.99; 8. 15)
8.50
(7. 14; 10.35)
5.96
(3.37; 10.94)
120.12
(83.85; 160.59)
16.76
(10.44; 39.41)
191.43
(140.48; 256.65)
1.92
(1 .49; 2.74)
0.33
(0.23; 0.49)
6.88
(6.44; 8.04)
0.99
(0.84; 1.21)
3.10
(2.66; 3.52)
4.56
(3.68; 5.36)
1.41
(1.18; 1.60)
6.40
(5.21 ; 7.78)
1.61
(1.41; 2. 17)
90th (CLs)c
57.20
(31. 18; 85.10)
18.50
(12.69; 30.31)
20.36
(12.60; 34.91)
84.23
(26.52; 442.63)
698.92
(411.84; 1,062.8)
439.73
(106.34; 4,436.2)
1 ,842.8
(1,1 51 .3; 2,782.5)
9.34
(7.87; 12.35)
1.85
(1.21; 3.04)
14.88
(10.78; 19.08)
2.47
(2.06; 3.59)
6.37
(4.89; 8.00)
12.61
(9.27; 16.38)
4.45
(3.1 5; 5.65)
16.05
(13.31; 18.85)
4.05
(3.24; 5.18)
Note: EPA Region 5 includes six states: Illinois, Indiana, Ohio, Michigan, Minnesota, and Wisconsin. Participants were enrolled
using a stratified, four-stage probability sampling design, and submitted questionnaire and physical measurements data. Summary
statistics (percentage above limit of detection (LOD), mean, median, 90th percentile) were computed using weighted sample data
analysis. The estimates apply to the larger Region 5 target population (all non-institutionalized residents residing in households).
"Estimates for indoor air, outdoor air, dust media, and water media apply to the target population of Region 5 households; estimates
for other media apply to the target population of Region 5 residents.
bPercentage of the target population of residents (or households) estimated to have Pb levels above limit of detection (LOD).
°The lower and upper bounds of the 95% confidence limits (CL) are provided.
dPM50.
Source: Reprinted with permission of Nature Publishing Group, Clayton et al. (1999)
November 2012
4-4
Draft - Do Not Cite or Quote
-------
4.1.1.1 Particle Size Distributions for Airborne-Pb, Dust-Pb, and
Soil-Pb
1 The size distribution of ingestible dust particles differs from the size distribution of
2 inhalable ambient air Pb particles and therefore cannot be directly compared. The
3 inhalability of airborne PM is a gradually decreasing function of particle size. Inhalability
4 criteria established from experimental data, obtained at wind speeds of 1-8
5 meters/second, describe PM inhalability of 77% for particles <10 jam (dae, aerodynamic
6 diameter). Inhalability of particles ranging in size from 40 to 100 (im dae is 50%; above
7 100 (im, inhalability data are lacking (Soderholm. 1989: ACGIH. 1985). The particles
8 that are not inhaled may settle to surfaces, making them available for subsequent
9 ingestion. The size distribution of soil and house dust particles tends to be much larger
10 than airborne PM. Que Hee et al. (1985) and U.S. EPA (1990b) observed that 50% or
11 more of the mass of house dust tends to be comprised of particles smaller than 150 pirn.
12 Gulson et al. (1995b) observed that the mode of the Pb house dust size distribution was in
13 the 38-53 nm range; they did not report the overall house dust size distribution. Given the
14 house dust Pb size distributions documented, dust Pb brought into homes with foot traffic
15 may be aerosolized but is likely to stay airborne for only a few seconds, since particles
16 larger than PM2 5 tend to settle from the air quickly; see Section 3.3.1.3 and Section
17 4.1.3.1. Siciliano et al. (2009) observed different size distributions for different types of
18 soils: agricultural sites had median soil Pb of 34 nm, and brownfields had median soil Pb
19 of 105 nm. These observations of larger particle sizes for soil and dust Pb support the
20 notion that exposure to Pb in dusts and soils would occur by ingestion rather than
21 inhalation following resuspension.
22 The main pathway for Pb ingestion by children is by hand to mouth contact (Lanphear et
23 al.. 1998). In a playground environment in London, U.K., Duggan et al. (1985) reported
24 that hand to mouth transfer was effectively limited to particles smaller than 10 urn, even
25 when the soil itself exhibited a much larger particle size distribution. More recently,
26 Yamamoto et al. (2006) reported for a cohort of children in Kanagawa Prefecture, Japan
27 (greater Tokyo area) that the mode of size distributions of particles adhering to children's
28 hands was 39 ± 26 (im, with the upper tail ranging from 200-300 (im. Kissel et al. (1996)
29 measured three size fractions of soil adhered to a hand via a hand press: < 150 pirn,
30 150-250 nm, and > 250 pirn and observed that, when soil was dry (<2% moisture
31 content), 43%-69% of the soil was in the smallest fraction. When the moisture content
32 was higher than 2%, 28-81% of the adhered soil was in larger than 250 pirn. Percentage
33 and mass adhered per area (mg/cm2) depended on soil type, with wet sand and loamy
34 sand adhering more to hands than sandy loam or silt loam. For dry soil, silt loam mass
35 produced the largest adherence in terms of mass per area. Differences among the size
November 2012 4-5 Draft - Do Not Cite or Quote
-------
1 distribution results may be related to differences in the soil type, soil moisture levels
2 between the locations, and/or to differences between the analytical methods used to
3 measure size distribution; Duggan and Inskip (1985) used optical microscopy of the dust
4 wipes, while Yamamoto et al. (2006) used a laser scattering device measuring sampled
5 particles suspended in an aqueous solution. Siciliano et al. (2009) regressed adhered
6 average soil size on hands bulk soil particle size and found a log-log relationship with
7 (3 = 0.66 using both brownfield and agricultural soils; the proportion of soil adhered
8 depended on organic content.
9 Several studies have found that Pb is enriched in the smaller fractions of the soil or
10 house-dust size distribution. Davies and White (1981) observed that enrichment
11 decreased linearly with increasing dust size bin, with dust particles smaller than 64 nm
12 having a Pb concentration of 76.1 mg/kg and particles in the 1,000-2,000 urn size range
13 having a Pb concentration of 16.4 mg/kg. Sheets and Bergquist (1999) also found that Pb
14 content decreased with increasing particle size. More recently, Ljung et al. (2006)
15 investigated childhood exposures to trace metals on playgrounds in Uppsala, Sweden and
16 observed that the Pb content in soil in the <50 pirn size fraction was 1.5 times higher than
17 that in the >4 mm or 50-100 pirn size fractions. Sheppard et al. (1995) measured
18 enrichment in different types of soils (sand and clay) and found that enrichment was
19 substantially higher in the sand.
20 Studies focusing on particle size distributions of house dust adhered to the hands are
21 lacking. Ingestion of house dust has been reported to be the major source of Pb intake
22 during early childhood (Lanphear et al.. 2002). If a similar particle size distribution holds
23 for household dust, then ingestion of indoor Pb of atmospheric origin could also be
24 strongly dependent on dust particle size. Therefore, larger particles of atmospheric origin,
25 which may not be considered relevant for exposure by inhalation exposure, are still
26 relevant for Pb exposure by ingestion. However, no studies in the literature have
27 presented information on the relative contributions of Pb from different PM size fractions
28 to blood Pb concentrations.
29 It should be noted that different measurement techniques are used for different
30 environmental media. For example, ambient air Pb-PM size distribution is measured by
31 one of the non-FRM instruments such as a MOUDI, described in Section 3.4. and its
32 measurement is subject to errors specific to the technique. Dust and soil size distribution
33 are typically measured with graduated sieves, and errors associated with these methods
34 occur more often in the smaller size fractions that are subject to agglomeration and
35 clogging if the particle shape is nonspherical.
November 2012 4-6 Draft - Do Not Cite or Quote
-------
4.1.1.2 Estimating Pb Exposure in the Integrated Exposure
Uptake Biokinetic (IEUBK) Model
1 Several studies have used a combination of measured values and default model values to
2 represent exposures and determine their relative contributions to blood Pb. For example,
3 Cornells et al. (2006) used the Integrated Exposure Uptake Biokinetic model (IEUBK),
4 described in detail in the 2006 Pb AQCD (U.S. EPA. 2006b) to model children's
5 exposures to Pb emissions from a non-ferrous smelter in Hoboken, Belgium. In deriving
6 the model input (annual averages) for ambient air Pb concentration, as well as soil and
7 dust, they employed weighting based on children's time spent in different locations in the
8 study area and air, soil and indoor dust measurements in those areas. In their results for
9 the area of the smelter, the ingestion of dust and soil pathways accounted for more than
10 70% of the exposure, while the inhalation pathway accounted for less than 2%. Similarly,
11 Carrizales et al. (2006) analyzed exposures to children living near a copper (Cu) smelter
12 in San Luis Potosi, Mexico. They employed the IEUBK default options for assignment of
13 Pb dust concentration as 70% of the soil Pb concentration, while air Pb concentration was
14 assigned based on measurements by the Mexican government. Based on these
15 assumptions, they attributed 87% of blood Pb to soil and dust exposure. These studies did
16 not estimate the air Pb contribution to the soil/dust Pb concentrations and consequently
17 did not estimate the portion of the ingestion pathway that derives from ambient air Pb.
18 Appendix I of the 2007 Pb Risk Assessment (U.S. EPA. 2007f) provides estimates of the
19 contribution of various pathways to the blood Pb of children simulated in several case
20 studies. Simulations provided estimates of contributions from outdoor ambient air Pb by
21 inhalation and by ingestion of indoor dust, including the fraction of indoor dust Pb
22 associated with recent penetration of ambient air Pb into the residence. Although ambient
23 air Pb may also contribute to Pb ingestion through other pathways (i.e., diet, soil), data
24 and tools to support a simulation of the linkage between air Pb concentrations and
25 concentrations in other media were limited. Accordingly, Pb concentrations pertaining to
26 other pathways (e.g., diet, outdoor soil, the component of indoor dust Pb other than that
27 derived from Pb recently in ambient air) were held constant across the different air
28 quality scenarios simulated. Table 4-2 provides estimates for the General Urban Case
29 Study in the 2007 Pb Risk Assessment (U.S. EPA. 2007f). The General Urban Case
30 Study, unlike the various location-specific case studies, was not based on any specific
31 urban location and reflected several simplifying assumptions including uniform ambient
32 air Pb levels across the simulated, hypothetical study area and a uniform study
33 population.
November 2012 4-7 Draft - Do Not Cite or Quote
-------
Table 4-2 Predicted concurrent blood Pb levels and source contributions for
children in their seventh year of life.
Pathway Contribution (%)
Air Pba
(ug/m3)
0.05
0.149
0.2
0.87 h
Median
Blood Pbb
(ug/dL)
1.7(5.7)'
1.9(6.5)
2.0 (6.9)
2.1 (7.2)
Diet0
32
28
26
25
Ingestion
Outdoor
Soil/Dust
44
38
36
33
Indoor
Otherd
11
6
5
4
Dust
Pb recently in
aire
12.6
28.3
32.7
37.2
Inhalation
Ambient
Air
0.1
0.5
0.7
0.9
"Concentrations are maximum calendar quarter averages of Pb in TSP with exception of 0.05 ug/m which is a maximum
monthly average
""Average of blood Pb concentrations at 75 and 81 months, assuming exposure concentrations were constant through
7 years of life
Includes food and drinking water
Includes indoor dust with Pb contributions from sources other than Pb recently in the air (e.g., indoor paint, outdoor
soil/dust, and additional sources including historical air Pb)
Includes contributions associated with outdoor ambient air Pb from ingestion of indoor dust predicted to be associated
with outdoor ambient Pb levels
Values in parentheses are the 95th percentile blood Pb for a geometric standard deviation of 2.1
9Mean of the maximum quarterly average concentrations of Pb in TSP (for period 2003 to 2005) among monitor locations
in urban areas having more than one million residents
h95th percentile of the maximum quarterly average concentration of Pb in TSP (for period 2003 to 2005) among monitor
locations in urban areas having more than one million residents
Source: Based on General Urban Case Study (Hybrid Dust Model) in Appendix I, 2007 Pb Risk Assessment (U.S. EPA,
2007 f}.
4.1.2 Environmental Exposure Assessment Methodologies
1 A number of monitoring and modeling techniques have been employed for exposure
2 assessment. These are detailed in either the 2006 Pb AQCD (U.S. EPA. 2006b) or in the
3 subsequent Risk and Exposure Assessment performed as part of the same NAAQS
4 review (U.S. EPA. 2007g). Some of these methods are briefly described here to provide a
5 context for the exposure studies described in Section 4.1.3. Blood Pb sampling is
6 described in detail in Section 4.3.2.
7 Data collection to assess Pb exposure pathways may involve air, soil, and dust samples.
8 Methods used for digesting air Pb samples are described in Section 3.4. as are ambient air
9 Pb monitoring techniques. Factors affecting collection of ambient air Pb samples are
10 described in detail in Section 3.4. For the monitors in the FRM network, the primary role
11 is compliance assessment. Accordingly, this network includes monitors in locations near
12 sources of air Pb emissions which are expected to or have been shown to contribute to
13 ambient air Pb concentrations in excess of the Pb NAAQS. In such locations, Pb may be
November 2012
4-8
Draft - Do Not Cite or Quote
-------
1 associated with relatively larger size particles, contributing to air Pb concentration
2 gradients with distance from the source and greater deposition in the near-source
3 locations. The FRM network also includes non-source-oriented monitors for which the
4 main objective is to gather information on neighborhood-scale Pb concentrations that are
5 typical in urban areas so to better understand ambient air-related Pb exposures for
6 populations in these areas. This part of the Pb NAAQS network, was required to be
7 operational as of December 27, 2011. These monitor locations are distributed across a
8 broad geographic area, representing approximately 63 large urban areas which contain
9 approximately half of the total U.S. population (based on recently published 2010 Census
10 Bureau data). In lieu of more detailed analysis of population proximity for these newly
11 established monitors, population counts were calculated near previously existing
12 monitors for which data are presented in Section 3.5. For the monitors in that limited
13 dataset, among the total population of 311,127,619 people in the 2010 Census (ESRI.
14 2011). 181,100 (0.06%) lived within 1 km of a source-oriented monitor, while 918,351
15 (0.30%) lived within 1 km of a non-source-oriented monitor.
16 Dust sampling has not changed drastically since it was first proposed by Sayre et al.
17 (1974). in which a disposable paper towel was soaked in 20% denatured alcohol and
18 1:750 benzalkonium chloride and then used to wipe a 1 ft2 sampling area in a systematic
19 fashion. Que Hee et al. (1985) and Sterling et al. (1999) compared wipe testing with
20 vacuum methods. Sampling efficiency for the first attempt varied between 53-76% with
21 vacuum pump flow rate and tube type and was 52% for the wipe method for the Que Hee
22 et al. (1985) study, with 100% efficiency after five consecutive samples were obtained.
23 Sterling et al. (1999) observed that two of three vacuuming methods had significantly
24 higher geometric mean collection (vacuum 1: 94.3 (ig/ft2; vacuum 2: 23.5 (ig/ft2)
25 compared with dust wipes (5.6 (ig/ft2).
26 Models may also be used in exposure assessment. For example, two dispersion models,
27 the American Meteorological Society/Environmental Protection Agency Regulatory
28 Model (AERMOD), and Industrial Source Complex-Plume Rise Model Enhancements
29 (ISC-PRIME) were employed to model dispersion of Pb emissions from specific
30 industrial facilities (Cimorelli et al.. 2005; Perry etal. 2005; EPRI. 1997). and to
31 estimate ambient air Pb concentrations at some of the case studies included in the 2007
32 Risk and Exposure Assessment (U.S. EPA. 2007g). These models assume plume
33 dispersion follows a Gaussian distribution from a point source. For the two point source
34 case studies included in the 2007 risk assessment, the plume models were used to track
35 emissions to ambient air near homes located within a few miles of emitting facilities.
36 However, dispersion models can also be used to track long distance transport of Pb
37 emissions, as performed by Krell and Roeckner (1988) to model the dispersion and
38 deposition of Pb and Cd from European nations into the North Sea.
November 2012 4-9 Draft - Do Not Cite or Quote
-------
1 Several models estimate blood Pb levels resulting from estimated exposure to Pb in
2 environmental media. These models, which are described in detail in the 2006 Pb AQCD
3 (U.S. EPA. 2006b) include the IEUBK model, and the EPA All Ages Lead Model
4 (AALM), which combines and expands the thorough exposure and absorption modules of
5 the IEUBK model with the comprehensive biokinetic model of Leggett (1993). As of the
6 writing of this assessment, the AALM is still in development.
7 The Stochastic Human Exposure and Dose (SHEDS) and NORMTOX models also are
8 capable of modeling metals exposures through various routes including inhalation,
9 ingestion, and dermal exposure (Loos etal. 2010; Burke et al.. 2002). Pb exposure
10 modeling can also be accomplished using the Modeling Environment for Total Risk
11 (MENTOR) framework, in which airborne Pb levels could be modeled using AQS,
12 dispersion modeling, or chemical transport modeling, while human exposure is modeled
13 with SHEDS or a similar exposure model (Georgopoulos and Lioy. 2006). Additionally,
14 housing data and time-activity data from the Consolidated Human Activity Database
15 (CHAD) are incorporated into MENTOR to develop refined estimates of Pb exposure and
16 tissue burden. However, a literature search did not produce any Pb exposure studies using
17 the SHEDS, NORMTOX, or MENTOR modeling systems. In general, these models take
18 input for several environmental Pb exposure media including soil, dust, food and water,
19 outdoor air, and indoor air. The models are designed to evaluate different exposure
20 scenarios based on specification of particular conditions.
4.1.3 Exposure Studies
4.1.3.1 Airborne Pb Exposure
21 Limited personal exposure monitoring data for airborne Pb were available for the
22 2006 Pb AQCD (U.S. EPA. 2006b). As described above, the NHEXAS study showed
23 personal air Pb concentrations to be significantly higher than indoor or outdoor air Pb
24 concentrations (Clayton et al.. 1999). Indoor air Pb concentration was moderately
25 correlated with floor dust and residential yard soil Pb concentration (Rabinowitz et al..
26 1985). Egeghy et al. (2005) performed multivariate fixed effects analysis of the
27 NHEXAS-Maryland data and found that Pb levels measured in indoor air were
28 significantly associated with log-transformed outdoor air Pb levels, ambient temperature,
29 number of hours in which windows were open, whether homes were built before 1950,
30 and frequency of fireplace usage (Table 4-3).
November 2012 4-10 Draft - Do Not Cite or Quote
-------
Table 4-3 Estimates of fixed effects multivariate modeling of Pb levels
measured during the NHEXAS-MD study.
Pb in Indoor Air
Fixed Effect pa p-value
Intercept -0.50 0.0051
Pb in Dust Dermal Pb Blood Pb
pa p-value pa p-value pa p-value
6.22 <0.0001 6.23 <0.0001 0.02 0.91
Outdoor Pb concentration11 0.51 <0.0001
Average weekly temperature (°F) 0.01 0.046
Open window periods (hr) 0.01 0.035
-0.03 0.0082
House pets (yes) -0.15 0.078
Air filter use (yes) -0.28 0.087
Home age (<1 950) 0.25 0.025
Fireplace (frequency of use) 0.11 0.045
Pb concentration in soilb
Interior Pb paint chipping/peeling (yes)
Cement at primary entryway (yes)
Indoor pesticide usage last 6 mo (yes)
Electrostatic air filter usage (yes)
Sex of participants (male)
Ethnic minority participants (yes)
Washing hands after lawn mowing (no)
Gasoline power- equipment usage (yes)
Bathing or showering activities (yes)
Dust level indoors (scale: 1-3)
Residing near commercial areas (yes)
Age of participants (yr)
Number cigarettes smoked (count)
Burning wood or trash (days)
Showering frequency (avg # days)
Work outside home (yes)
Health status (good)
Adherence to high fiber diet (yes)
Gas or charcoal grill usage (yes)
-0.12 0.088
0.96 0.029
0.46 0.0054
0.27 0.037
0.43 0.091
1 .97 0.0064
-0.78 0.0003
-0.91 0.062
0.41 0.0012 0.43 <0.0001
0.41 0.0063
1.04 0.0010
0.61 0.0072
-0.43 0.019
0.22 0.019
0.32 0.0087
0.02 <0.0001
0.03 <0.0001
0.58 0.0099
-0.29 0.0064
-0.26 <0.0001
0.23 0.0009
-0.15 0.040
-0.17 0.0002
Estimates of fixed effects in final multiple regression analysis models for Pb in the Maryland investigation data in the National Human Exposure Assessment Survey
(NHEXAS-MD).
bLog transform
Source: Reprinted with permission of Nature Publishing Group, Egeghy et al. (20051.
November 2012
4-11
Draft - Do Not Cite or Quote
-------
1 Some recent studies have shown that the ratio of indoor to outdoor Pb-PM varies from
2 site to site depending on factors including infiltration, indoor and outdoor Pb sources, and
3 meteorology. Adgate et al. (2007) measured the concentrations of several trace elements
4 in personal, indoor, and outdoor air samples of PM2 5 and found that average personal
5 Pb-PM2 5 concentration was roughly three times higher than outdoor air Pb-PM2 5
6 concentration and two times higher than indoor Pb-PM2 5 concentration (Table 4-4).
7 Another study of indoor and outdoor air concentrations of Pb was carried out by Molnar
8 et al. (2007). PM2 5 trace element concentrations were determined in homes, preschools
9 and schools in Stockholm, Sweden. In all sampled locations, Pb-PM2 5 concentrations
10 were higher in the outdoor environment than in the proximal indoor environment. The
11 indoor/outdoor ratios for Pb-PM2 5 suggest an outdoor Pb-PM2 5 net infiltration of -0.6 for
12 these buildings. Outdoor air Pb concentrations did not differ between the central and
13 more rural locations. Indoor air Pb concentrations were higher in spring than in winter,
14 which the authors attributed to greater resuspension of elements that had accumulated in
15 road dust over the winter period and increased roadwear on days with dry surfaces. Pekey
16 et al. (2010) measured indoor and outdoor trace element composition of PM2 5 and PMi0
17 in Kocaeli, an industrial region of Turkey, and found that average airborne Pb
18 concentrations were higher outdoors than indoors for both PM2 5 and PMi0 during
19 summer and for PMi0 during winter, but that indoor Pb concentration was higher than
20 outdoor Pb concentration for PM2 5 during winter. The indoor-to-outdoor ratio of airborne
21 Pb varied by environment; it tended to be less than one, but the ratio varied from one
22 microenvironment to another. In a pilot study in Windsor, Ontario, Rasmussen et al.
23 (2007) observed that the concentration of Pb in PM2 5 from a personal exposure sample
24 was roughly 40% higher than the concentration of Pb in outdoor PM2 5 and 150% higher
25 than Pb in indoor PM2 5. The three studies that included personal samples recorded
26 measurements that were consistently higher than indoor or outdoor levels, and outdoor
27 concentrations were higher than indoor concentrations.
28 Domestic wood burning is a potential source of Pb compounds (Section 3.2.2.5). Alves et
29 al. (2011) measured trace metals in woodstove and fireplace emissions and found that
30 PM25 contained Pb, with concentrations from wood burning ranging from 3.3-12.2 (ig/g
31 and 2.89-30.3 for woodstoves and fireplaces, respectively. When burning briquettes, the
32 PM2 5 measurements showed Pb enrichment above all other metal elements other than
33 potassium (woodstove: 1361 (ig/g; fireplace: 616 (ig/g). Molnar et al. (2005) measured
34 trace element concentration in indoor and personal exposure PM2 5 samples for homes in
35 which wood is burned and in a reference group where no wood burning occurs in the
36 home. For both indoor and personal samples, Molnar et al. (2005) observed that Pb
37 concentrations were higher for the wood burning group and nearly statistically significant
38 for the personal exposure samples (indoor concentration: 6.0 (ig/m3 versus 4.3 (ig/m3,
39 p = 0.26; personal exposure: 4.6 (ig/m3 versus 3.0 (ig/m3, p = 0.06).
November 2012 4-12 Draft - Do Not Cite or Quote
-------
1 Indoor activity has been associated with resuspension of settled dust, which could cause
2 airborne contact with particle-bound Pb. Qian et al. (2008) estimated a PMi0 resuspension
3 rate of 1.4xlO"4/hr for one person walking across a carpeted floor. Measurements of
4 submicron particles illustrated a roughly two-fold increase of airborne particle
5 concentration for particles smaller than 1.8 (im for activity versus low activity periods,
6 with maximum concentrations reaching 4-11 times the maximum value during low
7 activity periods. For PM10, average concentration was 2.5 times higher than background
8 levels during activity periods, while peak concentration was 4.5 times higher. Qian and
9 Ferro (2008) observed that resuspension rates depend on particle size, floor material, and
10 ventilation position. Increases in walking speed and weight of the walker did not
11 consistently produce increases in resuspension. 5-10 (im particles produced a higher
12 resuspension rate compared with smaller particles. Newly carpeted areas produced
13 significantly higher resuspension rates than vinyl floors. Zhang et al. (2008) modeled and
14 conducted experiments of particle dispersion from walking and observed that human
15 activity did affect resuspension. They found that larger particles were more readily
16 detached from the carpet by walking motion, but that smaller particles are more easily
17 resuspended once detached. Hunt and Johnson (2012) studied the duration and spatial
18 extent of resuspension of 0.3-5.0 (im particles following walking by a soiled shoe.
19 0.3-0.5 (im particle concentration remained increased over atime period of 23 min, while
20 1-5 (im particles declined in concentration over the same time period. Experiments and
21 computational fluid dynamics simulations by Eisner et al. (2010) for a mechanical foot
22 moving on carpeting suggested that the rotating motion of the moving foot on the carpet
23 induced rotating air movement beneath the foot that re-entrained the particles.
24 Several of the studies can be used to develop an understanding of how personal exposure
25 to PM-bound Pb varies with other exposures. Molnar et al. (2007) reported Spearman
26 correlations of Pb with PM25 and NO2 in three outdoor microenvironments (residence,
27 school, and preschool) and found that Pb and other trace metals were generally well
28 correlated with PM2 5 (r = 0.72-0.85), but Pb was only statistically significantly correlated
29 with NO2 in one of the three outdoor microenvironments (r = 0.24-0.75). Pb was
30 attributed by Molnar et al. (2007) to long range transport. Table 3-29 illustrates that Pb
31 concentrations in the four studies (summarized in the Chapter 3 Appendix [Section 3.81)
32 are typically well below the level of the NAAQS. The higher personal air concentratoins
33 occurred in a heavily industrialized area of Kocaeli, Turkey with an incinerator and
34 several industrial facilities including metal processing, cement, petroleum refining, and
35 agriculture processing. Otherwise, concentrations were all between 0.002 and
36 0.006 ug/m3. The proportion of Pb compared with other trace metals varied with location
37 and component. It was typically several times lower than S as well as crustal elements
38 such as Ca2+ and Fe. In the industrial area of Kocaeli, Pb comprised a greater proportion
39 of the PM compared with other areas.
November 2012 4-13 Draft - Do Not Cite or Quote
-------
1
2
o
J
4
5
6
7
Table 4-4
Study
Clayton et al.
(1999)
Adgate et al.
(2007)
Molnar et al.
(2007)
Tovalin-
Ahumada et al.
(2007)
Pekey et al.
(2010)
Rasmussen et
al. (2007)
Comparison of personal, indoor, and outdoor Pb-PM measurements
from several studies.
Location
IL, IN, Ml, MN,
OH, Wl
Minneapolis-
St. Paul, MN
Stockholm,
Sweden
Mexico City,
Mexico
Puebla, Mexico
Kocaeli, Turkey
Windsor,
Ontario,
Canada
4.1.3.2
The 2006 Pb
Pb Metric
Med. Pb-PMso
(ng/m3)
Avg. Pb-PM25
(ng/m3)
Avg. Pb-PM25
(ng/m3)
Med. Pb-PM25
(ng/m3)
Med. Pb-PM2.5
(ng/m3)
Avg. Pb-PM25
(ng/m3)
Avg. Pb-PM10
(ng/m3)
Med. Pb-PM2.5
(mg/kg)
Sampling
Period Personal Pb
July, 1995- 13
May, 1997
Spring, 6.2
Summer,
Fall, 1999
December,
2003-
July, 2004
April-May,
2002
April-May,
2002
May-June,
2006,
December,
2006-
January 2007
May-June,
2006,
December,
2006-
January 2007
April, 2004 31 1
Exposure to Pb in Soil and Dust
AOCD (U.S. EPA. 2006b) lists indoor Pb dust
Indoor Pb Outdoor Pb
6.6 8.5
3.4 2.0
Homes: 3.4 Homes: 4.5
Schools: 2.5 Schools: 4.6
Preschools: 1 .8 Preschools: 2.6
26 56
4 4
Summer: 34 Summer: 47
Winter: 85 Winter: 72
Summer: 57 Summer: 78
Winter: 125 Winter: 159
124 221
infiltrated from outdoors as a
potential source of exposure to Pb soil and dust. Thus, outdoor soil Pb may present an
inhalation exposure if resuspended indoors or an ingestion exposure during hand-to-
mouth contact. A detailed description of studies of outdoor soil Pb concentration is
provided in Section 3.6.1. Indoor measurements can reflect infiltrated Pb as well as Pb
dust derived from debrided paint, consumer products, or soil that has been transported
into the home via foot traffic. Table 4-5 presents indoor dust Pb concentrations for
2006-2011 observational studies in which indoor dust Pb was measured.
November 2012
4-14
Draft - Do Not Cite or Quote
-------
Table 4-5
Reference
Caravanos et al.
(2006b)
Khoder et al.
(2010)
Brattin and
Griffin (2011)
Yu et al. (2006)
Turner and
Simmonds
(2006)
Gaitenset al.
(2009)
Wilson et al.
(2007)
Zota et al. (201 1 )
Spalingeret al.
(2007)
Measurements of indoor dust Pb concentration from 2006-2011
studies.
Study Location
New York City, New York
Giza, Egypt (extensive leaded
gasoline use; industrial area)
Eureka, Utah near Eureka Mills
Superfund Site
Denver, CO, near VBI70 Superfund
Site
East Helena, MT, near East Helena
Superfund Site
Syracuse, New York
Birmingham, Plymouth,
and 2 rural sites, U.K.
U.S. (nationwide)
Milwaukee, Wisconsin
Ottawa County, Oklahoma
(area surrounding the Tar Creek
Superfund Site)
Rural towns, Idaho
Bunker Hill, Idaho Superfund site
Metric (units)
Weekly dust
loading (ug/m2)
Weekly dust
loading (ug/m2)
Dust concentration
(mg/kg)
Dust concentration
range (mg/kg)
Dust concentration
(mg/kg)
Dust loading
(ug/m2)
Dust concentration
(ug/m2)
Dust concentration
(mg/kg)
Dust concentration
(mg/kg)
Dust concentration
(mg/kg)
Sample Site
Glass plate next to open
window of academic
building
Glass plate in second-
floor living room of
apartments
Indoor home site (not
specified)
Indoor home site (not
specified)
Indoor home site (not
specified)
Floor
Floor
Smooth floor
Rough floor
Smooth windowsill
Rough windowsill
Central perimeter
Entry
Window
Indoor (site not specified)
Vacuum
Floor
Vacuum
Floor
Indoor Pb
Concentration
Median: 52
Median: 408
1 60-2000
11-660
68-1 000
Range: 209-1770
Median: 178
Median: 1.7
Avg.: 4.4
Median: 5.6
Avg.: 16
Median: 2.5
Avg.: 190
Median: 55
Avg.: 480
Avg.: 107
Avg.: 140
Avg.: 151
Avg.: 109
Median: 63
Max.: 881
Median: 120
Max: 830
Median: 95
Max: 1 ,300
Median: 470
Max: 2,000
Median: 290
Max: 4,600
November 2012
4-15
Draft - Do Not Cite or Quote
-------
1 Several studies suggested the infiltration of Pb dust into buildings. For example,
2 Caravanos et al. (2006b) collected dust on glass plates at an interior location near an open
3 window, a sheltered exterior location, and an open exterior location for a two-year period
4 in Manhattan, NY. Median weekly dust loading was reported to be 52 (ig/m2 for the
5 indoor site, 153 (ig/m2 for the unsheltered outdoor site, and 347 (ig/m2 for the sheltered
6 outdoor site. This paper demonstrated the likely role of outdoor Pb in influencing indoor
7 dust Pb loading and indicated that under quiescent conditions (e.g., no cleaning) near an
8 open second-story window, the indoor dust Pb level might exceed EPA's hazard level for
9 interior floor dust of 430 ug/m2 (40 ug/ft2). Khoder et al. (2010) used the same
10 methodology to study Pb dust deposition in residential households in the town of Giza,
11 Egypt, located between two industrial areas and where leaded gasoline is still in use; the
12 investigators reported a median weekly deposition rate of 408 (ig/m2 and an exterior
13 median deposition rate of 2,600 (ig/m2. In the latter study, Pb deposition rate correlated
14 with total dust deposition rate (R=0.92), Cd deposition rate (R=0.95), and Ni deposition
15 rate (R=0.90). Statistically significant differences in Pb deposition rates were observed
16 between old and new homes (p <0.01) in the Khoder et al. (2010) study, although the
17 only quantitative information provided regarding home age stated that the oldest home
18 was 22 years old when the study was performed in 2007. Khoder et al. (2010) found no
19 statistically significant difference between Pb loadings when segregating the data by
20 proximity to roadways. Recently, Brattin and Griffin (2011) performed linear regressions
21 of dust Pb on soil Pb based on data collected previously for outdoor soil Pb and indoor
22 dust Pb near mining and/or smelting Superfund sites in Utah, Colorado, and Montana
23 (U.S. EPA. 2005f: SRC. 2002: U.S. EPA. 2001). They observed that the dust Pb
24 concentration was 4-35% of outdoor soil Pb. Excluding outliers on the regression, dust
25 Pb concentration ranged from 160-2,000 mg/kg, 11-660 mg/kg, and 68-1,000 mg/kg at
26 three sites.
27 Correlations between indoor and outdoor Pb content in dust can be partially explained
28 with speciation. Beauchemin et al. (2011) used XANES to speciate in-home paint
29 samples to assess the contributions of indoor paint and outdoor material to indoor dust Pb
30 concentrations. In indoor dust samples of particles <150 (im in size, Pb oxide, Pb sulfate,
31 and Pb carbonate were measured. These materials commonly were used in white paint. In
32 the size fraction of particles <36 (im, half of the measured Pb was associated with
33 Fe-oxyhydroxides such as ferrihydrite and goethite and presumably adsorbed onto these
34 species. This finding suggested that a mix of indoor and outdoor sources may affect the
35 composition of dust in the smaller size fraction in houses with leaded paint.
36 Residual Pb dust contamination following cleaning activities has been documented. For
37 instance, Hunt et al. (2008) estimated Pb deposition and concentration from experiments
38 in which Herculaneum, MO yard soil samples that had been dried, ground, and sieved
November 2012 4-16 Draft - Do Not Cite or Quote
-------
1 were tracked onto a tile test surface and then repeatedly cleaned until visual inspection of
2 the tiles uncovered no surface discoloration. Cleaning resulted in a 5-6 fold decrease in
3 residual Pb, with 7,100 (ig/m2 measured after multiple walks across the sample floor prior
4 to cleaning. Yu et al. (2006) analyzed dust samples from 50 homes in northern
5 New Jersey (typically of older housing stock, although the study does not specify housing
6 age). The investigators found that total Pb concentration in carpet dust ranged from 209
7 to 1,770 mg/kg dust. Wilson et al. (2007) studied Pb dust samples from homes in
8 Milwaukee, WI, and in resident children with and without elevated blood Pb > 10 (ig/dL.
9 They found that Pb dust samples obtained from the floor were always significantly higher
10 in residences of children with elevated blood Pb, with the exception of samples from the
11 bathroom floor. Windowsill dust was not significantly higher in residences of children
12 with elevated blood Pb. Residual Pb dust in homes is a potential exposure source for
13 small children who use touch to explore their environments.
14 Pb dust on floors, windowsills, and other accessible surfaces is related to several
15 demographic, socioeconomic, and housing conditions. Gaitens et al. (2009) used National
16 Health and Nutrition Examination Survey (NHANES) data from 1999 through 2004 to
17 examine Pb in dust in homes of children ages 12-60 months. Floor Pb dust loading value
18 was modeled against several survey covariates and was significantly associated with
19 several covariates but with mixed sign (p <0.05). Floor Pb dust was positively associated
20 with windowsill Pb dust loading, being of non-Hispanic black race/ethnicity, and
21 presence of smokers in the home. Floor Pb dust was negatively associated with presence
22 of carpeting, poverty-to-income ratio, and living in a home built after 1950. It was nearly
23 significantly and positively associated (p = 0.056) with renovations made to pre-1950
24 homes. Windowsill Pb dust level was also significantly associated (p <0.05) with several
25 covariates. It was positively associated with being of non-Hispanic black race/ethnicity,
26 negatively associated with living in a home built after 1950, positively associated with
27 not smooth and cleanable window surface condition, positively associated with presence
28 of smokers in the home, and positively associated with deterioration of indoor paint. It
29 was nearly statistically significantly and positively associated (p = 0.076) with
30 deterioration of outdoor paint when homes were built prior to 1950. Dust Pb loading was
31 found by Egeghy et al. (2005) to be significantly and positively associated with the log-
32 transform of soil Pb concentration, cement content in the home entryway, frequency of
33 fireplace usage, and homes built before 1950. Dust Pb loading was significantly and
34 negatively associated with indoor pesticide use and number of hours in which windows
35 were open (Table 4-3).
36 Building demolition and renovation activities can create dust from interior and exterior
37 paints with Pb content. Mielke and Gonzales (2008) measured Pb content in paint chips
38 from paint applied prior to 1992 and found that median Pb levels were 420 mg/kg for
November 2012 4-17 Draft - Do Not Cite or Quote
-------
1 interior paint and 77,000 mg/kg for exterior paint. Maximum levels were 63,000 mg/kg
2 and 120,000 mg/kg for interior and exterior paint, respectively. Mielke et al. (2001)
3 compared dust samples from two New Orleans houses that were prepared for painting.
4 One home was power sanded without any confinement or control of removed material,
5 while the other was hand-scraped with containment and collection of paint chips.
6 Immediately after sanding, Pb dust samples ranged from <3 to 28,000 mg/kg at the
7 sanded house. Pb dust samples from the scraped house ranged from 7 to 1,200 mg/kg.
8 Dust Pb concentrations have also been reported for homes in the vicinity of historic and
9 active metals mining and smelting sources. As described in Section 3.6.1. soil Pb has
10 been found to be elevated near source of ambient air Pb. Near an active smelter in Port
11 Pirie, Australia, median hand dust Pb loadings increased with age among a cohort of
12 fourteen children followed over age 0-36 months (2-5 months: 54 (ig/m2, >15 months:
13 336 (ig/m2) (Simon et al.. 2007). Zota et al. (2011) studied Pb dust and indoor Pb-PM2 5
14 concentration in Ottawa County, OK near the Tar Creek Superfund Site, in which a
15 metals mine had closed. Statistically significant correlations among outdoor soil Pb
16 concentration, indoor dust Pb concentration, indoor dust Pb loading, and indoor air
17 Pb-PM2 5 concentrations were observed (r = 0.25-0.65), with an average dust Pb
18 concentration of 109 mg/kg, dust Pb loading of 54 (ig/m2, soil Pb concentration of
19 201 mg/kg, and indoor Pb-PM2 5 concentration of 1 ng/m3. House dust Pb concentrations
20 were found to increase significantly with residential proximity to two chat (i.e., dry
21 mining waste) sources and to decrease with distance to the street and presence of central
22 air conditioning. Spalinger et al. (2007) measured Pb in dust in homes in a 34 km2 area
23 surrounding a designated Superfund site where a Pb and Zn smelter formerly operated at
24 Bunker Hill, ID. During spring of 1999, vacuum and floor mat samples were taken from
25 homes in three towns within the 34 km2 area and five "background" towns further from
26 the Superfund site. For the background towns, Pb concentration in vacuum dust had a
27 median of 120 mg/kg, and Pb concentration in floor dust had a median of 95 mg/kg. The
28 median Pb dust loading rate was measured to be 40 (ig/m2 per day. In contrast, Pb in
29 vacuum dust and floor mats for the towns contained within the Bunker Hill Superfund
30 site had a median Pb concentration of 470 mg/kg and 290 mg/kg, respectively. The
31 median Pb loading rate for indoor dust in houses in these towns was 300 (ig/m2 per day.
32 These results suggest that those living in close proximity to large Pb and Zn smelters or
33 mines that are now Superfund sites are at much greater risk of exposure to Pb dust
34 compared to the general population.
35 Pb exposure has been reported on children's playgrounds. Mielke et al. (2011 a) reported
36 median soil Pb concentration of 558 mg/kg on playground soils at eleven New Orleans
37 day care or community centers. Following remediation efforts to cover playground soil
38 with clean soil, median concentration dropped to 4.1 mg/kg. Duggan et al. (1985)
November 2012 4-18 Draft - Do Not Cite or Quote
-------
1 reported on the concentration and size distribution of wipe samples on the hands of 368
2 pre-school children from eleven schools in London, U.K.. Hand Pb residue (PbH) values
3 were modeled as linear (p <0.05) and power functions (p <0.001) of Pb dust; linear slope
4 was 0.0064 (ig hand Pb residue per mg/kg Pb dust. Given that the Duggan et al. (1985)
5 study was performed when Pb additives were used in gasoline, dust Pb concentration
6 values are not reported here. As described in Section 4.1.1.1. exposure to Pb in soil and
7 dust may be related to size distribution of the soil or dust particles, with higher Pb
8 enrichment in the smaller particles.
4.1.3.3 Dietary Pb Exposure
9 This subsection covers several dietary Pb exposures from a diverse set of sources.
10 Included among those are drinking water, fish and meat, agriculture, urban gardening,
11 dietary supplements, tobacco, cultural food sources, and breastfeeding. The breadth of
12 dietary Pb exposures is illustrated in Figure 4-2. which illustrates the data obtained in the
13 2008 FDA Total Diet Study market basket survey (FDA. 2008). Among the highest Pb
14 concentrations were those for noodles, baby food carrots, baby food oatmeal, Swiss
15 cheese, beef tacos from a Mexican restaurant, and fruit-flavored cereal. Possible sources
16 of Pb in food samples include introduction during processing or preparation with drinking
17 water contaminated with Pb, deposition of Pb onto raw materials for each food, and Pb
18 exposure in livestock that produce dairy or meat ingredients. Manton et al. (2005) used
19 Pb isotope ratios to estimate sources of dietary Pb among a cohort of mothers and
20 children from Omaha, NE using a combination of food samples, hand wipes, house dust
21 wipes, and aerosol samples collected between 1990 and 1997. Drinking water Pb was not
22 included in this study. The authors cited results from Egan et al. (2002) that imported
23 vegetables contributed 55% of Pb dietary intake for infants, 30% for 2-6 year old
24 children, and 20% for 25-30 year old women. Imported candy contributed 10% of Pb
25 dietary intake for 2-6 year old children and 9% for 25-30 year old women. Isotopic data
26 from Manton et al. (2005) suggested that, with the exception of children age 0-12 mos,
27 house dust is a large contributor to dietary Pb. The pattern of certain Pb-isotope ratios
28 observed in the diet of children 0-12 mos are suggested to derive from Ca salts in
29 limestone that may have been used in dietary supplements in baby formula. The
30 contribution of ambient Pb aerosols to dietary Pb samples was not statistically significant
31 for this urban exposure study.
November 2012 4-19 Draft - Do Not Cite or Quote
-------
IL 1
INQOQISS ~~
BF(carrot) -
BF(oatmeal) -
Cheese -
Taco —
Cereal -
Egg'n'chs —
Peas -
Milk(2%) -
BF(grnbean) —
Cabbage -
BF(lamb) -
Beans —
Rice -
Coffee -
Strawberries —
Squash —
Potato —
Chicken -
Biscuits —
BF(cobbler) -
BF(beef) -
L J 1
M 1
i— i H
ED
r-|~|H
\L
h[J}--H
i nil
i ii
rflO
EH
\L
r| | |- - H
(0
h|~~f1-H
1 1 1 1
u
EH
i i i i i i i
0.00 0.02 0.04 0.06 0.08 0.10 0.12 014
Concentration (mg/kg)
Note: from the 2008 FDA Total Diet Study. "BF" denotes baby food.
Source Data: (FDA. 2008)
Figure 4-2
Market basket survey results for Pb concentration in foods.
November 2012
4-20
Draft - Do Not Cite or Quote
-------
Drinking Water
1 Pb concentrations in drinking water vary substantially. For example, Shotyk and Krachler
2 (2009) measured the Pb concentration in tap water, commercially bottled tap water and
3 bottled natural water. They found that, in many cases, tap water contained less Pb than
4 bottled water. Excluding bottled water in glass containers because Pb can be leached
5 from the glass, the median Pb concentration in the bottled water samples was 8.5 ng/L
6 (range < 1 to 761 ng/L). Pb in drinking water supplies can derive from atmospheric
7 deposition onto surface waters, runoff of atmospheric deposition as described in Section
8 3.3. or via corrosion of Pb in the distribution network exacerbated by contact with acidic
9 disinfection byproducts, as described in the following paragraphs.
10 It is now recognized that environmental nanoparticles (NPs) (-1-100 nm) can play a key
11 role in determining the chemical characteristics of treated drinking water as well as
12 natural waters (Wigginton et al., 2007). An important question is whether or not NPs
13 from source waters affect the quality of drinking water. For example, if Fe-oxide NPs are
14 not removed during the flocculation/coagulation stage of the treatment process, they may
15 become effective transporters of contaminants such as Pb, particularly if these
16 contaminants are leached from piping in the distribution system.
17 Corrosion byproducts can influence Pb concentrations in drinking water. Schock et al.
18 (2008) characterized Pb pipe scales from 91 pipes made available from 26 different
19 municipal water systems from across the northern U.S. They found a wide range of
20 elements including Cu, Zn and V as well as Al, Fe and Mn. Interestingly, V was present
21 at nearly one percent levels in pipes from many geographically diverse systems. In a
22 separate study, Gerke et al. (2009) identified the corrosion product, vanadinite
23 (Pb5(VO4)3Cl) in Pb pipe corrosion byproducts collected from 15 Pb or Pb-lined pipes
24 representing 8 different municipal drinking water distribution systems in the Northeastern
25 and Midwest regions of the U.S. Vanadinite was most frequently found in the surface
26 layers of the corrosion products. The vanadate ion, VO43", essentially replaces the
27 phosphate ion in pyromorphite and hydroxyapatite structures. It is not known whether the
28 application of orthophosphate as a corrosion inhibitor would destabilize vanadinite, but
29 this substitution would have implications for V release into drinking water. The stability
30 of vanadinite in the presence of monochloramine is also not known, and its stability
31 might have implications for both Pb and V release into drinking water.
32 In recent years, drinking water treatment plants in many municipalities have switched
33 from using chlorine to other disinfecting agents because their disinfection byproducts
34 may be less carcinogenic. However, chloramines are more acidic than chlorine and can
35 increase Pb solubility (Raab et al.. 1991) and increase Pb concentrations in tap water. For
36 example, after observing elevated Pb concentrations in drinking water samples, Kim and
November 2012 4-21 Draft - Do Not Cite or Quote
-------
1 Herrera (2010) observed Pb oxide corrosion scales occurring after using acidic alum as a
2 disinfection agent. Edwards and Dudi (2004) observed a red-brown particle-bound Pb in
3 Washington, B.C. water that could be confused with innocuous Fe. The source of the
4 particle-bound Pb was not known but was thought to originate from the source water. The
5 high Pb concentrations were attributed to leaching of Pb from Pb-bearing pipes promoted
6 by breakdown products of disinfection agents (Edwards and Dudi. 2004). Maas et al.
7 (2007) tested the effect of fluoridation and chlorine-based (chlorine and chloramines)
8 disinfection agents on Pb leaching from plumbing soldered with Pb. When using chlorine
9 disinfection agents alone, the Pb concentration in water samples doubled during the first
10 week of application (from 100 to 200 ppb) but then decreased over time. When adding
11 fluorosilicic acid and ammonia, the Pb concentration spiked to 900 ppb and increased
12 further over time. However, Macek et al. (2006) regressed blood Pb among children ages
13 1-16 years on fluoride treatment, adjusted for several demographic and socioeconomic
14 factors, and found no association when all data were combined into one model; when
15 stratifying by housing age, Macek et al. (2006) found statistically significant odds ratios
16 for those living in housing built before 1946 or for housing age unknown. Similarly,
17 Lasheen et al. (2008) observed Pb leaching from pipes in Egypt when exposed to an acid
18 of pH = 6. Exposure to basic solutions actually resulted in reduction of Pb concentration
19 in the drinking water. Leaching of Pb from pipes following disinfection with acidic
20 agents can lead to increased Pb exposure; Miranda et al. (2007b) observed a statistically
21 significant association between blood Pb levels among children living in Wayne County,
22 NC and use of chloramines (p <0.001) in a log-linear model, although the study did not
23 control for the presence of Pb paint in the dwellings, so it is difficult to distinguish the
24 influence of Pb pipes from Pb in paint on blood Pb levels.
25 Several chemical mechanisms may contribute to release of Pb during use of chloramine
26 disinfection agents. Edwards and Dudi (2004) hypothesized that Pb leaching occurs when
27 chloramines cause the breakdown of brass alloys and solder containing Pb. After
28 observing that nitrification also leads to increased Pb concentrations in water, they also
29 proposed that chloramines may trigger nitrification and hence cause decreasing pH,
30 alkalinity, and dissolved oxygen that leads to corrosion after observing that nitrification
31 also leads to increased Pb concentrations in water. However, Zhang et al. (2009b) found
32 no evidence that nitrification brought about significant leaching of Pb from Pb pipes.
33 Lytle et al. (2009) suggested that a lack of increased Pb(II) concentrations in drinking
34 water following a change from free chlorine to chloramine disinfection is attributed to the
35 formation of the Pb(II) mineral hydroxypyromorphite (Pb5(PO4)3OH) instead of Pb(IV)
36 oxide. Xie et al. (2010) further investigated the mechanisms by which Pb(II) release is
37 affected by chloramines. Two opposing mechanisms were proposed: Pb(IV)O2 reduction
38 by an intermediate species from decomposition of monochloramine; and increasing redox
39 potential which decreases the thermodynamic driving force for reduction. They suggest
November 2012 4-22 Draft - Do Not Cite or Quote
-------
1 that the contact time of monochloramine with PbO2 and the C12:N ratio in
2 monochloramine formation will determine which mechanism is more important. Free
3 chlorine can control Pb concentrations from dissolution under flowing conditions but for
4 long stagnation periods, Pb concentrations can exceed the action level: 4-10 days were
5 required for Pb concentrations to exceed 15 (ig/L (for relatively high loadings of PbO2 of
6 1 g/L). Thus, under less extreme conditions, it was concluded that chloramination was
7 unlikely to have a major effect on the release of Pb into drinking water.
Agriculture
8 The 2006 Pb AQCD (2006b) states that surface deposition "represents a significant
9 contribution to the total Pb in and on the plant", while uptake through a plant's roots can
10 also contribute to a plant's Pb concentration. Consequently, Pb content in plants may
11 contribute to human dietary exposure. Uptake of Pb by plants growing in contaminated
12 soil has been repeatedly demonstrated in some species during controlled potted plant
13 experiments (Del Rio-Celestino et al.. 2006). In this study, most species retained Pb in
14 the roots with little mobilization to the shoots of the plants. However, certain species of
15 grasses were able to mobilize Pb from the roots to the shoots of the plant; these specific
16 species could lead to human exposures through consumption of grazing animals. Lima et
17 al. (2009) conducted similar greenhouse experiments with several vegetable crops grown
18 in soil contaminated by Pb-containing residue from battery recycling waste. In this study,
19 carrots had high bioaccumulation, measured as the percent of Pb concentration measured
20 in the plant compared with the Pb concentration in the soil, with little translocation of the
21 Pb to the shoots. Conversely, beets, cabbages, sweet peppers, and collard greens had low
22 bioaccumulation but moderate to high translocation. Okra, tomatoes, and eggplants had
23 moderate bioaccumulation and moderate to high translocation. Sesli et al. (2008) also
24 noted uptake of Pb within wild mushrooms. Vandenhove et al. (2009) reviewed
25 bioaccumulation data for plant groupings and found that grasses had the highest uptake,
26 followed by leafy vegetables and root crops grown in sandy soils; see Table 4-6. These
27 references also suggested high transfer from roots to shoots among root crops, with
28 shoots having roughly four times higher Pb bioaccumulation than roots.
29 Sources of atmospheric Pb can lead to vegetable contamination. For example, Uzu et al.
30 (2010) found that Pb deposition from smelter emissions caused a linear increase in Pb
31 concentrations of 7.0 mg/kg per day (R2=0.96) in lettuce plants cultivated in the
32 courtyard of a smelter. They reported that lettuce grown 250-400 meters from the smelter
33 had concentrations that were 10-20 times lower, which is consistent with findings
34 described in Section 3.3 that deposition of Pb containing material drops off with distance
35 from a source. Pb contamination of crops may also occur through piston-engine aircraft
36 Pb emissions during aerial application of fertilizers and pesticides. In 2010, the U.S.
November 2012 4-23 Draft - Do Not Cite or Quote
-------
1 Federal Aviation Administration (FAA) recorded 396,000 hours of flight time for aerial
2 application. This term encompasses crop and timber production including seeding
3 cropland and fertilizer and pesticide application. It is estimated that 86% of these flight-
4 hours involved piston engine aircraft utilizing leaded fuel (FAA. 2010).
5 Some land use and soil characteristics have been shown to increase bioaccessibility of Pb
6 in soil, which could then lead to plant contamination. Fernandez et al. (2010; 2008; 2007)
7 measured Pb from atmospheric deposition in two adjacent plots of land having the same
8 soil composition but different uses: one was pasture land and one was agricultural. In the
9 arable land, size distributions of soil particle-bound Pb, were uniformly distributed. In
10 pasture land, size distributions of soil particle-bound Pb were bimodal with peaks around
11 2-20 urn and 50-100 urn (Fernandez etal., 2010). For the agricultural plot, Pb
12 concentration was constant around 70 mg/kg in samples taken over the first 30 cm of soil,
13 at which time it dropped below 10 mg/kg at soil depths between 35 and 100 cm. In
14 contrast, Pb concentration in pasture land peaked at a depth of 10 cm at a concentration
15 of roughly 70 mg/kg and then dropped off gradually to approach zero concentration at a
16 depth of approximately 50 cm. The sharp change in concentration for the arable land was
17 attributed to a combination of plowing the soil and use of fertilizers to increase the
18 acidity of the soil and solubility of Pb into the soil (Fernandez et al.. 2007). They found
19 that the surface layer was acidic (pH: 3.37-4.09), as was the subsurface layer (pH:
20 3.65-4.38). Jin et al. (2005) examined how soil characteristics affect Pb contamination of
21 crops by testing soil Pb, bioaccessibility of soil Pb (determined by CaCl2 extraction), and
22 Pb in tea samples from tea gardens. They observed that the Pb concentration in tea leaves
23 was proportional to the bioaccessible Pb in soil.
24 There is some evidence that Pb contamination of crops can originate with treatment of
25 crops. For example, compost produced from wastewater sludge has the potential to add
26 Pb to crops. Cai et al. (2007) demonstrated that production of compost from sludge
27 enriched the Pb content by 15-43% compared with the Pb content in sludge prior to
28 composting. Chen et al. (2008b) observed that the median concentration of Pb in
29 California crop soil samples was 16.2 mg/kg (range: 6.0-62.2 mg/kg). Chen et al. (2008a)
30 further observed that in three of the seven California agricultural regions sampled,
31 concentrations of Pb increased following addition of fertilizer, but the increase was less
32 than that for phosphorous (P) and Zn indicators of fertilizer. In four regions, there was no
33 increase of Pb at all. Furthermore, Tu et al. (2000) observed a decrease in Pb fraction
34 with increasing P application. Nziguheba and Smolders (2008) also surveyed phosphate-
35 based fertilizers sold in European markets to determine the contribution of these
36 fertilizers to heavy metal concentrations in agricultural products. They reported a median
37 fertilizer Pb concentration of 2.1 mg/kg based on total weight of the fertilizer, with a 95th
38 percentile concentration of 7.5 mg/kg. Across Europe, Nziguheba and Smolders (2008)
November 2012 4-24 Draft - Do Not Cite or Quote
-------
1 estimated that the amount of Pb applied via fertilizers to be only 2.6% of that resulting
2 from atmospheric deposition.
3 Although Pb in on-road vehicle gasoline has been phased out in the U.S., if imported
4 crops are produced in countries that still use Pb antiknock agents in on-road gasoline,
5 they have the potential to introduce dietary Pb to U.S. consumers. For example, high
6 concentrations of Pb have been found in chocolate from beans grown in Nigeria, during
7 the time when leaded gasoline was still legally sold. Rankin et al. (2005) observed that
8 the ratios of 207Pb to 206Pb and 208Pb to 207Pb were similar to those of Pb in gasoline.
9 Although this study showed that Pb concentration in the shelled cocoa beans was low (~1
10 ng/g), manufactured cocoa powder and baking chocolate had Pb concentrations similar to
11 those of the cocoa bean shells, on the order of 200 ng/g, and Pb concentration in
12 chocolate products was roughly 50 ng/g (Rankin et al.. 2005). It is possible that the
13 increases were attributed to contamination of the cocoa by the shells during storage or
14 manufacture, but the authors note that more research is needed to verify the source of
15 contamination.
16 Findings from Pb uptake studies have implications for urban gardening if urban soils may
17 be contaminated with Pb. For instance, Clark et al. (2006) tested the soil in 103 urban
18 gardens in two Boston neighborhoods. Using isotopic analysis, they found that Pb-based
19 paint contributed 40-80% of Pb in the urban garden soil samples, with the rest coming
20 from historical gasoline emissions. Furthermore, Clark et al. (2006) estimated that Pb
21 consumption from urban gardens can be equivalent to 10-25% of the exposure to Pb from
22 drinking water for children living in the Boston neighborhoods studied. Because soil Pb
23 levels in urban areas will depend on surrounding sources (Pruvot et al.. 2006). Pb
24 exposures in urban garden vegetables will vary.
November 2012 4-25 Draft - Do Not Cite or Quote
-------
Table 4-6 Pb bioaccumulation data for various plants. Bioaccumulation is
expressed as percent of Pb concentration in the plant to the Pb
concentration in the soil.
Plant
Plant Group Compartment
All
Cereals Grain
Straw
Maize Grain
Straw
Rice Grain
Leafy Vegetables
Non-Leafy Fruits
vegeiaoies ^^
Legumes Pods
Shoots
Root Crops Roots
Shoots
Tubers Tubers
Fruits Fruits
Leaves
Grasses
Natural Pastures
Leguminous Fodder
All Cereals
Pastures/Grasses
Fodder
Soil
All
All
All
All
All
All
Sand
Loam
Clay
All
All
All
Sand
Loam
Clay
All
All
Sand
Loam
All
All
Sand
Loam
All
All
All
All
All
All
Sand
Loam
Clay
All
All
Sand
Clay
n
210
9
4
9
3
2
31
4
3
7
5
2
17
3
5
4
1
27
5
5
12
30
5
17
5
1
17
34
1
20
5
8
6
51
24
4
4
GM
2.0%
1.0%
2.3%
0.12%
0.28%
8.0%
7.3%
82%
2.8%
1.5%
0.53%
0.27%
0.14%
0.080%
1.5%
6.4%
2.3%
6.3%
0.15%
0.64%
0.052%
0.77%
31%
92%
0.43%
0.61%
0.17%
0.90%
14%
2.5%
4.5%
0.82%
GSD
14
3.6
3.5
2.3
6.6
13
1.5
1.0
4.1
26
12
3.2
4.4
1.0
16
1.6
4.7
15
7.4
3.5
2.4
2.6
1.8
4.8
4.7
5.3
3.9
4.0
4.2
12
2.3
5.7
AM
63%
1.8%
3.8%
0.17%
0.85%
2.2%
210%
7.8%
82%
5.1%
78%
0.88%
34%
0.42%
0.42%
0.33%
0.080%
41%
7.0%
0.50%
250%
9.1%
1.2%
0.073%
1.0%
25%
36%
23%
1.6%
1.1%
1.3%
0.53%
1.8%
27%
130%
5.6%
2.7%
SD
290%
1.6%
4.0%
0.14%
1.3%
1.4%
610%
3.3%
3.5%
4.8%
170%
0.42%
120%
0.34%
0.34%
0.47%
98%
3.4%
0.68%
570%
48%
1.6%
0.062%
0.60%
22%
29%
1.4%
1.3%
1.1%
1.8%
27%
420%
4.0%
4.6%
Min
0.015%
0.19%
0.51%
0.052%
0.060%
1.2%
0.32%
4.9%
79%
0.41%
0.15%
0.58%
0.046%
0.065%
0.065%
0.046%
0.024%
4.2%
0.024%
0.30%
0.015%
0.16%
0.015%
0.15%
11%
0.22%
0.052%
0.052%
0.059%
0.22%
0.22%
0.060%
1.6%
0.16%
Max
2,500%
4.8%
9.6%
0.38%
2.3%
3.2%
2,500%
11%
86%
12%
390%
1.17%
490%
0.89%
0.89%
1.0%
330%
12%
1.7%
16%
260%
3.9%
0.23%
1.7%
100%
100%
4.8%
3.2%
3.2%
4.8%
100%
1 ,600%
11%
9.6%
Source: Reprinted with permission of Elsevier Publishers, Vandenhove et al.
November 2012
4-26
Draft - Do Not Cite or Quote
-------
Game
1 Atmospheric sources of Pb have also been shown to contaminate game meat, thus
2 potentially posing a risk of Pb exposure. In Pb mining or smelting areas, several studies
3 have documented Pb concentrations in game [e.g., (Nwude et al., 2010; Reglero et al.,
4 2009b)1.
5 Potential Pb exposure through consumption of animals exposed to or killed with Pb shot
6 has also been well documented (Hunt et al., 2009; Tsuji et al., 2009; Tsuji et al., 2008;
7 Hunt et al.. 2006). For example, Martinez-Haro et al. (2010) observed Pb in the feces of
8 mallards that ingested gunshot of 34-13,930 mg/kg with a median of 1,104 mg/kg, while
9 mallards that did not ingest gunshot had feces Pb levels <12.5 mg/kg. Mateo et al. (2011)
10 studied Pb bioaccessibility as a function of cooking method for breast meat from
11 partridges killed with gunshot. They observed that preparation in cold or hot vinegar
12 increased bioaccessibility compared with total Pb in the samples.
Fish
13 Pb content in fish could also lead to human exposure to Pb (U.S. EPA. 2006b. 1986a).
14 Ghosh et al. (2007) demonstrated in laboratory experiments that exposure to Pb in water
15 can lead to linearly increasing Pb levels in the kidneys, liver, gills, skeleton, and muscle
16 offish. Several studies have documented the potential for human Pb exposure through
17 fish and seafood. Welt et al. (2003) conducted a survey of individuals who fished in
18 Bayou St. John, Louisiana in conjunction with sampling Pb content in sediment. They
19 found that median sediment Pb concentrations ranged from 43 to 330 mg/kg in different
20 locations, while maximum sediment Pb concentrations ranged from 580 to 6,500 mg/kg.
21 In total, 65% of the surveyed individuals fished for food from the Bayou, with 86%
22 consuming fish from the Bayou each week. In a study of the effect of coal mining on
23 levels of metals in fish (measured as blood Pb) in northeastern Oklahoma, Schmitt et al.
24 (2005) found that fish blood Pb levels varied with respect to species of fish, but blood Pb
25 levels were higher in fish in areas close to mining activities. Similarly, Besser et al.
26 (2008) observed higher levels offish blood Pb close to mining activities in southeastern
27 Missouri. In a related study offish species in the same region of Missouri, fish blood Pb
28 levels were found to be statistically significantly higher in sites within 10 km downstream
29 of active Pb-Zn mines (p <0.01) compared with fish located further from the mines
30 (Schmitt et al.. 2007a). and elevated fish blood Pb levels were again noted near a Pb-Zn
31 mine (Schmitt et al.. 2009). It was noted that the Ozark streams where these studies were
32 performed were often used for recreational fishing.
November 2012 4-27 Draft - Do Not Cite or Quote
-------
Breast Milk
1 Studies of breastfeeding women suggest that infants may be exposed to Pb in breast milk.
2 Ettinger et al. (2004a) observed in a 1994-1995 study of Mexico City women that at
3 1 month postpartum, 88 women breastfeeding exclusively (with mean blood Pb level of
4 9.4 (ig/dL) had breast milk Pb concentrations of 1.4 ± 1.1 (ig/L, and 165 women
5 breastfeeding partially (with mean blood Pb level of 9.5 (ig/dL) had breast milk Pb
6 concentrations of 1.5 ± 1.2 (ig/L. During the same time period, Ettinger et al. (2006)
7 studied breastfeeding women in Mexico City over a child's first year of life and sampled
8 Pb concentration in breast milk at 1, 4, and 7 mo post-partum. They observed that mean
9 breast milk concentrations dropped from 1.4 (ig/L at 1 mo (mean maternal blood
10 Pb = 9.3 (ig/dL) to a mean of 1.2 (ig/L at 4 mo (mean maternal blood Pb = 9.0 (ig/dL) to
11 0.9 (ig/L at 7 mo (mean maternal blood Pb = 8.1 (ig/dL); this reduction was statistically
12 significant (p <0.00001). Among the 310 women included in the study, 181 had previous
13 pregnancies. In one study of nursing mothers living in Port Pirie, Australia near a Pb
14 smelter, 10 of the 11 mothers had breast milk concentrations <5 (ig/L (Simon et al..
15 2007). The authors hypothesized that breast milk concentration was too low to be a major
16 contributor to blood Pb level in these infants relative to other factors such as hand loading
17 of Pb. However, one mother with a blood Pb level of 25 (ig/dL had a breast milk Pb level
18 of 28 (ig/L (Simon et al.. 2007).
19 In summary, several sources of dietary Pb can originate from atmospheric Pb emissions,
20 including drinking water, vegetables, game, fish, and breast milk. Drinking water Pb
21 levels are affected by source strength and proximity, runoff, and water treatment
22 processes and chemicals. Among plants grown for agriculture, Pb content is highest in
23 grasses, followed by leafy vegetables, then root vegetables. Pb in soil or dust can also
24 collect on the surfaces of vegetables. Pb contamination of vegetables depends on a
25 number of factors, including presence of nearby sources of atmospheric Pb, soil type and
26 chemistry, land use, and land treatment. Other sources of Pb, such as international
27 consumer products or historic emissions, also have the potential to introduce Pb into the
28 U.S. diet. Pb contamination through the food chain potentially leads to elevated Pb levels
29 in meat. Likewise, Pb contamination of surface waters can lead to elevated levels of Pb in
30 fish used for consumption. Breastfeeding also presents a potential Pb exposure to
31 newborn babies, and that exposure drops off as the mothers nurse and as the babies age
32 and add more food to their diet.
November 2012 4-28 Draft - Do Not Cite or Quote
-------
4.1.3.4 Occupational
1 Occupational environments have the potential to expose individuals to Pb. Some modern
2 day occupational exposures are briefly discussed below in the context of understanding
3 potential exposures that are not attributed to ambient air. For example, Miller et al.
4 (2010) obtained personal and area samples of particle-borne Pb in a precious metals
5 refinery; year of the study was not reported. It was not stated explicitly, but it is likely
6 that Miller et al. (2010) measured the PM as TSP because the Occupational Safety and
7 Health Administration (OSHA) permissible exposure limit (PEL) for Pb is based on TSP
8 rather than a smaller size cut, and the OSHA PEL was used for comparison.
9 Concentrations measured by personal samples ranged from 2 to 6 (ig/m3, and
10 concentrations from area samples ranged from 4 to 14 (ig/m3. The OSHA PEL is 5 (ig/m3.
11 In steel production, sintering was found to be the largest source of airborne Pb exposure
12 in a survey of operations (Sammut et al.. 2010). with Pb enrichment in PM reported to be
13 20,000 mg/kg. Although total PM concentration was not reported by the authors, the PM
14 was reported to have 75% of its particulate mass at below the 2.5 (im diameter size.
15 Operations involving Pb-containing materials in various industries are a source of
16 occupational Pb exposure, in addition to a residential exposure. Rodrigues et al. (2010)
17 reported exposures to airborne Pb among New England painters, who regularly use
18 electric grinders to prepare surfaces for painting. Two-week averaged airborne Pb
19 concentrations, sampled with an Institute of Medicine inhalable PM sampler designed to
20 capture PM smaller than 100 (im, were reported to be 59 (ig/m3, with a maximum daily
21 value of 210 (ig/m3. The Pb concentrations reported here were corrected by the National
22 Institute for Occupational Safety and Health (NIOSH) respirator protection factors,
23 although the respirator protection factors were not reported by Rodrigues et al. (2010).
24 Information on the air Pb-blood Pb relationship can be found in Section 4.5.1.
4.1.3.5 Exposure to Pb from Consumer Products
25 Pb is present in varying amounts in several consumer products including alternative
26 medicines, candies, cosmetics, pottery, tobacco, toys, and vitamins (Table 4-7). Several
27 of these categories suggest children may incur regular exposures. Pb concentrations were
28 reported to range from non-detectable levels up to 77% by mass, for the case of one
29 medicinal product. Exposure to these products, which originate in a range of different
30 countries, can account for substantial influence on Pb body burden (Miodovnik and
31 Landrigan. 2009; Levin et al.. 2008).
November 2012 4-29 Draft - Do Not Cite or Quote
-------
Table 4-7 Pb content in various consumer products.
Product
Category
Alternative
and
Traditional
Medicines
Product
Cissus quadrangularis, Caulophyllum thalictroides,
Turners diffuse, Centella asiatica, Hoodia gordonii,
Sutherlandia frutescens, Curcuma longa,
fucoxanthin, Euterpe oleracea
(dietary supplements claimed to be from Hoodia
gordonii)
Malva sylvestris
Yugmijihwang-tang, Bojungigki-tang, Sibjeondaebo-
tang, Kuibi-tang, Ojeogsan
Lemongrass, licorice, holy basil, cloves, ginger
B-Success 28, Operation Sweep, Aloe Vera Plus
Location of
Purchase
U.S.
(Mississippi)3
Turkey
Korea
India
Nigeria
Pb Content (units)
Not detected (N.D.)b
to 4.21 mg/kg
1.1-2.0 mg/kg
7.9x1 0"6 to 2.5x1 0"5
mg/kg body weight/day
Average:
Lemongrass &
Holy Basil Leaves:
6.1 mg/kg;
Licorice Stolons:
6.1 mg/kg,
Clove Dried Flower
Buds: 7.8 mg/kg,
Ginger Rhizome:
5.8 mg/kg
925-27,000 ug
Reference
Avula et al.
(201 0)
Hicsbmnez et
al. (2009)
Kim et al.
(2009a)
Naithani and
Kakkar (2006)
Obi et al. (2006)
Bitter Aloes, Zarausmacine, Virgy-Virgy Computer
Worm-Expeller, Dorasine Powder, Sexual Energy,
U&DEE Infection Cleansing Powder, U&DEE Sweet
Bitter, Natural Power Stone, Chama Black Stone,
Portugal Antiseptic Soap, Edysol Antiseptic Soap,
H-Nal, M-Reg, Veins Flocher, Diabor, C-Candi, C-
Cysta, Firas, D-Diab, P-Pile, Infecta, Ribacin Forte,
Aloe Vera Cure Formula
Shell of Hen's Egg
Berberis (6. aristata, B. chitria, B. asiatica,
B. lyceum), Daruharidra
Greta powder
Candy Tamarind Candy
India
India
U.S.
(California)
U.S.
(Oklahoma)
1 4 mg/kg
Berberis:
Roots: 3.1-24.7 mg/kg
Stems: 8.0-23.8 mg/kg
Daruharidra:
16.9-49.8 mg/kg
770,000 ppm
Product: 0.15-3.61
mg/kg
Sharma et al.
(2009)
Srivastava et al.
(2006)
CDC (2002)
Lynch et al.
(2000)
Stems: 0.36-2.5 mg/kg
Wrappers: 459-27,125
mg/kg
Tamarind Candy
U.S.
(California)
Product: 0.2-0.3 mg/kg
Stems: 400 mg/kg
Wrappers:
16,000-21,000 mg/kg
Cosmetics
Lipsticks
Eye Shadows
U.S.
Nigeria
Average: 1.07 mg/kg
N.D. to 55 mg/kg
Heppet al.
(2009)
Omolaoye et al.
(201 Oa)
November 2012
4-30
Draft - Do Not Cite or Quote
-------
Product
Category
Pottery
Tobacco
Toys
Vitamins
Product
Foods prepared in Pb-glazed pottery
Smokeless Tobacco
Cigarette Tobacco (210Pb concentrations)
Red and yellow painted toy vehicles and tracks
535 PVC and non-PVC toys from day care centers
Soft plastic toys
Toy necklace
Soft plastic toys
Vitamins for young children, older children, and
pregnant or lactating women
Location of
Purchase
Mexico
U.K.
Pakistan
Brazil
U.S. (Nevada)
India
U.S.
Nigeria
U.S.
Pb Content (units)
N.D. to 3, 100 mg/kg
0.15-1.56mg/kg
Activity cone.: 7-20
Bq/kg
500-6,000 mg/kg
PVC: avg. 325 mg/kg
Non-PVC: avg. 89
mg/kg Yellow: 216
mg/kg
Non-yellow: 94 mg/kg
Average (by city):
21 -280 mg/kg
388,000 mg/kg
2.5-1 ,445 mg/kg
Average:
Young children:
2.9 ug/day
Older children:
1 .8 ug/day
Pregnant and lactating
women: 4.9 ug/day
Reference
Villalobos et al.
(2009)
McNeill et al.
(2006)
Tahir and
Alaamer (2008)
Godoi et al.
(2009)
Greenway and
Gerstenberger
(201 0)
Kumar and
Pastore (2007)
Meyer et al.
(2008)
Omolaoye et al.
(201 Ob)
Mindak et al.
(2008)
"Hoodia gordonii, from Eastern Cape, South Africa Euterpe oleracea from Ninole Orchard, Ninole, Hawaii
*Note that the country of origin is not provided because it was not published in the references cited.
4.2 Kinetics
1 This section summarizes the empirical basis for understanding Pb toxicokinetics in
2 humans. The large amount of empirical information on Pb biokinetics in humans and
3 animal models has been integrated into mechanistic biokinetics models (U.S. EPA.
4 2006b). These models support predictions about the kinetics of Pb in blood and other
5 selected tissues based on the empirically-based information about Pb biokinetics. In
6 Section 4.3 (and Section 4.2.2.1). Pb biokinetics is described from the context of model
7 predictions.
8 The discussion of Pb toxicokinetics emphasizes inorganic Pb since this comprises the
9 dominant forms of Pb to which humans in the U.S. are currently exposed as a result of
10 releases of Pb to the atmosphere and historic surface deposition of atmospheric Pb (see
11 Section 3.2.2). The toxicokinetics of organic Pb is only briefly described and a more
12 extensive discussion can be found in the 2006 Pb AQCD. Human exposures to organic
November 2012
4-31
Draft - Do Not Cite or Quote
-------
1 Pb could occur in occupational settings (e.g., during manufacturing of tetraethyl Pb or
2 aviation fuels); however, environmental exposures to organic Pb compounds rarely occur
3 in the U.S. other than in the limited circumstances of those involved in fueling piston-
4 driven aircraft that use leaded aviation gasoline.
4.2.1 Absorption
5 The major exposure routes of Pb in humans are inhalation and ingestion. Therefore, these
6 exposure routes are important in the discussion of Pb absorption (see Sections 4.2.1.1 and
7 4.2.1.2). The term "absorption" refers to the fraction of the amount of Pb ingested or
8 inhaled that is absorbed from the respiratory or gastrointestinal tract. The term
9 bioavailability, as it is used in this section, refers to the fraction of the amount of Pb
10 ingested or inhaled that enters the systemic circulation. If properly measured (e.g., time-
11 integrated blood Pb), under most conditions Pb bioavailability is equivalent (or nearly
12 equivalent) to Pb absorption. The time-integrated blood Pb (i.e., the integral of blood Pb
13 over time) provides a useful measure of bioavailability because it reflects both recent Pb
14 absorption as well as contributions from Pb sequestered in soft tissue and bone.
15 Bioaccessibility is a measure of the physiological solubility of Pb in the respiratory or
16 gastrointestinal tract. Pb must become bioaccessible in order for absorption to occur.
17 Processes that contribute to bioaccessibility include physical transformation of Pb
18 particles and dissolution of Pb compounds into forms that can be absorbed (e.g., Pb2+).
19 Bioaccessibility is typically assessed by measuring the fraction of Pb in a sample that can
20 be extracted into a physiological or physiological-like solution (e.g., gastric juice or
21 solution similar to gastric juice).
22 The 2006 Pb AQCD (U.S. EPA. 2006c) also presented dermal absorption of inorganic
23 and organic Pb compounds, which is generally considered to be much less than by
24 inhalation or ingestion. A study published subsequent to the 2006 Pb AQCD measured
25 rates of absorption of Pb in skin patches harvested from nude mice (PanetaL 2010).
26 Following application of 12 mg Pb as Pb acetate or Pb nitrate, the absorption rate
27 (measured over a 10-hour observation period) was approximately 0.02 (ig Pb/cm2 per
28 hour. Absorbed Pb was detected in liver and kidney of nude mice following a 120-hr
29 occluded dermal application of approximately 14 mg Pb as either Pb acetate or Pb nitrate.
30 Uptake of Pb into the skin at the site of application was greater when Pb acetate was
31 applied to the skin compared to Pb nitrate; however, liver and kidney Pb concentrations
32 observed at the conclusion of the study (120 hours following the application of Pb) were
33 not different for the two Pb compound. No additional information provides evidence of
34 dermal absorption being a major exposure route of environmental Pb.
November 2012 4-32 Draft - Do Not Cite or Quote
-------
4.2.1.1 Inhalation
1 Systemic absorption of Pb deposited in the respiratory tract is influenced by particle size
2 and solubility, as well as by the pattern of regional deposition within the respiratory tract.
3 Fine particles (<1 (im) deposited in the bronchiolar and alveolar region can be absorbed
4 after extracellular dissolution or can be ingested by phagocytic cells and transported from
5 the respiratory tract (Bailey and Roy. 1994). Larger particles (>2.5 (im) that are primarily
6 deposited in the ciliated airways (nasopharyngeal and tracheobronchial regions) can be
7 transferred by mucociliary transport into the esophagus and swallowed, thus being
8 absorbed via the gut.
9 Inhaled Pb lodging deep in the respiratory tract seems to be absorbed equally and totally,
10 regardless of chemical form (Morrow et al.. 1980; Chamberlain et al.. 1978; Rabinowitz
11 et al.. 1977). Absorption half-times (tl/2) have been estimated for radon decay progeny in
12 adults who inhaled aerosols of Pb and bismuth isotopes generated from decay of 220Rn or
13 222Rn. The absorption half-time for Pb from the respiratory tract to blood was estimated
14 to be approximately 10 hours in subjects who inhaled aerosols having an activity median
15 particle diameter of approximately 160 nm (range 50-500 nm) (Marsh and Birchall.
16 1999). and approximately 68 min for aerosols having diameters of approximately 0.3-
17 3 nm (Butterweck et al.. 2002). Given the submicron particle size of the exposure, these
18 rates are thought to represent, primarily, absorption from the bronchiolar and alveolar
19 regions of the respiratory tract.
20 Several studies have quantified the bioaccessibility of Pb in atmospheric PM, based on
21 various in vitro extraction methods. In a study of PMi0 and PM2 5 samples from
22 downtown Vienna, Austria, Falta et al. (2008) used synthetic gastric juice to investigate
23 the bioaccessibility of metals including Pb. The rationale was that inhaled particles in the
24 2.5-10 (im size range are mostly deposited in the tracheal and bronchial regions of the
25 lung from where they are transported within hours by mucociliary clearance, i.e., they are
26 mainly swallowed. In contrast, the <2.5 (im particles are deposited in the pulmonary
27 alveoli where they can stay for months to years. The study aimed to determine the
28 bioaccessibility of the 2.5-10 (im PM. It is important to note that they do not isolate the
29 2.5-10 (im size range; instead, they infer the characteristics from the difference between
30 the PM2 5 and PMi0 fractions. The Pb concentrations associated with the two fractions
31 were almost identical, as was the percentage extracted by synthetic gastric juice (86% and
32 83% Pb for PM2 5 and PMi0 fractions, respectively). The mean daily bioavailable mass
33 was calculated to be 16 ng for the PM25.10 size range. Since the quantitative clearance of
34 these particles to the stomach was assumed, this value represents an upper estimate for
35 the amount of bioavailable Pb. Niu et al. (2010) determined the bioaccessibility of Pb in
36 fine (100-1,000 nm) and ultrafine-sized (<100 nm) urban airborne PM from two sites
November 2012 4-33 Draft - Do Not Cite or Quote
-------
1 within the city of Ottawa, Canada. For all size fractions, the median Pb concentrations for
2 particles smaller than 10 um were 8,800 and 7,800 mg/kg for the two different locations.
3 The bioaccessibility was based on ammonium acetate extractability and it was found that,
4 within the fine and ultrafme-size ranges, 13-28% Pb was extracted. The Falta et al.
5 (2008) and Niu et al. (2010) results illustrate that different extraction techniques result in
6 different bioaccessible fractions. The main finding from Niu et al. (2010) was that the
7 highest values (-28% and -19% for the two different locations) were found for the
8 <57 nm particles, with percent bioaccessibility decreasing with increasing particle size.
9 This result indicated that Pb was potentially most bioaccessible in the ultrafme-size
10 range.
11 A recent study by Barrett et al. (2010) investigated the solid phase speciation of Pb in
12 urban road dust in Manchester, U.K., and considered the health implications of inhalation
13 and ingestion of such material. Human exposure via inhalation is likely to involve only
14 the finest grained fractions (up to 10 (im) and unfortunately this study characterized only
15 the <38 (im fraction. Pb-goethite and PbCrO4 comprised the largest fractions, 45% and
16 21% respectively, of Pb in the <38 (im fraction. These forms tend to be less bioaccessible
17 if ingested compared with PbO or Pb acetate because they are less soluble.
18 The above considerations indicate that the relationship between air Pb exposure and
19 blood Pb will depend on numerous exposure variables (e.g., particle size, solubility,
20 exposure frequency and duration) and physiological variables (age, activity level,
21 transport and absorption in the respiratory tract, blood Pb kinetics). For a detailed
22 discussion of factors affecting particle deposition and retention in the human respiratory
23 tract the reader is referred to Chapter 4 of 2009 PM ISA (U.S. EPA. 2009a). Section 4.2.4
24 of that document specifically addresses biological factors affecting particle deposition
25 such as activity level and age with an emphasis on children. Mechanistic models provide
26 one means for integrating these variables into predictions of blood Pb - air Pb
27 relationships; although, predictions are subject to simplifications and generalizations
28 made in constructing the models. As an example, the ICRP (Pounds and Leggett 1998;
29 ICRP. 1994; Leggett. 1993) model (see Section 4.3 for a brief description) can be used to
30 predict blood Pb - air Pb slopes for specific direct Pb inhalation exposure scenarios. For a
31 long-term continuous (24 hours/day) exposure of a typical adult male engaged in light
32 exercise (ventilation rate 20-22 mVday) to Pb-bearing particles having a 1 (im uniform
33 particle size, the predicted blood Pb - air Pb slopes range from 0.7 (ig/dL per (ig/m3 (for
34 low solubility particles; e.g., Pb oxide) to 3 (ig/dL per (ig/m3 (for highly soluble Pb;
35 e.g., Pb salts). These slopes were calculated by running ICRP model simulations with
36 varying air concentrations (0.1-6 (ig/m3) to achieve a range of blood Pb concentrations
37 up to 10 (ig/dL, starting with a baseline of 1.6 (ig/dL, and estimating the linear slope of
38 the relationship between blood Pb concentration and air Pb. Empirical estimates of blood
November 2012 4-34 Draft - Do Not Cite or Quote
-------
1 Pb - air Pb slopes for various populations, derived from epidemiological studies, are
2 summarized in Section 4.5.1.
Organic Pb
3 Alkyl Pb compounds can exist in ambient air as vapors. Inhaled tetraalkyl Pb vapor is
4 nearly completely absorbed following deposition in the respiratory tract. As reported in
5 the 2006 Pb AQCD (U.S. EPA. 2006c). a single exposure to vapors of radioactive (203Pb)
6 tetraethyl Pb resulted in 37% initially deposited in the respiratory tract, of which -20%
7 was exhaled in the subsequent 48 hours (Heard etal.. 1979). In a similar experiment
8 conducted with 203Pb tetramethyl Pb, 51% of the inhaled 203Pb dose was initially
9 deposited in the respiratory tract, of which -40% was exhaled in 48 hours (Heard et al..
10 1979).
11 Estimation of bioavailability of organic Pb is relevant to some aviation fuel exposures
12 (e.g., persons exposed to leaded gasoline used in piston-engine aircraft). Mahaffey (1977)
13 estimated that 40% of inhaled Pb in urban air (largely attributed to combustion of
14 gasoline containing tetraethyllead) is bioavailable to adults. Chamberlain et al. (1975)
15 suggested that 35% of inhaled combustion products of tetraethyl 203Pb fuel [likely to have
16 been a mixture dominated by inorganic Pb halides, but may also have include alkly Pb
17 species (U.S. EPA. 2006b)1 are deposited and then retained in adult lungs with a half-life
18 of 6 hours. Fifty percent of that 203Pb was detectable in the blood within 50 hours of
19 inhalation, and the rest was found to deposit in bone or tissue. Chamberlain et al. (1975)
20 estimated that continuous inhalation of Pb in engine exhaust from fuel containing
21 tetraethyllead at a concentration of 0.001 ug/m3 for a period of months could produce a
22 1 ug/dL increment in blood Pb.
4.2.1.2 Ingestion
23 The extent and rate of GI absorption of ingested inorganic Pb are influenced by
24 physiological states of the exposed individual (e.g., age, fasting, nutritional calcium
25 (Ca2+) and iron (Fe) status, pregnancy) and physicochemical characteristics of the
26 Pb-bearing material ingested (e.g., particle size, mineralogy, solubility). Pb absorption in
27 humans may be a capacity-limited process, in which case the percentage of ingested Pb
28 that is absorbed may decrease with increasing rate of Pb intake. Numerous observations
29 of nonlinear relationships between blood Pb concentration and Pb intake in humans
30 provide support for the likely existence of a saturable absorption mechanism or some
31 other capacity-limited process in the distribution of Pb in humans (Sherlock and Quinn,
32 1986: Sherlock et al.. 1984: Pococketal.. 1983: Sherlock et al.. 1982). While evidence
November 2012 4-35 Draft - Do Not Cite or Quote
-------
1 for capacity-limited processes at the level of the intestinal epithelium is compelling, the
2 dose at which absorption becomes appreciably limited in humans is not known.
3 In adults, estimates of absorption of ingested water-soluble Pb compounds
4 (e-g-, Pb chloride, Pb nitrate, Pb acetate) range from 3 to 10% in fed subjects (Maddaloni
5 etal.. 1998: Watson etal.. 1986: James etal.. 1985: Heard and Chamberlain. 1982:
6 Rabinowitz et al.. 1980). The absence of food in the GI tract increases absorption of
7 water-soluble Pb in adults. Reported estimates of soluble Pb absorption range from 26 to
8 70% in fasted adults (Maddaloni et al.. 1998: James etal.. 1985: Blake etal.. 1983: Heard
9 and Chamberlain. 1982: Rabinowitz et al.. 1980). Reported fed:fasted ratios for soluble
10 Pb absorption in adults range from 0.04 to 0.2 (James etal.. 1985: Blake etal.. 1983:
11 Heard and Chamberlain. 1982: Rabinowitz et al.. 1980).
12 Limited evidence demonstrates that GI absorption of water-soluble Pb is higher in
13 children than in adults. Estimates derived from dietary balance studies conducted in
14 infants and children (ages 2 weeks to 8 years) indicate that -40-50% of ingested Pb is
15 absorbed (Ziegler et al.. 1978: Alexander etal.. 1974). Experimental studies provide
16 further evidence for greater absorption of Pb from the gut in young animals compared to
17 adult animals (Aungst et al.. 1981: Kostial et al.. 1978: Pounds etal.. 1978: Forbes and
18 Reina. 1972). The mechanisms for an apparent age difference in GI absorption of Pb have
19 not been completely elucidated and may include both physiological and dietary factors
20 (Mushak. 1991). To further investigate the effects of the presence of food in the GI tract
21 on Pb absorption, children (3-5 years old) who ate breakfast had lower blood Pb levels
22 compared to children who did not eat breakfast (Liu etal.. 201 la). This difference
23 persisted after controlling for nutritional variables (blood iron [Fe], calcium [Ca2+],
24 copper [Cu], magnesium [Mg], zinc [Zn]). This observation may be explained by lower
25 GI absorption of Pb ingested with or in close temporal proximity to meals. Direct
26 evidence for meals lowering GI absorption of Pb has also been reported for adults
27 (Maddaloni et al.. 1998: James etal.. 1985).
28 Nutritional interactions of Pb with dietary elements (e.g., Fe, Ca2+, Zn) are complex. Pb
29 competes with other elements for transport and binding sites that can result in
30 adjustments of homeostatic regulators to absorb and retain needed elements.
31 Additionally, low levels of macronutrients may alter Pb bioaccessibility in the GI tract.
32 Genetic variation in absorption and metabolism may modify all of the above.
33 Children who are iron-deficient have higher blood Pb concentrations than similarly
34 exposed iron-replete children, suggesting that iron deficiency may result in higher Pb
35 absorption or, possibly, other changes in Pb biokinetics that contribute to altered blood
36 Pb concentrations (Schell et al.. 2004: Marcus and Schwartz. 1987: Mahaffey and
37 Annest. 1986). Studies conducted in animal models have provided direct evidence for
November 2012 4-36 Draft - Do Not Cite or Quote
-------
1 interactions between iron deficiency and increased Pb absorption, perhaps by enhancing
2 binding of Pb to iron-binding proteins in the intestine (Bannon et al.. 2003; Morrison and
3 Ouarterman. 1987; Barton etal.. 1978b). An analysis of data from a sample 448 woman
4 (age 20-55 years) did not find a significant association between iron body stores
5 (indicated from serum ferritin concentration) and blood Pb concentrations, although
6 depleted irons stores (serum ferritin of <12 (ig/L) was associated with higher blood
7 concentrations of Cd, cobalt (Co) and manganese (Mn) higher (Meltzer et al.. 2010).The
8 effects of iron nutritional status on blood Pb include changes in blood Pb concentrations
9 in association with genetic variation in genes involved in iron metabolism. For example,
10 genetic variants in the hemochromatosis (HFE) and transferrin genes are associated with
11 higher blood Pb concentrations in children (Hopkins et al.. 2008). In contrast, HFE gene
12 variants are associated with lower bone and blood Pb levels in elderly men (Wright et al..
13 2004).
14 Several studies have suggested that dietary Ca2+ may have a protective role against Pb by
15 decreasing absorption of Pb in the GI tract and by decreasing the mobilization of Pb from
16 bone stores to blood. In experimental studies of adults, absorption of a single dose of Pb
17 (100-300 (ig Pb chloride) was lower when the Pb was ingested together with
18 Ca2+ carbonate (0.2 g Ca2+ carbonate) than when the Pb was ingested without additional
19 Ca2+ (Blake and Mann. 1983; Heard and Chamberlain. 1982). A similar effect of Ca2+
20 occurs in rats (Barton et al.. 1978a). Similarly, an inverse relationship was observed
21 between dietary Ca2+ intake and blood Pb concentration in children, suggesting that
22 children who are Ca2+-deficient may absorb more Pb than Ca2+-replete children (Elias et
23 al.. 2007; Schell et al.. 2004; Mahaffev et al.. 1986; Ziegleretal.. 1978). These
24 observations suggest that Ca2+ and Pb share and may compete for common binding and
25 transport mechanisms in the small intestine which are regulated in response to dietary
26 Ca2+ and Ca2+-body stores (Fullmer and Rosen. 1990; Bronner et al.. 1986). However,
27 animal studies have also shown that multiple aspects of Pb toxicokinetics are affected by
28 Ca2+ nutritional status. For example, feeding rats a Ca2+ deficient diet is associated with
29 increased Pb absorption, decreased whole body Pb clearance, and increased volume of
30 distribution of Pb (Aungst and Fung. 1985). These studies suggest that associations
31 between Ca2+ nutrition and blood Pb that have been observed in human populations may
32 not be solely attributable to effects of Ca2+ nutrition on Pb absorption. Other potential
33 mechanisms by which Ca2+ nutrition may affect blood Pb and Pb biokinetics include
34 effects on bone mineral metabolism and renal function.
35 Blood Pb concentrations in young children have also been shown to increase in
36 association with lower dietary Zn levels (Schell et al., 2004). Mechanisms for how Zn
37 affects blood Pb concentration, i.e., whether it involves changes in absorption or changes
38 in distribution and/or elimination of Pb, have not been determined.
November 2012 4-37 Draft - Do Not Cite or Quote
-------
1 Dissolution of Pb from the soil/mineralogical matrix in the stomach appears to be the
2 major process that renders soil Pb bioaccessible for absorption in the GI tract. Absorption
3 of Pb has been shown to vary depending upon the Pb mineralogy and physical
4 characteristics of the Pb in the soil (e.g., encapsulated or exposed) and size of the
5 Pb-bearing grains. GI absorption of larger Pb-containing particles (MOO urn) tends to be
6 lower than smaller particles (Healy et al.. 1992; Barltrop and Meek. 1979). Absorption of
7 Pb in soils and dust has been most extensively studied in the in vivo swine model. Gastric
8 function of swine is thought to be sufficiently similar to that of humans to justify use of
9 swine as a model for assessing factors that may affect GI absorption of Pb from soils in
10 humans (Juhasz et al.. 2009; U.S. EPA. 2007b: Casteel et al.. 2006; Casteel et al.. 1997;
11 Weis and Lavelle. 1991). Other practical advantages of the swine model over rodent
12 models have been described, and include: absence of coprophagia; ease with which Pb
13 dosing can be administered and controlled; and higher absorption fraction of soluble Pb
14 (e-g-, Pb acetate) in swine, which is more similar to humans than rats (Smith et al.,
15 2009a). The swine studies measure blood and/or tissue Pb (e.g., kidney, liver, bone)
16 concentrations following oral dosing of swine with either soil or with a highly water
17 soluble and fully bioaccessible form of Pb (e.g., Pb acetate). A comparison of the internal
18 concentrations of Pb under these two conditions provides a measure of the bioavailability
19 (i.e., absorption) of Pb in soil relative to that of Pb acetate, which is typically referred to
20 as relative bioavailability (RBA). Relative bioavailability measured in the swine assay is
21 equivalent to the ratio of the absorbed fraction (AF) of ingested dose of soil Pb to that of
22 water-soluble Pb acetate (e.g., RBA = AFSoiipb/AFPbacetate)-
23 Collectively, published studies conducted in swine have provided 39 estimates of Pb
24 RBA for 38 different soil or "soil-like" test materials (Bannon et al.. 2009; Smith et al..
25 2009a; Casteel et al.. 2006; Marschner et al.. 2006). The mean of RBA estimates from 25
26 soils is 0.49 (± 0.29[SD]), median is 0.51, and 5th to 95th percentile range is 0.12 to
27 -0.89. RBA estimates for soils collected from 8 firing ranges were approximately 1.0
28 (Bannon et al.. 2009). The relatively high RBA for the firing range soils may reflect the
29 high abundance of relatively un-encapsulated Pb carbonate (30-90% abundance) and Pb
30 oxide (1-60%) in these soils. Similarly, a soil sample (low Pb concentration) mixed with
31 a NIST paint standard (55% Pb carbonate, 44% Pb oxide) also had a relatively high
32 bioavailability (0.72) (Casteel et al.. 2006). Samples of smelter slag, or soils in which the
33 dominant source of Pb was smelter slag, had relatively low RBA (0.14 - 0.40, n = 3), as
34 did a sample from a mine tailings pile (RBA = 0.06), and a sample of finely ground
35 galena mixed with soil (Casteel et al.. 2006).
36 Based on data for 18 soil materials assayed in swine, RBA of Pb mineral phases were
37 categorized into "low" (<0.25 [25%]), "medium" (0.25-0.75 [25 to 75%]), and "high"
38 (>0.75 [75%]) categories (Casteel et al.. 2006). Figure 4-3 shows some of the materials
November 2012 4-38 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
that fall into these three categories. Mineral phases observed in mineralogical wastes can
be expected to change overtime (i.e., weathering), which could change the RBA over
time. The above observations in swine are supported by various studies conducted in rats
that have found RBA of Pb in soils to vary considerably and to be less than 1.0 (Smith et
al.. 2009a. 2008; Freeman et al.. 1996; Freeman et al.. 1994; Freeman et al.. 1992).
CO
T3
re
0)
o
0)
Q.
Q.
3
O
O
-o
Q)
"re
E
m
Group
Note: based on results from juvenile swine assays.
Source: Casteel et al. (2006).
Figure 4-3 Estimated relative bioavailability (RBA, compared to Pb acetate)
of ingested Pb in mineral groups.
November 2012
4-39
Draft - Do Not Cite or Quote
-------
1 Drexler and Brattin (2007) developed an in vitro bioaccessibility (IVBA) assay for soil
2 Pb that utilizes extraction fluid comprised of glycine, deionized water, and hydrochloric
3 acid at a pH of 1.50 that is combined with sieved test material (<250 urn) for 1 hour. The
4 assay was tested for predicting in vivo RBA of 18 soil-like test materials that were
5 assayed in a juvenile swine assay (Casteel et al., 2006). A regression model relating
6 IVBA and RBA was derived based on these data (Equation 4-1):
RBA = (0.878 X IVBA) - 0.028
Equation 4-1
7 where RBA and IVBA are expressed as fractions (i.e., not as percent). The weighted r2
8 for the relationship (weighted for error in the IVBA and RBA estimates) was 0.924
9 (p <0.001). The IVBA assay reported in Drexler and Brattin (2007) has been identified by
10 the U.S. EPA as a validated method for predicting RBA of Pb in soils for use in risk
11 assessment (U.S. EPA. 2007e). A review of soil Pb RBA estimates made using the IVBA
12 assay described above and Equation 4-1 identified 270 estimates of Pb RBA in soils
13 obtained from 11 hazardous waste sites. The mean for the site-wide RBA estimates
14 (n = 11 sites) was 0.57 (SD 0.15), median was 0.63, and 5th to 95th percentile range was
15 0.34 to 0.71.
16 Equation 4-1 cannot be reliably extrapolated to other in vitro assays that have been
17 developed for estimating Pb bioaccessibility without validation against in vivo RBA
18 measurements made on the same test materials. Comparisons of outcomes among
19 different in vitro assays applied to the same soil test materials have found considerable
20 variability in IVBA estimates (Juhasz etal., 2011; Smith etal., 2011; Saikat et al., 2007;
21 Van de Wiele et al.. 2007). This variability has been attributed to differences in assay
22 conditions, including pH, liquid:soil ratios, inclusion or absence of food material, and
23 differences in methods used to separate dissolved and particle-bound Pb
24 (e.g., centrifugation versus filtration). Smith et al. (2011) found that algorithms for
25 predicting RBA based on two different IVBA assays did not yield similar predictions of
26 RBA when applied to the same material. Given the dependence of IVBA outcomes on
27 assay conditions, in vitro assays used to predict in vivo RBA should be evaluated against
28 in vivo RBA estimates to quantitatively assess uncertainty in RBA predictions (U.S.
29 EPA. 2007e).
30 Absorption of Pb in house dusts has not been rigorously evaluated quantitatively in
31 humans or in experimental animal models. The RBA for paint Pb mixed with soil was
32 reported to be approximately 0.72 (95% CI: 0.44, 0.98) in juvenile swine, suggesting that
33 paint Pb dust reaching the gastrointestinal tract maybe highly bioavailable (Casteel et al.,
November 2012 4-40 Draft - Do Not Cite or Quote
-------
1 2006). The same material yielded a bioaccessibility value (based on IVBA assay) of 0.75
2 (Drexler and Brattin. 2007). which corresponds to a predicted RBA of 0.63, based on
3 Equation 4-1. A review of indoor Pb RBA estimates made using the IVBA assay and
4 Equation 4-1 identified 100 estimates of Pb RBA in dusts obtained from two hazardous
5 waste sites. Mean Pb RBAs for the Herculaneum site were 0.47 (SD 0.07, 10 samples)
6 for indoor dust and 0.69 (SD 0.03, 12 samples) for soil. At the Omaha site, mean Pb
7 RBAs were 0.73 (SD 0.10, 90 samples) for indoor dust and 0.70 (SD 0.10, 45 samples)
8 for soil. Yu et al. (2006) applied an IVBA method to estimate bioaccessibility of Pb in
9 house dust samples collected from 15 urban homes. Homes were selected for inclusion in
10 this study based on reporting to the state department of health of at least on child with a
11 blood Pb concentration >15 (ig/dL and Pb paint dust may have contributed to indoor dust
12 Pb. The mean IVBA was 0.65 (SD 0.08, age: 52.5 to 77.2 months).
13 The above results, and the IVBA assays used in studies of interior dust, have not been
14 evaluated against in vivo RBA estimates for dust samples. Although, expectations are
15 that a validated IVBA methodology for soil would perform well for predicting RBA of
16 interior dust, this validation has not actually been experimentally confirmed. Factors that
17 may affect in vivo predictions of RBA of interior dust Pb could include particle size
18 distribution of interior dust Pb and the composition of the dust matrix, which may be
19 quite different from that of soil.
20 Other estimates of bioaccessibility of Pb in house dusts have been reported, based on
21 results from in vitro extraction assays that have not been validated for predicting in vivo
22 bioavailability. Bioaccessibility assays that sequentially extract soil at gastric pH
23 followed by intestinal pH tend to show higher bioaccessibility of soil and dust Pb when
24 incubated at gastric conditions (Juhasz et al., 2011; Lu et al.. 2011; Smith et al.. 2011;
25 Roussel et al.. 2010; Yu et al.. 2006). Yu et al. (2006) dissolved Pb dust, obtained from
26 vacuuming carpet samples into simulated gastric and intestinal acids (also
27 Section 4.1.3.2). The carpet samples were obtained from homes located in northern
28 New Jersey. Pb concentration in carpet ranged from 209 to 1,770 mg/kg dust, with
29 52-77% of Pb dissolving in simulated gastric acid and 5-32% dissolving in simulated
30 intestinal acids. In a similar test in the U.K., Turner and Simmonds (2006) observed
31 median Pb dust concentrations of 178 mg/kg with approximately 80% bioaccessibility in
32 simulated gastric acid. Jin et al. (2005) observed that bioaccessibility of Pb in soil was
33 proportional to the soil acidity and organic matter content of the soil.
November 2012 4-41 Draft - Do Not Cite or Quote
-------
4.2.2 Distribution
1 A simple conceptual representation of Pb distribution is that it contains a fast turnover
2 pool, comprising mainly soft tissue, and a slow pool, comprising mainly skeletal tissues
3 (Rabinowitz et al.. 1976). The highest soft tissue concentrations in adults occur in liver
4 and kidney cortex (Gerhardsson et al.. 1995; Oldereid et al.. 1993; Gerhardsson et al..
5 1986; Barry. 1975; Gross et al.. 1975). Pb in blood (i.e., plasma) exchanges with both of
6 these compartments.
4.2.2.1 Blood
7 Blood comprises ~1% of total Pb body burden. Pb in blood is found primarily (>99%) in
8 the RBCs (Smith et al.. 2002; Manton et al.. 2001; Bergdahl et al.. 1999; Bergdahl et al..
9 1998; Hernandez-Avila et al.. 1998; Bergdahl et al.. 1997a; Schutzetal. 1996).
10 5-aminolevulinic acid dehydratase (ALAD) is the primary binding ligand for Pb in
11 erythrocytes (Bergdahl et al.. 1998; Xieetal.. 1998; Bergdahl et al.. 1997a; Sakai et al..
12 1982). Two other Pb-binding proteins have been identified in the RBC, a 45 kDa protein
13 (Kmax 700 (ig/dL; Kd 5.5 (ig/L) and a smaller protein band having a molecular weight of
14 <10 kDa (Bergdahl et al.. 1998; Bergdahl et al.. 1997a; Bergdahl et al.. 1996). Of the
15 three principal Pb-binding proteins identified in RBCs, ALAD has the strongest affinity
16 for Pb (Bergdahl etal.. 1998) and appears to dominate the ligand distribution of Pb (35 to
17 84% of total erythrocyte Pb) at blood Pb levels below 40 (ig/dL (Bergdahl et al.. 1998;
18 Bergdahl et al.. 1996; Sakai etal.. 1982). Pb binding to ALAD is saturable; the binding
19 capacity was estimated to be -850 (ig/dL RBCs (or ~40 (ig/dL whole blood) and the
20 apparent dissociation constant has been estimated to be ~1.5 (ig/L (Bergdahl et al.. 1998).
21 Hematocrit is somewhat higher in the neonate at birth (51%) than in later infancy (35% at
22 6 months), which may lead to a decrease in the total binding capacity of blood over the
23 first 6 months of life that results in a redistribution of Pb among other tissues (Simon et
24 al.. 2007).
25 Saturable binding to RBC proteins contributes to an increase in the plasma/blood Pb ratio
26 with increasing blood Pb concentration and curvature to the blood Pb-plasma Pb
27 relationship (Rentschler etal.. 2012; Kang et al.. 2009; Jin et al.. 2008; Barbosaet al..
28 2006b; Smith et al.. 2002; Manton etal.. 2001; Bergdahl et al.. 1999; Bergdahl et al..
29 1998; Bergdahl et al.. 1997b; DeSilva. 1981). An example of this is shown in Figure 4-4.
30 Saturable binding of Pb to RBC proteins has several important consequences. As blood
31 Pb increases and the higher affinity binding sites for Pb in RBCs become saturated, a
32 larger fraction of the blood Pb is available in plasma to distribute to brain and other
33 Pb-responsive tissues. This change in distribution of Pb contributes to a curvature in the
November 2012 4-42 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
relationship between Pb intake (at constant absorption fraction) and blood Pb
concentration. Plasma Pb also exhibits faster kinetics. Following exposures of 5 adults
that resulted in relatively high blood Pb concentrations (56-110 (ig/dL), the initial (fast-
phase) elimination half-time for plasma Pb (38 ± 20 [SD] days) was approximately half
that of blood (81 ± 25 days) (Rentschler etal. 2012).
Typically, at blood Pb concentrations <100 (ig/dL, only a small fraction (<1%) of blood
Pb is found in plasma (Marcus. 1985; Manton and Cook. 1984; DeSilva. 1981). However,
as previously noted, plasma Pb may be the more biologically labile and lexicologically
effective fraction of the circulating Pb. Approximately 40-75% of Pb in the plasma is
bound to proteins, of which albumin appears to be the dominant ligand (Al-Modhefer et
al., 1991; Ong and Lee. 1980a). Pb in serum that is not bound to protein exists largely as
complexes with low molecular weight sulfhydryl compounds (e.g., cysteine,
homocysteine) and other ligands (Al-Modhefer et al., 1991).
0
oAdults • Children
20 40 60
Blood Pb (Mg/dL)
80
100
Source: Adapted with permission of Elsevier Publishing and the Finland Institute of Occupational Health, Bergdahl et al. (1999:
1997b).
Figure 4-4 Plot of blood and plasma Pb concentrations measured in adults
and children.
November 2012
4-43
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
As shown in Figure 4-4. the limited binding capacity of Pb binding proteins in RBCs
produces a curvilinear relationship between blood and plasma Pb concentration. The
limited binding capacity of RBC binding proteins also confers, or at least contributes, to a
curvilinear relationship between Pb intake and blood Pb concentration. A curvilinear
relationship between Pb intake and blood Pb concentration has been observed in children
(Sherlock and Quinn. 1986; Lacevetal.. 1985; Ryuetal.. 1983). As shown in Figure 4-5.
the relationship becomes pseudo-linear at relatively low daily Pb intakes
(i.e., <10 ug/day/kg) and at blood Pb concentrations <25 (ig/dL.
0
100 200 300
Pb Intake (|jg/day)
400
Data represent mean and standard errors for intake; the line is the regression model (blood Pb = 3.9 + 2.43 (Pb intake [|jg/week] ).
Source: Adapted with permission of Taylor & Francis Publishing, Sherlock and Quinn (1986).
Figure 4-5 Relationship between Pb intake and blood Pb concentration in
infants (n = 105, age 13 weeks, formula-fed).
November 2012
4-44
Draft - Do Not Cite or Quote
-------
1 Figure 4-6 shows the predicted relationship between quasi-steady state blood and plasma
2 Pb concentrations in a 4-year old child using the ICRP model [(Pounds and Leggett.
3 1998; ICRP. 1994; Leggett. 1993). see Section 43 for a brief description of the ICRP
4 model]. The abrupt inflection point that occurs at approximately 25 (ig/dL blood Pb is an
5 artifact of the numerical approach to simulate the saturation of binding using
6 discontinuous first-order rate constants for uptake and exit of Pb from the RBC. A
7 continuous function of binding sites and affinity, using empirical estimates of both
8 parameters, yield a similar but continuous curvature in the relationship (Bergdahl et al.
9 1998; O'Flaherty. 1995). Nevertheless, either approach predicts a pseudo-linear
10 relationship at blood Pb concentrations below approximately 25 (ig/dL which, in this
11 model, corresponds to an intake of approximately 100 (ig/day (absorption rate
12 ~ 30 (ig/day) (upper panel). An important consequence of the limited Pb-binding capacity
13 of RBC proteins is that the plasma Pb concentration will continue to grow at a linear rate
14 above the saturation point for RBC protein binding. One implication of this is that a
15 larger fraction of the Pb in blood will become available to distribute to brain and other
16 Pb-responsive tissues as blood Pb increases. This could potentially contribute to
17 non-linearity in dose-response relationships in studies in which blood Pb is the used as
18 the internal dose metric.
November 2012 4-45 Draft - Do Not Cite or Quote
-------
0.60
0.50 -
3) 0.40 -
°- 0.30 -
ra
E
ra 0.20 -
o.
0.10 -
0.00
10 20 30 40
Blood Pb (ng/dL)
50
60
50 -
IT
1) 40 -
^
si
£ 30 H
o
m 20 -
10 -
Blood
•Plasma
1.0
0.8
0)
tn
0.6 3
0)
TJ
0-4
0.2
0.0
100 200 300
Intake (tig/day)
400
Note: Model simulations are for a 4-year old having from birth a constant Pb intake of between 1 and 400 ug/day. Simulation based
on ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-6 Simulation of quasi-steady state blood and plasma Pb
concentrations in a child (age 4 years) associated with varying Pb
ingestion rates.
November 2012
4-46
Draft - Do Not Cite or Quote
-------
1 Studies conducted in swine provide additional evidence in support of RBC binding
2 kinetics influencing distribution of Pb to tissues. In these studies, the relationship
3 between the ingested dose of Pb and tissue Pb concentrations (e.g., liver, kidney, bone)
4 was linear, whereas, the relationship between dose and blood Pb was curvilinear with the
5 slope decreasing as the dose increased (Casteel et al.. 2006). Saturable binding of Pb to
6 RBC proteins also contributes to a curvilinear relationship between urinary Pb excretion
7 and plasma Pb concentration (Section 4.2.3) (Besser et al., 2008; Bergdahl et al., 1997b).
4.2.2.2 Bone
8 The dominant compartment for Pb in the body is in bone. In human adults, more than
9 90% of the total body burden of Pb is found in the bones, whereas bone Pb accounts for
10 just under 60% of the body burden in infants less than a year old and just over 70% of the
11 body burden in older children (Barry. 1975). Bone is comprised of two main types,
12 cortical (or compact) and trabecular (or spongy or cancellous). The proportion of cortical
13 to trabecular bone in the human body varies by age, but on average is about 80 to 20
14 percent (O'Flahertv. 1998; Leggett 1993; ICRP. 1973).
15 The exchange of Pb from plasma to the bone surface is a rapid process (i.e., adult ti/2
16 =0.19 and 0.23 hours for trabecular and cortical bone, respectively) (Leggett, 1993).
17 Some Pb diffuses from the bone surface to deeper bone regions (adult ti/2=150 days)
18 where it is relatively inert (in adults) and part of a "nonexchangeable" (removed only
19 through bone resorption/remodeling) pool of Pb in bone (Leggett. 1993).
20 Pb distribution in bone includes uptake into cells that populate bone (e.g., osteoblasts,
21 osteoclasts, osteocytes) and exchanges with proteins and minerals in the extracellular
22 matrix (Pounds etal. 1991). Pb forms highly stable complexes with phosphate and can
23 replace calcium in the calcium-phosphate salt, hydroxyapatite, which comprises the
24 primary crystalline matrix of bone (Meirer et al.. 2011; Bres etal.. 1986; Miyake. 1986;
25 Verbeeck et al., 1981). Several intracellular kinetic pools of Pb have been described in
26 isolated cultures of osteoblasts and osteoclasts which appear to reflect physiological
27 compartmentalization within the cell, including membranes, mitochondria, soluble
28 intracellular binding proteins, mineralized Pb (i.e., hydroxyapatite) and inclusion bodies
29 (Long etal.. 1990; Pounds and Rosen. 1986; Rosen. 1983). Approximately 70-80% of Pb
30 taken up into isolated primary cultures of osteoblasts or osteocytes is associated with
31 mitochondria and mineralized Pb (Pounds etal.. 1991).
32 Pb accumulates in bone regions having the most active calcification at the time of
33 exposure. Pb accumulation is thought to occur predominantly in trabecular bone during
34 childhood and in both cortical and trabecular bone in adulthood (Aufderheide and
November 2012 4-47 Draft - Do Not Cite or Quote
-------
1 Wittmers. 1992). Early Pb uptake in children is greater in trabecular bone due to its larger
2 surface area and higher metabolic rate. With continued exposure, Pb concentrations in
3 bone may increase with age throughout the lifetime beginning in childhood, indicative of
4 a relatively slow turnover of Pb in adult bone (Park et al.. 2009c: Barry and Connolly.
5 1981; Barry. 1975; Gross etal.. 1975; Schroeder and Tipton. 1968). The cortical and
6 trabecular bones have distinct rates of turnover and Pb release. For example, tibia has a
7 turnover rate of about 2% per year whereas trabecular bone has a turnover rate of more
8 than 8% per year in adults (Rabinowitz. 1991).
9 A high bone formation rate in early childhood results in the rapid uptake of circulating Pb
10 into mineralizing bone; however, bone Pb is also recycled to other tissue compartments
11 or excreted in accordance with a high bone resorption rate (O'Flaherty. 1995). Thus, most
12 of the Pb acquired early in life is not permanently fixed in the bone (60-65%)
13 (O'Flahertv. 1995; Leggett. 1993; ICRP. 1973). However, some Pb accumulated in bone
14 does persist into later life. McNeill et al. (2000) compared tibia Pb levels and cumulative
15 blood Pb indices in a population of 19- to 29-year-olds who had been highly exposed to
16 Pb in childhood from the Bunker Hill, Idaho smelter; they concluded that Pb from
17 exposure in early childhood had persisted in the bone matrix until adulthood.
18 Additional discussion of the Pb in bone and its mobilization are provided in other
19 sections of this chapter. Maternal mobilization of Pb from the bone to the fetus is
20 discussed in Section 4.2.2.4. The relationship between Pb in blood and bone is discussed
21 in Section 4.3.5.
4.2.2.3 Soft Tissues
22 Most of the Pb in soft tissue is in liver and kidney (Gerhardsson et al.. 1995; Oldereid et
23 al.. 1993; Gerhardsson et al.. 1986; Barry. 1981. 1975; Gross etal.. 1975). Presumably,
24 the Pb in these soft tissues (i.e., kidney, liver, and brain) exists predominantly bound to
25 protein. High affinity cytosolic Pb-binding proteins have been identified in rat kidney and
26 brain (DuVal and Fowler. 1989; Fowler. 1989). The Pb-binding proteins in rat are
27 cleavage products of a2(i globulin, a member of the protein superfamily known as
28 retinol-binding proteins that are generally observed only in male rats (Fowler and DuVal.
29 1991). Other high-affinity Pb-binding proteins (Kd ~14 nM) have been isolated in human
30 kidney, two of which have been identified as a 5 kDa peptide, thymosin 4 and a 9 kDa
31 peptide, acyl-CoA binding protein (Smith etal.. 1998). Pb also binds to metallothionein,
32 but does not appear to be a significant inducer of the protein in comparison with the
33 inducers Cd and Zn (Waalkes and Klaassen. 1985; Eaton et al.. 1980).
November 2012 4-48 Draft - Do Not Cite or Quote
-------
1 The liver and kidneys rapidly accumulate systemic Pb (ti/2=0.21 and 0.41 hours,
2 respectively), which amounts to 10-15% and 15-20% of intravenously injected Pb,
3 respectively (Leggett 1993). A linear relationship in dose-tissue Pb concentrations for
4 kidney and liver has been demonstrated in swine, dogs, and rats (Smith et al.. 2008;
5 Casteel et al.. 2006; Casteel et al., 1997; Azaretal.. 1973). In contrast to Pb in bone,
6 which accumulates Pb with continued exposure in adulthood, concentrations in soft
7 tissues (e.g., liver and kidney) are relatively constant in adults (Treble and Thompson.
8 1997; Barry. 1975). reflecting a faster turnover of Pb in soft tissue relative to bone.
4.2.2.4 Fetus
9 Evidence for maternal-to-fetal transfer of Pb in humans is derived from cord blood Pb to
10 maternal blood Pb ratios (i.e., cord blood Pb concentration divided by mother's blood Pb
11 concentration). Group mean ratios range from about 0.7 to 1.0 at the time of delivery for
12 mean maternal blood Pb levels ranging from 1.7 to 8.6 (ig/dL (Amaral et al.. 2010;
13 Kordas et al.. 2009; Patel and Prabhu. 2009; Carbone et al.. 1998; Gover. 1990; Graziano
14 et al.. 1990). In a study of 159 mothers having blood Pb levels of less than 14 (ig/dL,
15 based on a linear regression of maternal blood Pb and cord blood Pb, the ratio of cord
16 blood Pb to maternal blood Pb appeared to decrease with decreasing maternal blood Pb
17 from 1.0 at 10 (ig/dL to 0.34 at 3 (ig/dL (Carbone et al.. 1998). A ratio of 0.34 is lower
18 than reported based on mean data in other studies. However, consistent with other
19 studies, the ratio of mean cord blood Pb (4.87 ug/dL) to mean maternal blood Pb (5.81
20 ug/dL) was 0.84. In addition, the similarity of isotopic ratios in maternal blood and in
21 blood and urine of newly-born infants provide further evidence for placental transfer of
22 Pb to the fetus (Gulson et al.. 1999).
23 Transplacental transfer of Pb may be facilitated by an increase in the plasma/blood Pb
24 concentration ratio during pregnancy (Montenegro et al.. 2008; Lamadrid-Figueroa et al..
25 2006). Maternal-to-fetal transfer of Pb appears to be related partly to the mobilization of
26 Pb from the maternal skeleton. Evidence for transfer of maternal bone Pb to the fetus has
27 been provided by stable Pb isotope studies in cynomolgus monkeys exposed during
28 pregnancy. Approximately 7-39% of the maternal Pb burden transferred to the fetus was
29 derived from the maternal skeleton, with the remainder derived from contemporaneous
30 exposure (O'Flaherty. 1998; Franklin et al.. 1997). The upper value in the range (39%)
31 represented the one monkey with historical Pb exposure, but received only small amounts
32 of environmental Pb exposure during pregnancy; for the monkeys that received high
33 doses of Pb during pregnancy, the range was lower (7-25%) (O'Flahertv. 1998; Franklin
34 etal.. 1997).
November 2012 4-49 Draft - Do Not Cite or Quote
-------
4.2.2.5 Organic Pb
1 Information on the distribution of Pb in humans following exposures to organic Pb is
2 extremely limited. However, as reported in the 2006 Pb AQCD (U.S. EPA. 2006c). the
3 available evidence demonstrates near complete absorption following inhalation of
4 tetraalkyl Pb vapor and subsequent transformation to trialkyl Pb metabolites. One hour
5 following brief inhalation exposures to 203Pb tetraethyl or tetramethyl Pb (1 mg/m3),
6 -50% of the 203Pb body burden was associated with liver and 5% with kidney; the
7 remaining 203Pb was widely distributed throughout the body (Heard etal.. 1979). The
8 kinetics of 203Pb in blood showed an initial declining phase during the first 4 hours
9 (tetramethyl Pb) or 10 hours (tetraethyl Pb) after the exposure, followed by a
10 reappearance of radioactivity back into the blood after ~20 hours. The high level of
11 radioactivity initially in the plasma indicates the presence of tetraalkyl/trialkyl Pb. The
12 subsequent rise in blood radioactivity, however, probably represents water-soluble
13 inorganic Pb and trialkyl and dialkyl Pb compounds that were formed from the metabolic
14 conversion of the volatile parent compounds (Heard etal.. 1979).
15 Alkyl Pb compounds undergo oxidative dealkylation catalyzed by cytochrome P450 in
16 liver and, possibly, in other tissues. Trialkyl Pb metabolites have been found in the liver,
17 kidney, and brain following exposure to the tetraalkyl compounds in workers
18 (Bolanowska et al.. 1967); these metabolites have also been detected in brain tissue of
19 nonoccupational subjects (Nielsen et al.. 1978).
4.2.3 Elimination
20 The rapid-phase (30-40 days) of Pb excretion amounts to 50-60% of the absorbed
21 fraction (Chamberlain et al.. 1978; Rabinowitz et al., 1976; Kehoe. 1961a. b. c).
22 Absorbed Pb is excreted primarily in urine and feces, with sweat, saliva, hair, nails, and
23 breast milk being minor routes of excretion (Kehoe. 1987; Chamberlain et al.. 1978;
24 Rabinowitz et al.. 1976; Griffin etal.. 1975; Hurshetal.. 1969; Hursh and Suomela.
25 1968).
26 Approximately 30% of intravenously injected Pb in humans (40-50% in beagles and
27 baboons) is excreted via urine and feces during the first 20 days following administration
28 (Leggett 1993). The kinetics of urinary excretion following a single dose of Pb is similar
29 to that of blood (Chamberlain et al.. 1978). likely due to the fact that Pb in urine derives
30 largely from Pb in plasma. Evidence for this is the observation that urinary Pb excretion
31 is strongly correlated with the rate of glomerular filtration of Pb (Araki etal.. 1986) and
32 plasma Pb concentration (Rentschler et al., 2012; Bergdahl et al., 1997b) (i.e., glomerular
November 2012 4-50 Draft - Do Not Cite or Quote
-------
1 filtration rate x plasma Pb concentration), and both relationships are linear. While the
2 relationship between urinary Pb excretion and plasma Pb concentration is linear, the
3 plasma Pb relationship to blood Pb concentration is curvilinear (as described in Section
4 4.2.2.1 and demonstrated in Figure 4-6). This relationship contributes to an increase in
5 the renal clearance of Pb from blood with increasing blood Pb concentrations
6 (Chamberlain. 1983). Similarly, a linear relationship between plasma Pb concentration
7 and urinary excretion rate predicts a linear relationship between Pb intake (at constant
8 absorption fraction) and urinary Pb excretion rate, whereas the relationship with blood Pb
9 concentration would be expected to be curvilinear (Section 4.3.6).
10 Estimates of urinary filtration of Pb from plasma range from 13-22 L/day, with a mean of
11 18 L/day (Arakietal. 1986; Manton and Cook. 1984; Manton and Mallov. 1983;
12 Chamberlain et al.. 1978). which corresponds to half-time for transfer of Pb from plasma
13 to urine of 0.1-0.16 days for a 70-kg adult who has a plasma volume of ~3 L. The rate of
14 urinary excretion of Pb was less than the rate of glomerular filtration of ultrafilterable Pb,
15 suggesting that urinary Pb is the result of incomplete renal tubular re-absorption of Pb in
16 the glomerular filtrate (Araki etal. 1986): although, net tubular secretion of Pb has been
17 demonstrated in animals (Victery etal.. 1979; Vander et al.. 1977). On the other hand,
18 estimates of blood-to-urine clearance range from 0.03-0.3 L/day with a mean of 0.18
19 L/day (Diamond. 1992; Arakietal.. 1990; Bergeretal. 1990; Kosteretal.. 1989;
20 Manton and Malloy. 1983; Ryu etal.. 1983; Chamberlain et al.. 1978; Rabinowitz et al..
21 1973). consistent with a plasma Pb to blood Pb concentration ratio of-0.005-0.01 L/day
22 (Klotzback et al.. 2003). Based on the above differences, urinary excretion of Pb can be
23 expected to reflect the concentration of Pb in plasma and variables that affect delivery of
24 Pb from plasma to urine (e.g., glomerular filtration and other transfer processes in the
25 kidney).
26 The value for fecal:urinary excretion ratio (-0.5) was observed during days 2-14
27 following intravenous injection of Pb in humans (Chamberlain et al.. 1978; Booker et al..
28 1969; Hurshetal.. 1969). This ratio is slightly higher (0.7-0.8) with inhalation of
29 submicron Pb-bearing PM due to ciliary clearance and subsequent ingestion. The transfer
30 of Pb from blood plasma to the small intestine by biliary secretion in the liver is rapid
31 (adult ti/2 =10 days), and accounts for 70% of the total plasma clearance (O'Flahertv.
32 1995).
November 2012 4-51 Draft - Do Not Cite or Quote
-------
Organic Pb
1 Pb absorbed after inhalation of tetraethyl and tetramethyl Pb is excreted in exhaled air,
2 urine, and feces (Heard et al.. 1979). Fecal:urinary excretion ratios were 1.8 following
3 exposure to tetraethyl Pb and 1.0 following exposure to tetramethyl Pb (Heard et al.,
4 1979). Occupational monitoring studies of workers exposed to tetraethyl Pb showed that
5 tetraethyl Pb is excreted in the urine as diethyl Pb, ethyl Pb, and inorganic Pb (Vural and
6 Duvdu. 1995: Zhang etal.. 1994: Turlakiewicz and Chmielnicka. 1985).
4.3 Pb Biomarkers
7 This section describes the biological measurements of Pb and their interpretation as
8 indicators of exposure or body burden.
9 For any health endpoint of interest, the most useful biomarker of exposure is one that
10 provides information about the Pb dose at the critical target organ and, moreover, reflects
11 the exposure averaging time that is appropriate to the underlying pathogenetic processes
12 (e.g., instantaneous, cumulative over lifetime, or cumulative over a circumscribed age
13 range). In recent studies of Pb and health, the exposure biomarkers most frequently used
14 are Pb in blood and bone. For outcomes other than those relating to hematopoiesis and
15 bone health, these biomarkers provide information about Pb dose that is some distance
16 from the target organ. For example, given that the central nervous system is considered
17 the critical target organ for childhood Pb toxicity, it would be most helpful to be able to
18 measure, in vivo, the Pb concentrations at the cellular site(s) of action in the brain.
19 However, because such measurements are not currently feasible, investigators must rely
20 on measurements of Pb in the more readily accessible but peripheral tissues. The
21 relationship between brain Pb and Pb in each of these surrogate tissues is still poorly
22 understood, although the pharmacokinetics clearly differs among these compartments.
23 As an exposure biomarker, blood Pb concentration has other limitations. Only about 1%
24 of an individual's total body Pb burden resides in blood. Furthermore, blood consists of
25 several subcompartments. More than 90% of Pb in whole blood is bound to red cell
26 proteins such as ALAD, with the balance in plasma. From a toxicological perspective, the
27 unbound fraction is likely to be the most important subcompartment of blood Pb because
28 it distributes into soft tissues. The concentration of Pb in plasma is much lower than in
29 whole blood (<1%). The greater relative abundance of Pb in whole blood makes its
30 measurement much easier (and more affordable) than measurement of Pb in plasma. The
31 use of whole blood Pb as a surrogate for plasma Pb could be justified if the ratio of whole
32 blood Pb to plasma Pb were well characterized, but this is not so. At least some studies
33 suggest that it varies several-fold among individuals with the same blood Pb level.
November 2012 4-52 Draft - Do Not Cite or Quote
-------
1 Moreover, binding Pb in red blood cells is limited, so the ratio of blood Pb to plasma Pb
2 would be expected to be nonlinear. Thus, interpreting whole blood Pb level as a proxy for
3 plasma Pb level, which, itself, is a proxy for brain Pb level, will result in some exposure
4 misclassification.
5 Another limitation of blood Pb as an exposure biomarker is that the kinetics of Pb in
6 blood is relatively fast compared to the kinetics of Pb in bone, and therefore, of the whole
7 body burden. Thus, a high blood Pb concentration measured at any given time does not
8 necessarily indicate a high body Pb burden. Similarly, individuals who have the same
9 blood Pb level will not necessarily have similar body burdens or exposure histories. The
10 rate at which blood Pb changes with time/age depends on exposure history due to re-
11 equilibration of Pb stored in the various body pools.
12 The development of X-ray-fluorescence (XRF) methods for measuring Pb in mineralized
13 tissues offers another approach for characterization and reconstruction of exposure
14 history. Such tissues are long-term Pb storage sites, with a half-life measured in decades
15 and contain -90% of the total body Pb burden in adults and 70% in children. Thus, bone
16 Pb reflects a long exposure averaging time.
17 Mechanistic models are used throughout the section as a means to describe basic
18 concepts that derive from the wealth of information on Pb toxicokinetics. Although
19 predictions from models are inherently uncertain, models can serve to illustrate expected
20 interrelationships between Pb intake and tissue distribution that are important in
21 interpreting human clinical and epidemiologic studies. Thus, models serve as the only
22 means available for synthesizing the extensive, but incomplete, knowledge of Pb
23 biokinetics into a holistic representation of Pb biokinetics. Furthermore, models can also
24 be used to make predictions about biokinetics relationships that have not been thoroughly
25 evaluated in experiments or epidemiologic studies. In this way, models can serve as
26 heuristic tools for shaping data collection to improve understanding of Pb biokinetics.
27 Mechanistic toxicokinetics models can make predictions about hypothetical populations
28 and exposure scenarios. When a model is run as a single simulation, the output represents
29 average outcomes from what is in reality a distribution of possible outcomes that would
30 be expected in the population (or in any single individual) where intra-individual and
31 inter-individual variability in exposure and toxicokinetics exist. More realistic predictions
32 for the population can be developed by running a series of model simulations in which
33 ranges (i.e., distributions) of parameter values are considered that may better represent
34 the population of interest. In this section, only single simulations are used to demonstrate
35 relationships between various biomarkers (e.g., blood Pb and bone Pb) that would apply
36 to a population having "typical" or "average" exposure and toxicokinetics. These single
November 2012 4-53 Draft - Do Not Cite or Quote
-------
1 simulations are used for illustrative purposes to describe general concepts and patterns.
2 Variability would be expected in real populations.
3 Numerous mechanistic models of Pb biokinetics in humans have been proposed, and
4 these are described in the 2006 Pb AQCD (U.S. EPA. 2006b) and in the supporting
5 literature cited in that report. In this section, for simplicity and for internal consistency,
6 discussion is limited to predictions from a single model, the ICRP Pb biokinetics model
7 (Pounds and Leggett. 1998: ICRP. 1994: Leggett. 1993). The ICRP model consists of a
8 systemic biokinetics model (Leggett, 1993) and a human respiratory tract model (ICRP.
9 1994). The Leggett model simulates age-dependent kinetics of tissue distribution and
10 excretion of Pb ingestion and inhalation intakes. This model was originally developed for
11 the purpose of supporting radiation dosimetry predictions and it has been used to develop
12 cancer risk coefficients for internal radiation exposures to Pb and other alkaline earth
13 elements that have biokinetics similar to those of calcium (ICRP. 1993). Although the
14 ICRP model has not been validated by U.S. EPA as a regulatory model for Pb risk
15 assessment, it has been applied in Pb risk assessment (Abrahams et al.. 2006: Lorenzana
16 et al.. 2005: Khoury and Diamond. 2003). Portions of the model have been incorporated
17 into an AALM that is being developed by EPA (2005a). In addition to the above
18 considerations regarding previous applications of the ICRP model, the model was
19 selected for use in the ISA because it has several useful features for predicting exposure-
20 body burden relationships. The model simulates blood Pb and tissue Pb concentration
21 dynamics associated with the uptake and elimination phases of exposures of > 1 day in
22 duration; and it simulates age-dependent and particle size-dependent deposition and
23 clearance of inhaled Pb in the respiratory tract. These types of simulations can only be
24 approximated with the U.S. EPA IEUBK Model for Pb in children because it simulates
25 exposures in time steps of 1 year (i.e., age-year average exposures); lumps the simulation
26 of deposition, mechanical clearance, and absorption of inhaled Pb into a single absorption
27 term representing the combined processes of gastrointestinal and respiratory tract
28 absorption of inhaled Pb; simulates steady state blood Pb concentrations and was does
29 not allow access to the underlying simulations of tissue Pb concentrations which serve as
30 intermediate variables in the model for predicting steady state blood Pb concentrations.
31 Other models have been developed that allow simulations of tissue Pb concentrations
32 (e.g., O'Flahertv. 1995: Leggett. 1993) and comparisons of these models have been
33 previously described (Maddaloni et al.. 2005).
November 2012 4-54 Draft - Do Not Cite or Quote
-------
1 Pb biokinetics in adolescents is poorly characterized by all existing Pb biokinetics
2 models. Individuals undergo rapid changes in sexual development, growth, food and
3 water intake, bone growth and turnover, behavior, etc. during adolescence. There is a
4 paucity of experimental measurements of Pb biomarkers during this time developmental
5 window. The individual biological and kinetic parameters for adolescents are largely
6 interpolated rather than based on solid experimental and toxicological measurements.
7 These deficiencies limit the validity of model predictions in this age group.
4.3.1 Bone-Pb Measurements
8 For Pb measurements in bone, the most commonly examined bones are the tibia,
9 calcaneus, patella, and finger bone. For cortical bone, the midpoint of the tibia is
10 measured. For trabecular bone, both the patella and calcaneus are measured. The tibia
11 consists of more than 95% cortical bone, the calcaneus and patella comprise more than
12 95% trabecular bone, and finger bone is a mixed cortical and trabecular bone although
13 the second phalanx is dominantly cortical. Recent studies favor measurement of the
14 patella for estimating trabecular bone Pb, because it has more bone mass and may afford
15 better measurement precision than the calcaneus.
16 Bone Pb measurements are typically expressed in units of ug Pb per g bone mineral. This
17 convention may potentially introduce variability into the bone Pb measurements related
18 to variation in bone density. Typically, potential associations between bone density and
19 bone Pb concentration are not evaluated in epidemiologic studies (Theppeang et al..
20 2008a; Hu et al., 2007a). An important consequence of expressing bone Pb measures
21 relative to bone mineral content is that lower bone mineral density is associated with
22 greater measurement uncertainty in bone Pb. This can have important implications for
23 studies in older women for whom low bone mineral density is more common than in
24 other populations including men and younger adults.
25 Methods of direct analysis of bone tissue samples include flame atomic absorption
26 spectrometry (AAS), anode stripping voltammetry (ASV), inductively coupled plasma
27 atomic emission spectroscopy (ICP-AES), inductively coupled plasma mass spectrometry
28 (ICP-MS), laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS),
29 thermal ionization mass spectrometry (TIMS), synchrotron radiation induced X-ray
30 emission (SRIXE), particle induced X-ray emission (PIXE), and X-ray fluorescence
31 (XRF). Non-invasive, in vivo measurements of bone Pb is achieved with XRF. The
32 upsurge in popularity of the XRF method has paralleled a decline in the use of the other
33 methods. More information on the precision, accuracy, and variability in bone Pb
34 measurements can be found in the 2006 Pb AQCD (U.S. EPA. 2006b).
November 2012 4-55 Draft - Do Not Cite or Quote
-------
1 Two main approaches for XRF measurements have been used to measure Pb
2 concentrations in bone, the K-shell and L-shell methods. The K-shell method is the most
3 widely used, as there have been relatively few developments in L-shell devices since the
4 early 1990s. However, Nie et al. (2011 a) recently reported on the use of a new portable
5 L-shell device for human in vivo Pb measurements. Advances in L-shell device
6 technology resulted in much higher sensitivity than previous L-shell devices. The new
7 L-shell device showed sensitivity similar to that of K-shell methods (detection limit was
8 approximately 8 (ig/g bone mineral with 2 mm of soft tissue overlay targeted bone) and a
9 high correlation with results obtained from K-shell methods (intraclass
10 correlation = 0.65). Behinaen et al. (2011) described application of a 4-detector system
11 ("clover leaf array ") for the K-shell method that provided higher precision and lower
12 minimum detection limits (MDL) for tibia and calcaneus Pb measurements (3.25 and
13 4.78 (ig/g bone mineral, respectively) compared to measurements made with single
14 detectors (8-12 (ig/g and 14-15 (ig/g, respectively).
15 Since 1986, several investigators have reported refinements to hardware and software to
16 improve the precision and accuracy of XRF measurements and there have been a number
17 of investigations into the precision, accuracy and variability in XRF measurements [e.g.,
18 (Todd et al.. 2002: Toddetal.. 2001: Aro et al.. 2000: Todd et al.. 2000)1. Todd et al.
19 (2000) provided a detailed discussion of factors that influence the variability and
20 measurement uncertainty, including repositioning, sample measurement duration,
21 overlying tissue, operator expertise, detector resolution, and changes to measurement
22 process over time. Some of these aspects were also discussed by Hu et al. (1995). From
23 their cadaver and in vivo measurements, Todd et al. (2000) concluded that the uncertainty
24 in an individual measurement was an underestimate of the standard deviation of replicate
25 measurements, suggesting a methodological deficiency probably shared by most current
26 109Cd-based K-shell XRF Pb measurement systems. In examining the reproducibility of
27 the bone Pb measurements over a 4!/2 month period, Todd et al. (2000) also found the
28 average difference between the XRF results from short term and longer term
29 measurements was 1.2 (ig/g, indicating only a small amount of variability in the XRF
30 results over a sustained period of time.
31 In the epidemiologic literature, XRF bone Pb data have typically been reported in two
32 ways: one that involves a methodological approach to assessing the minimum detection
33 limit and the other termed an epidemiologic approach by Rosen and Pounds (1998). In
34 the former approach, a minimum detection limit is defined using various methods,
35 including two or three times the square root of the background counts; one, two, or three
36 times the standard deviation (SD) of the background; or two times the observed median
37 error. This approach relies upon the minimum detection limit to define a quantitative
38 estimate that is of sufficient precision to be included in the statistical analysis, as
November 2012 4-56 Draft - Do Not Cite or Quote
-------
1 demonstrated by Bellinger et al. Q994a), Gerhardsson et al. (1993). and Christoffersson
2 et al. (1986).
3 With the epidemiologic approach, all values are used (including negative values) to
4 determine the minimum detection limit of an instrument that results in extremely low
5 detection limits. Rosen and Pounds (1998) noted that this approach yields population
6 bone Pb averages that were artificially low. However, not including values that are
7 negative or below the detection limit, or assigning these values a fixed number is also of
8 concern. Using the epidemiologic approach of retaining all point estimates of measured
9 bone Pb concentrations provided the least amount of bias and the greatest efficiency in
10 comparing the mean or median levels of bone Pb of different populations (Kim et al..
11 1995).
4.3.2 Blood-Pb Measurements
12 Analytical methods for measuring Pb in blood include AAS, graphite furnace atomic
13 absorption spectrometry (GFAAS), ASV, ICP-AES, and ICP-MS. GFAAS and ASV are
14 generally considered to be the methods of choice (Flegal and Smith. 1995). Limits of
15 detection for Pb using AAS are on the order of 5-10 (ig/dL for flame AAS measurements
16 and approximately 0.1 (ig/dL for flameless AAS measurements (Flegal and Smith. 1995;
17 NIOSH. 1994). A detection limit of 0.005 (ig/dL has been achieved for Pb in blood
18 samples analyzed by GFAAS.
19 For measurement of Pb in plasma, ICP-MS provides sufficient sensitivity (Schutz et al..
20 1996). While the technique has been applied to assessing Pb exposures in adults, ICP-MS
21 has not received widespread use in epidemiologic studies.
22 The primary binding ligand for Pb in RBC, ALAD, is encoded by a single gene in
23 humans that is polymorphic in two alleles (ALAD1 and ALAD2) (Scinicariello et al..
24 2007). Since the ALAD1 and ALAD 2 alleles can be co-dominantly expressed, 3
25 different genotypes (ALAD 1-1, ALAD 1-2, and ALAD 2-2) are possible. The ALAD
26 1-1 genotype is the most common. Scinicariello et al. (2010) tested genotypes in civilian,
27 non-institutionalized U.S. individuals that participated as part of NHANES III from
28 1988-1994 and found that 15.6% of non-Hispanic whites, 2.6% non-Hispanic blacks, and
29 8.8% Mexican Americans carried the ALAD2 allele.
30 The 2006 Pb AQCD (U.S. EPA. 2006c) reported that many studies have shown that, with
31 similar exposures to Pb, individuals with the ALAD-2 allele have higher blood Pb levels
32 than those without (Kim et al.. 2004; Perez-Bravo et al.. 2004; Bergdahl et al.. 1997b:
33 Smith et al.. 1995a: Wetmur. 1994; Wetmuretal. 1991b: Astrinetal.. 1987). More
November 2012 4-57 Draft - Do Not Cite or Quote
-------
1 recent meta analyses provide further support for ALAD2 carriers having higher blood Pb
2 levels than ALAD1 -1 homozygotes (Scinicariello et al.. 2007; Zhao et al.. 2007). The
3 mechanism for this association may be higher Pb binding affinity of ALAD2. Although,
4 this interpretation would be consistent with the structural differences that result in greater
5 electronegativity of ALAD 1 compared to ALAD2 (Wetmur. 1994; Wetmur et al.. 199 la).
6 measurements of Pb binding affinity to ALAD1 and ALAD2 (i.e., Pb2+ displacement of
7 Zn2+ binding to recombinant ALAD 1 and ALAD2) have not revealed differences in Pb
8 binding affinity (Jaffe et al.. 2000). In a meta-analysis of 24 studies, Scinicariello et al.
9 (2007). observed the greatest differences for ALAD2 compared to ALAD1 in highly
10 exposed adults with little difference among environmentally-exposed adults; large
11 differences were also observed for children at low exposures. However, there are few
12 studies that evaluated children and the largest study contributing to the meta analysis may
13 have been influenced by selection bias (Scinicariello et al.. 2007). Individual studies find
14 similar results in occupationally-exposed adults, with blood Pb levels being higher in
15 individuals with ALAD2 alleles (Miyaki et al.. 2009; Shaik and Jamil. 2009). A
16 subsequent meta analysis of adult data from NHANES III did not find any differences in
17 blood Pb level between all carriers of either the ALAD 1-1 or ALAD 1-2/2-2 allele
18 (Scinicariello et al.. 2010). Other studies provide further support for no blood Pb
19 differences among ALAD1 and ALAD2 carriers (Sobin et al.. 2009; Rabstein et al..
20 2008; Montenegro et al.. 2006; Wananukul et al.. 2006) or lower blood Pb levels for
21 individuals with ALAD 1 -2/2-2 (Krieg et al.. 2009; Chia et al.. 2006).
22 Genetic polymorphism in the gene that encodes for peptide transporter 2 (PEPT2) has
23 been associated with variability in blood Pb concentrations in children (Sobin et al..
24 2009). PEPT2 expression in the brain and renal proximal tubule has been associated with
25 transport of di- and tri-peptides and may function in the transport of 5-ALA into brain
26 and renal tubular re-absorption of peptides. The PRPT2*2 polymorphism was associated
27 with increased blood Pb concentrations in a sample of 116 children of Mexican-
28 American/Hispanic heritage (age 4-12 years, mean blood Pb concentration 3-6 (ig/dL).
29 Analyses of serial blood Pb concentrations measured in longitudinal epidemiologic
30 studies found relatively strong correlations (e.g., r = 0.5-0.8) among each child's
31 individual blood Pb concentrations measured after 6-12 months of age (Schnaas et al..
32 2000; Dietrich et al.. 1993a; McMichael et al.. 1988; Ottoetal.. 1985; Rabinowitz et al..
33 1984). These observations suggest that, in general, exposure characteristics of an
34 individual child (e.g., exposure levels and/or exposure behaviors) tend to be relatively
35 constant across age. However, a single blood Pb measurement may not distinguish
36 between a history of long-term lower-level Pb exposure from a history that includes
37 higher acute exposures (Mushak. 1998). This concept is illustrated in Figure 4-7. Two
38 hypothetical children are simulated. Child A has a relatively constant Pb intake from
November 2012 4-58 Draft - Do Not Cite or Quote
-------
1 birth, whereas Child B has the same Pb intake as Child A for the first two years of life,
2 then a 1-year elevated intake beginning at age 24 months (Figure 4-7. upper panel) that
3 returns to the same intake as Child A at 36 months. The absorption fraction is assumed to
4 be the same for both children. Blood Pb samples 1 and 5 for Child A and B, or 2 and 4
5 for Child B, will yield similar blood Pb concentrations (~3 or 10 (ig/dL, respectively), yet
6 the exposure contexts for these samples are very different. Two samples (e.g., 1 and 2, or
7 4 and 5), at a minimum, are needed to ascertain if the blood Pb concentration is changing
8 over time. The rate of change can provide information about the magnitude of change in
9 exposure, but not necessarily about the time history of the change (Figure 4-7. lower
10 panel). Time-integrated measurements of Pb concentration may provide a means for
11 accounting for some of these factors and, thereby, provide a better measure of long-term
12 Pb exposure.
November 2012 4-59 Draft - Do Not Cite or Quote
-------
0 12 24 36 48 60 72 84
Age (months)
O)
o>
40
30
20 ^
m
0
0 12 24 36 48 60
Age (months)
72 84
Note: Child A and Child B had a constant basal Pb intake (10 |jg/day) from birth; Child B experienced an elevated intake of
5.5 |jg/day/kg for 1 year beginning at 24 months of age (upper panel). Blood Pb measurements 1 and 5 for Child A and B, or 2 and
4 for Child B, will yield similar blood Pb concentrations (~3 or 10 ug/dL, respectively), yet the exposure scenarios for these samples
are very different. As shown in the example of Child C and Child D, two blood Pb measurements can provide information about the
magnitude of change in exposure, but not necessarily the temporal history of the change (lower panel). Child C and D had a
constant basal Pb intake (10 ug/day) from birth. Child C experienced an elevated intake of 13 ug/day starting at 12 months of age
for 1 year, whereas, Child D experienced an elevated intake of 5.5 ug/day starting at 24 months of age for 1 year. Simulation based
on ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-7 Simulation of temporal relationships between Pb exposure and
blood Pb concentration in children.
November 2012
4-60
Draft - Do Not Cite or Quote
-------
4.3.3 Urine-Pb Measurements
1 Standard methods that have been reported for urine Pb analysis are, in general, the same
2 as those analyses noted for determination of Pb in blood. Reported detection limits are
3 -50 (ig/L for AAS, 5-10 (ig/L for ICP AES, and 4 (ig/L for ASV for urine Pb analyses.
4 The concentration of Pb in urine is a function of the urinary Pb excretion (Section 4.2.3)
5 and the urine flow rate. Urine flow rate requires collection of a timed urine sample, which
6 is often problematic in epidemiologic studies. Collection of untimed ("spot") urine
7 samples, a common alternative to timed samples, requires adjustment of the Pb
8 measurement in urine to account for variation in urine flow (Diamond. 1988). Several
9 approaches to this adjustment have been explored, including adjusting the measured urine
10 Pb concentration by the urine creatinine concentration, urine osmolality, or specific
11 gravity (Fukui et al., 1999; Araki et al., 1990). Urine flow rate can vary by a factor or
12 more than 10, depending on the state of hydration and other factors that affect glomerular
13 filtration rate and renal tubular reabsorption of the glomerular filtrate. All of these factors
14 can be affected by Pb exposure at levels that produce nephrotoxicity (i.e., decreased
15 glomerular filtration rate, impaired renal tubular transport function). Therefore, urine Pb
16 concentration measurements provide little reliable information about exposure (or Pb
17 body burden), unless they can be adjusted to account for unmeasured variability in urine
18 flow rate (Araki etal. 1990).
19 Urinary Pb concentration reflects, mainly, the concentration of Pb in the blood. As such,
20 urinary concentrations reflect both recent and past exposures to Pb (see Section 4.3.5). A
21 single urinary Pb measurement cannot distinguish between a long-term low level of
22 exposure or a higher acute exposure. Urinary Pb measurements would be expected to
23 correlate with concurrent blood Pb (see Section 4.3.6 for additional discussion of the
24 relationship between blood and urine Pb). Chiang et al. (2008) reported a significant, but
25 relatively weak correlation between urinary Pb levels ((ig/dg creatinine) and individual
26 Pb intakes ((ig/day) estimated in a group of 10- to 12-year-old children (|3: 0.053,
27 R = 0.320, p = 0.02, n = 57). A contributing factor to the relatively weak correlation may
28 have been the temporal displacement between the urine sampling and measurements used
29 to estimate intake, which may have been as long as 6 months for some children.
30 Thus, a single urine Pb measurement, or a series of measurements taken over short-time
31 span, is likely a relatively poor index of Pb body burden for the same reasons that blood
32 Pb is not a good indicator of body burden. On the other hand, long-term average
33 measurements of urinary Pb can be expected to be a better index of body burden (Figure
34 4;8.
November 2012 4-61 Draft - Do Not Cite or Quote
-------
10
re
5
O)
8 -
6 -
£1
0.
0)
•I 4
2 -
"" '
— j
20
15 CD
o
Q.
<
CD
10 I.
CD
3
(Q
- 5
25 30 35 40 45 50 55 60 65 70
Age (year)
o
I 4
Its
SI
o
2 -
1 -I
20
15 oo
o
0.
<
03
10 a
o
3
(Q
- 5
25 30 35 40 45 50 55 60 65 70
Age (year)
Note: A change in Pb uptake results in a relatively rapid change in urinary excretion of Pb, to a new quasi-steady state, and a
relatively small change in body burden (upper panel). Baseline ingestion was 20 ug/day from age 0 to 30 yrs, then intake increased
to 120 ug/day from age 30 to 50 with a subsequent decrease in intake to the baseline of 20 ug/day at age 50. The long-term
average urinary Pb excretion more closely tracks the pattern of change in body burden (lower panel). Simulation based on ICRP Pb
biokinetics model (Leggett. 1993).
Figure 4-8 Simulation of relationship between urinary Pb excretion and body
burden in adults.
November 2012
4-62
Draft - Do Not Cite or Quote
-------
4.3.4 Pb in Other Biomarkers
1 There was extensive discussion in the 2006 Pb AQCD (U.S. EPA. 2006c) regarding the
2 utility of other Pb biomarkers as indicators of exposure or body burden. Due to the fact
3 that most epidemiologic studies continue to use blood Pb or bone Pb, and other potential
4 biomarkers (i.e., teeth, hair, and saliva) have not been established to the same extent as
5 blood or bone Pb, only summaries are provided below.
4.3.4.1 Teeth
6 Tooth Pb is a minor contributor to the total body burden of Pb. As teeth accumulate Pb,
7 tooth Pb levels are generally considered an estimate of cumulative Pb exposure. The
8 tooth Pb-blood Pb relationship is more complex than the bone Pb-blood Pb relationship
9 because of differences in tooth type, location, and analytical method. Although
10 mobilization of Pb from bone appears well established, this is not the case for Pb in teeth.
11 Conventional wisdom has Pb fixed once it enters the tooth. Although that may be the case
12 for the bulk of enamel, it is not true for the surface of the enamel and dentine (Gulson et
13 al.. 1997; Rabinowitz et al., 1993). Limited studies have demonstrated moderate-to-high
14 correlations between tooth Pb levels and blood Pb levels (Rabinowitz. 1995; Rabinowitz
15 etal.. 1989).
16 Teeth are composed of several tissues formed pre- and postnatal. Therefore, if a child's
17 Pb exposure during the years of tooth formation varied widely, different amounts of Pb
18 would be deposited at different rates (Rabinowitz et al., 1993). This difference may allow
19 investigators to elucidate the history of Pb exposure in a child. Robbins et al. (2010)
20 found a significant association between environmental Pb measures that correlated with
21 leaded gasoline use and tooth enamel Pb in permanent teeth. Costa de Almeida et al.
22 (2007) discerned differences between tooth enamel Pb concentration in biopsy samples
23 from children who lived in areas having higher or lower levels of Pb contamination.
24 Gulson and Wilson (1994) advocated the use of sections of enamel and dentine to obtain
25 additional information compared with analysis of the whole tooth (e.g., (Tvinnereim et
26 al.. 1997; Fosse etal.. 1995). For example, deciduous tooth Pb in the enamel provides
27 information about in utero exposure whereas that in dentine from the same tooth provides
28 information about postnatal exposure until the tooth exfoliates at about 6-7 years of age.
November 2012 4-63 Draft - Do Not Cite or Quote
-------
4.3.4.2 Hair
1 The 2006 Pb AQCD (U.S. EPA. 2006c) discussed applications of hair Pb measurements
2 for assessing Pb body burden or exposure and noted methodological limitations
3 (e.g., external contamination) and lack of a strong empirical basis for relating hair Pb
4 levels to body burden or exposure. No new methodological or conceptual advances
5 regarding hair Pb measurements have occurred since 2006, and widespread application of
6 hair Pb measurements in epidemiologic studies has not occurred.
7 Pb is incorporated into human hair and hair roots (Bos et al.. 1985; Rabinowitz et al.,
8 1976) and has been explored as a noninvasive approach for estimating Pb body burden
9 rWilhelm et al.. 2002; Gerhardsson et al.. 1995; Wilhelm et al.. 1989). Hair Pb
10 measurements are subject to error from contamination of the surface with environmental
11 Pb and contaminants in artificial hair treatments (i.e., dyeing, bleaching, permanents) and
12 are a relatively poor predictor of blood Pb concentrations, particularly at blood Pb levels
13 less than 10-12 (ig/dL (Rodrigues et al.. 2008; Campbell and Toribara. 2001; Esteban et
14 al.. 1999; Drasch et al.. 1997). Temporal relationships between Pb exposure and hair Pb
15 levels, and kinetics of deposition and retention of Pb in hair have not been evaluated.
16 Although hair Pb measurements have been used in some epidemiologic studies (Shah et
17 al.. 2011; U.S. EPA. 2006b). an empirical basis for interpreting hair Pb measurements in
18 terms of body burden or exposure has not been firmly established.
4.3.4.3 Saliva
19 A growing body of literature on the utility of measurements of salivary Pb has developed
20 since the completion of the 2006 Pb AQCD (U.S. EPA. 2006b). Earlier reports suggested
21 a relatively strong correlation between salivary Pb concentration and blood Pb
22 concentration (Omokhodion and Crockford. 1991; Brodeur et al.. 1983; P'an. 1981);
23 however, more recent assessments have shown relatively weak or inconsistent
24 associations (Costa de Almeida et al.. 2011; Costa de Almeida et al.. 2010; Costa de
25 Almeida et al., 2009; Barbosa et al., 2006a; Nriagu et al., 2006). The differences in these
26 outcomes may reflect differences in blood Pb concentrations, exposure history and/or
27 dental health (i.e., transfer of Pb between dentin and saliva) and possibly methods for
28 determining Pb in saliva. Barbosa et al. (2006a) found a significant but relatively weak
29 correlation (logfblood PB] versus logfsaliva Pb], r = 0.277, p = 0.008) in a sample of
30 adults, ages 18-60 years (n = 88). The correlation was similar for salivary and plasma Pb.
31 Nriagu et al. (2006) found also found a relatively weak association (R2 ~ 0.026) between
32 blood Pb ((ig/dL) and salivary Pb ((ig/L) in a sample of adults who resided in Detroit, MI
33 (n = 904). Costa de Almeida et al. (2009) found a significant correlation between salivary
November 2012 4-64 Draft - Do Not Cite or Quote
-------
1 and blood Pb concentrations in children in a Pb-contaminated region in Sao Paulo State,
2 Brazil (r = 0.76. p = 0.04, n = 7) prior to site remediation; however, the correlation
3 degenerated (r = 0.03, p = 0.94, n = 9) following remediation. Nevertheless, salivary Pb
4 concentrations in the group of children who lived in the contaminated area were
5 significantly elevated compared to a reference population. It is possible, that salivary Pb
6 measurements may be a useful non-invasive biomarker for detecting elevated Pb
7 exposure; however, it is not clear based on currently available data, if salivary Pb
8 measurements would be a more reliable measure of exposure than blood Pb
9 measurements.
4.3.4.4 Serum 5-ALA and ALAD
10 The association between blood Pb and blood ALAD activity and serum 5-aminolevulinic
11 acid (5-ALA) levels was recognized decades ago as having potential use as a biomarker
12 of Pb exposure (Mitchell et al., 1977; Hernberg et al., 1970). More recently reference
13 values for blood ALAD activity ratio (the ratio of ALAD activity in the blood sample to
14 that measured after fully activating the enzyme in the sample) have been reported
15 (Gultepe et al.. 2009). Inhibition of erythrocyte ALAD by Pb results in a rise in the
16 plasma concentration of the ALAD substrate 5-ALA. The 5-ALA biomarker can be
17 measured in serum and has been used as a surrogate for Pb measurements in studies in
18 which whole blood samples or adequately prepared plasma or serum samples were not
19 available for Pb measurements (Opleretal., 2008; Opler et al.. 2004).
4.3.5 Relationship between Pb in Blood and Pb in Bone
20 The kinetics of elimination of Pb from the body reflects the existence of multiple pools of
21 Pb in the body that have different elimination kinetics. The dominant washout phase of
22 Pb from the blood, exhibited shortly after a change in exposure occurs, has a half-life of
23 -20-30 days (Leggett. 1993; Rabinowitz et al.. 1976). Studies of a limited number of
24 adults (four individuals with hip or knee replacement, a married couple, and 10 female
25 Australian immigrants) in which the Pb exposure was from historical environmental
26 sources have found that bone Pb stores can contribute 40-70% to blood Pb (Smith et al..
27 1996; Gulsonetal. 1995a: Manton. 1985). Bone Pb burdens in adults are slowly lost by
28 diffusion (heteroionic exchange) as well as by resorption (O'Flaherty. 1995). Half-times
29 for the release of Pb in bone are dependent on age and intensity of exposure. Bone
30 compartments are much more labile in infants and children than in adults as reflected by
31 half-times for movement of Pb from bone into the plasma (e.g., cortical ti/2 = 0.23 years
November 2012 4-65 Draft - Do Not Cite or Quote
-------
1 at birth, 1.2 years at 5 years of age, 3.7 years at 15 years of age, and 23 years in adults;
2 trabecular ti/2 = 0.23 years at birth, 1.0 years at 5 years of age, 2.0 years at 15 years of
3 age, and 3.9 years in adults) (Leggett 1993). Slow transfer rates for the movement of Pb
4 from nonexchangeable bone pools to the plasma are the dominant transfer process
5 determining long-term accumulation and elimination of bone Pb burden.
6 When blood Pb concentrations are monitored in individuals over periods of years
7 following a cessation or decrease in exposure, the decrease in blood Pb concentration
8 exhibits complex kinetics that can be disaggregated into components having faster and
9 slower rates. The slower rates of clearance of Pb from the blood over months and years
10 following the cessation or reduction in exposures is thought to primarily reflect
11 elimination of Pb stores in bone. Nilsson et al. (1991) reported a tri-exponential decay in
12 the blood Pb concentrations of 14 individuals having a median occupational exposure
13 period of 26 years. Thirteen of these 14 individuals had been temporarily removed from
14 work because of excessive exposures (blood levels > 70 ug/dL or high urinary
15 5-aminolevulinic acid levels). Representing 22% of blood Pb, the fast compartment had a
16 clearance half time of 34 days. The intermediate compartment, 27% of blood Pb, had a
17 clearance half time of 1.12 year. And, the slow compartment, 50% of blood Pb, had a
18 clearance half time of 13 years. The authors attributed the fast, intermediate, and slow
19 compartment clearance to elimination of Pb from blood and some soft tissues, from
20 trabecular bone, and cortical bone, respectively. Rentschler et al. (2012) also observed a
21 slow terminal phase of Pb elimination from blood in five adults who had Pb poisoning
22 due to either occupational or non-occupational exposures that ranged from approximately
23 1 month to 12 years and resulted in blood Pb concentrations of 70-110 (ig/dL. In this
24 study, the blood Pb monitoring period extended from 1 to 74 days following cessation of
25 exposure to approximately 800 days following the diagnosis of poisoning; however, it
26 was not of sufficient duration to estimate the terminal half-time. When the terminal half-
27 time estimated by Nilsson et al. (1991) was used (13 years) to fit data for these Pb
28 poisoning cases to a two-component exponential decay model, the initial faster phase
29 represented approximately 80% of the blood Pb and the half-time was estimated to range
30 from 60 to 120 days. The relatively longer fast phase half-time reported by Rentschler et
31 al. (2012) compared to Nilsson et al. (1991) may reflect the relatively high blood Pb
32 concentrations in these poisoning cases that resulted in temporary anemia and subsequent
33 reestablishment of a normal erythrocyte levels. Also, the use of a two-compartment
34 model, with an assumed slow half-time of 13 years, as well as uncertainty about the
35 actual time of cessation of exposure may have prevented discerning a third, faster
36 elimination compartment in these data.
November 2012 4-66 Draft - Do Not Cite or Quote
-------
1 Children who have been removed from a relatively brief exposure to elevated
2 environmental Pb also exhibit faster slow-phase kinetics than children removed from
3 exposures that lasted several years, with half-times of 10 and 20-38 months, respectively
4 (Manton et al.. 2000). Rothenberg et al. (1998) also showed that exposures in the first 6
5 months of life could contribute to elevated blood Pb levels through at least 3 years
6 relative to children with lower early life exposures, despite similar environmental
7 exposures at later time points. In both adults and children, the longer half-times measured
8 under the latter conditions reflect the contribution of bone Pb stores to blood Pb
9 following a change in exposure.
10 The longer half-life of Pb in bone compared to blood Pb, allows a more cumulative
11 measure of long-term Pb exposure. Pb in adult bone can serve to maintain blood Pb levels
12 long after external exposure has ceased (Fleming et al.. 1997; Inskip etal. 1996; Smith et
13 al.. 1996; Kehoe. 1987; O'Flaherty et al., 1982). even for exposures that occurred during
14 childhood (McNeill et al.. 2000). The more widespread use of in vivo XRF Pb
15 measurements in bone and indirect measurements of bone processes with stable Pb
16 isotopes have enhanced the use of bone Pb as a biomarker of Pb body burden.
17 Several studies have found a stronger relationship between patella Pb and blood Pb than
18 tibia Pb and blood Pb (Park et al.. 2009c; Huetal.. 1998; Hernandez-Avilaetal.. 1996;
19 Hu et al.. 1996a). Hu et al. (1998) suggest that trabecular bone is the predominant bone
20 type providing Pb back into circulation under steady-state and pathologic conditions. The
21 stronger relationship between blood Pb and trabecular Pb compared with cortical bone is
22 probably associated with the larger surface area of trabecular bone allowing for more Pb
23 to bind via ion exchange mechanisms and more rapid turnover making it more sensitive
24 to changing patterns of exposure.
25 Relationships between Pb in blood and bone in children and adults are discussed in
26 greater detail below (Sections 4.3.5.1. and 4.3.5.2). In these discussions, simulations
27 based on a biokinetics model are shown to illustrate general patterns in the relationships
28 between bone Pb and blood Pb that can be predicted based on the current understanding
29 of Pb biokinetics in children and adults. However, these simulations reflect assumptions
30 in the model and may not accurately represent the observed blood Pb kinetics in
31 individuals or variability in blood Pb kinetics observed in specific populations. The
32 simulations include two metrics of blood Pb, the blood Pb concentration at each time
33 point in the simulation and the time-integrated blood Pb for the period preceding each
34 time point in the simulation (also referred to as the cumulative blood Pb index [CBLI]).
35 The time-integrated blood Pb metric has been used to estimate long-term average and
36 cumulative absorbed Pb doses in epidemiologic studies (e.g., Nie et al.. 20lib; Healev et
November 2012 4-67 Draft - Do Not Cite or Quote
-------
1 al.. 2008; Hu et al. 2007a: Chuang et al.. 2000; McNeill et al.. 2000; Fleming et al..
2 1997: Roelsetal. 1995: Gerhardsson et al.. 1993: Armstrong et al.. 1992).
4.3.5.1 Children
3 As mentioned in Section 4.2.2.2. bone growth in children will contribute to accumulation
4 of Pb in bone, which will comprise most of the Pb body burden. As a result, Pb in bone
5 will more closely reflect Pb body burden than blood Pb. However, changes in blood Pb
6 concentration in children (i.e., associated with changing exposures) are thought to more
7 closely parallel changes in total body burden. Figure 4-9 shows a biokinetics model
8 simulation of the temporal profile of Pb in blood and bone in a child who experiences a
9 period of constant Pb intake (from age 2-5) via ingestion (|ig Pb/day) followed by an
10 abrupt decline in intake. The figure illustrates several important general concepts about
11 the relationship between Pb in blood and bone. While blood Pb approaches a quasi-steady
12 state after a period of a few months with a constant rate of Pb intake (as demonstrated by
13 the vertical dashed line), Pb continues to accumulate in bone with continued Pb intake
14 after the quasi-steady state is achieved in blood. The model also predicts that the rate of
15 release of Pb from bone after a reduction in exposure is faster than in adults. This
16 difference has been attributed to accelerated growth-related bone mineral turnover in
17 children, which is the primary mechanism for release of Pb that has been incorporated
18 into the bone mineral matrix.
19 Empirical evidence in support of this conclusion comes from longitudinal studies in
20 which relatively high correlations were found between concurrent (r = 0.75) or average
21 lifetime (obtained at 6-month intervals from birth to age 10 or 12) blood Pb
22 concentrations (r = 0.85) and tibia bone Pb concentrations (measured by XRF) in a
23 sample of children in which the group mean concurrent blood Pb concentration exceeded
24 20 (ig/dL; the correlations was much weaker (r <0.15) among the group of children with
25 a mean concurrent blood Pb concentration <10 (ig/dL (Wasserman et al.. 2003).
26 Time-integrated blood Pb metrics display rates of change in response to the exposure
27 event that more closely approximate the slower kinetics of bone Pb and body burden,
28 than the kinetics of blood Pb concentration, with notable differences (Figure 4-9). The
29 time-integrated blood PB concentration is a cumulative function and increases throughout
30 childhood; however, the slope of the increase is higher during the exposure event than
31 prior to or following the event. Following cessation of the enhanced exposure period, the
32 time-integrated blood Pb and body burden diverge. This result is expected, as the time-
33 integrated blood Pb curve is a cumulative function which cannot decrease over time and
34 bone Pb levels will decrease with reduction in exposure.
November 2012 4-68 Draft - Do Not Cite or Quote
-------
1 The time-integrated blood Pb concentration will be a better reflection of the total amount
2 of Pb that has been absorbed, than the body burden at any given time. The time-
3 integrated blood Pb concentration will also reflect cumulative Pb absorption, and
4 cumulative exposure if the absorption fraction is constant. This is illustrated in the
5 hypothetical simulations of an exposure event experienced by a child (Figure 4-10). This
6 pattern is similar for adults.
November 2012 4-69 Draft - Do Not Cite or Quote
-------
10
10
50
246
Age (year)
10
Note: Blood Pb concentration is thought to parallel body burden more closely in children than in adults, due to more rapid turnover of
bone and bone-Pb stores in children (upper panel). Baseline Pb intake is 3.2 ug/day from birth until age 2, followed by a period of
increased intake (38.2 ug/day) from age 2 until age 5, with a return to baseline intake of 3.2 ug/day at age 5. The time-integrated
blood Pb concentration increases overtime (lower panel). Simulation based on ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-9 Simulation of relationship between blood Pb concentration and
body burden in children, with an elevated constant Pb intake from
age 2 to 5 years.
November 2012
4-70
Draft - Do Not Cite or Quote
-------
(0
o>
_i
T3
O
CD
•o
50
40 -
20 -
10 -
•Blood
•Absorption
246
Age (year)
8
20
O
15 I
Q)
S-.
(D
>
CT
(/>
O
•3
a-.
o
3
-- 10
-- 5 --
10
(Q
Note: The simulations include a 3-year period of elevated constant Pb intake during ages 2-5 years. Baseline Pb intake is
3.2 ug/day from birth until age 2, followed by a period of increased intake (38.2 ug/day) from age 2 until age 5, with a return to
baseline intake of 3.2 ug/day at age 5. The time-integrated blood Pb concentration closely parallels cumulative Pb absorption.
Simulation based on ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-10 Simulation of relationship between time-integrated blood Pb
concentration and cumulative Pb absorption in children.
4.3.5.2
Adults
i
2
o
J
4
5
6
7
8
9
10
11
12
In adults, where a relatively large fraction of the body burden residing in bone has a
slower turnover compared to blood, a constant Pb uptake (or constant intake and
fractional absorption) gives rise to a quasi-steady state blood Pb concentration, while the
body burden continues to increase over a much longer period, largely as a consequence of
continued accumulation of Pb in bone. This pattern is illustrated in Figure 4-11 that
depicts a hypothetical simulation of an exposure consisting of a 20-year period of daily
ingestion of Pb in an adult. The exposure shown in the simulations gives rise to a
relatively rapid increase in blood Pb concentration from a baseline of approximately
2 (ig/dL, to a new quasi-steady state of approximately 9 (ig/dL, achieved in -75-100 days
(i.e., approximately 3-4 times the blood elimination half-life). In contrast, the body
burden exhibits a steady increase across the full exposure period of 70 yr. Following
cessation of the enhanced exposure period, blood Pb concentration declines relatively
November 2012
4-71
Draft - Do Not Cite or Quote
-------
1 rapidly compared to the slower decline in body burden. Careful examination of the
2 simulation shown in Figure 4-11 reveals that the accumulation and elimination phases of
3 blood Pb kinetics are not symmetrical; elimination is slower than accumulation as a result
4 of the gradual release of bone Pb stores to blood. This response, known as the prolonged
5 terminal elimination phase of Pb from blood, has been observed in retired Pb workers and
6 in workers who continued to work after improved industrial hygiene standards reduced
7 their exposures. In the adult simulation shown in Figure 4-11, following cessation of the
8 enhanced exposure period at age 50, the blood Pb concentration is reduced by half in
9 approximately 75 days. Following this relatively short elimination period, the half-time
10 of the subsequent 4-year period is approximately 14 years; however, the half-time
11 increases to approximately 50 years during the period 5-20 years after the reduction in
12 exposure.
13 These model predictions are consistent with the slow elimination of Pb from blood and
14 elimination half-times of several decades for bone Pb (e.g., 16-98 years) that have been
15 estimated from observations made on Pb workers (Wilker et al., 2011; Fleming et al.,
16 1997; Gerhardsson et al.. 1995). Based on this hypothetical simulation, a blood Pb
17 concentration measured 1 year following cessation of a period of increased Pb uptake
18 would be elevated by only a relatively small amount over the baseline measured prior to
19 the exposure (3 (ig/dL versus the 2 (ig/dL), whereas, the body burden would remain
20 elevated. These simulations in Figure 4-11 illustrate how a single blood Pb concentration
21 measurement, or a series of measurements taken over a short-time span, could be a
22 relatively poor index of Pb body burden. The simulation shown in Figure 4-11 represents
23 an exposure that resulted in a quasi-steady state blood Pb concentration of approximately
24 10 (ig/dL. Exposures that achieve higher blood Pb concentrations, more indicative of
25 poisoning or historic occupational exposures will result in a more prolonged elevation of
26 blood Pb concentration following cessation of the enhanced exposure period. Figure 4-12
27 shows a model simulation of an adult exposed to Pb that results in a quasi-steady state
28 blood Pb concentration of approximately 90 (ig/dL. In this case, the blood Pb
29 concentration remains substantially elevated 1 year following the exposure event
30 (42 (ig/dL versus 2 (ig/dL) and 20 years following the exposure event (11 (ig/dL).
31 One important potential implication of the profoundly different kinetics of Pb in blood
32 and bone is that, for a constant Pb exposure, Pb in bone will increase with increasing
33 duration of exposure and, therefore, with age. In contrast, blood Pb concentration will
34 achieve a quasi-steady state. As a result, the relationship between blood Pb and bone Pb
35 will diverge with increasing exposure duration and age. This divergence can impart
36 different degrees of age-confounding when either blood Pb or bone Pb is used as an
37 internal dose metric in dose-response models. In a review of epidemiologic studies that
38 evaluated the associations between blood Pb, bone Pb and cognitive function, the
November 2012 4-72 Draft - Do Not Cite or Quote
-------
1 association was stronger for bone Pb than blood Pb (particularly for longitudinal studies)
2 for older individuals with environmental Pb exposures and low blood Pb levels (Shih et
3 al., 2007). In contrast, occupational workers with high current Pb exposures had the
4 strongest associations for blood Pb levels with cognitive function, thus providing
5 evidence for this divergence (Shih et al., 2007).
6 The aforementioned expectation for an increase in bone Pb and body burden with age
7 applies to scenarios of constant exposure but not necessarily to real world populations in
8 which individual and population exposures have changed overtime. Longitudinal studies
9 of blood and bone Pb trends have not always found strong dependence on age (Nie et al.
10 2009; Kimetal. 1997). Kim et al. (1997) found that bone Pb levels increased with
11 increasing age in elderly adults (age 52-83) years), only when the data were analyzed
12 cross-sectionally. When analyzed longitudinally, the trend for individual patella Pb was a
13 23% decrease over a 3-year period (approximate ti/2 of 8 years), whereas tibia Pb levels
14 did not change with over the same period. Therefore, although older individuals tended to
15 have higher bone Pb levels, the 3-year temporal trend for individuals was a loss of Pb
16 from the more labile Pb stores in trabecular bone. Nie et al. (2011 a) observed that
17 longitudinal observations of blood and bone Pb in elderly adults did not show a
18 significant age effect on the association between blood Pb and bone Pb (patella and tibia),
19 when the sample population (n=776) was stratified into age tertiles (mean age 62, 69 or
20 77 years). Nie et al. (2009) did find that regressed function bone Pb and appeared to level
21 off at bone Pb levels >20 (ig/g bone mineral.
November 2012 4-73 Draft - Do Not Cite or Quote
-------
10
O)
~ "H
CL
i "^
2 -
20
CD
« 1
O
•^
CD
O
10 <
CD
c.
a
(D
3
-5 •=•
CD
25 30 35 40 45 50 55 60 65 70
Age (year)
350
.0
Q.
300 -
•o 250 -
^ > 200 -
||15°-
•T 100 -
E
'£ 50 H
- 15
C>-i 10
- 5
20
03
o
D
(D
O
CD
O
<
03
c.
a
(D
3
CD
25 30 35 40 45 50 55 60 65 70
Age (year)
Note: A constant baseline intake of 20 |jg/day from age 0-30 results in a quasi-steady state blood Pb concentration and body
burden. An increase in Pb intake to 120 ug/day from age 30 to 50 gives rise to a relatively rapid increase in blood Pb, to a new
quasi-steady state, and a slower increase in body burden (upper panel). At age 50, intake returns to the baseline of 20 ug/day.
Following the long period of elevated Pb intake, there is a rapid decline in blood Pb from 9 to 3 ug/dL over the first year and a more
gradual decline in blood Pb to less than 2 ug/dL by age 60. The time-integrated blood Pb concentration increases over the lifetime,
with a greater rate of increase during periods of higher Pb uptake (lower panel). Simulation based on ICRP Pb biokinetics model
(Leggett. 1993).
Figure 4-11 Simulation of relationship between blood Pb concentration, bone
Pb and body burden in adults with relatively low Pb intake.
November 2012
4-74
Draft - Do Not Cite or Quote
-------
600
25 30 35 40 45 50 55 60 65 70
Age (year)
2500
600
25 30 35 40 45 50 55 60 65 70
Age (year)
Note: A constant baseline intake of 20 |jg/day from age 0-30 results in a quasi-steady state blood Pb concentration and body
burden. A increase in Pb intake to 6020 ug/day from age 30 to 50 gives rise to a relatively rapid increase in blood Pb, to a new
quasi-steady state, and a slower increase in body burden (upper panel). At age 50, intake returns to the baseline of 20 ug/day.
Following the long period of high Pb intake, there is a rapid decline in blood Pb from 90 to 40 ug/dL over the first a year followed by
a more gradual decline in blood Pb to 20 ug/dL by age 60 and 10 ug/dL at age 70. The time-integrated blood Pb concentration
increases over the lifetime, with a greater rate of increase during periods of higher Pb uptake (lower panel). Simulation based on
ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-12 Simulation of relationship between blood Pb concentration, bone
Pb and body burden in adults with relatively high Pb intake.
November 2012
4-75
Draft - Do Not Cite or Quote
-------
1 Tibia bone Pb is correlated with time-integrated blood Pb concentration (i.e., CBLI).
2 McNeill et al. (2000) compared tibia Pb levels and cumulative blood Pb indices in a
3 population of 19- to 29-year-olds who had been highly exposed to Pb in childhood from
4 the Bunker Hill, Idaho smelter. They concluded that Pb from exposure in early childhood
5 had persisted in the bone matrix until adulthood. The bone Pb/CBLI slopes from various
6 studies range from 0.022 to 0.067 ug/g bone mineral per ug-year/dL (Healev et al.. 2008;
7 Hu et al.. 2007a). Because the CBLI is a cumulative function which cannot decrease over
8 time, CBLI and bone Pb would be expected to diverge following cessation of exposure,
9 as bone Pb levels decrease. This divergence was observed as a lower bone Pb/CBLI slope
10 in retired Pb workers compared to active workers and in worker populations whose
11 exposures declined over time as a result of improved industrial hygiene (Fleming et al..
12 1997; Gerhardsson et al.. 1993).
13 Although differences in kinetics of blood and bone Pb degrade the predictive value of
14 blood Pb as a metric of Pb body burden, within a population that has similar exposure
15 histories and age demographics, blood and bone Pb may show relatively strong
16 associations. A recent analysis of a subset of data from the Normative Aging Study (an
17 all male cohort) showed that cross-sectional measurements of blood Pb concentration
18 accounted for approximately 9% (tibia) to 13% (patella) of the variability in bone Pb
19 levels. Inclusion of age in the regression model accounted for an additional 7-10% of the
20 variability in bone Pb (Park et al.. 2009c).
Mobilization of Pb from Bone in Adulthood
21 In addition to changes in exposure (e.g., declines in exposure discussed in prior sections),
22 there are physiological processes during different life circumstances that can increase the
23 contribution of bone Pb to blood Pb. These life circumstances include times of
24 physiological stress associated with enhanced bone remodeling such as during pregnancy
25 and lactation (Hertz-Picciotto et al.. 2000; Silbergeld. 1991; Manton. 1985). menopause
26 or in the elderly (Silbergeld et al.. 1988). extended bed rest (Markowitz and Weinberger.
27 1990). hyperparathyroidism (Kessler et al.. 1999) and severe weight loss (Riedt et al..
28 2009).
29 During pregnancy, bone Pb can serve as a Pb source as maternal bone is resorbed for the
30 production of the fetal skeleton (Gulson et al.. 2003; Gulson et al.. 1999; Franklin et al..
31 1997; Gulson etal.. 1997). Increased blood Pb during pregnancy has been demonstrated
32 in numerous studies and these changes have been characterized as a "U-shaped" pattern
33 of lower blood Pb concentrations during the second trimester compared to the first and
34 third trimesters (Lamadrid-Figueroa et al.. 2006; Gulson et al.. 2004a: Hertz-Picciotto et
35 al.. 2000; Gulson etal.. 1997; Lagerkvist et al.. 1996; Schuhmacher et al.. 1996;
November 2012 4-76 Draft - Do Not Cite or Quote
-------
1 Rothenberg et al., 1994a). The U-shaped relationship reflects the relatively higher impact
2 of hemodilution in the second trimester versus the rate of bone Pb resorption
3 accompanying Ca2+ releases for establishing the fetal skeleton. In the third trimester, fetal
4 skeletal growth on calcium demand is greater, and Pb released from maternal skeleton
5 offsets hemodilution. Gulson et al. Q998b) reported that, during pregnancy, blood Pb
6 concentrations in the first immigrant Australian cohort (n = 15) increased by an average
7 of about 20% compared to non-pregnant migrant controls (n = 7). Skeletal contribution to
8 blood Pb, based on the isotopic composition for the immigrant subjects, increased in an
9 approximately linear manner during pregnancy. The mean increases for each individual
10 during pregnancy varied from 26% to 99%. Interestingly, the percent change in blood Pb
11 concentration was significantly greater during the post-pregnancy period than during the
12 second and third trimesters. This is consistent with Hansen et al. (20 lib) that
13 demonstrated the greatest blood Pb levels at 6 weeks postpartum compared to the second
14 trimester in 211 Norwegian women. Increased calcium demands of lactation (relative to
15 pregnancy) may contribute to the greater change in blood Pb observed post pregnancy
16 compared to the second and third trimesters. The contribution of skeletal Pb to blood Pb
17 during the post-pregnancy period remained essentially constant at the increased level of
18 Pb mobilization.
19 Gulson et al. (2004a) observed that calcium supplementation was found to delay
20 increased mobilization of Pb from bone during pregnancy and halved the flux of Pb
21 release from bone during late pregnancy and postpartum. In another study, women whose
22 daily Ca2+ intake was 850 mg per day showed lower amounts of bone resorption during
23 late pregnancy and postpartum than those whose intake was 560 mg calcium per day
24 (Manton et al.. 2003). Similarly, calcium supplementation (1,200 mg/day) in pregnant
25 Mexican women resulted in an 11% reduction in blood Pb level compared to placebo and
26 a 24% average reduction for the most compliant women (Ettinger et al.. 2009). When
27 considering baseline blood Pb levels in women who were more compliant in taking
28 calcium supplementation, the reductions were similar for those <5 (ig/dL and those
29 > 5 (ig/dL (14% and 17%, respectively). This result is in contrast to a study of women
30 who had blood Pb concentrations <5 (ig/dL, where calcium supplementation had no
31 effect on blood Pb concentrations (Gulson et al., 2006b). These investigators attributed
32 their results to changes in bone resorption with decoupling of trabecular and cortical bone
33 sites.
34 Miranda et al. (2010) studied blood Pb level among pregnant women aged 18-44 years
35 old. The older age segments in the study presumably had greater historic Pb exposures
36 and associated stored Pb than the younger age segments. Compared with the blood Pb
37 levels of a reference group in the 25-29 years old age category, pregnant women
38 > 30 years old had significant odds of having higher blood Pb levels (aged 30-34:
November 2012 4-77 Draft - Do Not Cite or Quote
-------
1 OR = 2.39, p <0.001; aged 35-39: OR = 2.98, p <0.001; aged 40-44: OR = 7.69,
2 p <0.001). Similarly, younger women had less chance of having higher blood Pb levels
3 compared with the reference group (aged 18-19: OR = 0.60, p = 0.179; aged 20-24:
4 OR = 0.54, p = 0.015). These findings indicate that maternal blood Pb levels are more
5 likely the result of Pb mobilization of bone stores from historic exposures as opposed to
6 contemporaneous exposures.
7 Blood Pb levels increase during lactation due to alterations in the endogenous bone Pb
8 release rate. After adjusting for patella Pb concentration, an increase in blood Pb levels of
9 12.7% (95% CI: 6.2, 19.6) was observed for women who practiced partial lactation and
10 an increase of 18.6% (95% CI: 7.1, 31.4) for women who practiced exclusive lactation
11 compared to those who stopped lactation (Tellez-Rojo et al., 2002). In another Mexico
12 City study, Ettinger et al. (2006; 2004b) concluded that an interquartile increase in patella
13 Pb was associated with a 14% increase in breast milk Pb, whereas for tibia Pb the
14 increase was ~5%. Breast milk:maternal blood Pb concentration ratios are generally <0.1,
15 although values of 0.9 have been reported (Kovashiki et al., 2010; Ettinger et al., 2006;
16 Gulsonet al.. 1998a). Dietary intake of polyunsaturated fatty acids (PUFA) has been
17 shown to weaken the association between Pb levels in patella and breast milk, perhaps
18 indicating decreased transfer of Pb from bone to breast milk with PUFA consumption
19 (Arora et al., 2008). Breast milk as a source of infant Pb exposure was also discussed in
20 Section 4.1.3.3 on dietary Pb exposure.
21 The Pb content in some bones (i.e., mid femur and pelvic bone) plateau at middle age and
22 then decreases at older ages (Drasch etal.. 1987). This decrease is most pronounced in
23 females and may be due to osteoporosis and release of Pb from resorbed bone to blood
24 (Gulson et al., 2002). Two studies indicate that the endogenous release rate in
25 postmenopausal women ranges from 0.13-0.14 (ig/dL in blood per (ig/g bone and is
26 nearly double the rate found in premenopausal women (0.07-0.08 (ig/dL per (ig/g bone)
27 (Popovic et al.. 2005; Garrido Latorre et al.. 2003). An analysis of data on blood Pb
28 concentrations and markers of bone formation (serum alkaline phosphatase) and
29 resorption (urinary cross-linked N-telopeptides, NTx) in a sample of U.S. found that
30 blood Pb concentrations were higher in women (pre- or post-menopausal) who exhibited
31 the highest bone formation or resorption activities (Jackson et al.. 2010). Calcium or
32 vitamin D supplementation decreased the blood Pb concentrations in the highest bone
33 formation and resorption tertiles of the population of post-menopausal women.
34 Significant associations between increasing NTx and increasing blood Pb levels
35 (i.e., increased intercept of regression model relating the change in blood Pb per change
36 in bone Pb) has also been observed in elderly males (Nie et al., 2009).
November 2012 4-78 Draft - Do Not Cite or Quote
-------
1 Studies of the effect of hormone replacement therapy on bone Pb mobilization have
2 yielded conflicting results (Popovic et al.. 2005; Berkowitz et al.. 2004; Garrido Latorre
3 et al.. 2003; Korrick et al.. 2002; Webber etal., 1995). In women with severe weight loss
4 (28% of BMI in 6 months) sufficient to increase bone turnover, increased blood Pb levels
5 of approximately 2.1 (ig/dL (250%) were reported, and these blood Pb increases were
6 associated with biomarkers of increased bone turnover (e.g., urinary pyridinoline cross-
7 links) (Riedt et al.. 2009).
4.3.6 Relationship between Pb in Blood and Pb in Soft Tissues
8 Figure 4-13 shows simulations of blood and soft tissues Pb (including brain) for the same
9 exposure scenarios previously displayed. Pb uptake and elimination in soft tissues is
10 much faster than bone. As a result, following cessation of a period of elevated exposure,
11 Pb in soft tissues is more quickly returned to blood. The terminal elimination phase from
12 soft tissue mimics that of blood, and it is similarly influenced by the contribution of bone
13 Pb returned to blood and being redistributed to soft tissue.
14 Information on Pb levels in human brain is limited to autopsy data. These data indicate
15 brain/blood Pb ratios of approximately 0.5 in infancy which remain relatively constant
16 over the lifetime (range 0.3 to 1.1) (Barry. 1981. 1975). The simulation of brain Pb
17 shown in Figure 4-14 reflects general concepts derived from observations made in
18 non-human primates, dogs and rodents. These observations suggest that peak Pb levels in
19 the brain are reached 6 months following a bolus exposure and within two months
20 approximately 80% of steady state brain Pb levels are reached (Leggett 1993). There is a
21 relatively slow elimination of Pb from brain (ti/2 ~ 2 years) compared to other soft tissues
22 (Leggett 1993). This slow elimination rate is reflected in the slower elimination phase
23 kinetics is shown in Figure 4-14. Although in this model, brain Pb to blood Pb transfer
24 half-times are assumed to be the same in children and adults, uptake kinetics are assumed
25 to be faster during infancy and childhood, which achieves a higher fraction of the soft
26 tissue burden in brain, consistent with higher brain/body mass relationships. The uptake
27 half times predicted by Leggett (1993) vary from 0.9 to 3.7 days, depending on age.
28 Brain Pb kinetics represented in the simulations are simple outcomes of modeling
29 assumptions and cannot currently be verified with available observations in humans.
November 2012 4-79 Draft - Do Not Cite or Quote
-------
10
8 -
O)
^ 6 H
.a
Q.
•o
m
2 -
, Blood
"" \
•Soft Tissue
1.0
-- 0.8
0.0
4 6
Age (year)
10
10
0.0
25 30 35 40 45 50 55 60 65 70
Age (year)
Note: For the child simulation (upper panel), baseline Pb intake is 3.2 ug/day from birth until age 2, followed by a period of increased
intake to 38.2 ug/day from age 2 until 5, with a return to baseline intake at age 5. For the adult simulation (lower panel), baseline
intake is 20 ug/day from age 0-30, followed by a 20-year period of increased intake to 120 ug/day from age 30 to 50, with a return to
baseline intake at age 50. Simulation based on ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-13 Simulation of blood and soft tissue (including brain) Pb in
children and adults who experience a period of increased Pb
intake.
November 2012
4-80
Draft - Do Not Cite or Quote
-------
10
„ 8
en
6
CL
•D
8 4H
CO
2 -
m
Blood
— Brain
4 6
Age (year)
30
25
o
20 sr
03
15 I
T3
tr
10 g
5
10
25 30 35 40 45 50 55 60 65 70
Age (year)
Note: For the child simulation (upper panel), baseline Pb intake is 3.2 ug/day from birth until age 2, followed by a period of increased
intake to 38.2 ug/day from age 2 until 5, with a return to baseline intake at age 5. For the adult simulation (lower panel), baseline
intake is 20 ug/day from age 0-30, followed by a 20-year period of increased intake to 120 ug/day from age 30 to 50, with a return to
baseline intake at age 50. Simulation based on ICRP Pb biokinetics model (Leggett. 1993).
Figure 4-14 Simulation of blood and brain Pb in children and adults who
experience a period of increased Pb intake.
November 2012
4-81
Draft - Do Not Cite or Quote
-------
1 Urinary filtering and excretion of Pb is associated with plasma Pb concentrations. Given
2 the curvilinear relationship between blood Pb and plasma Pb, a secondary expectation is
3 for a curvilinear relationship between blood Pb and urinary Pb excretion that may
4 become evident only at relatively high blood Pb concentrations (e.g., >25 (ig/dL). Figure
5 4-15 shows these relationships predicted from the model. In this case, the exposure
6 scenario shown is for an adult (age 40 years) at a quasi-steady state blood Pb
7 concentration; the same relationships hold for children. At lower blood Pb concentrations
8 (<25 (ig/dL), urinary Pb excretion is predicted to closely parallel plasma Pb concentration
9 for any given blood Pb level (Figure 4-15. top panel). It follows from this that, similar to
10 blood Pb, urinary Pb will respond much more rapidly to an abrupt change in Pb exposure
11 than will bone Pb. One important implication of this relationship is that, as described
12 previously for blood Pb, the relationships between urinary Pb and bone Pb will diverge
13 with increasing exposure duration and age, even if exposure remains constant.
14 Furthermore, following an abrupt cessation of exposure, urine Pb will quickly decrease
15 while bone Pb will remain elevated (Figure 4-15. lower panel).
November 2012 4-82 Draft - Do Not Cite or Quote
-------
80
20 30 40
Blood Pb (ng/dL)
0.00
50
IZ. •
10 •
"> j"
WTJ
5 o> 8
0 3.
3.—
""""^
£ °- 6 •
a>^
•El
Dm 4 .
2
n
Blood
^x"**1!'.1- • • • • J •'
f**'*'
* •
t
1 •
*
/
r
*
r
*
^— Urine
. Bone
•
•
•
• .
V
^-*— „
• ^u
- 15
CD
0
- 10 m
(Q
*™*
- 5
n
25 30 35 40 45 50 55 60 65 70
Age (year)
Note: Upper panel, model simulations are for a 40-year old having a constant intake from birth of between 1 and 1,000 ug/day. For
the lower panel, baseline intake is 20 ug/day from age 0-30, followed by a 20-year period of increased intake to 120 ug/day from
age 30 to 50, with a return to baseline intake at age 50. Simulation based on ICRP Pb biokinetics model (Leggett, 1993).
Figure 4-15 Relationship between Pb in urine, plasma, blood and bone.
November 2012
4-83
Draft - Do Not Cite or Quote
-------
4.4 Studies of Pb Biomarker Levels
4.4.1 Pb in Blood
1 Overall, trends in blood Pb levels have been decreasing among U.S. residents over the
2 past 35 years. Blood Pb concentrations in the U.S. general population have been
3 monitored in the NHANES. Analyses of these data show a progressive downward trend
4 in blood Pb concentrations during the period 1976-2010, with the most dramatic declines
5 coincident with the phase out of leaded gasoline and reductions in point source Pb
6 emissions described in Section 3.2 (Pirkle et al.. 1998; Brody et al.. 1994; Pirkle et al..
7 1994; Schwartz and Pitcher. 1989). The temporal trend for the period 1988-2010 is
8 shown in Figure 4-16. Summary statistics from the most recent publicly available data
9 (1999-2010) are presented in Table 4-8 (CDC. 2011 a). The geometric mean Pb
10 concentration among children 1-5 years of age, based on the sample collected during the
11 period 2009-2010, was 1.17 (ig/dL (95% CI: 1.08, 1.26), which was decreased from
12 2007-2008(1.51 jig/dL, 95% CI: 1.37, 1.66). Figure 4-17 uses NHANES data to illustrate
13 temporal trends in the distribution of blood Pb levels among U.S. children aged 1-5 years.
14 For 2005-2010, the 95th percentile of blood Pb levels for children aged 1-5 years was less
15 than 5 ng/dL. The geometric mean blood Pb concentration among adults > 20 years of
16 age was 1.23 (ig/dL (95% CI: 1.19, 1.28) for the sample collected during the period
17 2009-2010 (CDC. 2011 a). Based on these same data, the geometric mean for all males
18 (aged > 1 year) was 1.31 (ig/dL (95% CI: 1.25, 1.36), and for females (aged > 1 year) was
19 0.97 (ig/dL (95% CI: 0.93, 1.01).
20 There has been a steep decline in mean blood Pb levels from 1975 through 2010 among
21 all birth cohorts from 1975 to 2010 (Figure 4-18). For all cohorts, blood Pb generally
22 decreases with age during childhood until adolescence; following adolescence (in the
23 early 20s), blood Pb generally levels off or even increases with age. It is possible that
24 bone growth in young people and occupational exposure for adults influences the shape
25 of these curves. For the 1960 to 1970 birth cohort, the mean blood Pb is the highest of the
26 cohorts in the 1970s, but beginning in 1993 the mean blood Pb is one of the lowest of the
27 cohorts. This interaction between time and cohort may be due to the faster release of Pb
28 from bone in younger people (Rabinowitz. 1991). This interaction is also apparent for
29 some of the other more recently born cohorts. In comparison, the slopes of blood Pb over
30 time are nearly parallel among the cohorts born before 1960. This suggests that the time-
31 cohort-interaction diminishes among older people. Also, the leveling of the blood Pb in
32 the 2000s could be due to aging of the birth cohort and consequent slowing of their Pb
33 release from bone.
November 2012 4-84 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
When race/ethnicity groups were compared for years 1999-2004, geometric means (GM)
of blood Pb levels in children were highest in the ethnicity category non-Hispanic black
(GM 2.8, 95% CI: 2.5, 3.0) compared to the categories Mexican-American (GM 1.9, 95%
CI: 1.7, 2.0) and non-Hispanic white (GM 1.7, 95% CI: 1.6, 1.8) (Jones et al.. 2009a).
Figure 4-19 demonstrates the change in percent of children (aged 1-5 years) with various
blood Pb levels by race/ethnicity between the survey during 1988-1991 and that during
1999-2004. When these data for children aged 1-5 years were aggregated for all survey
years from 1988 to 2004, residence in older housing, poverty, age, and being
non-Hispanic black were significant predictors of higher Pb levels (Jones et al., 2009a).
O)
T3
TO
0)
4 -
3 -
1 -
Children 1-5 yrs
Children 6-11 yrs
Teens 12-19 yrs
Adults>20 yrs
88-91 91-94 99-00 01-02 03-04 05-06 07-08 09-10
Survey Period
Note: Shown are geometric means and 95% CIs based on data from NHANES III Phase 1 (Brodvet al.. 1994: Pirkleetal.. 1994):
NHANES III Phase 2 (Pirkleetal.. 1998): and NHANES IV (CDC. 2011 a). Data for adults during the period 1988-1994 are for ages
20-49 years, and > 20 years for the period 1999-2008.
Figure 4-16 Temporal trend in blood Pb concentration.
November 2012
4-85
Draft - Do Not Cite or Quote
-------
Table 4-8 Blood Pb concentrations in the U.S. population.
Survey Stratum Period Geometric Mean (ug/dL) 95% Confidence Interval
All 1999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
1-5yr 1999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
6-1 1 yr 1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
12-1 9 yr 1999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
> 20 yr 1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
Males 1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
1.66
1.45
1.43
1.29
1.27
1.12
2.23
1.70
1.77
1.46
1.51
1.17
1.51
1.25
1.25
1.02
0.99
0.84
1.10
0.94
0.95
0.80
0.80
0.68
1.75
1.56
1.52
1.41
1.38
1.23
2.01
1.78
1.69
1.52
1.47
1.31
1.60, 1.72
1.39, 1.51
1.36, 1.50
1.23, 1.36
1.21, 1.34
1.08, 1.16
1 .96, 2.53
1.55, 1.87
1.60, 1.95
1.36, 1.57
1.37, 1.66
1.08, 1.26
1.36, 1.66
1.14, 1.36
1.12, 1.39
0.95, 1.01
0.91, 1.07
0.79, 0.89
1.04, 1.17
0.90, 0.99
0.88, 1.02
0.75, 0.85
0.74, 0.86
0.64, 0.73
1.68, 1.81
1.49, 1.62
1.45, 1.60
1 .34, 1 .48
1.31, 1.46
1.19, 1.28
1 .93, 2.09
1.71, 1.86
1.62, 1.75
1.42, 1.62
1.39, 1.56
1.25, 1.36
Number of Subjects
7,970
8,945
8,373
8,407
8,266
8,793
723
898
911
968
817
836
905
1,044
856
934
1,011
1,009
2,135
2,231
2,081
1,996
1,074
1,183
4,207
4,772
4,525
4,509
5,364
5.765
3,913
4,339
4,132
4,092
4,147
4,366
November 2012
4-86
Draft - Do Not Cite or Quote
-------
Survey Stratum Period Geometric Mean (ug/dL) 95% Confidence Interval Number of Subjects
Females 1999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
Mexican - Americans 1999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
Non-Hispanic blacks 1999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
Non-Hispanic whites 1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
2009-201 0
1.37
1.19
1.22
1.11
1.11
0.97
1.83
1.46
1.55
1.29
1.25
1.14
1.87
1.65
1.69
1.39
1.39
1.24
1.62
1.43
1.37
1.28
1.24
1.10
1.32,
1.14,
1.14,
1.05,
1.06,
0.93,
1.75,
1.34,
1.43,
1.21,
1.15,
1.03,
1.75,
1.52,
1.52,
1.26,
1.30,
1.18,
1.55,
1.37,
1.32,
1.19,
1.16,
1.04,
1.43
1.25
1.31
1.17
1.16
1.01
1.91
1.60
1.69
1.38
1.36
1.28
2.00
1.80
1.89
1.53
1.48
1.30
1.69
1.48
1.43
1.37
1.33
1.16
4,057
4,606
4,241
4,315
4,119
4.427
2,742
2,268
2,085
2,236
1,712
1,966
1,842
2,219
2,293
2,193
1,746
1,593
2,716
3,806
3,478
3,310
3,461
3,760
Age strata correspond to the NHANES study design.
Source: Adapted from data from the NHANES (CDC. 2011 a).
November 2012
4-87
Draft - Do Not Cite or Quote
-------
10 -
Note: Top: all data. Bottom: data for subjects having blood Pb levels less than 15 ug/dL.
Source: Adapted from data from the NHANES (NCHS. 2010)
Figure 4-17 Box plots of blood Pb levels among U.S. children (1-5 years old at
baseline) from the NHANES survey, 1988-2010.
November 2012
4-8
Draft - Do Not Cite or Quote
-------
o
.2 LO
C
ro
0) CO
o
o
0)
O)
cohort birth yrs
1900 to 1930<
1930to1940<
1940 to 1950 '
1950 to 1960i
1960 to 1970<
1970 to 1975<
1975 to 1980J
1980 to 1985i
1985 to 1990
1990 to 1995
1995 to 2000
2000 to 2005
2005 to 2008 «
1970
1980
1990
exam year
2000
2010
Note: The means of logged blood Pb were weighted to represent national averages. Data were from the publicly available
NHANES II, NHANES II for 1988-1991 and 1992-1994, and the continuous NHANES in 1999-2000, 2003-2004, 2005-2006,
2007-2008. Continuous NHANES data from 2001-2002 and 2009-2010 are not included because there were only 551 blood Pb
samples in each of those data sets. The year plotted for exam year was the reported exam year for NHANES II, the middle year of
each of the phases of NHANES III, and the second year of each of the continuous NHANES.
Source: Adapted from data from the NHANES (NCHS. 2010)
Figure 4-18 Blood Pb cohort means versus year of exam.
November 2012
4-89
Draft - Do Not Cite or Quote
-------
10
1999 - 2004
0
<1 l-<2.5 2.5-<5 5-<7.5 7.5 - <10 > 10
Blood Pb Level (ug/dL)
...+.. Non-Hispanic black^^^—Mexican American —^— Non-Hispanic white
Source: Data used with permission of the American Academy of Pediatrics, Jones et al. (2009a)
Figure 4-19 Percent distribution of blood Pb levels by race/ethnicity among
U.S. children (1-5 years) from the NHANES survey, 1988-1991 (top)
and 1999-2004 (bottom).
November 2012
4-90
Draft - Do Not Cite or Quote
-------
1 In agreement with the 1986 AQCD (U.S. EPA. 1986a). several studies have shown
2 seasonal variation in blood Pb concentrations in children (e.g., Havlena et al.. 2009;
3 Gulson et al.. 2008; Kemp et al.. 2007; Laidlaw et al.. 2005; Haley and Talbot. 2004;
4 Johnson and Bretsch. 2002; Yiin et al.. 2000; Johnson et al.. 1996). with elevated
5 concentrations during the warm season and lower levels in the cold season. Seasonal
6 dynamics of blood Pb concentrations in children appear to be caused at least in part by
7 seasonal patterns in access of children to soils and soil properties (e.g., moisture content)
8 that may contribute to seasonal variation in entrainment of soil and dust Pb into breathing
9 zone air (Laidlaw et al.. 2012; Laidlaw et al.. 2005; Johnson and Bretsch. 2002). Seasonal
10 variation in blood Pb concentrations occur with strong associations with soil Pb
11 concentrations (Johnson and Bretsch. 2002). Laidlaw et al. (2012) observed that air Pb in
12 the PM2 5 fraction and PM2 5 attributed to soil were elevated in the warm season compared
13 with the cold season. Yiin et al. (2000) found that geometric mean for blood Pb, floor Pb
14 loading and concentration, and carpet dust loading were statistically significantly higher
15 in the cold season compared with the hot season. However, regression of blood Pb on
16 floor and windowsill dust with and without adjustment for the hot, warm, and cool
17 seasons showed no statistically significant effect of the seasons directly on blood Pb.
18 Meteorological factors appear to contribute to blood Pb seasonality. Laidlaw et al. (2005)
19 analyzed the temporal relationships between child blood Pb concentrations and various
20 atmospheric variables in three cities (Indianapolis, IN: 1999-2002; Syracuse, NY:
21 1994-1998; New Orleans, LA: 1998-2003). Blood Pb data was obtained from public
22 health screening programs conducted in the three cities. Blood Pb samples were
23 dominated by children <5 years of age and age distribution varied across the three cities.
24 The temporal variation in blood Pb concentrations in each city was predicted by
25 multivariate regression models that included the following significant variables: PMi0,
26 wind speed, air temperature, and soil moisture; as well as dummy variables accounting
27 for temporal displacement of the effects of each independent variable on blood Pb.
28 Laidlaw et al. (2005) reported R2 values for the regression models, but did not report the
29 actual regression coefficients. The R2 values were as follows: Indianapolis 0.87
30 (p = 0.004); Syracuse 0.61 (p = 0.0012); New Orleans 0.59 (p O.OOOOl).
31 Studies have examined the change in blood Pb with changes in potential Pb sources.
32 Gulson et al. (2004b) observed that children living near a Zn-Pb smelter in Australia had
33 blood Pb levels ranging from 10 to 42 (ig/dL, with 55-100% of Pb attributed to the
34 smelter based on isotope ratio analysis. Rubio-Andrade et al. (2011) followed a cohort of
35 6-8 year old children living within 3.5 km of a Mexican smelter at 0, 6, 12, and
36 60 months after environmental intervention including removal of 100,000 kg of
37 Pb-containing dust from roads and homes using high efficiency vacuums. Soil Pb was
38 concurrently obtained but not reported at 6, 12, or 60 months. Median blood Pb level at
39 initiation of the study was 10.1 (ig/dL for the 598 initial participants (average age: 7.2 y),
November 2012 4-91 Draft - Do Not Cite or Quote
-------
1 and median soil Pb was 3,300 mg/kg at the start of the study. After 60 months, median
2 blood Pb level was 4.4 ug/dL for the remaining 232 participants (average age: 12.2 y),
3 and median soil Pb concentration was 370 mg/kg at that time. Bonnard and McKone
4 (2009) modeled blood Pb of French children ages 21-74 months living within a village
5 containing a Pb smelter and estimated blood Pb levels of 3.2-10.9 ug/dL. Lanphear et al.
6 (1998) noted that the probability of children having blood Pb > 10 ug/dL increases both
7 with exterior soil Pb content and interior Pb dust loading. Mielke et al. (20lib) noted
8 significant increases in percentages of children younger than 7-years old with blood Pb
9 level > 10 ug/dL for those living in inner city New Orleans housing developments
10 (22.9%) compared with children living in communities located on the city outskirts
11 (9.1%). At the same time, median soil Pb was significantly higher in the inner city
12 (438 mg/kg) compared with the city outskirts (117 mg/kg).
13 For infants <1 year old, very little data are available on blood Pb levels. Simon et al.
14 (2007) followed a cohort of 13 children living near an Australian smelter from birth
V / O
15 through 36 months. In general, except for children born with low blood Pb levels of ~1 to
16 2 ug/dL, immediately after birth blood Pb levels fell for 1-2 months to approximately
17 47% of birth blood Pb level. After this initial fall, all infants' blood Pb levels rose with
18 age until approximately 12 months old for children living in a high risk area and until
19 approximately 18 months for children living in a low risk area (Simon et al.. 2007).
20 Median blood Pb level among the children was 1.9 ug/dL at 2 months and increased to
21 13.6 ug/dL at 16 months. Geometric mean hand-Pb loading of the child and the mother
22 were significant contributors to the area under the curve for infant blood Pb, with 46%
23 (infant hand loading) and 60% (mother hand loading) of the variance being explained by
24 these variables, respectively; geometric mean of the mothers' blood Pb explained 46% of
25 the variance (Simon et al.. 2007). Across all the data, there was a good correlation
26 between child blood Pb level and child hand Pb loading (R2 = 0.70). In another study
27 (Carbone et al.. 1998). blood Pb levels of 15 infants aged 6-12 months were statistically
28 significantly lower than their neonatal cord blood Pb levels (2.24 ug/dL versus
29 4.87 ug/dL). Additionally, 3 infants born with blood Pb levels of greater than 7 ug/dL
30 were followed for a week, there was a dramatic drop in the blood Pb of from an average
31 of 7.6 ug/dL on Day 1 to 2.4 ug/dL on Day 7 (Carbone et al.. 1998).
32 Pb body burden has been reported among individuals known to consume wild game
33 hunted with Pb shot. For example, fifty men from Nuuk, Greenland participated in a
34 study in which they recorded their diet and produced blood samples (Johansen et al..
35 2006). Men who regularly ate hunted sea birds had an average blood Pb concentration of
36 12.8 ug/dL, in contrast with those who did not and had an average blood Pb
37 concentration of 1.5 ug/dL. Umbilical cord blood was collected from a cohort of Inuit
38 newborns from northern Quebec, where the Inuit population consumes game killed with
November 2012 4-92 Draft - Do Not Cite or Quote
-------
1 Pb shot (Levesque et al., 2003). The geometric mean cord blood Pb level was
2 0.19 (imol/L [3.9 (ig/dL], with a range of 0.01-1.31 (imol/L [0.2-27 (ig/dL]; the Canadian
3 level of concern for cord blood Pb is 0.48 (imol/L [10 jig/dL]. The authors contrasted the
4 finding that 7% of Inuit newborns had cord blood Pb concentration > 0.48 (imol/L
5 [10 (ig/dL] in contrast with 0.16% of the Caucasian population in southern Quebec.
6 Recent studies have sought to characterize human exposure to Pb from piston-engine
7 aircraft emissions. Section 3.2.2.1 describes a study by Carr et al. (2011) in which Pb
8 concentrations, both modeled and monitored, extended beyond airport property. Miranda
9 et al. (2011) used GIS to study the association between blood Pb level and distance from
10 airports in six North Carolina Counties. They observed that the trend in blood Pb level
11 decreases monotonically with distance class from the airports, with subjects within 500
12 meters of the airports having significantly increased blood Pb levels ((3 = 0.043, 95% CI:
13 (0.006,0.080), p <0.05) compared with the general population for a given county after
14 controlling for proportion of black, Hispanic, percent receiving public assistance, and
15 household median income at the census block group level and including dummy variables
16 for season during which the children were screened for blood Pb. In this study, children
17 living within 500 meters of an airport had blood Pb levels that were, on average, 4.4%
18 higher than those at distance. Note that the authors did not include Pb emissions in their
19 model.
20 Trends in blood Pb levels have been accompanied by changes in Pb isotope ratios for
21 blood Pb. Isotopic ratios, described in Sections 3.2 and 3.3 as a tool for source
22 apportionment, have been used to associate blood Pb measurements with anthropogenic
23 sources of Pb in the environment. Changes in Pb isotopic ratios in blood samples reflect
24 the changing influence of sources of Pb following the phase-out of tetraethyl Pb
25 antiknock agents in automotive gasoline and changes in Pb usage in paints and other
26 industrial and consumer products (Gulson et al., 2008; Ranft et al.. 2008; Gulson et al..
27 2006a; Ranft et al.. 2006). Gulson et al. (2006a) illustrated how a linear increase in the
28 isotopic ratio 206Pb/204Pb occurred in concert with a decrease in blood Pb levels among
29 selected study populations in Australia during the period 1990-2000 (Figure 4-20).
30 Gulson et al. (2006a) point out that the isotopic signature of 206Pb/204Pb derived from
31 Australian mines (median -16.8) differs from that of European and Asian mines, where
32 206Pb/204Pb varies between -17.4 and -18.1. Liang et al. (2010) also examined the trends
33 in blood Pb level over the period 1990 to 2006 in Shanghai and saw a reduction
34 corresponding to the phase out of Pb in gasoline. A plot of 208Pb/206Pb to 207Pb/206Pb for
35 blood and environmental samples showed overlap between the isotopic signature for coal
36 combustion ash and that measured in blood. This result suggests a growing influence of
37 Pb from coal ash in Shanghai in the absence of Pb in automobile emissions. Oulhote et al.
38 (2011) examined Pb isotope ratios in blood Pb samples of 125 French children aged 6
November 2012 4-93 Draft - Do Not Cite or Quote
-------
1 mo-6 yr. The study found that Pb isotope ratios could be used to attribute Pb exposure to
2 one source for 32% of children and to eliminate an unlikely source of Pb exposure in
3 30% of children.
source
O Adelaide
-s, BK Adult
" females
D BK children
A Broken Hill
V Hobart
<| PbinCa
[> Port Pirie
r Sydney
' children
17.60-
.a 17.20-
Q.
1
*d
B.
1
16.80-
16.40-
D
Q
+ 1
\yf*°
I I 4
R Sq Linear = 0.53
A A
12.5-
10.0-
7.5-
ffl
£
5.0-
2.5-
0.0-
1 1 1 1 1 1
1990 1992 1994 1996 1998 2000
>
>
O
> 1
Q" * 0 ^
^'"""""'"•••^ ^
7 § "~~~~~
-------
4.4.2 Pb in Bone
1 An extensive national database (i.e., NHANES) is available for blood Pb concentrations
2 in children and adults, as described in Section 4.4.1. Bone Pb concentrations are less well
3 characterized. Table 4-9 and Table 4-10 are compilations of data from epidemiologic
4 studies that provided bone Pb concentrations by K-XRF and/or variability in
5 concentrations among individuals without reported occupational exposure and those with
6 occupational exposures, respectively. In non-occupationally exposed individuals, typical
7 group mean tibia bone Pb concentrations ranged from 10 to 30 ug/g. Patella bone Pb
8 levels are typically higher than tibia bone Pb levels in the studies considered (Table 4-9).
9 For example, in the Normative Aging Study, patella bone Pb concentrations were
10 approximately 32 ug/g, whereas tibia bone Pb concentrations were about 22 ug/g.
11 Occupationally exposed individuals generally had greater bone Pb concentrations than
12 seen in control groups (i.e., unexposed). Bone Pb data in Table 4-10 for occupationally
13 exposed individuals were also generally higher compared to non-occupationally exposed
14 individuals (Table 4-9).
November 2012 4-95 Draft - Do Not Cite or Quote
-------
Table 4-9
Reference
Bandeen-
Roche et al.
(2009)
Bellinger et
al. (|994a)
Cheng et al.
(2001 )
Coon et al.
(2006)
Epidemiologic studies that provide bone Pb
occupationally exposed populations.
Prior Pb Bone Pb
Study Methods Exposure biomarker
Cohort: Cumulative Tibia
Baltimore Memory Study cohort
Age (yrs): 50-70
N:1,140
Location: Baltimore, MD
Study Period: 2001 -2005
Cohort: Not reported Cumulative Tibia
Age (yrs): 5-8 (recruited); Patella
19-20 (follow-up)
N:79
Location: Boston, MA
Study Period: 1989-1990
Cohort: Cumulative Tibia
Normative Aging Study cohort Patella
Age (yrs): Mean ± SD:
Normotensive:
65.49 ±7.1 7
Borderline hypertension:
68.3 ± 7.79
Definite hypertension:
67.93 ± 6.79
N: 833 males
Location: Boston, MA
Study Period:
8/1/1991-12/31/1997
Cohort: Cumulative Tibia
Participants from Henry Ford Calcaneus
Health System (HFHS)
Age (yrs): > 50; Mean: 69.9
N: 121 cases; 41 4 controls
Location: Southeastern Michigan
Study Period:
1995-1999 (participants received
primary health care services)
measurements
Bone Pb Cone.
(Hg/g)
Meant SD
Tibia: 18.8 ± 11.6
Mean (Range):
Tibia: 5.4 (3-1 6)
Patella: 9.2 (4-1 8)
Meant SD
Tibia:
Normotensive:
20.27 ±11. 55
Borderline
hypertension:
23.46 ±15.02
Definite
hypertension:
22.69 ± 14.71
Patella:
Normotensive:
28.95 ±18.01
Borderline
hypertension:
33.73 ± 21 .76
Definite
hypertension:
32.72 ± 19.55
Mean± SD:
Tibia: 12.5 ±7.8
Calcaneus: 20.5 ±
10.2
for non-
Distribution
of Bone Pb
(ug/g)
Not reported
High
exposure:
>24
Low exposure:
<8.7
Lowest
quintile: Tibia:
8.5
Patella: 12.0
Highest
quintile:
Tibia: 36.0
Patella: 53.0
Tibia
Q1 : 0-5.91
Q2:
5.92-10.40
Q3:
10.41-15.50
Q4:> 15.51
Calcaneus
Q1: 0-1 1.70
Q2:
11.71-19.07
Q3:
19.08-25.28
Q4: > 25.29
November 2012
4-96
Draft - Do Not Cite or Quote
-------
Reference
Elmarsafawy
et al. (2006)
Glass et al.
(2009)
Hsieh et al.
(2009b)
Huetal.
(1996a)
[As reported
in Navas-
Acien et al.,
(2008)1
Study Methods
Cohort:
Normative Aging Study
Age (yrs): Not reported
N: 471 elderly males
Location: Greater Boston area,
MA
Study Period: 6/1 991-1 2/1 994
Cohort:
Baltimore Memory Study
Age (yrs): Mean: 59.4;
Range: 50-70
N: 1,001
Location: Baltimore, MD
Study Period: 2001 -2005
Cohort:Not reported
Age (yrs): Mean: Control: 46.06
N: 18 controls
Location: Not reported
Study Period: Not reported
Cohort:
Normative Aging Study
Age (yrs): 48-92; Mean ± SD:
66.6 ± 7.2
N: 590 males
Location: Boston, MA
Study Period: 8/1 991-1 2/1 994
Prior Pb
Exposure
Not reported
Cumulative
(lifetime)
Control group for
occupational
exposure group
Cumulative
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Meanni SD:
Patella Tibia: 21.6±1 2.0
Patella: 31 .7 ±18.3
Tibia Mean ± SD:
Tibia: 18.8 ±11.1
Tibia Mean ± SD
Patella Tibia Control: 18.51
± 22.40
Patella Control: 7.14
±9.81
Tibia Mean ± SD:
Patella Tibia: 21.8 ± 12.1
Patella: 32.1 ±18.7
Range:
Tibia: <1-96
Patella: 1-142
Distribution
of Bone Pb
(ug/g)
Not reported
NPH Scale:
Lowest tertile:
Mean Tibia
level: 16.3±
11.0
Middle tertile:
Mean Tibia
level: 19.3±
10.7
Highest tertile:
Mean Tibia
level: 20.3 ±
11.4
Not reported
Figures 1 and
2 show both
types of bone
Pb levels
increasing
with age
November 2012
4-97
Draft - Do Not Cite or Quote
-------
Reference
Jain et al.
(2007)
Kamel et al.
(2002):
Kamel et al.
(2005):
Kamel et al.
(2008)
Prior Pb
Study Methods Exposure
Cohort: Not reported
VA-Normative Aging Study
Age (yrs): Not reported
N: 837 males
Location: Greater Boston, MA
Study Period:
9/1/1991-12/31/2001
Cohort:Not reported Cumulative
Age (yrs): 30-80 Control group for
N: 256 controls (Bone samples occupational
collected from 41 controls) exposure group
Location: New England (Boston,
MA)
Study Period: 1993-1996
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Mean ± SD
Patella Tibia:
Non-Cases: 21 .4 ±
13.6
Cases: 24.2 ± 15.9
Patella:
Non-cases:
30.6±19.7
Cases: 36.8 ± 20.8
Range:
Tibia:
Noncases:
-3-1 26
Cases: -5-75
Patella:
Noncases: -10-165
Cases: 5-101
Tibia Mean ± SE
Patella Tibia Controls: 11.1 ±
1.6
Patella Controls:
16.7 ±2.0
Distribution
of Bone Pb
(ug/g)
Mean ± SD
(Range):
Tibia:
Non-cases:
Tertile 1: 10.2
±3.8 (-3-1 5)
Tertile 2: 19.1
±2.3(16-23)
Tertile 3: 35.5
± 14.4
(24-1 26)
Cases:
Tertile 1: 10.1
±5.3 (-5-1 5)
Tertile 2: 19.8
±2.2(16-23)
Tertile 3: 39.5
±14.9(25-75)
Patella:
Non-cases:
Tertile 1 :
13.9±4.9
(-10-20)
Tertile 2:
27.1±4.1
(21 -34)
Tertile 3:
52.5± 20.7
(35-165)
Cases:
Tertile 1 :
15.3±4.3
(5-19)
Tertile 2: 25.7
±3.8(21-33)
Tertile 3: 53.3
±17.3
(35-101)
Controls
Tibia: N (%)
-7-7: 14(34)
8-1 4: 1 2 (29)
15-61: 15(37)
Patella: N (%)
-4-9: 14(34)
10-20:14(34)
21-107: 13
(32)
November 2012
4-98
Draft - Do Not Cite or Quote
-------
Reference
Khalil et al.
(2009a)
Korrick et al.
(1999) [As
reported in
Navas-Acien
etal.,
(2008)]
Study Methods
Cohort:
1982 Lead Occupational Study
Age (yrs): Control mean: 55
N: 51 controls
Location: Eastern Pennsylvania
Study Period: 1982-2004
Cohort:
Nurses' Health Study
Age (yrs): Combined: 47-74;
Mean ± SD:
Combined: 58.7 ± 7.2;
Cases: 61.1 ±7.1;
High controls: 61.1 ± 7.2;
Low controls: 58. 7 ± 7.1
Prior Pb
Exposure
Control group for
occupational
exposure group
Nonoccupationally
exposed
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Median (IQR)
Tibia Control: 12
(-8-32)
Tibia Mean ± SD
Patella Tibia:
Combined: 13.3 ±
9.0
Cases: 13.0 ±9.4
High controls: 14.7 ±
10
Distribution
of Bone Pb
(ug/g)
Not reported
Patella:
10th
percentile: 6
90th
percentile: 31
N: 284 females; (89 cases;
195 controls)
Location: Boston, MA
Study Period: 7/1993-7/1995
Low controls: 12.7 ±
8.1
Patella:
Combined: 17.3 ±
11.1
Cases: 19.5± 12.9
High controls: 17.2 ±
9
Low controls: 15.8 ±
10.6
Lee et al.
(2001 a) [As
reported in
Navas-Acien
etal.,
(2008)1
Martin et al.
(2006)
Cohort: Not reported
Age (yrs):
22.0-60.2
Mean ± SD: Controls:
34.5 ±9.1
N: 135 controls
Location: South Korea
Study Period:
10/24/1997-8/19/1999
Cohort:
Baltimore Memory Study
Control group for Tibia
occupational
exposure group
Cumulative Tibia
(lifetime)
Range
Tibia Combined:
-5-69
Patella Combined:
-5-87
Mean ± SD
Tibia Controls: 5.8 ±
7.0
Range
Tibia Controls:
-11-27
Mean ± SD
Tibia: 18.8 ±12.4
Not reported
Tibia IQR:
1 1 .9-24.8
Age (yrs): 50-70; Mean: 59.4
N:964
Location: Baltimore, MD
Study Period: 5/2001-9/2002
(1st study visit)
8/2002-3/2004 (2nd study visit -
tibia Pb measured)
November 2012
4-99
Draft - Do Not Cite or Quote
-------
Reference
Needleman
et al. (2002)
Osterberg et
al. (1997)
[As reported
in Shih et
al., (2007)1
Parket al.
(2006)
Prior Pb
Study Methods Exposure
Cohort: Not reported Not reported
Age (yrs):
12-18; Mean age ± SD:
African American cases:
15.8 ±1.4
African American controls:
15.5 ± 1 .1;
White cases: 15.7± 1.3;
White controls: 1 5.8 ± 1.1
N: 194 male youth cases; 146
male youth controls
Location: Allegheny County, PA
(cases); Pittsburgh, PA (controls)
Study Period: 4/1 996-8/1 998
Cohort: Not reported Control group for
Age (yrs): Median: 41 .5 occupational
exposure group
N: 19 male controls
Location: Not reported
Study Period: Not reported
Cohort: Not reported
Normative Aging Study
Age (yrs): Mean: 72.9 ± 6.5
N: 41 3 males
Location: Greater Boston, MA
Study Period:
11/14/2000-12/22/2004; (HRV
measurements taken); 1991-2002
(bone Pb measurements taken)
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Mean ± SD
Tibia Cases (ppm):
All subjects: 1 1 .0 ±
32.7
African American:
9.0 ±33.6
White: 20 ± 27.5
Tibia Controls (ppm):
All subjects: 1 .5 ±
32.1
African American:
-1 .4± 31 .9
White: 3.5 ± 32.6
Finger bone Median (range)
Finger Bone
Controls:
4 (-19-1 8)
Tibia Median (IQR)
Patella
Tibia: 19.0(11-28)
Patella: 23.0 (15-34)
Estimated Patella3:
16.3(10.4-25.8)
Distribution
of Bone Pb
(ug/g)
Table 4 of
paper
distributes
bone Pb by
> 25 or <25 for
race, two
parental
figures, and
parent
occupation
Not reported
Median (IQR)
for No. of
metabolic
abnormalities:
Tibia:
0: 18.5
(10.5-23)
1:19(11 -28)
2: 19(12-26)
Patella:
0:22(13.5-32)
1:25(16-36)
2:20(15-32)
Estimated
Patella:
0:16.3
(10.8-24.8)
1: 17.1
(11-29.3)
2:15.1
(9.4-22.1)
November 2012
4-100
Draft - Do Not Cite or Quote
-------
Reference
Parket al.
(2009b)
Parket al.
(2010)
Payton et al.
(1998)
Peters et al.
(2007)
Rajan et al.
(2007)
Prior Pb
Study Methods Exposure
Cohort: Not reported
Normative Aging Study
Age (yrs): Mean: 67.3 ± 7.2
N: 613 males
Location: Greater Boston, MA
Study Period: 8/1 991-1 2/1 995
Cohort: Cumulative (chronic
VA Normative Aging Study cohort exposure)
Age (yrs): Mean: 64.9 (at bone
Pb measurement)
N: 448 males
Location: Eastern Massachusetts
Study Period: 1991-1996
Cohort: Not reported
VA Normative Aging Study cohort
Age (yrs): Mean: 66.8
N: 141 males
Location: Boston, MA
Study Period: 4/1 993-3/1 994
Cohort: Cumulative
Normative Aging Study cohort
Age (yrs): Mean: 66.9
N: 513 male cases
Location: Boston, MA
Study Period: 1991 -1996
Cohort: Not reported
VA Normative Aging Study Cohort
Age (yrs): Mean: 67.5 (at bone
scan)
N: 1075 males
Location: Boston, MA
Study Period: 1991 -2002
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Median (IQR)
Patella
Tibia: 19(14-27)
Patella: 26 (18-37)
Tibia Mean ± SD
Patella Tibia: 22.5 ± 14.2
Patella: 32.5 ±20.4
Tibia Mean ± SD
Patella Tibia: 22.5 ± 12.2
Patella: 31 .7 ± 19.2
Tibia Mean ± SD
Patella Tibia: 21 .5 ± 13.4
Patella: 31 .5 ± 19.3
Tibia Mean ± SD
Patella Tibia: 22.1 ± 13.8
Patella: 31 .4 ± 19.6
Distribution
of Bone Pb
(ug/g)
Table 1 of
paper
distributes
tibia and
patella Pb by
genotype;
Table 2 of
paper
distributes
tibia and
patella Pb by
number of
gene variants
Tibia IQR: 15
Patella IQR:
21
Table 2 of
paper
provides age-
adjusted mean
bone Pb levels
(age, race,
education,
smoking
[pack-yr],
occupational
noise, noise
notch, BMI,
hypertension,
diabetes)
Not reported
Not reported
Not reported
November 2012
4-101
Draft - Do Not Cite or Quote
-------
Reference
Rajan et al.
(2008)
Rhodes et
al. (2003)
Roelset al.
(1994)
Rothenberg
etal.
(2002a) [as
reported in
Navas-Acien
et al. (2008)1
Study Methods
Cohort:
VA Normative Aging Study Cohort
Age (yrs): a 45
N: 720 males
Location: Boston, MA
Study Period: 1993-2001
Cohort:
VA Normative Aging Study Cohort
Age (yrs): Mean: 67.1
N: 526 males
Location: Boston, MA
Study Period:
1/1/1991-12/31/1995
Cohort: Not reported
Age (yrs): 30-60
N: 68 males
Location: Belgium
Study Period: Not reported
Cohort: Not reported
Age (yrs): 15-44; Mean ± SD:
31.0 ±7.7
N: 720 females
Location: Los Angeles, CA
Study Period: 6/1995-5/2001
Prior Pb
Exposure
Current and
cumulative
Not reported
Control group for
occupational
exposure group
Not reported
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Mean ± SD
Patella ALAD1-1
Tibia: 21 .9 ±13.8
Patella: 29.3 ± 19.1
ALAD 1 -2/2-2
Tibia: 21 .2 ±11. 6
Patella: 27.9 ± 17.3
Tibia Mean ± SD
Patella Tibia: 21 .9 ± 13.5
Patella: 32.1 ±19.8
Tibia Geometric Mean
(Range)
Tibia Controls:
Normotensive: 21.7
(<1 5.2-69.3)
Hypertensive: 20.2
(<1 5.2-52.9)
Total: 21 .4
(<1 5.2-69.3)
Tibia Mean ± SD
Calcaneus Tibia: 8.0 ± 11 .4
Calcaneus: 10.7 ±
11.9
Distribution
of Bone Pb
(ug/g)
Not reported
No. of
participants
Tibia:
<1-15: 173
(33)
16-24: 186
(35)
25-126: 167
(32)
Patella:
<1-22: 189
(36)
23-35: 165
(31)
36-165:172
(33)
Not reported
Tibia quartiles:
Q1 : -33.7-0.9
Q2: 1 .0-8.0
Q3: 8. 1-16.1
Q4: 16.2-42.5
Calcaneus
quartiles:
Q1 : -30.6-3.0
Q2: 3. 1-10.0
Q3: 10.1 -18.7
Q4: 18.8-49.0
November 2012
4-102
Draft - Do Not Cite or Quote
-------
Reference Study Methods
Prior Pb
Exposure
Bone Pb
biomarker
Bone Pb Cone.
Distribution
of Bone Pb
(ug/g)
Shihetal.,
(2006)
Cohort:
Baltimore Memory Study cohort
Age (yrs): Mean: 59.39
N:985
Location: Baltimore, MD
Study Period: Not reported
Not reported
Tibia
Meant SD:
Tibia: 18.7 ± 11.2
Not reported
Stokes et al.
(1998) [as
reported in
Shihetal.
/onnvM
(ZUU/ )l
Cohort: Not reported
Age (yrs): 19-29 (in 1994); Mean
± SD: Cases: 24.3 ± 3.18
Control: 24.2 ± 3.02
Cases: 9 months-9 yr
(during 1 /1 /1 974-1 2/31 /1 975)
N: 257 cases; 276 controls
Location: Silver Valley, ID;
Spokane, WA
Study Period:
7/10/1994-8/7/1994
Cumulative Tibia
(lifelong)
Environmental
(resided near Pb
smelter during
childhood)
Mean (Range):
Tibia Cases: 4.6
(-28.9-37)
Tibia Controls: 0.6
(-46.4-1 7.4)
Tibia
No. of Cases:
<1 ug/g:
31 .5%
1-5 ug/g:
24.4%
5-10 ug/g:
22.3%
>10 ug/g:
21 .8%
No. of
Controls:
<1 ug/g:
50.4%
1-5 ug/g:
25.6%
5-10 ug/g:
19.4%
>10 ug/g:
4.7%
Mean ± SD
Tibia
concentration
by age group:
Cases:
19-21: 1.47±
8.35
22-24: 4.48 ±
7.45
25-27: 4.82 ±
8.92
28-30: 6.64 ±
9.53
Controls:
19-21: 1.27±
6.60
22-24: -0.61 ±
6.19
25-27: 0.60 ±
8.60
28-30: 1.74 ±
6.42
November 2012
4-103
Draft - Do Not Cite or Quote
-------
Reference
Van
Wijngaarden
et al. (2009)
Wasserman
et al. (2003)
Weisskopf et
al. (2004).
[as reported
in Shih et al.
(2007)1
Weisskopf et
al. (2007a)
Prior Pb
Study Methods Exposure
Cohort: Not reported Cumulative
Age (yrs): Mean: 61 .5
N:47
Location: Rochester, NY
Study Period: Not reported
Cohort: Cumulative
Yugoslavia Prospective Study of (lifetime)
Environmental Pb Exposure Environmental
Age (yrs): 1 0-1 2 (Pb smelter,
N: 167 children refinery, battery
plant)
Location: Kosovska, Mitrovica,
Kosovo, Yugoslavia;
Pristina, Kosovo, Yugoslavia
Study Period: 5/1 985-1 2/1 986
(mother's enrollment); 1986-1999
(follow-up through age 1 2 yr);
Tibia Pb measured 1 1 -1 3 yr old
Cohort: Environmental
Normative Aging Study
Age (yrs): Mean ± SD: 67.4 ± 6.6
N: 466 males
Location: Boston, MA
Study Period: 1991 -2002
Cohort: Not reported
VA Normative Aging Study cohort
Age (yrs): Mean:
Lowest Patella quintile: 73.2;
Highest Patella quintile: 80.7
N: 31 males
Location: Boston, MA
Study Period: Bone Pb
measured: 1994-1999 Scans
performed: 2002-2004
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Mean ± SD
Calcaneus Tibia: 2.0 ± 5.2
Calcaneus: 6.1 ± 8.5
Tibia Mean ± SD:
Tibia
Pristina: 1.36 ±6.5
Mitrovica: 39.09±
24.55
Tibia Median (IQR)
Patella Tibia: 19 (12,26)
Patella: 23 (15, 35)
Tibia Median (IQR)
Patella Tibia
Lowest quintile: 13
(9-17)
Highest quintile: 41
(38-59)
Patella
Lowest quintile: 9
(5-15)
Highest quintile: 63
(43-86)
Distribution
of Bone Pb
(ug/g)
Not reported
Tibia quartiles:
Q1: -14.4-1 .85
Q2: 1.85-1 0.5
Q3: 10.5-35
Q4: 35-1 93.5
Table 3 of
paper
distributes
tibia Pb by
sex, ethnicity,
address at
birth relative to
factory, and
maternal
education
Tibia IQR: 14
Patella IQR:
20
Table 3 of
paper shows
mean Pb
levels across
categorical
variables (yr of
education,
smoking
status,
computer
experience,
first language
English)
Not reported
November 2012
4-104
Draft - Do Not Cite or Quote
-------
Reference
Weisskopf et
al. (2007b)
Weisskopf et
al. (2009)
Weisskopf et
al. (201 0)
Prior Pb
Study Methods Exposure
Cohort: Concurrent and
VA Normative Aging Study cohort cumulative
Age (yrs): Mean: 68.7
N: 1,089 males
Location: Boston, MA
Study Period: 1993-2001
Cohort: Cumulative
Normative Aging Study; (95%
white)
Age (yrs): Mean ± SD (at Patella
baseline); Tertile 1 : 65.2 ± 7.1 ;
Tertile 2: 66.5 ± 6.5
Tertile 3: 70.2 ± 7.2
N: 868 males
Location: Greater Boston area,
MA
Study Period: 1991-1999
Cohort: Cumulative
BUMC, BWH, BIDMC, HVMA,
Normative Aging Study (NAS),
Harvard Cooperative Program on
Aging (HCPOA)
Age (yrs): Mean:
Cases: 66.5; Controls: 69.4
N: 330 cases; 308 controls
Location: Boston, MA
Study Period: 2003-2007
1991-1999 (NAS patients bone Pb
measured)
Distribution
Bone Pb Bone Pb Cone. of Bone Pb
biomarker (ug/g) (ug/g)
Tibia Median (IQR) Table 1 of
Patella Tibia: 20 (13-28) paper shows
distribution of
Patella: 25 (17-37) Pb biomarkers
by categories
of covariates
(age,
education,
smoking
status, alcohol
intake,
physical
activity,
computer
experience,
first language
English)
Tibia Mean ± SD Patella tertiles:
Patella Tibia: 21 .8 ± 13.6 1:<22
Patella: 31 .2 ±19.4 2:22-35
3: >35
Tibia Mean ± SD: Tibia quartiles:
Patella Tibia: 10.7 ± 12.1 Q1:<3.1
Patella: 13.6 ±15.9 Q2: 3.5-9.6
Q3: 10.0-17.0
Q4:>17.3
Patella
quartiles:
Q1:<2.7
Q2: 3.5-1 1.0
Q3:1 1.3-20.9
Q4: >20.9
November 2012
4-105
Draft - Do Not Cite or Quote
-------
Reference
Weuve et al.
(2006)
Weuve et al.
(2009)
Wright et al.
(2003) [as
reported in
Shihetal.
(2007)1
Prior Pb
Study Methods Exposure
Cohort: Cumulative
VA Normative Aging Study cohort
Age (yrs): a 45
N: 720 males
Location: Boston, MA
Study Period: 1991 (measuring
bone Pb levels)
End date not reported
Cohort: Recent and
Nurses' Health Study cohort cumulative
Age (yrs): 47-74
N: 587 females
Location: Boston, MA
Study Period: 1995-2005
Cohort: Environmental
Normative Aging Study
Age (yrs): Mean ± SD: 68.2 ± 6.9
N: 736 males
Location: Boston, MA
Study Period: 1991 -1997
Bone Pb Bone Pb Cone.
biomarker (ug/g)
Tibia Median (1st-3rd
Patella quartile):
Tibia: 19(13-28)
Patella: 27 (18-39)
Tibia Mean ± SD:
Patella Tibia: 10.5 ± 9.7
Patella: 1 2.6 ± 11.6
Tibia Mean ± SD:
Patella
Tibia: 22.4 ±15.3
Patella: 29.5 ±21 .2
Distribution
of Bone Pb
(ug/g)
Table 1 of
paper shows
distribution of
mean Pb bio-
marker levels
by
characteristics
of participants
(age,
education,
computer
experience,
smoking
status, alcohol
consumption,
fertile of Ca2+
intake, fertile
of physical
activity,
diabetes)
Not reported
Tibia:
Difference in
mean from
Lowest-
highest
quartile: 34.2
Patella:
Difference in
mean from
lowest-highest
quartile: 47
November 2012
4-106
Draft - Do Not Cite or Quote
-------
Table 4-10 Epidemiologic studies that provide bone Pb measurements for
occupationally exposed populations.
Reference
Bleecker et
al. (1997)
[as reported
in Shin et
al. (2007)1
Bleecker et
al.(2QQ7b)
Caffo et al.
(2008)
Dorsey et
al. (2006)
Glenn et al.
(2003) [as
reported in
Navas-
Acien et al.
(2008)1
Study Methods
Cohort: Canada Lead Study
Age (yrs): Cumulative: 24-64
Younger: 24-43
Older: 44-64
Mean ± SD:
Cumulative: 44.1 ± 8.36
Younger: 37.2 ± 4.57
Older: 50.9 ± 4.86
N: 80 males
Location: Canada
Study Period: Not Reported
Cohort: Not reported
Age (yrs): Mean: 39.7
N:61
Location: Northern Canada
Study Period: Not Reported
Cohort: Not reported
Age (yrs): Mean: 60.39
N: 51 3 males
Location: Delaware and New
Jersey, U.S.
Study Period: 1994-1997
(Phase 1 recruitment);
2001 -2003 (Phase 2
recruitment)
Cohort: Not reported
Age (yrs): Mean: 43.4
N:652
Location: Korea
Study Period:
10/24/1997-8/19/1999
(enrolled)
Cohort: Not reported
Age (yrs): 40-70; Mean: 55.8
(baseline)
N: 496 males
Location: Eastern U.S.
Study Period: 6/1 994-6/1 996
(enrolled); 6/1998 (follow-up
period ended)
Prior Pb
Exposure
Occupational
(Pb smelter
workers)
Occupational
(primary Pb smelter
workers)
Cumulative
Occupational
(Former organolead
manufacturing
workers)
Occupational (Pb
workers)
Occupational
(Chemical
manufacturing
facility; inorganic
and organic Pb)
Bone Pb
Bone Pb Concentration
biomarker (ug/g)
Tibia Mean ± SD (Tibia):
Cumulative: 41 .0 ±
24.44
Younger: 35 ±
24.11
Older: 46.9 ± 23.59
Range (Tibia):
Cumulative: -12-90
Younger: -12-80
Older: 3-90
Tibia Mean:
Tibia: 38.6
Tibia Mean ± SD:
Peak Tibia: 23.99 ±
18.46
Tibia Mean ± SD:
Patella Tibia: 33.5 ± 43.4
Patella: 75.1 ±
101.1
Tibia Mean ± SD:
Tibia: 14.7 ±9.4 (at
yr3)
Peak Tibia: 24.3 ±
18.1
Range:
Tibia: -1.6-52 (at
year 3)
Peak Tibia:
-2.2-118.8
Distribution of
Bone Pb
(ug/g)
Not reported
Not reported
Not reported
Not reported
Not reported
November 2012
4-107
Draft - Do Not Cite or Quote
-------
Reference
Glenn et al.
(2006)
Hanninen et
al. (1998)
[as reported
in Shin et
al. (2007)1
Study Methods
Cohort: Not reported
Age (yrs): 0-36.2 (baseline);
Mean ± SD: 41 .4 ±9.5
(baseline)
N: 575; (76% male; 24%
female)
Location: South Korea
Study Period: 10/1997-6/2001
Cohort: Not reported
Age (yrs): Mean ± SD:
Male: 43; Female: 48
Blood Pb (max) < 2.4 umol/L:
41 .7 ±9.3
Blood Pb (max) >2.4 umol/L:
46.6 ±6.2
N: 54; (43 males, 11 females)
Location: Helsinki, Finland
Study Period: Not reported
Prior Pb
Exposure
Cumulative and
recent
Occupational (Pb-
using facilities)
Occupational (Pb
acid battery factory
workers)
Bone Pb Distribution of
Bone Pb Concentration Bone Pb
biomarker (ug/g) (ug/g)
Tibia Mean ± SD: Not reported
Tibia: 38.4 ±42.9
Tibia-Women:
Visit 1: 28.2±19.7
Visit 2: 22.8±20.9
Tibia-Men:
Visit 1 : 41 .7±47.6
Visit 2: 37.1 ±48.1
Tibia Mean ± SD: Not reported
Calcaneus
Tibia:
Blood Pb (max)
< 2.4 umol/L: 19.8 ±
13.7
Blood Pb (max)
>2.4 umol/L: 35.3 ±
16.6
f'alr^a nai ic'
Blood Pb (max)
< 2.4 umol/L: 78.6 ±
62.4
Blood Pb (max)
>2.4 umol/L: 100.4
±43.1
Hsiehetal.
2009
(2009b)
Cohort: Not reported Occupational Tibia
Age (yrs): Mean: (Pb paint factory Patella
Cases: 45.71 workers)
Controls: 46.06
N: 22 cases; 18 controls
Location: Not Reported
Study Period: Not reported
Mean ± SD
Tibia
Case: 61 .55 ± 30.21
Control: 18.51 ±
22.40
Patella
Not reported
Case: 66.29 ± 19.48
Control: 7.14 ±9.81
November 2012
4-108
Draft - Do Not Cite or Quote
-------
Reference
Kamel et al.
(2002):
Kamel et al.
(2005):
Kamel et al.
(2008)
Khalil et al.
(2009a)
Prior Pb
Study Methods Exposure
Cohort: Not reported Cumulative
Age (yrs): 30-80 Occupational (Pb
N: 109 cases; 256 controls; fumes' dust' or
(Bone samples collected from particles)
104 cases and 41 controls)
Location: New England
(Boston, MA)
Study Period: 1993-1996
Cohort: 1982 Pb Occupational Occupational (Pb
Study cohort battery plant
Age (yrs): Mean: workers)
Cases: 54
Bone Pb
Bone Pb Concentration
biomarker (ug/g)
Tibia Mean ± SE
Patella Tibia
Cases: 14.9± 1.6
Controls: 11.1 ±1.6
Patella
Cases: 20. 5 ± 2.1
Controls: 16.7± 2.0
Tibia Median (IQR)
Tibia
Cases: 57 (20-86)
Controls: 12 (-8-32)
Distribution of
Bone Pb
(ug/g)
Cases
Tibia Pb: N (%)
-7-7: 21 (20)
8-14:35(34)
15-61: 48(46)
Patella Pb: N (%)
-4-9: 27 (26)
1 0-20: 40 (38)
21-107:37(36)
Controls
Tibia Pb: N (%)
-7-7: 14(34)
8-14: 12(29)
15-61: 15(37)
Patella Pb: N (%)
-4-9: 14(34)
10-20: 14(34)
21-107:13(32)
Not reported
Osterberg
et al. (1997)
[as reported
in Shih et
al. (2007)1
Controls: 55
N: 83 cases; 51 controls
Location: Eastern
Pennsylvania
Study Period: 1982-2004
Cohort: Not reported
Age (yrs): Median: 41.5
N: 38 male cases; 19 male
controls
Location: Not reported
Study Period: Not Reported
Occupational
(secondary Pb
smelter- inorganic
Pb)
Finger bone
Median
Finger Bone:
High Cases: 32
Low cases: 16
Control: 4
Range
Finger Bone:
High Cases: 17-101
Low cases: -7-49
Control:-19-18
Not reported
November 2012
4-109
Draft - Do Not Cite or Quote
-------
Reference
Roelset al.
(1994)
Schwartz
etal.
(2000b) [as
reported in
Shihetal.,
(2007)1
Schwartz et
al. (2000c)
[as reported
in Navas-
Acien et al.
(2008)1
Schwartz et
al. (2001):
Lee et al.
(2001 a)
Study Methods
Cohort: Not reported
Age (yrs): 30-60
N: 76 male cases; 68 male
controls
Location: Belgium
Study Period: Not Reported
Cohort: U.S. Organolead Study
Age (yrs): Mean ± SD:
Cases: 55.6 ± 7.4
Controls: 58.6 ± 7.0
N: 535 male cases
118 male controls
Location: Eastern U.S.
Study Period: 6/1 994-1 0/1 997
(enrolled); Completed 2-4
annual follow-up visits; Tibia
Pb taken in 3rd year
Cohort: Not reported
Age (yrs): 41 .7-73.7
(Combined)
Mean ± SD:
Combined: 57.6 ± 7.6
Hypertensive: 60.2 ± 6.9
Nonhypertensive: 56.6 ± 7.5
N: 543 males
Location: Eastern U.S.
Study Period: 1995 (recruited);
1996-1 997 (Tibia Pb
taken during the 3rd yr)
Cohort: Not reported
Age (yrs): Mean:
Exposed: 40.4
Control: 34.5
N: 803 cases; 135 controls
Location: South Korea
Study Period:
10/24/1997-8/19/1999
Prior Pb
Exposure
Occupational (Pb
smelter workers)
Mean case
exposure: 18 yr
(range: 6 to 36 yr)
Occupational
(tetraethyl and
tetramethyl Pb
manufacturing
facility)
Occupational
(former organolead
manufacturing
workers)
Occupational
(battery
manufacturing,
secondary smelting,
Pb oxide
manufacturing, car
radiator
manufacturing)
Bone Pb
Bone Pb Concentration
biomarker (ug/g)
Tibia Geometric Mean
(Range)
Tibia Cases:
Normotensive: 64.0
(19.6-167.1)
Hypertensive: 69.0
(21.7-162.3)
Total: 65.8
(19.6-167.1)
Tibia Controls:
Normotensive: 21.7
(<1 5.2-69.3)
Hypertensive: 20.2
(<1 5.2-52.9)
Total: 21 .4
(<1 5.2-69.3)
Tibia Mean ± SD
Current Tibia:
Cases: 14.4 ±9.3
Peak Tibia:
Cases: 22.6 ± 16.5
Tibia Mean ± SD
Tibia:
Combined: 14.4±
9.3
Hypertensive: 15.4
±9.1
Nonhypertensive:
14.0 ±9.3
Range Tibia:
Combined: -1.6-52
Tibia Mean ± SD
Tibia
Cases: 37.1 ±40.3
Control: 5.8 ± 7.0
Range:
Tibia
Cases: -7-338
Controls: -11-27
Distribution of
Bone Pb
(ug/g)
Not reported
Not reported
Not reported
Not reported
November 2012
4-110
Draft - Do Not Cite or Quote
-------
Reference
Schwartz et
al. (2005)
Stewart et
al. (1999)
[as reported
in Shih et
al., (2007)1
Stewart et
al. (2006)
Weaver et
al. (2008)
Study Methods
Cohort: Not reported
Age (yrs): Mean at 1st visit:
41.4
N:576
Location: South Korea
Study Period: 10/1997-6/2001
Cohort: U.S. Organolead Study
Age (yrs): 40-70 (in 1995)
38% > 60 yrs
Mean: 58
N: 534 males
Location: Eastern U.S.
Study Period: Not Reported
Cohort: Not reported
Age (yrs): Mean: 56.1
N: 532 males
Location: Eastern U.S.
Study Period: 1994-1997;
2001-2003
Cohort: Not reported
Age (yrs): Mean ± SD: 43.3 ±
9.8
N:652
Location: South Korea
Study Period: 12/1999-6/2001
Prior Pb
Exposure
Occupational
(current and former
Pb workers)
Occupational
(tetraethyl and
tetramethyl Pb
manufacturing
facility)
Cumulative
Occupational
(Organolead
workers - not
occupationally
exposed to Pb at
time of enrollment)
Occupational
(Current and former
Pb workers; plants
produced Pb
batteries, Pb oxide,
Pb crystal, or
radiators, or were
secondary Pb
smelters)
Bone Pb
Bone Pb Concentration
biomarker (ug/g)
Tibia Mean ± SD
Tibia: 38.4 ±43
Tibia Mean ± SD
Tibia:
Current: 14.4 ±9.3
Peak: 23.7 ± 17.4
Range: Tibia
Current: -1 .6-52
Peak: -2.2-105.9
Tibia Mean ± SD
Current Tibia: 14.5
±9.6
Peak Tibia: 23.9 ±
18.3
Patella Mean ± SD
Patella: 37.5 ± 41 .8
Distribution of
Bone Pb
(ug/g)
Tibia:
25th percentile at
V1: 14.4
75th percentile at
V1:47.1
Current Tibia Pb:
N (%)
<5: 77 (14.2)
5-9.99: 113
(20.8)
10-14.99: 119
(21.9)
15-19.99: 117
(21.5)
>20: 118(21.7)
Peak Tibia Pb: N
<5:49(9.1)
5-9.99: 64 (1 1 .8)
10-14.99: 70
(12.9)
15-19.99:87
(16.1)
20-24.99: 79
(14.6)
25-29.99: 55
(10.2)
>30: 137(26.1)
Not reported
Not reported
November 2012
4-111
Draft - Do Not Cite or Quote
-------
4.4.3
Pb in Urine
i
2
o
J
4
5
6
Urine Pb concentrations in the U.S. general population have been monitored in the
NHANES. Data from the most recent survey (CDC. 2011 a) are shown in Table 4-11. The
geometric mean for the entire sample
creatinine (95% CI: 0.48, 0.55). The §
(n = 1,300) were 0.50 (ig/g creatinine
CI: 0.49, 0.57), respectively.
for the period 2007-2008 (n =
geometric means for males (n =
(95% CI: 0.47, 0.53) and 0.53
2,627) was 0.52 ug/g
= 1,327) and females
(ig/g creatinine (95%
Table 4-11 Urine Pb concentrations in the U.S. population.
Survey Stratum Period Geometric Mean (ug/g CR)a 95% Confidence Interval
All 1999-2000 0.721
2001-2002 0.639
2003-2004 0.632
2005-2006 0.546
2007-2008 0.515
6-1 1 yr 1999-2000 1.170
2001-2002 0.918
2003-2004 0.926
2005-2006 0.628
2007-2008 0.644
12-1 9 yr 1999-2000 0.496
2001-2002 0.404
2003-2004 0.432
2005-2006 0.363
2007-2008 0.301
> 20 yr 1 999-2000 0.720
2001-2002 0.658
2003-2004 0.641
2005-2006 0.573
2007-2008 0.546
Males 1 999-2000 0.720
2001-2002 0.639
2003-2004 0.615
2005-2006 0.551
2007-2008 0.502
Females 1999-2000 0.722
2001-2002 0.639
0.700, 0.742
0.603, 0.677
0.603, 0.662
0.502, 0.573
0.483, 0.549
0.975, 1.41
0.841 , 1 .00
0.812,1.06
0.563, 0.701
0.543, 0.763
0.460, 0.535
0.380, 0.428
0.404, 0.461
0.333, 0.395
0.270, 0.336
0.683, 0.758
0.617,0.703
0.606, 0.679
0.548, 0.600
0.513, 0.580
0.679, 0.763
0.607, 0.673
0.588, 0.644
0.522, 0.582
0.471,0.534
0.681, 0.765
0.594, 0.688
Number of Subjects
2,465
2,689
2,558
2,576
2,627
340
368
290
355
394
719
762
725
701
376
1,406
1,559
1,543
1,520
1,857
1,227
1,334
1,281
1,271
1,327
1,238
1,355
November 2012
4-112
Draft - Do Not Cite or Quote
-------
Survey Stratum
Mexican -Americans
Non-Hispanic blacks
Non-Hispanic whites
Period Geometric Mean (ug/g CR)a 95% Confidence Interval
2003-2004
2005-2006
2007-2008
1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
1 999-2000
2001-2002
2003-2004
2005-2006
2007-2008
aValues are in ug Pb/g creatinine (CR)
Source: Based on data from the NHANES
0.648
0.541
0.527
0.940
0.810
0.755
0.686
0.614
0.722
0.644
0.609
0.483
0.452
0.696
0.615
0.623
0.541
0.506
(CDC. 2011 a)
0.601,0.698
0.507, 0.577
0.489, 0.568
0.876,1.01
0.731, 0.898
0.681, 0.838
0.638, 0.737
0.521,0.722
0.659, 0.790
0.559, 0.742
0.529, 0.701
0.459, 0.508
0.414, 0.492
0.668, 0.725
0.579, 0.654
0.592, 0.655
0.500, 0.585
0.466, 0.550
Number of Subjects
1,277
1,305
1,300
884
682
618
652
515
568
667
723
692
589
822
1,132
1,074
1,041
1,095
4.4.4 Pb in Teeth
1 The influence of historical Pb exposures was recently studied by Robbins et al. (2010).
2 Tooth enamel samples from 127 subjects born between 1936 and 1993 were analyzed for
3 Pb concentration and Pb isotope ratios of the tooth enamel and compared with those
4 parameters for sediment cores and estimates of Pb emissions from gasoline during the
5 years when 50% enamel formation was estimated to occur. They found that the log-
6 transform of tooth enamel concentration was significantly predicted by the log-transform
7 of Lake Erie sediment core data obtained by Graney et al. (1995) (p <0.00001) and by the
8 log-transform of U.S. consumption of Pb in gasoline (p <0.00001); see Figure 4-21.
9 Additionally, Robbins et al. (2010) found that 207Pb/206Pb was significantly predicted by
10 the 207Pb/206Pb observed in the Lake Erie sediment cores obtained by Graney et al. (1995)
11 (p O.OOOl) and forthis study (p <0.0002).
November 2012
4-113
Draft - Do Not Cite or Quote
-------
100
g
I
8. 50
25
o-
1930 1940 1950
1960 1970 1980 1990
Year
Note: The lines and symbols on the plot represent Pb in study participant teeth (solid line), newly obtained Pb sediment Lake Erie
cores (open triangles), Pb in previously obtained Lake Erie sediment [open circles, Graney et al. (1995)1, and U.S. gasoline usage
(closed circles). All values are normalized by the peak observation for that parameter.
Source: Reprinted with permission of Elsevier Publishing, Robbins et al. (2010).
Figure 4-21 Comparison of relative temporal changes in tooth enamel Pb
concentration.
i
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
Several Brazillian studies have found increased levels of Pb in teeth in areas where Pb
sources are present. For example, Costa de Almeida et al. (2007) reported Pb
concentration in tooth enamel among 4-6 year old kindergarteners in Sao Paulo, Brazil to
be significantly higher for children living near a Pb-acid battery processing plant in the
Baruru neighborhood compared with 4-6 year old children in other parts of the city
(non-exposed median: 206 mg/kg, n = 247; exposed median: 786 mg/kg, n = 26;
p <0.0001). Subsequent analysis revealed that 55% of 4-6 year old children from Baruru
had tooth enamel Pb concentrations greater than 600 mg/kg, forming a significant
comparison with other neighborhoods having 0-33% of 4-6 year old children with tooth
enamel Pb greater than 600 mg/kg (p <0.0001) (de Almeida et al., 2008). The authors did
not describe controlling for additional factors, such as socioeconomics or housing
conditions. Arruda-Neto et al. (2009) studied Pb in tooth samples among Sao Paulo
children to compare exposures of children age 4-12 years, living near a dam with heavy
metal sediments with those of children ages 4-13 years, living in a control area thought to
have few exposures. They observed a significant comparison (near dam: avg 1.28 ± 0.11
mg/kg, n = 50; control region: avg 0.91 mg/kg, n = 24). In a related study of Pb measures
in teeth among the general population ages 7-60 years, Arruda-Neto et al.(2010) observed
that 10-year old children had the highest teeth Pb concentrations, which were 115% of
the teeth Pb concentrations in 7-year olds. Twenty-year old subjects had teeth
Pb concentrations at roughly 50% of the 7-year olds' teeth Pb concentrations. Tooth Pb
November 2012
4-114
Draft - Do Not Cite or Quote
-------
1 concentrations stayed fairly constant throughout adulthood but then dropped to just above
2 30% among 65-year old subjects. Note that the authors did not clarify if average or
3 median values were presented, nor did they adjust for potentially confounding factors.
4.5 Empirical Models of Pb Exposure-Blood Pb Relationships
4 Multivariate regression models, commonly used in epidemiology, provide estimates of
5 the contribution of variance in the internal dose metric to various determinants or control
6 variables (e.g., air Pb concentration, surface dust Pb concentration). Structural equation
7 modeling links several regression models together to estimate the influence of
8 determinants on the internal dose metric. Regression models can provide estimates of the
9 rate of change of blood or bone Pb concentration in response to an incremental change in
10 exposure level (i.e., slope factor). One strength of regression models for this purpose is
11 that they are empirically verified within the domain of observation and have quantitative
12 estimates of uncertainty imbedded in the model structure. However, regression models
13 are based on (and require) paired predictor-outcome data, and, therefore, the resulting
14 predictions are confined to the domain of observations and are typically not generalizable
15 to other populations. Regression models also frequently exclude numerous parameters
16 that are known to influence human Pb exposures (e.g., soil and dust ingestion rates) and
17 the relationship between human exposure and tissue Pb levels, parameters which are
18 expected to vary spatially and temporally. Thus, extrapolation of regression models to
19 other spatial or temporal contexts, which is often necessary for regulatory applications of
20 the models, can be problematic.
21 A variety of factors may potentially affect estimates of blood Pb-air Pb slope factors.
22 Simultaneous changes in other (nonair) sources of Pb exposure can affect the relationship
23 indicated for air Pb. For example, remedial programs (e.g., community and home-based
24 dust control and education) may be responsible for partial blood Pb reduction seen in
25 some studies. The effect of remedial programs may lead to an overestimation of declines
26 in blood Pb due to changes in air Pb and a corresponding positive bias in blood Pb-air Pb
27 slopes. However, model adjustment for remedial programs and other factors (e.g., soil Pb
28 concentrations) may also cause a negative bias in blood Pb-air Pb slopes. A tendency
29 over time for children with lower blood Pb levels to not return for follow-up testing has
30 been reported. The follow-up of children with higher blood Pb levels would likely lead to
31 an underestimation of reductions in blood Pb following reductions in air Pb and cause a
32 negative bias in blood Pb-air Pb slopes. Another factor is the extent to which all the air
33 Pb exposure pathways are captured by the data set and its analysis. For example, some
34 pathways (such as exposure through the diet or surface soils) may respond more slowly to
35 changes in air Pb than others (such as inhalation). Additionally, some studies may include
November 2012 4-115 Draft - Do Not Cite or Quote
-------
1 adjustments for variables that also reflect an influence from air Pb (e.g., SES or soil Pb).
2 Studies may also vary in the ages of subjects, which given age-related changes in blood
3 Pb can also influence estimates. Many studies have utilized TSP measurements of air Pb
4 concentrations. The sampling efficiency of TSP samplers is affected by particle size
5 distribution, wind speed, and wind direction as described in Section 3.4.1. For example,
6 especially for larger particles (aerodynamic diameter of 20 urn or more), TSP sampling
7 efficiency decreases with increasing wind speed. Such effects on TSP sampling
8 efficiency can, in areas where such large particles are a substantial portion of airborne Pb,
9 lead to uncertainties in the comparability of air Pb concentrations between samples within
10 a study and across studies. A uniformly low bias in air Pb concentrations in a study could
11 positively bias estimated blood Pb-air Pb slopes for that study. Moreover, variability in
12 TSP samples is likely to result from temporal variation in wind speed, wind direction, and
13 source strength; see Sections 3.3 and 3.5. Such temporal variability would tend to
14 increase uncertainty and reduce the statistical strength of the relationship between air Pb
15 and blood Pb but may not necessarily affect the slope of this relationship. A number of
16 factors including those described above cause uncertainty in the magnitude of estimated
17 blood Pb-air Pb slope factors and may lead to both positive and negative biases in the
18 estimates from individual studies.
4.5.1 Air Pb-Blood Pb Relationships in Children
19 The 1986 Pb AQCD (U.S. EPA. 1986a) described epidemiological studies of
20 relationships between air Pb and blood Pb. Of the studies examined, the aggregate blood
21 Pb-air Pb slope factor (when considering both air Pb and Pb in other media derived from
22 air Pb) was estimated to be approximately double the slope estimated from the
23 contribution due to inhaled air alone (U.S. EPA. 1986a).
24 Much of the pertinent earlier literature (e.g., prior to 1984) on children's blood Pb levels
25 was summarized by Brunekreef (1984). Based on meta-analysis of data from studies of
26 urban or industrial-urban populations in 18 different locations, Brunekreef (1984)
27 estimated the blood Pb-air Pb slope for children to be 0.3485 ln[(ig/dL blood Pb] per
28 ln[(ig/m3 air Pb] (R2 = 0.69; see Figure 4-22). This slope corresponds to an increase of
29 4.6 (ig/dL blood Pb per (ig/m3 air Pb at an air Pb concentration of 1.5 (ig/m3 for all
30 groups included in the analysis. The 1.5 (ig/m3 value is the median of the air Pb
31 concentrations that match the blood Pb concentrations in 96 different child populations in
32 Figure 3 of Brunekreef et al. (1984). taken from the Appendix to the same paper. When
33 the analysis was limited to child populations whose mean blood Pb concentrations were
34 <20 (ig/dL (n=43), the slope was 0.2159 (R2=0.33), which corresponds to an increase of
35 4.8 (ig/dL blood Pb per (ig/m3 air Pb at the median air concentration (0.54 (ig/m3).
November 2012 4-116 Draft - Do Not Cite or Quote
-------
1 Newer studies that provide estimates for the blood Pb-air Pb slope factor are described in
2 the sections that follow. Those studies that have at least three data points are included, as
3 fewer than that contributes little to the understanding of the shape of the blood Pb-air Pb
4 relationship. A tabular summary of the major outcomes is provided in Table 4-12. In
5 some studies, the blood Pb-air Pb relationship was described with a nonlinear regression
6 function, in which the blood Pb-air Pb slope factor varied with air Pb concentration.
7 Studies also varied with regard to the use of simple or multivariate regression and, for the
8 latter, with regard to variables included. In Table 4-12. with the exception of Ranft et al.
9 (2008). slopes corresponding to a central estimate of the air Pb concentrations are
10 provided, to represent each study. These were calculated by evaluating each regression
11 function at ± 0.01 (ig/m3 from the central estimate of the air Pb concentration. Air Pb
12 concentration ranges and central estimates varied across studies, making it difficult to
13 interpret comparisons based solely on the central estimates of the slope factors.
14 Therefore, Figure 4-23 depicts the relationship between the blood Pb-air Pb slope factor
15 as a function of air Pb concentration for the range of air Pb concentrations evaluated in
16 those studies that provided the regression equation (the central estimate is also shown).
17 Figure 4-23 provides a more informative picture of the extent to which slope estimates
18 vary (and overlap) within and between studies. The Ranft et al. (2008) study includes a
19 separate term for soil Pb, so the blood Pb-air Pb slope factor presented for that study
20 underestimates the slope factor that would reflect all air-related pathways, since soil Pb
21 encompasses deposited ambient air Pb. A few studies used a log-log model that predict
22 an increase in the blood Pb-air Pb slope factor with decreasing air Pb concentration, and
23 the remainder of the studies used linear models that predict a constant blood Pb-air Pb
24 slope factor across all air Pb concentrations.
November 2012 4-117 Draft - Do Not Cite or Quote
-------
Table 4-12 Summary of estimated slopes for blood Pb to air Pb slope factors
in humans.
Reference
Study Methods
Model Description
Blood Pb-
AirPb
Slope3
Children Populations-Air
Location: Various countries
Years: 1974-1983
Brunekreef (1984) Subjects: Children (varying age ranges; n>190,000)
Analysis: Meta analysis of 96 child populations from
18 study locations
Model: Log-Log
Blood Pb: 5-76 ug/dL
(mean range for study
populations)
Air Pb: 0.1-24 ug/m3
(mean range for study locations)
All children:
4.6(1.5)b
Children
<20 ug/dL:
4.8 (0.54)°
Location: Chicago, IL
Years: 1974-1988
Hayes et al. (1994) Subjects: 0.5-5 yr (n = 9,604)
Analysis: Regression of quarterly median blood Pb
and quarterly mean air Pb
Model: Log-Log
Blood Pb: 10-28 ug/dL
(quarterly median range)
Air Pb: 0.05-1.2 ug/m3
(quarterly mean range)
8.2 (0.62)d
Location: Trail, BC
Years: 1989-2001
Subjects: 0.5-6 yr (Estimated n = 220-460, based on
292-536 blood Pb measurements/yr with 75-85%
participation)
Analysis: Regression of blood Pb screening and
community air Pb following upgrading of a local
smelter
Model: Linear
Blood Pb: 4.7-11.5 ug/dL
(annual geometric mean range)
AirPb: 0.03-1.1 ug/m3
(annual geometric mean range)
7.0 (0.48)e
Schwartz and
Pitcher (1989). U.S.
EPA (1986a)
Location: Chicago, IL
Years: 1976-1980
Subjects: Black children, 0-5 yr (n = 5,476)
Analysis: Multivariate regression of blood Pb with
mass of Pb in gasoline (derived from gasoline
consumption data and Pb concentrations in gasoline
for the U.S.)
Model: Linear
Blood Pb: 18-27 ug/dL(mean
range)'
Air Pb: 0.36-1.22 ug/m3
(annual maximum quarterly
mean)h
8.6 (0.75)9
Tripathi et al.
(2001)
Location: Mumbai, India (multiple residential
locations)
Years: 1984-1996
Subjects: 6-10 yr (n = 544)
Analysis: Regression of residential location-specific
average blood Pb and air Pb data
Model: Linear
Blood Pb: 8.6-14.4 ug/dL
(GM range for residential
locations)
Air Pb: 0.10-1.18 ug/m3
(GM range for residential
locations)
3.6 (0.45)'
November 2012
4-118
Draft - Do Not Cite or Quote
-------
Reference
Study Methods
Model Description
Blood Pb-
AirPb
Slope3
Children Populations-Air and Soilj
Ranft et al. (2008)
Location: Germany
Years: 1983-2000 (blood Pb and air Pb), 2000-2001
(soil Pb)
Subjects: 6-11 yr (n = 843)
Analysis: Pooled multivariate regression of 5 cross-
sectional studies
Model: Log-Linear
Blood Pb: 2.2-13.6 ug/dL
(5th-95th percentile)
Air Pb: 0.03-0.47 ug/m3
(5th-95th percentile)
3.2, 6.4R
Mixed Child-Adult Populations
Schwartz and
Pitcher (1989). U.S.
EPA (1986a)
Location: U.S.
Years: 1976-1980
Subjects: NHANES II, 0.5-74 yr, whites (n = 9,987)
Analysis: Multivariate regression of blood Pb with
mass of Pb in gasoline (derived from gasoline
consumption data and Pb concentrations in gasoline
for the U.S.)
Model: Linear
Blood Pb: 11-18 ug/dL9
(mean range)'
Air Pb: 0.36-1.22 ug/m3
(annual maximum quarterly
mean)h
9.3(0.75)'
a Slope is predicted change in blood Pb (ug/dL per ug/m3) evaluated at ± 0.01 ug/m3 from central estimate of air Pb for the study
(shown in parentheses), with the exception of Ranft et al. (2008) in which the slope from the paper is provided because the
regression equation was not available. The central estimate for Brunekreef (1984) is the median of air Pb concentrations since it
was a meta-analysis; for all other studies the mean is presented. For multiple regression models, this is derived based only on air
Pb coefficient and intercept. Depending on extent to which other variables modeled also represent air Pb, this method may
underestimate the slope attributable to air pathways. In single regression models, the extent to which non-modeled factors,
unrelated to air Pb exposures, exert an impact on blood Pb that covaries with air Pb may lead to the slope presented here to over
represent the role of air Pb.
b In(PbB) = In(PbA) x 0.3485 + 2.853
0 In(PbB) = In(PbA) x 0.2159 + 2.620
d In(PbB) = In(PbA) x 0.24 + 3.17
6PbB = PbAx7.0
f Observed blood Pb values not provided; data are for regressed adjusted blood Pb.
9PbB = PbAx8.6
h Based on air Pb data for U.S. (1986 PbAQCD) as a surrogate for Chicago.
!PbB = PbAx3.6
J Study that considered air Pb and soil Pb where the air Pb-blood Pb relationship was adjusted for soil Pb.
k Slope provided in paper with background blood Pb level of 1.5 and 3 ug/dL, respectively, and GMR of 2.55 for ambient air.
'PbB = PbAx9.63
GM, geometric mean; GSD, geometric standard deviation; PbB, blood Pb concentration (ug/dL); PbA, air Pb concentration (ug/m3)
November 2012
4-119
Draft - Do Not Cite or Quote
-------
0
10 15
AirPb (|jg/m3)
20
25
Note: The regression model is: (ln[|jg/dL blood Pb] = 0.3485-ln[|jg/m air Pb] + 2.85) for all children (n=96 subject groups) and
(ln[ug/dL blood Pb] = 0.2159-ln[ug/m3air Pb] + 2.62) when the sample was restricted to populations that had blood Pb
concentrations <20 ug/dL (n=44 subject groups).
Data provided from Brunekreef (1984).
Figure 4-22 Predicted relationship between air Pb and blood Pb based on a
meta analysis of 18 studies.
November 2012
4-120
Draft - Do Not Cite or Quote
-------
40
35 -
CO^
0)30
o>
-25
O)
a 20
o>
Q.
O
w 15
.a
CL
E 10
00
5 -
0.0
0.5
1.0
AirPb (|jg/m3)
1.5
ABrunekreef84<20
AHayes94
»Hilts03
oSchwartz89(Chi)
CSchwartz89(US)
DTripathiOl
2.0
Note: Slopes are calculated for a change in air Pb (±0.01 ug/m ) over ranges of air Pb concentrations reported in each study (lines).
The air Pb axis is truncated at 2 ug/m3; the actual range for the Brunkreef et al. (1984) study was 0.1-6.4 ug/dL per ug/m3. The slope
axis has been truncated at 40; the actual range for the Hayes et al. (1994) study was 5-56 ug/dL per ug/m3 (the high end of the
range was estimated for the minimum annual average air Pb of 0.05 ug/m3). The two estimates for Schwartz and Pitcher (1989)
represent data for U.S. and Chicago. Models are log-log (solid lines) and linear (dotted lines). Symbols show the slope at the central
estimate of air Pb (e.g., median for Brunereef and mean for the other studies).
Figure 4-23 Blood Pb - air Pb slopes (ug/dL per ug/m3) predicted from
epidemiologic studies.
1 Hilts et al. (2003) reported child blood Pb and air Pb trends for the city of Trail, British
2 Columbia, over a period preceding and following installation of a new smelter process in
3 1997 which resulted in lower air Pb concentrations. Blood Pb data were obtained from
4 annual (1989-2001) surveys of children 6-60 months of age who lived within 4 km from
5 the smelter (n: 292-536 eligible per year, 75-85% participation). Air Pb concentrations
6 were obtained from high volume suspended particulate samplers placed within 2 km of
7 the smelter that operated 24 hours every 6th day. Data on Pb levels in air, residential soil,
8 interior dust, and blood for three sampling periods are summarized in Table 4-13. Based
9 on these data, blood Pb decreased 6.5 (ig/dL per 1 (ig/m3 air Pb and by 0.068 (ig/dL per
10 mg/kg soil Pb (based on linear regression with air or soil Pb as the sole independent
11 variable) for the entire period. When considering a 9-month weighted mean of
12 0.13 (ig/m3) for 2001 (3 months when the smelter was closed, 0.03 (ig/m3; 6 months
13 when it was open, 0.18 (ig/m3), the slope is 7.0. Several uncertainties apply to these
November 2012
4-121
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
estimates. Potential mismatching of air Pb concentrations (often termed misclassification)
with individual blood Pb levels may have occurred as a result of air Pb being measured
within 2 km of the smelter, whereas, the blood Pb data included children who resided
>2 km from the smelter. The regression estimates were based on group mean estimates
for three sampling dates, rather than on the individual blood Pb estimates, which included
repeated measures on an unreported fraction of the sample. The limited number of data
pairs (three) constrained parameter estimates to simple regression coefficients. Other
important factors probably contributed to blood Pb declines in this population that may
have been correlated with air, soil and dust Pb levels. These factors include aggressive
public education and exposure intervention programs (Hilts et al.. 1998; Hilts. 1996).
Therefore, the coefficients shown in Table 4-13 are likely to overestimate the influence of
air, dust, or soil Pb on blood Pb concentrations at this site.
Table 4-13 Environmental Pb levels and blood Pb levels in children in Trail,
British Columbia.
Date
Blood Pb (ug/dL)
Air Pb (|jg/m3)
Soil Pb (mg/kg)
Interior Dust Pb (mg/kg)
1996 a
11.5
1.1
844
758
1999
5.9
0.3
756
583
2001
4.7
0.13b
750°
580°
Regression Coefficient
NA
7.01 ± 0.009 (R2=1.00, p=0.001)
0.069 ± 0.008 (R2=0.99, p=0.069)
0.035 ± 0.005 (R2=0.98, p=0.097)
A new smelter process began operation in 1997. Values for air, soil and dust Pb are annual geometric means; values for blood Pb
are annual geometric means. Regression coefficients are for simple linear regression of each exposure variable on blood Pb.
a Values for air Pb, soil Pb, and interior dust Pb are actually for period of 1994-1996.
bNine month time-weighted average of 0.03 ug Pb/m3for3 months and 0.18 ug Pb/m3for6 months.
0 Values assumed by study authors.
Source: Data from Hilts et al. (2003).
13
14
15
16
17
18
19
20
21
22
23
24
Ranft et al. (2008) reported a meta-analysis of five cross-sectional surveys of air and soil
Pb levels and blood Pb concentrations in children living in Duisburg, Germany. The
analysis included observations on 843 children (6-11 years of age) made during the
period 1983-2000. Children recruited in 1983 were an average of 9.1 yrs of age, whereas
children recruited in later years of the study averaged 6.3 to 6.4 yrs of age. The 1983 air
Pb concentrations were based on two monitoring stations, while a combination of
dispersion modeling and monitoring data was used in the later years to estimate Pb in PM
in a 200 meter by 200 meter grid that encompassed the city. Pb in surface soil (0-10 cm)
was measured at 145 locations in the city in 2000 and 2001. Air and soil Pb
concentrations were assigned to each participant by spatial interpolation from the
sampling grid data to each home residence. The 5th-95th percentile ranges were
0.025-0.465 (ig Pb/m3 for air and 72-877 mg Pb/kg for soil. The results of multivariate
November 2012
4-122
Draft - Do Not Cite or Quote
-------
1 regression analyses were reported in terms of the relative increase (the geometric mean
2 blood Pb ratio, GMR) for an increase in air or soil Pb from the 5th to 95th percentile
3 value. In a multivariate regression model (equation not provided) that included air and
4 soil Pb in the same model and adjusted for covariates, the GMR values were: 2.55 per
5 0.44 (ig/m3 increase in air Pb (95% CI: 2.40, 2.71, R2=0.484, p <0.001) and 1.30 per
6 800 mg/kg soil Pb (95% CI: 1.19, 1.43, R2 = 0.017, p <0.001). Based on the values for R2,
7 the regression model accounted for approximately 59% of the total variance in blood Pb
8 and, of this, 83% was attributed to air Pb. Values for GMR for soil Pb ranged from 1.41
9 to 2.89, with most recent blood Pb data (from the year 2000) yielding a value of 1.63 per
10 800 mg/kg increase in soil Pb. The GMR values can be converted to regression slopes
11 (slope = [starting blood Pbxln(GMR)]/[95th - 5th percentile air or soil Pb]) for
12 calculating equivalent airblood Pb ratios. The model predicts an increase of 3.2 (ig/dL
13 blood Pb per 1 (ig/m3 increase in air Pb at the median air Pb concentration for the study
14 (0.1 (ig/m3) and assuming a background blood Pb concentration of 1.5 (ig/dL. Based on
15 the GMR estimate of 1.63 for soil Pb, a 1,000 mg/kg increase in soil Pb would be
16 associated with an increase in blood Pb of 0.9 (ig/dL per mg/kg soil at the median soil Pb
17 concentration of 206 mg/kg and assuming a background blood Pb concentration of
18 1.5 (ig/dL. The degree of confounding of the GMR and estimates resulting from the air
19 and soil Pb correlation was not reported, although the correlation coefficient for the two
20 variables was 0.136 for the whole data set and 0.703 when data collected in 1983 was
21 omitted. Because the model also included Pb levels in soil, the blood Pb-air Pb ratio may
22 be underestimated since some of the Pb in soil was likely derived from air. The blood
23 Pb-air Pb slope does not include the portion of the soil/dust Pb ingestion pathway that
24 derives from air Pb, such as recently airborne Pb deposited to soil and dust which remains
25 available for inhalation and ingestion.
26 To estimate the blood Pb-air Pb ratio that included all air-related pathways, data for
27 median of blood Pb and air Pb among the cohort of children studied were extracted from
28 Table 2 in Ranft et al. (2008) for each of the five study years. The median blood Pb and
29 air Pb were used in regressions employing linear, log-log, and log-linear (i.e., similar to
30 authors' approach with ln[blood Pb] against air Pb) fits. The linear model obtained was:
31 PbB = 12.2>
-------
1 Schnaas et al. (2004) analyzed data on blood Pb and air Pb concentrations during and
2 after the phase out of leaded gasoline use in Mexico (1986-1997) in children as part of a
3 prospective study conducted in Mexico City. The sample included 321 children born
4 during the period 1987 through 1992. Repeated blood Pb measurements were made on
5 each child at 6-month intervals up to age 10 years. Air Pb measurements (annual average
6 of quarterly means) were derived from three area monitors which represented distinct
7 study zones. Children were assigned to study zones based on their current address.
8 Associations between lifetime (across the first 10 years of life) blood Pb concentration,
9 air Pb concentration (mean annual for each calendar year of study) and other variables
10 (e-g-, age, year of birth, family use of glazed pottery) were evaluated using multivariate
11 regression models. The largest air Pb coefficient occurred in the cohort born in 1987, who
12 experienced the largest decline in air Pb (from 2.8 to about 0.25 (ig/m3); the air Pb
13 coefficient for this group of children was 0.213 (95% CI: 0.114-0.312) In [(ig/dL blood]
14 per ln[(ig/m3 air]. The smallest, statistically significant air Pb coefficient occurred for the
15 1990 birth year cohort, who experienced a decline in air Pb from 1.5 to about 0.1 (ig/m3.
16 The air Pb coefficient for the 1990 cohort was 0.116 (95% CI: 0.035-0.196). Based on
17 these air Pb coefficients, children in the 1987 and 1990 cohorts were estimated to have
18 24% and 12% decreases in lifetime (across the first 10 years of life) blood Pb levels,
19 respectively, per natural log decrease in air Pb. Table 4-14 provides predicted blood Pb
20 and blood Pb-air Pb slopes as a function of age for the 1987 and 1990 cohorts. The values
21 in Table 4-14 are for children having complete datasets that lived in the Merced (study
22 region having medium air Pb concentrations) of Mexico City in medium SES families
23 (for the study population) that did not use clay pottery. Higher estimated blood Pb and
24 blood Pb-air Pb slopes than those in Table 4-14. would be predicted for low SES families
25 living in Xalostoc (study region having highest air Pb) that use clay pottery (e.g., 2-yr-
26 olds predicted blood Pb of 11-14 ug/dL and blood Pb-air Pb slope of 5.1-12 ug/dL
27 per ug/m3). Conversely, lower estimates would be predicted for high SES families living
28 in Pedregal (study region having lowest air Pb) that did not use clay pottery (e.g., 2-yr-
29 olds predicted blood Pb of 7-9 ug/dL and blood Pb-air Pb slope of 3.8-4.8 ug/dL
30 per ug/m3). The effect of air Pb on blood Pb may have been underestimated in this study
31 due to inclusion of location and SES terms in the regression model. It was specifically
32 noted by the authors that air Pb differed significantly between the locations and the
33 poorer residential areas were usually the more industrialized areas with higher pollution.
34 Hence, the inclusion of these terms may have accounted for some of the variance in blood
35 Pb attributable to air Pb.
November 2012 4-124 Draft - Do Not Cite or Quote
-------
Table 4-14 Predicted blood Pb levels and blood-air slopes for Mexico City
children (1987 and 1990 cohorts).
Age (in years)
1
2
3
4
5
6
7
8
9
10
Blood Pb
(|ig/dL)
7.4-
8.9-
8.4-
7.9-
7.7-
6.9-
6.8-
6.1 -
5.8-
5.6-
8.5 a
10.2
9.7
9.1
8.8
7.9
7.8
7.1
6.7
6.5
Blood Pb-Air Pb Slope (pg/dL
per |jg/m3)
2.1 -4.5a
2.6-
2.4-
2.3-
2.2-
2.0-
2.0-
1.8-
1.7-
1.6-
5.5
5.2
4.9
4.7
4.2
4.2
3.8
3.6
3.5
'Values are for 1990 and 1987 cohorts, respectively, at an air Pb concentration of 0.4 ug/m3 which is the median and geometric mean of the
annual air Pb concentrations over the course of the study based on data in Figure 1 of Schnaas et al. (2004)
Source: Based on Table 4 of Schnaas et al. (2004)
1 For an approach that considers all the Schnaas et al. (2004) cohorts simultaneously, data
2 for annual geometric mean of blood Pb and air Pb were extracted from Figure 1 in
3 Schnaas et al. (2004). However, in employing this approach, blood Pb is confounded by
4 age and year because in the early years of the study, only younger children were available
5 and in the later years of the study, only older children contributed data. The extracted
6 values of the geometric mean of blood Pb and mean air Pb were used in regressions
7 employing linear and log-log models for comparison to other studies. The linear model
8 obtained was: PbB = 2.50xPbA + 5.61 (R2 = 0.84), i.e., the linear model produced a
9 constant slope of 2.50 (ig/dL per (ig/m3. However, inspection of the graph (not shown
10 here) suggested a bi-linear fit. Regression of the data over the interval 0.1-0.4 (ig/m3
11 produced a slope of 9.0 (ig/dL per (ig/m3 (R2 = 0.83), and regression of the data over the
12 interval 0.4-2.8 (ig/m3 produced a slope of 1.52 (ig/dL per (ig/m3 (R2 = 0.83). The log-log
13 model was: In(PbB) = 0.26xm(PbA) + 2.20 (R2 = 0.94), resulting in an inverse curve for
14 dPbB/dPb, versus PbA, with a slope of 4.5 (ig/dL per (ig/m3 at PbA = 0.4 (ig/m3.
15 Schwartz and Pitcher (1989) reported a multivariate regression analysis of associations
16 between U.S. gasoline Pb consumption (i.e., sales) and blood Pb concentrations in the
17 U.S. population during the period 1976-1980 when use of Pb in gasoline was being
18 phased out. Although this analysis did not directly derive a slope for the air Pb-blood Pb
19 relationships, other analyses have shown a strong correlation between U.S. gasoline Pb
20 consumption and ambient air Pb levels during this same period (U.S. EPA. 1986a).
21 Therefore, it is possible to infer an air Pb-blood Pb relationship from these data. Two
November 2012 4-125 Draft - Do Not Cite or Quote
-------
1 sources of blood Pb data were used in Schwartz and Pitcher (1989): NHANES II
2 provided measurements for U.S. individuals 6 months to 74 years of age (n = 9,987
3 subjects) between February 1976 and February 1980, and the Chicago blood Pb screening
4 program provided measurements in black children aged birth to 5 years (n = 5476
5 subjects) for the period from 1976 to mid-1980. Observed blood Pb levels were not
6 provided. Gasoline Pb consumption for the U.S. was estimated as the product of monthly
7 gasoline sales and quarterly estimates of Pb concentrations in gasoline reported to U.S.
8 EPA. Based on the NHANES blood Pb data for whites, the regression coefficient for
9 blood Pb on the previous month's gasoline Pb usage, adjusted for age, race, sex, income,
10 degree of urbanization, nutrient intake, smoking, alcohol consumption, occupational
11 exposure, and other significant covariates was 2.14 (ig/dL blood per 100 metric tons of
12 gasoline Pb/day (SE=0.19, p=0.0000); the authors reported that the results for blacks
13 were essentially identical. Based on the Chicago blood Pb data, the age-adjusted
14 regression coefficient was 16.12 ((ig/dL per 1,000 metric tons gasoline Pb/quarter
15 [SE=1.37, p=0.0001]). When the coefficient was scaled by the ratio of Chicago's
16 gasoline use to the nation's and converted to units of 100 metric tons per day, the
17 gasoline Pb coefficient was 1.97 (ig/dL blood per 100 metric tons of gasoline Pb/day),
18 which is similar to the coefficient reported for the NHANES cohort. U.S. EPA (1986a)
19 reported data on gasoline Pb consumption (sales) and ambient Pb levels in the U.S.
20 during the period 1976-1984 (Table 4-15). Based on these data, air Pb concentrations
21 decreased in association with gasoline Pb consumption. The linear regression coefficient
22 for the air Pb decrease was 0.23 (ig/m3 per 100 metric tons gasoline Pb/day (SE = 0.02,
23 R2~0.95, p <0.0001). If this regression coefficient is used to convert the blood Pb slopes
24 from Schwartz and Pitcher (1989). the corresponding air Pb-blood Pb slopes would be
25 9.3 and 8.6 (ig/dL per (ig/m3, based on the NHANES and Chicago data, respectively
26 (e.g., 2.14/0.23 = 9.3 and 1.97/0.23=8.6).
November 2012 4-126 Draft - Do Not Cite or Quote
-------
Table
Date
1976
1977
1978
1979
1980
1981
1982
1983
1984
4-15 U.S. gasoline Pb consumption and air Pb levels.
Total Gasoline Pb
(103 metric tons/year)
171.4
168.9
153.0
129.4
78.8
60.7
59.9
52.3
46.0
Total Gasoline Pb
(102 metric tons/day)3
4.70
4.63
4.19
3.53
2.16
1.66
1.64
1.43
1.26
The linear regression coefficient is 0.23 |jg/m3air per 100 metric tons/day (SE= 0.020, R2 = 0.95, p
Conversion factor is 10/365 days/year.
bAnnual mean of per site maximum quarterly means (1 984 Trend Report (Available online:
httD://www.eDa.aov/air/airtrends/Ddfs/Trends Report 1984.pdf
AirPb
(M9/m3)b
1.22
1.20
1.13
0.74
0.66
0.51
0.53
0.40
0.36
O.0001)
Source: Table 5-5, U.S. EPA 1986 Pb AQCD Q986a).
1 Tripathi et al. (2001) reported child blood Pb and air Pb for the city and suburbs of
2 Mumbai, India over the period 1984-1996. Pb-free petroleum was introduced in India
3 beginning in late 1996, which was outside the period of this study. Blood Pb data were
4 obtained from children 6-10 years of age (n = 544) who lived in 13 locations within the
5 Mumbai area. Air Pb concentrations were measured from high volume PM samplers
6 (with the majority of Pb in the respirable size range) placed at a height of 1.6 meters that
7 operated 24 hours. Data on Pb concentrations in air and blood are summarized in Table
8 4-16. An additional 16 children from two regions of Mumbai were excluded from the
9 analysis because of their high blood Pb levels (geometric means: 69.2 and 20.8 (ig/dL)
10 and proximity to industrial Pb sources with high air Pb concentrations (geometric means:
11 41.2 and 6.7 (ig/m3). Based on the data from residential locations presented in Table 4-16.
12 blood Pb increased 3.6 (ig/dL per 1 (ig/m3 air Pb (based on linear regression with air Pb
13 as the sole independent variable). Several uncertainties apply to these estimates,
14 including potential exposure misclassification since the mean air Pb concentration was
15 used for each suburb over the entire study period. In addition, the regression estimates
16 were based on group mean blood Pb estimates for the 13 sampling locations, rather than
17 on the individual blood Pb estimates. Ingestion of Pb-containing food was estimated in
18 this study, but was not considered in the regression equation for estimating blood Pb,
19 despite the author's conclusion that the ingestion route is important for the intake of Pb
20 by children in Mumbai.
November 2012 4-127 Draft - Do Not Cite or Quote
-------
Table 4-16 Air Pb concentrations and blood Pb levels in children in Mumbai,
India.
Blood Pb (Hg/dL)
Location
Borivilli
Byculla
Deonar
Goregaon
Govandi
Jogeshwari
Khar
Parel
Sion
Thans (SS)
Vile Parle
Colaba
Vakola
N
12
117
46
21
20
20
17
168
34
37
19
12
21
GM
10.4
11.0
9.5
9.1
8.9
8.6
9.0
10.4
9.6
12.0
9.1
9.2
14.4
GSD
1.67
1.99
2.29
1.30
1.42
1.32
1.53
1.91
1.49
1.86
1.46
1.86
1.64
N
10
30
93
24
10
24
22
37
96
4
7
9
7
Air Pb (|jg/m3)
GM
0.32
0.99
0.11
0.35
0.10
0.11
0.18
0.44
0.39
1.18
0.37
0.14
1.12
GSD
1.51
1.73
3.21
1.77
1.52
2.47
3.15
1.48
1.75
1.04
1.34
1.63
1.12
The linear regression coefficient is 3.62 ug/dL blood per ug/m3 air (SE= 0.61, R2= 0.76, p <0.001).
GM, geometric mean; GSD, geometric standard deviation; N, number of subjects.
Source: Data are from Tripathi et al. (2001).
1 Hayes et al. (1994) analyzed data collected as part of the Chicago, IL blood Pb screening
2 program for the period 1974-1988, following the phase-out of leaded gasoline. The data
3 included 9,604 blood Pb measurements in children (age: 6 months to 5 years) and
4 quarterly average air Pb concentrations measured at 12 monitoring stations in Cook
5 County, IL. Annual median blood Pb levels declined from 30 ug/dL in 1968 to 12 ug/dL
6 in 1988. During most of the years of the study, blood Pb measurements at or below
7 10 ug/dL were recorded as 10 ug/dL because of concerns over measurement accuracy of
8 the instrument below these levels. Quarterly median blood Pb levels declined in
9 association with quarterly mean air Pb concentrations. The regression model predicted a
10 slope of 0.24 In [ug/dL blood] per ln[ug/m3 air], as illustrated in Figure 4-24. This slope
11 corresponds to an increase of 8.2 ug/dL blood Pb per ug/m3 at the average annual mean
12 air Pb concentration of 0.62 ug/m3. As shown in Figure 4-25. with decreasing air Pb
13 concentration, the slope increases. The study reports a slope of 5.6 associated with
14 ambient air Pb levels near 1 ug/m3 and a slope of 16 for ambient air Pb levels in the range
15 of 0.25 ug/m3, indicating a pattern of higher ratios with lower ambient air Pb and blood
16 Pb levels.
November 2012 4-128 Draft - Do Not Cite or Quote
-------
30
25 -
20 -
15 -
O)
_Q
Q_
T3
O
= 10 H
5 -
0
0.0 0.2 0.4
0.6 0.8 1.0
AirPb (|jg/m3)
1.4 1.6
Note: The regression model is: (ln[|jg/dL blood Pb] = 0.24-ln[|jg/m air Pb] + 3.17).
Modified from Hayes et al. (1994).
Figure 4-24 Predicted relationship between air Pb and blood Pb based on data
from Chicago, IL in children age 0-5 years (1974-1988).
1 The evidence on the quantitative relationship between air Pb and blood Pb is now, as in
2 the past, limited by the circumstances in which the data are collected. These estimates are
3 generally developed from studies of populations in various Pb exposure circumstances.
4 The 1986 Pb AQCD (U.S. EPA. 1986a) discussed the studies available at that time that
5 addressed the relationship between air Pb and blood Pb, recognizing that there is
6 significant variability in air-to-blood ratios for different populations exposed to Pb
7 through different air-related exposure pathways and at different exposure levels. The
8 1986 Pb AQCD noted that ratios derived from studies involving higher blood and air Pb
9 levels are generally smaller than ratios from studies involving lower blood and air Pb
10 levels [see the 1986 Pb AQCD, Chapter 11, pp 99 (U.S. EPA. 1986a)1. In consideration
11 of this factor, slopes in the range of 3 to 5 for children generally reflected study
12 populations with blood Pb levels in the range of approximately 10-30 ug/dL [see
13 Chapter 11, pp 100 of the 1986 Pb AQCD, Table 11-36, from (Brunekreef. 1984)1. much
14 higher than those common in today's population. The slope of 3.6 from Tripathi et al.
15 (2001) is consistent with this observation, given that the blood Pb levels were at the lower
16 end of this range (i.e., 10-15 ug/dL).
November 2012
4-129
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
There are fewer studies that evaluate the air Pb-blood Pb relationship in conditions that
are more reflective of the current state. Hilts et al. (2003) is one such study that provides
insight because the blood Pb and air Pb levels were relatively lower than those studies
mentioned above; the slope reported was 6.5, but could be as high as 7.0. Similarly,
Hayes et al. (1994) demonstrates greater slopes observed with decreasing air Pb
concentrations. These studies provide evidence that air-to-blood slopes relevant for
today's population of children would likely extend higher than the 3 to 5 range identified
in the 1986 Pb AQCD (U.S. EPA. 1986a). In the 2008 final rule for the Pb NAAQS (73
FR 66964), with recognition of uncertainty and variability in the absolute value of an air-
to-blood relationship, the air-to-blood slopes of 5, 7, and 10 (ig/dL per (ig/m3 were
utilized in evaluating air-related IQ loss of children. Figure 4-25 illustrates the impact of
these air-to-blood slopes on the estimated change in air-related blood Pb as a function of
change in air Pb.
M
.Q
Q.
~O
O
_o
CO
0)
M
re
1.5 H
1 -
0.5 •
Slope
(ng/dLperng/m3)
•10
7
5
0
0.05 0.1
Change in Air Pb (u,g/m3)
0.15
14
Figure 4-25 Effect of air-to-blood slope on estimated change in air-related
blood Pb with change in air Pb.
November 2012
4-130
Draft - Do Not Cite or Quote
-------
4.5.2 Air Pb-Blood Pb Relationships in Occupational Cohorts
1 At the time of the 1986 Pb AQCD, there was a great deal of information on blood Pb
2 responses to air Pb exposures of workers in Pb-related occupations (U.S. EPA, 1986a).
3 Almost all such exposures were at air Pb exposures far in excess of typical non-
4 occupational exposures and typically did not account for other potential sources of Pb
5 exposure. The air Pb-blood Pb slopes in these studies were generally much less
6 (i.e., 0.03-0.2; pg 11-106) than those observed in children when considering aggregate air
7 Pb contributions (i.e., 3-5; pg 11-106). In addition, the air Pb concentrations in
8 occupational studies are typically collected at much shorter durations (e.g., over an 8-hr
9 workday) compared to ambient Pb monitoring, making it difficult to draw comparisons
10 between occupationally and non-occupationally exposed populations. Therefore, only a
11 few occupational studies are presented below to demonstrate that more recent air Pb and
12 blood Pb levels remain much higher in these studies compared to those conducted in the
13 general population.
14 Rodrigues et al. (2010) examined factors contributing to variability in blood Pb
15 concentration in New England bridge painters, who regularly use electric grinders to
16 prepare surfaces for painting. The study included 84 adults (83 males, 1 female) who
17 were observed during a 2-week period in 1994 or 1995. The geometric mean air Pb
18 concentration obtained from personal PM samplers worn over the workday was 58 (ig/m3
19 (GSD 2.8), with a maximum daily value of 210 (ig/m3. Hand wipe samples were
20 collected and analyzed for Pb (GM = 793 jig, GSD 3.7). Blood Pb samples were
21 collected at the beginning of the 2-week period (GM =16.1 (ig/dL, GSD 1.7) and at the
22 end of the period (GM=18.2 (ig/dL, GD=1.6). Associations between exposure variables
23 and blood Pb concentrations were explored with multivariate regression models. When
24 the model excluded hand-wipe data, the regression coefficient for the relationship
25 between ln[blood Pb concentration (ng/dL)] and ln[air Pb ((ig/m3)] was 0.11 (SE = 0.05,
26 p = 0.03). This corresponds to a slope of 0.009 (ig/dL per (ig/m3 at the geometric mean
27 air Pb concentration for the study. A second regression model included hand wipe Pb
28 (n = 54) and yielded a regression coefficient of 0.05 (SE = 0.07, p = 0.45), which
29 corresponds to a slope of 0.02 (ig/dL per (ig/m3 at the geometric mean air Pb
30 concentration for the study.
31 Two other studies that examined the air Pb-blood Pb relationship in occupational settings
32 at higher air Pb concentrations (geometric mean of 82 and 111 (ig/m3) for Pb battery and
33 crystal workers, respectively (Pierre et al., 2002; Lai etal. 1997). Blood Pb levels for the
34 Pb battery workers averaged 56.9 (ig/dL (SD 25.3) and for the crystal workers was
35 21.9 (ig/dL. Both studies employed log-log regression models, resulting in slopes of 0.49
36 (Pierre et al.. 2002) and 0.08 (Laietal.. 1997V
November 2012 4-131 Draft - Do Not Cite or Quote
-------
4.5.3 Environmental Pb-Blood Pb Relationships
1 Empirically-based relationships between blood Pb levels and Pb intakes and/or Pb
2 concentrations in environmental media have provided the basis for what has become
3 known as slope factor models. Slope factor models are highly simplified representations
4 of empirically based regression models in which the slope parameter represents the
5 change in blood Pb concentration projected to occur in association with a change in Pb
6 intake or uptake. The slope parameter is factored by exposure parameters (e.g., exposure
7 concentrations, environmental media intake rates) that relate exposure to blood Pb
8 concentration (Maddaloni etal., 2005; U.S. EPA. 2003c; Abadin and Wheeler. 1997;
9 Stern. 1996; Bowers et al.. 1994; Stern. 1994; Carlisle and Wade. 1992). In slope factor
10 models, Pb biokinetics are represented as a linear function between the blood Pb
11 concentration and either Pb uptake (uptake slope factor, USF) or Pb intake (intake slope
12 factor, ISF). The models take the general mathematical forms:
PbB = E x ISF
Equation 4-2
PbB = E x AF x USF
Equation 4-3
13 where PbB is the blood Pb concentration, E is an expression for exposure (e.g., soil
14 intake x soil Pb concentration) and AF is the absorption fraction for Pb in the specific
15 exposure medium of interest. Intake slope factors are based on ingested rather than
16 absorbed Pb and, therefore, integrate both absorption and biokinetics into a single slope
17 factor, whereas models that utilize an uptake slope factor include a separate absorption
18 parameter. In contrast to mechanistic models, slope factor models predict quasi-steady
19 state blood Pb concentrations that correspond to time-averaged daily Pb intakes (or
20 uptakes) that occur over sufficiently long periods to produce a quasi-steady state
21 (i.e., >75 days, ~3 times the ti/2 for elimination of Pb in blood).
November 2012 4-132 Draft - Do Not Cite or Quote
-------
1 The U.S. EPA Adult Lead Methodology (ALM) is an example of a slope factor model
2 that has had extensive regulatory use in the EPA Superfund program for assessing health
3 risks to adults associated with non-residential exposures to Pb in contaminated soils
4 (Maddaloni et al.. 2005; U.S. EPA. 1996a). The model was developed to predict maternal
5 and fetal blood Pb concentrations that might occur in relation to maternal exposures to
6 contaminated soils. The model assumes an uptake slope factor of 0.4 (ig/dL blood
7 per (ig/day Pb uptake. Additional discussion of slope factor models that have been used
8 or proposed for regulatory use can be found in the 2006 Pb AQCD (U.S. EPA. 2006b).
9 Previous studies included in the 2006 Pb AQCD (U.S. EPA. 2006b) explored the
10 relationship between blood Pb in children and environmental Pb concentrations. In a
11 pooled analysis of 12 epidemiologic studies, interior dust Pb loading, exterior soil/dust
12 Pb, age, mouthing behavior, and race were all statistically significant variables included
13 in the regression model for blood Pb concentration (Lanphear et al., 1998). Significant
14 interactions were found for age and dust Pb loading, mouthing behavior and exterior
15 soil/dust level, and SES and water Pb level. In a meta-analysis of 11 epidemiologic
16 studies, among children the most common exposure pathway influencing blood Pb
17 concentration in structural equation modeling was exterior soil, operating through its
18 effect on interior dust Pb and hand Pb (Succop et al.. 1998). Similar to Lanphear et al.
19 (1998). in the linear regression model, interior dust Pb loading had the strongest
20 relationships with blood Pb concentration. Individual studies conducted in Rochester,
21 NY, Cincinnati, OH, and Baltimore, MD report similar relationships between children's
22 blood Pb and interior dust concentrations (Lanphear and Roghmann. 1997; U.S. EPA.
23 1996b: Bornschein et al.. 1985).
24 Dixon et al. (2009) reported a multivariate analysis of associations between
25 environmental Pb concentrations and blood Pb concentrations, based on data collected in
26 the NHANES (1999-2004). The analyses included 2,155 children, age 12-60 months. The
27 population-weighted geometric mean blood Pb concentration was 2.03 (ig/dL
28 (GSD 1.03). A linear model applied to these data yielded an R2 of 40% (Table 4-17). The
29 regression coefficient for the relationship between ln[blood Pb concentration ((ig/dL)]
30 and ln[floor dust Pb concentration ((ig/ft2)] was 0.386 (SE 0.089) for "not smooth and
31 cleanable" surfaces (e.g., high-pile carpets) and 0.205 (SE 0.032) for "smooth and
32 cleanable" surfaces (e.g., uncarpeted or low-pile carpets). These coefficients correspond
33 to a 2.4-fold or 1.6-fold increase in blood Pb concentration, respectively, for a 10-fold
34 increase in floor dust Pb concentration.
November 2012 4-133 Draft - Do Not Cite or Quote
-------
Table 4-17 Linear model relating environmental Pb
concentration in children.
Variables Overall p-value
Intercept 0.172
Age (in years) <0.001
Year of construction 0.014
PIR <0.001
Race/ethnicity <0.001
Country of birth 0.002
Floor surface/condition x log floor <0.001
PbD
Floor surface/condition
x (log floor PbD)2
Floor surface/condition
x (log floor PbD)3
Log windowsill PbD 0.002
Home-apartment type <0.001
Anyone smoke inside the home 0.015
Log cotinine concentration (ng/dL) 0.004
Window, cabinet, or wall renovation 0.045
in a pre-1 978 home
Levels3
Age
Age2
Age3
Age4
Intercept for missing
1990-present
1978-1989
1960-1977
1950-1959
1940-1949
Before 1940
Intercept for missing
Slope
Non-Hispanic white
Non-Hispanic black
Hispanic
Other
Missing
U.S."
Mexico
Elsewhere
Intercept for missing
Not smooth and cleanable
Smooth and cleanable or carpeted
Not smooth and cleanable
Smooth and cleanable or carpeted
Uncarpeted not smooth and
cleanable
Smooth and cleanable or carpeted
Intercept for missing
Slope
Intercept for missing
Mobile home or trailer
One family house, detached
One family house, attached
Apartment (1-9 units)
Apartment (> 10 units)
Missing
Yes
No
Intercept for missing
Slope
Missing
Yes
No
exposure and blood Pb
Estimate (SE)
-0.517(0.373)
2.620 (0.628)
-1.353(0.354)
0.273 (0.083)
-0.019 (0.007)
-0.121 (0.052)
-0.198(0.058)
-0.196(0.060)
-0.174(0.056)
-0.207 (0.065)
-0.012(0.072)
0.000
0.053 (0.065
-0.053(0.012)
0.000
0.247 (0.035
-0.035 (0.030)
0.128(0.070)
-0.077 (0.219)
0.000
0.353 (0.097)
0.154(0.121
0.178(0.094)
0.386 (0.089)
0.205 (0.032)
0.023(0.015)
0.027 (0.008)
-0.020 (0.014)
-0.009 (0.004)
0.053 (0.040
0.041 (0.011
-0.064 (0.097
0.127(0.067)
-0.025 (0.046)
0.000
0.069 (0.060)
-0.133(0.056)
0.138(0.140)
0.100(0.040)
0.000
-0.150(0.063)
0.039(0.012)
-0.008(0.061)
0.097 (0.047)
0.000
p-Value
0.172
<0.001
<0.001
0.002
0.008
0.024
0.001
0.002
0.003
0.003
0.870
0.420
<0.001
<0.001
0.251
0.073
0.728
<0.001
0.209
0.065
<0.001
<0.001
0.124
0.001
0.159
0.012
0.186
<0.001
0.511
0.066
0.596
0.256
0.022
0.331
0.015
0.023
0.002
0.896
0.045
'Children: n = 2,155 (age 10-60 months); R2 = 40%
'includes the 50 states and the District of Columbia
Source: Dixon et al. (20091.
November 2012
4-134
Draft - Do Not Cite or Quote
-------
1 Mielke et al. (2007a) analyzed blood Pb and soil Pb concentration data collected as part
2 of a blood Pb screening program in New Orleans, Louisiana (2000-2005). The data set
3 included 55,551 blood Pb measurements for children 0-6 years of age and 5,467 soil Pb
4 measurements. Blood Pb and soil Pb concentrations were matched at the level of census
5 tracts. The association between blood Pb concentration and soil Pb concentration was
6 evaluated using non-parametric permutation methods. The resulting best-fit model was:
PbB = 2.038 +(0.172 xPbS05)
Equation 4-4
7 where PbB is the median blood Pb concentration and PbS is the median soil Pb
8 concentration. Although the overall association between blood Pb and soil Pb was strong
9 (R2=0.528), there was considerable scatter in the data. For example, at the median soil Pb
10 levels of 100 and 500 mg/kg, median blood Pb ranged from 2 to 8 (ig/dL and 3 to
11 12 (ig/dL, respectively. The resulting curvilinear relationship predicts a twofold increase
12 in blood Pb concentration for an increase in soil Pb concentration from 100 to 1,000 ppm
13 (Figure 4-26).
November 2012 4-135 Draft - Do Not Cite or Quote
-------
O)
.Q
CL
•a
o
_o
m
Blood=2.038+0.172xSoil°-5
400 800 1200
SoilPb(ppm)
1600
2000
Note: The data set included 55,551 blood Pb measurements for children 0-6 years of age and 5,467 soil Pb measurements. Blood
Pb and soil Pb concentrations were matched at the level of census tracts (Mielke et al.. 2007a).
Figure 4-26 Predicted relationship between soil Pb concentration and blood
Pb concentration in children based on data collected in
New Orleans, Louisiana: 2000-2005.
1 In a subsequent re-analysis of the New Orleans (2000-2005) data, individual child blood
2 Pb observations were matched to census tract soil concentrations (Zahran etal. 2011).
3 This analysis confirmed the association between blood Pb and both soil Pb and age
4 reported in Mielke et al. (2007a). Regression coefficients for soil Pb (random effects
5 generalized least squares regression) ranged from 0.217 to 0.214 (per soil Pb05), which is
6 equivalent to approximately a 2-fold increase in blood Pb concentration for an increase in
7 soil Pb concentration from 100 to 1,000 ppm.
8 Several studies have linked elevated blood Pb levels to residential soil exposures for
9 populations living nearby industrial or mining facilities. Gulson et al. (2009) studied the
10 blood Pb and isotopic Pb ratios of children younger than 5-years old and adults older than
11 18-years old living in the vicinity of a mine producing Magellan Pb ore in western
12 Australia. They observed a median blood Pb level of 6.6 (ig/dL for the children, with
13 isotopic ratios indicating contributions from the mine ranging from 27 to 93%. A weak
14 but significant linear association between blood Pb level and percent Magellan Pb was
15 observed (R2 = 0.12, p = 0.018). Among children with blood Pb levels over 9 (ig/dL and
16 among adults, the isotopic ratios revealed Pb exposures from a variety of sources.
17 Garavan et al. (2008) measured soil Pb and blood Pb levels among children aged 1-month
18 to 17.7-years old in an Irish town near a coal mine. The blood Pb measurements were
November 2012
4-136
Draft - Do Not Cite or Quote
-------
1 instituted as part of a screening and community education program given that the
2 presence of Pb had been documented in the environment. Garavan et al. (2008) found that
3 over 3 years of the screening period, median blood Pb levels reduced by roughly 22%
4 from 2.7 to 2.1 ng/dL.
5 An extensive discussion of the relationships between environmental Pb levels and blood
6 Pb concentrations in children at the Bunker Hill Superfund Site location, a former Pb
7 mining and smelting site, was provided in the 2006 Pb AQCD (U.S. EPA. 2006c). In the
8 most recent analysis (TerraGraphics Environmental Engineering. 2004) of the data on
9 environmental Pb levels and child blood Pb concentrations (1988-2002), blood Pb
10 concentrations (annual GM) ranged from 2.6 to 9.9 (ig/dL. Environmental Pb levels
11 (e-g-, dust, soil, paint Pb levels) data were collected at -3,000 residences, with interior
12 dust Pb concentrations (annual GM) ranging from -400 to 4,200 mg/kg and yard soil Pb
13 concentration (annual GM) ranging from -150 to 2,300 mg/kg. Several multivariate
14 regression models relating environmental Pb levels and blood Pb concentration were
15 explored; the model having the highest R2 (0.26) is shown in Table 4-18. The model
16 predicts significant associations between blood Pb concentration, age, interior dust, yard
17 soil, neighborhood soil (geometric mean soil Pb concentration for areas within 200 ft of
18 the residence), and community soil Pb concentration (community GM). Based on the
19 standardized regression coefficients, the community soil Pb concentration had the largest
20 effect on blood Pb concentration, followed by neighborhood soil Pb concentration,
21 interior dust Pb concentration, and yard soil Pb concentration (Table 4-18). The model
22 predicted a 1.8 (ig/dL decrease in blood Pb concentration in association with a decrease
23 in community soil Pb concentration from 2,000 to 1,000 mg/kg. The same decrease in
24 neighborhood soil Pb concentration, interior dust Pb concentration, or yard soil Pb
25 concentration was predicted to result in a 0.8, 0.5, or 0.2 (ig/dL decrease in blood Pb
26 concentration, respectively. Note that the soil Pb component of the model was similar to
27 that derived by Lewin et al. (1999). in which a model of blood Pb as a function of soil Pb
28 among 0-6 year old children living near one of four industrial sites was given as
29 PbB = 0.24381n(PbS} + 0.2758.
November 2012 4-137 Draft - Do Not Cite or Quote
-------
Table 4-18 General linear model relating blood Pb concentration in children and
environmental Pb levels—Bunker Hill Superfund Site.
Parameter
Intercept
Age (yr)
ln(interiordust Pb); (mg/kg)
ln(yard soil Pb); (mg/kg)
GM soil Pb within 200 ft of residence (mg/kg)
GM community soil Pb (mg/kg)
Coefficient
-0.1801
-0.4075
0.7288
0.2555
0.0008
0.0018
P-value
0.7916
<0.0001
<0.0001
0.0002
<0.0001
<0.0001
Standardized
Coefficient
0.00000
-0.2497
0.1515
0.0777
0.1380
0.2250
R2 = 0.264; p <0.0001; based on data from Bunker Hill Superfund Site, collected over the period 1988-2002.
GM: geometric mean; In: natural log.
Source: TerraGraphics (2004).
1 Malcoe et al. (2002) analyzed 1997 data on blood Pb and environmental Pb
2 concentrations in a representative sample of Native American and white children
3 (n = 224, age 1-6 years) who resided in a former Pb mining region in Ottawa County,
4 OK. The data set included measurements of blood Pb, yard soil Pb, residential interior
5 dust Pb loading, first-draw water Pb, paint Pb assessment and other behavioral (i.e., hand-
6 to-mouth activity, hygiene rating) and demographic variables (i.e., hygiene rating,
7 poverty level, caregiver education). A multivariate regression model accounted for 34%
8 of the observed variability in blood Pb. Yard soil Pb and interior dust Pb loading
9 accounted for 10% and 3% of the blood Pb variability, respectfully. The regression model
10 predicted a slope of 0.74 (ig/dL blood Pb per ln[(ig/g soil Pb] and a slope of 0.45 (ig/dL
11 blood Pb per ln[(ig/ft2] dust Pb loading.
4.6 Biokinetic Models of Pb Exposure-Blood Pb Relationships
12 An alternative to regression models are mechanistic models, which attempt to specify all
13 parameters needed to describe the mechanisms (or processes) of transfer of Pb from the
14 environment to human tissues. Such mechanistic models are more complex than
15 regression models; this added complexity introduces challenges in terms of their
16 mathematical solution and empirical verification. However, by incorporating parameters
17 that can be expected to vary spatially or temporally, or across individuals or populations,
18 mechanistic models can be extrapolated to a wide range of exposure scenarios, including
19 those that may be outside of the domain of paired predictor-outcome data used to develop
20 the model. Exposure-intake models, a type of mechanistic models, are highly simplified
21 mathematical representations of relationships between levels of Pb in environmental
November 2012 4-138 Draft - Do Not Cite or Quote
-------
1 media and human Pb intakes (e.g., (ig Pb ingested per day). These models include
2 parameters representing processes of Pb transfer between environmental media (e.g., air
3 to surface dust) and to humans, including rates of human contact with the media and
4 intakes of the media (e.g., g soil ingested per day). Intake-biokinetic models provide the
5 analogous mathematical representation of relationships between Pb intakes and Pb levels
6 in body tissues (e.g., blood Pb concentration). Biokinetic models include parameters that
7 represent processes of Pb transfer (a) from portals of entry into the body and (b) from
8 blood to tissues and excreta. Linked together, exposure-intake and intake-biokinetics
9 models (i.e., integrated exposure-intake-biokinetics models) provide an approach for
10 predicting blood Pb concentrations (or Pb concentrations in other tissues) that
11 corresponds to a specified exposure (medium, concentration, and duration). Detailed
12 information on exposure and internal dose can be obtained from controlled experiments,
13 but almost never from epidemiological observations or from public health monitoring
14 programs. Exposure intake-biokinetics models can provide these predictions in the
15 absence of complete information on the exposure history and blood Pb concentrations for
16 an individual (or population) of interest. Therefore, these models are critical to applying
17 epidemiologic-based information on blood Pb-response relationships to the quantification
18 and characterization of human health risk. They are also critical for assessing the
19 potential impacts of public health programs directed at mitigation of Pb exposure or of
20 remediation of contaminated sites.
21 However, they are not without their limitations. Human exposure-biokinetics models
22 include large numbers of parameters, which are required to describe the many processes
23 that contribute to Pb intake, absorption, distribution, and elimination. The large number
24 of parameters complicates the assessment of confidence in parameter values, many of
25 which cannot be directly measured. Statistical procedures can be used to evaluate the
26 degree to which model outputs conform to "real-world" observations and values of
27 influential parameters can be statistically estimated to achieve good agreement with
28 observations. Still, large uncertainty can be expected to remain about many, or even
29 most, parameters in complex exposure-biokinetic models. Such uncertainties need to be
30 identified and their impacts on model predictions quantified (i.e., sensitivity analysis or
31 probabilistic methods).
32 Modeling of human Pb exposures and biokinetics has advanced considerably during the
33 past several decades, although there have been relatively few developments since the
34 2006 Pb AQCD was published. Still in use is the Integrated Exposure Uptake Biokinetic
35 (IEUBK) Model for Lead in Children (U.S. EPA. 1994) and models that simulate Pb
36 biokinetics in humans from birth through adulthood (O'Flahertv, 1995; Leggett 1993;
37 O'Flahertv. 1993). The EPA AALM is still in development. A complete and extensive
38 discussion of these models can be found in the 2006 Pb AQCD (U.S. EPA. 2006b).
November 2012 4-139 Draft - Do Not Cite or Quote
-------
4.7 Summary and Conclusions
4.7.1 Exposure
1 Exposure data considered in this assessment build upon the conclusions of the
2 2006 Pb AQCD (2006b), which found that air Pb concentrations in the U.S. and
3 associated biomarkers of exposure to Pb have decreased substantially following
4 reductions in industrial point sources of Pb, and restrictions on Pb in gasoline, house-hold
5 paints, and solder. Pb exposure is difficult to assess because Pb has multiple sources in
6 the environment and passes through various media. The atmosphere is the main
7 environmental transport pathway for Pb, and, on a global scale, atmospheric Pb is
8 primarily associated with fine particulate matter, which can deposit to soil and water. In
9 addition to primary emission of particle-bearing or gaseous Pb to the atmosphere, Pb can
10 be suspended to the air from soil or dust. Air-related pathways of Pb exposure are the
11 focus of this assessment. In addition to inhalation of Pb from ambient air, air-related Pb
12 exposure pathways include inhalation and ingestion of Pb from indoor dust and/or
13 outdoor soil that originated from recent or historic ambient air (e.g., air Pb that has
14 penetrated into the residence either via the air or tracking of soil), ingestion of Pb in
15 drinking water contaminated from atmospheric deposition onto surface waters or from
16 indirect surface runoff of deposition of ambient Pb, and ingestion of Pb in dietary sources
17 after uptake by plants or grazing animals. Non-air-related Pb exposures may include
18 occupational exposures, hand-to-mouth contact with Pb-containing consumer goods,
19 hand-to-mouth contact with dust or chips of peeling Pb-containing paint, or ingestion of
20 Pb in drinking water conveyed through Pb pipes. Pb can cycle through multiple media
21 prior to human exposure. Given the multitude of possible air-related exposure scenarios
22 and the related difficulty of constructing Pb exposure histories, most studies of Pb
23 exposure through air, water, and soil can be informative to this review. Other exposures,
24 such as occupational exposures, contact with consumer goods in which Pb has been used,
25 or ingestion of Pb in drinking water conveyed through Pb pipes may also contribute to Pb
26 body burden.
27 A number of monitoring and modeling techniques have been employed for ambient Pb
28 exposure assessment. Environmental Pb concentration data can be collected from
29 ambient air Pb monitors, soil Pb samples, dust Pb samples, and dietary Pb samples to
30 estimate human exposure. Exposure estimation error depends in part on the collection
31 efficiency of these methods; collection efficiency for ambient air Pb FRM samplers is
32 described in Section 3.4. Additionally, high spatial variability of the Pb concentrations in
33 various media also can contribute to exposure error, as described in the 2009 PM ISA
34 (U.S. EPA. 2009a). Models, such as the Integrated Exposure Uptake Biokinetic (IEUBK)
November 2012 4-140 Draft - Do Not Cite or Quote
-------
1 model, simulate human exposure to Pb from multiple sources and through various routes
2 including inhalation, ingestion, and dermal exposure. IEUBK model inputs include soil
3 Pb concentration, air Pb concentration, dietary Pb intake including drinking water, Pb
4 dust ingestion, human activity, and biokinetic factors. Measurements and/or assumptions
5 can be utilized when formulating the model inputs; errors in measurements and
6 assumptions thus have the potential to propagate through the exposure models.
7 Section 4.1 presents data illustrating potential exposure pathways. Soil can act as a
8 reservoir for deposited Pb emissions, and exposure to soil contaminated with deposited
9 Pb can occur through resuspended PM as well as shoe tracking and hand-to-mouth
10 contact, which is the main pathway of childhood exposure to Pb. Airborne particles
11 containing Pb tend to be small (much of the distribution <10 nm) compared with Pb in
12 soil or dust particles (~50 pirn to several hundred nm); Pb deposition to soil is described
13 in Section 3.3. Hence, hand-to-mouth contact with Pb-bearing soil or dust and/or tracking
14 Pb contaminated soil or dust into homes are more common means for human exposure to
15 Pb. Infiltration of Pb dust into indoor environments has been observed, and Pb dust has
16 been shown to persist in indoor environments even after repeated cleanings.
17 Measurements of particle-bound Pb exposures reported in this assessment have shown
18 that personal exposure measurements for Pb concentration are typically higher than
19 indoor or outdoor ambient Pb concentrations.
4.7.2 Toxicokinetics
20 The majority of Pb in the body is found in bone (roughly 90% in adults, 70% in children);
21 only about 1% of Pb is found in the blood. Pb in blood is primarily (-99%) bound to red
22 blood cells (RBCs). It has been suggested that the small fraction of Pb in plasma (<1%)
23 may be the more biologically labile and lexicologically active fraction of the circulating
24 Pb. The relationship between Pb in blood and plasma is pseudo-linear at relatively low
25 daily Pb intakes (i.e., <10 ug/day/kg) and at blood Pb concentrations <25 (ig/dL, and
26 becomes curvilinear at higher blood Pb concentrations due to saturable binding to RBC
27 proteins. As blood Pb level increases and the higher affinity binding sites for Pb in RBCs
28 become saturated, a larger fraction of the blood Pb is available in plasma to distribute to
29 brain and other Pb-responsive tissues.
30 The burden of Pb in the body may be viewed as divided between a dominant slow
31 (i.e., uptake and elimination) compartment (bone) and smaller fast compartment(s) (soft
32 tissues). Pb uptake and elimination in soft tissues is much faster than in bone. Pb
33 accumulates in bone regions undergoing the most active calcification at the time of
34 exposure. During infancy and childhood, bone calcification is most active in trabecular
November 2012 4-141 Draft - Do Not Cite or Quote
-------
1 bone (e.g., patella); whereas, in adulthood, calcification occurs at sites of remodeling in
2 cortical (e.g., tibia) and trabecular bone (Aufderheide and Wittmers. 1992). A high bone
3 formation rate in early childhood results in the rapid uptake of circulating Pb into
4 mineralizing bone; however, in early childhood bone Pb is also recycled to other tissue
5 compartments or excreted in accordance with a high bone resorption rate (O'Flaherty.
6 1995). Thus, much of the Pb acquired early in life is not permanently fixed in the bone.
7 The exchange of Pb from plasma to the bone surface is a relatively rapid process. Pb in
8 bone becomes distributed in trabecular and the more dense cortical bone. The proportion
9 of cortical to trabecular bone in the human body varies by age, but on average is about
10 80% cortical to 20% trabecular. Of the bone types, trabecular bone is more reflective of
11 recent exposures than is cortical bone due to the slow turnover rate and lower blood
12 perfusion of cortical bone. Some Pb diffuses to deeper bone regions where it is relatively
13 inert, particularly in adults. These bone compartments are much more labile in infants
14 and children than in adults as reflected by half-times for movement of Pb from bone into
15 to the plasma (e.g., cortical half-time = 0.23 years at birth, 3.7 years at 15 years of age,
16 and 23 years in adults; trabecular half-time = 0.23 years at birth, 2.0 years at 15 years of
17 age, and 3.8 years in adults) (Leggett 1993).
18 Evidence for maternal-to-fetal transfer of Pb in humans is derived from cord blood to
19 maternal blood Pb ratios. Group mean ratios range from about 0.7 to 1.0 at the time of
20 delivery for mean maternal blood Pb levels ranging from 1.7 to 8.6 (ig/dL. Transplacental
21 transfer of Pb may be facilitated by an increase in the plasma/blood Pb concentration
22 ratio during pregnancy. Maternal-to-fetal transfer of Pb appears to be related partly to the
23 mobilization of Pb from the maternal skeleton.
24 The dominant elimination phase of Pb kinetics in the blood, exhibited shortly after a
25 change in exposure occurs, has a half-life of-20-30 days. An abrupt change in Pb uptake
26 gives rise to a relatively rapid change in blood Pb, to a new quasi-steady state, achieved
27 in -75-100 days (i.e., 3-4 times the blood elimination half-life). A slower phase of Pb
28 clearance from the blood may become evident with longer observation periods following
29 a decrease in exposure due to the gradual redistribution of Pb among bone and other
30 compartments.
4.7.3 Pb Biomarkers
31 Overall, trends in blood Pb levels have been decreasing among U.S. children and adults
32 over the past 20 years (Section 4.4). The median blood Pb level for the entire U.S.
33 population is 1.2 ug/dL and the 95th percentile blood Pb level was 3.7 ug/dL, based on
34 the 2007-2008 NHANES data (NCHS. 2010). Among children aged 1-5 years, the
November 2012 4-142 Draft - Do Not Cite or Quote
-------
1 median and 95th percentiles were slightly higher at 1.4 ug/dL and 4.1 ug/dL,
2 respectively.
3 Blood Pb is dependent on both the recent exposure history of the individual, as well as
4 the long-term exposure history that determines body burden and Pb in bone. The
5 contribution of bone Pb to blood Pb changes depending on the duration and intensity of
6 the exposure, age, and various other physiological stressors that may affect bone
7 remodeling (e.g., nutritional status, pregnancy, menopause, extended bed rest,
8 hyperparathyroidism) beyond that which normally and continuously occurs. In children,
9 largely due to faster exchange of Pb to and from bone, blood Pb is both an index of recent
10 exposure and potentially an index of body burden. In adults and children, where exposure
11 to Pb has effectively ceased or greatly decreased, a slow decline in blood Pb
12 concentrations over the period of years is most likely due to the gradual release of Pb
13 from bone. Bone Pb is an index of cumulative exposure and body burden. Even bone
14 compartments should be recognized as reflective of differing exposure periods with Pb in
15 trabecular bone exchanging more rapidly than Pb in cortical bone with the blood. This
16 difference in the compartments makes Pb in cortical bone a better marker of cumulative
17 exposure and Pb in trabecular bone more likely to be correlated with blood Pb, even in
18 adults.
19 Sampling frequency is an important consideration when evaluating blood Pb and bone Pb
20 levels in epidemiologic studies, particularly when the exposure is not well characterized.
21 It is difficult to determine what blood Pb is reflecting in cross-sectional studies that
22 sample blood Pb once, whether recent exposure or movement of Pb from bone into blood
23 from historical exposures. In contrast, cross-sectional studies of bone Pb and longitudinal
24 samples of blood Pb concentrations overtime provide more of an index of cumulative
25 exposure and are more reflective of average Pb body burdens overtime. The degree to
26 which repeated sampling will reflect the actual long-term time-weighted average blood
27 Pb concentration depends on the sampling frequency in relation to variability in
28 exposure. High variability in Pb exposures can produce episodic (or periodic) oscillations
29 in blood Pb concentration that may not be captured with low sampling frequencies.
30 Furthermore, similar blood Pb concentrations in two individuals (or populations),
31 regardless of their age, do not necessarily translate to similar body burdens or similar
32 exposure histories.
33 The concentration of Pb in urine follows blood Pb concentration, in that it mainly reflects
34 the exposure history of the previous few months and therefore, is likely a relatively poor
35 index of Pb body burden. There is added complexity with Pb in urine because
36 concentration is also dependent upon urine flow rate, which requires timed urine samples
November 2012 4-143 Draft - Do Not Cite or Quote
-------
1 that is often not feasible in epidemiologic studies. Other biomarkers have been utilized to
2 a lesser extent (e.g., Pb in teeth).
4.7.4 Air Lead-Blood Lead Relationships
3 The 1986 Pb AQCD (U.S. EPA. 1986a) described epidemiological studies of
4 relationships between air Pb and blood Pb. Much of the pertinent earlier literature for
5 child populations described in the 1986 Pb AQCD was also included in a meta-analysis
6 by Brunekreef (1984). Based on the studies available at that time, the 1986 Pb AQCD
7 concluded that "the blood Pb versus air Pb slope (3 is much smaller at high blood and air
8 levels." This is to say that the slope (3 was much smaller for occupational exposures
9 where high blood Pb levels (>40 ug/dL) and high air Pb levels (much greater than
10 10 ug/m3) prevailed relative to lower environmental exposures which showed lower
11 blood Pb and air Pb concentrations (<30 ug/dL and <3 ug/m3). For those environmental
12 exposures, it was concluded that the relationship between blood Pb and air Pb "... for
13 direct inhalation appears to be approximately linear in the range of normal ambient
14 exposures (0.1-2.0 ug/m3)" (Chapter 1, pp 98 of the 1986 Pb AQCD). In addition to the
15 meta-analysis of Brunekreef (1984). more recent studies have provided data from which
16 estimates of the blood Pb-air Pb slope can be derived for children (Table 4-12). The range
17 of estimates from these studies is 4-9 ug/dL per ug/m3, which encompasses the estimate
18 from the Brunekreef (1984) meta-analysis. Most studies have described the blood Pb-air
19 Pb relationship as either log-log (Schnaas et al.. 2004; Hayes et al.. 1994; Brunekreef,
20 1984). which predicts an increase in the blood Pb-air Pb slope with decreasing air Pb
21 concentration or linear (Hilts. 2003; Tripathi et al. 2001; Schwartz and Pitcher. 1989).
22 which predicts a constant blood Pb-air Pb slope regardless of air Pb concentrations. These
23 differences may simply reflect model selection by the investigators; alternative models
24 are not reported in these studies. The blood Pb-air Pb slope may also be affected in some
25 studies by the inclusion of parameters (e.g., soil Pb) that may account for some of the
26 variance in blood Pb attributable to air Pb. Other factors that likely contribute to the
27 derived blood Pb-air Pb slope include differences in the populations examined and Pb
28 sources, which varied among individual studies.
November 2012 4-144 Draft - Do Not Cite or Quote
-------
References for Chapter 4
Abadin. HG: Wheeler. JS. (1997). Guidance for risk assessment of exposure to lead: A site-specific,
multi-media approach. In JS Andrews; H Frumkin; BL Johnson; MA Mehlman; C Xintaras; JA
Bucsela (Eds.), Hazardous waste and public health: International congress on the health effects of
hazardous waste (pp. 477-485). Princeton, NJ: Princeton Scientific Publishing.
Abrahams. PW; Follansbee. MH; Hunt. A; Smith. B; Wragg. J. (2006). Iron nutrition and possible lead
toxicity: An appraisal of geophagy undertaken by pregnant women of U.K. Asian communities. Appl
Geochem 21: 98-108. http://dx.doi.Org/10.1016/i.apgeochem.2005.09.015
ACGIH (American Conference of Governmental Industrial Hygienists). (1985). Particle size-selective
sampling in the workplace: Report of the ACGIH Technical Committee on Air Sampling Procedures.
Cincinnati, OH.
Adgate. JL; Mongin. SJ; Pratt. GC; Zhang. J; Field. MP; Ramachandran. G; Sexton. K. (2007).
Relationships between personal, indoor, and outdoor exposures to trace elements in PM2.5. Sci Total
Environ 386: 21-32. http://dx.doi.Org/10.1016/i.scitotenv.2007.07.007
Al-Modhefer. AJ; Bradbury. MW; Simons. TJ. (1991). Observations on the chemical nature of lead in
human blood serum. Clin Sci (Lond) 81: 823-829.
Alexander. FW; Clayton. BE; Delves. HT. (1974). Mineral and trace-metal balances in children receiving
normal and synthetic diets. QJM 43: 89-111.
Alves. C; Goncalves. C; Fernandes. AP; Tarelho. L; Pio. C. (2011). Fireplace and woodstove fine particle
emissions from combustion of western Mediterranean wood types. Atmos Res 101: 692-700.
http://dx.doi.0rg/10.1016/i.atmosres.2011.04.015
Amaral. JH; Rezende. VB; Quintana. SM; Gerlach. RF; Barbosa. F. Jr; Tanus-Santos. JE. (2010). The
relationship between blood and serum lead levels in peripartum women and their respective umbilical
cords. Basic Clin Pharmacol Toxicol 107: 971-975.
http://dx.doi.0rg/10.llll/i.1742-7843.2010.00616.x
Araki. S; Aono. H; Yokoyama. K; Murata. K. (1986). Filterable plasma concentration, glomerular
filtration, tubular balance, and renal clearance of heavy metals and organic substances in metal
workers. Arch Environ Occup Health 41: 216-221. http://dx.doi.org/10.1080/00039896.1986.9938336
Araki. S; Sata. F; Murata. K. (1990). Adjustment for urinary flow rate: an improved approach to
biological monitoring. Int Arch Occup Environ Health 62: 471-477.
http://dx.doi.org/10.1007/BF00379066
Armstrong. R; Chettle. PR; Scott. MC; Somervaille. LJ; Pendlington. M. (1992). Repeated measurements
of tibia lead concentrations by in vivo X-ray fluorescence in occupational exposure. Occup Environ
Med49: 14-16.
Aro. A; Amarasiriwardena. C; Lee. ML; Kim. R; Hu. H. (2000). Validation of K x-ray fluorescence bone
lead measurements by inductively coupled plasma mass spectrometry in cadaver legs. Med Phys 27:
119-123. http://dx.doi.0rg/10.1118/l.598863
Arora. M; Ettinger. AS; Peterson. KE; Schwartz. J; Hu. H; Hernandez-Avila. M; Tellez-Rojo. MM;
Wright. RO. (2008). Maternal dietary intake of polyunsaturated fatty acids modifies the relationship
between lead levels in bone and breast milk. J Nutr 138: 73-79.
Arruda-Neto. JDT; de Oliveira. MCC; Sarkis. JES; Sordini. P; Manso-Guevara. MV; Garcia. F; Prado.
GR; Krug. FJ; Mesa. J; Bittencourt-Oliveira. MC; Garcia. C; Rodrigues. TE; Shtejer. K; Genofre. GC.
(2009). Study of environmental burden of lead in children using teeth as bioindicator. Environ Int 35:
614-618. http://dx.doi.0rg/10.1016/i.envint.2008.12.005
November 2012 4-145 Draft - Do Not Cite or Quote
-------
Arruda-Neto. JD; Geraldo. LP; Prado. GR; Garcia. F; Bittencourt-Oliveira. MC: Sarkis. JE; Martinez-
Lusardo. F; Lima-Cazorla. L; Rosa-Medero. D; Rodrigues. TE; Genofre. GC. (2010). Study of metals
transfer from environment using teeth as biomonitor. Environ Int 36: 243-246.
http://dx.doi.0rg/10.1016/i.envint.2009.12.003
Astrin. KH; Bishop. DF; Wetmur. JG: Kaul. B; Davidow. B; Desnick. RJ. (1987). 5-aminolevulinic acid
dehydratase isozymes and lead toxicity. Ann N Y Acad Sci 514: 23-29.
http://dx.doi.org/10.1111/i. 1749-6632.1987.tb48757.x
Aufderheide. AC: Wittmers. LE. Jr. (1992). Selected aspects of the spatial distribution of lead in bone
[Review]. Neurotoxicology 13: 809-819.
Aungst. BJ; Dolce. JA; Fung. H. (1981). The effect of dose on the disposition of lead in rats after
intravenous and oral administration. Toxicol Appl Pharmacol 61: 48-57.
http://dx.doi.org/10.1016/0041-008X(81)90006-5
Aungst. BJ; Fung. H. (1985). The effects of dietary calcium on lead absorption, distribution, and
elimination kinetics in rats. J Toxicol Environ Health 16: 147-159.
http://dx.doi.org/10.1080/15287398509530726
Avula. B; Wang. YH; Smillie. TJ; Duzgoren-Aydin. NS: Khan. IA. (2010). Quantitative determination of
multiple elements in botanicals and dietary supplements using ICP-MS. J Agric Food Chem 58:
8887-8894. http://dx.doi.org/10.1021/ifl01598g
Azar. A; Trochimowicz. HJ; Maxfield. ME. (1973). Review of lead studies in animals carried out at
Haskell Laboratory: Two year feeding study and response to hemorrhage study. In D Earth; A Berlin;
R Engel; P Recht; J Smeets (Eds.), Environmental health aspects of lead: Proceedings of an
international symposium (pp. 199-210). Luxembourg: Commission of the European Communities.
Bailey. MR; Roy. M. (1994). Clearance of particles from the respiratory tract. In Human respiratory tract
model for radiological protection: A report of a task group of the International Commission on
Radiological Protection (pp. 301-413). Oxford, U.K.: Pergamon.
Bandeen-Roche. K; Glass. TA; Bolla. KI; Todd. AC; Schwartz. BS. (2009). Cumulative lead dose and
cognitive function in older adults. Epidemiology 20: 831-839.
http://dx.doi.org/10.1097/EDE.Ob013e3181b5flOO
Bannon. PI; Abounader. R; Lees. PSJ; Bressler. JP (2003). Effect of DMT1 knockdown on iron,
cadmium, and lead uptake in Caco-2 cells. Am J Physiol Cell Physiol 284: C44-C50.
Bannon. PI; Drexler. JW; Pent. GM; Casteel. SW; Hunter. PJ; Brattin. WJ; Major. MA. (2009).
Evaluation of small arms range soils for metal contamination and lead bioavailability. Environ Sci
Technol 43: 9071-9076. http://dx.doi.org/10.1021/es901834h
Barbosa. F. Jr; Heloisa. M; Rodrigues. C; Buzalaf. MR; Krug. FJ; Gerlach. RF; Tanus-Santos. JE.
(2006a). Evaluation of the use of salivary lead levels as a surrogate of blood lead or plasma lead levels
in lead exposed subjects. Arch Toxicol 80: 633-637. http://dx.doi.org/10.1007/s00204-006-0096-y
Barbosa. F. Jr; Ramires. I; Rodrigues. MHC; Saint' Pierre. TD; Curtius. AJ; Buzalaf. MR; Gerlach. RF;
Tanus-Santos. JE. (2006b). Contrasting effects of age on the plasma/whole blood lead ratio in men and
women with a history of lead exposure. Environ Res 102: 90-95.
http://dx.doi.0rg/10.1016/i.envres.2006.03.007
Barltrop. D; Meek. F. (1979). Effect of particle size on lead absorption from the gut. Arch Environ Occup
Health 34: 280-285.
Barrett. J; Taylor. K; Hudson-Edwards. K; Charnock. J. (2010). Solid-phase speciation of Pb in urban
road dust sediment: A XANES and EXAFS study. Environ Sci Technol 44: 2940-2946.
http://dx.doi.org/10.1021/es903737k
Barry. PSI. (1975). A comparison of concentrations of lead in human tissues. Occup Environ Med 32:
119-139.
Barry. PSI. (1981). Concentrations of lead in the tissues of children. Occup Environ Med 38: 61-71.
November 2012 4-146 Draft - Do Not Cite or Quote
-------
Barry. PSI; Connolly. R. (1981). Lead concentrations in mediaeval bones. Int Arch Occup Environ Health
48: 173-177. http://dx.doi.org/10.1007/BF00378438
Barton. JC: Conrad. ME; Harrison. L; Nubv. S. (1978a). Effects of calcium on the absorption and
retention of lead. Transl Res 91: 366-376.
Barton. JC: Conrad. ME; Nubv. S: Harrison. L. (1978b). Effects of iron on the absorption and retention of
lead. Transl Res 92: 536-547.
Beauchemin. S: Maclean. LC: Rasmussen. PE. (2011). Lead speciation in indoor dust: A case study to
assess old paint contribution in a Canadian urban house. Environ Geochem Health 33: 343-352.
http://dx.doi.org/10.1007/sl0653-011-9380-8
Behinaein. S: Chettle. PR; Atanackovic. J: Egden. LM; Fleming. DE; Nie. LH; Richard. N; Stever. S.
(2011). In vivo measurement of lead in the bones of smelter workers using the four-element 'clover-
leaf geometry detector system. Phys Med Biol 56: 653-665.
http://dx.doi.Org/10.1088/0031-9155/56/3/008
Bellinger. D; Hu. H; Titlebaum. L; Needleman. HL. (1994a). Attentional correlates of dentin and bone
lead levels in adolescents. Arch Environ Health 49: 98-105.
http://dx.doi.org/10.1080/00039896.1994.9937461
Bergdahl. IA; Grubb. A; Schiitz. A; Desnick. RJ; Wetmur. JG: Sassa. S: Skerfving. S. (1997a). Lead
binding to delta-aminolevulinic acid dehydratase (ALAD) in human erythrocytes. Basic Clin
Pharmacol Toxicol 81: 153-158. http://dx.doi.org/10.1111/i.1600-0773.1997.tb02061.x
Bergdahl. IA; Schutz. A; Gerhardsson. L; Jensen. A; Skerfving. S. (1997b). Lead concentrations in human
plasma, urine and whole blood. Scand J Work Environ Health 23: 359-363.
Bergdahl. IA; Schutz. A; Grubb. A. (1996). Application of liquid chromatography-inductively coupled
plasma mass spectrometry to the study of protein-bound lead in human erythrocytes. J Anal At
Spectrom 11: 735-738. http://dx.doi.org/10.1039/JA9961100735
Bergdahl. IA; Sheveleva. M; Schutz. A; Artamonova. VG: Skerfving. S. (1998). Plasma and blood lead in
humans: Capacity-limited binding to delta-aminolevulinic acid dehydratase and other lead-binding
components. Toxicol Sci 46: 247-253. http://dx.doi.Org/10.1093/toxsci/46.2.247
Bergdahl. IA; Vahter. M; Counter. SA; Schutz. A; Buchanan. LH; Ortega. F; Laurell. G: Skerfving. S.
(1999). Lead in plasma and whole blood from lead-exposed children. Environ Res 80: 25-33.
http://dx.doi.org/10.1006/enrs.1998.3880
Berger. OG: Gregg. DJ: Succop. PA. (1990). Using unstimulated urinary lead excretion to assess the need
for chelation in the treatment of lead poisoning. J Pediatr 116: 46-51.
http://dx.doi.org/10.1016/S0022-3476(05)81643-9
Berkowitz. GS: Wolff. MS: Lapinski. RH; Todd. AC. (2004). Prospective study of blood and tibia lead in
women undergoing surgical menopause. Environ Health Perspect 112: 1673-1678.
Besser. JM; Brumbaugh. WG: Ivev. CD; Ingersoll. CG: Moran. PW. (2008). Biological and chemical
characterization of metal bioavailability in sediments from Lake Roosevelt, Columbia River,
Washington, USA. Arch Environ Contam Toxicol 54: 557-570.
http://dx.doi.org/10.1007/s00244-007-9074-5
Blake. KCH; Barbezat. GO: Mann. M. (1983). Effect of dietary constituents on the gastrointestinal
absorption of 203Pb in man. Environ Res 30: 182-187.
http://dx.doi.org/10.1016/0013-9351(83)90178-0
Blake. KCH: Mann. M. (1983). Effect of calcium and phosphorus on the gastrointestinal absorption of
203Pb in man. Environ Res 30: 188-194. http://dx.doi.org/10.1016/0013-9351(83)90179-2
Bleecker. ML; Ford. DP; Vaughan. CG: Walsh. KS: Lindgren. KN. (2007b). The association of lead
exposure and motor performance mediated by cerebral white matter change. Neurotoxicology 28:
318-323. http://dx.doi.0rg/10.1016/i.neuro.2006.04.008
November 2012 4-147 Draft - Do Not Cite or Quote
-------
Bleecker. ML; Lindgren. KN; Ford. DP. (1997). Differential contribution of current and cumulative
indices of lead dose to neuropsychological performance by age. Neurology 48: 639-645.
Bolanowska. W: Piotrowski. J; Garczynski. H. (1967). Triethyllead in the biological material in cases of
acute tetraethyllead poisoning. Arch Toxicol 22: 278-282. http://dx.doi.org/10.1007/BF00577718
Bonnard. R; Mckone. TE. (2009). Integration of the predictions of two models with dose measurements in
a case study of children exposed to the emissions from a lead smelter. Hum Ecol Risk Assess 15:
1203-1226. http://dx.doi.org/10.1080/10807030903304849
Booker. DV; Chamberlain. AC: Newton. D; Stott. ANB. (1969). Uptake of radioactive lead following
inhalation and injection. Br J Radiol 42: 457-466. http://dx.doi.org/10.1259/0007-1285-42-498-457
Bornschein. RL; Succop. P; Dietrich. KN; Clark. CS: Que Hee SHammond. PB. (1985). The influence of
social and environmental factors on dust lead, hand lead, and blood lead levels in young children.
Environ Res 38: 108-118. http://dx.doi.org/10.1016/0013-9351(85)90076-3
Bos. AJJ; Van der Stap. CCA. H; Valkovic. V: Vis. RD; Verheul. H. (1985). Incorporation routes of
elements into human hair: Implications for hair analysis used for monitoring. Sci Total Environ 42:
157-169. http://dx.doi.org/10.1016/0048-9697(85)90015-4
Bowers. TS: Beck. BD; Karam. HS. (1994). Assessing the relationship between environmental lead
concentrations and adult blood lead levels. Risk Anal 14: 183-189.
httrj://dx.doi.org/10.1111/i.l539-6924.1994.tb00043.x
Brattin. W: Griffin. S. (2011). Evaluation of the contribution of lead in soil to lead in dust at superfund
sites. Hum Ecol Risk Assess 17: 236-244. http://dx.doi.org/10.1080/10807039.2011.538638
Bres. EF; Voegel. JC: Barry. JC: Waddington. WG: Frank. RM. (1986). Feasibility study for the detection
of lead substitution sites in the hydroxyapatite crystal structure using high-resolution electron
microscopy (HREM) at optimum focus. Journal of Applied Crystallography 19: 168-173.
http://dx.doi.org/10.1107/S0021889886089719
Brodeur. J: Lacasse. Y; Talbot. D. (1983). Influence of removal from occupational lead exposure on blood
and saliva lead concentrations. Toxicol Lett 19: 195-199.
http://dx.doi.org/10.1016/0378-4274(83)90282-5
Brodv. DJ: Pirkle. JL; Kramer. RA; Flegal. KM; Matte. TD; Gunter. EW: Paschal. DC. (1994). Blood
lead levels in the US population: Phase 1 of the third National Health and Nutrition Examination
Survey (NHANES III, 1988 to 1991). JAMA 272: 277-283.
http://dx.doi.org/10.1001/iama.1994.03520040039038
Bronner. F; Pansu. D: Stein. W. (1986). An analysis of intestinal calcium transport across the rat intestine
[Review]. Am J Physiol 250: G561- G569.
Brunekreef. B. (1984). The relationship between air lead and blood lead in children: A critical review
[Review]. Sci Total Environ 38: 79-123. http://dx.doi.org/10.1016/0048-9697(84)90210-9
Burke. JM; Rea. AW: Suggs. J: Williams. R; Xue. JP; Ozkavak. H. (2002). Ambient paniculate matter
exposures: A comparison of sheds-PM exposure model predictions and estimations derived from
measurements collected during NERL's RTF PM panel study. Epidemiology 13: S83.
Butterweck. G: Schuler. C: Vezzu. G: Miiller. R; Marsh. J: Thrift. S: Birchall. A. (2002). Experimental
determination of the absorption rate of unattached radon progeny from respiratory tract to blood.
Radiat Prot Dosimetry 102: 343- 348.
Caffo. B; Chen. S: Stewart. W: Bolla. K; Yousem. D; Davatzikos. C: Schwartz. BS. (2008). Are brain
volumes based on magnetic resonance imaging mediators of the associations of cumulative lead dose
with cognitive function? Am J Epidemiol 167: 429-437. http://dx.doi.org/10.1093/aje/kwm326
Cai. QY; Mo. CH; Wu. QT; Zeng. QY; Katsoyiannis. A. (2007). Concentration and speciation of heavy
metals in six different sewage sludge-composts. J Hazard Mater 147: 1063-1072.
http://dx.doi.0rg/10.1016/i.ihazmat.2007.01.142
November 2012 4-148 Draft - Do Not Cite or Quote
-------
Campbell. JR; Toribara. TY. (2001). Hair-root lead to screen for lead toxicity. J Trace Elem Exp Med 14:
69-72. http://dx.doi.org/10.1002/1520-670X(2001)14:l<69::AID-JTRA1009>3.0.CO:2-A
Caravanos. J: Weiss. AL; Jaeger. RJ. (2006b). An exterior and interior leaded dust deposition survey in
New York City: Results of a 2-year study. Environ Res 100: 159-164.
http://dx.doi.0rg/10.1016/i.envres.2005.08.005
Carbone. R; Laforgia. N; Crollo. E; Mautone. A; lolascon. A. (1998). Maternal and neonatal lead
exposure in southern Italy. Neonatology 73: 362-366.
Carlisle. JC: Wade. MJ. (1992). Predicting blood lead concentrations from environmental concentrations.
Regul Toxicol Pharmacol 16: 280-289. httrj://dx.doi.org/10.1016/0273-2300(92)90008-W
Carr. E; Lee. M; Marin. K; Holder. C: Hover. M; Pedde. M; Cook. R; Touma. J. (2011). Development and
evaluation of an air quality modeling approach to assess near-field impacts of lead emissions from
piston-engine aircraft operating on leaded aviation gasoline. Atmos Environ 45: 5795-5804.
http://dx.doi.0rg/10.1016/i.atmosenv.2011.07.017
Carrizales. L; Razo. I; Tellez-Hernandez. JI; Torres-Nerio. R; Torres. A; Batres. LE; Cubillas. AC: Diaz-
Barriga. F. (2006). Exposure to arsenic and lead of children living near a copper-smelter in San Luis
Potosi, Mexico: Importance of soil contamination for exposure of children. Environ Res 101: 1-10.
http://dx.doi.0rg/10.1016/i.envres.2005.07.010
Casteel. SW: Cowart. RP; Weis. CP; Henningsen. GM; Hoffman. E; Brattin. WJ: Guzman. RE; Starost.
MF; Payne. JT; Stockham. SL; Becker. SV: Drexler. JW: Turk. JR. (1997). Bioavailability of lead to
juvenile swine dosed with soil from the Smuggler Mountain NPL site of Aspen, Colorado. Toxicol Sci
36: 177-187. http://dx.doi.org/10.1006/faat. 1997.2296
Casteel. SW: Weis. CP: Henningsen. GM; Brattin. WL. (2006). Estimation of relative bioavailability of
lead in soil and soil-like materials using young swine. Environ Health Perspect 114: 1162-1171.
http://dx.doi.org/10.1289/ehp.8852
CDC (Centers for Disease Control and Prevention). (2002). Childhood lead poisoning associated with
tamarind candy and folk remedies-California, 1999-2000. MMWR Recomm Rep 51: 684-686.
CDC (Centers for Disease Control and Prevention). (2011 a). Fourth national report on human exposure to
environmental chemicals: Updated tables, February 2011. Atlanta, GA.
http://www.cdc.gov/exposurereport/pdf/FourthReport.pdf
Chamberlain. AC. (1983). Effect of airborne lead on blood lead. Atmos Environ 17: 677-692.
http://dx.doi.org/10.1016/0004-6981(83)90415-8
Chamberlain. AC: Clough. WS: Heard. MJ: Newton. D; Stott. ANB; Wells. AC. (1975). Uptake of lead by
inhalation of motor exhaust. Proc Biol Sci 192: 77-110.
Chamberlain. AC: Heard. MJ: Little. P; Newton. D; Wells. AC: Wiffin. RD. (1978). Investigations into
lead from motor vehicles. (AERE-R9198). Berkshire, England: Transportation and Road Research
Laboratory.
Chen. WP; Krage. N; Wu. LS: Page. AL; Chang. AC. (2008a). Fertilizer applications and trace elements
in vegetable production soils of California. Water Air Soil Pollut 190: 209-219.
http://dx.doi.org/10.1007/sll270-007-9594-7
Chen. WP: Krage. N; Wu. LS: Pan. GX; Khosrivafard. M; Chang. AC. (2008b). Arsenic, cadmium, and
lead in California cropland soils: Role of phosphate and micronutrient fertilizers. J Environ Qual 37:
689-695. http://dx.doi.org/10.2134/iea2007.0444
Cheng. Y; Schwartz. J: Sparrow. D: Aro. A; Weiss. ST; Hu. H. (2001). Bone lead and blood lead levels in
relation to baseline blood pressure and the prospective development of hypertension: The Normative
Aging Study. Am J Epidemiol 153: 164-171. http://dx.doi.Org/10.1093/aje/153.2.164
November 2012 4-149 Draft - Do Not Cite or Quote
-------
Chia. SE; Zhou. HJ; Yap. E; Tham. MT; Dong. NV; Hong Tu. NT; Chia. KS. (2006). Association of renal
function and delta-aminolevulinic acid dehydratase polymorphism among Vietnamese and Singapore
workers exposed to inorganic lead. Occup Environ Med 63: 180-186.
http://dx.doi.org/10.1136/oem.2005.021154
Chiang. WF: Yang. HJ: Lung. SCC: Huang. S: Chiu. CY: Liu. IL: Tsai. CL: Kuo. CY. (2008). A
comparison of elementary schoolchildren's exposure to arsenic and lead. J Environ Sci Health C
Environ Carcinog Ecotoxicol Rev 26: 237-255. http://dx.doi.org/10.1080/10590500802343958
Christoffersson. JO; Ahlgren. L; Schwartz. A; Skerfving. S: Mattsson. S. (1986). Decrease of skeletal lead
levels in man after end of occupational exposure. Arch Environ Occup Health 41: 312-318.
http://dx.doi.org/10.1080/00039896.1986.9936703
Chuang. HY; Schwartz. J; Tsai. SY; M-LT. L; Wang. JD; Hu. H. (2000). Vibration perception thresholds
in workers with long term exposure to lead. Occup Environ Med 57: 588-594.
Cimorelli. AJ; Perry. SG: Venkatram. A; Weil. JC: Paine. R; Wilson. RB; Lee. RF; Peters. WD; Erode.
RW. (2005). AERMOD: A dispersion model for industrial source applications. Part I: General model
formulation and boundary layer characterization. J Appl Meteorol 44: 682-693.
http://dx.doi.Org/10.1175/JAM2227.l
Clark. HF; Brabander. DJ; Erdil. RM. (2006). Sources, sinks, and exposure pathways of lead in urban
garden soil. J Environ Qual 35: 2066-2074. http://dx.doi.org/10.2134/ieq2005.0464
Clayton. CA; Pellizzari. ED; Whitmore. RW: Perritt RL Quackenboss. JJ. (1999). National human
exposure assessment survey (NHEXAS): Distributions and associations of lead, arsenic, and volatile
organic compounds in EPA Region 5. J Expo Sci Environ Epidemiol 9: 381-392.
http://dx.doi.org/10.1038/sj.jea.7500055
Coon. S: Stark. A; Peterson. E; Gloi. A; Kortsha. G: Pounds. J: Chettle. D; Gorell. J. (2006). Whole-body
lifetime occupational lead exposure and risk of Parkinson's disease. Environ Health Perspect 114:
1872-1876. http://dx.doi.org/10.1289/ehp.9102
Cornelis. C: Berghmans. P; van Sprundel. M; Van der Auwera. JC. (2006). Use of the IEUBK model for
determination of exposure routes in view of site remediation. Hum Ecol Risk Assess 12: 963-982.
http://dx.doi.org/10.1080/10807030600826979
Costa de Almeida. GR; de Freitas Tavares. CF; de Souza. AM; Sampaio de Sousa. T; Rodrigues
Funavama. CA; Barbosa. F. Jr; Tanus-Santos. JE; Gerlach. RF. (2010). Whole blood, serum, and saliva
lead concentrations in 6- to 8-year-old children. Sci Total Environ 408: 1551-1556.
http://dx.doi.0rg/10.1016/i.scitotenv.2009.12.034
Costa de Almeida. GR: de Sousa Guerra. C: de Angelo Souza Leite. G: Antonio. RC: Barbosa. F; Tanus-
Santos. JE; Gerlach. RF. (2011). Lead contents in the surface enamel of primary and permanent teeth,
whole blood, serum, and saliva of 6- to 8-year-old children. Sci Total Environ 409: 1799-1805.
http://dx.doi.0rg/10.1016/i.scitotenv.2011.01.004
Costa de Almeida. GR: Pereira Saraiva Mda. C: Barbosa F. J..r; Krug. FJ; Cury. JA; Rosario de Sousa
Mda. L; Rabelo Buzalaf. MA; Gerlach. RF. (2007). Lead contents in the surface enamel of deciduous
teeth sampled in vivo from children in uncontaminated and in lead-contaminated areas. Environ Res
104: 337-345. http://dx.doi.Org/10.1016/i.envres.2007.03.007
Costa de Almeida. GR: Umbelino de Freitas. C: Barbosa. F. Jr; Tanus-Santos. JE; Gerlach. RF. (2009).
Lead in saliva from lead-exposed and unexposed children. Sci Total Environ 407: 1547-1550.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.10.058
Davies. BE: White. HM. (1981). ENVIRONMENTAL-POLLUTION BY WIND BLOWN LEAD MINE
WASTE - A CASE-STUDY IN WALES, U.K.. Sci Total Environ 20: 57-74.
http://dx.doi.org/10.1016/0048-9697(81)90036-X
November 2012 4-150 Draft - Do Not Cite or Quote
-------
de Almeida. GRC: de Souza Guerra. C: Tanus-Santos. JE; Barbosa. F. Jr; Gerlach. RF. (2008). A plateau
detected in lead accumulation in subsurface deciduous enamel from individuals exposed to lead may
be useful to identify children and regions exposed to higher levels of lead. Environ Res 107: 264-270.
http://dx.doi.0rg/10.1016/i.envres.2008.01.002
Del Rio-Celestino. M; Font. R; Moreno-Roj as. R; De Haro-Bailon. A. (2006). Uptake of lead and zinc by
wild plants growing on contaminated soils. Ind Crop Prod 24: 230-237.
http://dx.doi.0rg/10.1016/i.indcrop.2006.06.013
DeSilva. PE. (1981). Determination of lead in plasma and studies on its relationship to lead in
erythrocytes. Br J Ind Med 38: 209-217.
Diamond. GL. (1992). Review of default value for lead plasma-to-urine transfer coefficient (TPLUR) in
the U.S. EPAuptake/biokinetic model. (SRC TR-92-135). Syracuse, NY: Syracuse Research
Corporation.
Diamond. GL. (1988). Biological monitoring of urine for exposure to toxic metals. In TW Clarkson; L
Friberg; GF Nordberg; PR Sager (Eds.), Biological monitoring of toxic metals (pp. 515-529). New
York, NY: Plenum Press.
Dietrich. KN; Berger. OG: Succop. PA. (1993 a). Lead exposure and the motor developmental status of
urban six-year-old children in the Cincinnati Prospective Study. Pediatrics 91: 301-307.
Dixon. SL; Gaitens. JM; Jacobs. DE; Strauss. W: Nagaraia. J: Pivetz. T; Wilson. JW: Ashley. PJ. (2009).
Exposure of US children to residential dust lead, 1999-2004: II. The contribution of lead-contaminated
dust to children's blood lead levels. Environ Health Perspect 117: 468-474.
http://dx.doi.org/10.1289/ehp.11918
Dorsev. CD: Lee. BK: Bolla. KI: Weaver. VM: Lee. SS: Lee. GS: Todd. AC: Shi. W: Schwartz. BS.
(2006). Comparison of patella lead with blood lead and tibia lead and their associations with
neurobehavioral test scores. J Occup Environ Med 48: 489-496.
http://dx.doi.org/10.1097/01.iom.0000199678.86629.3b
Drasch. G: Wanghofer. E; Roider. G. (1997). Are blood, urine, hair, and muscle valid biomonitors for the
internal burden of men with the heavy metals mercury, lead and cadmium? An investigation on 150
deceased. Trace Elem Electroly 14: 116-123.
Drasch. GA; Bohm. J: Baur. C. (1987). Lead in human bones. Investigations on an occupationally non-
exposed population in southern Bavaria (F.R.G.): I. Adults. Sci Total Environ 64: 303-315.
http://dx.doi.org/10.1016/0048-9697(87)90252-X
Drexler. JW: Brattin. WJ. (2007). An in vitro procedure for estimation of lead relative bioavailability:
With validation. Hum Ecol Risk Assess 13: 383-401. http://dx.doi.org/10.1080/10807030701226350
Duggan. MJ; Inskip. MJ; Rundle. SA; Moorcroft. JS. (1985). Lead in playground dust and on the hands
of schoolchildren. Sci Total Environ 44: 65-79. http://dx.doi.org/10.1016/0048-9697(85)90051-8
DuVal. G: Fowler. BA. (1989). Preliminary purification and characterization studies of a low molecular
weight, high affinity cytosolic lead-binding protein in rat brain. Biochem Biophys Res Commun 159:
177-184. http://dx.doi.org/10.1016/0006-291X(89)92420-0
Eaton. PL; Stacev. NH; Wong. KL; Klaassen. CD. (1980). Dose-response effects of various metal ions on
rat liver metallothionein, glutathione, heme oxygenase, and cytochrome P-450. Toxicol Appl
Pharmacol 55: 393-402.
Edwards. M; Dudi. A. (2004). Role of chlorine and chloramine in corrosion of lead-bearing plumbing
materials. J Am Water Works Assoc 96: 69-81.
Egan. SK; SS-H. T; Pennington. JAT; Bolger. PM. (2002). US Food and Drug Administration's Total Diet
Study: intake of nutritional and toxic elements, 1991-96. Food Addit Contam 19: 103-125.
Egeghy. PP; Quackenboss. JJ: Catlin. S: Ryan. PB. (2005). Determinants of temporal variability in
NHEXAS-Maryland environmental concentrations, exposures, and biomarkers. J Expo Sci Environ
Epidemiol 15: 388-397. http://dx.doi.org/10.1038/sj.jea.7500415
November 2012 4-151 Draft - Do Not Cite or Quote
-------
Eisner. AD; Rosati. J: Wiener. R. (2010). Experimental and theoretical investigation of particle-laden
airflow under a prosthetic mechanical foot in motion. Build Environ 45: 878-886.
http://dx.doi.0rg/10.1016/i.buildenv.2009.09.007
Elias. SM; Hashim. Z; Marian. ZM; Abdullah. AS: Hashim. JH. (2007). Relationship between blood lead
concentration and nutritional status among Malay primary school children In Kuala Lumpur,
Malaysia. Asia Pac J Public Health 19: 29-37. http://dx.doi.org/10.1177/101053950701900306
Elmarsafawv. SF; Jain. NB; Schwartz. J; Sparrow. D; Nie. HL; Hu. H. (2006). Dietary calcium as a
potential modifier of the relationship of lead burden to blood pressure. Epidemiology 17: 531-537.
http://dx.doi.org/10.1097/01.ede.0000231285.86968.2b
EPRI (Electric Power Research Institute). (1997). Results of the independent evaluation of ISCST3 and
ISC-PRIME. (TR-2460026 WO3527-02). Palo Alto, CA.
ESRI (Environmental Systems Research Institute Inc.). (2011). Updated Demographics Data.
http://www.esri.com/data/esri data/demographic.html
Esteban. E; Rubin. CH; Jones. RL; Noonan. G. (1999). Hair and blood substrates for screening children
for lead poisoning. Arch Environ Occup Health 54: 436-440.
Ettinger. AS: Lamadrid-Figueroa. H; Tellez-Rojo. MM; Mercado-Garcia. A; Peterson. KE; Schwartz. J:
Hu. H; Hernandez-Avila. M. (2009). Effect of calcium supplementation on blood lead levels in
pregnancy: A randomized placebo-controlled trial. Environ Health Perspect 117: 26-31.
http://dx.doi.org/10.1289/ehp.11868
Ettinger. AS: Tellez-Rojo. MM; Amarasiriwardena. C: Bellinger. D; Peterson. K; Schwartz. J: Hu. H;
Hernandez-Avila. M. (2004a). Effect of breast milk lead on infant blood lead levels at 1 month of age.
Environ Health Perspect 112: 1381-1385.
Ettinger. AS: Tellez-Rojo. MM; Amarasiriwardena. C: Gonzalez-Cossio. T; Peterson. KE; Aro. A; Hu. H;
Hernandez-Avila. M. (2004b). Levels of lead in breast milk and their relation to maternal blood and
bone lead levels at one month postpartum. Environ Health Perspect 112: 926-931.
Ettinger. AS: Tellez-Rojo. MM; Amarasiriwardena. C: Peterson. KE: Schwartz. J: Aro. A; Hu. H;
Hernandez-Avila. M. (2006). Influence of maternal bone lead burden and calcium intake on levels of
lead in breast milk over the course of lactation. Am J Epidemiol 163: 48-56.
http://dx.doi.org/10.1093/aie/kwj010
FAA (U.S. Federal Aviation Administration). (2010). General aviation and part 135 activity surveys - CY
2010. http://www.faa.gov/data research/aviation data statistics/general aviation/CY2010/
Falta. T; Limbeck. A; Koellensperger. G: Hann. S. (2008). Bioaccessibility of selected trace metals in
urban PM2.5 and PM10 samples: A model study. Anal Bioanal Chem 390: 1149-1157.
http://dx.doi.org/10.1007/s00216-007-1762-5
FDA (U.S. Food and Drug Administration). (2008). Total diet study.
http://www.fda.gov/Food/FoodSafetv/FoodContaminantsAdulteration/TotalDietStudv/default.htm
Fernandez. C: Labanowski. J: Cambier. P; Jongmans. AG: Van Port. F. (2007). Fate of airborne metal
pollution in soils as related to agricultural management. 1. Zn and Pb distributions in soil profiles. Eur
J Soil Sci 58: 547-559. http://dx.doi.Org/10.llll/j.1365-2389.2006.00827.x
Fernandez. C: Labanowski. J: Jongmans. T; Bermond. A; Cambier. P; Lamy. I; Van Port. F. (2010). Fate
of airborne metal pollution in soils as related to agricultural management: 2. Assessing the role of
biological activity in micro-scale Zn and Pb distributions in A, B and C horizons. Eur J Soil Sci 61:
514-524. http://dx.doi.0rg/10.llll/j.1365-2389.2010.01256.x
Fernandez. C: Monna. F; Labanowski. J: Loubet. M; van Port. F. (2008). Anthropogenic lead distribution
in soils under arable land and permanent grassland estimated by Pb isotopic compositions. Environ
Pollut 156: 1083-1091. http://dx.doi.Org/10.1016/i.envpol.2008.04.014
November 2012 4-152 Draft - Do Not Cite or Quote
-------
Flegal. AR; Smith. DR. (1995). Measurements of environmental lead contamination and human exposure.
In GW Ware (Ed.), Reviews of environmental contamination and toxicology, continuation of residue
reviews, v 143 (pp. 1-45). New York, NY: Springer.
Fleming. DEB: Boulav. D: Richard. NS: Robin. JP: Gordon. CL: Webber. CE: Chettle. DR. (1997).
Accumulated body burden and endogenous release of lead in employees of a lead smelter. Environ
Health Perspect 105: 224-233.
Forbes. GB; Reina. JC. (1972). Effect of age on gastrointestinal absorption (Fe, Sr, Pb) in the rat. J Nutr
102: 647-652.
Fosse. G: Wesenberg. GR; Tvinnereim HM Eide. R; Kristoffersen. O; Nag. OH; Wierzbicka. M; Banoczv.
J; De Oliveira. AA; Srisopak. C: Zamudio. A. (1995). Lead in deciduous teeth from larger cities of
some countries. Int J Environ Stud 47: 203-210.
Fowler. BA. (1989). Biological roles of high affinity metal-binding proteins in mediating cell injury.
Comments Toxicol 3: 27-46.
Fowler. BA; DuVal. G. (1991). Effects of lead on the kidney: Roles of high-affinity lead-binding proteins.
Environ Health Perspect 91: 77-80. http://dx.doi.org/10.1289/ehp.919177
Franklin. CA; Inskip. MJ; Baccanale. CL; Edwards. CM; Manton. WI; Edwards. E; O'Flahertv. EJ.
(1997). Use of sequentially administered stable lead isotopes to investigate changes in blood lead
during pregnancy in a nonhuman primate (Macaca fascicularis). Toxicol Sci 39: 109-119.
Freeman. GB; Dill. JA; Johnson. JD; Kurtz. PJ; Parham. F; Matthews. HB. (1996). Comparative
absorption of lead from contaminated soil and lead salts by weanling Fischer 344 rats. Toxicol Sci 33:
109-119.
Freeman. GB: Johnson. JD; Killinger. JM; Liao. SC: Feder. PI; Davis. AO; Ruby. MV; Chanev. RL;
Lovre. SC: Bergstrom. PP. (1992). Relative bioavailability of lead from mining waste soil in rats.
Toxicol Sci 19: 388-398.
Freeman. GB: Johnson. JD: Liao. SC: Feder. PI; Davis. AO: Ruby. MV: Schoof. RA; Chanev. RL;
Bergstrom. PP. (1994). Absolute bioavailability of lead acetate and mining waste lead in rats.
Toxicology 91: 151-163.
Fukui. Y; Miki. M; Ukai. H; Okamoto. S: Takada. S: Higashikawa. K; Ikeda. M. (1999). Urinary lead as a
possible surrogate of blood lead among workers occupationally exposed to lead. Int Arch Occup
Environ Health 72: 516-520. http://dx.doi.org/10.1007/s004200050409
Fullmer. CS: Rosen. JF. (1990). Effect of dietary calcium and lead status on intestinal calcium absorption.
Environ Res 51: 91-99.
Gaitens. JM; Pixon. SL; Jacobs. PE; Nagaraia. J: Strauss. W: Wilson. JW: Ashley. PJ. (2009). Exposure
of US children to residential dust lead, 1999-2004:1. Housing and demographic factors. Environ
Health Perspect 117: 461-467. http://dx.doi.org/10.1289/ehp.11917
Garavan. C: Breen. J: Moles. R; ORegan. B. (2008). A case study of the health impacts in an abandoned
lead mining area, using children's blood lead levels. International Journal of Mining, Reclamation and
Environment 22: 265-284. http://dx.doi.org/10.1080/17480930802109885
Garrido Latorre. F; Hernandez-Avila. M; Orozco. JT; Medina. CAA; Aro. A; Palazuelos. E; Hu. H.
(2003). Relationship of blood and bone lead to menopause and bone mineral density among middle-
age women in Mexico City. Environ Health Perspect 111: 631-636.
Georgopoulos. PG: Lioy. PJ. (2006). From theoretical aspects of human exposure and dose assessment to
computational model implementation: the Modeling ENvironment for TOtal Risk studies (MENTOR).
J Toxicol Environ Health B Crit Rev 9: 457-483.
Gerhardsson. L; Attewell. R: Chettle. PR; Englyst. V: Lundstrom. NG: Nordberg. GF; Nyhlin. H; Scott.
MC: Todd. AC. (1993). In vivo measurements of lead in bone in long-term exposed lead smelter
workers. Arch Environ Occup Health 48: 147-156.
November 2012 4-153 Draft - Do Not Cite or Quote
-------
Gerhardsson. L; Brune. D; Nordberg. GF; Wester. PO. (1986). Distribution of cadmium, lead and zinc in
lung, liver and kidney in long-term exposed smelter workers. Sci Total Environ 50: 65-85.
Gerhardsson. L; Englyst. V: Lundstrom. NG: Nordberg. G: Sandberg. S: Steinvall. F. (1995). Lead in
tissues of deceased lead smelter worker. J Trace Elem Med Biol 9: 136-143.
Gerke. TL; Scheckel. KG: Schock. MR. (2009). Identification and Distribution of Vanadinite
(Pb-5(V5+O4)3Cl) in Lead Pipe Corrosion By-Products. Environ Sci Technol 43: 4412-4418.
http://dx.doi.org/10.1021/es900501t
Ghosh. L; Adhikari. S: Ayyappan. S. (2007). Assessment of toxic interactions of heavy metals and their
effects on accumulation in tissues of freshwater fish. Research Journal of Environmental Toxicology
1: 37-44. http://dx.doi.org/10.3923/riet.2007.37.44
Glass. TA; Bandeen-Roche. K; McAtee. M; Bolla. K; Todd. AC: Schwartz. BS. (2009). Neighborhood
psychosocial hazards and the association of cumulative lead dose with cognitive function in older
adults. Am J Epidemiol 169: 683-692. http://dx.doi.org/10.1093/aie/kwn390
Glenn. BS: Bandeen-Roche. K; Lee. BK; Weaver. VM; Todd. AC: Schwartz. BS. (2006). Changes in
systolic blood pressure associated with lead in blood and bone. Epidemiology 17: 538-544.
http://dx.doi.org/10.1097/01.ede.0000231284.19078.4b
Glenn. BS: Stewart. WF; Links. JM; Todd. AC: Schwartz. BS. (2003). The longitudinal association of
lead with blood pressure. Epidemiology 14: 30-36.
Godoi. Q: Santos. D; Nunes. LC: Leme. FO; Rufini. IA; Agnelli. JAM; Trevizan. LC: Krug. FJ. (2009).
Preliminary studies of laser-induced breakdown spectrometry for the determination of Ba, Cd, Cr and
Pb in toys. Spectrochimica Acta Part B: Atomic Spectroscopy 64: 573-581.
http://dx.doi.0rg/10.1016/i.sab.2009.05.003
Gover. RA. (1990). Transplacental transport of lead. Environ Health Perspect 89: 101-105.
Granev. JR; Hallidav. AN; Keeler. GJ: Nriagu. JO: Robbins. JA; Norton. SA. (1995). Isotopic record of
lead pollution in lake sediments from the northeastern United States. Geochim Cosmo Act 59:
1715-1728.
Graziano. JH; Popovac. D: Factor-Litvak. P; Shrout P; Kline. J: Murphy. MJ; Zhao. YH; Mehmeti. A;
Ahmedi. X; Raiovic. B; Zvicer. Z; Nenezic. DU: Lolacono. NJ; Stein. Z. (1990). Determinants of
elevated blood lead during pregnancy in a population surrounding a lead smelter in Kosovo,
Yugoslavia. Environ Health Perspect 89: 95-100. http://dx.doi.org/10.1289/ehp.908995
Greenwav. JA; Gerstenberger. S. (2010). An evaluation of lead contamination in plastic toys collected
from day care centers in the Las Vegas Valley, Nevada, USA. Bull Environ Contam Toxicol 85:
363-366. http://dx.doi.org/10.1007/s00128-010-0100-3
Griffin. TB; Coulston. F; Wills. H; Russell. JC. (1975). Clinical studies on men continuously exposed to
airborne paniculate lead. In TB Griffin; JH Knelson (Eds.), Lead (pp. 221-240). Stuttgart, Germany:
Georg Thieme Publishers.
Gross. SB; Pfitzer. EA; Yeager. DW: Kehoe. RA. (1975). Lead in human tissues. Toxicol Appl Pharmacol
32: 638-651.
Gulson. B; Jameson. CW: Mahaffev. KR; Mizon. KJ; Patison. N; Law. AJ; Korsch. MJ: Salter. MA.
(1998a). Relationships of lead in breast milk to lead in blood, urine, and diet of the infant and mother.
Environ Health Perspect 106: 667-674. http://dx.doi.org/10.1289/ehp.98106667
Gulson. B; Korsch. M; Matisons. M; Douglas. C: Gillam. L; McLaughlin. V (2009). Windblown lead
carbonate as the main source of lead in blood of children from a seaside community: An example of
local birds as "canaries in the mine". Environ Health Perspect 117: 148-154.
http://dx.doi.org/10.1289/ehp.11577
Gulson. B; Mahaffev. KR; Jameson. CW: Mizon. KJ: Korsch. MJ: Cameron. MA; Eisman. JA. (1998b).
Mobilization of lead from the skeleton during the postnatal period is larger than during pregnancy.
TranslRes 131: 324-329.
November 2012 4-154 Draft - Do Not Cite or Quote
-------
Gulson. B; Mahaffev. KR: Jameson. CW: Patison. N; Law. AJ; Mizon. KJ; Korsch. MJ; Pederson. D.
(1999). Impact of diet on lead in blood and urine in female adults and relevance to mobilization of
lead from bone stores. Environ Health Perspect 107: 257-263. http://dx.doi.org/10.1289/ehp.99107257
Gulson. B; Mahaffev. KR; Mizon. KJ: Korsch. MJ: Cameron. MA; Vimpani. G. (1995a). Contribution of
tissue lead to blood lead in adult female subjects based on stable lead isotope methods. Transl Res
125: 703-712.
Gulson. B; Mizon. K; Korsch. M; Taylor. A. (2006a). Changes in the lead isotopic composition of blood,
diet and air in Australia over a decade: Globalization and implications for future isotopic studies.
Environ Res 100: 130-138. http://dx.doi.Org/10.1016/j.envres.2005.03.006
Gulson. B; Mizon. K; Smith. H; Eisman. J; Palmer. J; Korsch. M; Donnelly. J: Waite. K. (2002). Skeletal
lead release during bone resorption: Effect of bisphosphonate treatment in a pilot study. Environ
Health Perspect 110: 1017-1023.
Gulson. B; Mizon. K; Taylor. A; Korsch. M; Stauber. J: Davis. JM; Louie. H; Wu. M; Antin. L. (2008).
Longitudinal monitoring of selected elements in blood of healthy young children. J Trace Elem Med
Biol 22: 206-214. http://dx.doi.Org/10.1016/i.itemb.2008.04.001
Gulson. B; Mizon. KJ: Korsch. MJ: Palmer. JM; Donnelly. JB. (2003). Mobilization of lead from human
bone tissue during pregnancy and lactation: A summary of long-term research. Sci Total Environ 303:
79-104. http://dx.doi.org/10.1016/S0048-9697(02)00355-8
Gulson. B; Mizon. KJ: Korsch. MJ: Taylor. AJ. (2006b). Low blood lead levels do not appear to be
further reduced by dietary supplements. Environ Health Perspect 114: 1186-1192.
http://dx.doi.org/10.1289/ehp.8605
Gulson. B; Mizon. KJ: Palmer. JM: Korsch. MJ: Taylor. AJ: Mahaffev. KR. (2004a). Blood lead changes
during pregnancy and postpartum with calcium supplementation. Environ Health Perspect 112:
1499-1507. http://dx.doi.org/10.1289/ehp.6548
Gulson. B; Wilson. D. (1994). History of lead exposure in children revealed from isotopic analyses of
teeth. Arch Environ Occup Health 49: 279-283. http://dx.doi.org/10.1080/00039896.1994.9937480
Gulson. BL; Davis. JJ: Mizon. KJ: Korsch. MJ: Bawden-Smith. J. (1995b). Sources of lead in soil and
dust and the use of dust fallout as a sampling medium. Sci Total Environ 166: 245-262.
http://dx.doi.org/10.1016/0048-9697(95)04505-U
Gulson. BL; Jameson. CW: Mahaffev. KR: Mizon. KJ: Korsch. MJ: Vimpani. G. (1997). Pregnancy
increases mobilization of lead from maternal skeleton. Transl Res 130: 51-62.
http://dx.doi.org/10.1016/S0022-2143(97)90058-5
Gulson. BL; Mizon. KJ: Davis. JD; Palmer. JM: Vimpani. G. (2004b). Identification of sources of lead in
children in a primary zinc-lead smelter environment. Environ Health Perspect 112: 52-60.
http://dx.doi.org/10.1289/ehp.6465
Gultepe. I: Pearson. G: Milbrandt. JA; Hansen. B; Platnick. S: Taylor. P; Gordon. M; Oakley. JP; Cober.
SG. (2009). The fog remote sensing and modeling field project. Bull Am Meteorol Soc 90: 341-359.
httrj://dx.doi.org/10.1175/2008bams2354.1
Haley. VB; Talbot TO. (2004). Seasonality and trend in blood lead levels of New York State children.
BMC Pediatr 4:8. http://dx.doi.org/10.1186/1471-2431-4-8
Hanninen. H; Aitio. A; Kovala. T; Luukkonen. R; Matikainen. E; Mannelin. T; Erkkila. J: Riihimaki. V.
(1998). Occupational exposure to lead and neuropsychological dysfunction. Occup Environ Med 55:
202-209.
Hansen. S: Nieboer. E; Sandanger. TM; Wilsgaard. T; Thomassen. Y; Veyhe. AS: Odland. JO. (20lib).
Changes in maternal blood concentrations of selected essential and toxic elements during and after
pregnancy. J Environ Monit 13: 2143-2152. http://dx.doi.org/10.1039/cleml0051c
Havlena. J: Kanarek. MS: Coons. M. (2009). Factors associated with the seasonality of blood lead levels
among preschool Wisconsin children. WMJ 108: 151-155.
November 2012 4-155 Draft - Do Not Cite or Quote
-------
Haves. EB; McElvaine. MD; Orbach. HG: Fernandez. AM; Lyne. S: Matte. TD. (1994). Long-term trends
in blood lead levels among children in Chicago: Relationship to air lead levels. Pediatrics 93: 195-200.
Healev. N; Chettle. PR; Mcneill. FE; Fleming. DE. (2008). Uncertainties in the relationship between tibia
lead and cumulative blood lead index [Letter]. Environ Health Perspect 116: A109.
http://dx.doi.org/10.1289/ehp. 10778
Healv. MA; Harrison. PG: Aslam. M; Davis. SS: Wilson. CG. (1992). Lead sulphide and traditional
preparations: Routes for ingestion, and solubility and reactions in gastric fluid. J Clin Hosp Pharm 7:
169-173. http://dx.doi.0rg/10.llll/j.1365-2710.1982.tb01019.x
Heard. MJ; Chamberlain. AC. (1982). Effect of minerals and food on uptake of lead from the
gastrointestinal tract in humans. Hum Exp Toxicol 1: 411-415.
http://dx.doi.org/10.1177/096032718200100407
Heard. MJ: Wells. AC: Newton. D; Chamberlain. AC. (1979). Human uptake and metabolism of terra
ethyl and tetra methyl lead vapour labelled with 203Pb. In International Conference on Management
and Control of Heavy Metals in the Environment, London, England, September. Edinburgh, United
Kingdom: CEP Consultants, Ltd.
Hepp. NM; Mindak. WR; Cheng. J. (2009). Determination of total lead in lipstick: Development and
validation of a microwave-assisted digestion, inductively coupled plasma-mass spectrometric method.
J Cosmet Sci 60: 405-414. http://dx.doi.Org/10.llll/i.1468-2494.2010.00577 2.x
Hernandez-Avila. M; Gonzalez-Cossio. T; Palazuelos. E; Romieu. I; Aro. A; Fishbein. E; Peterson. KE;
Hu. H. (1996). Dietary and environmental determinants of blood and bone lead levels in lactating
postpartum women living in Mexico City. Environ Health Perspect 104: 1076-1082.
http://dx.doi.org/10.1289/ehp.961041076
Hernandez-Avila. M; Smith. D; Meneses. F; Sanin. LH; Hu. H. (1998). The influence of bone and blood
lead on plasma lead levels in environmentally exposed adults. Environ Health Perspect 106: 473-477.
http://dx.doi.org/10.1289/ehp.98106473
Hernberg. S: Nikkanen. J: Mellin. G: Lilius. H. (1970). Delta-aminolevulinic acid dehydrase as a measure
of lead exposure. Arch Environ Occup Health 21: 140-145.
Hertz-Picciotto. I; Schramm. M; Watt-Morse. M; Chantala. K; Anderson. J: Osterloh. J. (2000). Patterns
and determinants of blood lead during pregnancy. Am J Epidemiol 152: 829-837.
http://dx.doi.0rg/10.1093/aie/152.9.829
Hicsonmez. U; Erees. FS: Ozdemir. C: Ozdemir. A; Cam. S. (2009). Determination of major and minor
elements in the Malva sylvestris L. from Turkey using ICP-OES techniques. Biol Trace Elem Res 128:
248-257. http://dx.doi.org/10.1007/sl2011-008-8270-0
Hilts. SR. (1996). A co-operative approach to risk management in an active lead/zinc smelter community.
Environ Geochem Health 18: 17-24. http://dx.doi.org/10.1007/BF01757215
Hilts. SR. (2003). Effect of smelter emission reductions on children's blood lead levels. Sci Total Environ
303: 51-58. http://dx.doi.org/10.1016/S0048-9697(02)00357-l
Hilts. SR: Bock. SE; Oke. TL; Yates. CL; Copes. RA. (1998). Effect of interventions on children's blood
lead levels. Environ Health Perspect 106: 79-83. http://dx.doi.org/10.1289/ehp.9810679
Hopkins. MR; Ettinger. AS: Hernandez-Avila. M; Schwartz. J: Tellez-Rojo. MM; Lamadrid-Figueroa. H;
Bellinger. D; Hu. H; Wright. RO. (2008). Variants in iron metabolism genes predict higher blood lead
levels in young children. Environ Health Perspect 116: 1261-1266.
http://dx.doi.org/10.1289/ehp.11233
Hsieh. TJ: Chen. YC: Li. CW: Liu. GC: Chiu. YW: Chuang. HY. (2009b). A proton magnetic resonance
spectroscopy study of the chronic lead effect on the basal ganglion and frontal and occipital lobes in
middle-age adults. Environ Health Perspect 117: 941-945. http://dx.doi.org/10.1289/ehp.0800187
November 2012 4-156 Draft - Do Not Cite or Quote
-------
Hu. H; Aro. A; Pavton. M; Korrick. S: Sparrow. (1996a). The relationship of bone and blood lead to
hypertension: The Normative Aging Study. JAMA 275: 1171-1176.
http://dx.doi.org/10.1001/iama.1996.03530390037031
Hu. H; Aro. A; Rotnitzky. A. (1995). Bone lead measured by X-ray fluorescence: Epidemiologic methods.
Environ Health Perspect 103: 105-110. http://dx.doi.org/10.1289/ehp.95103sll05
Hu. H; Rabinowitz. M; Smith. D. (1998). Bone lead as a biological marker in epidemiologic studies of
chronic toxicity: Conceptual paradigms [Review]. Environ Health Perspect 106: 1-8.
http://dx.doi.org/10.1289/ehp.981061
Hu. H; Shih. R; Rothenberg. S: Schwartz. BS. (2007a). The epidemiology of lead toxicity in adults:
Measuring dose and consideration of other methodologic issues [Review]. Environ Health Perspect
115: 455-462. http://dx.doi.org/10.1289/ehrj.9783
Hunt. A; Johnson. PL. (2012). Suspension and resuspension of dry soil indoors following track-in on
footwear. Environ Geochem Health 34: 355-363. http://dx.doi.org/10.1007/sl0653-011-9400-8
Hunt. A; Johnson. PL; Brooks. J: Griffith. DA. (2008). Risk remaining from fine particle contaminants
after vacuum cleaning of hard floor surfaces. Environ Geochem Health 30: 597-611.
http://dx.doi.org/10.1007/sl0653-008-9183-8
Hunt. WG: Burnham. W: Parish. CN; Burnham. KK; Mutch. B; Oaks. JL. (2006). Bullet fragments in
deer remains: Implications for lead exposure in avian scavengers. Wildlife Society Bulletin 34:
167-170. http://dx.doi.org/10.2193/0091-7648(2006)34ri67:BFIDPJ12.0.CO:2
Hunt. WG: Watson. RT: Oaks. JL: Parish. CN: Burnham. KK: Tucker. RL: Belthoff. JR: Hart. G. (2009).
Lead bullet fragments in venison from rifle-killed deer: Potential for human dietary exposure. PLoS
ONE 4: e5330. http://dx.doi.org/10.1371/iournal.pone.0005330
Hursh. JB; Schraub. A; Sattler. EL; Hofmann. HP. (1969). Fate of 212Pb inhaled by human subjects.
Health Phys 16: 257-267.
Hursh. JB; Suomela. J. (1968). Absorption of 212Pb from the gastrointestinal tract of man. Acta Radiol 7:
108-120. http://dx.doi.org/10.3109/02841866809133184
ICRP (International Commission on Radiological Protection). (1973). Alkaline earth metabolism in adult
man. Oxford, U.K.: Pergamon Press.
ICRP (International Commission on Radiological Protection). (1993). Age-specific biokinetic models for
the alkaline earth elements and lead. In Age-dependent doses to members of the public from intake of
radionuclides: Part 2: Ingestion dose coefficients (pp. 95-120). Elsevier: New York.
ICRP (International Commission on Radiological Protection). (1994). Human respiratory tract model for
radiological protection: A report of a task group of the International Commission on Radiological
Protection. ICRP Publication 66 [Review]. Ann ICRP 24: 1-482.
Inskip. MJ; Franklin. CA; Baccanale. CL; Manton. WI; O'Flahertv. EJ; Edwards. CMH; Blenkinsop. JB:
Edwards. EB. (1996). Measurement of the flux of lead from bone to blood in a nonhuman primate
(Macaca fascicularis) by sequential administration of stable lead isotopes. Toxicol Sci 33: 235-245.
http://dx.doi.org/10.1006/faat.1996.0161
Jackson. LW: Cromer. BA; Panneerselvam. A. (2010). Association between bone turnover, micronutrient
intake and blood lead levels among pre- and post-menopausal women, NHANES 1999-2002. Environ
Health Perspect 118: 1590-1596. http://dx.doi.org/10.1289/ehp.1002158
Jaffe. E; Volin. M; Bronson-Mullins. C: Dunbrack. R; Kervinen. J: Martins. J: Quinlan. J: Sazinsky. M;
Steinhouse. E; Yeung. A. (2000). An artificial gene for human porphobilinogen synthase allows
comparison of an allelic variation implicated in susceptibility to lead poisoning. J Biol Chem 275:
2619- 2626. http://dx.doi.Org/10.1074/ibc.275.4.2619
Jain. NB; Potula. V: Schwartz. J: Vokonas. PS: Sparrow. D; Wright. RO: Nie. H; Hu. H. (2007). Lead
levels and ischemic heart disease in a prospective study of middle-aged and elderly men: The VA
Normative Aging Study. Environ Health Perspect 115: 871-875. http://dx.doi.org/10.1289/ehp.9629
November 2012 4-157 Draft - Do Not Cite or Quote
-------
James. HM; Hilburn. ME; Blair. JA. (1985). Effects of meals and meal times on uptake of lead from the
gastrointestinal tract of humans. Hum Exp Toxicol 4: 401-407.
Jin. C: Li. Y; Li. YL; Zou. Y; Zhang. GL; Normura. M; Zhu. GY. (2008). Blood lead: Its effect on trace
element levels and iron structure in hemoglobin. Nucl Instrum Methods Phys Res B 266: 3607-3613.
http://dx.doi.0rg/10.1016/i.nimb.2008.05.087
Jin. CW: Zhang. SJ: He. YF; Zhou. GD; Zhou. ZX. (2005). Lead contamination in tea garden soils and
factors affecting its bioavailability. Chemosphere 59: 1151-1159.
http://dx.doi.0rg/10.1016/i.chemosphere.2004.ll.058
Johansen. P; Pedersen. HS: Asmund. G: Riget. F. (2006). Lead shot from hunting as a source of lead in
human blood. Environ Pollut 142: 93-97. http://dx.doi.Org/10.1016/i.envpol.2005.09.015
Johnson. D; Bretsch. J. (2002). Soil lead and children's blood lead levels in Syracuse, NY, USA. Environ
Geochem Health 24: 375-385. http://dx.doi.Org/10.1023/A:1020500504167
Johnson. PL; McDade. K; Griffith. D. (1996). Seasonal variation in paediatric blood lead levels in
Syracuse, NY, USA. Environ Geochem Health 18: 81-88. http://dx.doi.org/10.1007/BF01771136
Jones. RL; Homa. DM; Meyer. PA; Brodv. DJ: Caldwell. KL; Pirkle. JL; Brown. MJ. (2009a). Trends in
blood lead levels and blood lead testing among US children aged 1 to 5 Years, 1988-2004. Pediatrics
123: e376-e385. http://dx.doi.org/10.1542/peds.2007-3608
Juhasz. AL; Weber. J; Smith. E. (2011). Impact of soil particle size and bioaccessibility on children and
adult lead exposure in peri-urban contaminated soil. J Hazard Mater 186: 1870-1879.
http://dx.doi.0rg/10.1016/i.jhazmat.2010.12.095
Juhasz. AL; Weber. J: Smith. E; Naidu. R; Marschner. B; Rees. M; Rofe. A; Kuchel. T; Sansom. L.
(2009). Evaluation of SBRC-gastric and SBRC-intestinal methods for the prediction of in vivo relative
lead bioavailability in contaminated soils. Environ Sci Technol 43: 4503-4509.
http://dx.doi.org/10.1021/es803238u
Kamel. F; Umbach. DM; Hu. H; Munsat. TL; Shefner. JM; Taylor. JA; Sandier. DP. (2005). Lead
exposure as a risk factor for amyotrophic lateral sclerosis. Neurodegener Dis 2: 195-201.
http://dx.doi.org/10.1159/000089625
Kamel. F; Umbach. DM; Munsat. TL; Shefner. JM; Hu. H; Sandier. DP. (2002). Lead exposure and
amyotrophic lateral sclerosis. Epidemiology 13: 311-319.
Kamel. F; Umbach. DM; Stallone. L; Richards. M; Hu. H; Sandier. DP. (2008). Association of lead
exposure with survival in amyotrophic lateral sclerosis. Environ Health Perspect 116: 943-947.
http://dx.doi.org/10.1289/ehp.11193
Kang. HG: Jeong. SH; Cho. MR; Cho. JH; Bischoff. K. (2009). Time-dependent changes in lead and
delta-aminolevulinic acid after subchronic lead exposure in rats. Hum Exp Toxicol 28: 647-654.
http://dx.doi.org/10.1177/0960327109107046
Kehoe. RA. (196la). The Harben Lectures, 1960: The metabolism of lead in man in health and disease. 1.
The normal metabolism of lead. J R Inst Public Health 24: 81-97.
Kehoe. RA. (1961b). The Harben Lectures, 1960: The metabolism of lead in man in health and disease.
2(2). The metabolism of lead under abnormal conditions. J R Inst Public Health 24: 129-143.
Kehoe. RA. (1961c). The Harben Lectures, 1960: The metabolism of lead in man in health and disease. 3.
Present hygienic problems relating to the absorption of lead. J R Inst Public Health 24: 177-203.
Kehoe. RA. (1987). Studies of lead administration and elimination in adult volunteers under natural and
experimentally induced conditions over extended periods of time. Food Chem Toxicol 25: 425-493.
Kemp. FW: Neti. PVS. V: Howell. RW: Wenger. P; Louria. DB; Bogden. JD. (2007). Elevated blood lead
concentrations and vitamin D deficiency in winter and summer in young urban children. Environ
Health Perspect 115: 630-635. http://dx.doi.org/10.1289/ehp.9389
November 2012 4-158 Draft - Do Not Cite or Quote
-------
Kessler. M; Durand. PY; Huu. TC: Royer-Morot MJ; Chanliau. J: Netter. P; Due. M. (1999).
Mobilization of lead from bone in end-stage renal failure patients with secondary
hyperparathyroidism. Nephrol Dial Transplant 14: 2731-2733.
http://dx.doi.org/10.1093/ndt/14.ll.2731
Khalil. N; Morrow. LA; Needleman. H; Talbott. EO; Wilson. JW: Caulev. JA. (2009a). Association of
cumulative lead and neurocognitive function in an occupational cohort. Neuropsychology 23: 10-19.
http://dx.doi.org/10.1037/a0013757
Khoder. MI; Hassan. SK; El-Abssawv. AA. (2010). An evaluation of loading rate of dust, Pb, Cd, and Ni
and metals mass concentration in the settled surface dust in domestic houses and factors affecting
them. Indoor Built Environ 19: 391-399. http://dx.doi.org/10.1177/1420326X10367284
Khourv. GA; Diamond. GL. (2003). Risks to children from exposure to lead in air during remedial or
removal activities at Superfund sites: a case study of the RSR lead smelter superfund site. J Expo Sci
Environ Epidemiol 13: 51-65. http://dx.doi.org/10.1038/sj.jea.7500254
Kim. EJ; Herrera. JE. (2010). Characteristics of lead corrosion scales formed during drinking water
distribution and their potential influence on the release of lead and other contaminants. Environ Sci
Technol 44: 6054-6061. http://dx.doi.org/10.1021/esl01328u
Kim. HK; Yoon. EK; Jang. J: Hwang. M; Kim. J: Ha. JH; Jang. DP; Yoo. TM; Park. KL. (2009a).
Assessment of heavy metal exposure via the intake of oriental medicines in Korea. J Toxicol Environ
HealthA72: 1336-1342. http://dx.doi.org/10.1080/15287390903212485
Kim. HS: Lee. SS: Lee. GS: Hwangbo. Y; Ahn. KD; Lee. BK. (2004). The protective effect of delta-
aminolevulinic acid dehydratase 1-2 and 2-2 isozymes against blood lead with higher hematologic
parameters. Environ Health Perspect 112: 538-541. http://dx.doi.org/10.1289/ehp.6464
Kim. R; Aro. A; Rotnitzky. A; Amarasiriwardena. C: Hu. H. (1995). K x-ray fluorescence measurements
of bone lead concentration: the analysis of low-level data. Phys Med Biol 40: 1475-1485.
http://dx.doi.Org/10.1088/0031-9155/40/9/007
Kim. R; Landrigan. C: Mossmann. P; Sparrow. D; Hu. H. (1997). Age and secular trends in bone lead
levels in middle-aged and elderly men: Three-year longitudinal follow-up in the Normative Aging
Study. Am J Epidemiol 146: 586-591.
Kissel. JC: Richter. KY; Fenske. RA. (1996). Field measurement of dermal soil loading attributable to
various activities: implications for exposure assessment. Risk Anal 16: 115-125.
Klotzback. JM; Follansbee. MH; Diamond. GL. (2003). Evaluation of the ICRP lead biokinetics model:
Empirical comparisons with observations of plasma-blood lead concentration relationships in humans
(draft final). Washington, DC: U.S. Environmental Protection Agency.
Kordas. K; Ettinger. AS: Lamadrid-Figueroa. H; Tellez-Rojo. MM; Hernandez-Avila. M; Hu. H; Wright.
RO. (2009). Methylenetetrahydrofolate reductase (MTHFR) C677T, A1298C and Gl 793 A genotypes,
and the relationship between maternal folate intake, tibia lead and infant size at birth. Br J Nutr 102:
907-914. http://dx.doi.org/10.1017/s0007114509318280
Korrick. SA; Hunter. DJ; Rotnitzky. A; Hu. H; Speizer. FE. (1999). Lead and hypertension in a sample of
middle-aged women. Am J Public Health 89: 330-335. http://dx.doi.Org/10.2105/AJPH.89.3.330
Korrick. SA; Schwartz. J: Tsaih. SW: Hunter. DJ: Aro. A; Rosner. B; Speizer. FE; Hu. H. (2002).
Correlates of bone and blood lead levels among middle-aged and elderly women. Am J Epidemiol
156: 335-343. http://dx.doi.org/10.1093/aje/kwf042
Koster. J: Erhardt. A; Stoeppler. M; Mohl. C: Ritz. E. (1989). Mobilizable lead in patients with chronic
renal failure. Eur J Clin Invest 19: 228-233. http://dx.doi.Org/10.llll/i.1365-2362.1989.tb00222.x
Kostial. K; Kello. D; Jugo. S: Rabar. I; Maljkovic. T. (1978). Influence of age on metal metabolism and
toxicity. Environ Health Perspect 25: 81-86. http://dx.doi.org/10.1289/ehp.782581
November 2012 4-159 Draft - Do Not Cite or Quote
-------
Koyashiki. GA; Paoliello. MM; Matsuo. T; de Oliveira. MM; Mezzaroba. L; de Fatima Carvalho. M;
Momoyo Sakuma. A; Turini. C: Terezinha Oliveira Vannuchi. M; Barbosa. CS. (2010). Lead levels in
milk and blood from donors to the breast milk bank in Southern Brazil. Environ Res 110: 265-271.
http://dx.doi.0rg/10.1016/i.envres.2009.12.001
Krell. U: Roeckner. E. (1988). Model simulation of the atmospheric input of lead and cadmium into the
North Sea. Atmos Environ 22: 375-381. http://dx.doi.org/10.1016/0004-6981(88)90043-l
Krieg. EF. Jr; Butler. MA; Chang. MH; Liu. TB; Yesupriya. A; Lindegren. ML; Dowling. N. (2009). Lead
and cognitive function in ALAD genotypes in the Third National Health and Nutrition Examination
Survey. Neurotoxicol Teratol 31: 364-371. http://dx.doi.Org/10.1016/i.ntt.2009.08.003
Kumar. A; Pastore. P. (2007). Lead and cadmium in soft plastic toys. Curr Sci 93: 818-822.
Lacev. RF; Moore. MR; Richards. WN. (1985). Lead in water, infant diet and blood: The Glasgow
Duplicate Diet Study. Sci Total Environ 41: 235-257.
Lagerkvist BJ; Ekesrydh. S: Englyst. V: Nordberg. GF; Soderberg. HA; Wiklund. DE. (1996). Increased
blood lead and decreased calcium levels during pregnancy: A prospective study of Swedish women
living near a smelter. Am J Public Health 86: 1247-1252.
Lai. JS: Wu. TN: Liou. SH: Shen. CY: Guu. CF: Ko. KN: Chi. HY: Chang. PY. (1997). A study of the
relationship between ambient lead and blood lead among lead battery workers. Int Arch Occup
Environ Health 69: 295-300.
Laidlaw. MAS: Mielke. HW: Filippelli. GM; Johnson. PL; Gonzales. CR. (2005). Seasonality and
children's blood lead levels: Developing a predictive model using climatic variables and blood lead
data from Indianapolis, Indiana, Syracuse, New York, and New Orleans, Louisiana (USA). Environ
Health Perspect 793-800: 113. http://dx.doi.org/10.1289/ehp.7759
Laidlaw. MAS: Zahran. S: Mielke. HW: Taylor. MP; Filippelli. GM. (2012). Re-suspension of lead
contaminated urban soil as a dominant source of atmospheric lead in Birmingham, Chicago, Detroit
and Pittsburgh, USA. Atmos Environ 49: 302-310. http://dx.doi.Org/10.1016/i.atmosenv.2011.ll.030
Lamadrid-Figueroa. H; Tellez-Rojo. MM; Hernandez-Cadena. L; Mercado-Garcia. A; Smith. D; Solano-
Gonzalez. M; Hernandez-Avila. M; Hu. H. (2006). Biological markers of fetal lead exposure at each
stage of pregnancy. J Toxicol Environ Health A 69: 1781-1796.
http://dx.doi.org/10.1080/15287390600630195
Lanphear. BP; Hornung. R; Ho. M; Howard. CR: Eberly. S: Knauf. K. (2002). Environmental lead
exposure during early childhood. J Pediatr 140: 40-47.
Lanphear. BP; Matte. TD; Rogers. J: Clickner. RP; Dietz. B; Bornschein. RL; Succop. P; Mahaffev. KR;
Dixon. S: Galke. W: Rabinowitz. M; Farfel M; Rohde. C: Schwartz. J: Ashley. P; Jacobs. DE. (1998).
The contribution of lead-contaminated house dust and residential soil to children's blood lead levels: A
pooled analysis of 12 epidemiologic studies. Environ Res 79: 51-68.
http://dx.doi.org/10.1006/enrs.1998.3859
Lanphear. BP: Roghmann. KJ. (1997). Pathways of lead exposure in urban children. Environ Res 74:
67-73.
Lasheen. MR; Sharabv. CM; El-Kholy. NG: Elsherif. IY; El-Wakeel. ST. (2008). Factors influencing lead
and iron release from some Egyptian drinking water pipes. J Hazard Mater 160: 675-680.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.03.040
Lavton. DW: Beamer. PI. (2009). Migration of contaminated soil and airborne particulates to indoor dust.
Environ Sci Technol 43: 8199-8205. http://dx.doi.org/10.1021/es9003735
Lee. BK; Lee. GS: Stewart. WF; Ahn. KD; Simon. D; Kelsev. KT; Todd. AC: Schwartz. BS. (200la).
Associations of blood pressure and hypertension with lead dose measures and polymorphisms in the
vitamin D receptor and delta-aminolevulinic acid dehydratase genes. Environ Health Perspect 109:
383-389.
November 2012 4-160 Draft - Do Not Cite or Quote
-------
Leggett. RW. (1993). An age-specific kinetic model of lead metabolism in humans. Environ Health
Perspect 101: 598-616.
Levesque. B; Duchesne. JF; Gariepv. C: Rhainds. M; Dumas. P; Scheuhammer. AM; Proulx. JF; Dery. S:
Muckle. G: Dallaire. F; Dewailly. E. (2003). Monitoring of umbilical cord blood lead levels and
sources assessment among the Inuit. Occup Environ Med 60: 693-695.
Levin. R; Brown. MJ; Kashtock. ME; Jacobs. DE; Whelan. EA; Rodman. J; Schock. MR; Padilla. A;
Sinks. T. (2008). Lead exposures in US children, 2008: Implications for prevention. Environ Health
Perspect 116: 1285-1293. http://dx.doi.org/10.1289/ehp.11241
Lewin. MD; Sarasua. S: Jones. PA. (1999). A multivariate linear regression model for predicting
children's blood lead levels based on soil lead levels: A study at four superfund sites. Environ Res 81:
52-61. http://dx.doi.org/10.1006/enrs.1998.3952
Liang. F; Zhang. G: Tan. M; Yan. C: Li. X; Li. Y; Zhang. Y; Shan. Z. (2010). Lead in children's blood is
mainly caused by coal-fired ash after phasing out of leaded gasoline in Shanghai. Environ Sci Technol
44: 4760-4765. http://dx.doi.org/10.1021/es9039665
Lima. FD; do Nascimento. CWA; da Silva. FBV; de Carvalho. VGB; Ribeiro. MR. (2009). Lead
concentration and allocation in vegetable crops grown in a soil contaminated by battery residues.
Horticulture Brasileira 27: 362-365. http://dx.doi.org/10.1590/S0102-05362009000300019
Liu. J; McCaulev. L; Compher. C: Yan. C: Shen. X; Needleman. H; Pinto-Martin. JA. (2011 a). Regular
breakfast and blood lead levels among preschool children. Environ Health Global Access Sci Source
10: 28. http://dx.doi.org/10.1186/1476-069X-10-28
Ljung. K; Selinus. O: Otabbong. E; Berglund. M. (2006). Metal and arsenic distribution in soil particle
sizes relevant to soil ingestion by children. Appl Geochem 21: 1613-1624.
http://dx.doi.0rg/10.1016/i.apgeochem.2006.05.005
Long. GJ: Rosen. JF: Pounds. JG. (1990). Cellular lead toxicity and metabolism in primary and clonal
osteoblastic bone cells. Toxicol Appl Pharmacol 102: 346-361.
Loos M. SA. (2010). Receptor-oriented approaches in wildlife and human exposure modelling: A
comparative study. Environ Modell Softw 25: 369-382.
Loos, M; Schipper, AM; Schlink, U; Strebel, K; Ragas, AJM. (2010). Receptor-oriented approaches in
wildlife and human exposure modelling: A comparative study. Environ Modell Softw 25: 369-382.
Lorenzana. RM; Troast R: Klotzbach. JM; Follansbee. MH; Diamond. GL. (2005). Issues related to time
averaging of exposure in modeling risks associated with intermittent exposures to lead. Risk Anal 25:
169-178. http://dx.doi.0rg/10.llll/i.0272-4332.2005.00576.x
Lu. Y; Yin. W: Huang. L; Zhang. G: Zhao. Y. (2011). Assessment of bioaccessibility and exposure risk of
arsenic and lead in urban soils of Guangzhou City, China. Environ Geochem Health 33: 93-102.
http://dx.doi.org/10.1007/sl0653-010-9324-8
Lynch. RA; Boatright. DT; Moss. SK. (2000). Lead-contaminated imported tamarind candy and children's
blood lead levels. Public Health Rep 115: 537-543. http://dx.doi.Org/10.1093/phr/115.6.537
Lytle. DA; Schock. MR; Scheckel. K. (2009). The inhibition of Pb(IV) oxide formation in chlorinated
water by orthophosphate. Environ Sci Technol 43: 6624-6631. http://dx.doi.org/10.1021/es900399m
Maas. RP; Patch. SC: Christian. AM; Coplan. MJ. (2007). Effects of fluoridation and disinfection agent
combinations on lead leaching from leaded-brass parts. Neurotoxicology 28: 1023-1031.
http://dx.doi.0rg/10.1016/i.neuro.2007.06.006
Macek. MD; Matte. TD; Sinks. T; Malvitz. DM. (2006). Blood lead concentrations in children and
method of water fluoridation in the United States, 1988-1994. Environ Health Perspect 114: 130-134.
http://dx.doi.org/10.1289/ehp.8319
November 2012 4-161 Draft - Do Not Cite or Quote
-------
Maddaloni. M; Ballew. M; Diamond. G: Follansbee. M; Gefell. D; Goodrum. P; Johnson. M; Koporec. K;
Khoury. G: Luev. J: Odin. M; Troast. R; Van Leeuwen. P; Zaragoza. L. (2005). Assessing lead risks at
non-residential hazardous waste sites. Hum Ecol Risk Assess 11: 967-1003.
http://dx.doi.org/10.1080/10807030500257838
Maddaloni. M; Lolacono. N; Manton. W: Blum. C: Drexler. J; Graziano. J. (1998). Bioavailability of
soilborne lead in adults, by stable isotope dilution. Environ Health Perspect 106: 1589-1594.
http://dx.doi.org/10.1289/ehp.98106s61589
Mahaffev. KR. (1977). Quantities of lead producing health effects in humans: Sources and bioavailability.
Environ Health Perspect 19: 285-295. http://dx.doi.org/10.1289/ehp.7719285
Mahaffev. KR; Annest. JL. (1986). Association of erythrocyte protoporphyrin with blood lead level and
iron status in the second national health and nutrition examination survey, 1976-1980. Environ Res 41:
327-338. http://dx.doi.org/10.1016/80013-9351(86)80194-3
Mahaffev. KR: Gartside. PS: Glueck. CJ. (1986). Blood lead levels and dietary calcium intake in 1-11
year-old children: The second national health and nutrition examination survey, 1976-1980. Pediatrics
78: 257-262.
Malcoe. LH; Lynch. RA; Kegler. MC: Skaggs. VJ. (2002). Lead sources, behaviors, and socioeconomic
factors in relation to blood lead of Native American and white children: A community-based
assessment of a former mining area. Environ Health Perspect 110: 221-231.
http://dx.doi.org/10.1289/ehp.02110s2221
Manton. WI. (1985). Total contribution of airborne lead to blood lead. Occup Environ Med 42: 168-172.
http://dx.doi.0rg/10.1136/oem.42.3.168
Manton. WI: Angle. CR; Krogstrand. KLS. (2005). Origin of lead in the United States diet. Environ Sci
Technol 39: 8995-9000. http://dx.doi.org/10.1021/es051145e
Manton. WI: Angle. CR: Stanek. KL; Kuntzelman. D; Reese. YR; Kuehnemann. TJ. (2003). Release of
lead from bone in pregnancy and lactation. Environ Res 92: 139-151.
http://dx.doi.org/10.1016/80013-9351(03)00020-3
Manton. WI: Angle. CR: Stanek. KL; Reese. YR; Kuehnemann. TJ. (2000). Acquisition and retention of
lead by young children. Environ Res 82: 60-80. http://dx.doi.org/10.1006/enrs.1999.4003
Manton. WI: Cook. JD. (1984). High accuracy (stable isotope dilution) measurements of lead in serum
and cerebrospinal fluid. Occup Environ Med 41: 313-319. http://dx.doi.Org/10.1136/oem.41.3.313
Manton. WI: Mallov. CR. (1983). Distribution of lead in body fluids after ingestion of soft solder. Occup
Environ Med 40: 51-57. http://dx.doi.Org/10.1136/oem.40.l.51
Manton. WI: Rothenberg. SJ: Manalo. M. (2001). The lead content of blood serum. Environ Res 86:
263-273. http://dx.doi.org/10.1006/enrs.2001.4271
Marcus. AH. (1985). Multicompartment kinetic model for lead, III: Lead in blood plasma and
erythrocytes. Environ Res 36: 473-489. http://dx.doi.org/10.1016/0013-9351(85)90039-8
Marcus. AH; Schwartz. J. (1987). Dose-response curves for erythrocyte protoporphyrin vs blood lead:
Effects of iron status. Environ Res 44: 221-227. http://dx.doi.org/10.1016/S0013-9351(87)80230-X
Markowitz. ME; Weinberger. HL. (1990). Immobilization-related lead toxicity in previously lead-
poisoned children. Pediatrics 86: 455-457.
Marschner. B; Welge. P: Hack. A; Wittsiepe. J: Wilhelm. M. (2006). Comparison of soil Pb in vitro
bioaccessibility and in vivo bioavailability with Pb pools from a sequential soil extraction. Environ Sci
Technol 40: 2812-2818. http://dx.doi.org/10.1021/es051617p
Marsh. J: Birchall. A. (1999). Determination of lung-to-blood absorption rates for lead and bismuth which
are appropriate for radon progeny. Radiat Prot Dosimetry 83: 331- 337.
November 2012 4-162 Draft - Do Not Cite or Quote
-------
Martin. D; Glass. TA; Bandeen-Roche. K; Todd. AC: Shi. WP; Schwartz. BS. (2006). Association of
blood lead and tibia lead with blood pressure and hypertension in a community sample of older adults.
Am J Epidemiol 163: 467-478. http://dx.doi.org/10.1093/aje/kwj060
Martinez-Haro. M; Taggart. MA; Mateo. R. (2010). Pb-Al relationships in waterfowl feces discriminate
between sources of Pb exposure. Environ Pollut 158: 2485-2489.
http://dx.doi.0rg/10.1016/i.envpol.2010.03.018
Mateo. R; Baos. AR; Vidal. D; Camarero. PR; Martinez-Haro. M; Taggart. MA. (2011). Bioaccessibility
of Pb from ammunition in game meat is affected by cooking treatment. PLoS ONE 6: e!5892.
http://dx.doi.org/10.1371/iournal.pone.0015892
McMichael. AJ; Baghurst. PA; Wigg. NR; Vimpani. GV: Robertson. EF; Roberts. RJ. (1988). Port Pirie
cohort study: Environmental exposure to lead and children's abilities at the age of four years. N Engl J
Med 319: 468-475. http://dx.doi.org/10.1056/NEJM19880825319080
McNeill. A; Bedi. R: Islam. S: Alkhatib. MN; West. R. (2006). Levels of toxins in oral tobacco products
in the U.K.. Tob Control 15: 64-67. http://dx.doi.org/10.1136/tc.2005.013011
McNeill. FE; Stokes. L; Brito. JA; Chettle. PR; Kave. WE. (2000). 109Cd K x-ray fluorescence
measurements of tibial lead content in young adults exposed to lead in early childhood. Occup
Environ Med 57: 465-471. http://dx.doi.Org/10.1136/oem.57.7.465
Meirer. F; Pemmer. B; Pepponi. G: Zoeger. N; Wobrauschek. P; Sprio. S: Tampieri. A; Goettlicher. J:
Steininger. R; Mangold. S: Roschger. P; Berzlanovich. A; Hofstaetter. JG: Streli. C. (2011).
Assessment of chemical species of lead accumulated in tidemarks of human articular cartilage by X-
ray absorption near-edge structure analysis. J Synchrotron Radiat 18: 238-244.
http://dx.doi.org/10.1107/S0909049510052040
Meltzer. HM; Brantsaeter. AL; Borch-Iohnsen. B; Ellingsen. DG: Alexander. J: Thomassen. Y; Stigum.
H; Ydersbond. TA. (2010). Low iron stores are related to higher blood concentrations of manganese,
cobalt and cadmium in non-smoking, Norwegian women in the HUNT 2 study. Environ Res 110:
497-504. http://dx.doi.0rg/10.1016/i.envres.2010.03.006
Meyer. PA; Brown. MJ; Falk. H. (2008). Global approach to reducing lead exposure and poisoning
[Review]. Mutat Res Rev Mutat Res 659: 166-175. http://dx.doi.Org/10.1016/i.mrrev.2008.03.003
Mielke. HW: Covington. TP: Mielke. PW: Wolman. FJ: Powell. ET: Gonzales. CR. (2011 a). Soil
intervention as a strategy for lead exposure prevention: The New Orleans lead-safe childcare
playground project. Environ Pollut 159: 2071-2077. http://dx.doi.Org/10.1016/i.envpol.2010.ll.008
Mielke. HW: Gonzales. C. (2008). Mercury (Hg) and lead (Pb) in interior and exterior New Orleans
house paint films. Chemosphere 72: 882-885. http://dx.doi.Org/10.1016/i.chemosphere.2008.03.061
Mielke. HW: Gonzales. CR: Mielke. PW. (20 lib). The continuing impact of lead dust on children's blood
lead: Comparison of public and private properties in New Orleans. Environ Res 111: 1164-1172.
http://dx.doi.0rg/10.1016/i.envres.2011.06.010
Mielke. HW: Gonzales. CR: Powell. E; Jartun. M; Mielke. PW. Jr. (2007a). Nonlinear association
between soil lead and blood lead of children in metropolitan New Orleans, Louisiana: 2000-2005. Sci
Total Environ 388: 43-53. http://dx.doi.Org/10.1016/i.scitotenv.2007.08.012
Mielke. HW: Powell. ET; Shah. A; Gonzales. CR: Mielke. PW. (2001). Multiple metal contamination
from house paints: Consequences of power sanding and paint scraping in New Orleans. Environ
Health Perspect 109: 973-978. http://dx.doi.org/10.1289/ehp.01109973
Miller. A; Drake. P: Hintz. P; Habian. M. (2010). Characterizing exposures to airborne metals and
nanoparticle emissions in a refinery. Ann Occup Hyg 54: 504-513.
http://dx.doi.org/10.1093/annhyg/meq032
Mindak. WR; Cheng. J: Canas. BJ; Bolger. PM. (2008). Lead in women's and children's vitamins. J Agric
Food Chem 56: 6892-6896. http://dx.doi.org/10.1021/if801236w
November 2012 4-163 Draft - Do Not Cite or Quote
-------
Miodovnik. A; Landrigan. PJ. (2009). The U.S. Food and Drug Administration risk assessment on lead in
women's and children's vitamins is based on outdated assumptions. Environ Health Perspect 117:
1021-1022. http://dx.doi.org/10.1289/ehp.0900573
Miranda. ML; Anthopolos. R; Hastings. D. (2011). Ageospatial analysis of the effects of aviation
gasoline on childhood blood lead levels. Environ Health Perspect 119: 1513-1516.
http://dx.doi.org/10.1289/ehp. 1003231
Miranda. ML; Edwards. SE; Swamy. GK; Paul. CJ: Neelon. B. (2010). Blood lead levels among pregnant
women: Historical versus contemporaneous exposures. Int J Environ Res Public Health 7: 1508-1519.
http://dx.doi.org/10.3390/iierph7041508
Miranda. ML; Kim. D; Hull. AP; Paul. CJ: Galeano. MAO. (2007b). Changes in blood lead levels
associated with use of chloramines in water treatment systems. Environ Health Perspect 115: 221-225.
http://dx.doi.org/10.1289/ehp.9432
Mitchell. RA; Drake. JE; Wittlin. LA; Rejent. TA. (1977). Erythrocyte porphobilinogen synthase (delta-
aminolaevulinate dehydratase) activity: A reliable and quantitative indicator of lead exposure in
humans. Clin Chem 23: 105-111.
Miyake. M. (1986). Structure refinements of Pb2+ ion-exchanged apatites by x-ray powder pattern-
fitting. J Solid State Chem 61: 230-235. http://dx.doi.org/10.1016/0022-4596(86)90026-5
Miyaki. K; Lwin. H; Masaki. K; Song. YX; Takahashi. Y; Muramatsu. M; Nakavama. T. (2009).
Association between a polymorphism of aminolevulinate dehydrogenase (ALAD) gene and blood lead
levels in Japanese subjects. Int J Environ Res Public Health 6: 999-1009.
http://dx.doi.org/10.3390/ijerph6030999
Molnar. P; Bellander. T; Sallsten. G: Boman. J. (2007). Indoor and outdoor concentrations of PM2.5 trace
elements at homes, preschools and schools in Stockholm, Sweden. J Environ Monit 9: 348-357.
http://dx.doi.org/10.1039/b616858b
Molnar. P; Gustafson. P; Johannesson. S: Boman. J: Barregard. L; Sallsten. G. (2005). Domestic wood
burning and PM2.5 trace elements: Personal exposures, indoor and outdoor levels. Atmos Environ 39:
2643-2653. http://dx.doi.Org/10.1016/i.atmosenv.2005.01.016
Montenegro. MF; Barbosa. F. Jr; Sandrim. VC: Gerlach. RF; Tanus-Santos. JE. (2006). A polymorphism
in the delta-aminolevulinic acid dehydratase gene modifies plasma/whole blood lead ratio. Arch
Toxicol 80: 394-398. http://dx.doi.org/10.1007/s00204-005-0056-v
Montenegro. MF; Barbosa. F. Jr; Tanus-Santos. JE. (2008). Assessment of how pregnancy modifies
plasma lead and plasma/whole blood lead ratio in ALAD 1-1 genotype women. Basic Clin Pharmacol
Toxicol 102: 347-351. http://dx.doi.Org/10.llll/i.1742-7843.2007.00205.x
Morrison. JN; Quarterman. J. (1987). The relationship between iron status and lead absorption in rats.
Biol Trace Elem Res 14: 115-126. http://dx.doi.org/10.1007/BF02795602
Morrow. PE; Beiter. H; Amato. F; Gibb. FR. (1980). Pulmonary retention of lead: An experimental study
in man. Environ Res 21: 373-384. http://dx.doi.org/10.1016/0013-9351(80)90040-7
Mushak. P. (1991). Gastro-intestinal absorption of lead in children and adults: Overview of biological and
biophysico-chemical aspects. Chem Speciation Bioavailability 3: 87-104.
Mushak. P. (1998). Uses and limits of empirical data in measuring and modeling human lead exposure
[Review]. Environ Health Perspect 106: 1467-1484. http://dx.doi.org/10.1289/ehp.98106s61467
Naithani. V; Kakkar. P. (2006). Effect of ecological variation on heavy metal content of some medicinal
plants used as herbal tea ingredients in India. Bull Environ Contam Toxicol 76: 285-292.
http://dx.doi.org/10.1007/s00128-006-0919-9
Navas-Acien. A; Schwartz. BS: Rothenberg. SJ: Hu. H; Silbergeld. EK; Guallar. E. (2008). Bone lead
levels and blood pressure endpoints: Ameta-analysis. Epidemiology 19: 496-504.
http://dx.doi.org/10.1097/EDE.Ob013e31816a2400
November 2012 4-164 Draft - Do Not Cite or Quote
-------
NCHS (National Center for Health Statistics). (2010). National health and nutrition examination survey:
Questionnaires, datasets, and related documentation. Available online at
http://www.cdc.gov/nchs/nhanes/nhanes questionnaires.htm (accessed November 19, 2010).
Needleman. HL; McFarland. C: Ness. RB; Fienberg. SB; Tobin. MJ. (2002). Bone lead levels in
adjudicated delinquents: A case control study. Neurotoxicol Teratol 24: 711-717.
http://dx.doi.org/10.1016/S0892-0362(02)00269-6
Nie. H; Sanchez. BN; Wilker. E; Weisskopf. MG: Schwartz. J; Sparrow. D; Hu. H. (2009). Bone lead and
endogenous exposure in an environmentally exposed elderly population: The Normative Aging Study.
J Occup Environ Med 51: 848-857. http://dx.doi.org/10.1097/JOM.Ob013e3181aa0106
Nie. L; Sanchez. S: Newton. K; Grodzins. L; Cleveland. R; Weisskopf. M. (2011 a). In vivo quantification
of lead in bone with a portable x-ray fluorescence system: Methodology and feasibility. Phys Med
Biol 56: N39-N51. http://dx.doi.Org/10.1088/0031-9155/56/3/N01
Nie. LH; Wright. RO: Bellinger. DC: Hussain. J: Amarasiriwardena. C: Chettle. PR; Pejovic-Milic. A;
Woolf. A; Shannon. M. (20lib). Blood lead levels and cumulative blood lead index (CBLI) as
predictors of late neurodevelopment in lead poisoned children. Biomarkers 16: 517-524.
http://dx.doi.org/10.3109/1354750X.2011.604133
Nielsen. T; Jensen. KA; Grandjean. P. (1978). Organic lead in normal human brains. Nature 274:
602-603. http://dx.doi.org/10.1038/274602aO
Nilsson. U; Attewell. R; Christoffersson. JO; Schutz. A; Ahlgren. L; Skerfving. S: Mattsson. S. (1991).
Kinetics of lead in bone and blood after end of occupational exposure. Basic Clin Pharmacol Toxicol
68: 477-484.
NIOSH (National Institute for Occupational Safety and Health). (1994). NIOSH manual of analytical
methods: Method no. 7105: Lead by GFAAS. (NIOSH 94-113). Cincinnati, OH: U.S. Department of
Health and Human Services, National Institute for Occupational Safety and Health.
Niu. J; Rasmussen. P; Hassan. N; Vincent. R. (2010). Concentration distribution and bioaccessibility of
trace elements in nano and fine urban airborne paniculate matter: Influence of particle size. Water Air
Soil Pollut 213: 211-225. http://dx.doi.org/10.1007/sll270-010-0379-z
Nriagu. J: Burt. B; Linder. A; Ismail. A; Sohn. W. (2006). Lead levels in blood and saliva in a low-income
population of Detroit, Michigan. Int J Hyg Environ Health 209: 109-121.
http://dx.doi.0rg/10.1016/i.iiheh.2005.ll.005
Nwude. DO: Okoye. PAC: Babavemi. JO. (2010). Blood heavy metal levels in cows at slaughter at awka
abattoir, Nigeria. International Journal of Dairy Science 5: 264-270.
http://dx.doi.org/10.3923/iids.2010.264.270
Nziguheba. G: Smolders. E. (2008). Inputs of trace elements in agricultural soils via phosphate fertilizers
in European countries. Sci Total Environ 390: 53-57. http://dx.doi.Org/10.1016/j.scitotenv.2007.09.031
O'Flahertv. EJ. (1993). Physiologically based models for bone-seeking elements: IV. Kinetics of lead
disposition in humans. Toxicol Appl Pharmacol 118: 16-29. http://dx.doi.org/10.1006/taap.1993.1004
O'Flahertv. EJ. (1995). Physiologically based models for bone-seeking elements: V. Lead absorption and
disposition in childhood [Review]. Toxicol Appl Pharmacol 131: 297-308.
http://dx.doi.org/10.1006/taap.1995.1072
O'Flahertv. EJ. (1998). A physiologically based kinetic model for lead in children and adults [Review].
Environ Health Perspect 106: 1495-1503. http://dx.doi.org/10.1289/ehp.98106s61495
O'Flahertv. EJ: Hammond. PB; Lerner. SI. (1982). Dependence of apparent blood lead half-life on the
length of previous lead exposure in humans. Toxicol Sci 2: 49-54.
http://dx.doi.org/10.1093/toxsci/2.1.49
O'Rourke. MK; Rogan. SP; Jin. S: Robertson. GL. (1999). Spatial distributions of arsenic exposure and
mining communities from NHEXAS Arizona. National Human Exposure Assessment Survey. J Expo
Anal Environ Epidemiol 9: 446-455. http://dx.doi.org/10.1038/sj.jea.7500050
November 2012 4-165 Draft - Do Not Cite or Quote
-------
Obi. E; Akunyili. DN; Ekpo. B; Orisakwe. OE. (2006). Heavy metal hazards of Nigerian herbal remedies.
Sci Total Environ 369: 35-41. http://dx.doi.Org/10.1016/i.scitotenv.2006.04.024
Oldereid. NB; Thomassen. Y; Attramadal. A; Olaisen. B; Purvis. K. (1993). Concentrations of lead,
cadmium and zinc in the tissues of reproductive organs of men. Reprod Fertil Dev 99: 421-425.
http://dx.doi.org/10.1530/irf.0.0990421
Omokhodion. FO; Crockford. GW. (1991). Lead in sweat and its relationship to salivary and urinary
levels in normal healthy subjects. Sci Total Environ 103: 113-122.
http://dx.doi.org/10.1016/0048-9697(91)90137-4
Omolaove. JA; Uzairu. A; Gimba. CE. (2010a). Heavy metal assessment of some eye shadow products
imported into Nigeria from China. Arch Appl Sci Res 2: 76-84.
Omolaove. JA; Uzairu. A; Gimba. CE. (201 Ob). Heavy metal assessment of some soft plastic toys
imported into Nigeria from China. Journal of Environmental Chemistry and Ecotoxicology 2:
126-130.
Ong. CN; Lee. WR. (1980a). Distribution of lead-203 in human peripheral blood in vitro. Br J Ind Med
37: 78-84. http://dx.doi.Org/10.1136/oem.37.l.78
Opler. MGA; Brown. AS: Graziano. J; Desai. M; Zheng. W: Schaefer. C: Factor-Litvak. P; Susser. ES.
(2004). Prenatal lead exposure, delta-aminolevulinic acid, and schizophrenia. Environ Health Perspect
112: 548-552. http://dx.doi.org/10.1289/ehp.10464
Opler. MGA: Buka. SL; Groeger. J: McKeague. I; Wei. C: Factor-Litvak. P; Bresnahan. M; Graziano. J:
Goldstein. JM; Seidman. LJ; Brown. AS: Susser. ES. (2008). Prenatal exposure to lead, delta-
aminolevulinic acid, and schizophrenia: Further evidence. Environ Health Perspect 116: 1586-1590.
http://dx.doi.org/10.1289/ehp.10464
Osterberg. K; Borjesson. J: Gerhardsson. L; Schutz. A; Skerfving. S. (1997). A neurobehavioural study of
long-term occupational inorganic lead exposure. Sci Total Environ 201: 39-51.
http://dx.doi.org/10.1016/80048-9697(97)84051-X
Otto. DA; Robinson. G: Baumann. S: Schroeder. S: Mushak. P; Kleinbaum. D; Boone. L. (1985). 5-year
follow-up study of children with low-to-moderate lead absorption: Electrophysiological evaluation.
Environ Res 38: 168-186. http://dx.doi.org/10.1016/0013-9351(85)90082-9
Oulhote. Y; Bot. BL; Poupon. J: Lucas. JP; Mandin. C: Etchevers. A; Zmirou-Navier. D; Glorennec. P.
(2011). Identification of sources of lead exposure in French children by lead isotope analysis: A cross-
sectional study. Environ Health Global Access Sci Source 10: 75.
http://dx.doi.org/10.1186/1476-069X-10-75
P'an. AYS. (1981). Lead levels in saliva and in blood. J Toxicol Environ Health 7: 273- 280.
http://dx.doi.org/10.1080/15287398109529978
Pan. TL; Wang. PW: Al-Suwaveh. SA; Chen. CC: Fang. JY. (2010). Skin toxicology of lead species
evaluated by their permeability and proteomic profiles: A comparison of organic and inorganic lead.
Toxicol Lett 197: 19-28. http://dx.doi.Org/10.1016/i.toxlet.2010.04.019
Park. SK; Elmarsafawv. S: Mukherjee. B; Spiro. A. Ill; Vokonas. PS: Nie. H; Weisskopf MG: Schwartz.
J; Hu. H. (2010). Cumulative lead exposure and age-related hearing loss: The VA Normative Aging
Study. Hear Res 269: 48-55. http://dx.doi.Org/10.1016/i.heares.2010.07.004
Park. SK; Hu. H; Wright. RO: Schwartz. J: Cheng. Y; Sparrow. D: Vokonas. PS: Weisskopf. MG.
(2009b). Iron metabolism genes, low-level lead exposure, and QT interval. Environ Health Perspect
117: 80-85. http://dx.doi.org/10.1289/ehp.11559
Park. SK: Mukherjee. B; Xia. X; Sparrow. D: Weisskopf. MG: Nie. H; Hu. H. (2009c). Bone lead level
prediction models and their application to examine the relationship of lead exposure and hypertension
in the third National Health and Nutrition Examination Survey. J Occup Environ Med 51: 1422-1436.
http://dx.doi.org/10.1097/JOM.Ob013e3181bf6c8d
November 2012 4-166 Draft - Do Not Cite or Quote
-------
Park. SK; Schwartz. J; Weisskopf. M; Sparrow. D; Vokonas. PS: Wright. RO: Coull. B; Nie. H; Hu. H.
(2006). Low-level lead exposure, metabolic syndrome, and heart rate variability: The VA Normative
Aging Study. Environ Health Perspect 114: 1718-1724.
Patel. AB; Prabhu. AS. (2009). Determinants of lead level in umbilical cord blood. Indian Pediatr 46:
791-793.
Pavton. M; Puggs. KM; Spiro. A. Ill; Weiss. ST; Hu. H. (1998). Relations of bone and blood lead to
cognitive function: The VA Normative Aging Study. Neurotoxicol Teratol 20: 19-27.
Pekev. B; Bozkurt. ZB; Pekev. H; Dogan. G: Zararsiz. A; Efe. N; Tuncel. G. (2010). Indoor/outdoor
concentrations and elemental composition of PM10/PM2.5 in urban/industrial areas of Kocaeli City,
Turkey. Indoor Air 20: 112-125. http://dx.doi.org/10. Illl/i.l600-0668.2009.00628.x
Perez-Bravo. F; Ruz. M; Moran-Jimenez. MJ; Olivares. M; Rebolledo. A; Codoceo. J: Sepulveda. V:
Jenkin. A; Santos. JL; Fontanellas. A. (2004). Association between aminolevulinate dehydrase
genotypes and blood lead levels in children from a lead-contaminated area in Antofagasta, Chile. Arch
Environ Contam Toxicol 47: 276-280. http://dx.doi.org/10.1007/s00244-004-2215-l
Perry. SG: Cimorelli. AJ; Paine. RJ: Erode. RW: Weil. JC: Venkatram. A; Wilson. RB: Lee. RF; Peters.
WD. (2005). AERMOD: A dispersion model for industrial source applications. Part II: Model
performance against 17 field study databases. J Appl Meteorol 44: 694-708.
http://dx.doi.Org/10.1175/JAM2228.l
Peters. JL; Kubzansky. L; McNeely. E; Schwartz. J: Spiro. A. Ill; Sparrow. D; Wright. RO: Nie. H; Hu. H.
(2007). Stress as a potential modifier of the impact of lead levels on blood pressure: The Normative
Aging Study. Environ Health Perspect 115: 1154-1159. http://dx.doi.org/10.1289/ehp.10002
Pierre. F: Vallaver. C: Baruthio. F: Peltier. A; Pale. S: Rouver. J: Goutet. P; Aubrege. B; Lecossois. C:
Guillemin. C: Elcabache. JM; Verelle. B; Fabries. JF. (2002). Specific relationship between blood lead
and air lead in the crystal industry. Int Arch Occup Environ Health 75: 217-223.
Pirkle. JL: Brodv. DJ: Gunter. EW: Kramer. RA; Paschal. DC: Flegal. KM; Matte. TD. (1994). The
decline in blood lead levels in the United States: The National Health and Nutrition Examination
Surveys (NHANES). JAMA 272: 284-291. http://dx.doi.org/10.1001/jama.1994.03520040046039
Pirkle. JL; Kaufmann. RB; Brodv. DJ: Hickman. T; Gunter. EW: Paschal. DC. (1998). Exposure of the
U.S. population to lead, 1991-1994. Environ Health Perspect 106: 745-750.
Pocock. SJ: Shaper. AG: Walker. M; Wale. CJ: Clayton. B; Delves. T; Lacev. RF: Packham. RF; Powell.
P (1983). Effects of tap water lead, water hardness, alcohol, and cigarettes on blood lead
concentrations. J Epidemiol Community Health 37: 1-7. http://dx.doi.Org/10.1136/jech.37.l.l
Popovic. M; McNeill. FE; Chettle. PR; Webber. CE; Lee. CV: Kave. WE. (2005). Impact of occupational
exposure on lead levels in women. Environ Health Perspect 113: 478-484.
http://dx.doi.org/10.1289/ehp.7386
Pounds. JG: Leggett. RW. (1998). The ICRP age-specific biokinetic model for lead: Validations, empirical
comparisons, and explorations [Review]. Environ Health Perspect 106: 1505-1511.
Pounds. JG: Long. GJ: Rosen. JF. (1991). Cellular and molecular toxicity of lead in bone [Review].
Environ Health Perspect 91: 17-32. http://dx.doi.org/10.1289/ehp.919117
Pounds. JG: Marlar. RJ: Allen. JR. (1978). Metabolism of lead-210 in juvenile and adult rhesus monkeys
(Macaca mulatta). Bull Environ Contam Toxicol 19: 684-691.
Pounds. JG: Rosen. JF. (1986). Cellular metabolism of lead: Akinetic analysis in cultured osteoclastic
bone cells. Toxicol Appl Pharmacol 83: 531-545. http://dx.doi.org/10.1016/0041-008X(86)90236-X
Pruvot. C: Douav. F; Herve. F; Waterlot. C. (2006). Heavy metals in soil, crops and grass as a source of
human exposure in the former mining areas. Journal of Soils and Sediments 6: 215-220.
http://dx.doi.org/10.1065/iss2006.10.186
November 2012 4-167 Draft - Do Not Cite or Quote
-------
Qian. J; Ferro. AR. (2008). Resuspension of dust particles in a chamber and associated environmental
factors. Aerosol Sci Technol 42: 566-578. http://dx.doi.org/10.1080/02786820802220274
Qian. J; Ferro. AR; Fowler. KR. (2008). Estimating the resuspension rate and residence time of indoor
particles. J Air Waste Manag Assoc 58: 502-516. http://dx.doi.Org/10.3155/1047-3289.58.4.502
Que Hee. SS: Peace. B; Clark. CS: Boyle. JR; Bornschein. RL; Hammond. PB. (1985). Evolution of
efficient methods to sample lead sources, such as house dust and hand dust, in the homes of children.
Environ Res 38: 77-95. http://dx.doi.org/10.1016/0013-9351(85)90074-X
R.. ZA; A.. SL; S.. EA. (2011). Metal sources and exposures in the homes of young children living near a
mining-impacted Superfund site. J Expo Sci Environ Epidemiol 21: 495-505.
http://dx.doi.org/10.1038/ies.2011.21
Raab. GM; Laxen. D; Lindsay. F; Anderson. N; Fulton. M. (1991). The influence of pH and household
plumbing on water lead levels in Edinburgh. In JG Farmer (Ed.), International conference: Heavy
metals in the environment, v 1; September; Edinburgh, United Kingdom (v. 1 ed., pp. 127-130).
Edinburgh, United Kingdom: CEP Consultants, Ltd.
Rabinowitz. M; Leviton. A; Needleman. H; Bellinger. D; Waternaux. C. (1985). Environmental correlates
of infant blood lead levels in Boston. Environ Res 38: 96-107.
http://dx.doi.org/10.1016/0013-9351(85)90075-1
Rabinowitz. M; Needleman. H; Burlev. M; Finch. H; Rees. J. (1984). Lead in umbilical blood, indoor air,
tap water, and gasoline in Boston. Arch Environ Health 39: 299-301.
Rabinowitz. MB. (1991). Toxicokinetics of bone lead [Review]. Environ Health Perspect 91: 33-37.
httrj://dx.doi.org/10.1289/ehp.919133
Rabinowitz. MB. (1995). Relating tooth and blood lead levels in children. Bull Environ Contam Toxicol
55: 853-857. http://dx.doi.org/10.1007/BF00209464
Rabinowitz. MB; Kopple. JD; Wetherill. GW. (1980). Effect of food intake and fasting on gastrointestinal
lead absorption in humans. Am J Clin Nutr 33: 1784-1788.
Rabinowitz. MB; Leviton. A; Bellinger. D. (1993). Relationships between serial blood lead levels and
exfoliated tooth dentin lead levels: Models of tooth lead kinetics. Calcif Tissue Int 53: 338-341.
http://dx.doi.org/10.1007/BF01351840
Rabinowitz. MB; Leviton. A; Bellinger. DC. (1989). Blood lead - tooth lead relationship among Boston
children. Bull Environ Contam Toxicol 43: 485-492. http://dx.doi.org/10.1007/BF01701924
Rabinowitz. MB; Wetherill. GW: Kopple. JD. (1973). Lead metabolism in the normal human: Stable
isotope studies. Science 182: 725-727. http://dx.doi.org/10.1126/science.182.4113.725
Rabinowitz. MB; Wetherill. GW: Kopple. JD. (1976). Kinetic analysis of lead metabolism in healthy
humans. J Clin Invest 58: 260-270. http://dx.doi.org/10.1172/JCI108467
Rabinowitz. MB; Wetherill. GW: Kopple. JD. (1977). Magnitude of lead intake from respiration by
normal man. Transl Res 90: 238-248.
Rabstein. S: Unfried. K; Ranft. U: Illig. T; Kolz. M; Mambetova. C: Vlad. M; Roman. C: Weiss. T;
Becker. D; Bruning. T; Pesch. B. (2008). Lack of association of delta-aminolevulinate dehydratase
polymorphisms with blood lead levels and hemoglobin in Romanian women from a lead-contaminated
region. J Toxicol Environ Health A 71: 716-724. http://dx.doi.org/10.1080/15287390801985190
Rajan. P; Kelsev. KT; Schwartz. JD: Bellinger. DC: Weuve. J: Sparrow. D: Spiro. A. Ill; Smith. TJ; Nie.
H; Hu. H; Wright. RO. (2007). Lead burden and psychiatric symptoms and the modifying influence of
the delta-aminolevulinic acid dehydratase (ALAD) polymorphism: The VA Normative Aging Study.
Am J Epidemiol 166: 1400-1408. http://dx.doi.org/10.1093/aie/kwm220
November 2012 4-168 Draft - Do Not Cite or Quote
-------
Raian. P; Kelsev. KT; Schwartz. JD; Bellinger. DC: Weuve. J: Spiro. A. Ill; Sparrow. D; Smith. TJ; Nie.
H; Weisskopf. MG: Hu. H; Wright. RO. (2008). Interaction of the delta-aminolevulinic acid
dehydratase polymorphism and lead burden on cognitive function: The VA Normative Aging Study. J
Occup Environ Med 50: 1053-1061. http://dx.doi.org/10.1097/JOM.Ob013e3181792463
Ranft. U: Delschen. T; Machtolf. M; Sugiri. D; Wilhelm. M. (2006). Lead concentration in the blood of
children and its association with lead in soil and ambient air: Trends between 1983 and 2000 in a
German industrial city [Abstract]. Epidemiology 17: S122-S122.
Ranft. U: Delschen. T; Machtolf. M; Sugiri. D; Wilhelm. M. (2008). Lead concentration in the blood of
children and its association with lead in soil and ambient air: Trends between 1983 and 2000 in
Duisburg. J Toxicol Environ Health A 71: 710-715. http://dx.doi.org/10.1080/15287390801985117
Rankin. C: Nriagu. JO: Aggarwal. JK; Arowolo. TA; Adebavo. K; Flegal. AR. (2005). Lead
contamination in cocoa and cocoa products: Isotopic evidence of global contamination. Environ
Health Perspect 113: 1344-1348. http://dx.doi.org/10.1289/ehp.8009
Rasmussen. PE; Wheeler. AJ; Hassan. NM; Filiatreault. A; Lanouette. M. (2007). Monitoring personal,
indoor, and outdoor exposures to metals in airborne paniculate matter: Risk of contamination during
sampling, handling and analysis. Atmos Environ 41: 5897-5907.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.03.018
Reglero. MM; Taggart. MA; Monsalve-Gonzalez. L; Mateo. R. (2009b). Heavy metal exposure in large
game from a lead mining area: Effects on oxidative stress and fatty acid composition in liver. Environ
Pollut 157: 1388-1395. http://dx.doi.Org/10.1016/i.envpol.2008.ll.036
Rentschler. G: Broberg. K; Lundh. T; Skerfving. S. (2012). Long-term lead elimination from plasma and
whole blood after poisoning. Int Arch Occup Environ Health 85: 311-316.
http://dx.doi.org/10.1007/s00420-011-0673-0
Rhodes. D: Spiro. A. Ill; Aro. A; Hu. H. (2003). Relationship of bone and blood lead levels to psychiatric
symptoms: The Normative Aging Study. J Occup Environ Med 45: 1144-1151.
http://dx.doi.org/10.1097/01.iom.0000094995.23808.7b
Riedt. CS: Buckley. BT; Brolin. RE; Ambia-Sobhan. H; Rhoads. GG: Shapses. SA. (2009). Blood lead
levels and bone turnover with weight reduction in women. J Expo Sci Environ Epidemiol 19: 90-96.
http://dx.doi.0rg/10.1038/ies.2008.5
Robbins. N; Zhang. ZF; Sun. J: Ketterer. ME; Lalumandier. JA; Shulze. RA. (2010). Childhood lead
exposure and uptake in teeth in the Cleveland area during the era of leaded gasoline. Sci Total Environ
408: 4118-4127. http://dx.doi.Org/10.1016/i.scitotenv.2010.04.060
Rodrigues. EG: Virji. MA; McClean. MD; Weinberg. J: Woskie. S: Pepper. LD. (2010). Personal
exposure, behavior, and work site conditions as determinants of blood lead among bridge painters. J
Occup Environ Hyg 7: 80-87. http://dx.doi.org/10.1080/15459620903418316
Rodrigues. JL; Batista. BL; Nunes. JA; Passos. CJS: Barbosa. F. (2008). Evaluation of the use of human
hair for biomonitoring the deficiency of essential and exposure to toxic elements. Sci Total Environ
405: 370-376. http://dx.doi.Org/10.1016/i.scitotenv.2008.06.002
Roels. H; Konings. J: Green. S: Bradley. D: Chettle. D: Lauwerys. R. (1995). Time-integrated blood lead
concentration is a valid surrogate for estimating the cumulative lead dose assessed by tibial lead
measurement. Environ Res 69: 75-82.
Roels. H; Lauwervs. R: Konings. J: Buchet JP; Bernard. A; Green. S: Bradley. D: Morgan. W: Chettle.
EX (1994). Renal function and hyperfiltration capacity in lead smelter workers with high bone lead.
Occup Environ Med 51: 505-512. http://dx.doi.Org/10.1136/oem.51.8.505
Rosen. JF. (1983). The metabolism of lead in isolated bone cell populations: Interactions between lead
and calcium. Toxicol Appl Pharmacol 71: 101-112. http://dx.doi.org/10.1016/0041-008X(83)90049-2
November 2012 4-169 Draft - Do Not Cite or Quote
-------
Rosen. JF; Pounds. JG. (1998). Severe chronic lead insult that maintains body burdens of lead related to
those in the skeleton: Observations by Dr Clair Patterson conclusively demonstrated [Review].
Environ Res 78: 140-151. http://dx.doi.org/10.1006/enrs.1997.3830
Rothenberg. SJ: Karchmer. S: Schnaas. L; Perroni. E; Zea. F; Alba. JF. (1994a). Changes in serial blood
lead levels during pregnancy. Environ Health Perspect 102: 876-880.
http://dx.doi.org/10.1289/ehp.94102876
Rothenberg. SJ: Kondrashov. V; Manalo. M; Jiang. J; Cuellar. R; Garcia. M; Reynoso. B; Reyes. S: Diaz.
M; Todd. AC. (2002a). Increases in hypertension and blood pressure during pregnancy with increased
bone lead levels. Am J Epidemiol 156: 1079-1087. http://dx.doi.org/10.1093/aie/kwfl63
Rothenberg. SJ: Schnaas. L; Perroni. E; Hernandez. RM; Karchmer. S. (1998). Secular trend in blood lead
levels in a cohort of Mexico City children. Arch Environ Health 53: 231-235.
http://dx.doi.org/10.1080/00039899809605700
Roussel. H; Waterlot. C: Pelfrene. A; Pruvot. C: Mazzuca. M; Douav. F. (2010). Cd, Pb and Zn oral
bioaccessibility of urban soils contaminated in the past by atmospheric emissions from two lead and
zinc smelters. Arch Environ Contam Toxicol 58: 945-954.
http://dx.doi.org/10.1007/s00244-009-9425-5
Rubio-Andrade. M; Valdes-Perezgasga. F; Alonso. J: Rosado. JL; Cebrian. ME; Garcia-Vargas. GG.
(2011). Follow-up study on lead exposure in children living in a smelter community in northern
Mexico. Environ Health Global Access Sci Source 10: 66. http://dx.doi.org/10.1186/1476-069X-10-66
Ryu. JE; Ziegler. EE; Nelson. SE; Fomon. SJ. (1983). Dietary intake of lead and blood lead concentration
in early infancy. Am J Dis Child 137: 886-891.
Saikat. S: Barnes. B; Westwood. D. (2007). A review of laboratory results for bioaccessibility values of
arsenic, lead and nickel in contaminated U.K. soils. J Environ Sci Health A Tox Hazard Subst Environ
Eng42: 1213-1221. http://dx.doi.org/10.1080/10934520701435486
Sakai. T; Yanagihara. S: Kunugi. Y; Ushio. K. (1982). Relationships between distribution of lead in
erythrocytes in vivo and in vitro and inhibition of ALA-D. Br J Ind Med 39: 382-387.
Sammut. ML; Noack. Y; Rose. J: Hazemann. JL; Proux. O; Depoux. M; Ziebel. A; Fiani. E. (2010).
Speciation of Cd and Pb in dust emitted from sinter plant. Chemosphere 78: 445-450.
http://dx.doi.0rg/10.1016/i.chemosphere.2009.10.039
Sayre. JW: Charnev. E; Vostal. J: Pless. IB. (1974). House and hand dust as a potential source of
childhood lead exposure. Am J Dis Child 127: 167-170.
Schell. LM; Denham. M; Stark. AD; Ravenscroft. J: Parsons. P; Schulte. E. (2004). Relationship between
blood lead concentration and dietary intakes of infants from 3 to 12 months of age. Environ Res 96:
264-273. http://dx.doi.0rg/10.1016/i.envres.2004.02.008
Schmitt. CJ: Brumbaugh. WG: May. TW. (2007a). Accumulation of metals in fish from lead-zinc mining
areas of southeastern Missouri, USA. Ecotoxicol Environ Saf 67: 14-30.
http://dx.doi.0rg/10.1016/i.ecoenv.2006.ll.002
Schmitt. CJ: Brumbaugh. WG: May. TW. (2009). Concentrations of cadmium, cobalt, lead, nickel, and
zinc in blood and fillets of northern hog sucker (Hypentelium nigricans) from streams contaminated
by lead-zinc mining: Implications for monitoring. Arch Environ Contam Toxicol 56: 509-524.
http://dx.doi.org/10.1007/s00244-009-9288-9
Schmitt. CJ: Whyte. JJ: Brumbaugh. WG: Tillitt. DE. (2005). Biochemical effects of lead, zinc, and
cadmium from mining on fish in the Tri-States District of northeastern Oklahoma, USA. Environ
Toxicol Chem 24: 1483-1495. http://dx.doi.Org/10.1897/04-332R.l
Schnaas. L; Rothenberg. SJ: Flores. MF; Martinez. S: Hernandez. C: Osorio. E; Perroni. E. (2004). Blood
lead secular trend in a cohort of children in Mexico City (1987-2002). Environ Health Perspect 112:
1110-1115. http://dx.doi.org/10.1289/ehp.6636
November 2012 4-170 Draft - Do Not Cite or Quote
-------
Schnaas. L; Rotfaenberg. SJ: Perroni. E; Martinez. S: Hernandez. C: Hernandez. RM. (2000). Temporal
pattern in the effect of postnatal blood lead level on intellectual development of young children.
Neurotoxicol Teratol 22: 805-810. http://dx.doi.org/10.1016/S0892-0362(00)00101-X
Schock. MR; Hyland. RN; Welch. MM. (2008). Occurrence of contaminant accumulation in lead pipe
scales from domestic drinking-water distribution systems. Environ Sci Technol 42: 4285-4291.
http://dx.doi.org/10.1021/es702488v
Schroeder. HA; Tipton. IH. (1968). The human body burden of lead [Review]. Arch Environ Health 17:
965-978.
Schuhmacher. M; Hernandez. M; Domingo. JL; Fernandez-Ballart. JD; Llobet. JM; Corbella. J. (1996). A
longitudinal study of lead mobilization during pregnancy: Concentrations in maternal and umbilical
cord blood. Trace Elem Electroly 13: 177-181.
Schutz. A; Bergdahl. IA; Ekholm. A; Skerfving. S. (1996). Measurement by ICP-MS of lead in plasma
and whole blood of lead workers and controls. Occup Environ Med 53: 736-740.
http://dx.doi.org/10.1136/oem.53.ll.736
Schwartz. BS: Lee. BK; Bandeen-Roche. K; Stewart. W: Bolla. K; Links. J: Weaver. V; Todd. A. (2005).
Occupational lead exposure and longitudinal decline in neurobehavioral test scores. Epidemiology 16:
106-113. http://dx.doi.org/10.1097/01.ede.0000147109.62324.51
Schwartz. BS: Lee. BK: Lee. GS: Stewart. WF: Lee. SS: Hwang. KY: Ahn. KD: Kim. YB: Bolla. KI:
Simon. D; Parsons. PJ: Todd. AC. (2001). Associations of blood lead, dimercaptosuccinic acid-
chelatable lead, and tibia lead with neurobehavioral test scores in South Korean lead workers. Am J
Epidemiol 153: 453-464. http://dx.doi.Org/10.1093/aie/153.5.453
Schwartz. BS: Stewart. WF; Bolla. KI; Simon. MS: Bandeen-Roche. K; Gordon. B; Links. JM; Todd. AC.
(2000b). Past adult lead exposure is associated with longitudinal decline in cognitive function.
Neurology 55: 1144-1150.
Schwartz. BS: Stewart. WF: Todd. AC: Simon. D; Links. JM. (2000c). Different associations of blood
lead, meso 2,3-dimercaptosuccinic acid (DMSA)-chelatable lead, and tibial lead levels with blood
pressure in 543 former organolead manufacturing workers. Arch Environ Occup Health 55: 85-92.
http://dx.doi.org/10.1080/00039890009603392
Schwartz. J: Pitcher. H. (1989). The relationship between gasoline lead and blood lead in the United
States. J Offic Stat 5: 421-431.
Scinicariello. F; Murray. HE; Moffett DB; Abadin. HG: Sexton. MJ; Fowler. BA. (2007). Lead and delta-
aminolevulinic acid dehydratase polymorphism: Where does it lead? Ameta-analysis. Environ Health
Perspect 115: 35-41. http://dx.doi.org/10.1289/ehp.9448
Scinicariello. F; Yesupriya. A; Chang. MH; Fowler. BA. (2010). Modification by ALAD of the
association between blood lead and blood pressure in the U.S. population: Results from the Third
National Health and Nutrition Examination Survey. Environ Health Perspect 118: 259-264.
http://dx.doi.org/10.1289/ehp.0900866
Sesli. E; Tuzen. M; Soylak. M. (2008). Evaluation of trace metal contents of some wild edible
mushrooms from Black Sea region, Turkey. J Hazard Mater 160: 462-467.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.03.020
Shah. F; Kazi. TG: Afridi. HI; Khan. S: Kolachi. NF; Arain. MB; Baig. JA. (2011). The influence of
environmental exposure on lead concentrations in scalp hair of children in Pakistan. Ecotoxicol
Environ Saf 74: 727-732. http://dx.doi.Org/10.1016/j.ecoenv.2010.10.036
Shaik. AP; Jamil. K. (2009). Individual susceptibility and genotoxicity in workers exposed to hazardous
materials like lead. J Hazard Mater 168: 918-924. http://dx.doi.Org/10.1016/i.jhazmat.2009.02.129
Sharma. A; ChY. R; Tiwari. RK; Kumar Tyagi. L; Kori. ML; Singh. V; Gaur. K; Shankar. K. (2009). Role
of Aloe barbadensis Linn in the removal of toxic heavy metal of Kukkutandatwak (shell of hen's egg):
A drug used in Indian system of medicine (Ayurveda). Adv Biol Res 3: 79-83.
November 2012 4-171 Draft - Do Not Cite or Quote
-------
Sheets. RG: Bergquist. BA. (1999). Laboratory treatability testing of soils contaminated with lead and
PCBs using particle-size separation and soil washing. J Hazard Mater 66: 137-150.
Sheppard. SC. (1995). Parameter values to model the soil ingestion pathway. Environ Monit Assess 34:
27-44. http://dx.doi.org/10.1007/BF00546244
Sherlock. J; Smart. G: Forbes. GI; Moore. MR; Patterson. WJ: Richards. WN; Wilson. TS. (1982).
Assessment of lead intakes and dose-response for a population in Ayr exposed to a plumbosolvent
water supply. Hum Exp Toxicol 1: 115-122. http://dx.doi.org/10.1177/096032718200100203
Sherlock. JC: Ashbv. D; Delves. HT; Forbes. GI: Moore. MR; Patterson. WJ: Pocock. SJ: Quinn. MJ;
Puchards. WN; Wilson. TS. (1984). Reduction in exposure to lead from drinking water and its effect
on blood lead concentrations. Hum Exp Toxicol 3: 383-392.
http://dx.doi.org/10.1177/096032718400300503
Sherlock. JC: Quinn. MJ. (1986). Relationship between blood and lead concentrations and dietary lead
intake in infants: The Glasgow Duplicate Diet Study 1979-1980. Food Addit Contam 3: 167-176.
http://dx.doi.org/10.1080/02652038609373579
Shih. RA; Glass. TA; Bandeen-Roche. K; Carlson. MC: Bolla. KI; Todd. AC: Schwartz. BS. (2006).
Environmental lead exposure and cognitive function in community-dwelling older adults. Neurology
67: 1556-1562. http://dx.doi.org/10.1212/01.wnl.0000239836.26142.c5
Shih. RA; Hu. H; Weisskopf. MG: Schwartz. BS. (2007). Cumulative lead dose and cognitive function in
adults: A review of studies that measured both blood lead and bone lead [Review]. Environ Health
Perspect 115: 483-492. http://dx.doi.org/10.1289/ehp.9786
Shotyk. W: Krachler. M. (2009). Determination of trace element concentrations in natural freshwaters:
How low is "low," and how low do we need to go? J Environ Monit 11: 1747-1753.
http://dx.doi.org/10.1039/b917090c
Siciliano. SD; James. K; Zhang. GY; Schafer. AN; Peak. JD. (2009). Adhesion and Enrichment of Metals
on Human Hands from Contaminated Soil at an Arctic Urban Brownfield. Environ Sci Technol 43:
6385-6390. http://dx.doi.org/10.1021/es901090w
Silbergeld. EK. (1991). Lead in bone: Implications for toxicology during pregnancy and lactation
[Review]. Environ Health Perspect 91: 63-70.
Silbergeld. EK; Schwartz. J: Mahaffev. K. (1988). Lead and osteoporosis: Mobilization of lead from bone
in postmenopausal women. Environ Res 47: 79-94. http://dx.doi.org/10.1016/80013-9351(88)80023-9
Simon. PL; Maynard. EJ; Thomas. KD. (2007). Living in a sea of lead changes in blood- and hand-lead
of infants living near a smelter. J Expo Sci Environ Epidemiol 17: 248-259.
http://dx.doi.org/10.1038/sj.jes.7500512
Smith. CM; Hu. H; Wang. X; Kelsev. KT. (1995a). ALA-D genotype is not associated with HT or HB
levels among workers exposed to low levels of lead. Med Lav 86: 229-235.
Smith. D; Hernandez-Avila. M; Tellez-Rojo. MM; Mercado. A; Hu. H. (2002). The relationship between
lead in plasma and whole blood in women. Environ Health Perspect 110: 263-268.
http://dx.doi.org/10.1289/ehp.02110263
Smith. DM; Mielke. HW: Heneghan. JB. (2008). Subchronic lead feeding study in male rats. Arch
Environ Contam Toxicol 55: 518-528. http://dx.doi.org/10.1007/s00244-008-9138-l
Smith. DM; Mielke. HW: Heneghan. JB. (2009a). Subchronic lead feeding study in male rats and
micropigs. Environ Toxicol 24: 453-461. http://dx.doi.org/10.1002/tox.20448
Smith. PR; Kahng. MW: Quintanilla-Vega. B; Fowler. BA. (1998). High-affinity renal lead-binding
proteins in environmentally-exposed humans. Chem Biol Interact 115: 39-52.
http://dx.doi.org/10.1016/S0009-2797(98)00060-X
November 2012 4-172 Draft - Do Not Cite or Quote
-------
Smith. PR; Osterloh. JD; Flegal. AR. (1996). Use of endogenous, stable lead isotopes to determine
release of lead from the skeleton. Environ Health Perspect 104: 60-66.
http://dx.doi.org/10.1289/ehp.9610460
Smith. E; Weber. J; Naidu. R; McLaren. RG: Juhasz. AL. (2011). Assessment of lead bioaccessibility in
peri-urban contaminated soil. J Hazard Mater 186: 300-305.
http://dx.doi.0rg/10.1016/i.jhazmat.2010.10.lll
Sobin. C: Gutierrez. M; Alterio. H. (2009). Polymorphisms of delta-aminolevulinic acid dehydratase
(ALAD) and peptide transporter 2 (PEPT2) genes in children with low-level lead exposure.
Neurotoxicology 30: 881-887. http://dx.doi.Org/10.1016/i.neuro.2009.08.006
Soderholm. SC. (1989). Proposed international conventions for particle size-selective sampling. Ann
Occup Hyg 33: 301-320. http://dx.doi.Org/10.1093/annhyg/33.3.301
Spalinger. SM; von Braun. MC: Petrosvan. V; von Lindern. IH. (2007). Northern Idaho house dust and
soil lead levels compared to the Bunker Hill superfund site. Environ Monit Assess 130: 57-72.
http://dx.doi.org/10.1007/sl0661-006-9450-z
SRC (Syracuse Research Corporation). (2002). Baseline human health risk assessment: Eureka Mills -
Eureka, Utah. Denver, CO: U.S. Environmental Protection Agency.
http://www.epa.gov/region8/r8risk/pdf/hhra_eureka.pdf
Srivastava. SK; Rai. V: Srivastava. M; Rawat. AKS: Mehrotra. S. (2006). Estimation of heavy metals in
different Berberis species and its market samples. Environ Monit Assess 116: 315-320.
http://dx.doi.org/10.1007/sl0661-006-7395-x
Sterling. DA; Roegner. KG: Lewis. RD; Luke. DA; Wilder. LC: Burchette. SM. (1999). Evaluation of
four sampling methods for determining exposure of children to lead-contaminated household dust.
Environ Res 81: 130-141. http://dx.doi.org/10.1006/enrs.1999.3962
Stern. AH. (1994). Derivation of a target level of lead in soil at residential sites corresponding to a de
minimis contribution to blood lead concentration. Risk Anal 14: 1049-1056.
http://dx.doi.0rg/10.llll/i.1539-6924.1994.tb00075.x
Stern. AH. (1996). Derivation of a target concentration of Pb in soil based on elevation of adult blood
pressure. Risk Anal 16: 201-210. http://dx.doi.Org/10.llll/i.1539-6924.1996.tb01450.x
Stewart. WF; Schwartz. BS: Davatzikos. C: Shen. D; Liu. D; Wu. X; Todd. AC: Shi. W: Bassett. S:
Youssem. D. (2006). Past adult lead exposure is linked to neurodegeneration measured by brain MRI.
Neurology 66: 1476-1484. http://dx.doi.org/10.1212/01.wnl.0000216138.69777.15
Stewart. WF; Schwartz. BS: Simon. D; Bolla. KI; Todd. AC: Links. J. (1999). Neurobehavioral function
and tibial and chelatable lead levels in 543 former organolead workers. Neurology 52: 1610-1617.
Stokes. L; Letz. R; Gerr. F; Kolczak. M; McNeill. FE; Chettle. PR; Kave. WE. (1998). Neurotoxicity in
young adults 20 years after childhood exposure to lead: the Bunker Hill experience. Occup Environ
Med 55: 507-516.
Succop. P; Bornschein. R; Brown. K; Tseng. CY. (1998). An empirical comparison of lead exposure
pathway models. Environ Health Perspect 1066: 1577-1583.
Tahir. SN; Alaamer. AS. (2008). PB-210 concentrations in cigarettes tobaccos and radiation doses to the
smokers. Radiat Prot Dosimetry 130: 389-391. http://dx.doi.org/10.1093/rpd/ncn097
Tellez-Rojo. MM; Hernandez-Avila. M; Gonzalez-Cossio. T; Romieu. I; Aro. A; Palazuelos. E; Schwartz.
J; Hu. H. (2002). Impact of breastfeeding on the mobilization of lead from bone. Am J Epidemiol 155:
420-428. http://dx.doi.0rg/10.1093/aje/155.5.420
TerraGraphics Environmental Engineering (TerraGraphics Environmental Engineering Inc). (2004).
Human health remedial evaluation report for the Bunker Hill Superfund site box. Moscow, ID.
November 2012 4-173 Draft - Do Not Cite or Quote
-------
Theppeang. K; Glass. TA; Bandeen-Roche. K; Todd. AC: Rohde. CA; Links. JM; Schwartz. BS. (2008a).
Associations of bone mineral density and lead levels in blood, tibia, and patella in urban-dwelling
women. Environ Health Perspect 116: 784-790. http://dx.doi.org/10.1289/ehp.10977
Todd. AC: Buchanan. R; Carroll. S: Moshier. EL; Popovac. D; Slavkovich. V; Graziano. JH. (2001). Tibia
lead levels and methodological uncertainty in 12-year-old children. Environ Res 86: 60-65.
http://dx.doi.org/10.1006/enrs.2001.4251
Todd. AC: Carroll S: Geraghtv. C: Khan. FA; Moshier. EL; Tang. S: Parsons. PJ. (2002). L-shell x-ray
fluorescence measurements of lead in bone: accuracy and precision. Phys Med Biol 47: 1399-1419.
http://dx.doi.org/10.1088/0031-9155/47/8/312
Todd. AC: Carroll. S: Godbold. JH; Moshier. EL; Khan. FA. (2000). Variability in XRF-measured tibia
lead levels. Phys Med Biol 45: 3737-3748. http://dx.doi.org/10.1088/0031-9155/45/12/316
Tovalin-Ahumada. H; Whitehead. L; Blanco. S. (2007). Personal exposure to PM2.5 and element
composition—A comparison between outdoor and indoor workers from two Mexican cities. Atmos
Environ 41: 7401-7413. http://dx.doi.Org/10.1016/i.atmosenv.2007.05.059
Treble. RG: Thompson. TS. (1997). Preliminary results of a survey of lead levels in human liver tissue.
Bull Environ Contam Toxicol 59: 688-695. http://dx.doi.org/10.1007/s001289900535
Tripathi. RM; Raghunath. R; Kumar. AV; Sastry. VN; Sadasivan. S. (2001). Atmospheric and children's
blood lead as indicators of vehicular traffic and other emission sources in Mumbai, India. Sci Total
Environ 267: 101-108. htrp://dx.doi.org/10.1016/S0048-9697(00)00770-l
Tsuji. LJS: Wainman. BC: Javasinghe. RK; VanSpronsen. EP; Liberda. EN. (2009). Determining tissue-
lead levels in large game mammals harvested with lead bullets: Human health concerns. Bull Environ
Contam Toxicol 82: 435-439. http://dx.doi.org/10.1007/s00128-009-9647-2
Tsuji. LJS: Wainman. BC: Martin. ID; Sutherland. C: Weber. JP; Dumas. P; Nieboer. E. (2008). The
identification of lead ammunition as a source of lead exposure in First Nations: The use of lead
isotope ratios. Sci Total Environ 393: 291-298. http://dx.doi.Org/10.1016/i.scitotenv.2008.01.022
Tu. C: Zheng. CR; Chen. HM. (2000). Effect of applying chemical fertilizers on forms of lead and
cadmium in red soil. Chemosphere 41: 133-138. http://dx.doi.org/10.1016/S0045-6535(99)00400-2
Turlakiewicz. Z; Chmielnicka. J. (1985). Diethyllead as a specific indicator of occupational exposure to
tetraethyllead. Occup Environ Med 42: 682-685. http://dx.doi.org/10.1136/oem.42.10.682
Turner. A; Simmonds. L. (2006). Elemental concentrations and metal bioaccessibility in U.K. household
dust. Sci Total Environ 371: 74-81. http://dx.doi.Org/10.1016/i.scitotenv.2006.08.011
Tvinnereim. HM; Eide. R: Riise. T; Wesenberg. GR; Fosse. G: Steinnes. E. (1997). Lead in primary teeth
from Norway: Changes in lead levels from the 1970s to the 1990s. Sci Total Environ 207: 165-177.
htrp://dx.doi.org/10.1016/S0048-9697(97)00263-5
U.S. EPA (U.S. Environmental Protection Agency). (1986a). Air quality criteria for lead [EPA Report].
(EPA/600/8-83/028aF-dF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
U.S. EPA (U.S. Environmental Protection Agency). (1990b). Field evaluation of a high volume surface
sampler for pesticides in floor dust [EPA Report]. (EPA/600/S3-90/030). Research Triangle Park.
U.S. EPA (U.S. Environmental Protection Agency). (1994). Guidance manual for the integrated exposure
uptake biokinetic model for lead in children [EPA Report]. (EPA/540/R-93/081). Washington, D.C.
http://nepis.epa.gov/Exe/ZvPURL.cgi?Dockev=2000WN4R.txt
U.S. EPA (U.S. Environmental Protection Agency). (1996a). Recommendations of the Technical Review
Workgroup for Lead for an interim approach to assessing risks associated with adult exposures to lead
in soil [EPA Report]. Washington, DC: U.S. Environmental Protection Agency, Technical Review
Workgroup for Lead.
November 2012 4-174 Draft - Do Not Cite or Quote
-------
U.S. EPA (U.S. Environmental Protection Agency). (1996b). Urban soil lead abatement demonstration
project: Volume I: EPA integrated report [EPA Report]. (EPA/600/P-93/001aF). Washington, D.C.
http://nepis.epa.gov/Exe/ZyPURL.cgi?Dockev=30002663.txt
U.S. EPA (U.S. Environmental Protection Agency). (2001). Baseline human health risk assessment:
Vasquez Boulevard and 1-70 Superfund site, Denver, CO [EPA Report]. Denver, CO.
http://nepis.epa.gov/Exe/ZvPURL.cgi?Dockev=P1006STM.txt
U.S. EPA (U.S. Environmental Protection Agency). (2003c). Recommendations of the Technical Review
Workgroup for Lead for an approach to assessing risks associated with adult exposures to lead in soil
[EPA Report]. (EPA-540-R-03-001). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2005a). All ages lead model (AALM) (Version Draft
1.05) [Computer Program]. Research Triangle Park, NC: U.S. Environmental Protection Agency,
National Center for Environmental Assessment.
U.S. EPA (U.S. Environmental Protection Agency). (2005f). Re-evaluation of the lEUBK-based clean-up
level for lead in soil in East Helena, Montana [EPA Report]. Denver, CO.
U.S. EPA (U.S. Environmental Protection Agency). (2006b). Air quality criteria for lead: Volume I of II
[EPA Report]. (EPA/600/R-05/144aF). Research Triangle Park, NC.
http ://cfpub. epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
U.S. EPA (U.S. Environmental Protection Agency). (2006c). Air quality criteria for lead: Volume II of II
[EPA Report]. (EPA/600/R-05/144bF). Research Triangle Park, NC.
http ://cfpub .epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
U.S. EPA (U.S. Environmental Protection Agency). (2007b). Estimation of relative bioavailability of lead
in soil and soil-like materials using in vivo and in vitro methods [EPA Report]. (OSWER 9285.7-77).
Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2007e). Guidance for evaluating the oral
bioavailability of metals in soils for use in human health risk assessment [EPA Report]. (OSWER
9285.7-80). Washington, D.C.
http://www.epa.gov/superfund/health/contaminants/bioavailabilitv/guidance.htm
U.S. EPA (U.S. Environmental Protection Agency). (20071). Lead: Human exposure and health risk
assessments for selected case studies. Volume II: Appendices [EPA Report]. (452R07014b). Research
Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2007g). Lead: Human exposure and health risk
assessments for selected case studies: Volume 1: Human exposure and health risk assessments - full-
scale [EPA Report]. (EPA-452/R-07-014a). Research Triangle Park, NC.
http://www.ntis.gov/search/product.aspx?ABBR=PB2008102573
U.S. EPA (U.S. Environmental Protection Agency). (2009a). Integrated science assessment for paniculate
matter [EPA Report]. (EPA/600/R-08/139F). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=216546
Uzu. G: Sobanska. S: Sarret. G: Munoz. M; Dumat. C. (2010). Foliar lead uptake by lettuce exposed to
atmospheric fallouts. Environ Sci Technol 44: 1036-1042. http://dx.doi.org/10.1021/es902190u
Van de Wiele. TR; Oomen. AG: Wragg. J: Cave. M; Minekus. M; Hack. A; Cornells. C: Rompelberg.
CJM; De Zwart. LL; Klinck. B; Van Wijnen. J; Verstraete. W: Sips. A. (2007). Comparison of five in
vitro digestion models to in vivo experimental results: Lead bioaccessibility in the human
gastrointestinal tract. J Environ Sci Health A Tox Hazard Subst Environ Eng 42: 1203-1211.
htrp://dx.doi.org/10.1080/10934520701434919
van Wijngaarden. E; Campbell. JR; Cory-Slechta. DA. (2009). Bone lead levels are associated with
measures of memory impairment in older adults. Neurotoxicology 30: 572-580.
http://dx.doi.0rg/10.1016/i.neuro.2009.05.007
November 2012 4-175 Draft - Do Not Cite or Quote
-------
Vandenhove. H; Olyslaegers. G: Sanzharova. N; Shubina. O: Reed. E; Shang. Z; Velasco. H. (2009).
Proposal for new best estimates of the soil-to-plant transfer factor of U, Th, Ra, Pb and Po. J Environ
Radioact 100: 721-732. http://dx.doi.Org/10.1016/i.jenvrad.2008.10.014
Vander. AJ; Taylor. PL; Kalitis. K; Mouw. PR; Victery. W. (1977). Renal handling of lead in dogs:
Clearance studies. Am J Physiol 233: F532-F538.
Verbeeck. RMH; Lassuvt. CJ: Heijligers. HJM; Driessens. FCM; Vrolijk. JWG. A. (1981). Lattice
parameters and cation distribution of solid solutions of calcium and lead hydroxyapatite. Calcif Tissue
Int 33: 243-247. http://dx.doi.org/10.1007/BF02409444
Victery. W: Vander. AJ: Mouw. DR. (1979). Effect of acid-base status on renal excretion and
accumulation of lead in dogs and rats. Am J Physiol 237: F398-F407.
Villalobos. M; Merino-Sanchez. C: Hall. C: Grieshop. J: Gutierrez-Ruiz. ME; Handlev. MA. (2009). Lead
(II) detection and contamination routes in environmental sources, cookware and home-prepared foods
from Zimatlan, Oaxaca, Mexico. Sci Total Environ 407: 2836-2844.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.12.059
Vural. N: Duvdu. Y. (1995). Biological monitoring of lead in workers exposed to tetraethyllead. Sci Total
Environ 171: 183-187. http://dx.doi.org/10.1016/0048-9697(95)04676-6
Waalkes. MP; Klaassen. CD. (1985). Concentration of metallothionein in major organs of rats after
administration of various metals. Toxicol Sci 5: 473-477. http://dx.doi.Org/10.1093/toxsci/5.3.473
Wananukul. W: Sura. T; Salaitanawatwong. P. (2006). Polymorphism of delta-aminolevulinic acid
dehydratase and its effect on blood lead levels in Thai workers. Arch Environ Occup Health 61: 67-72.
http://dx.doi.Org/10.3200/AEOH.61.2.67-72
Wasserman. GA; Factor-Litvak. P; Liu. X; Todd. AC: Kline. JK; Slavkovich. V; Popovac. D; Graziano.
JH. (2003). The relationship between blood lead, bone lead and child intelligence. Child Neuropsychol
9: 22-34. http://dx.doi.Org/10.1076/chin.9.l.22.14497
Watson. WS: Morrison. J: Bethel. MIF; Baldwin. NM; Lyon. DTB; Dobson. H; Moore. MR; Hume. R.
(1986). Food iron and lead absorption in humans. Am J Clin Nutr 44: 248-256.
Weaver. VM; Ellis. LR; Lee. BK; Todd. AC: Shi. W: Ahn. KD; Schwartz. BS. (2008). Associations
between patella lead and blood pressure in lead workers. Am J Ind Med 51: 336-343.
http://dx.doi.org/10.1002/aiim.20573
Webber. CE; Chettle. PR; Bowins. RJ: Beaumont. LF; Gordon. CL; Song. X; Blake. JM; McNutt. RH.
(1995). Hormone replacement therapy may reduce the return of endogenous lead from bone to the
circulation. Environ Health Perspect 103: 1150-1153.
Weis. CP; Lavelle. JM. (1991). Characteristics to consider when choosing an animal model for the study
of lead bioavailability. Chem Speciation Bioavailability 3: 113-119.
Weisskopf. MG: Hu. H; Sparrow. D; Lenkinski. RE; Wright. RO. (2007a). Proton magnetic resonance
spectroscopic evidence of glial effects of cumulative lead exposure in the adult human hippocampus.
Environ Health Perspect 115: 519-523. http://dx.doi.org/10.1289/ehp.9645
Weisskopf. MG: Jain. N; Nie. HL; Sparrow. D: Vokonas. P; Schwartz. J: Hu. H. (2009). A prospective
study of bone lead concentration and death from all causes, cardiovascular diseases, and cancer in the
department of veterans affairs normative aging study. Circulation 120: 1056-1064.
http://dx.doi.org/10.1161/circulationaha.108.827121
Weisskopf. MG: Proctor. SP; Wright. RO: Schwartz. J: Spiro. A. Ill; Sparrow. D: Nie. HL; Hu. H.
(2007b). Cumulative lead exposure and cognitive performance among elderly men. Epidemiology 18:
59-66. http://dx.doi.org/10.1097/01.ede.0000248237.35363.29
Weisskopf. MG: Weuve. J: Nie. H; Saint-Hilaire. MH; Sudarsky. L; Simon. DK; Hersh. B; Schwartz. J:
Wright. RO: Hu. H. (2010). Association of cumulative lead exposure with Parkinson's Disease.
Environ Health Perspect 118: 1609-1613. http://dx.doi.org/10.1289/ehp.1002339
November 2012 4-176 Draft - Do Not Cite or Quote
-------
Weisskopf MG: Wright. RO: Schwartz. J; Spiro. A. Ill: Sparrow. D; Aro. A; Hu. H. (2004). Cumulative
lead exposure and prospective change in cognition among elderly men: The VA Normative Aging
Study. Am J Epidemiol 160: 1184-1193. http://dx.doi.org/10.1093/aje/kwh333
Welt. M: Mielke. HW: Gonzales. C: Cooper. KM: Batiste. CG: Cresswell. LH. Ill: Mielke. PW. (2003).
Metal contamination of sediments and soils of bayou Saint John: A potential health impact to local
fishermen? Environ Geochem Health 25: 387-396.
Wetmur. JG. (1994). Influence of the common human delta-aminolevulinate dehydratase polymorphism
on lead body burden [Review]. Environ Health Perspect 102: 215-219.
Wetmur. JG: Kava. AH; Plewinska. M; Desnick. RJ. (1991a). Molecular characterization of the human 5-
aminolevulinate dehydratase-2 (ALAD2) allele: Implications for molecular screening of individuals
for genetic susceptibility to lead poisoning. Am J Hum Genet 49: 757-763.
Wetmur. JG: Lehnert. G: Desnick. RJ. (1991b). The delta-aminolevulinate dehydratase polymorphism:
Higher blood lead levels in lead workers and environmentally exposed children with the 1-2 and 2-2
isozymes. Environ Res 56: 109-119. http://dx.doi.org/10.1016/80013-9351(05)80001-5
Weuve. J: Kelsev. KT; Schwartz. J: Bellinger. D; Wright. RO: Raian. P; Spiro. A. Ill; Sparrow. D; Aro. A;
Hu. H. (2006). Delta-aminolevulinic acid dehydratase polymorphism and the relation between low
level lead exposure and the Mini-Mental Status Examination in older men: The Normative Aging
Study. Occup Environ Med 63: 746-753. http://dx.doi.org/10.1136/oem.2006.027417
Weuve. J: Korrick. SA; Weisskopf. MA; Ryan. LM; Schwartz. J: Nie. HL; Grodstein. F; Hu. H. (2009).
Cumulative exposure to lead in relation to cognitive function in older women. Environ Health
Perspect 117: 574-580. http://dx.doi.org/10.1289/ehp.11846
Wigginton. NS: Haus. KL; Hochella. MF. (2007). Aquatic environmental nanoparticles [Review]. J
Environ Monit 9: 1306-1316. http://dx.doi.org/10.1039/b712709i
Wilhelm. M; Lombeck. I; Hafner. D; Ohnesorge. FK. (1989). Hair lead levels in young children from the
FRG. J Trace Elem Med Biol 3: 165-170.
Wilhelm. M; Pesch. A; Rostek. U; Begerow. J: Schmitz. N; Idel. H; Ranft. U. (2002). Concentrations of
lead in blood, hair and saliva of German children living in three different areas of traffic density. Sci
Total Environ 297: 109-118.
Wilker. E; Korrick. S: Nie. LH; Sparrow. D: Vokonas. P; Coull. B; Wright. RO: Schwartz. J: Hu. H.
(2011). Longitudinal changes in bone lead levels: The VA Normative Aging Study. J Occup Environ
Med 53: 850-855. http://dx.doi.org/10.1097/JOM.Ob013e31822589a9
Wilson. J; Dixon. S: Galke. W: McLaine. P. (2007). An investigation of dust lead sampling locations and
children's blood lead levels. J Expo Sci Environ Epidemiol 17: 2-12.
http://dx.doi.org/10.1038/sj.jes.7500514
Wright. RO: Silverman. EK; Schwartz. J: Tsaih. SW: Senter. J: Sparrow. D: Weiss. ST; Aro. A; Hu. H.
(2004). Association between hemochromatosis genotype and lead exposure among elderly men: The
Normative Aging Study. Environ Health Perspect 112: 746-750.
Wright. RO: Tsaih. SW: Schwartz. J: Spiro. A; McDonald. K; Weiss. ST; Hu. H. (2003). Lead exposure
biomarkers and mini-mental status exam scores in older men. Epidemiology 14: 713-718.
http://dx.doi.org/10.1097/01.EDE.0000081988.85964.db
Xie. Y; Chiba. M; Shinohara. A; Watanabe. H; Inaba. Y. (1998). Studies on lead-binding protein and
interaction between lead and selenium in the human erythrocytes. Ind Health 36: 234-239.
http://dx.doi.org/10.2486/indhealth.36.234
Xie. Y; Wang. Y; Giammar. DE. (2010). Impact of chlorine disinfectants on dissolution of the lead
corrosion product PbO2. Environ Sci Technol 44: 7082-7088. http://dx.doi.org/10.1021/esl016763
Yamamoto. N; Takahashi. Y; Yoshinaga. J: Tanaka. A; Shibata. Y. (2006). Size distributions of soil
particles adhered to children's hands. Arch Environ Contain Toxicol 51: 157-163.
http://dx.doi.org/10.1007/s00244-005-7012-y
November 2012 4-177 Draft - Do Not Cite or Quote
-------
Yiin. LM; Rhoads. GG: Lioy. PJ. (2000). Seasonal influences on childhood lead exposure. Environ Health
Perspect 108: 177-182.
Yu. CH; Yiin. LM; Lioy. PJ. (2006). The bioaccessibility of lead (Pb) from vacuumed house dust on
carpets in urban residences. Risk Anal 26: 125-134.
http:/7dx.doi.org/10.1111/i.l539-6924.2006.00710.x
Zahran. S: Mielke. HW: Weiler. S: Gonzales. CR. (2011). Nonlinear associations between blood lead in
children, age of child, and quantity of soil lead in metropolitan New Orleans. Sci Total Environ 409:
1211-1218. http://dx.doi.0rg/10.1016/j.scitotenv.2010.ll.036
Zhang. W: Zhang. GG: He. HZ; Bolt. HM. (1994). Early health effects and biological monitoring in
persons occupationally exposed to tetraethyl lead. Int Arch Occup Environ Health 65: 395-399.
http://dx.doi.org/10.1007/BF00383250
Zhang. X; Ahmadi. G: Qian. J; Ferro. A. (2008). Particle detachment, resuspension and transport due to
human walking in indoor environments. J Adhes Sci Tech 22: 591-621.
http://dx.doi.org/10.1163/156856108X305624
Zhang. Y; Griffin. A; Rahman. M; Camper. A; Baribeau. H; Edwards. M. (2009b). Lead contamination of
potable water due to nitrification. Environ Sci Technol 43: 1890-1895.
http://dx.doi.org/10.1021/es802482s
Zhao. Y; Wang. L; Shen. HB; Wang. ZX; Wei. QY; Chen. F. (2007). Association between delta-
aminolevulinic acid dehydratase (ALAD) polymorphism and blood lead levels: A meta-regression
analysis. J Toxicol Environ Health 70: 1986-1994. http://dx.doi.org/10.1080/15287390701550946
Ziegler. EE; Edwards. BB; Jensen. RL; Mahaffev. KR; Fomon. SJ. (1978). Absorption and retention of
lead by infants. Pediatr Res 12: 29-34. http://dx.doi.org/10.1203/00006450-197801000-00008
November 2012 4-178 Draft - Do Not Cite or Quote
-------
5 INTEGRATED HEALTH EFFECTS OF LEAD
EXPOSURE
5.1 Introduction
1 This chapter summarizes, integrates, and evaluates the evidence for the broad spectrum of
2 health effects associated with exposure to Pb. The chapter begins (Section 5.2) with a
3 discussion of the evidence for the modes of action that mediate the health effects of Pb,
4 including those modes of action that are shared by all of the health effects evaluated in
5 this ISA and those modes of action that are specific to particular endpoints. Subsequent
6 sections comprise evaluations of the epidemiologic and toxicological evidence for the
7 effects of Pb exposure on health outcomes related to nervous system effects (Section 5.3).
8 cardiovascular effects (Section 5.4). renal effects (Section 5.5). immune effects
9 (Section 5.6). hematological effects (Section 5.7). and reproductive and developmental
10 effects (Section 5.8). Section 5.9 reviews the evidence for the effects of Pb on other
11 noncancer health outcomes, for which the cumulative bodies of evidence are smaller,
12 including those related to the hepatic system (Section 5.9.1). gastrointestinal system
13 (Section 5.9.2). endocrine system (Section 5.9.3). bone and teeth (Section 5.9.4). ocular
14 health (Section 5.9.5). and respiratory system (Section 5.9.6). Chapter 5 concludes with a
15 discussion of the evidence for Pb effects on cancer (Section 5.10).
16 Individual sections for major outcome categories (e.g., nervous system, cardiovascular,
17 renal) begin with a brief summary of conclusions from the 2006 Pb AQCD (U.S. EPA.
18 2006c) followed by an evaluation of recent (i.e., published since the completion of the
19 2006 Pb AQCD) studies that is intended to build upon evidence from previous reviews.
20 Within each of these sections, results are organized by endpoint (e.g., cognitive function,
21 behavior, neurodegenerative diseases) then by specific scientific discipline
22 (i.e., epidemiology, toxicology). This chapter evaluates evidence for both short- and long-
23 term Pb exposures, which are defined as less than four weeks and greater than
24 four weeks, respectively, in animal toxicological studies and less than one year and
25 greater than one year, respectively, in epidemiologic studies (Section 2.1).
26 Sections for each of the major outcome categories (e.g., nervous system, cardiovascular,
27 renal effects) conclude with an integrated summary of the evaluation of evidence and a
28 conclusion regarding causality. Based upon the framework (described in the Preamble to
29 this ISA), a determination of causality was made for a group of related endpoints within a
30 major outcome category (e.g., cognitive function, attention-related behavioral problems).
31 In judgments regarding causality, emphasis was placed on studies with relevant Pb
32 exposure routes and concentrations in toxicological studies and internal dose measures in
November 2012 5-1 Draft - Do Not Cite or Quote
-------
1 epidemiologic studies (generally one order of magnitude above blood Pb levels in the
2 current U.S. population as described in Section 2.1). Studies that examined higher Pb
3 concentrations were evaluated particularly to inform mode of action. Further, evidence
4 was evaluated for consistency of findings across multiple studies and the extent to which
5 chance, confounding (i.e., bias due to a correlation with Pb biomarker level and causal
6 association with the outcome), and other biases could be ruled out with reasonable
7 confidence. Such evidence included high quality epidemiologic studies with
8 representative population-based groups or samples, prospective versus cross-sectional or
9 ecologic design; rigorous statistical analysis (i.e., multivariate regression) with
10 assessment of potential confounding factors; information on the concentration-response
11 relationship; and supporting toxicological evidence. The extent of consideration for
12 potential confounding varied among epidemiologic studies. Because no single study
13 considered all potential confounding factors, and not all potential confounding factors
14 were examined in the collective body of evidence, residual confounding by unmeasured
15 factors is possible. However, the examination of factors well documented in the literature
16 to be associated with Pb exposure and health outcomes and supporting toxicological
17 evidence help to minimize the undue influence of confounding bias on the observed
18 epidemiologic associations. The biological plausibility provided by the coherence of
19 evidence between toxicology and epidemiology and across a spectrum of related
20 endpoints, including evidence for modes of action, was used as support to address
21 uncertainties in the epidemiologic evidence due to biases from factors such as selective
22 publication, recruitment or participation of subjects; reverse causality; or confounding.
5.2 Modes of Action
5.2.1 Introduction
23 The diverse health effects associated with Pb exposure are dependent on multiple factors,
24 including the concentration and duration of exposure, the particular Pb compounds
25 constituting the exposure, and which tissues are affected. Pb exposure is linked to
26 downstream health effects by various modes of action. A mode of action (MOA) is the
27 common set of biochemical, physiological, or behavioral responses (i.e., empirically
28 observable precursor steps) that can cumulatively result in the formation of negative
29 health outcomes. Although the effects of Pb exposure appear to be mediated through
30 multiple modes of action, alteration of cellular ion status (including disruption of calcium
31 homeostasis, altered ion transport mechanisms, and perturbed protein function through
32 displacement of metal cofactors) seems to be the major unifying mode of action
33 underlying all subsequent modes of action (Figure 5-1). This section draws information
November 2012 5-2 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
from all of the subsequent health effects sections in Chapter 5, and identifies the major
modes of action operating at the molecular, cellular, and tissue/organ level. In turn, the
individual health effect sections bridge these MOA effects to those observed on the
organismal level. Each of the individual health effect sections includes a more detailed
description of the mechanisms specific to the individual health effect. Accordingly, this
section differs in structure and content from other health effects sections as it does not
primarily focus on the literature published since the 2006 Pb AQCD, but rather
incorporates recent information with earlier studies (which together represent the current
state of the science) on the possible modes of action of Pb. Higher concentrations of Pb
are often utilized in mode of action studies. This section includes some studies that are
conducted at concentrations greater than one order of magnitude above the upper end of
the blood Pb distribution of the general U.S. population when it is likely that the mode of
action does not differ at higher concentrations.
Oxidative
Stress
(5.2.4)
Cell Death
Genotoxicity
(5.2.7)
Inflammation
(5.2.5)
Endocrine
Disruption
(5.2.6)
Note: The subsections where these MOAs are discussed are indicated in parentheses.
(Section 5.2.2: Section 5.2.3: Section 5.2.4: Section 5.2.5: Section 5.2.6: and Section 5.2.7).
Figure 5-1 Schematic representation of the relationships between the
various MOAs by which Pb exposure exerts its health effects.
November 2012
5-3
Draft - Do Not Cite or Quote
-------
5.2.2 Altered Ion Status
1 Physiologically-relevant metal ions (e.g., Ca2+, Mg, Zn, Fe) are known to have a
2 multitude of functions in biological systems, including roles as charge carriers,
3 intermediates in enzymatically-catalyzed reactions, and structural elements in the proper
4 maintenance of tertiary protein conformations (Garza et al.. 2006). It is through
5 disruption of these biological functions that Pb exerts its negative actions, ultimately
6 interfering with such tightly regulated processes as cell signaling, intracellular ion
7 homeostasis, ion transport, energy metabolism, and enzymatic function.
5.2.2.1 Disruption of Ca2+ Homeostasis
8 Calcium (Ca2+) is one of the most important carriers of cell signals and regulates virtually
9 all aspects of cell function, including energy metabolism, signal transduction, hormonal
10 regulation, cellular motility, and apoptosis (Carafoli. 2005). Ca2+ homeostasis is
11 maintained through a tightly regulated balance of cellular transport and intracellular
12 storage (Pentvala et al.. 2010). Disruption of Ca2+ homeostasis by Pb has been observed
13 in a number of different cell types and cell-free environments, indicating that this is a
14 major mode of action for Pb-induced toxicity on a cellular level.
15 Ca2+ homeostasis is particularly important in bone cells, as the skeletal system serves as
16 the major dynamic reservoir of Ca2+ in the body (Wiemann et al., 1999; Long et al.,
17 1992). Bone cells also are unique in that they exist in a microenvironment that is high in
18 Ca2+, and potentially high in Pb concentrations. This may increase their relative exposure
19 to Pb and thus Pb-induced effects (Long et al.. 1992). A series of studies from the
20 laboratory of Long, Dowd, and Rosen have indicated that exposure of cultured
21 osteoblastic bone cells to Pb alters intracellular Ca2+ levels ([Ca2+]0. Exposure of
22 osteoblasts to 1, 5, or 25 (iM Pb for 40-300 minutes resulted in prolonged increases in
23 [Ca2+]j of 36, 50 and 120% over baseline, respectively (Schanne etal.. 1997; Schanne et
24 al.. 1989). Long et al. (1992) observed that exposure of osteoblasts to either 400 ng
25 parathyroid hormone (PTH)/mL culture medium for 1 hour or 25 (iM Pb for 20 hours
26 increased [Ca2+]j. Pb-exposed cells pretreated with PTH increased [Ca2+]j above
27 concentrations observed in either single exposure (Pb alone or PTH alone), indicating
28 that Pb may disrupt the ability of bone cells to respond to normal hormonal control. A
29 similar increase in [Ca2+]j was also observed when bone cells were co-treated with
30 epidermal growth factor (EGF, 50 ng/mL) plus Pb (5 (iM), versus EGF alone (Long and
31 Rosen. 1992). Pb-induced increases in [Ca2+]j were blocked by a protein kinase C (PKC)
32 inhibitor, indicating that PKC activation may serve as one mechanism by which Pb
33 perturbs [Ca2+]j (Schanne etal.. 1997). Schirrmacher et al. (1998) also observed
November 2012 5-4 Draft - Do Not Cite or Quote
-------
1 alterations in Ca2+ homeostasis in osteoblasts exposed to 5 (iM Pb for 50 minutes due to
2 potential disruption of Ca2+ATPases. However, Wiemann et al. (1999) demonstrated that
3 exposure to 5 or 12.5 (iM Pb inhibited the Ca2+-release-activated calcium influx of Ca2+
4 independently of any inhibitory effect on Ca2+ATPases.
5 Ca2+ homeostasis has also been shown to be disturbed in erythrocytes exposed to Pb
6 (Quintanar-Escorza et al., 2010; Quintanar-Escorza et al., 2007; Shin et al.. 2007). In
7 blood samples taken from Pb-exposed workers (mean [SD] blood Pb level: 74.4
8 [21.9] (ig/dL), the [Ca2+]j was approximately 2.5-fold higher than that seen in nonexposed
9 workers (mean [SD] blood Pb level: 9.9 [2.0] (ig/dL) (Ouintanar-Escorza et al.. 2007).
10 The increase in [Ca2+]j was associated with higher osmotic fragility and modifications in
11 erythrocyte shape. In a separate investigation, when erythrocytes from 10 healthy
12 volunteers were exposed (in vitro) at concentrations of 0.2 to 6.0 (iM Pb for 24 or 120
13 hours, concentration-related increases in [Ca2+]j were observed across all concentrations
14 for both durations of exposure (Quintanar-Escorza et al.. 2010). Subsequent exposures of
15 erythrocytes to either 0.4 or 4.0 (iM Pb [corresponding to 10 or 80 (ig/dL in exposed
16 workers (Quintanar-Escorza et al.. 2007)] for 12-120 hours resulted in duration-related
17 increases with durations >12 hours. Osmotic fragility (measured as percent hemolysis)
18 was increased in erythrocytes exposed to 0.4 (iM Pb for 24 hours. Co-incubation with a
19 vitamin E analog mitigated these effects, indicating that the increase in [Ca2+]j is
20 dependent on the oxidative state of the erythrocytes. Shin et al. (2007) observed that
21 incubation of human erythrocytes with 5 (iM Pb for 1 hour resulted in a 30-fold increase
22 in [Ca2+]j in vitro, inducing the pro-coagulant activity of exposed erythrocytes. Induction
23 of pro-coagulant activity in erythrocytes could lead to thrombus formation and negatively
24 contribute to overall cardiovascular health; whereas increased osmotic fragility could
25 substantially reduce erythrocyte life span and ultimately lead to anemic conditions.
26 Similar to effects seen in erythrocytes, Pb has been observed to interfere with Ca2+
27 homeostasis in platelets and white blood cells. Dowd and Gupta (1991) observed that
28 1 (iM Pb (for 3.5 hours) was the lowest exposure concentration to result in increases in
29 [Ca2+]j in human platelets (in vitro). The observed increase in [Ca2+]j levels was attributed
30 to the increased influx of external Ca2+, possibly through ligand-gated Ca2+ channels. In
31 mouse splenic lymphocytes, 1 (iM Pb was the lowest exposure concentration found to
32 increase [Ca2+]j with incubation periods of 10 minutes or greater (Li et al., 2008c). These
33 increases in [Ca2+]j appeared to be reversible as [Ca2+]j returned to baseline after one
34 hour. Pretreatment with a calmodulin antagonist slightly mitigated the effects of Pb
35 exposure, indicating a role for calmodulin in disruption of Ca2+ homeostasis by Pb
36 exposure in lymphocytes. In rat tail arteries exposed to 1.2 (iM Pb acetate for 1 hour,
37 [Ca2+]j increased over controls, possibly through increased transmembrane influx of
38 external Ca2+ (Piccinini et al.. 1977).
November 2012 5-5 Draft - Do Not Cite or Quote
-------
1 Exposure of the microsomal fraction (prepared from rat brain cells) to as little as 0.25
2 Pb for 2 minutes resulted in increased release of Ca2+ into the culture medium (Pentvala
3 et al., 2010). Further, Pb exposure also decreased the activity of microsomal Ca2+ATPase,
4 thus decreasing the sequestration of Ca2+ into microsomes. The results of this study
5 suggest that disruption of microsomal release and re-uptake of Ca2+ may alter Ca2+
6 homeostasis, ultimately leading to altered signal transduction and neuronal dysfunction.
7 However, Ferguson et al. (2000) observed that [Ca2+]j was decreased in rat hippocampal
8 neurons in response to exposure to 0.1 (iM Pb for 1-48 hours; although the observed
9 decreases were not time-dependent. The decrease in [Ca2+]j was shown to be due to
10 increased efflux of Ca2+ out of the neuron via a calmodulin-regulated mechanism,
11 possibly through stimulated Ca2+ efflux via Ca2+ATPase.
12 Pb exposure has been shown to disrupt [Ca2+]j levels in multiple cell types including
13 osteoblasts, erythrocytes, platelets, and neuronal cells. This alteration in Ca2+ homeostasis
14 could potentially affect cell signaling and disrupt the normal physiological function of
15 these cells.
5.2.2.2 Disruption of Ion Transport Mechanisms
16 As described above, deregulation of Ca2+ homeostasis can result in negative effects in
17 multiple organ systems. Under normal conditions in the life cycle of most cells, cytosolic
18 concentrations of free Ca2+ fluctuate between approximately 100 to 200 nM and Ca2+that
19 has entered the cell must be removed in order to maintain normal homeostatic
20 concentrations (Carafoli, 2005). An important component in the maintenance of Ca2+
21 homeostasis is transmembrane transport of Ca ions via Ca2+ATPase and voltage-gated
22 Ca2+ channels (Carafoli. 2005). Pb has been shown to disrupt the normal movement of
23 Ca2+ ions, as well as other physiologically important ions through interactions with these
24 transport mechanisms.
25 Multiple studies have reported alterations in the activity of Na+/K+ATPase, Ca2+ATPase,
26 and Mg2+ATPases after Pb exposure in animal models. Decreases in the activity of all
27 three ATPases were observed in the kidneys and livers of rats exposed to 750 ppm Pb in
28 drinking water for 11 weeks (mean [SD] blood Pb level: 55.6 [6.3] (ig/dL) (Kharoubi et
29 al., 2008a) and in erythrocytes from rats exposed to 2,000 ppm Pb in drinking water for
30 5 weeks (mean [SD] blood Pb level: 97.56 [11.8] jig/dL) (Sivaprasad et al.. 2003).
31 Increases in lipid peroxidation were seen in both studies, and the decrements in ATPase
32 activities may be explained by generation of free radicals in Pb-exposed animals. A
33 decrease in the activity of Na+/K+ATPase was observed in rabbit kidney membranes
34 exposed to 0.01 to 10 (iM Pb, possibly due to Pb inhibiting the hydrolytic cleavage of
November 2012 5-6 Draft - Do Not Cite or Quote
-------
1 phosphorylated intermediates in the K-related branch of the pump (Gramigni et al.,
2 2009). Similar decreases in Na+/K+ATPase activity were observed in brain synaptosomes
3 isolated from rats that were exposed to 200 ppm Pb in drinking water for 3 months (blood
4 Pb level: 37.8 ng/dL) (Rafalowska et al.. 1996) or 15 mg Pb/kg injected (i.p.) for 7 days
5 (blood Pb level: 112.5 jig/dL) (Struzynska et al.. 1997a). Inhibition of Na+/K+ATPase
6 activity was also observed in primary cerebellar granule neuronal cultures obtained from
7 rat pups that were pre- and post-natally (to PND8) exposed to Pb (1,000 ppm Pb acetate
8 in dams' drinking water, resulting in blood Pb level of 4 (ig/dL) (Baranowska-Bosiacka
9 et al.. 20 lib). The activity of Ca2+ATPase in the sarcoplasmic reticulum of rabbits
10 exposed to 0.01 (iM Pb was similarly decreased (Hechtenberg and Beyersmann. 1991).
11 The inhibitory effect of Pb was diminished in the presence of high Mg-ATP
12 concentrations. The activity of generic ATPase was reported to be altered in the testes of
13 rat pups exposed to 300 ppm (mg/L) Pb acetate, both during lactation and in drinking
14 water after weaning to the age of 6, 8, 10, or 12 weeks (Liu et al., 2008). In pregnant rats
15 fed a Pb-depleted (20 ± 5 ng/kg) or control (1 mg/kg) diet during gestation and lactation,
16 no difference was observed in the activity of Na+/K+ATPase and Ca2+/Mg2+ATPase in the
17 parental generation (Eder et al.. 1990). However, the offspring (exposed via placental and
18 lactational transfer of Pb) of Pb-depleted rats displayed decreased activities in both
19 enzymes compared with offspring of rats with higher Pb exposures. An increase in the
20 Na+/K+ATPase activity was observed in rats treated (i.p.) with 20 mg/kg Pb for
21 14 consecutive days (Jehan and Motlag. 1995). Co-exposure of Pb with Zn and Cu
22 greatly attenuated the increase in ATPase activity. Although the precise mechanism was
23 not investigated, Navarro-Moreno et al. (2009) reported that Ca2+ uptake was diminished
24 in proximal renal tubule cells in rats chronically exposed to 500 ppm Pb in drinking water
25 for 7 months (mean [SD] blood Pb level: 43.0 [7.6] (ig/dL).
26 In vitro studies of ATPase activities in human erythrocyte ghosts have also shown that Pb
27 affects the transport of metal ions across membranes. Calderon-Salinas et al. (1999a)
28 observed that 1-5 x 103 (iM Pb and Ca2+ were capable of inhibiting the passive transport
29 of each other in human erythrocyte ghosts incubated with both cations. Subsequent
30 inhibition experiments indicated that both cations share the same electrogenic transport
31 pathway (Sakuma et al., 1984). Further study by this group (Calderon-Salinas et al.,
32 1999b) demonstrated that Pb can noncompetitively block the transport of Ca2+ by
33 inhibiting the activity of Ca2+/Mg2+ATPase at concentrations of 1-5 x 103 (iM. Mas-Oliva
34 (1989) demonstrated that the activity of Ca2+/Mg2+ATPase in human erythrocyte ghosts
35 was inhibited by incubation with 0.1-100 (iM Pb. The inhibitory action was most likely
36 due to direct reaction with sulfhydryl groups on the ATPase enzyme at Pb concentrations
37 greater than 1 \iM, but due to the action of Pb on calmodulin at lower concentrations.
38 Grabowska and Guminska (1996) observed that 10 (ig/dL was the lowest
39 Pb concentration to decrease the activity of Na+/K+ATPase in erythrocyte ghosts; activity
November 2012 5-7 Draft - Do Not Cite or Quote
-------
1 of Ca2+/Mg2+ATPase was less sensitive to Pb exposure, and Mg2+ATPase activity was not
2 affected.
3 Effects on ATPase activity are also observed in association with blood Pb levels in
4 human populations. In a study investigating ATPase activities in Pb-exposed workers in
5 Nigeria, Abam et al. (2008) observed that the activity of erythrocyte membrane-bound
6 Ca2+/Mg2+ATPase was decreased by roughly 50% in all occupational groups (range of
7 mean [SD] blood Pb level across nine occupational groups: 28.75 [11.31] to 42.07
8 [12.01] (ig/dL) compared to nonexposed controls (mean [SD] blood Pb level: 12.34
9 [2.44] in males and 16.85 [6.01] (ig/dL in females). Higher membrane concentrations of
10 Ca2+ and Mg2+ were also observed, indicating that Pb prevented the efflux of those
11 cations from the cell, most likely by substituting for those metals in the active site of the
12 ATPase. In a study of 247 mother-newborn pairs, Campagna et al. (2000) observed that
13 newborn (cord) blood Pb (geometric mean [5th-95th percentile]: 4.8 [2.8-9.2] (ig/dL) was
14 negatively and significantly associated with maternal blood Ca2+ pump activities;
15 however, newborn (cord) blood Pb was not significantly associated with newborn (cord)
16 blood Ca2+ pump activities. Newborn hair Pb (geometric mean [5th-95th percentile]: 1.1
17 [0.1-8.0] (ig/g) was negatively and significantly associated with both maternal and
18 newborn (cord) blood Ca2+ pump activities. In a population of 81 newborns, Huel et al.
19 (2008) found that newborn hair and newborn (cord) blood Pb levels (mean [SD] newborn
20 hair Pb and newborn [cord] blood Pb levels: 1.22 [1.41] (ig/g and 3.54 [1.72] (ig/dL)
21 were negatively associated with Ca2+ATPase activity in plasma membranes of
22 erythrocytes isolated from newborn (cord) blood; newborn hair Pb levels were more
23 strongly associated with newborn (cord) Ca2+ pump activity than were newborn (cord)
24 blood Pb levels.
25 Pb has also been shown to disrupt cation transport mechanisms through direct action on
26 voltage-gated cation channels. Audesirk and Audesirk (1993. 1991) demonstrated that
27 extracellular free Pb inhibits the action of multiple voltage-gated Ca2+ channels, with free
28 Pb IC50 (half maximal inhibitory concentration) values of 0.7 (iM for L-type channels and
29 1.3 (iM for T-type channels in neuroblastoma cells maintained in culture media, and IC50
30 values as low as 0.03 (iM for L-type channels in cultured hippocampal neurons. Sun and
31 Suszkiw (1995) corroborated the inhibitory action of extracellular Pb on voltage-gated
32 Ca2+ channels, demonstrating an IC50 value of 0.3 (iM in bovine adrenal chromaffin cells.
33 The observed disruption of the voltage-gated Ca2+ channels most likely reflects
34 competition between Pb and Ca2+ for the extracellular Ca2+ binding domain of the
35 channel. Research by other laboratories supported these findings: Pb inhibited the action
36 of multiple Ca2+ channels in human embryonic kidney cells transfected with L-, N-, and
37 R-type channels (IC50 values of 0.38 (JVI, 1.31 (iM, and 0.10 (iM, respectively) (Peng et
38 al.. 2002) and P-type channels in cultured hippocampal neurons at concentrations up to
November 2012 5-8 Draft - Do Not Cite or Quote
-------
1 3 (iM (Ujihara et al., 1995). However, in bovine adrenal chromaffin cells, intracellular Pb
2 was observed to enhance Ca2+ currents through attenuation of the Ca2+ dependent
3 deactivation of Ca2+ channels at an EC50 value of 200 (iM, possibly through blocking the
4 intracellular Ca2+ binding domain, or through Ca2+ dependent dephosphorylation of the
5 channel (Sun and Suszkiw. 1995). Recently, Pb has also been shown to enter cells
6 (HEK293, HeLa, and PC 12 cell lines) through store-operated Ca2+ channels (Chiu et al..
7 2009; Chang et al., 2008b). In particular, the Orail-STIMl complex was shown to be
8 critical in the entry of Pb ions into cells, and increased Pb permeation was directly related
9 to decreased [Ca2+]j concentrations at exposure concentrations as low as 0.1
10 Pb also has been found to disrupt the action of Ca2+-dependent K+ channels. Alvarez et al.
1 1 (1986) observed that Pb promoted the efflux of K+ from inside-out erythrocyte vesicles in
12 a concentration-dependent manner at concentrations of 1-300 (iM, either through action
13 on a Mg2+ modulatory site or through direct interaction with the Ca2+ binding site. Fehlau
14 et al. (1989) also demonstrated Pb-induced activation of the K+ channel in erythrocytes.
15 However, Pb only activated the K+ channels at concentrations below 10 (iM; higher
16 concentrations of Pb completely inhibited channel activity, indicating the modulation of
17 K+ permeability is due to concentration dependent alterations in channel gating. Silken et
18 al. (2001) observed that Pb activated K+ channels in erythrocytes from the marine teleost
19 Scorpaena porcus in a concentration-dependent manner after a 20-minute incubation;
20 minor loss of K+ was seen at Pb concentrations of 1-2 (iM, whereas exposure to
21 20-50 (iM Pb resulted in approximately 70% K+ loss. Competitive and inhibitory binding
22 assays suggest that Pb directly activates K+ channels in S. porcus.
Disruption of Neurotransmitter Release
23 Pb has been shown to inhibit the evoked release of neurotransmitters by inhibiting Ca2+
24 transport through voltage-gated channels in in vitro experiments (Cooper and Manalis.
25 1984; Suszkiw et al.. 1984). However, in these same experiments, concentrations of Pb
26 > 5 (iM were also observed to actually increase the spontaneous release of
27 neurotransmitters. Subsequent research by other groups affirmed that Pb demonstrates
28 Ca2+-mimetic properties in enhancing neurotransmitter release from cells in the absence
29 of Ca2+ and Ca2+-induced depolarization. Tomsig and Suszkiw (1993) reported that Pb
30 exposure induced the release of norepinephrine (NE) from bovine adrenal chromaffin
3 1 cells, and was considerably more potent (as measured by half-maximal metal -dependent
32 release [K0 5]) than was Ca2+ (K0 5 of 4.6 x 10'3 jiM for Pb versus 2.4 jiM for Ca2+).
33 Activation of PKC was observed to enhance the Pb-induced release of NE (Tomsig and
34 Suszkiw. 1995). Westerink and Vijverberg (2002) observed that Pb acted as a high
35 affinity substitute for Ca2+, and triggered enhanced catecholamine release from PC 12
36 cells at 10 (iM in intact cells and 0.03 (iM in permeabilized cells. The suppression of
November 20 12 5-9 Draft - Do Not Cite or Quote
-------
1 Ca2+-evoked release of neurotransmitters combined with the ability of Pb to enhance
2 spontaneous releases could result in higher noise observed in the synaptic transmission of
3 nerve impulses in Pb-exposed animals.
4 In rats exposed to Pb at concentrations of 1,000-10,000 ppm in drinking water beginning
5 at gestational days GDI5-GDI6 and continuing to postnatal days PND120, decreases in
6 total K+-stimulated hippocampal gamma aminobutyric acid (GABA) release were seen at
7 exposure levels of 1,000-5,000 ppm (range of mean [SD] blood Pb levels: 26.8 [1.3] -
8 61.8 [2.9] (ig/dL) (Lasley and Gilbert. 2002). Maximal effects were observed at
9 2,000 ppm Pb in drinking water, but effects were less evident at 5,000 ppm, and were
10 absent at 10,000 ppm. In the absence of Ca2+, K+-induced GABA release was increased
11 with the two highest Pb exposure concentrations, suggesting a Pb-induced enhancement
12 of K+-evoked release of GABA. The authors suggest that this pattern of response
13 indicates that Pb is a potent suppressor of K+-evoked release at low concentrations, but a
14 Ca2+ mimic in regard to independently inducing exocytosis and evoking neurotransmitter
15 release at higher concentrations (Lasley and Gilbert. 2002). Suszkiw (2004) reports that
16 augmentation of spontaneous release of neurotransmitters may involve Pb-induced
17 activation of CaMKII-dependent phosphorylation of synapsin I or direct activation of
18 synaptotagmin I. Further, Suszkiw (2004) suggests that unlike the intracellularly
19 mediated effects of Pb on spontaneous release of neurotransmitters, Pb-induced inhibition
20 of evoked transmitter releases is largely due to extracellular blockage of the voltage-gated
21 Ca2+ channels.
22 In summary, Pb has been shown to disrupt ion transport mechanisms in toxicological and
23 epidemiologic studies. Specific mechanisms disrupted include various cation-specific
24 ATPases and voltage-gated cation channels. Alterations in ion transport functions have
25 also been shown to disrupt neurotransmitter release in both in vivo and in vitro
26 experiments.
5.2.2.3 Displacement of Metal Ions and Perturbed Protein
Function
27 The binding of metal ions to proteins causes specific changes in protein shape, and these
28 conformational changes may alter specific cellular function of many proteins (Kirberger
29 and Yang. 2008). Metal binding sites on proteins are generally ion-specific and are
30 influenced by multiple factors, including binding geometries, ligand preferences, ionic
31 radius, and metal coordination numbers (Kirberger and Yang. 2008; GarzaetaL 2006).
32 The coordination chemistry that normally regulates metal-protein binding makes many
33 proteins particularly susceptible to perturbation from Pb, as it is able to function with
November 2012 5-10 Draft - Do Not Cite or Quote
-------
1 flexible coordination numbers and can bind multiple ligands (Kirberger and Yang. 2008;
2 Garza et al.. 2006). However, due to differences in its physical properties, Pb induces
3 abnormal conformational changes when it binds to proteins (Kirberger and Yang, 2008;
4 Bitto et al.. 2006; Garza et al.. 2006; Magyar et al.. 2005). and these structural changes
5 elicit altered protein function. It is known that [Ca2+]j is an important second messenger
6 in cell signaling pathways, and operates by binding directly to and activating proteins
7 such as calmodulin and PKC (Goldstein. 1993). Alterations in the functions of both of
8 these proteins due to direct interaction with Pb have been well documented in the
9 literature.
10 PKC is a family of serine/threonine protein kinases critical for cell signaling and
11 important for cellular processes, including growth and differentiation (Goldstein. 1993).
12 PKC contains a "C2" Ca2+-binding domain and requires binding of the cation, as well as
13 the presence of diacylglycerol and phospholipids, for proper cellular activity (Garza et
14 al.. 2006). Markovac and Goldstein (1988b) observed that, in the absence of Ca2+,
15 exposure to 10"6 (iM concentrations of Pb for 5 minutes directly activated PKC purified
16 from rat brains. The activation of PKC by Pb was more potent than was Ca2+-dependent
17 activation by five orders of magnitude. Long et al. (1994) affirmed these findings,
18 reporting that Pb had a Kact 4,800 times smaller than that of Ca2+ (5.5 x 10"5 (iM versus
19 25 (iM, following a 3 minute exposure). However, Ca2+ had a higher maximal activation
20 of PKC than did Pb. This possibly indicates the presence of multiple Ca2+-binding sites
21 on the protein, and that Pb may bind the first site more efficiently than does Ca2+, but not
22 subsequent sites. Tomsig and Suszkiw (1995) further demonstrated the ability of Pb to
23 activate PKC in bovine adrenal chromaffin cells incubated with 10"6 (iM concentrations
24 of Pb for 10 minutes but also reported that activation of PKC by Pb was only partial
25 (approximately 40% of the maximum activity induced by Ca2+) and tended to decrease at
26 concentrations >1 x 10"3 (iM.
27 Contrary to the above findings, Markovac and Goldstein (1988a) observed that Pb and
28 Ca2+ activated PKC at equivalent concentrations and efficacies when broken cell
29 preparations of rat brain microvessels were incubated with either cation for 45 minutes.
30 However, when PKC activation was investigated in whole vessel preparations, no
31 activation was observed, but PKC did become redistributed from the cytosolic to the
32 particulate fraction after centrifugation. This suggests that Pb redistributes PKC at (iM
33 concentrations, but does not activate the protein in brain microvessels. In human
34 erythrocytes exposed to Pb acetate for 60 minutes, the amount of PKC found in
35 erythrocyte membranes and total PKC activity was increased at concentrations greater
36 than 0.1 (iM (Belloni-Olivi et al., 1996). The observation that neither Ca2+ nor
37 diacylglycerol concentrations were increased due to Pb exposure, indicates that
38 Pb-induced activation of PKC is due to direct interaction with the protein. Pb-induced
November 2012 5-11 Draft - Do Not Cite or Quote
-------
1 alterations in PKC have also been observed in other tissues, including increased activity
2 in rabbit mesenteric arteries at 10~6 (iM concentrations of Pb (Watts et al.. 1995; Chai and
3 Webb, 1988) and human erythrocytes from Pb-exposed workers (range of blood Pb
4 levels: 5.4 to 69.3 (ig/dL) (Hwang et al.. 2002). and decreased activity in mouse
5 macrophages and the rat brain cortex at (iM concentrations (Murakami et al., 1993; Lison
6 etal.. 1990).
7 Calmodulin is another important protein essential for proper Ca2+-dependent cell
8 signaling. Calmodulin contains an "EF-hand" Ca2+ binding domain, which is dependent
9 on the cation for proper activity (GarzaetaL 2006). Calmodulin regulates events as
10 diverse as cellular structural integrity, gene expression, and maintenance of membrane
11 potential (Vetter and Leclerc, 2003; Saimi and Kung. 2002). Habermann et al. (1983)
12 observed that exposure to Pb altered numerous cellular functions of calmodulin,
13 including activation of calmodulin-dependent phosphodiesterase activity after 10 minutes
14 incubation (minimal activation at 0.1 (iM, EC50 = 0.5-1.0 (JVI), stimulation of brain
15 membrane phosphorylation at Pb concentrations greater than 0.4 (iM after 1 minute
16 incubation, and increased binding of calmodulin to brain membranes at Pb concentrations
17 greater than 1 (iM after 10 minutes incubation. Habermann et al. (1983) reported that the
18 affinity of Pb for Ca2+-binding sites on calmodulin was approximate to that of Ca2+ itself
19 (Kd ~20 (JVI), whereas Richardt et al. (1986) observed that Pb was slightly more potent
20 than Ca2+ was at binding calmodulin (IC50 =11 and 26 (iM, respectively). Both studies
21 indicated that Pb was much more effective at binding to calmodulin than was any other
22 metal cation investigated (e.g., Hg, Cd, Fe). Kern et al. (2000) observed that Pb was more
23 potent in binding to, and affecting conformational changes in, calmodulin compared to
24 Ca2+ (EC50 values of 4-5.5 x 10'4 jiM [threshold = 1 x 10'4 jiM] and 0.45-0.5 jiM
25 [threshold = 0.1 |iM], respectively). Pb, in the absence of Ca2+, was also observed to
26 activate calmodulin-dependent cyclic nucleotide phosphodiesterase activity at much
27 lower concentrations compared to Ca2+ (EC50 value 4.3 x 10"4 (iM [threshold = 3 x
28 10"4 (iM] versus EC50 1.2 x 10"3 \iM [threshold = 0.2 (iM; 50 minute incubation]). When
29 incubated with physiological concentrations of Ca2+, Pb induced phosphodiesterase
30 activity at concentrations as low as 5 x 10"5 (iM. Pb activated calcineurin, a Ca2+-
31 dependent phosphatase with widespread distribution in the brain and immune system, at
32 threshold concentrations as low as 2 x 10"5 (iM in the presence of Ca2+ (incubation
33 time = 30 minutes), but inhibited its activity at concentrations greater than 2 x 10"4 (iM
34 (Kern and Audesirk. 2000). Thus, 10"6 (iM concentrations of intracellular Pb appear to
35 amplify the activity of calmodulin and thus can be expected to alter intracellular Ca2+
36 signaling in exposed cells (Kern et al.. 2000). Mas-Oliva (1989) observed that
37 low-exposure (<1 (iM, 20 minute incubation) stimulatory effects of Pb exposure on the
38 activity of Ca2+/Mg2+ATPase was due to Pb binding to calmodulin and subsequent
39 activation of the ion pore. Ferguson et al. (2000) observed that exposure of rat
November 2012 5-12 Draft - Do Not Cite or Quote
-------
1 hippocampal neurons to Pb for 1 to 48 hours resulted in increased activation of a
2 calmodulin-dependent Ca2+ extrusion mechanism.
3 Pb has also been observed to alter the activity of other proteins that rely on Ca2+ binding
4 for normal cellular function. Osteocalcin is a matrix protein important in bone resorption,
5 osteoclast differentiation, and bone growth; and has three Ca2+-binding sites (Dowd et al..
6 2001). Incubation of osteocalcin in solution with Ca2+ and Pb resulted in the competitive
7 displacement of Ca2+ by Pb (Dowdetal.. 1994). Pb was found to bind to osteocalcin
8 more than 1,000 times more tightly than was Ca2+ (Kd = 1.6 x 10"2 (iM versus 7.0 (iM,
9 respectively), and analysis with nuclear magnetic resonance (NMR) indicated that Pb
10 induced similar, though slightly different, secondary structures in osteocalcin, compared
11 to Ca2+. The authors hypothesized that the observed difference in Pb-bound osteocalcin
12 structure may explain previous findings in the literature that Pb exposure reduced
13 osteocalcin adsorption to hydroxyapatite (Dowd etal. 1994). Further research by this
14 group also found that Pb binded osteocalcin approximately 10,000-times more tightly
15 than did Ca2+ (Kd = 8.5 x 1Q"2 jiM versus 1.25 x 1Q3 jiM, respectively) (Dowd et al..
16 2001). However, the authors reported that Pb exposure actually caused increased
17 hydroxyapatite adsorption at concentrations 2-3 orders of magnitude lower than that seen
18 with Ca2+. Additionally, Pb can displace Ca2+ in numerous other Ca2+-binding proteins,
19 such as proteins important in muscle contractions, renal Ca2+ transport and
20 neurotransmission, including troponin C, parvalbumin, CaBP I and II, phospholipase A2,
21 and synaptotagmin I, at concentrations as low as the 10"3 (iM range (Bouton etal.. 2001;
22 Osterode and Ulberth. 2000: Richardt et al.. 1986).
23 Pb can displace metal cations other than Ca2+that are requisite for protein function. One
24 of the most researched targets for molecular toxicity of Pb is the second enzyme in the
25 heme synthetic pathway, aminolevulinic acid dehydratase (ALAD). ALAD contains four
26 Zn-binding sites and all four need to be occupied to confer full enzymatic activity
27 (Simons. 1995). ALAD has been identified as the major protein binding target for Pb in
28 human erythrocytes (Bergdahl et al., 1997a). and blood Pb levels are associated with
29 inhibition of the enzyme in the erythrocytes of Pb-exposed workers and adolescents
30 (blood Pb levels > 10 (ig/dL) (Ahamed et al.. 2006; Ademuyiwa et al.. 2005b). in lysed
31 human erythrocytes exposed to Pb in vitro for 60 minutes (K; = 7 x 10"6 (iM) (Simons.
32 1995). and in rats exposed to 25 mg/kg Pb once a week for 4 weeks (mean [SD] blood Pb
33 level: 6.56 [0.98] (ig/dL) (Lee et al.. 2005). Additional experiments indicated that lower
34 concentrations of Zn result in greater inhibition of enzyme activity by Pb, suggesting a
35 competitive inhibition between Zn and Pb at a single site (Simons. 1995).
36 Zn-binding domains are also found in transcription factors and proteins necessary for
37 gene expression, including GATA proteins and transcription factors TFIIIA, Spl, and
November 2012 5-13 Draft - Do Not Cite or Quote
-------
1 Erg-1 (Gheringetal.. 2005; Huang et al.. 2004; Reddv and Zawia. 2000; Hanas et al..
2 1999; Zawia et al.. 1998). Pb was found to form tight complexes with the cysteine
3 residues in GATA proteins (Pb stoichiometric stability constant (CF(31pb) = 6.4 (± 2.0)x
4 109 M'1 for single C-terminal GATA Zn finger from chicken and DFp2Pb2 = 6.3 (± 6.3) x
5 1019 M"2 for double-GATA Zn finger from human), and was able to displace bound Zn
6 from the protein under physiologically relevant conditions (Ghering et al.. 2005). Once
7 Pb was bound to GATA proteins, they displayed decreased ability to bind to DNA (Pb
8 concentrations > 1.25 (iM) and activate transcription. Pb at a minimum concentration of
9 10 (iM also binds to the Zn domain of TFIIIA, inhibiting its ability to bind DNA at
10 concentrations (Huang et al.. 2004; Hanas et al.. 1999). Huang et al. (2004) also reported
11 that exposure to Pb caused the dissociation of TFIIIA-DNA adducts and using NMR
12 spectroscopy, found that altered TFIIIA activity was the result of a Pb-induced abnormal
13 protein conformation.
14 Pb exposure modulated the DNA-binding profiles of the transcription factors Spl and
15 Erg-1 in rat pups exposed to 2,000 ppm Pb acetate via lactation, resulting in a shift in
16 DNA-binding toward early development (i.e., the first week following birth) (Reddy and
17 Zawia. 2000; Zawia et al.. 1998). The shifts in Spl DNA-binding profiles were shown to
18 be associated with abnormal expression of genes related to myelin formation
19 (Section 5.2.7.5). Further mechanistic research utilizing a synthetic peptide containing a
20 Zn finger motif demonstrated that Pb can bind the histidine and cysteine residues of the
21 Zn finger motif, thus displacing Zn and resulting in an increase in the DNA-binding
22 efficiency of the synthetic peptide (Razmiafshari et al.. 2001; Razmiafshari and Zawia.
23 2000). However, in DNA-binding assays utilizing recombinant Spl (which has three Zn
24 finger motifs, opposed to only one in the synthetic peptide), 37 (iM Pb was the lowest
25 concentration observed to abolish the DNA-binding capabilities of Spl (Razmiafshari
26 and Zawia. 2000).
27 Pb has also been reported to competitively inhibit Mg binding and thus inhibit the
28 activities of adenine and hypoxanthine/guanine phosphoribosyltransferase in erythrocyte
29 lysates from rats exposed to 1,000 ppm Pb in drinking water for 9 months (mean [SD]
30 blood Pb level: 7.01 [1.64] (ig/dL) and in in vitro human erythrocyte lysates exposed to
31 0.1 (iM Pb for as little as 5 minutes (Baranowska-Bosiacka et al.. 2009). and cGMP
32 phosphodiesterase at 10"6 (iM concentrations in homogenized bovine retinas (Srivastava
33 et al.. 1995). Pb was also reported to inhibit pyrimidine 5'-nucleotidase through
34 competitive inhibition of Mg binding, resulting in conformational changes and improper
35 amino acid positioning in the active site (Bitto et al.. 2006).
November 2012 5-14 Draft - Do Not Cite or Quote
-------
1 In summary, Pb has been shown to displace metal cations from the active sites of
2 multiple enzymes and proteins, and thus to alter the functions of those proteins in
3 occupationally-exposed humans with blood Pb levels of 5.4-69.3 (ig/dL, in adult rodents
4 with blood Pb levels of 6.5 (ig/dL (exposure 4 weeks), in suckling rats exposed to
5 2,000 ppm Pb via lactation, and in cell-free and cellular in vitro experiments conducted at
6 exposure concentrations ranging from 10"6 (iM to 1 (iM. These alterations in protein
7 function have implications for numerous cellular and physiological processes, including
8 cell signaling, growth and differentiation, gene expression, energy metabolism, and
9 biosynthetic pathways. Table 5-1 provides a list of enzymes and proteins whose function
10 may be perturbed by Pb exposure.
November 2012 5-15 Draft - Do Not Cite or Quote
-------
Table 5-1 Enzymes and proteins potentially affected by exposure to Pb and the
metal cation cofactors necessary for their proper physiological
activity.
Enzymes
Ion Channels/
Transport
Signal
Transduction
Pb Binding
DMA Binding
Metalloprotein/Enzyme
Aminolevulinic acid
dehydratase
Ferrochelatase
Superoxide dismutase
Catalase
Glutathione peroxidase
Guanylate cyclase
cGMP phosphodiesterase
NAD synthase
NAD(P)H oxidase
Pyrimidine 5'-nucleotidase
Erythrocyte
phosphoribosyltransferase
ATPase
Mitochondrial
transmembrane pore
Calcium-dependent
potassium channel
Protein kinase C
Calmodulin
Metallothionein
GATAtranscriptional factors
a^ indicates increased activity; J, indicates decreased
Direction
1
1
IT
IT
IT
1
1
1
T
1
1
IT
T
T
IT
T
T
1
activity; J,f
of Action3 Metal Cation; Reference
Zn; Simons (1995)
Fe (2Fe-2S Cluster);
Crooks et al. (2010)
Mn, Cu, Zn, Fe;
Antonyuk et al. (2009),
Borgstahl et al. (1992)
Fe (Heme); Putnam et al. (2000)
Se; Rotruck et al. (1973)
Fe (Heme);
Boerrigter and Burnett (2009)
Mg, Zn; Ke (2004)
Mg; Hara et al. (2003)
Ca2+; Leseney et al. (1999)
Mg, Ca2+;
Bitto et al. (2006).
Amici et al. (1997).
Paglia and Valentine (1975)
Mg (Mn, Ca2+, Co, Ni, Zn);
Dengetal. (2010),
Arnold and Kelley (1978)
Ca2+, Mg, Na/K; Technische
Universitat Braunschweig (2011)
Ca2+; He et al. (2000)
Ca2+;
Silkin et al. (2001),
Alvarez et al. (1986)
Ca2+; Garza et al. (2006)
Ca2+; Garza et al. (2006)
Zn, Cu; Yu et al. (2009)
Zn;
Hanas et al. (1999),
Huang et al. (2004)
indicates activity can be alternatively increased or
decreased.
November 2012
5-16
Draft - Do Not Cite or Quote
-------
5.2.2.4 Mitochondrial Abnormality
1 Alterations in mitochondrial function, including disruptions in ion transport,
2 ultrastructural changes, altered energy metabolism, and perturbed enzyme activities due
3 to Pb exposure are well documented in the scientific literature. Exposure of rats to Pb in
4 feed (10,000 ppm Pb for 4, 6, 8, 10, 12, or 20 weeks) or drinking water (300 ppm for
5 8 weeks, 500 ppm for 7 months, or 10,000 ppm Pb for 9 months) resulted in gross
6 ultrastructural changes in renal tubule mitochondria and epididymal mitochondria
7 characterized as a general swollen appearance with frequent rupture of the outer
8 membrane, distorted cristae, loss of cristae, frequent inner compartment vacuolization,
9 observation of small inclusion bodies, and fusion with adjacent mitochondria (Wang et
10 al..2010d: Marchlewicz et al.. 2009; Navarre-Moreno et al.. 2009; Gover. 1968; Gover et
11 al.. 1968).
12 Transmembrane mitochondrial ion transport mechanisms have been found to be
13 perturbed by exposure to Pb. Pb inhibits the uptake of Ca2+ into mitochondria (Parr and
14 Harris. 1976). while simultaneously stimulating the efflux of Ca2+ out of the organelle
15 (Simons. 1993a). thus disrupting intracellular/mitochondrial Ca2+ homeostasis. Pb
16 exposure has also been shown to decrease the mitochondrial transmembrane potential in
17 primary cerebellar granule neuronal cultures from rats exposed to 1,000 ppm Pb in
18 drinking water throughout gestation and lactation (Baranowska-Bosiacka et al., 201 Ib).
19 astroglia incubated with 0.1 or 1.0 (iM Pb for 14 days (Legare etal.. 1993). proximal
20 tubule cells exposed to 0.25, 0.5, and 1.0 (iM for 12 hours (Wang et al.. 2009c). and
21 retinal rod photoreceptor cells incubated with 0.01 to 10 (iM for 15 minutes (He et al..
22 2000). Further research indicated that Pb-induced mitochondrial swelling and decreased
23 membrane potential is the result of the opening of a mitochondrial transmembrane pore
24 (MTP), possibly by directly binding to the metal (Ca2+)-binding site on the matrix side of
25 the pore (Bragadin et al.. 2007; He et al.. 2000). Opening of the MTP is the first step of
26 the mitochondrial-regulated apoptotic cascade pathway in many cells (Rana. 2008;
27 Lidsky and Schneider. 2003). He et al. (2000) additionally observed other indicators of
28 apoptosis including, cytochrome c release from mitochondria, and caspase-9 and -3
29 activation following exposure of retinal rod cells to Pb. Induction of mitochondrially-
30 regulated apoptosis via stimulation of the caspase cascade following exposure to Pb has
31 also been observed in rat hepatic oval cells (Agarwal et al.. 2009).
Altered Energy Metabolism
32 Pb has been reported to alter normal cellular bioenergetics. In mitochondria isolated from
33 the kidneys of rats exposed to 10,000 ppm Pb in feed for 6 weeks, the rate of oxygen
34 uptake during ADP-activated (state 3) respiration was lower compared to controls (Gover
November 2012 5-17 Draft - Do Not Cite or Quote
-------
1 et al.. 1968). The rate of ATP formation from exposed mitochondria was observed to be
2 approximately 50% that of control mitochondria. A decrease in state 3 respiration and
3 respiratory control ratios (state 3/state 4 [succinate or pyruvate/malate-activated]) was
4 also observed in kidney mitochondria from rats exposed continuously from conception to
5 six or nine months of age (i.e., gestationally, lactationally, and via drinking water after
6 weaning) to 50 or 250 ppm Pb (Fowler et al.. 1980). Pb-induced decreases in ATP and
7 adenylate energy charge (AEC) were observed concurrently with increases in ADP,
8 AMP, and adenosine in adult rats exposed to 10,000 ppm Pb in drinking water for
9 9 months (Marchlewicz et al., 2009). Similarly, ATP and AEC were decreased, and AMP
10 increased, in primary cerebellar granule neuronal cultures from rats exposed to
1 1 1,000 ppm Pb in drinking water throughout gestation and lactation (Baranowska-
12 Bosiacka et al., 201 Ib). One (iM Pb (48 hours) was the lowest concentration observed to
13 decrease cellular ATP levels in NGF-differentiated PC- 12 cells, and these changes were
14 correlated with a Pb-induced decrease in the expression of the mitochondrial voltage-
15 dependent anion channel, which maintains cellular ATP levels in neurons (Prins et al..
16 2010). Dowd et al. (1990) reported that oxidative phosphorylation was decreased up to
17 74% after exposure of osteoblasts to 10 (iM Pb. Parr and Harris (1976) reported that Pb
18 inhibited both coupled and uncoupled respiratory oxygen use in mitochondria, and that
19 Pb prevented pyruvate, but not malate, uptake. Mitochondrial levels of ATP were
20 diminished after Pb exposure, and the authors compared the effects of Pb on the energy
21 supply to the actions of classic respiratory inhibitors, low temperature, and chemical
22 uncouplers. Bragadin et al. (1998) supported this view by demonstrating that alkylated Pb
23 compounds acted as a chemical uncoupler of respiration by abolishing the proton gradient
24 necessary for oxidative phosphorylation. Further, the enzymatic activities of complex I
25 and IV of the respiratory chain have been shown to be decreased in the peroneous longus
26 muscle of rats exposed to 250 ppm Pb (or 5 ppm thallium) in drinking water for 90 days
27 (Mendez-Armenta et al.. 201 1). Contrary to the above findings, Rafalowska et al. (1996)
28 reported that, although ATP levels did decrease in the forebrain synaptosomes prepared
29 from rats exposed to 200 ppm Pb in water for 3 months, this chronic exposure to Pb did
30 not inhibit oxidative phosphorylation in the synaptosomal mitochondria. Similar effects
3 1 with regard to the activity of the mitochondrial oxidative chain were observed in rats
32 injected with 15 mg Pb/kg (i.p.) daily for seven days, as reported by Struzynksa et al.
33 (1997a). although ATP levels were reported to increase after exposure to Pb.
34 Pb has also been shown to decrease glycolysis in osteoblasts exposed to 10 (iM Pb and in
35 human erythrocytes exposed (in vitro) to 30 (ig/dL Pb (Grabowska and Guminska. 1996;
36 Dowd et al.. 1990). Contrary to these findings, Antonowicz et al. (1990) observed higher
37 levels of glycolytic enzymes in erythrocytes obtained from Pb workers directly exposed
38 to Pb, compared to workers exposed to lower concentrations of Pb (blood Pb levels: 82.1
39 versus 39.9 (ig/dL), and suggested that Pb activated anaerobic glycolysis. In vitro
November 2012 5-18 Draft - Do Not Cite or Quote
-------
1 exposure of human umbilical cord erythrocytes to 100-200 (ig/dL Pb for 20 hours was
2 observed to lower the cellular pools of adenine and guanine nucleotide pools, including
3 NAD and NADPH (Baranowska-Bosiacka and Hlynczak. 2003). These decreases in
4 nucleotide pools were accompanied by an increase in purine degradation products
5 (adenosine, etc.). Similar decreases in cellular nucleotide pools were observed when rats
6 were exposed to 10,000 ppm Pb in drinking water for four weeks (Baranowska-Bosiacka
7 and Hlynczak. 2004). In erythrocytes, nucleotides are synthesized via salvage pathways
8 such as the adenine pathway, which requires adenine phosphoribosyltransferase. The
9 activity of this enzyme is inhibited by exposure to Pb in human and rat erythrocytes (see
10 above for concentration and duration) (Baranowska-Bosiacka et al.. 2009).
11 Disruptions in erythrocyte energy metabolism have been observed in adults
12 occupationally exposed to Pb. Nikolova and Kavaldzhieva (1991) reported higher ratios
13 of ATP/ADP in Pb-exposed workers with an average duration of exposure of 8.4 years
14 (blood Pb not reported) than in unexposed controls. Morita et al. (1997) evaluated the
15 effect of Pb on NAD synthetase in the erythrocytes of Pb-exposed workers (mean [SD]
16 blood Pb level: 34.6 [20.7] (ig/dL) and observed an apparent concentration-dependent
17 decrease in NAD synthetase activity with increased blood Pb level. The blood Pb level
18 associated with 50% inhibition of NAD synthetase, which requires a Mg2+ cation for
19 activity (Hara et al., 2003), was 43 (ig/dL.
Altered Heme Synthesis
20 Exposure to Pb is demonstrated to inhibit two key steps in the synthesis of heme:
21 porphobilinogen synthase (i.e., 5-aminolevulinic acid dehydratase), a cytoplasmic
22 enzyme requiring Zn for enzymatic activity that condenses two molecules of
23 aminolevulinic acid into porphobilinogen, and ferrochelatase, a mitochondrial iron-sulfur
24 containing enzyme that incorporates Fe2+ into protoporphyrin IX to create heme. Farant
25 and Wigfield (1990. 1987) observed that Pb inhibits the activity of porphobilinogen
26 synthase in rabbit and human erythrocytes, and that the effect on the enzyme was
27 dependent on the affinity for thiol groups at its active site. Taketani et al. (1985)
28 examined the activity of Pb on ferrochelatase in rat liver mitochondria and observed that
29 10 (iM Pb (30 minute incubation) reduced NAD(P)H-dependent heme synthesis by half
30 when ferric, but not ferrous, iron was used. Pb inhibits the insertion of Fe2+ into the
31 protoporphyrin ring and instead, Zn is inserted into the ring creating Zn-protoporphyrin
32 (ZPP). While not directly measuring the activity of ferrochelatase, numerous studies have
33 shown that blood Pb levels are associated with increased erythrocyte ZPP levels in
34 humans (mean blood Pb levels ranging from 21.92 to 53.63 (ig/dL) (Mohammad et al.,
35 2008; Counter et al.. 2007; Patil et al.. 2006b: Ademuyiwa et al.. 2005b) and in animals
36 (blood Pb level: 24.7 (ig/dL) (Rendon-Ramirez et al.. 2007).
November 2012 5-19 Draft - Do Not Cite or Quote
-------
1 In summary, Pb has been shown to disrupt mitochondrial function including
2 transmembrane mitochondrial ion transport mechanisms and has been shown to perturb
3 normal cell bioenergetics. These effects have not only been demonstrated in in vitro
4 toxicological studies but also exposed worker populations.
5.2.3 Protein Binding
5 Evidence indicates that Pb binds to proteins within cells through interactions with side
6 group moieties (e.g., thiol residues) to form inclusion bodies and can thereby potentially
7 disrupt cellular function (Sections 5.2.2.3 and 5.2.2.4). However, some proteins are also
8 able to bind Pb and protect against its negative effects through sequestration. The ability
9 of Pb to bind proteins was first reported by Blackman (1936). In children exposed to high
10 levels of Pb, formation of intranuclear inclusion bodies in the liver and kidney was
11 observed. Since that time, further research has been conducted into characterizing the
12 composition of intranuclear inclusion bodies and identifying specific Pb-binding proteins.
5.2.3.1 Intranuclear and Cytoplasmic Inclusion Bodies
13 Goyer (1968) and Goyer et al. (1968) observed the formation of intranuclear inclusion
14 bodies in the renal tubules of rats fed 10,000 ppm Pb in food for up to 20 weeks. The
15 observation of inclusion bodies was accompanied by altered mitochondrial structure and
16 reduced rates of oxidative phosphorylation. Pb has further been observed to form
17 cytoplasmic inclusion bodies preceding the formation of the intranuclear bodies, and to
18 be concentrated within the subsequently induced intranuclear inclusion bodies following
19 i.p. injection, drinking water, and dietary exposures (Navarre-Moreno et al., 2009;
20 Oskarsson and Fowler. 1985: Fowler etal.. 1980: McLachlin et al.. 1980: Choie and
21 Richter. 1972: Goyer etal.. 1970b: Goyer etal.. 1970a). Inclusion bodies have also been
22 observed in the mitochondria of kidneys and the perinuclear space in the neurons of rats
23 exposed to 500 ppm Pb acetate in drinking water for 60 days or 7 months (Navarro-
24 Moreno et al., 2009: Deveci, 2006). Intranuclear and cytoplasmic inclusions have also
25 been found in organs other than the kidney, including liver, lung, and glial cells (Singh et
26 al.. 1999: Gover and Rhvne. 1973). Pb found within nuclei has also been shown to bind
27 to the nuclear membrane and histone fractions (Sabbioni and Marafante. 1976).
28 Upon denaturing intranuclear inclusion bodies with strong denaturing agents, Moore et
29 al. (1973) observed that proteins included in the bodies were rich in aspartic and glutamic
30 acid, glycine, and cysteine. Further work by Moore and Goyer (1974) characterized the
31 protein as a 27.5 kilodalton (kDa) protein that migrates as a single band on
November 2012 5-20 Draft - Do Not Cite or Quote
-------
1 polyacrylamide gel electrophoresis. In contrast with the findings of Moore and Goyer,
2 Shelton and Egle (1982) identified a 32 kDa protein with an isoelectric point of 6.3 from
3 the kidneys of rats exposed to 10,000 ppm Pb acetate in feed or 7,500 ppm in drinking
4 water. This protein, dubbed p32/6.3, was not found in control rats, indicating that the
5 protein was induced by Pb exposure. This finding was in agreement with studies that
6 indicated the formation of intranuclear inclusion bodies required protein synthesis
7 (McLachlin et al., 1980; Choie et al., 1975). In addition to its presence in kidneys of
8 Pb-exposed animals, p32/6.3 has been observed to be present and highly conserved in the
9 brains of rats, mice, dogs, chickens, and humans (Egle and Shelton. 1986). Exposure of
10 neuroblastoma cells to 50 or 100 (iM Pb glutamate for 1 or 3 days increased the
11 abundance of p32/6.3 (Klann and Shelton. 1989). Shelton et al. (1990) determined that
12 p32/6.3 was enriched in the basal ganglia, diencephalon, hippocampus, cerebellum,
13 brainstem, spinal cord, and cerebral cortex, and that it contained a high percentage of
14 glycine, aspartic, and glutamic acid residues. Selvin-Testa et al. (1991) and Harry et al.
15 (1996) reported that pre- and post-natal exposure of rats to 2,000-10,000 ppm Pb in
16 drinking water increased the levels of another brain protein, glial fibrillary acidic protein,
17 in developing astrocytes; and that this increase may be indicative of a demand for
18 astrocytes to sequester Pb.
5.2.3.2 Cytosolic Pb Binding Proteins
19 Numerous studies have also identified cytosolic Pb-binding proteins. Two binding
20 proteins, with molecular weights (MW) of 11.5 and 63 kDa, were identified by Oskarsson
21 et al. (1982) in the kidney postmitochondrial cytosolic fraction prepared from adult male
22 rats after i.p. injection with 50 mg Pb acetate/kg, followed by i.p. injection of 50 (iCi
23 203Pb acetate 6 days later. The two proteins were also found in the brain, but not the liver
24 or lung. Mistry et al. (1985) identified three proteins (MW = 11.5, 63, and >200 kDa) in
25 rat kidney cytosol, two of which, the 11.5 and 63 kDa proteins, were able to translocate
26 into the nucleus. The 11.5 kDa kidney protein was also able to reverse Pb binding to
27 ALAD through chelation of Pb and donation of a Zn cation to ALAD (Goering and
28 Fowler. 1985. 1984). Cd and Zn, but not Ca2+ or Fe, prevented the binding of Pb to the 63
29 and 11.5 kDa cytosolic proteins, which agrees with previous observations that Cd is able
30 to reduce total kidney Pb and prevent the formation of intranuclear inclusion bodies
31 (Mistry et al.. 1986; Mahaffev et al.. 1981; Mahaffev and Fowler. 1977). Additional
32 cytosolic Pb-binding proteins have been identified in the kidneys of Pb-exposed rats and
33 humans, including the cleavage product of a2-microglobulin, acyl-CoA binding protein
34 (MW = 9 kDa), and thymosin |34 (MW = 5 kDa) (Smith etal.. 1998: Fowler and DuVal.
35 1991).
November 2012 5-21 Draft - Do Not Cite or Quote
-------
1 Cytosolic Pb-binding proteins distinct from kidney proteins have also been identified in
2 the brain of exposed rats, and in human brain homogenates exposed to Pb in vitro
3 (Quintanilla-Vega et al.. 1995; DuVal and Fowler. 1989; Goering et al.. 1986). One
4 protein (MW =12 kDa) was shown to alleviate hepatic ALAD inhibition due to Pb
5 exposure through competitive binding with Pb and donation of Zn to ALAD. Cytosolic
6 Pb-binding proteins have been shown to be high in glutamic acid, aspartic acid, and
7 cysteine residues (Fowler et al.. 1993; DuVal and Fowler. 1989). Some evidence exists
8 that cytosolic Pb-binding proteins directly target Pb and compartmentalize intracellular
9 Pb as a protective measure against toxicity (Oian etal. 2005; Oian et al., 2000).
5.2.3.3 Erythrocytic Pb Binding Proteins
10 The majority (94%) of Pb in whole blood is found in erythrocytes (Ong and Lee. 1980a).
11 Originally, the major Pb-binding protein in erythrocytes was identified as hemoglobin
12 (Cohen et al.. 2000; Lolin and O'Gorman. 1988; Ong and Lee. 1980a. b; Raghavan and
13 Gonick. 1977). However, Bergdahl et al. (1997b) observed the principal Pb-binding
14 protein to be 240 kDa and identified it as ALAD. Two smaller Pb-binding proteins were
15 observed, but not identified (MW = 45 and <10 kDa). ALAD levels are inducible by Pb
16 exposure; the total concentration of the enzyme, but not the activity, is higher in both Pb-
17 exposed humans (blood Pb = 30-75 (ig/dL) and rats (Pb exposure = 2.5 x 10"4 (iM in
18 drinking water) (Boudene et al.. 1984; Fujitaetal.. 1982; Fujitaetal.. 1981).
19 ALAD is a polymorphic gene with three isoforms: ALAD 1-1, ALAD 1-2, or ALAD 2-2.
20 Carriers of the ALAD-2 allele have been shown to have higher blood Pb levels than
21 carriers of the homozygous ALAD-1 allele (Scinicariello et al.. 2007; Zhao et al.. 2007;
22 Kim et al., 2004; Perez-Bravo et al., 2004; Smith etal.. 1995a; Wetmur. 1994; Wetmur et
23 al.. 1991b; Astrin et al.. 1987). Some recent studies, however, either observed lower
24 blood Pb levels in carriers of the ALAD-2 allele or no difference in Pb levels among the
25 different allele carriers (Scinicariello et al.. 2010; Krieg et al.. 2009; Chen et al.. 2008c;
26 Chia et al.. 2007; Chia et al.. 2006; Wananukul et al.. 2006).
27 The ALAD-2 protein binds Pb more tightly than the ALAD-lform: in Pb-exposed
28 workers carrying the ALAD-2 gene, 84% of blood Pb was bound to ALAD versus 81%
29 in carriers of the ALAD-1 gene (p = 0.03) (Bergdahl et al.. 1997a). This higher affinity
30 for Pb in ALAD-2 carriers may sequester Pb and prevent its bioavailability for reaction
31 with other enzymes or cellular components. This is supported by the observation that
32 carriers of the ALAD-2 gene have higher levels of hemoglobin (Scinicariello et al..
33 2007). decreased plasma levulinic acid (Schwartz et al.. 1997b). decreased levels of Zn
34 protoporphyrin (Scinicariello et al.. 2007; Kim et al.. 2004). lower cortical bone Pb
November 2012 5-22 Draft - Do Not Cite or Quote
-------
1 (Smith et al.. 1995b). and lower amounts of DMSA-chelatable Pb (Scinicariello et al..
2 2007; Schwartz et al. 2000a: Schwartz et al.. 1997a). However, the findings, that
3 ALAD-2 polymorphisms reduced the bioavailability of Pb, are somewhat equivocal. Wu
4 et al. (2003a) observed that ALAD-2 carriers had lower blood Pb level (5.8 ± 4.2 (ig/dL)
5 than carriers of the ALAD-1 gene (blood Pb level = 6.2 ± 4.1 (ig/dL), and that ALAD-2
6 carriers demonstrated decreased renal function at lower patellar Pb concentrations than
7 those associated with decreased renal function in ALAD-1 carriers. This potentially
8 indicates that ALAD-2 carriers have enhanced Pb bioavailability. Weaver et al. (2003b)
9 observed that ALAD-2 polymorphisms were associated with higher DMSA-chelatable Pb
10 concentrations, when normalized to creatinine levels. Further, Montenegro et al. (2006)
11 observed that compared with individuals with the ALAD 1-1 genotype, individuals with
12 the ALAD 1-2/2-2 genotypes had a higher amount of Pb in the plasma (0.44 (ig/L versus
13 0.89 (ig/L, respectively) and in the percent plasma/blood ratio (0.48% versus 1.45%,
14 respectively). This potentially suggests that individuals with the ALAD 1-2/2-2 genotype
15 are at increased risk of Pb-induced health effects due to lower amounts of Pb
16 sequestration by erythrocyte ALAD, although this study did not specifically investigate
17 the clinical implications of ALAD polymorphisms.
18 ALAD has the estimated capacity to bind Pb at 85 (ig/dL in erythrocytes and 40 (ig/dL in
19 whole blood (Bergdahl et al.. 1998). The 45 kDa and <10 kDa Pb-binding proteins bound
20 approximately 12-26% and <1% of the blood Pb, respectively. At blood Pb
21 concentrations greater than 40 (ig/dL, greater binding to these components would likely
22 be observed. Bergdahl et al. (1998) tentatively identified the 45 kDa protein as
23 pyrimidine-5'-nucleotidase and the <10 kDa protein as acyl-CoA binding protein. Smith
24 et al. (1998) previously identified acyl-CoA binding protein as a Pb-binding protein
25 found in the kidney.
5.2.3.4 Metallothionein
26 In adults occupationally exposed to Pb, the presence of an inducible, low-molecular
27 weight (approximately 10 kDa) Pb-binding protein was identified in multiple early
28 studies (Gonick et al.. 1985; Raghavan et al.. 1981. 1980; Raghavan and Gonick. 1977).
29 The presence of this low molecular weight protein seemed to have a protective effect, as
30 workers who exhibited toxicity at low blood Pb concentrations were observed to have
31 lowered expression of this protein or low levels of Pb bound to it (Raghavan et al.. 1981.
32 1980). The presence of low molecular weight Pb-binding proteins in exposed workers
33 was corroborated by Lolin and O'Gorman (1988) and Church et al. (1993a. b). Further
34 Lolin and O'Gorman (1988) reported that the observed protein was only present when
35 blood Pb levels were greater than 39 (ig/dL, in agreement with the Pb-binding capacity of
November 2012 5-23 Draft - Do Not Cite or Quote
-------
1 ALAD, identified by Bergdahl et al. (1998). Xie et al. (1998) confirmed this, observing
2 the presence of a second low molecular weight protein with greater affinity than ALAD,
3 only at higher blood Pb levels. Church et al. Q993a, b) observed the presence of a 6-7
4 kDa protein in the blood of two Pb workers (blood Pb >160 (ig/dL); approximately 67%
5 of Pb was bound to the protein in the blood of the asymptomatic worker, whereas only
6 22% of the Pb was bound to it in the symptomatic (tremor, ataxia, extremity numbness)
7 worker. The reported protein was rich in cysteine residues and tentatively identified as
8 metallothionein.
9 Metallothionein is a low-MW metal-binding protein, most often binding Zn or Cu, that is
10 rich in cysteine residues and plays an important role in protecting against heavy metal
11 toxicity, maintaining trace element homeostasis, and scavenging free radicals (Yu et al..
12 2009). Exposure to Pb acetate induced the production of Pb- and Zn-metallothionein in
13 mice treated via i.p. or intravenous (i.v.) injection at 30 mg/kg (Maitani et al.. 1986). in
14 mice treated via i.p. injection at 300 (imol/kg (Yu et al.. 2009). or in rats treated via i.p.
15 injection at 24 (imol/lOOg (Ikebuchi et al.. 1986). The induced Pb-metallothionein
16 consisted of 28% half-cysteine and reacted with an antibody for Zn-metallothionein II
17 (Ikebuchi et al.. 1986). In contrast, exposure of rats to Pb via drinking water (200 or
18 300 ppm) failed to induce metallothionein in the kidneys or intestines (Wang et al..
19 2009b; Jamieson et al.. 2007). Goering and Fowler (1987a. b) observed that pretreatment
20 of rats with Zn before injection with Pb resulted in Pb and Zn co-eluting with Zn-
21 thionein, and that Zn-thionein I and II were able to bind Pb in vitro (Goering and Fowler.
22 1987a. b). Further, Goering and Fowler (1987a) found that kidney and liver Zn-thionein
23 decreased binding of Pb to liver ALAD and was able to donate Zn to ALAD, thus
24 attenuating the inhibition of ALAD due to Pb exposure. These findings are in agreement
25 with Goering et al. (1986) and DuVal and Fowler (1989) who demonstrated that rat brain
26 Pb-binding proteins attenuated Pb-induced inhibition of ALAD.
27 Metallothionein has been reported to be important in the amelioration of Pb-induced
28 toxicity effects. Liu et al. (1991) reported that Zn-metallothionein reduced Pb-induced
29 membrane leakage and loss of K+ in cultured hepatocytes incubated with 600-3,600 (iM
30 Pb. Metallothionein-null mice exposed to 1,000, 2,000, or 4,000 ppm Pb for 20 weeks
31 suffered renal toxicity described as nephromegaly and decreased renal function compared
32 to Pb-treated wild-type mice (Qu et al.. 2002). Interestingly, metallothionein-null mice
33 were unable to form intranuclear inclusion bodies and accumulated less renal Pb than did
34 the wild-type mice (Qu et al.. 2002). Increased metallothionein levels were induced by Pb
35 exposure in non-null mice. Exposure to Pb (1,000, 2,000, or 4,000 ppm), both for
36 104 weeks as adults and from GD8 to early adulthood, resulted in increased preneoplastic
37 lesions and carcinogenicity in the testes, bladder, and kidneys of metallothionein-null rats
38 compared to wild type mice (Tokaretal.. 2010; Waalkes et al.. 2004). Inclusion bodies
November 2012 5-24 Draft - Do Not Cite or Quote
-------
1 were not observed in null mice. The authors concluded that metallothionein is important
2 in the formation of inclusion bodies and in the mitigation of Pb-induced toxic effects, and
3 that those with polymorphisms in metallothionein coding genes that are associated with
4 reduced inducibility may be at greater susceptibility to Pb. In support of this theory, Chen
5 et al. (2010a) observed that Pb-exposed workers with a mutant metallothionein allele had
6 higher blood Pb levels than did carriers of the normal allele (24.17 and 21.27 versus
7 17.03 (ig/dL), and had larger blood Pb-associated changes in systolic BP and serum renal
8 function parameters.
9 In summary, a number of proteins have been identified that can bind and sequester Pb
10 including ALAD and metallothionein. Additionally, evidence suggests that certain
11 polymorphisms that alter the binding capability or inducibility of these proteins can
12 increase the risk of Pb induced health effects.
5.2.4 Oxidative Stress
13 Oxidative stress occurs when free radicals or reactive oxygen species (ROS) exceed the
14 capacity of antioxidant defense mechanisms. This Oxidative imbalance results in
15 uncontained ROS, such as superoxide (O2~), hydroxyl radical (OH), and hydrogen
16 peroxide (H2O2), which can attack and denature functional/structural molecules and,
17 thereby, promote tissue damage, cytotoxicity, and dysfunction. Pb exposure has been
18 shown to cause Oxidative damage to the heart, liver, kidney, reproductive organs, brain,
19 and erythrocytes, which may be responsible for a number of Pb-induced health effects
20 (Salawu et al.. 2009; Shan et al.. 2009; Vaziri. 2008b: Gonicketal.. 1997; Sandhir and
21 Gill. 1995: Khalil-Manesh et al.. 1994: Khalil-Manesh et al.. 1992aV The origin of ROS
22 (produced after Pb exposure) is likely a multipathway process, resulting from oxidation
23 of 5-aminolevulinic acid (ALA), membrane and lipid oxidation, NAD(P)H oxidase
24 activation, and antioxidant enzyme depletion, as discussed below. Some of these
25 processes result from the disruption of functional metal ions within oxidative stress
26 enzymes, such as superoxide dismutase (SOD), catalase (CAT), and glutathione
27 peroxidase (GPx). Interestingly, Pb exposure in many species of plants, invertebrates, and
28 vertebrates discussed in ChapterJ? (Ecological Effects of Lead) results in upregulation of
29 antioxidant enzymes and increased lipid peroxidation. Oxidative stress is a common
30 mode of action for a number of other metals (e.g., Cd, Mn, As, Co, Cr) that are often
31 found with Pb and by which possible interactions with Pb have been suggested to occur
32 (Jomova and Valko. 2011; Jomovaet al.. 2011; Matovic et al.. 2011; HaMai and Bondy.
33 2004). Not all of these co-occurring metals directly produce ROS or redox cycle, but
34 instead may suppress the free radical scavenging ability of the organism thus leading to
35 oxidative stress.
November 2012 5-25 Draft - Do Not Cite or Quote
-------
5.2.4.1 5-ALA Oxidation
1 The majority of Pb present in the blood accumulates in erythrocytes where it enters
2 through passive carrier-mediated mechanisms including a vanadate-sensitive Ca2+ pump.
3 Once Pb enters erythrocytes, it is predominantly found in the protein-bound form, with
4 hemoglobin and 5-ALAD both identified as targets (Bergdahl et al., 1997a). Through its
5 sulfhydryl and metal ion disrupting properties, Pb incorporates with and inhibits a
6 number of enzymes in the heme biosynthetic process, including 5-ALA synthetase,
7 5-ALAD, and ferrochelatase. Pb has been shown to be able to disrupt the Zn ions
8 requisite for the activity of 5-ALAD, the rate limiting step in heme synthesis, leading to
9 enzyme inhibition at 10"6 (iM concentrations (Simons. 1995). Additionally, blood Pb
10 levels (mean: 7 (ig/dL) have been associated with inhibited activity of 5-ALAD in
11 humans, and the lowest blood Pb level observed to be associated with lower 5-ALAD
12 activity in these studies was 5 (ig/dL (Ahamed et al., 2005; Sakai and Morita. 1996). A
13 negative correlation (r = -0.6) was found between blood Pb levels in adolescents (range
14 of blood Pb levels: 4 to 20 (ig/dL) and blood 5-ALAD activity (Ahamed et al.. 2006).
15 This inhibition of 5-ALAD results in the accumulation of 5-ALA in blood and urine,
16 where 5-ALA undergoes tautomerization and autoxidation. Oxidized 5-ALA generates
17 ROS through reduction of ferricytochrome c and electron transfer from oxyHb, metHb,
18 and other ferric and ferrous iron complexes (Hermes-Lima et al.. 1991; Monteiro et al..
19 1991). The autoxidation of 5-ALA produces O2~, OH, H2O2, and an ALA radical
20 (Monteiro et al.. 1989; Monteiro et al.. 1986).
5.2.4.2 Membrane and Lipid Peroxidation
21 A large number of studies in humans and experimental animals have indicated that
22 exposure to Pb can lead to membrane and lipid peroxidation. It is possible that ROS
23 produced from 5-ALA oxidation, as described above, interacts with and disrupts
24 membrane lipids (Oteizaet al., 1995; Bechara et al.. 1993). Additionally, Pb has the
25 capacity to stimulate ferrous ion initiated membrane lipid peroxidation serving as a
26 catalyst for these events (Adonavlo and Oteiza. 1999; Quinlan et al.. 1988). The extent of
27 peroxidation of lipids varies based on the number of double bonds present in unsaturated
28 fatty acids, since double bonds weaken the C-H bonds on the adjacent carbon, making
29 hydrogen (H) removal easier (Yiin and Lin. 1995). After the essential unsaturated fatty
30 acid solutions were incubated with Pb (4-12 (ig/dL, 24 hours), the production of
31 malondialdehyde (MDA), a marker of oxidative stress and lipid oxidation end product,
32 increased relative to the number of double bonds of the fatty acid (Yiin and Lin. 1995). In
33 the absence of Fe2+, Pb has not been shown to promote lipid peroxidation; however, it
34 may accelerate peroxidation by H2O2 (Quinlan et al., 1988). This could be due to altering
November 2012 5-26 Draft - Do Not Cite or Quote
-------
1 membrane structure, restricting phospholipid movement, and facilitating the propagation
2 of peroxidation.
3 Pb induces changes in the fatty acid composition of a membrane, which could lead to
4 oxidative damage. Exposure to Pb (>62.5 ppm in drinking water, 3 weeks) in chicks
5 promoted an increase in arachidonic acid (20:4) as a percentage of total fatty acids, and
6 decreased the relative proportion of shorter chain fatty acids (linoleic acid, 18:2) (Lawton
7 and Donaldson. 1991). It is possible that Pb depressed the desaturation of saturated fatty
8 acids to the corresponding monoenoic fatty acids, while stimulating elongation and
9 desaturation of linoleic acid to arachidonic acid. Since fatty acid chain length and
10 unsaturation are related to the oxidative potential, changes in fatty acid membrane
11 composition may result in enhanced lipid peroxidation. In addition, changes in fatty
12 acids, thus membrane composition, can result in altered membrane fluidity (Donaldson
13 and Knowles. 1993). Changes in membrane fluidity will disturb the conformation of the
14 active sites of membrane associated enzymes, disrupt metabolic regulation, and alter
15 membrane permeability and function.
16 A number of recent studies report increased measures of lipid peroxidation in various
17 organs, tissues, and species in association with Pb. Occupational Pb exposure resulting in
18 elevated blood Pb levels (means >8 (ig/dL) reported in various countries provides
19 evidence of lipid peroxidation, including increased plasma MDA levels (Ergurhan-Ilhan
20 et al.. 2008; Khan et al.. 2008; Mohammad et al.. 2008; Quintanar-Escorza et al.. 2007;
21 Patil et al.. 2006a: Patil et al.. 2006b). One study found a correlation between the MDA
22 levels and blood Pb levels even in the unexposed workers, although they had blood Pb
23 levels higher than the mean blood Pb level of the current U.S. population
24 (i.e., <12 (ig/dL) (Quintanar-Escorza et al., 2007). Other studies found evidence of
25 increased lipid peroxidation among the general population, including children, with
26 elevated blood Pb levels (means >10 (ig/dL) (Ahamed et al.. 2008; Ahamed et al.. 2006;
27 Jin et al.. 2006). In adolescents, Ahamed et al. (2006) found a blood MDA levels to be
28 positively correlated (r = 0.7) with concurrent blood Pb levels ranging between 4 and
29 20 (ig/dL. Similar results have been shown after Pb exposure in animal studies (Abdel
30 Moneimetal. 20lib; Pandvaet al.. 2010; Dogru et al.. 2008; Yu et al.. 2008; Adegbesan
31 and Adenuga. 2007; Lee et al.. 2005). Enhanced lipid peroxidation has been found in Pb
32 treated (50 (ig, 1-4 hours) rat brain homogenates (Rehman et al.. 1995). rat proximal
33 tubule cells (0.5-1 (JVI, 12 hours) (Wang etal. 20lib), and in specific brain regions,
34 hippocampus and cerebellum, after Pb exposure (500 ppm, 8 weeks) to rats (Bennet et al..
35 2007). Overall, there was a correlation between the blood Pb level and measures of lipid
36 peroxidation often measured by MDA levels.
November 2012 5-27 Draft - Do Not Cite or Quote
-------
1 In summary, studies in humans and animals provide evidence for increased lipid and
2 membrane oxidation following Pb exposure. Interestingly, many species of plants,
3 invertebrates, and other vertebrates also exhibit increased lipid peroxidation with Pb
4 exposure (Sections 7.3.12.6 and 7.4.12.6). The increase in lipid peroxidation following
5 Pb exposure observed across species and kingdoms demonstrate an evolutionarily
6 conserved oxidative response following Pb exposure.
5.2.4.3 NAD(P)H Oxidase Activation
7 NAD(P)H oxidase is a membrane bound enzyme that requires Ca2+ in order to catalyze
8 the production of O2 from NAD(P)H and molecular oxygen (Leseney et al., 1999). Two
9 studies provide evidence for increased activation of NAD(P)H oxidase that may
10 contribute to the production of ROS after Pb exposure (Ni et al., 2004; Vaziri et al.,
11 2003). Vaziri et al. (2003) found increased protein expression of the NAD(P)H subunit
12 gp91phox in the brain, heart, and renal cortex of Pb-treated rats (100 ppm in drinking
13 water, 12 weeks). This upregulation was present in Pb-treated (1-10 ppm) human
14 coronary artery endothelial cells, but not vascular smooth muscle cells (VSMC), which
15 do not express the protein (Ni et al., 2004). It is possible that NAD(P)H oxidase serves as
16 a potential source of ROS in cells that express this protein.
5.2.4.4 Antioxidant Enzyme Disruption
17 Oxidative stress can result not only from the increased production of ROS, but also from
18 the decreased activity of antioxidant defense enzymes. Pb has been shown to alter the
19 function of several antioxidant enzymes, including SOD, CAT, glucose-6-phosphate
20 dehydrogenase (G6PD), and the enzymes involved in glutathione metabolism, GPx,
21 glutathione-S-transferase (GST), and glutathione reductase (GR). These changes in the
22 antioxidant defense system could be due to the high affinity of Pb for sulfhydryl groups
23 contained within proteins and its metal ion mimicry. However changes could also be a
24 consequence of increased oxidative damage by Pb.
25 Studies of the effects of Pb exposure on the activities of SOD and CAT give divergent
26 results. These metalloprotein enzymes require various essential trace elements for proper
27 structure and function, making them a target for Pb toxicity. CAT is a heme containing
28 protein that requires Fe ions to function (Putnam et al.. 2000). SOD exists in multiple
29 isoforms that require Cu and Zn (SOD 1 and SOD3) (Antonyuk et al.. 2009) or Mn
30 (SOD2) (Borgstahl et al.. 1992). A number of studies have found decreased activity of
31 these enzymes (Pandya et al., 2010; Ergurhan-Ilhan et al., 2008; Mohammad et al., 2008;
November 2012 5-28 Draft - Do Not Cite or Quote
-------
1 Yu et al.. 2008; Patil et al.. 2006a: Patil et al.. 2006b: Conterato et al.. In Press), whereas
2 others have observed increased activity following Pb exposure (Ahamed et al.. 2008; Lee
3 et al.. 2005). The heterogeneity in species examined, (i.e., humans, rodents, boars), and
4 Pb exposure metrics reported did not permit evaluation of whether a nonlinear
5 concentration-response relationship could explain heterogeneity in findings. Pb exposure
6 (500 ppm Pb acetate, 1, 4, and 8 weeks) in adult male rats showed that SOD and CAT
7 activity responded differently depending on the brain region analyzed and length of
8 exposure (Bennet et al.. 2007). Another study found that the brain had consistently
9 decreased SOD activity, irrespective of dose in prenatally-exposed animals (0.3 and
10 3.0 ppm, blood Pb level 20.4 and 24.5 (ig/dL); however hepatic SOD activity increased at
11 low level Pb administration and decreased after high level exposure (Uzbekov et al..
12 2007). It is possible that the increased activity of the SOD and CAT proteins is due to
13 activation by ROS, while decreased enzyme activity is the result of metal ion substitution
14 by Pb, causing enzyme inactivation.
15 Glutathione is a tripeptide antioxidant containing a cysteine with a reactive thiol group
16 that can act nonenzymatically as a direct antioxidant or as a cofactor in enzymatic
17 detoxification reactions by GST. Glutathione will donate an electron while in its reduced
18 state (GSH), which leads to conversion to the oxidized form, glutathione disulfide
19 (GSSG). Pb binds to the thiol and can both interfere with the antioxidant capacity of
20 GSH, and can decrease levels of GSH. Short-term administration of Pb in vitro (0.1 (iM)
21 and observed biomarkers of Pb exposure in humans (18 (ig/dL mean blood Pb level) have
22 been associated with decreased blood and organ GSH and cysteine content, which may be
23 due to increased GSH efflux from tissues (Pandya et al.. 2010; Pillai et al.. 2010; Ahamed
24 et al.. 2009; Ahamed et al.. 2008; Flora et al.. 2007; Ahamed etal. 2005; Chettv et al..
25 2005; Nakagawa. 1991. 1989). Long-term Pb exposure has elicited a compensatory
26 upregulation in the biosynthesis of GSH in the attempt to overcome Pb toxicity, thus
27 often manifesting as an increase in Pb-induced GSH in animals (Daggett et al.. 1998;
28 Corongiu and Milia. 1982; Hsu. 1981) and occupationally exposed adults (SRC. 2002;
29 Conterato et al.. In Press). However, other studies have found that long-term Pb exposure,
30 resulting in mean blood Pb levels between 6.6 and 22 (ig/dL, causes the depletion of GSH
31 in animals (Lee et al.. 2005; Ercal etal.. 1996) and occupationally exposed adults
32 (Mohammad et al.. 2008). Thus, the duration of Pb exposure is important to consider
33 when measuring GSH levels.
34 Glutathione reductase is able to reduce GSSG back to GSH. Therefore, an increased
35 GSSG/GSH ratio is considered to be indicative of oxidative stress. Epidemiologic studies
36 have found higher blood Pb levels to be associated with increases in the GSSG/GSH ratio
37 (Mohammad et al.. 2008; Ercal etal.. 1996; Sandhir and Gill. 1995). In one study, this
38 association was observed in a population of children with a mean blood Pb level below
November 2012 5-29 Draft - Do Not Cite or Quote
-------
1 10 (ig/dL (Diouf etal.. 2006). Studies have found mixed effects on GR activation. GR
2 possesses a disulfide at its active site that is a target for inhibition by Pb. Studies in
3 animals and cells have reported decreased (Bokara et al.. 2009; Sandhir and Gill, 1995;
4 Sandhiretal.. 1994). increased (Sobekova et al. 2009; Howard. 1974). and no change
5 (Hsu. 1981) in GR activity after Pb exposure. This could be because the effect of Pb on
6 GR varies depending on sex (Sobekova et al.. 2009). and organ or organ region (Bokara
7 et al.. 2009). The heterogeneity in species examined, (i.e., humans, rodents), and Pb
8 exposure duration and metrics reported did not permit evaluation of whether a nonlinear
9 concentration-response relationship could explain heterogeneity in findings.
10 GSH is used as a cofactor for peroxide reduction and detoxification of xenobiotics by the
11 enzymes GPx and GST. GPx requires Se (selenium) for peroxide decomposition (Rotruck
12 et al.. 1973). whereas GST functions via a sulfhydryl group. Evidence indicates that by
13 reducing the uptake of Se, depleting cellular GSH, and disrupting protein thiols, Pb
14 decreases the activity of GPx and GST (Pillai etal.. 2010; Yu et al.. 2008; Lee et al..
15 2005; Nakagawa. 1991; Schrauzer. 1987). Similar to other antioxidant enzymes,
16 compensatory upregulation of these enzymes was observed after Pb exposure in animals
17 and in Pb-exposed workers (painters with a mean blood Pb level of 5.4 (ig/dL) (Bokara et
18 al.. 2009; Ergurhan-Ilhan et al.. 2008; Conterato et al.. 2007; Daggett et al.. 1998;
19 Conterato et al.. In Press). However, in another study, these enzymes were not able to
20 compensate for the increased Pb-induced ROS, further contributing to the oxidative
21 environment (Farmand et al.. 2005).
22 Recently, y-glutamyltransferase (GGT) within its normal range has been regarded as an
23 early and sensitive marker of oxidative stress. This may be because cellular GGT
24 metabolizes extracellular GSH to be used in intracellular GSH synthesis. Thus, cellular
25 GGT acts as an antioxidant enzyme by increasing the intracellular GSH pool. However,
26 the reasons for the association between GGT and oxidative stress have not been fully
27 realized (Lee et al.. 2004). In one study, occupational Pb exposure (mean blood Pb level
28 of 29.1 (ig/dL) was associated with increased serum GGT levels (Khan et al.. 2008).
29 Interestingly, higher blood Pb level was similarly associated with higher serum GGT
30 levels in a sample of the U.S. adult population (NHANES III) (Lee et al.. 2006a). In this
31 study of nonoccupationally-exposed individuals, a concentration-dependent relationship
32 was observed with blood Pb levels <7 (ig/dL.
33 In summary, Pb has been shown to alter the function of several antioxidant enzymes,
34 including SOD, CAT, G6PD, and the enzymes involved in glutathione metabolism, GPx,
35 GST, and GR in human populations and experimental animal models. Alteration of these
36 enzymes may lead to further oxidative stress following Pb exposure.
November 2012 5-30 Draft - Do Not Cite or Quote
-------
5.2.4.5 Nitric Oxide Signaling
1 NO (nitric oxide radical), also known as endothelium-derived relaxing factor, is a potent
2 endogenous signaling molecule involved in vasodilation. Short- and long-term Pb
3 exposure in animals have been found to decrease the biologically active NO, not through
4 reduction in NO-production capacity (Vaziri and Ding. 2001; Vaziri etal.. 1999a). but as
5 a result of inactivation and sequestration of NO by ROS (Malvezzi et al.. 2001; Vaziri et
6 al.. 1999b). Endogenous NO can interact with ROS, specifically O2~, produced following
7 exposure to Pb to form the highly cytotoxic reactive nitrogen species, peroxynitrite
8 (ONOO"). This reactive compound can damage cellular DNA and proteins, resulting in
9 the formation of nitrotyrosine among other products. Overabundance of nitrotyrosine in
10 plasma and tissues is present after exposure to Pb (Vaziri et al.. 1999b). NO is also
11 produced by macrophages in the defense against certain infectious agents, including
12 bacteria. Studies have indicated that Pb exposure can significantly reduce production of
13 NO in human (Pineda-Zavaleta et al.. 2004) and animal immune cells (Lee et al.. 200Ib;
14 Tian and Lawrence. 1995). possibly leading to reduced host resistance (Tian and
15 Lawrence. 1996).
16 Production of NO is catalyzed by a family of enzymes called nitric oxide synthases
17 (NOS), including endothelial NOS (eNOS), neuronal NOS (nNOS), and inducible NOS
18 (iNOS), which require a heme prosthetic group and a Zn cation for enzymatic activity
19 (Messerschmidt et al.. 2001). Paradoxically, the reduction in NO availability in vascular
20 tissue following Pb exposure is accompanied by statistically significant upregulation in
21 NOS isotypes (Vaziri and Ding. 2001; Vaziri etal.. 1999a; Gonicketal. 1997). A direct
22 inhibitory action of Pb on NOS enzymatic activity has been rejected (Vaziri et al..
23 1999a). Instead, the upregulation of NOS occurs as compensation for the decreased NO
24 resulting from ROS inactivation (Vaziri et al.. 2005; Vaziri and Ding. 2001; Vaziri and
25 Wang. 1999).
Soluble Guanylate Synthase
26 Many biological actions of NO, such as vasorelaxation, are mediated by cyclic guanosine
27 monophosphate (cGMP), which is produced by soluble guanylate cyclase (sGC) from the
28 substrate guanosine triphosphate. Soluble guanylate cyclase is a heterodimer requiring
29 one molecule of heme for enzymatic activity (Boerrigter and Burnett. 2009). In VSMC,
30 sGC serves as the NO receptor. Marked reduction in plasma concentrations and urinary
31 excretion of cGMP is observed after Pb exposure to rats [5 ppm Pb in drinking water for
32 30 days (Marques et al.. 2001)] and [100 ppm Pb acetate in drinking water for 3 months,
33 resulting in a mean blood Pb level of 29.4 (ig/dL) (Khalil et al.. 2008)1 (Marques et al..
34 2001; Khalil-Manesh et al.. 1993b). In addition, Pb exposure downregulated the protein
November 2012 5-31 Draft - Do Not Cite or Quote
-------
1 abundance of sGC in vascular tissue (Farmand et al., 2005; Courtois etal. 2003;
2 Marques et al.. 2001). This downregulation in sGC was prevented by antioxidant therapy
3 (ascorbic acid) suggesting that oxidative stress also plays a role in Pb-induced
4 downregulation of sGC (no change in blood Pb level was observed after ascorbic acid
5 treatment) (Marques et al., 2001).
5.2.5 Inflammation
6 Misregulated inflammation represents one of the major hallmarks of Pb-induced immune
7 effects. It is important to note that this can manifest in any tissue where immune cell
8 mobilization and tissue insult occurs. Enhanced inflammation and tissue damage occurs
9 through the modulation of inflammatory cell function and production of pro-
10 inflammatory cytokines and metabolites. Overproduction of ROS and an apparent
11 depletion of antioxidant protective enzymes and factors (e.g., Se) accompany this
12 immunomodulation (Chetty et al., 2005).
13 Traditional immune-mediated inflammation can be seen with bronchial
14 hyperresponsiveness, asthma, and respiratory infections, some of which have been
15 associated with exposure to Pb. But it is important to recognize that any tissue or organ
16 can be affected by immune-mediated inflammatory dysfunction given the distribution of
17 immune cells as both permanent residents and infiltrating cell populations (Mudipalli.
18 2007; Carmignani et al., 2000). Pb has been associated with multiple indicators of
19 inflammation in multiple cell types. Pb has also induced renal tubulointerstitial
20 inflammation (100 ppm exposure for 14 weeks) (Rodriguez-Iturbe et al., 2005)
21 (24.6 (ig/dL blood Pb level, 150 ppm for 16 weeks) (Roncal et al.. 2007). Renal
22 tubulointerstitial inflammation has been coupled with activation of the redox sensitive
23 nuclear transcription factor kappa B (NFKB) and lymphocyte and macrophage infiltration
24 in rats (100 ppm for 14 weeks resulting in mean blood Pb levels ranging 23.7 (ig/dL)
25 (Bravo et al.. 2007). These events could be in response to the oxidative environment
26 arising from Pb exposure, since Pb-induced inflammation and NFxB activation can be
27 ameliorated by antioxidant therapy (Rodriguez-Iturbe et al.. 2004). Pb spheres implanted
28 in the brains of rats produced neutrophil-driven inflammation with apoptosis and
29 indications of neurodegeneration (Nakao et al.. 2010).
30 Inflammation can be mediated by the production of chemical messengers such as
31 prostaglandins (PG). Pb exposure has been associated with increased arachidonic acid
32 (AA) metabolism, thus elevating the production of PGE2, PGF2, and thromboxane in
33 occupationally-exposed humans (mean blood Pb level 48 (ig/dL) (Cardenas et al., 1993)
34 and animal and cell models (e.g., 0.01 (JVI, 48 hours) (Chetty et al.. 2005; Flohe et al..
November 2012 5-32 Draft - Do Not Cite or Quote
-------
1 2002; Knowles and Donaldson. 1997; Lee and Battles. 1994). Dietary Pb exposure of
2 animals (500 ppm, 19 days) can increase the percentage of cell membrane AA, the
3 precursor of cyclooxygenase and lipoxygenase metabolism to PGs and leukotrienes
4 (Knowles and Donaldson. 1990). Additionally, Pb (1 (iM) may promote the release of
5 AA via activation of phospholipase A2, as shown in isolated VSMC (Dorman and
6 Freeman. 2002).
7 Inflammation may be the result of increased pro-inflammatory signaling or may stimulate
8 these signaling pathways. Pb can elevate the expression of the pro-inflammatory
9 transcription factors NFxB and activator protein-1 (AP-1), as well as the AP-1
10 component c-Jun (Korashy and El-Kadi. 2008; Korashy and Ei-Kadi. 2008; Bravo et al..
11 2007; RameshetaL 1999; Pvatt et al.. 1996). Pb exposure (25 (iM) to dendritic cells
12 stimulated phosphorylation of the Erk/MAPK pathway, but not p38, STATS or 5, or
13 CREB (Gao et al.. 2007)
5.2.5.1 Cytokine Production
14 There are three modes by which Pb has been shown to affect immune cytokine
15 production. First, Pb can act on macrophages to elevate the production of pro-
16 inflammatory cytokines such as TNF-a and interleukin (IL)-6 (Cheng et al.. 2006; Chen
17 et al.. 1999; Dentener et al.. 1989). This can result in local tissue damage during the
18 course of immune responses affecting such targets as the liver. Second, Pb can skew the
19 ratio of IL-12/IL-10 such that T-derived lymphocyte helper (Th)l responses are
20 suppressed and Th2 responses are promoted (Chen et al., 2004; Miller et al.. 1998)
21 possibly by affecting dendritic cells. Third, when acquired immune responses occur
22 following exposure to Pb, Thl lymphocyte production of cytokines is suppressed
23 (e-g-, IFN-y) (Lynes et al.. 2006; Heo et al.. 1996); in contrast, Th2 cytokines such as
24 IL-4, IL-5, and IL-6 are elevated (Gao et al.. 2007; Kim and Lawrence. 2000). The
25 combination of these three modes of cytokine changes induced by Pb can create a
26 hyperinflammatory state among innate immune cells and skew acquired immunity toward
27 Th2 responses.
28 lavicoli et al. (2006b) reported that low blood Pb concentrations produced significant
29 changes in cytokine levels in mice. At a low dietary Pb concentration (0.11 ppm, blood
30 Pb level of 1.6 (ig/dL), IL-2 and IFN-y were decreased compared to the controls
31 (0.02 ppm, 0.8 (ig/dL), indicating a suppressed Thl response. As the dietary and blood Pb
32 concentrations increased (resulting in blood Pb levels 12-61 (ig/dL), a Th2 phenotype
33 was observed with suppressed IFN-y and IL-2 and elevated IL-4 production. These
34 findings support the notion that the immune system is remarkably sensitive to Pb-induced
November 2012 5-33 Draft - Do Not Cite or Quote
-------
1 functional alterations and that nonlinear effects may occur at low Pb exposures. TGF-(3
2 production was also altered by Pb exposure to transfected mouse limb bud mesenchymal
3 stem cells (1 (JVI, 3 days) (Zuscik et al., 2007). IL-2 is one of the more variable cytokines
4 with respect to Pb-induced changes. Depending upon the protocol it can be slightly
5 elevated in production or unchanged. Recently, Gao et al. (2007) found that Pb-treated
6 dendritic cells (25 (iM) promoted a slight but statistically significant increase in IL-2
7 production among lymphocytes. Proinflammatory cytokines have been measured in other
8 organs and cell types after Pb exposure. Elevation of IL-1(3 and TNF-a were observed in
9 the hippocampus after Pb treatment (15 ppm, i.p., daily for 2 weeks, blood Pb level of
10 30.8 (ig/dL) and increased IL-6 was found in the forebrain (Struzvnska et al., 2007).
11 Consistent with animal studies, epidemiologic studies also found higher concurrent blood
12 Pb levels in children and occupationally-exposed adults to be associated with a shift
13 toward production of Th2 cytokines relative to Thl cytokines. The evidence in children
14 was based on comparisons of serum cytokine levels among groups with different blood
15 Pb levels without consideration of potential confounding factors. Among children ages
16 9 months to 6 years in Missouri, Lutz et al. (1999) found that children with concurrent
17 blood Pb levels 15-19 (ig/dL had higher serum levels of IL-4 and IgE (Section 5.6.3) than
18 did children with lower blood Pb levels. These results were consistent with the mode of
19 action for IL-4 to activate B cells to induce B cell class switching to IgE. Concurrent
20 blood Pb levels did not differ by residence in old versus new homes or by urban versus
21 rural residence (means: 3.2-3.8 (ig/dL) but were higher among children living near an oil
22 refinery, in particular, among children with known respiratory allergies (mean:
23 8.8 (ig/dL). This latter group of children also had the lowest serum levels of IFN-y and
24 highest levels of IL-4. There was no direct comparison of cytokine levels between blood
25 Pb level groups in the population overall; however, cytokine levels were similar between
26 healthy and allergy groups in the other Pb source groups that had similar blood Pb levels.
27 Thus, the differences in cytokine levels between healthy and allergic children living near
28 the oil refinery may have been influenced by differences in their blood Pb levels or other
29 factors related to residence near an oil refinery.
30 Evidence of association between blood Pb levels and cytokine levels in
31 nonoccupationally-exposed adults was unclear. Among healthy adult university students
32 in Incheon, Korea, Kim et al. (2007) found associations of concurrent blood Pb level with
33 serum levels of TNF-a and IL-6 that were larger among male students with blood Pb
34 levels 2.51-10.47 (ig/dL. Notably, the relative contributions of lower recent versus higher
35 past Pb exposures to these cytokine effects is not known. In models that adjusted for age,
36 sex, BMI, and smoking status, a 1 (ig/dL increase in blood Pb level was associated with a
37 23% increase (95% CI: 4, 55%) in log of TNF-a and a 26% increase in log of IL-6 (95%
38 CI: 0, 55%). The association between levels of blood Pb and plasma TNF-a was greater
November 2012 5-34 Draft - Do Not Cite or Quote
-------
1 among men who were GSTM1 null or had the TNF-a GG genotype. For the association
2 between blood Pb level and plasma IL-6, the effect estimate was slightly elevated in
3 TNF-a GG genotype but not elevated in the GSTM1 positive group. The effects of Pb on
4 several physiological systems have been hypothesized to be mediated by the generation
5 of ROS (Daggett et al.. 1998). Thus, the null variant of GSTM1, which is associated with
6 reduced elimination of ROS, may increase the risk of Pb-associated immune effects. The
7 results for the TNF-a polymorphism are difficult to interpret. The GG genotype is
8 associated with lower expression of TNF-a, and the literature is mixed with respect to
9 which variant increases risk of inflammation-related conditions. A study of adults in Italy
10 did not provide quantitative results and only reported a lack of statistically significant
11 correlation between blood Pb levels with Th2 or Thl cytokine levels in men (Boscolo et
12 al., 1999) and women (Boscolo et al., 2000).
13 Results from studies of occupationally-exposed adults also suggested that Pb exposure
14 may be associated with decreases in Thl cytokines and increases in Th2 cytokines;
15 however, analyses were mostly limited to comparisons of levels among different
16 occupational groups with different mean blood Pb levels (Pi Lorenzo et al.. 2007;
17 Valentino et al., 2007; Yiicesoy et al., 1997a) without consideration for potential
18 confounding factors including other occupational exposures. An exception was a study of
19 male foundry workers, pottery workers, and unexposed workers by Valentino et al.
20 (2007). Although quantitative regression results were not provided, higher blood Pb level
21 was associated with higher IL-10 and TNF-a with adjustment for age, BMI, smoking, and
22 alcohol consumption. In analyses of blood Pb groups, levels of IL-2, IL-10, and IL-6 also
23 increased from the lowest to highest blood Pb group. In contrast with most other studies,
24 both exposed worker groups had lower IL-4 levels compared with controls. In a similar
25 analysis, DiLorenzo et al. (2007) separated exposed workers into intermediate
26 (9.1-29.4 (ig/dL) and high (29.4-81.1 (ig/dL) blood Pb level groups, with unexposed
27 workers comprising the low exposure group (blood Pb levels 1-11 (ig/dL). Mean TNF-a
28 levels showed a monotonic increase from the low to high blood Pb group. Levels of
29 granulocyte colony-stimulating factor (G-CSF) did not differ between the intermediate
30 and high blood Pb groups among the Pb recyclers; however, G-CSF levels were higher in
31 the Pb recyclers than in the unexposed controls. Furthermore, among all subjects, blood
32 Pb showed a strong, positive correlation with G-CSF. Yucesoy et al. (1997a) found lower
33 serum levels of the Thl cytokines, IL-lp and IFN-y, in workers (mean blood Pb level of
34 59.4 (ig/dL) compared with controls (mean blood Pb level of 4.8 (ig/dL); however levels
35 of the Th2 cytokines, IL-2 and TNF-a levels, were similar between groups. As most
36 occupationally-exposed cohorts represent populations highly exposed to Pb (with mean
37 blood Pb levels >22 (ig/dL), effects observed within these cohorts may not be
38 generalizable to the population as a whole.
November 2012 5-35 Draft - Do Not Cite or Quote
-------
1 In summary, animal, general population, and occupational studies suggest that exposure
2 to Pb increases the production of pro-inflammatory cytokines, skews the ratio of Thl and
3 Th2 cytokines to favor Th2 responses, and suppresses lymphocyte cytokine production.
5.2.6 Endocrine Disruption
5.2.6.1 Hypothalamic-Pituitary-Gonadal Axis
4 Evidence indicates that Pb is a potent endocrine disrupting chemical found to be
5 associated with reproductive and developmental effects in both male and female animal
6 models (see Section 5.8). Pb may act both at multiple points along the hypothalamic-
7 pituitary-gonadal (HPG) axis and directly at gonadal sites. The HPG axis functions in a
8 closely regulated manner to produce circulating sex steroids and growth factors required
9 for normal growth and development. Long-term Pb exposure in animals has been shown
10 to alter serum levels of follicle-stimulating hormone (FSH), luteinizing hormone (LH),
11 testosterone, and estradiol (Biswas and Ghosh. 2006; Rubio et al. 2006). Similar changes
12 in serum HPG hormones have been observed after high-level Pb exposure in animals,
13 resulting in blood Pb levels >20 (ig/dL (Dearth et al.. 2002; Ronisetal.. 1998b: Foster.
14 1992; Sokol and Berman. 1991). Increases in serum LH and FSH have been associated
15 with increasing concurrent blood Pb levels in adult women from the NHANES cohort
16 (Krieg. 2007). The change in HPG hormones likely occurs through the inhibition of LH
17 secretion and the reduction in the expression of the steroidogenic acute regulatory protein
18 (StAR) (Huang and Liu. 2004; Srivastavaet al.. 2004; Huang et al.. 2002; Ronis et al..
19 1996). StAR expression is the rate-limiting step essential in maintaining gonadotropin-
20 stimulated steroidogenesis, which results in the formation of testosterone and estradiol.
21 Prenatal and lactational Pb exposure (resulting in 3 (ig/dL blood Pb in the female rat
22 offspring at PND31) was found to decrease basal StAR synthesis, but not gonadotropin-
23 stimulated StAR synthesis, suggesting that Pb may not directly affect ovarian
24 responsiveness to gonadotropin stimulation (Srivastava et al.. 2004). Instead, Pb may act
25 at the hypothalamic-pituitary level to alter LH secretion, which is necessary to drive
26 StAR production and subsequent sex hormone synthesis. Release of LH and FSH from
27 the pituitary is controlled by gonadotropin-releasing hormone (GnRH). Pb exposure
28 (10 (iM, 90 min) in rat brain median eminence cells can block GnRH release (Bratton et
29 al.. 1994). Pb may also interfere with release of pituitary hormones through interference
30 with cation-dependent secondary messenger systems that mediate hormone release and
31 storage.
November 2012 5-36 Draft - Do Not Cite or Quote
-------
1 Endocrine disruption may also be a result of altered hormone binding to endocrine
2 receptors. Prenatal and postnatal Pb exposure (20 ppm in drinking water) to rats was able
3 to decrease the number of estrogen, LH, and FSH receptors found in the uterus or ovaries
4 and receptor binding affinity ("Wiebe etal.. 1988; Wiebe and Barr. 1988). Altered
5 hormone binding ability may be due to the ion binding properties of Pb, resulting in
6 changes in receptor tertiary structure that will disrupt ligand binding. In addition,
7 Pb-induced changes in hormone levels that act as inducing agents for receptor synthesis
8 may affect the number of hormone receptors produced.
9 Some of these endocrine disrupting effects of Pb have been related to the generation of
10 ROS. Treatment with antioxidants has been able to counteract a number of the endocrine
11 disrupting effects of Pb, including apoptosis and decreased sperm motility and production
12 (Salawu et al.. 2009: Shan et al. 2009: Madhavi et al.. 2007: Rubio et al.. 2006: Wang et
13 al.. 2006a: Hsu etal.. 1998b). Direct generation of ROS in epididymal spermatozoa was
14 observed after Pb treatment in rats (i.p. 20 or 50 ppm, 6 weeks) (Hsu etal.. 1998a). In
15 addition, lipid peroxidation has been observed in Pb-treated rats (i.p. 0.025 ppm, 15 days)
16 (Pandya et al.. 2012). Lipid peroxidation in the seminal plasma was significantly
17 increased in a group of Pb-exposed workers with high blood Pb levels (>40 (ig/dL) than
18 in unexposed controls (Kasperczyk et al.. 2008).
19 The liver is often associated with the HPG axis due in part to its production of insulin-
20 like growth factor 1 (IGF-1). Children with higher concurrent blood Pb levels (>4 (ig/dL)
21 (Huseman et al.. 1992) and Pb-exposed animals (blood Pb level of 14 (ig/dL) (Pine etal..
22 2006: Dearth et al., 2002) and gonadal cells (46 ppm Pb exposure) (Kolesarova et al.,
23 2010) have shown a decrease in plasma IGF-1, which may be the result of decreased
24 translation or secretion of IGF-1 (Dearth et al., 2002). IGF-1 also induces LH-releasing
25 hormone release, such that IGF-1 decrements may explain decreased LH and estradiol
26 levels. IGF-1 production is stimulated by growth hormone (GH) secreted from the
27 pituitary gland and could be the result of GH depletion.
28 A number of studies have revealed that Pb exposure affects the dynamics of growth (see
29 Section 5.8.1). Decreased growth after Pb exposure could be the result of Pb-induced
30 decreased GH levels (Berry et al.. 2002: Camoratto et al.. 1993: Huseman et al.. 1992:
31 Huseman et al., 1987). This decrease in GH could be a result of decreased release of GH
32 releasing hormone (GHRH) from the hypothalamus or disrupted GHRH binding to its
33 receptor, which has been reported in vitro after Pb treatment (IC50 free Pb in solution 5.2
34 x 10"5 (iM, 30 minutes) (Lauet al.. 1991). GH secretion may also be altered from
35 decreased testosterone, a result of Pb exposure.
November 2012 5-37 Draft - Do Not Cite or Quote
-------
5.2.6.2 Hypothalamic-Pituitary-Thyroid Axis
1 The evidence for the effects of Pb exposure on the hypothalamic-pituitary-thyroid (HPT)
2 axis is mixed. Pb exposure impacts a variety of components in the thyroid hormone
3 system. A number of occupational studies (blood Pb levels >7.3 (ig/dL) have shown that
4 elevated blood Pb are associated with lower thyroxine (T4) (and free T4 levels) without
5 alteration in triiodothyronine (T3), suggesting that long-term Pb exposure may depress
6 thyroid function in workers (Dundar et al.. 2006; Tuppurainen et al.. 1988; Robins et al..
7 1983). However, animal studies on thyroid hormones have shown mixed results.
8 Pb-exposed cows (blood Pb levels >51 (ig/dL) were reported to have an increase in
9 plasma T3 and T4 levels (Swarup et al.. 2007). whereas mice and chickens manifested
10 decreased serum T3 concentrations after Pb exposure, which was accompanied by
11 increased lipid peroxidation (Chaurasia et al.. 1998; Chaurasia and Kar. 1997). Both
12 decreased serum T3 and increased lipid peroxidation were restored by vitamin E
13 treatment, suggesting the disruption of thyroid hormone homeostasis could be a result of
14 altered membrane architecture and oxidative stress; however, no data were provided to
15 exclude changes in Pb kinetics as the mechanism of protection (Chaurasia and Kar.
16 1997).
17 Decreased T4 and T3 may be the result of altered pituitary release of thyroid stimulating
18 hormone (TSH). However, several studies have reported higher TSH levels in high-level
19 Pb-exposed workers (blood Pb levels >39 (ig/dL) (Lopez et al.. 2000; Singh et al.. 2000;
20 Gustafson et al.. 1989). which would result in increased T4 levels. Overall, results on the
21 effects of Pb on the HPT axis are inconclusive.
November 2012 5-38 Draft - Do Not Cite or Quote
-------
5.2.7 Cell Death and Genotoxicity
1 A number of studies have attempted to characterize the genotoxicity of inorganic Pb in
2 human populations, laboratory animals, and cell cultures. Endpoints investigated include
3 DNA damage (single- and double-strand breaks, DNA-adduct formation), mutagenicity,
4 clastogenicity (sister chromatid exchange, micronucleus formation, chromosomal
5 aberrations), and epigenetic changes (changes in gene expression, DNA methylation,
6 mitogenesis). It is important to note that numerous studies have utilized exposure to
7 Pb chromate to investigate genotoxicity endpoints; some studies have specifically
8 attributed the observed increases in DNA damage and clastogenicity to the chromate ion
9 while others have not. Due to the uncertainty regarding whether observed genotoxic
10 effects are due to chromate or Pb in studies using this form of inorganic Pb, only studies
11 utilizing other forms of inorganic Pb (e.g., Pb nitrate, acetate, chloride, sulfate) are
12 discussed below. Overall, evidence indicates that in vitro or in vivo exposure to various
13 Pb compounds can increase risk of genotoxic effects, including DNA damage,
14 clastogenicity, and mutagenicity.
5.2.7.1 DNA Damage
15 A number of studies in human populations have observed associations between indicators
16 of Pb exposure and increased DNA damage, as measured as DNA strand breaks. Most of
17 these associations have been observed in occupationally-exposed populations (Grover et
18 al..201Q: Minozzo etal.. 2010: Shaik and Jamil. 2009: Danadevi et al.. 2003: Hengstler
19 etal.. 2003: Palus etal.. 2003: Fracasso et al.. 2002: de Restrepo et al.. 2000). Evidence
20 overall was equivocal in regard to how blood Pb levels correlated with DNA damage:
21 Fracasso et al. (2002) observed that DNA damage increased with increasing blood Pb
22 levels (blood Pb levels, <25, 25-35, and >35 (ig/dL), whereas Palus et al. (2003) (mean
23 blood Pb level: 50.4 (ig/dL [range: 28.2 to 65.5 ng/dL]) and Minozzo et al. (2010) (mean
24 [SD]: 59.43 (ig/dL [28.34]) observed no correlation. Hengstler et al. (2003) examined
25 workers exposed to Pb, Cd, and Co and observed that neither blood (mean: 4.4 [IQR:
26 2.84-13.6] (ig/dL) nor air Pb levels (mean: 3.0 [IQR: 1.6-50.0] (ig/m3) were associated
27 with DNA damage when examined alone, but that blood Pb influenced the occurrence of
28 single strand DNA breaks when included in a multiple regression model along with Cd in
29 air and blood and Co in air.
30 A few studies were found that investigated Pb-induced DNA damage resulting from
31 nonoccupational exposures. Mendez-Gomez (2008) observed that children attending
32 grade schools at close and intermediate distances to a Pb smelter had mean (range) blood
33 Pb levels of 28.6 (11.4 to 47.5) and 19.5 (11.3 to 49.2) ng/dL, respectively, compared to
November 2012 5-39 Draft - Do Not Cite or Quote
-------
1 blood Pb level of 4.6 (0.1 to 8.7) (ig/dL for children living distant to the smelter. DNA
2 damage in lymphocytes was higher in children living nearest to the smelter, compared to
3 the children at the intermediate distance, but was not different from children living
4 farthest away from the smelter. Multivariate analysis (which considered children urinary
5 As levels, highest in children farthest from the smelter), revealed no statistically
6 significant associations between DNA damage and blood Pb level. Further, DNA repair
7 ability was also observed to be unrelated to blood Pb levels. Alternatively, Yanez et al.
8 (2003) observed that children living close to a mining complex (mean [range] blood Pb
9 level: 11.6 [3.0 to 19.5] (ig/dL) did have higher levels of DNA damage compared to
10 control children who lived further away from the mining facility (mean [range] blood Pb
11 level: 8.3 [3.0 to 25.0] (ig/dL).
12 Pb-induced DNA damage was observed in multiple animal studies. In mice exposed to Pb
13 (blood Pb level of 0.68 ug/dL) via inhalation for up to 4 weeks, differential levels of
14 DNA damage were observed in different organ systems, with only the lung and the liver
15 demonstrating statistically greater DNA damage compared to the respective organ
16 controls after acute exposure (Valverde et al.. 2002). Statistically elevated levels of DNA
17 damage were observed in the kidneys, lungs, liver, brain, nasal cavity, bone marrow, and
18 leukocytes of mice exposed to Pb over a period of 4 weeks, although variability was high
19 in all groups. The magnitude of the DNA damage was characterized as weak and did not
20 increase with increasing durations of exposure. In mice given Pb nitrate (0.7 to
21 89.6 mg/kg) by gavage for 24, 48, or 72 hours, or 1 or 2 weeks, single strand DNA breaks
22 in white blood cells were observed but did not increase with increasing concentration
23 (Devi et al.. 2000). The three highest concentrations had responses that were similar in
24 magnitude to each other and were actually lower than the responses to the lower
25 concentrations tested. Xu et al. (2008) exposed mice to 10-100 mg/kg Pb acetate via
26 gavage for four weeks and observed a concentration-dependent increase in DNA single
27 strand breaks in white blood cells that was statistically significant at 50 and 100 mg/kg.
28 The authors characterized the observed DNA damage as severe. Pb nitrate induced DNA
29 damage in primary spermatozoa in rats (blood Pb levels of 19.5 and 21.9 (ig/dL) over that
30 in control rats (Nava-Hernandez et al.. 2009). The level of DNA damage was not
31 concentration dependent and was comparable in both exposure groups. Narayana and Al-
32 Bader (2011) observed no increase in DNA damage in the livers of rats exposed to 5,000
33 or 10,000 ppm Pb nitrate in drinking water for 60 days. Interestingly, although the results
34 were not statistically significant and were highly variable within exposure groups, DNA
35 fragmentation appeared to be lower in the exposed animals.
36 Studies investigating Pb-induced DNA damage in human cell cultures were
37 contradictory. Pb acetate did not induce DNA strand breaks in human HeLa cells when
38 exposed in vitro to 500 (iM Pb acetate for 20-25 hours or 100 (iM for 0.5-4 hours
November 2012 5-40 Draft - Do Not Cite or Quote
-------
1 (Hartwig et al., 1990; Snyder and Lachmann. 1989). Pb nitrate, administered to
2 lymphoma cells in vitro at 1,000-10,000 (iM for 6 hours, did not result in any
3 DNA-protein crosslinks (Costa et al.. 1996). Pb acetate was observed by Wozniak and
4 Blasiak (2003) to result in DNA single and double strand breaks in primary human
5 lymphocytes exposed in vitro to 1-100 (iM for 1 hour, although the pattern of damage
6 was peculiar. DNA damage was greater in cells exposed to 1 or 10 (iM, compared to
7 those exposed to 100 (iM. DNA-protein crosslinks were only observed in the 100 (iM
8 exposure group, suggesting that the decreased strand breaks observed in the high
9 exposure group may be a result of increased crosslinking in this group. Pasha Shaik et al.
10 (2006) also observed DNA damage in human lymphocytes exposed in vitro to
11 2,100-3,300 (iM Pb nitrate for 2 hours. Although there was a concentration-dependent
12 increase in DNA damage from 2,100-3,300 (iM, no statistics were reported and no
13 unexposed control group was included, making it difficult to interpret these results.
14 Gastaldo et al. (2007) observed that in vitro exposure of human endothelial cells to
15 1-1,000 uM Pb nitrate for 24 hours resulted in a concentration-dependent increase in
16 DNA double strand breaks.
17 Studies in animal cell lines collectively were equally as ambiguous as those using human
18 cell lines. Zelikoff et al. (1988) and Roy and Rossman (1992) reported that Pb acetate
19 (concentration not stated and 1,000 uM, respectively) did not induce single or double
20 DNA strand breaks or DNA-protein or DNA-DNA crosslinks in CHV79 cells. However,
21 both Xu et al. (2006) and Kermani et al. (2008) reported Pb acetate-induced DNA
22 damage in undifferentiated PC12 cells exposed to 0.1, 1, or 10 (iM for 24 hours; and in
23 bone marrow mesenchymal stem cells exposed to 60 (iM for 48 hours, respectively.
24 Wedrychowski et al. (1986) reported that DNA-protein crosslinks were induced in a
25 concentration-dependent manner in hepatoma cells exposed to 50-5,000 (iM Pb nitrate
26 for 4 hours. Pb acetate and Pb nitrate increased the incidence of nick translation in
27 CHV79 cells when a bacterial DNA polymerase was added.
28 Pb exposure has also been shown to inhibit DNA repair mechanisms. Pb acetate did not
29 induce single strand DNA breaks in HeLa cells exposed to 500 uM for 20-25 hours
30 (Hartwig etal., 1990). However, exposure to both Pb acetate and UV light resulted in
31 increased persistence of UV-induced strand breaks, compared with exposure to UV light
32 alone. Similar effects were seen in hamster V79 cells: UV-induced mutation rates and
33 SCE frequency was exacerbated by co-incubation with Pb acetate. Taken together, these
34 data suggest that Pb acetate interferes with normal DNA repair mechanisms triggered by
35 UV exposure alone. Pb nitrate was observed to affect different DNA double strand break
36 repair pathways in human endothelial cells exposed in vitro to 100 uM for 24 hours.
37 Exposure to Pb inhibited nonhomologous end joining repair, but increased two other
38 repair pathways, MRE11 -dependent and Rad51 -related repair (Gastaldo et al.. 2007).
November 2012 5-41 Draft - Do Not Cite or Quote
-------
1 Interestingly, in contrast to the above studies, exposure of lung carcinoma cells to 100,
2 300, or 500 uM Pb acetate for 24 hours resulted in an increase in nucleotide excision
3 repair efficiency (Li et al., 2008a). Roy and Rossman (1992) observed an increase in UV-
4 induced mutagenicity when CHV79 cells were co-exposed to 400 uM Pb acetate (a
5 nonmutagenic concentration of Pb acetate), indicating an inhibition of DNA repair.
6 Treatment of Chinese hamster ovary cells to 0.5-500 uM Pb acetate resulted in a
7 concentration-dependent accumulation of apurinic/apyrimidinic site incision activity,
8 indicating that DNA repair was diminished (McNeill et al.. 2007).
5.2.7.2 Mutagenicity
9 Only one human study was found that investigated Pb-induced mutagenicity. Van
10 Larebeke et al. (2004) investigated the frequency of mutations in the hypoxanthine
11 phosphoribosyltransferase (HPRT) gene in Flemish women without occupational
12 exposures to Pb or to a number of other heavy metals and organic contaminants. Higher
13 blood Pb level (1 Oth-90th percentile: 1.6 to 5.2 (ig/dL) was associated with greater HPRT
14 mutation frequency than found in the total population. Also, women with high blood Pb
15 levels (i.e., greater than the population median, not reported) demonstrated a greater
16 mutation frequency compared to women with lower blood Pb levels.
17 Pb-induced mutagenicity was investigated in a few studies using human cell cultures.
18 Ye (1993) exposed human keratinocytes to 100 (iM to 1 * 105 (iM Pb acetate for 2-24
19 hours. This study did not measure HPRT mutations directly, but rather measured the
20 amount of tritium (3H) incorporated into DNA as an indicator of mutation. In the
21 presence of 6-thioguanine, tritium incorporation was increased in exposed cells,
22 indicating weak mutagenicity. Hwua and Yang (1998) reported that Pb acetate was not
23 mutagenic in human foreskin fibroblasts exposed to 500-2,000 (iM for 24 hours.
24 Pb acetate remained nonmutagenic in the presence of 3-aminotriazole, a catalase
25 inhibitor, indicating that oxidative metabolism did not play a part in potential
26 mutagenicity of Pb. Exposure to Pb acetate alone did not induce mutagenicity in lung
27 carcinoma cells (100-500 (iM for 24 hours) or fibroblasts (300-500 (iM for 24 hours) (U
28 et al., 2008a; Wang et al., 2008c). However, pretreatment with PKC inhibitors before Pb
29 treatment did result in statistically significant increases in mutagenicity in both cell lines.
30 Results from investigations into Pb-induced mutagenicity using animal cell lines were as
31 equivocal as were the findings from human cell line studies, although the mixed findings
32 may be reflective of specific Pb compounds used. Pb acetate was observed to be
33 nonmutagenic (HPRT assay) in CHV79 cells exposed to 1-25 (iM of the compound for
34 24 hours (Hartwig et al.. 1990). but elicited a mutagenic response in CHV79 cells (gpt
November 2012 5-42 Draft - Do Not Cite or Quote
-------
1 assay) exposed to 1,700 (iM for 5 days (Roy and Rossman. 1992). Pb acetate was
2 observed to be nonmutagenic (HPRT assay) in Chinese hamster ovary cells exposed to
3 5 (jJVI for 6 hours (McNeill et al.. 2007). The implication of mutagenicity in the latter
4 study is complicated by the concurrent observation of severe cytotoxicity at the same
5 concentration. Pb nitrate was alternatively found to be nonmutagenic in CHV79 cells (gpt
6 assay) exposed to 0.5-2,000 (iM for 5 days (Roy and Rossman. 1992) but mutagenic in
7 the same cell line (HPRT assay) exposed to 50-5,000 (iM for 5 days (Zelikoff et al..
8 1988). However, mutagenicity was only observed at 500 (JVI, and was higher than that
9 observed at higher Pb concentrations. Pb sulfate was also observed to be mutagenic in
10 CHV79 cells (HPRT assay) exposed to 100-1,000 (iM for 24 hours, but as with
11 Pb nitrate, it was not concentration-dependent (Zelikoff etal. 1988). Pb chloride was the
12 only Pb compound tested in animal cell lines that was consistently mutagenic: three
13 studies from the same laboratory observed concentration-dependent mutagenicity in the
14 gpt assay in Chinese hamster ovary cells exposed to 0.1-1 (iM Pb chloride for one hour
15 (Ariza and Williams. 1999: Arizaetal.. 1998: Ariza and Williams. 1996V
5.2.7.3 Clastogenicity
16 Clastogenicity is the ability of a compound to induce chromosomal damage, and is
17 commonly observed as sister chromatid exchange (SCE), micronuclei formation, or
18 incidence of chromosomal aberrations (i.e., breaks or gaps in chromosomes). Pb has been
19 shown to increase sister chromatid exchange, micronuclei formation, and chromosomal
20 aberrations in human populations, exposed animal models, and in vitro experiments.
Sister Chromatid Exchange
21 An association between blood Pb levels (means: 10.48 - 86.9 (ig/dL) and sister chromatid
22 exchange (SCE) was observed in a number of occupational studies (Wiwanitkit et al..
23 2008: Duvdu etal.. 2005: Palus etal.. 2003: Duvdu etal.. 2001: Pinto et al.. 2000:
24 Bilban. 1998: Anwar and Kamal. 1988: Huang etal.. 1988). In most studies that
25 attempted to investigate the concentration-response relationship in workers, no
26 association was observed between increasing blood Pb levels and the number of SCE
27 (Palus etal.. 2003: Duvdu etal.. 2001: Pinto et al.. 2000V However, Huang et al. (1988)
28 did observe increased SCE in exposed workers in the two highest blood Pb groups (52.1
29 and 86.9 (ig/dL), with a statistically significant association observed in the 86.9 (ig/dL
30 group. Pinto et al. (2000) did report an association with duration of exposure (range of
31 years exposed: 1.6-40). Two studies reported no correlation between occupational
32 exposure to Pb and number of SCE (Rajah and Ahuja. 1996: Rajah and Ahuja. 1995).
November 2012 5-43 Draft - Do Not Cite or Quote
-------
1 Mielzynska et al. (2006) found no association between blood Pb level and SCEs in
2 children in Poland. Children had an average blood Pb level of 7.69 (ig/dL and 7.87
3 SCEs/cell.
4 Pb exposure has been observed to induce SCEs in multiple laboratory animal studies. In
5 mice treated with up to 100 mg/kg Pb acetate i.p., Pb induced SCEs with 50 and
6 100 mg/kg (Fahmy. 1999). Pb nitrate, also administered i.p. induced the formation of
7 increased SCE levels in a concentration-dependent manner (10-40 mg/kg) in the bone
8 marrow of exposed mice (Dhir et al., 1993). Nayak et al. (1989b) treated pregnant mice
9 with 100-200 mg/kg Pb nitrate via i.v. injection and observed an increase in the number
10 of SCE in dams at 150 and 200 mg/kg; no increases in SCE levels were observed in the
11 fetuses. Tapisso et al. (2009) treated rats with 21.5 mg/kg Pb acetate (l/10th the LD50) via
12 i.p. injection on alternating days for 11 or 21 days, for a total of 5 or 10 treatments.
13 Induction of SCE in the bone marrow of exposed rats was increased over controls in a
14 statistically significant duration-dependent manner. It is important to note that all of these
15 studies utilized an injection route of exposure that may not be relevant to routes of
16 exposure in the human population (e.g., air, drinking water exposure).
17 Few studies were found that investigated SCE formation due to Pb exposure in human
18 cell lines. Statistically significant, concentration-dependent increases in SCE were
19 observed in human lymphocytes obtained from a single donor when incubated with 1,5,
20 10, or 50 (iM Pb nitrate (Ustundag and Duydu. 2007). Melatonin and N-acetylcysteine
21 were reported to ameliorate these effects, indicating Pb may induce increases in SCE
22 levels through increased oxidative stress. Pb chloride was also observed to increase SCE
23 levels in human lymphocytes exposed to 3 or 5 ppm (Turkez et al.. 2011).
24 Evidence from studies investigating SCE in rodent cells was more equivocal than that in
25 human cells. Pb sulfate, acetate, and nitrate were found not to induce SCE in CHV79
26 cells (Hartwig et al.. 1990; Zelikoff et al.. 1988). Both of these studies only examined
27 25-30 cells per concentration, reducing their power to detect Pb-induced increases in SCE
28 levels. Cai and Arenaz (1998). on the other hand, used 100 cells per treatment and
29 observed that exposure to 0.05-1 (iM Pb nitrate for 3-12 hours resulted in a weak,
30 concentration-dependent increase in SCE levels in Chinese hamster ovary cells. Lin et al.
31 (1994) also observed a concentration-dependent increase in SCE levels in Chinese
32 hamster cells exposed to 3-30 (iM Pb nitrate for 2 hours.
November 2012 5-44 Draft - Do Not Cite or Quote
-------
Micronucleus Formation
1 Pb-induced micronucleus formation was observed in numerous occupational studies
2 (Groveretal..201Q: Khanetal.. 2010b: Minozzo etal.. 2010: Shaik and Jamil. 2009:
3 Minozzo et al.. 2004: Palus etal.. 2003: Vaglenov etal.. 2001: Pinto et al.. 2000: Bilban.
4 1998: Vaglenov et al.. 1998). Pinto et al. (2000) observed increased micronuclei in
5 exposed workers with an average blood Pb level of 10.48 (ig/dL compared with
6 unexposed controls. In studies investigating the correlation between blood Pb levels and
7 micronucleus formation, no association was observed (Minozzo et al.. 2010: Minozzo et
8 al.. 2004: Palus etal.. 2003: Pinto et al.. 2000). although Pinto et al. (2000). Grover et al.
9 (2010). and Minozzo et al. (2010) did report an association between micronuclei
10 formation and duration of exposure. Mielzynska et al. (2006) investigated micronucleus
11 formation in a nonworker population and reported a statistically significant positive
12 correlation between blood Pb levels and micronuclei frequency in children in Poland.
13 Children, with an average blood Pb level of 7.69 (ig/dL, were observed to have 4.44
14 micronucleated cells per 1,000 cells analyzed. Children with blood Pb levels greater than
15 10 (ig/dL had significantly more micronucleated cells than did children with blood Pb
16 levels less than 10 (ig/dL.
17 Micronucleus formation in response to Pb exposure has been observed in rodent animal
18 studies. Celik et al. (2005) observed that exposure of female rats to Pb acetate (140, 250,
19 or 500 mg/kg once per week for 10 weeks) resulted in statistically significant increases in
20 numbers of micronucleated polychromatic erythrocytes (PCEs) compared to controls.
21 Similarly, Alghazal et al. (2008b) exposed rats to Pb acetate (100 ppm daily for 125 days)
22 and observed statistically significant increases in micronucleated PCEs in both sexes.
23 Tapisso et al. (2009) treated rats with Pb acetate (21.5 mg/kg; l/10th the LD50) via i.p.
24 injection on alternating days for 11 or 21 days, for a total of 5 or 10 exposures. Formation
25 of micronuclei in the bone marrow of exposed rats was increased over formation in
26 controls in a significant duration-dependent manner. Two further studies investigated
27 formation of micronuclei in the bone marrow of exposed mice: Roy et al. (1992) treated
28 mice with Pb nitrate (10 or 20 mg/kg, i.p.) and observed a concentration-dependent
29 increase in micronuclei, whereas Jagetia and Aruna (1998) observed an increase in
30 micronuclei in mice treated with Pb nitrate (0.625-80 mg/kg, i.p.), though the increase
31 was not concentration-dependent. Mice exposed to Pb acetate (0.1 (ig/L via drinking
32 water, a more environmentally relevant route of exposure, for 90 days) had statistically
33 significant increases in micronucleated PCEs (Marques et al.. 2006).
34 A few studies were found that reported increased micronucleus formation in human cell
35 lines treated with Pb. Concentration-dependent micronucleus formation was observed in
36 human lymphocytes when exposed in vitro to either 1, 5, 10, or 50 (iM Pb nitrate or 3 or
37 5 ppm Pb chloride (Turkez etal.. 2011: Ustundag and Duydu. 2007). Gastaldo et al.
November 2012 5-45 Draft - Do Not Cite or Quote
-------
1 (2007) also observed a concentration-dependent increase in micronuclei in human
2 endothelial cells exposed in vitro to 1-1,000 (iM Pb nitrate for 24 hours. Animal cell
3 culture studies investigating micronuclei formation produced contrasting results. One
4 study observed that micronuclei were not induced in Chinese hamster cells exposed to
5 3-30 (iM Pb nitrate for 2 hours (Lin et al.. 1994). whereas the other observed that
6 Pb acetate induced a concentration-dependent increase in Chinese hamster cells when
7 administered at 0.03-10 (iM for 18 hours (Bonacker et al., 2005).
Chromosomal Aberrations
8 Chromosomal aberrations (e.g., chromosome breaks, nucleoplasmic bridges, di- and a-
9 centric chromosomes, and rings) were examined in a number of occupational studies
10 (Groveretal..201Q: Shaik and Jamil. 2009; Pinto et al.. 2000; Bilban. 1998; De et al..
1 1 1995; Huang etal.. 1988). No correlation was observed between increasing blood Pb
12 level and the number of chromosomal aberrations, although an association was observed
13 between duration of exposure and chromosomal damage (Grover et al.. 2010; Pinto et al..
14 2000). Other studies reported no association between occupational exposure to Pb and
15 chromosomal aberrations (Anwar and Kamal. 1988; Andreae. 1983). Smejkalova (1990)
16 observed greater chromosomal damage and aberrations in children living in a heavily
17 Pb-contaminated area of Czechoslovakia compared with children living in an area with
18 less contamination, although the difference between the two areas was not statistically
19 significant. Blood Pb levels were comparable between children living in the
20 Pb-contaminated area and children living in the less contaminated area (low 30s versus
21 high 20s (ig/dL, respectively), indicating there may not be enough of a dose contrast to
22 detect a significant difference in aberration rates.
23 The majority of animal studies investigating Pb-induced genotoxicity focused on the
24 capacity of Pb to produce chromosomal damage. Fahmy (1999) treated mice with
25 Pb acetate (25-400 mg/kg i.p.), either as a single dose or repeatedly for 3, 5, or 7 days.
26 Chromosomal damage was observed to increase in bone marrow cells (100-400 mg/kg)
27 and spermatocytes (50-400 mg/kg) in a concentration-dependent manner after both
28 dosing regimens. Pb nitrate was also observed to produce concentration-dependent
29 chromosomal damage in mice treated i.p. to a single dosage of 5, 10, or 20 mg/kg (Dhir
30 et al.. 1992b). In a similar experiment, Dhir et al. (1990) treated mice with Pb nitrate (10,
3 1 20, or 40 mg/kg) and saw an increase in chromosomal aberrations, although there was no
32 concentration-dependent response as the response was similar in all concentrations tested.
33 Nayak et al. (1989b) treated pregnant mice with Pb nitrate (100, 150, or 200 mg/kg via
34 i.v. injection) and observed no chromosomal gaps or breaks in dams or fetuses but did
35 report some karyotypic chromosomal damage and weak aneuploidy at the low dose. In a
36 similar experiment, low levels of chromosomal aberrations were observed in dams and
November 20 12 5-46 Draft - Do Not Cite or Quote
-------
1 fetuses injected with Pb nitrate (12.5, 50, or 75 mg/kg), but there was no concentration-
2 dependent response reported and few cells were analyzed (Navaketal.. 1989a). In rats
3 given Pb acetate (2.5 mg/100 g body weight, i.p. daily for 5-15 days or 10-20 mg/100 g
4 once and analyzed after 15 days), Pb-induced chromosomal aberrations were observed
5 (Chakraborty et al.. 1987). The above studies all are limited by the use of a route of
6 exposure that may not be relevant to human environmental exposures. However, studies
7 utilizing oral exposures also observed increases in chromosomal damage. Aboul-Ela
8 (2002) exposed mice to Pb acetate (200 or 400 mg/kg by gavage for 5 days) and reported
9 that chromosomal damage was present in the bone marrow cells and spermatocytes of
10 animals exposed to both concentrations. Dhir et al. (1992a) also observed a
11 concentration-dependent increase in chromosomal damage in mice exposed via gavage,
12 albeit at much lower concentrations: either 5 or 10 mg/kg. Nehez et al. (2000) observed a
13 Pb-induced increase in aneuploidy and percent of cells with damage after exposure to
14 10 mg/kg administered by gavage 5 days a week for 4 weeks. In the only study that
15 investigated dietary exposure, El-Ashmawy et al. (2006) exposed mice to 5,000 ppm
16 Pb acetate in feed, and observed an increase in abnormal cells and frequency of
17 chromosomal damage.
18 In the few studies that investigated the capacity of Pb to induce chromosomal damage in
19 human cell lines, Pb exposure did not induce chromosomal damage. Wise et al. (2005;
20 2004) observed that Pb glutamate was not mutagenic in human lung cells exposed in vitro
21 to 250-2,000 (iM for 24 hours. Pasha Shaik et al. (2006) observed that Pb nitrate did not
22 increase chromosomal aberrations in primary lymphocytes (obtained from healthy
23 volunteers) when incubated with 1,200 or 2,000 (iM for 2 hours. Studies utilizing animal
24 cell lines generally supported the finding of no Pb-induced chromosomal damage in
25 human cell lines. Pb nitrate was found to induce no chromosomal damage in Chinese
26 hamster ovary cells exposed to 500-2,000 (iM for 24 hours (Wiseetal.. 1994). 3-30 (iM
27 for 2 hours (Linetal.. 1994). or 0.05 -1 (JVI for 3 -12 hours (Cai and Arenaz. 1998). Wise
28 et al. (1994) did observe increased chromosomal damage in Chinese hamster ovary cells
29 exposed to 1,000 (iM Pb glutamate for 24 hours, but did not see any damage in cells
30 exposed to higher concentrations (up to 2,000
November 2012 5-47 Draft - Do Not Cite or Quote
-------
5.2.7.4 Epigenetic Effects
1 Epigenetic effects are heritable changes in gene expression resulting without changes in
2 the underlying DNA sequence. A prime example of an epigenetic effect is the abnormal
3 methylation of DNA, which could lead to altered gene expression and cell proliferation
4 and differentiation. Possible indications of Pb-induced epigenetic changes include
5 alterations in methylation patterns in exposed rats, and alterations in mitogenesis and cell
6 proliferation in exposed humans and animals, as well as human and animal cell cultures.
DNA Methylation
7 A single i.v. injection of Pb nitrate (75 umol/kg) resulted in global hypomethylation of
8 hepatic DNA in rats (Kanduc et al.. 1991). The observed hypomethylation in the liver
9 was associated with an increase in cell proliferation. A few additional studies in humans
10 observed that higher bone Pb levels were associated with lower global DNA methylation
11 patterns in adults and cord blood of newborns (Wright et al.. 2010; Pilsner et al.. 2009).
12 Hypomethylation specifically is associated with increased gene expression. Changes in
13 DNA methylation patterns could potentially lead to dysregulation of gene expression and
14 altered tissue differentiation.
Mitogenesis
15 Conflicting results have been reported regarding Pb-induced effects on mitogenesis, with
16 both increased and decreased cell growth and mitogenesis. A discernible pattern of
17 effects is difficult to detect when analyzing effects across human, in vivo animal, and in
18 vitro studies. Only a few studies have investigated the mitogenic effects of Pb exposure
19 in human populations indirectly by examining mitogenesis or the induction of cell
20 proliferation, which can be a consequence of epigenetic changes. These studies (Minozzo
21 et al.. 2010; Minozzo et al.. 2004; Rajah and Ahuja. 1995) reported reduced mitogenesis
22 in two groups of Pb-exposed workers compared with unexposed controls (mean blood Pb
23 levels: 35.4 (ig/dL, 59.4 (ig/dL, and not reported, respectively). The observation of
24 decreased cell division in exposed workers may indicate that cells suffered DNA damage
25 and died during division, or that division was delayed to allow for DNA repair to occur. It
26 is also possible that Pb exerts an aneugenic effect and arrests the cell cycle.
27 Many studies have investigated the ability of Pb to induce mitogenesis in animal models,
28 and have consistently shown that Pb nitrate can stimulate DNA synthesis and cell
29 proliferation in the liver of animals treated with 100 (iM Pb per kg body weight, via i.v.
30 injection (Nakaiima et al.. 1995; Conietal.. 1992; Ledda-Columbano et al.. 1992;
31 Columbano et al., 1990; Columbano et al., 1987). Shinozuka et al. (1996) observed that
November 2012 5-48 Draft - Do Not Cite or Quote
-------
1 Pb-induced hepatocellular proliferation was similar in magnitude to that induced by
2 TNF-a at 100 (iM/kg; and Pb was observed to induce TNF-a in glial and nerve cells in
3 mice (and NF-KB, TNF-a, and iNOS in rat liver cells) from mice treated with Pb at
4 12.5 mg/kg and 100 (imol/kg, respectively (Cheng et al.. 2002; Menegazzi et al. 1997).
5 The only study that examined Pb exposure via inhalation (Pb acetate, 10,000 (iM for
6 4 weeks) resulted in increased cellular proliferation in murine lungs (Fortoul et al.. 2005).
7 Extensive research has been conducted investigating the potential effects of Pb on
8 mitogenesis in human and animal cell cultures. In human cell cultures, Pb acetate
9 inhibited cell growth in hepatoma cells (0.1-100 (iM for 2-6 days) (Heiman and Tonner.
10 1995) and primary oligodendrocyte progenitor cells (1 (iM for 24 hours) (Deng and
11 Poretz. 2002) but had no observable effects on growth in glioma cells (0.01-10 (iM for
12 12-72 hours) (Liu et al.. 2000). Pb glutamate had no effect on cell growth in human lung
13 cells in vitro, but did increase the mitotic index (250-1,000 (iM exposure for 24 hours)
14 (Wise et al.. 2005). The increase in the mitotic index was attributed to an arrest of the cell
15 cycle at M-phase, and was not attributed to an actual increase of cell growth and
16 proliferation. Gastaldo et al. (2007) also reported S and G2 cell cycle arrests in human
17 endothelial cells following exposure to 100 uM Pb nitrate for 24 hours. Conflicting
18 results with regard to DNA synthesis were reported, with a concentration-dependent
19 inhibition of DNA synthesis reported in hepatoma cells (1-100 (iM for 72 hours) (Heiman
20 and Tonner. 1995). but an induction of synthesis observed in astrocytoma cells (1-50 (iM
21 for 24 hours) (Lu et al.. 2002).
22 In rat fibroblasts and epithelial cells, Pb acetate, Pb chloride, Pb oxide, and Pb sulfate
23 were all observed to inhibit cell growth (10-1,000 (iM for 1-7 days and 0.078-320 (iM for
24 48 hours, respectively) (lavicoli et al.. 2001; Apostoli et al.. 2000). lavicoli et al. (2001)
25 observed that in addition to inhibiting cell growth in rat fibroblasts, Pb acetate caused
26 GS/M and S-phase arrest. Pb acetate decreased cell proliferation in mouse bone marrow
27 mesenchymal stem cells when administered at 20-100 uM for 48 hours (Kermani et al..
28 2008). Pb nitrate was alternatively reported to increase (Lin et al.. 1994) and decrease
29 (Cai and Arenaz. 1998) the mitotic index in Chinese hamster ovary cells exposed to 1 uM
30 Pb nitrate. Lin et al. (1994) did not consider cell cycle arrest when measuring the mitotic
31 index and did not observe a decrease at higher concentrations; in fact, the highest
32 concentration tested, 30 uM, had a mitotic index equal to that in the untreated control
33 cells.
November 2012 5-49 Draft - Do Not Cite or Quote
-------
5.2.7.5 Gene Expression
1 A few animal studies have investigated the ability of Pb exposure to alter gene expression
2 in regard to phase I and II metabolizing enzymes. Suzuki et al. (1996) treated rats with
3 Pb acetate or Pb nitrate (100 ug/kg via i.p. injection) and observed an induction of GST-P
4 with both Pb compounds. The induction of GST-P by Pb was observed to occur on the
5 transcriptional level and to be dependent on the direct activation of the cis-element GPEI
6 enhancer. Degawa et al. (1993) reported that Pb nitrate (20, 50, or 100 umol/kg, via i.v.)
7 selectively inhibited CYP1A2 levels. Pb was shown not to inhibit CYP1A2 by direct
8 enzyme inhibition, but rather to decrease the amount of CYP1A2 mRNA. In contrast,
9 Korashy and El Kadi (2004) observed that exposure of murine hepatoma cells to
10 Pb nitrate (10-100 uM for 24 hours) increased the amount of CYP1A1 mRNA while not
11 influencing the activity of the enzyme. NAD(P)H:quinone oxidoreductase and GST Ya
12 activities and mRNA levels were increased after exposure to Pb. Incubation of primary
13 human bronchial epithelial cells with Pb acetate (500 ug/L for 72 hours) resulted in the
14 up-regulation of multiple genes associated with cytochrome P450 activity, glutathione
15 metabolism, the pentose phosphate pathway, and amino acid metabolism (Glahn et al..
16 2008).
17 Additional animal studies provide further evidence that exposure to Pb compounds can
18 perturb gene expression. Zawia and Harry (1995) investigated whether the observed
19 Pb-induced disruption of myelin formation in rat pups exposed postnatally was due to
20 altered gene expression. In pups exposed to 2,000 ppm Pb acetate via lactation from
21 PND1-PND20, the expression of proteolipid protein, a major structural constituent of
22 myelin, was elevated (statistically significant) at PND20, compared to controls. The
23 expression of another structural element of myelin (myelin basic protein) was similarly
24 elevated in exposed animals, although not significantly so. The expression of both genes
25 returned to control levels 5 days following the termination of exposure. These data
26 suggest that altered gene expression in structural myelin proteins due to Pb exposure may
27 be responsible for observed alterations in abnormal conduction of nerve impulses. Long
28 et al. (2011) investigated the Pb-induced increase in ABCC5, an ATP-binding cassette
29 transporter, in embryonic and adult zebrafish. In the initial in vitro portion of the study,
30 exposure of zebrafish fibroblasts to 20 (iM Pb nitrate for 24 hours significantly increased
31 the induction of ABBC5 mRNA 2.68-fold over controls. Similar levels of induction were
32 observed when embryonic zebrafish were exposed to 5 (iM for 24 to 96 hours;
33 specifically, induction of ABCC5 was seen in the livers of developing embryos. In adult
34 fish, induction of ABCC5 was observed in the brains, intestines, and kidneys of exposed
35 fish, but a decrease was found in their livers. Induction of ABCC5 in adult fish was
36 observed to attenuate the toxicity of Cd (but not Hg or As); however, in developing
37 embryos, the attenuation of Pb-induced toxicity was not investigated. These findings
November 2012 5-50 Draft - Do Not Cite or Quote
-------
1 indicate that increased expression of ABCC5 due to heavy metal exposure may play a
2 part in cellular defense mechanisms.
5.2.7.6 Apoptosis
3 Occupational exposure to Pb and induction of apoptosis in various cell types was
4 investigated in a few studies. The study that directly measured apoptosis reported that
5 exposure to Pb increased apoptosis of lymphocytes compared to nonexposed controls
6 (Minozzo et al.. 2010). whereas the others reported that two early indicators of apoptosis,
7 karyorrhexis and karyolysis, were elevated in occupationally exposed workers (Grover et
8 al.. 2010; Khan et al.. 2010b). Pb nitrate was also observed to induce apoptosis in the
9 liver of exposed animals (Columbano et al.. 1996; Nakaiima et al.. 1995). Apoptosis was
10 observed in rat fibroblasts exposed in vitro to Pb acetate and rat alveolar macrophages
11 exposed to Pb nitrate davicoli et al.. 2001; Shabani and Rabbani. 2000). Observation of
12 Pb-induced apoptosis may represent the dysregulation of genetically-controlled cell
13 processes and tissue homeostasis.
5.2.8 Summary
14 The diverse health effects of Pb are mediated through multiple, interconnected modes of
15 action. Each of the modes of action discussed here has the potential to contribute to the
16 development of a number of Pb-induced health effects (Table 5-2). While this section
17 draws from earlier literature as well as newer lines of evidence, the inclusion of recent
18 evidence does not qualitatively change the previous conclusions regarding individual
19 modes of action. Rather, the more recent evidence agrees with, and thus strengthens these
20 conclusions. Evidence for the majority of these modes of action is observed with blood
21 Pb levels in humans ranging between 2 and 17 (ig/dL, with supporting evidence from
22 animal and in vitro assays. As many of these studies examined adults, with likely higher
23 past than current Pb exposures, uncertainty exists as to the Pb exposure level, duration,
24 frequency, and timing associated with these modes of action. The blood Pb levels or in
25 vitro concentrations presented in Table 5-2 reflect the current evidence for these modes
26 of action and are not intended to convey conclusions regarding specific thresholds. Also,
27 the data presented in this table do not inform the exposure frequency and duration
28 required to elicit a particular MOA.
November 2012 5-51 Draft - Do Not Cite or Quote
-------
Table 5-2 MOAs, their related health effects, and information on concentrations
eliciting the MOAs.
Mode of Action
[Related Health Effects
(ISA Section)]
Concentrations or Doses (Conditions)
Blood Pb
Dose
Altered Ion Status
[All Health Effects of Pb]
3.5 ug/dL
(Mean in cord blood; association with cord
blood Ca2+ATPase pump activity)
Hueletal. (2008)
0.00005 uM free Pb2+
(In vitro; 30 minutes; calmodulin activation
assay)
Kern et al. (2000)
Protein Binding
[Renal (5.5). Hematological
Effects (5.7)1
17.0ug/dL
(Concurrent mean in adult workers with
wildtype metallothionein expression;
increased BP susceptibility)
Chen et al. (201 Oa)
50 uM Pb glutamate
(In vitro; 24 hours; increased nuclear protein
in neurological cell)
Klann and Shelton (1989)
Oxidative Stress
[All Heath Effects of Pb]
5.4 ug/dL
(Concurrent mean in adult male workers;
decreased CAT activity in blood)
Conterato et al. (In Press)
0.1 uM Pb acetate
(In vitro; 48 hours; decreased cellular GSH in
neuroblastoma cells)
Chetty et al. (2005)
Inflammation
[Nervous System (5.3).
Cardiovascular (5.4). Renal
(5.5). Immune (5.6). Respiratory
(5.9.6). Hepatic (5.9.1)1
Among males with concurrent blood Pb
> 2.5 ug/dL
(Increased serum TNF-a and blood WBC
count)
Kim et al. (2007)
0.01 uM Pb acetate
(In vitro; 48 hours; increased cellular PGE2 in
neuroblastoma cells)
Chetty et al. (2005)
Endocrine Disruption
[Reproductive and
Developmental Effects (5.8).
Endocrine System (5.9.3). Bone
and Teeth (5.9.4)1
1.7 ug/dL
(lowest level at which a relationship could be
detected in adult women with both ovaries
removed; increased serum FSH)
Krieg (2007)
10 uM Pb nitrate
(In vitro; 30 minutes; displaced GHRH
binding to rat pituitary receptors)
Lau et al. (1991)
Cell Death/Genotoxicity
[Cancer (5.10). Reproductive
and Developmental Effects
(5.8). Bone and Teeth (5.9.4)1
3.3 ug/dL
(concurrent median in adult women;
increased rate of HPRT mutation frequency)
Van Larebeke et al. (2004)
0.03 uM Pb acetate
(In vitro; 18 hours; increased formation of
micronuclei)
Bonacker et al. (2005)
aThis table provides examples of studies that report effects with low Pb dosages or concentrations; they are not the full body of
evidence used to characterize the weight of the evidence. In addition, the levels cited are reflective of the data and methods
available and do not imply that these modes of action are not acting at lower Pb exposure or blood Pb levels or that these doses
represent the threshold of the effect. Additionally, the blood concentrations and doses (indicating Pb concentrations from in vitro
systems) refer to the concentrations and doses at which these modes of action were observed. While the individual modes of
action are related back to specific health effects sections (e.g., Nervous System, Cardiovascular), the concentrations and doses
given should not be interpreted as levels at which those specific health effects occur.
1
2
o
J
4
5
6
7
8
9
10
11
The alteration of cellular ion status (including disruption of Ca2+ homeostasis, altered ion
transport mechanisms, and perturbed protein function through displacement of metal
cofactors) appears to be the major unifying mode of action underlying all subsequent
modes of action (Figure 5-1). Pb is well characterized to interfere with endogenous Ca2+
homeostasis (necessary as a cell signal carrier mediating normal cellular functions).
[Ca2+]j has been shown to increase after Pb exposure in a number of cell types including
bone, erythrocytes, brain cells, and white blood cells, due to the increased flux of
extracellular Ca2+ into the cell. This disruption of ion transport is due in part to the
alteration of the activity of transport channels and proteins, such as Na+/K+ATPase and
voltage-gated Ca2+ channels. Pb can interfere with these proteins through direct
competition between Pb and the native metals present in the protein metal binding
November 2012
5-52
Draft - Do Not Cite or Quote
-------
1 domain or through disruption of proteins important in Ca2+-dependent cell signaling, such
2 as PKC or calmodulin.
3 Disruption of ion transport not only leads to altered Ca2+ homeostasis, but it can also
4 result in perturbed neurotransmitter function. Pb has been shown to displace metal ions,
5 (such as Zn2+, Mg2+, and Ca2+) from proteins due to the flexible coordination number of
6 Pb and multiple ligand binding ability, leading to abnormal conformational changes in
7 proteins and altered protein function. Evidence for this metal ion displacement and
8 protein perturbation has been shown at t 10~6 (iM concentrations of Pb. Additional effects
9 of altered cellular ion status are the inhibition of heme synthesis and decreased cellular
10 energy production due to perturbation of mitochondrial function.
11 Although Pb can bind to proteins within cells through interactions with side group
12 moieties, thus potentially disrupting cellular function, protein binding of Pb may
13 represent a mechanism by which cells protect themselves against the toxic effects of Pb.
14 Intranuclear and intracytosolic inclusion body formation has been observed in the kidney,
15 liver, lung, and brain following Pb exposure. A number of unique Pb binding proteins
16 have been detected, constituting the observed inclusion bodies. The major Pb binding
17 protein in blood is ALAD with carriers of the ALAD-2 allele potentially exhibiting
18 higher Pb binding affinity. Additionally, metallothionein is an important protein in the
19 formation of inclusion bodies and mitigation of the toxic effects of Pb.
20 A second major mode of action of Pb is its role in the development of oxidative stress,
21 due in many instances to the antagonism of normal metal ion functions. The origin of
22 oxidative stress produced after Pb exposure is likely a multipathway process, resulting
23 from oxidation of 5-ALA, NAD(P)H oxidase activation, membrane and lipid
24 peroxidation, and antioxidant enzyme depletion. Through the inhibition of 5-ALAD (due
25 to displacement of Zn by Pb), accumulated 5-ALA goes through an auto-oxidation
26 process to produce ROS. Additionally, Pb can induce the production of ROS through the
27 activation of NAD(P)H oxidase. Pb-induced ROS can interact with membrane lipids to
28 cause a membrane and lipid peroxidation cascade. Enhanced lipid peroxidation can also
29 result from Pb potentiation of Fe2+ initiated lipid peroxidation and alteration of membrane
30 composition after Pb exposure. Increased Pb-induced ROS can also sequester and
31 inactivate biologically active NO, leading to the increased production of the toxic product
32 nitrotyrosine, increased compensatory NOS, and decreased sGC protein. Pb-induced
33 oxidative stress not only can result from increased ROS production but also through the
34 alteration and reduction in activity of the antioxidant defense enzymes. The biological
35 actions of a number of these enzymes are antagonized due to the displacement of the
36 protein functional metal ions by Pb.
November 2012 5-53 Draft - Do Not Cite or Quote
-------
1 In a number of organ systems, Pb-induced oxidative stress is accompanied by
2 misregulated inflammation. Pb exposure can modulate inflammatory cell function,
3 production of pro-inflammatory cytokines and metabolites, inflammatory chemical
4 messengers, and pro-inflammatory signaling cascades. Cytokine production is skewed
5 toward the production of pro-inflammatory cytokines like TNF-a and IL-6 as well as
6 toward the promotion of a Th2 response and suppression of a Thl response accompanied
7 by decreased production of related cytokines.
8 Evidence indicates that Pb is a potent endocrine disrupting chemical. Pb can disrupt the
9 HPG axis evidenced by altered serum hormone levels, such as FSH, LH, testosterone,
10 and estradiol. Pb can interact with the hypothalamic-pituitary level hormone control
11 causing a decrease in pituitary hormones, alteration of growth dynamics due to decreased
12 IGF-1, inhibition of LH secretion, and reduction in StAR protein. Pb has also been shown
13 to alter hormone receptor binding likely due to interference of metal cations with
14 secondary messenger systems and receptor ligand binding and through generation of
15 ROS. Pb also may disrupt the HPT axis by alteration of a number of thyroid hormones,
16 possibly due to oxidative stress. However, the results of these studies investigating HPT
17 are mixed.
18 The association of Pb with increased genotoxicity and cell death has been investigated in
19 humans, animals, and cell models. Occupational Pb exposure in humans has been
20 associated with increased DNA damage; however, lower blood Pb and exposure levels
21 have been associated with these effects in experimental animals and cells. While not
22 entirely consistent, a number of studies reported decreased repair processes following Pb
23 exposure. There is evidence of mutagenesis and clastogenicity in highly-exposed
24 humans; however, weak evidence has been shown in animals and cell based systems.
25 Human occupational studies provide limited evidence for micronucleus formation (blood
26 Pb levels >10 (ig/dL) and are supported by Pb-induced effects in both animal and cell
27 studies at higher exposure levels. Animal studies have also provided evidence for
28 Pb-induced chromosomal aberrations. The observed increases in clastogenicity may be
29 the result of increased oxidative damage to DNA due to Pb exposure, as co-exposures
30 with antioxidants ameliorate the observed toxicities. Limited evidence of epigenetic
31 effects is available, including abnormal DNA methylation, mitogenesis, and gene
32 expression. Pb may alter gene expression by displacing Zn from multiple transcriptional
33 factors, thus perturbing their normal cellular activities. Consistently positive results have
34 provided evidence of increased apoptosis following Pb exposure.
35 Similar to Pb, other polyvalent metal ions (e.g., Cd, Cr, Be, Ba, Se, Sr, As, Al, Cu) have
36 demonstrated molecular mimicry and displacement of biological cations (Garza et al..
37 2006). In this manner, these metal ions share with Pb a common central mode of action of
November 2012 5-54 Draft - Do Not Cite or Quote
-------
1 disruption of ion status. Specifically, these metals have been shown to disrupt cellular
2 processes as diverse as Ca2+ homeostasis, cell signaling, neurotransmitter release, cation
3 membrane channel function, protein-DNA binding, and cellular membrane structure
4 (Pentyalaetal.. 2010; Huang et al.. 2004; Atchison. 2003; Jehan and Motlag. 1995;
5 Richardt et al.. 1986; Cooper and Manalis. 1984; Habermann et al., 1983). Additionally,
6 presumably through their shared central mode of action, some of these metal ions also
7 display corresponding downstream modes of actions such as oxidative stress, apoptosis,
8 and genotoxicity (Jomova and Valko. 2011; Jomovaet al.. 2011; Matovic et al.. 2011;
9 Agarwal et al., 2009; Mendez-Gomez et al.. 2008; Rana. 2008; Hengstler et al., 2003).
10 Overall, Pb-induced health effects can occur through a number of interconnected modes
11 of action that generally originate with the alteration of ion status.
5.3 Nervous System Effects
5.3.1 Introduction
12 The 2006 Pb AQCD concluded that the "overall weight of the available evidence
13 provides clear substantiation of neurocognitive decrements being associated in young
14 children with blood-Pb concentrations..." (U.S. EPA. 2006b). This conclusion was based
15 on evidence from several prospective and cross-sectional epidemiologic studies
16 conducted in diverse populations with adjustment for potential confounding by
17 socioeconomic status (SES), parental intelligence, and parental caregiving quality and
18 stimulation. The association between blood Pb levels and cognitive function decrements
19 was substantiated in an international pooled analysis of children, ages 4.8 to 10 years,
20 participating in seven prospective studies (Boston, MA; Cincinnati, OH; Rochester, NY;
21 Cleveland, OH; Mexico City, Mexico; Port Pirie, Australia; and Kosovo, Yugoslavia)
22 (Lanphear et al.. 2005). Across all previously evaluated studies, associations between
23 blood Pb levels and decrements in full-scale intelligence quotient (FSIQ), infant mental
24 development, memory, learning, and executive function were found in children ages 2 to
25 17 years with population mean blood Pb levels (measured at various lifestages and time
26 periods) 5-10 (ig/dL; however, several results indicated associations in groups of children
27 (ages 2-10 years) with mean blood Pb levels in the lower range of 3-5 (ig/dL (Bellinger.
28 2008; Canfield. 2008; Hornung. 2008; Tellez-Roio. 2008). Based on fewer available
29 studies, the 2006 Pb AQCD described evidence from prospective and cross-sectional
30 epidemiologic studies for associations of childhood blood Pb levels with attention-related
31 behavioral problems in children ages 6-13 years and misconduct and delinquent behavior
32 in children ages 7-17 years and young adults ages 21-22 years (U.S. EPA. 2006b).
November 2012 5-55 Draft - Do Not Cite or Quote
-------
1 Biological plausibility for epidemiologic evidence in children was provided by similarly
2 consistent toxicological findings for Pb-induced impairments in learning and behavior in
3 rodents and monkeys (U.S. EPA. 2006b). Pb exposure was not found consistently to
4 affect the memory of animals. In animals, learning impairments were demonstrated
5 largely as poorer performance in maze tests, shorter interresponse times on schedule
6 controlled behavior tasks, and response perseveration errors in discrimination reversal
7 tests. Some results from these tests also indicated Pb-induced increases in inattention.
8 Pb-induced impulsivity in animals was demonstrated as increased response rates on the
9 Fixed Ratio (FR)/waiting for reward test. These effects on learning and behavioral
10 problems in animals were found predominately with Pb exposures that resulted in blood
11 Pb levels 20-50 (ig/dL; however, some studies observed these impairments in rodents
12 (pre- and/or post-natal Pb exposure) and monkeys (postnatal Pb exposure) with blood Pb
13 levels 14-25 (ig/dL (Kuhlmann et al.. 1997; Altmann et al.. 1993; Rice and Karpinski.
14 1988; Gilbert and Rice. 1987). Toxicological studies further provided biological
15 plausibility for Pb-induced learning impairments and behavioral problems by
16 characterizing modes of action. Evidence for Pb affecting neuronal development and
17 function at the cellular and subcellular level (e.g., blood brain barrier integrity, synaptic
18 architecture during development, neurite outgrowth, glial growth, neurotransmitter
19 release, oxidative stress), provided biological plausibility for associations observed
20 between blood Pb levels and deficits in multiple functional domains such as cognitive
21 function, motor function, memory, mood, and behavioral problems in children.
22 Additional biological plausibility was provided by associations observed of childhood
23 blood Pb levels with changes indicative of neuronal damage and altered brain physiology
24 assessed in small groups of children (Meng etal. 2005; Trope etal.. 2001) and young
25 adults (Yuan et al.. 2006; Cecil et al.. 2005) using magnetic resonance imaging
26 techniques.
27 A common finding across several different populations of children was a supralinear
28 concentration-response relationship between blood Pb level and cognitive function
29 decrements, i.e., a larger decrement in cognitive function per unit increase in blood Pb
30 level in children in the lower range of the population blood Pb level distribution (Kordas
31 et al.. 2006; Schnaas et al., 2006; Tellez-Rojo et al.. 2006; Bellinger and Needleman.
32 2003; Canfield et al.. 2003a). Most of these epidemiologic results were based on the
33 analysis of concurrent blood Pb levels and a cut-point of 10 (ig/dL to define lower and
34 higher blood Pb levels. These findings were corroborated in pooled analyses of seven
35 cohorts, which indicated that a nonlinear relationship fit the data better than a linear
36 relationship (Lanphear et al.. 2005; Rothenberg and Rothenberg. 2005). Explanations for
37 the supralinear concentration-response were not well characterized.
November 2012 5-56 Draft - Do Not Cite or Quote
-------
1 Another area of focus was the comparison of various lifestages and time periods of Pb
2 exposure with respect to increasing risk of neurodevelopmental deficits. Toxicological
3 studies clearly demonstrated that gestational Pb exposure with or without additional early
4 postnatal exposure resulted in neurodevelopmental impairments. Nonetheless, not all
5 neurodevelopmental effects in animals had a single defined window of risk; for example,
6 postnatal-only and lifetime Pb exposures also were shown to impair learning and
7 behavior. Epidemiologic studies observed decrements in cognitive function in children
8 ages 3 to 17 years in association with prenatal, peak childhood, cumulative childhood,
9 and concurrent blood Pb levels. Although examined in few studies, tooth or bone Pb
10 levels were associated with cognitive function decrements and behavioral problems in
11 children and adolescents (Wasserman et al.. 2003; Bellinger etal.. 1994b: Fergusson et
12 al., 1993; Needleman et al., 1979). also pointing to an effect of cumulative childhood Pb
13 exposure. Among studies of children (ages 3-10 years) that examined blood Pb levels
14 measured at multiple lifestages and time periods, several found that concurrent blood Pb
15 was associated with a similar magnitude or larger decrement in FSIQ than blood Pb
16 levels measured earlier in childhood or averaged over multiple years (Lanphear et al..
17 2005; Wasserman et al.. 1994; Dietrich et al.. 1993). A common limitation of prospective
18 studies of children was the high correlation among blood Pb levels at different ages,
19 making it difficult to identify an individual critical lifestage or duration of Pb exposure
20 associated with risk of neurodevelopmental decrements (Lanphear et al.. 2005). Some
21 evidence indicated the persistence of neurodevelopmental effects of Pb exposure, by
22 associations of biomarkers of earlier childhood Pb exposure (e.g., deciduous tooth, blood
23 at age 2 or 6 years) with cognitive function decrements and behavioral problems in
24 adolescents and young adults (Ris et al.. 2004; Wasserman et al.. 2003; Bellinger et al..
25 1994a: 1994b: Fergusson et al.. 1993: Baghurst et al.. 1992: Needleman et al.. 1979).
26 Persistence of effects also was demonstrated by findings in some studies of rats and
27 monkeys that gestational and/or early postnatal Pb exposures were associated with
28 impairments in cognitive function and behavior in animals evaluated as adults
29 (Kuhlmann et al.. 1997: Altmann et al.. 1993: Rice. 1992b. 1990).
30 In epidemiologic studies of adults, a range of nervous system effects (e.g., impaired
31 memory, attention, reaction time, visuomotor tasks and reasoning, alterations in visual or
32 brainstem evoked potentials, postural sway) were mostly clearly indicated in Pb-exposed
33 workers with blood Pb levels in the range of 14 to 40 ug/dL (Iwataet al.. 2005; Bleecker
34 et al.. 1997: Baker etal.. 1979: Cantarow and Trumper. 1944). In the smaller body of
35 studies examining nonoccupationally-exposed adults, poorer cognitive performance was
36 associated with bone Pb levels (Weisskopf et al.. 2004: Wright et al.. 2003) but not
37 concurrent blood Pb levels (Krieg etal.. 2005; Nordberg et al.. 2000; Pavtonetal.. 1998;
38 Muldoon et al.. 1996). These findings suggested the influence of past or cumulative Pb
39 exposures on cognitive function decrements in nonoccupationally-exposed adults. With
November 2012 5-57 Draft - Do Not Cite or Quote
-------
1 regard to neurodegenerative diseases, whereas a few toxicological studies found
2 Pb-induced amyloid plaques, a pathology commonly found in the brains of adults with
3 Alzheimer's disease (Bashaet al., 2005; Zawia and Basha. 2005), epidemiologic studies
4 did not indicate that Pb exposure was associated with Alzheimer's Disease in adults.
5 Blood and bone Pb levels were inconsistently associated with amyotrophic lateral
6 sclerosis (ALS) in adults in the general population; however, in some case-control
7 studies, history of occupational Pb exposure was more prevalent among ALS cases than
8 controls (Kamel et al. 2002; Chancellor etal.. 1993). Associations were reported for
9 essential tremor and symptoms of anxiety and depression in adults, but each was
10 examined in only a few studies.
11 As discussed throughout this section, recent epidemiologic and toxicological studies
12 continued to demonstrate associations of Pb exposure and biomarkers of Pb exposure
13 with nervous system effects. The strongest evidence continued to be derived from
14 associations observed for Pb exposure and blood Pb levels in young animals and children,
15 respectively, with cognitive function decrements. Several recent studies in children
16 expanded the evidence for associations between concurrent blood Pb levels and attention-
17 related behavioral problems. Recent epidemiologic studies in adults focused primarily on
18 cognitive function decrements but provided additional evidence for Pb-associated
19 psychopathological effects, ALS, Parkinson's disease, and essential tremor. Recent
20 toxicological studies supported evidence for the effects of prenatal and postnatal Pb
21 exposure on learning, memory, and impulsivity in animals and examined interactions
22 between Pb exposure and stress. New or expanded areas of toxicological research related
23 to Pb exposure included, neurofibrillary tangle formation and neurodegenerative effects
24 after early life Pb exposures and effects potentially related to psychopathological effects.
25 Recent toxicological studies added to the large extant evidence base for Pb-induced
26 effects on endpoints describing modes of action, including neurotransmitters, synapses,
27 glia, neurite outgrowth, the blood brain barrier, and oxidative stress. The data detailed in
28 the subsequent sections continue to enhance the understanding of the spectrum of nervous
29 system effects associated with Pb exposure.
November 2012 5-58 Draft - Do Not Cite or Quote
-------
5.3.2 Cognitive Function
1 Epidemiologic studies have assessed cognitive function extensively by FSIQ and its
2 verbal and performance subscale components in children ages 3 to 17 years. FSIQ has
3 strong psychometric properties (i.e., reliability, consistency, validity), is among the most
4 rigorously standardized cognitive function measures, is relatively stable in school-age,
5 and has been predictive of life success. In children ages 6 months to 3 years, mental
6 development has been assessed with the Bayley Scales of Infant Development. A large
7 body of evidence also comprises associations of blood and tooth Pb levels with memory
8 and learning, executive function, language, and visuospatial processing. Several of these
9 domains of cognitive function are evaluated in the subtests of FSIQ, and some are more
10 comparable to endpoints examined in tests in animals. Fewer studies have examined
11 academic performance and achievement; however, these outcomes may provide
12 information on the impact of Pb exposure on life success. In the subsequent sections, the
13 epidemiologic evidence for each of these categories of outcomes is reviewed separately
14 in order of strength of evidence as assessed by the following parameters. Emphasis was
15 placed on prospective studies with repeated measurements of blood Pb levels and
16 cognitive function and on studies that examined blood Pb levels more similar to those of
17 contemporary U.S. children (i.e., <5 (ig/dL) and children whose blood Pb levels were less
18 influenced by higher past Pb exposures. Studies of chelation in children generally were
19 not included because the high pre-chelation blood Pb levels may limit generalizability of
20 results, and chelation itself has been linked to neurodevelopmental effects.
21 Many factors have been shown to influence the cognitive function of children, including
22 parental SES, parental education, parental IQ, quality and stability of parental caregiving
23 environment (often measured as Home Observation for the Measurement of Environment
24 inventory [HOME]), nutritional status, and birth weight (Nation and Gleaves. 2001;
25 Wasserman and Factor-Litvak. 2001). These and other influences on neurodevelopment
26 often are correlated with blood Pb levels. Thus, due to their association with both blood
27 Pb level and causal association with outcome, these other risk factors potentially may
28 bias or confound the associations observed between blood Pb level and indices of
29 cognitive function. In the evaluation of the effects of Pb independent from the effects of
30 the other risk factors, greater weight was given to studies that more extensively accounted
31 for potential confounding in the study design or in statistical analyses. A detailed
32 evaluation of control for potential confounding in associations between indicators of Pb
33 exposure and neurodevelopmental effects is located in Section 5.3.14.
November 2012 5-59 Draft - Do Not Cite or Quote
-------
5.3.2.1 Full Scale IQ in Children
Evidence from Prospective Studies
1 Prospective cohort studies that were initiated in the 1980s addressed some limitations of
2 cross-sectional studies, including better characterizing the temporal sequence between
3 blood Pb levels and cognitive function, examining the persistence of cognitive function
4 decrements to older ages, and comparing associations among blood Pb levels measured at
5 various life stages or representing various time periods. Recruitment of participants before
6 or at birth without consideration of Pb exposure or maternal IQ, high follow-up
7 participation (>70%), and nonselective loss-to-follow-up in most studies increase
8 confidence that the observed associations are not due to selection bias. Moreover,
9 cooperation among investigators to adopt similar study protocols (e.g., similar tests of IQ
10 and consideration of similar potential confounding factors) strengthened inferences
11 regarding the consistency of associations with blood Pb level by facilitating pooled
12 analyses and by reducing sources of heterogeneity in evaluating results across
13 populations that varied in geographic location, proximity to Pb sources, blood Pb level
14 range, race/ethnicity, and SES.
15 Individual cohort studies of varying sample sizes (n = 148-375) conducted in several
16 different populations (e.g., Boston, MA; Cincinnati, OH; Rochester, NY; Cleveland, OH;
17 Mexico City, Mexico; Port Pirie and Sydney, Australia; and Kosovo, Yugoslavia) were
18 consistent in demonstrating associations of higher blood Pb measured prenatally
19 (maternal or umbilical cord), earlier in childhood, or averaged over childhood with lower
20 FSIQ measured later in childhood, i.e., 4 to 17 years (Schnaas et al. 2006; Ris et al..
21 2004; Canfield et al.. 2003a: Schnaas et al.. 2000; Factor-Litvak et al.. 1999; Tong et al..
22 1996: Wasserman et al.. 1994: Dietrich et al.. 1993b: Baghurst et al.. 1992: Bellinger et
23 al.. 1992: Bellinger et al.. 1991: McMichael et al.. 1988) (Figure 5-2 and Table 5-3). Null
24 or weak associations were limited to a few cohorts, namely, the Cleveland and Sydney
25 cohorts (Greene et al.. 1992; CooneyetaL 1991; 1989a. b; Ernhart et al.. 1988). In the
26 prospective studies, lower FSIQ also was associated with higher concurrent blood Pb
27 levels (Figure 5-2 and Table 5-3) and tooth Pb levels. These latter results were based on
28 cross-sectional analyses; however, the pattern of associations observed for blood Pb
29 levels measured at various lifestages or time periods does not indicate that reverse
30 causation explains the FSIQ decrements observed in association with concurrent blood
31 Pb or tooth Pb levels.
32 In addition to better characterizing the temporal sequence between Pb exposure and
33 decrements in FSIQ, a common strength of most prospective studies was the adjustment
34 for several of the potential confounding factors noted above, including maternal IQ and
November 2012 5-60 Draft - Do Not Cite or Quote
-------
1 education, child sex and birth weight, SES, and HOME score (Table 5-3). Although not
2 considered as frequently, some studies also indicated lack of confounding by parental
3 smoking, birth order, and nutritional factors. Multiple testing of associations with blood
4 Pb levels and/or FSIQ was common in prospective studies that found and did not find
5 associations between blood Pb level and FSIQ. However, higher probability of
6 associations due to chance alone does not appear to unduly influence the evidence
7 because in studies that found associations, there was a consistent pattern of blood
8 Pb-associated cognitive function decrements across the various ages of blood Pb level
9 and/or cognitive assessments evaluated (Table 5-3). Studies finding null or weak
10 associations also tended to show a consistent pattern across the various analyses
11 conducted.
November 2012 5-61 Draft - Do Not Cite or Quote
-------
Study Blood PbTiming Blood PbMean(SD) Blood PbInterval FSIQage(yr)
(ug/dL) examined"(ug/dL)
Prospective Studies
Lanphearetal. (2005) Concurrent, peak< 7.5 3.2
Canfield etal. (2003) Concurrent, peak< 10 3.3
Juskoetal. (2008)" Peak
Bellingeretal. (1992) Age2 yr, peak< 10
Dietrich etal. (1993) Concurrent
Schnaasetal. (2006) Prenatal (maternal)
Cooney etal.(1991)b Age3-5yravg
Wasserman etal. (1997) Oto7yravg
Tongetal. (1996) Oto 11-13yravg
11.4(7.3)
3.8
11.8 (6.3) (Age 5)
1.3-6
0.5-8.4
2.1-10
1.9.3
5.5-10
7.8 geometric mean 3.2-10
NR NR
16.2 geometric mean6.0-10
14.0(1.2) geometric 12.7-18.1
Kordasetal. (2011)
Greene etal. (1992)
Minetal. (2009)
Prenatal (cord)
Concurrent
Age2yr
Concurrent
4 yr
4 yr
6.6(3.3)
8.7(4.4)
3.2-10.8
4.2-10
15.6 (1.4) geometric 10.1-24.0
7.0(4.1) 3.0-10
Cross-sectional Studies
Kim etal. (2009) Concurrent, low Mn 1.73(0.80)
Concurrent, high Mn
0.9-2.8
Fultonetal. (1987) Concurrent 11.5(range:3.3-34) 5.6-10
Royetal. (2011) Concurrent 11.4(5.4) 5.8-10
FSIQage(yr)
6 O
7 D
11-13 -D-
4.8 O
6-9 —*—
Change in FSIQ (95% Cl) per 1 ug/dL increase in various intervals
of blood Pb level
aSee Table 5-3 for explanation of the blood Pb level interval examined. Where possible, effect estimates were calculated for the
lowest range examined or the 10th percentile of blood Pb level to a blood Pb level of 10 ug/dL.
bSufficient data were not provided to calculate 95% Cl.
Note: Results are presented for most of the cohorts examined in the literature and generally are presented in order of strength of
study design and representativeness of study population. Evidence usually is presented for the oldest age examined in cohorts.
Multiple results from a cohort are grouped together. To facilitate comparisons among effect estimates across studies with different
distributions of blood Pb levels and model structures (e.g., linear, log-linear), effect estimates are standardized to a 1 ug/dL increase
for the lowest range of blood Pb levels examined or the interval from the 10th percentile of blood Pb level to 10 ug/dL. For
populations with 10th percentile near or above 10 ug/dL, the effect estimate was calculated for the 10th to 90th percentile of blood
Pb level. The percentiles are estimated using various methods and are only approximate values. Effect estimates are assumed to
be linear within the blood Pb level interval evaluated. The various tests used to measure FSIQ are scored on a similar scale
(approximately 40-160 FSIQ points). Black diamonds, blue circles, orange triangles, and gray squares represent effect estimates for
concurrent, earlier childhood, prenatal, and lifetime average blood Pb levels, respectively. The lines represent 95% confidence
intervals (Cl).
Figure 5-2 Associations of blood Pb levels with full-scale IQ (FSIQ) among
children.
November 2012
5-62
Draft - Do Not Cite or Quote
-------
Table 5-3 Additional characteristics and quantitative results for studies
represented in Figure 5-2
Study
Study Population and Methodological Details
Blood Pb Data
(Ma/dL)
FSIQ
Testing3
Effect
Estimate
(95% CIV3
Prospective Studies:
Lanphear et
al. (2005)
Canfield et al.
(2003a)
Jusko et al.
(2008)
103 children pooled from Boston, Cincinnati,
Cleveland, Mexico City, Port Pirie, Rochester, and
Yugoslavia cohorts.
Uniform analysis of cohorts from diverse locations
and SES. Blood Pb levels and FSIQ measured at
different ages. Several sensitivity analyses to
examine heterogeneity of results by cohort, model
specification, and confounding. Linear regression
model adjusted for HOME score, birth weight,
maternal IQ, maternal education. Also considered
potential confounding by child sex, birth order,
maternal age, marital status, prenatal smoking status
and alcohol use.
101 children born 1994-1995 followed from age 6 mo
to 5 yr, Rochester, NY
Recruitment from study of dust control. 73%
nonwhite. High follow-up participation, no selective
attrition. Linear regression model adjusted for
maternal race, IQ, education, and prenatal smoking
status, household income, HOME score, child sex,
Fe status, birth weight.
1 74 children born 1 994-1 995 followed from age 6 mo
to 6 yr, Rochester, NY
Same cohort as above. High follow-up participation.
Participants had higher maternal IQ. Nonparametric
regression model adjusted for maternal race, IQ,
education, and prenatal smoking status, HOME
score, family income, child sex, birth weight, and
Fe status.
Concurrent,
children with peak
<7.5
Mean: 3.2
Interval analyzed:
1.3-6.0 =
5th-95th
percentiles
Concurrent,
children with peak
<10
Mean: 3.3
Interval analyzed:
<1-8.4 = range
(1 = detection
limit)
Peak
Mean (SD):
11.4(7.3)
Interval analyzed:
2.1-10
WISC-III, -2.94
WISC-R, (-5.16, -0.71)
WPPSI,
WISC-S
Ages
4.8-1 Oyr
Stanford- -1 .8
Binet (-3.0, -0.60)
Age 5 yr
WPPSI-R -1.2°
Age 6 yr
Bellinger and
Needleman
(2003)
48 children followed from birth (1979-1981) to age
10yr, Boston, MA area
Recruitment at birth hospital. Moderate follow-up
participation. Participants had higher SES and HOME
scores. 95% white. Linear regression model adjusted
for HOME score (age 10 and 5 yr), maternal race, IQ,
and marital status, SES, child sex, birth order, and
stress, and number of residence changes. Also
considered potential confounding by family stress,
maternal age, psychiatric factors, child serum ferritin
levels.
Earlier childhood
(age 2 yr),
children with peak
<10
Mean: 3.8
Interval analyzed:
1-9.3 = range
WISC-R
Age 10 yr
-1.56
(-2.9, -0.20)
Mazumdar et 43 adults followed from birth (1979-1981) to age
al. (2011) 28-30 yr, Boston, MA area
Same cohort as above. Small proportion of original
cohort but no selective attrition. 93% white. Linear
regression model adjusted for maternal IQ. Also
found associations adjusted for maternal marital
status at birth, education at birth, prenatal smoking
status, or alcohol use, HOME score (mean across
ages), subject sex, birth weight, birth order,
gestational age, race, concussion history, or current
smoking status. Also considered potential
confounding by subject alcohol use.
Earlier childhood
avg (age 6 mo-
10yr):NR
Mean (SD), [age]:
8.0 (5.3) [6 mo],
10.0 (6.7) [1 yr],
7.7 (4.0) [2 yr],
6.7 (3.6) [4 yr],
3.0 (2.7) [1 Oyr].
WAIS
Age 28-30
yr
-1.1
(-2.29, 0.06)d
November 2012
5-63
Draft - Do Not Cite or Quote
-------
Study
Dietrich et al.
(1993b)
Schnaas et al.
(2006)
Cooney et al.
(1991)
Wasserman et
al. (1997)
long et al.
(1996)
Kordaset al.
(2011)
Study Population and Methodological Details
253 children followed from birth (1979-1985) to age
6. Syr, Cincinnati, OH
Recruitment at prenatal clinic. High follow-up
participation. Participants had slightly higher age 1 yr
blood Pb levels. Primarily African-American. Linear
regression model adjusted for HOME score, maternal
IQ and prenatal cigarette smoking, child birth weight,
birth length, and sex. Also considered potential
confounding by perinatal complications, prenatal
maternal substance abuse, nutritional status.
150 children followed from birth (1987-1992) to age
10 yr, Mexico City, Mexico.
Recruitment at prenatal clinic. Low follow-up
participation. Participants had higher SES, FSIQ,
higher blood Pb level before age 5 yr, lower at older
ages. Log linear mixed effects model adjusted for
SES, maternal IQ, HOME score, child sex, birth
weight, and postnatal blood Pb, indicator of first
FSIQ, random slope for subject. Most covariates
assessed in pregnancy or within age 6 mo.
175 children followed from birth (1983) to age 7 yr,
Sydney, Australia
Recruitment at birth hospital. Moderate follow-up
participation but no selective attrition. 100% white.
Linear regression adjusted for maternal education
and IQ, paternal education and occupation, HOME
score, child gestational age.
258 children followed prenatally (1984-1985) to age
7 yr, Kosovo, Yugoslavia
50% subjects live near Pb sources. Low follow-up
participation. Participants had lower HOME score,
maternal IQ, higher early childhood blood Pb levels
and fewer subjects lived in town with Pb sources.
Generalized estimating equations with log blood Pb
adjusted for maternal age, education, and IQ, child
age, sex, sibship size, and birth weight, language
spoken in home, HOME score.
375 children followed from birth (1979-1982) to age
1 1-13 yr, Port Pirie, Australia
Residence near Pb smelter. Moderate follow-up
participation. Participants had higher parental
occupational prestige. Regression model adjusted for
maternal IQ and age, parental occupational prestige,
smoking, marital status, and education, HOME score,
family functioning score, family size, child sex, age,
school grade, birth weight, birth order, feeding
method, breastfeeding duration, life events,
prolonged absences from school. Also considered
potential confounding by maternal psychopathology,
child Fe status, medication use in previous 2 weeks,
length of residence in area.
186 children followed from birth (1994-1995) to age
4 yr, Mexico City, Mexico
Recruitment at prenatal clinic. Low follow-up
participation but no selective attrition. Linear
regression model adjusted for maternal
age, education, IQ, smoking status, and marital
status, crowding in home, type of floor in home, child
sex, birth weight, gestational age. Did not consider
potential confounding by parental caregiving quality.
Blood Pb Data
(ug/dL)
Concurrent NR
Age 5 yr
Mean (SD):
11.8(6.3)
Interval analyzed:
5.5 (10th
percentile)-10
Prenatal (maternal
28-36 weeks)
Geometric mean
(5th-95th): 7.8
(2.5-24.5)
Interval analyzed:
3.2 (10th
percentile)-10
Age 3-5 yr avg:
NR
Age 5 yr
Mean (Max):
8.3 (27)
Lifetime avg
(to age 7 yr)
Geometric mean:
16.2
Interval analyzed:
6.0 (10th
percentile)-10
Lifetime avg
(to age 11-1 Syr)
Geometric mean
(GSD): 14.0(1.2)
Interval analyzed:
12.7-18.1 =
10th-90th
percentiles
Prenatal (cord)
mean (SD):
6.6 (3.3),
Interval analyzed:
3.2-10.8 =
10th-90th
percentiles
Concurrent mean
(SD):
8.7 (4.4) ,
Interval analyzed:
4.2 (1 Oth
percentile)-10
FSIQ
Testing3
WISC-R
Age 6.5 yr
McCarthy
GCI
Ages
6-1 Oyr
WISC-R
Age 7 yr
WISC-III
Age 7 yr
WISC-R
Age
11-13yr
Prenatal
Concurrent
McCarthy
GCI
Age 4 yr
Effect
Estimate
(95% CIV3
-0.33
(-0.60, -0.06)
-1.05
(-1 .67, -0.43)
-0.07°
-0.48
(-0.68, -0.27)
-0.12
(-0.24, -0.003)
-0.20
(-0.79, 0.39)
-0.60
(-0.99, -0.21)
November 2012
5-64
Draft - Do Not Cite or Quote
-------
Study
Greene et al.
(1992)
Min et al.
(2009)
Study Population and Methodological Details
270 children followed from with age 4 yr 10 mo,
Cleveland, OH
Recruitment at birth hospital. High prevalence of
prenatal alcohol and drug exposure. High follow-up
participation. Participants tended to be Black,
exposed to marijuana. Log linear regression adjusted
for maternal IQ, weight, street drug use,
cigarettes/day, alcohol use, and age, parental
education, authoritarian scale, race, parity, gestation
duration, date of first prenatal visit, HOME score,
quality of physical environment peeling paint, home
cleanliness, pica behavior
267 children followed from birth (1994-1996) to age
11 yr, Cleveland, OH
Recruitment at birth hospital. 86% African American
with high prevalence of prenatal drug and alcohol
exposure. Moderate follow-up participation to age 4
yr, high retention to age 1 1 yr. Participation tended to
be African American and had married mothers. Linear
regression model adjusted for HOME score, head
circumference at birth (all ages), current caregiver
vocabulary score, maternal marital status, parity,
child sex (age 4 yr), maternal vocabulary score at
birth (age 9 and 1 1 yr), average prenatal cocaine use
(age 9 yr), prenatal 1 st trimester marijuana use (age
1 1 yr). Also considered potential confounding by
maternal education, Fe deficiency, maternal
psychological distress, and race.
Blood Pb Data
(ug/dL)
Earlier childhood
(age 2 yr)
Geometric mean
(GSD):
15.6(1.4)
Interval analyzed:
10.1-24.0 =
10th-90th
percentiles
Age 4 yr
Mean (range):
7.0 (1 .3-23.8)
Interval analyzed:
3.0 (10th
percentile) -10
FSIQ
Testing3
WPPSI
Age 4.8 yr
WPPSI,
age 4yr
(concurrent)
WISC-R,
age 9 yr
WISC-R,
age 11 yr
Effect
Estimate
(95% CIV3
-0.11
(-0.51,0.29)
-0.50
(-0.89, -0.11)
-0.41
(-0.78, -0.04)
-0.54
(-0.91, -0.17)
Cross-sectional Studies:
Fulton et al.
(1987)
Surkan et al.
(2007)
Kim et al.
(2009b)
Roy et al.
(2011)
501 children, ages 6-9 yr, Edinburgh, Scotland
Recruitment at schools. High participation rate,
representative of area population. Log linear
regression model adjusted for parental SES,
education, marital status, health, mental health,
cigarettes smoked, vocabulary and matrices test
scores, involvement, interest, communication, and
participation with child, family size and structure, child
age, sex, handedness, height, gestation length, birth
weight, medical history, absence from school, recent
school change, grade, and time of day of test, people
per room in home, car/phone ownership, consumer
goods ownership.
389 children, ages 6-10 years, Boston, MA,
Farmington, ME
Recruitment from trial of amalgam fillings. High
participation rate. Higher participation of white children
in Maine. Analysis of covariance adjusted for caregiver
IQ, child age, SES, race, and birth weight, Also
considered potential confounding by site, sex, birth
order, caregiver education and marital status,
parenting stress, and maternal utilization of prenatal
and annual health care but not parental caregiving
quality.
279 children (born 1996-1999) ages 8-1 1 yr, Seoul,
Seongnam, Ulsan, and Yeoncheon, Korea
Recruitment at schools. Moderate participation rate.
Log linear regression model adjusted for maternal
age, education and prenatal smoking status, paternal
education, yearly income, smoking exposure status
after birth, child age, sex, and birth weight. Did not
consider potential confounding by parental caregiving
quality or IQ.
71 7 children ages 3-7 yr, Chennai, India
Recruitment at schools. High participation rate. Log
linear model adjusted for mid-arm circumference, age,
sex, family income, parental education and IQ, family
size. Did not consider potential confounding by
parental caregiving quality.
Concurrent
Geometric mean
(range):
1 1 .5 (3.3- 34)
Interval analyzed:
5.6 (mean of 1 st
decile)-10
Concurrent
Group! : 1-2
Group 2: 3-4
GroupS: 5-10
Concurrent
Mean (SD):
1 .73 (0.80)
Interval analyzed:
n Q OR
u.y -z.o -
1 0th-90th
percentiles
Concurrent
Mean (SD):
1 1 .4 (5.4)
Interval analyzed:
5.8 (10th
percentile)-! 0
BASC
Age 6-9 yr
WISC-III
Age 6-1 0 yr
KEDI-WISC
Ages 8-1 1 yr
Blood Mn:
<1.4ug/dL
Blood Mn:
>1.4ug/dL
Binet-Kamat
Ages 3-7 yr
-0.22
(-0.37, -0.06)
Reference
-0.1 2 (-3.3, 3.1 )e
-6.0 (-10.7, -1.4)e
-2 4
(-6.0,1.1)
-3.2
(-6.1, -0.23)
-1.16
(-1 .94, -0.37)
November 2012
5-65
Draft - Do Not Cite or Quote
-------
Study
Chiodo et al.
(2007)
Chiodo et al.
(2004)
Study Population and Methodological Details
506 children (born, 1982-1984) age 7 yr, Detroit, Ml
area
Recruitment at prenatal clinic. 100% African -
American. High prevalence of prenatal drug exposure.
High follow-up participation. Linear regression model
adjusted for maternal concurrent psychopathology, IQ,
prenatal cigarettes/day, and prenatal use of marijuana,
SES, HOME score, caretaker education, number
children in home, child sex. Also considered potential
confounding by child age, caretaker marital status,
maternal age, custody, cocaine use, prenatal alcohol
use, concurrent alcohol/week, concurrent
cigarettes/day, and concurrent marijuana use.
237 children, age 7.5 yr, Detroit, Ml area
Recruitment at prenatal clinic. 100% African-
American. High prevalence of prenatal alcohol
exposure. High participation rate. Log linear
regression model adjusted for SES, maternal
education and vocabulary score, # children <18 yr,
HOME score, parity, family environment scale, child
sex. Also considered potential confounding by
prenatal alcohol, marijuana, smoking, or cocaine use,
crowding, child age and life stress, caregiver life
stress, conflict tactics.
Blood Pb Data FSIQ
(ug/dL) Testing3
Concurrent WISC-III
Mean(SD): Age 7 yr
5.0 (3.0)
Interval analyzed:
2.1-8.7 =
1 0th-90th
percentiles
Concurrent WISC-III
Mean(SD): Age 7.5 yr
5.4(3.3)
Interval analyzed:
2.3-9.5 =
1 0th-90th
percentiles
Effect
Estimate
ic\co/ /"*i\k
(95% Cl)
-0.19
(-0.30, -0.08)e'f
Standardized
regression
coefficient
-0.22
(-0.38, -0.05)e'f
Standardized
regression
coefficient
aWISC = Wechsler Intelligence Scale for Children, WPPSI = Wechsler Preschool and Primary Scale of Intelligence,
WAIS = Wechsler Adult Intelligence Scale, GCI = General Cognitive Index, BASC = British Ability Scales Combined,
KEDI = Korean Educational Development Institute
bEffect estimates are standardized to a 1 ug/dL increase in blood Pb level within the lowest range examined in study or 10th
percentile to 10 ug/dL For populations with 10th percentiles near or above 10 ug/dL, effect estimates were calculated for the
10th-90th percentile interval of blood Pb level. Effect estimates are assumed to be linear within the evaluated interval of blood Pb
level. The percentiles are estimated using various methods and are only approximate values.
""Sufficient data were not provided to calculate 95% Cl.
dResults not included in Figure 5-2 because FSIQ assessed in adults.
eResults not included in Figure 5-2 because blood Pb level analyzed as categorical variable or because standardized regression
coefficient reported.
f95% CIs were constructed using a standard error that was estimated for the reported p-value of 0.01.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
Across the cohort studies, blood Pb-associated FSIQ decrements were found in
populations with mean blood Pb levels 5-10 (ig/dL. In analyses restricted to children in
the lower range of the blood Pb distribution (e.g., peak <10 (ig/dL), associations were
observed in groups of children with mean blood Pb levels 3-4 (ig/dL (Bellinger. 2008;
Canfield. 2008; Hornung. 2008). The analysis of the Rochester cohort is particularly
informative for lower blood Pb levels of children (mean at age 5 years: 5.8 (ig/dL)
compared to other cohorts and the greater consideration for potential confounding by
factors such as sex, race, family income, maternal education, race, prenatal maternal
smoking, birth weight, maternal IQ, HOME score, and transferrin saturation which
indicates iron status and for providing unadjusted and covariate-adjusted results (Canfield
et al.. 2003a). At age 5 years, higher age 6-24 month average, peak, concurrent, and
lifetime average blood Pb levels (area under the curve calculation using repeat
measurements between age 6 months and 5 years) were associated with lower FSIQ, and
while effect estimates in the covariate-adjusted model were 40-45% smaller than
estimated in the unadjusted models, they remained statistically significant. A larger effect
was estimated for the 101 (59%) children whose peak blood Pb levels never exceeded
November 2012
5-66
Draft - Do Not Cite or Quote
-------
1 10 (ig/dL, i.e., -1.8 points (95% CI: -3.0, -0.60) per 1 (ig/dL in concurrent blood Pb level.
2 Similarly, Bellinger and Needleman (2003) estimated a larger effect in the subset of the
3 Boston cohort (n = 48, 32%) with peak blood Pb levels <10 (ig/dL, i.e., -1.6 points (95%
4 CI: -2.9, -0.2) per 1 (ig/dL increase in age 2-year blood Pb level. The mean blood Pb
5 levels in these subsets of children were 3.3 (Rochester) and 3.8 (ig/dL (Boston) closer to
6 that of current U.S. children compared with other prospective studies.
7 Analyses of the Cincinnati and Port Pirie, Australia cohort also indicated associations
8 between blood Pb level and FSIQ decrements with as extensive consideration for
9 potential confounding (Table 5-3) albeit in populations with higher blood Pb levels
10 (i.e., mean at age 5 years: 11.8 (ig/dL, lifetime average geometric mean: 14.0 (ig/dL)
11 (Tong et al.. 1996; Dietrich et al.. 1993b). In contrast with other studies, in the Cleveland
12 cohort, associations of blood Pb level (ages 2 and 3 years) and tooth Pb level with FSIQ
13 (ages 3 and 4.8 years), became attenuated or were too imprecise to be informative with
14 adjustment for a large number of potential confounding factors, including maternal
15 substance abuse, home cleanliness, and pica behavior, which were not considered widely
16 in other studies (Greene etal.. 1992; Ernhart et al.. 1988). HOME score accounted for a
17 large proportion of the variance in FSIQ and was the major factor accounting for the
18 attenuation of the effect estimates for Pb biomarkers. The association between tooth Pb
19 level and FSIQ at age 4.8 years was attenuated with additional adjustment for HOME
20 score but was estimated with similar precision (-3.0 points [95% CI: -6.4, 0.32] per
21 1 (ig/g increase in tooth Pb level) (Greene and Ernhart. 1993). The few weak or null
22 associations do not mitigate the otherwise strong evidence provided by other studies. The
23 Cleveland (Greene et al., 1992) and Sydney (Cooney et al., 1991) studies were not
24 outliers with respect to population mean blood Pb levels or the specific confounding
25 factors considered (Table 5-3). and the Cleveland cohort had high prevalence of maternal
26 prenatal substance abuse which may limit the representativeness of results. Further, the
27 blood Pb-FSIQ association in children was substantiated in a pooled analysis of seven
28 prospective studies by Lanphear et al. (2005). which included the Cleveland cohort, as
29 well as multiple meta-analyses that combined results across various prospective and
30 cross-sectional studies, including those from the Cleveland and Sydney cohorts (Pocock
31 et al.. 1994; Schwartz. 1994; Needleman and Gatsonis. 1990). The meta-analysis by
32 Schwartz (1994) demonstrated the robustness of evidence to potential publication bias.
33 The addition of eight hypothetical studies with a zero effect and with the average weight
34 of the eight published studies resulted in a 50% lower but still negative and precise
35 (p <0.001) blood Pb-FSIQ effect estimate.
36 The pooled analysis of seven prospective studies included individual-level data from
37 1,333 children ages 4.8-10 years of age with a median (5th-95th percentile) concurrent
38 blood Pb level of 9.7 (ig/dL (2.5-33.2 (ig/dL) (Lanphear et al.. 2005). In multivariate
November 2012 5-67 Draft - Do Not Cite or Quote
-------
1 models that adjusted for study site, maternal IQ, HOME score, birth weight, and maternal
2 education, higher concurrent, peak, lifetime average, and early childhood blood Pb levels
3 were associated with lower FSIQ measured at age 4.8-10 years. Various models were
4 investigated to characterize the shape of the blood Pb-FSIQ concentration-response
5 relationship. Consistent with the supralinear concentration-response relationship found in
6 several individual cohort studies, Lanphear et al. (2005) found that a nonlinear (i.e., log-
7 linear) model fit the data better than a linear model. The nonlinear relationship was
8 indicated further by observations of a greater decrease in FSIQ for a 1 (ig/dL increase in
9 concurrent blood Pb for the 244 (18%) children who had peak blood Pb levels <10 (ig/dL
10 (-0.80 points [95% CI: -1.74, -0.14]) and the 103 (8%) children with peak blood Pb levels
11 <7.5 (ig/dL (-2.9 points [95% CI: -5.2, -0.71]). Among children with peak blood Pb
12 <10 (ig/dL and <7.5 (ig/dL, the median concurrent blood Pb levels were 4.2 (ig/dL and
13 3.2 (ig/dL, respectively (Hornung. 2008).
14 An additional strength of the pooled analysis by Lanphear et al. (2005) was the
15 examination of several potential confounding factors related to SES and the caregiving
16 environment. Variables such as HOME score, birth weight, maternal IQ, and maternal
17 education were selected for inclusion in the final model with blood Pb level based on
18 their statistically significant association with FSIQ. Child sex, maternal prenatal tobacco
19 or alcohol use, maternal age at delivery, marital status, and birth order were not
20 statistically significantly associated with FSIQ and did not alter the effect estimate for
21 concurrent blood Pb level adjusted for the four aforementioned covariates. While a
22 smaller decrement in FSIQ was estimated for concurrent blood Pb level in the adjusted
23 model than in the unadjusted model (-4.7 points [95% CI: -5.7, -3.6] versus -2.7 points
24 [95% CI: -3.7, -1.7] per log increase in concurrent blood Pb level), the adjusted blood Pb
25 level effect estimate did not lose precision.
26 The few prospective studies published since the 2006 Pb AQCD continued to
27 demonstrate associations between higher blood Pb level and lower FSIQ, in some cases,
28 with additional follow-up of previous cohorts. Similar to studies reviewed in the
29 2006 Pb AQCD, most recent studies demonstrated associations between blood Pb level
30 and lower FSIQ in populations with mean blood Pb level between 5 to 10 (ig/dL. Jusko et
31 al. (2008) affirmed the findings in the Rochester cohort previously reported by Canfield
32 et al. (2003a), who examined the cohort at age 5 years. Jusko et al. (2008) examined 174
33 Rochester cohort subjects at age 6 years and similar to Canfield et al. (2003a), found that
34 an increase in peak blood Pb level was associated with a larger decrease in FSIQ among
35 children with peak blood Pb levels <10 (ig/dL than among children with peak blood Pb
36 levels 10-20 (ig/dL (-1.2 points versus -0.32 points per 1 (ig/dL increase in blood). The
37 age 6 year analysis had similarly extensive consideration for potential confounding as did
November 2012 5-68 Draft - Do Not Cite or Quote
-------
1 Canfield et al. (2003a) (Table 5-3) and also indicated associations with higher concurrent,
2 infancy average, and lifetime average blood Pb levels (effect estimates not reported).
3 Additional evidence was provided for children in Mexico City, albeit in a separate cohort
4 of children born later with lower blood Pb levels at corresponding ages. Among 150
5 children born 1987-1992, Schnaas et al. (2006) previously reported larger Pb-associated
6 decrements in FSIQ for prenatal maternal (28-36 weeks) blood Pb levels than for child
7 concurrent blood Pb levels between ages 1 and 10 years. In contrast, Kordas et al. (2011)
8 found that an increase in concurrent blood Pb level was associated with a larger
9 decrement in FSIQ at age 4 years than was an increase in cord blood Pb level with
10 adjustment for several potential confounding factors (HOME score not examined) (Table
11 5-3). The 186 children in the latter study were born 1994-1995 and at age 4 years had a
12 mean blood Pb level of 8.7 (ig/dL. In Schnaas et al. (2006). the geometric mean blood Pb
13 level at age 4 years was 10.3 (ig/dL. It is not clear whether different temporal patterns of
14 Pb exposure or age of FSIQ assessment may have contributed to the contrasting
15 associations for prenatal and concurrent blood Pb levels in these two Mexico City
16 cohorts.
17 Mazumdar et al. (2011) followed the Boston cohort (Bellinger etal.. 1992) to age
18 28-30 years, and indicated that the effect of childhood Pb exposures may persist to
19 adulthood. Only 43 of the original 249 subjects enrolled at birth were examined at age
20 28-30 years, but they did not differ from the original cohort in demographic
21 characteristics or blood Pb history. Higher blood Pb levels measured at age 6 months,
22 4 years, 10 years, and averaged over childhood (to age 10 years) (means: 3 (ig/dL at age
23 10 years to 8 (ig/dL at age 6 months) were associated with lower FSIQ in adults with
24 adjustment for sex, birth weight, birth order, gestational age, maternal marital status,
25 maternal education, maternal IQ, race, maternal smoking and alcohol use in pregnancy,
26 average of childhood HOME score, concussion, and subject current smoking status. The
27 effect estimates were similar in magnitude for all childhood blood Pb measures, except
28 for age 6 month blood Pb level, which was associated with a smaller FSIQ decrement.
29 Min et al. (2009) found higher earlier childhood blood Pb levels (age 4 year) to be
30 associated with decrements in FSIQ in another cohort of children in Cleveland, OH
31 between ages 4 and 11 years, indicating persistence of effects (Figure 5-2 and Table 5-3).
32 However, similar to the other Cleveland cohort (Greene etal.. 1992). the recent cohort
33 had high prevalence of prenatal alcohol and drug exposure. These exposures were weakly
34 associated with FSIQ or did not influence the blood Pb-FSIQ association, indicating lack
35 of strong confounding bias. However, because the population lacks representativeness,
36 these findings are less of a consideration in drawing conclusions regarding the effects of
37 Pb exposure on FSIQ of children.
November 2012 5-69 Draft - Do Not Cite or Quote
-------
1 An important consideration in the evaluation of epidemiologic evidence is the precision
2 of effect estimates, both within and among studies. There was variability in precision
3 among studies, which appeared to be influenced by sample size rather than the age of
4 subjects or the extent of adjustment for potential confounding factors. Analyses of the
5 Port Pirie (n = 375, ages 11-13 years) and Yugoslavia (n = 258, age 7 years) cohorts
6 ("Wasserman et al.. 1997; Tong et al.. 1996) and the pooled analysis of 1,333 children
7 (Lanphear et al., 2005) estimated more precise effects compared to the Boston (n = 148,
8 age 10 years) and Rochester cohorts (n = 172, age 5 years) (Canfield et al.. 2003a:
9 Bellinger et al.. 1992). Analyses of the Yugoslavia and Port Pirie cohorts did not
10 necessarily have more or less extensive adjustment for potential confounding.
11 Among prospective studies, a wide range of blood Pb-associated FSIQ decrements was
12 estimated (Figure 5-2 and Table 5-3). This wide range is not unexpected, given
13 differences among studies in blood Pb level ranges, model specification (linear versus log
14 linear), lifestage or time period of blood Pb level examined, and distribution of potential
15 confounding factors. The pooled analysis examined study populations of diverse SES,
16 maternal education, and cultural backgrounds with the same model and indicated
17 precision of effect (Lanphear et al.. 2005). A series of sensitivity analyses, in which one
18 cohort was excluded at a time, revealed that no single study was responsible for the
19 results. Per log increase in blood Pb level, effect estimates excluding one study at a time
20 fell within a narrow range, -2.36 to -2.94 (blood Pb level ranges for sensitivity analyses
21 not reported). Precision of effect also was indicated by the similar effect estimates found
22 with similar model specifications, population blood Pb levels, and sample sizes in the
23 Boston and Rochester cohorts (Table 5-3). but very different SES and racial distributions
24 of the cohorts, and different ages of blood Pb level and FSIQ examined (Canfield et al..
25 2003a; Bellinger etal.. 1992). These estimates were larger than those found in the
26 Cincinnati, Port Pirie, and Yugoslavia cohorts but were based on similar extent of
27 adjustment for potential confounding factors. Several of the smaller blood Pb-associated
28 FSIQ decrements were based on log-linear models that estimated effects at higher blood
29 Pb levels (10th percentiles >5.5 versus 2 (ig/dL). The widely different null associations
30 found in the Cleveland cohort have weaker implications because of the
31 nonrepresentativeness of the cohort due to their high prevalence of prenatal alcohol and
32 drug exposure (Greene etal.. 1992; Ernhart et al.. 1988).
November 2012 5-70 Draft - Do Not Cite or Quote
-------
Evidence from Cross-sectional Studies
1 The smaller body of cross-sectional studies reviewed in the 2006 Pb AQCD (U.S. EPA.
2 2006b) found associations of higher concurrent blood (Fulton et al.. 1987) or tooth
3 (Needleman et al., 1979) Pb levels with FSIQ decrements in children ages 6-9 years, and
4 associations also were found in the few recent studies in children ages 3-11 years (Figure
5 5-2 and Table 5-3). Several cross-sectional studies had larger sample sizes (n = 279-717)
6 than the prospective studies and produced effect estimates with similar precision.
7 Previous meta-analyses produced similar combined blood Pb-FSIQ effect estimates for
8 prospective and cross-sectional studies (Pocock etal. 1994; Schwartz. 1994). However,
9 in this ISA, the cross-sectional findings were given less weight in conclusions regarding
10 Pb-associated effects on cognitive function. The temporal sequence between Pb exposure
11 and decreases in FSIQ is difficult to establish. Some studies had population-based
12 recruitment, high participation rates, and did not indicate undue selection bias, but the
13 evidence overall had less consideration for potential confounding by parental caregiving
14 quality and/or parental IQ (Roy etal.. 2011: Kim et al.. 2009b: Zailina et al.. 2008:
15 Surkan et al., 2007: Needleman et al., 1979). The meta-analysis by Pocock et al. (1994)
16 noted the lack of adequate control for potential confounding factors in previous cross-
17 sectional studies. Other cross-sectional studies lacked representative study populations
18 because of high prevalence of prenatal alcohol (Chiodo et al.. 2004) or drug exposure
19 (Chiodo et al.. 2007).
20 Among the cross-sectional studies, Fulton et al. (1987) had more extensive consideration
21 for potential confounding. Among 501 children, ages 6-9 years, in Edinburgh, Scotland, a
22 1 (ig/dL increase in concurrent blood Pb level in the interval between 5.6 and 10 (ig/dL
23 was associated with a 0.22-point decrease (-0.37, -0.06) in FSIQ, after adjustment for
24 several factors related to SES, parental health and mental health, child health, and
25 parental caregiving quality (Table 5-3). The effect estimate from this study was among
26 the smallest produced by cross-sectional studies. The study population was representative
27 of the source population but had much higher blood Pb levels (geometric mean:
28 11.4 (ig/dL) than those of most of the current U.S. population of children.
29 Recent cross-sectional studies examined potential confounding by parental IQ and
30 education and SES but not parental caregiving quality. Studies that examined populations
31 with mean concurrent blood Pb levels <4 (ig/dL did not conclusively indicate
32 associations with FSIQ decrements at lower blood Pb levels. Among 389 children from
33 urban Boston, Massachusetts and rural Farmington, Maine with mean concurrent blood
34 Pb level 2.2 (ig/dL, lower FSIQ was limited to children with blood Pb levels 5-10 (ig/dL,
35 with children with blood Pb levels 1-2 (ig/dL serving as the referent group (Table 5-3)
36 (Surkan et al., 2007). There was consideration for potential confounding by several
37 factors, including age, race/ethnicity, birth weight, SES, primary caregiver IQ, SES,
November 2012 5-71 Draft - Do Not Cite or Quote
-------
1 education and marital status, parenting stress, and maternal utilization of prenatal or
2 annual health care. Other recent cross-sectional studies found associations at lower
3 concurrent blood Pb levels; however, some of the children likely had higher earlier
4 childhood Pb exposures, which may have contributed to the associations observed with
5 relatively low concurrent blood Pb levels. In a group of 279 children ages 8-11 years
6 from four Korean cities with a mean concurrent blood Pb level 1.73 (ig/dL, Kim et al.
7 (2009b) found an association between higher concurrent blood Pb level and lower FSIQ
8 with adjustment for parental education, yearly income, prenatal and postnatal smoking
9 exposure, birth weight, age, and sex. The adjusted effect estimate was attenuated but
10 similarly precise as the unadjusted estimate. The concurrent blood Pb-FSIQ and verbal
11 IQ relationship was modified by concurrent blood manganese (Mn) levels. Blood Pb and
12 Mn levels were not correlated (r = 0.03, p = 0.64). Higher concurrent blood Pb level was
13 associated with lower FSIQ in both children in high Mn (above the median of 1.4 (ig/dL)
14 and low Mn group (below the median of 1.4 (ig/dL) but a larger FSIQ decrement in the
15 130 children in the high Mn group (-3.2 points [95% CI: -6.1, -0.23] per 1 (ig/dL increase
16 in the 10th to 90th percentile interval 0.9-2.8 (ig/dL) compared with children in the low
17 Mn group (-2.4 points [95% CI: -6.0, 1.1]). The biological plausibility for the Pb-Mn
18 interaction is provided by observations that Mn has similar modes of action and cellular
19 targets as does Pb, i.e., altering Ca2+ metabolism, inducing oxidative damage in neuronal
20 cells, diminishing dopamine transmission. Among 169 children in Malaysia, ages 6-8
21 years, concurrent blood Pb (mean ~4 (ig/dL) level but not parental education or family
22 income was associated with FSIQ decrements, producing uncertainty as to whether these
23 potential confounding factors were measured adequately (Zailinaet al.. 2008).
24 Other recent cross-sectional studies found associations in populations of children with
25 relatively higher concurrent blood Pb levels (means >8 (ig/dL). Evidence demonstrates
26 that Pb affects dopaminergic neurons and dopamine release (Section 5.3.11.8). Further,
27 dopaminergic activity is a key mediator of cognitive function. These findings suggest that
28 variants in dopamine-related genes may modify Pb-associated effects on cognition.
29 Epidemiologic evidence for effect modification is not consistent; however, subgroup
30 analyses are subject to higher probability of findings by chance. The larger of these
31 studies (n = 717 children ages 3-7 years in Chennai, India) with higher concurrent blood
32 Pb levels (mean: 11.4 (ig/dL) found that a 1 (ig/dL higher blood Pb level was associated
33 with a larger decrease in FSIQ among the 72 children with the Taq A1/A1 dopamine
34 receptor (DRD2) variant (-2.5 points [95% CI: -5.0, -0.04] within the blood Pb level
35 interval 5.8-10 (ig/dL) than among the 651 children with the Taq A2/A2 variant (-1.1
36 points [-2.3, -0.12]) (Roy et al.. 2011). Kordas et al. (2011) did not find effect
37 modification in a smaller study of 186 children in Mexico City with a mean concurrent
38 blood Pb level of 8.7 (ig/dL. Another difference between studies that may have
39 contributed to the difference in effect modification by the DRD2 variant was an
November 2012 5-72 Draft - Do Not Cite or Quote
-------
1 association observed between Taq A1/A1 genotype and higher mean FSIQ score in the
2 group in Mexico but no association in the group in India.
3 In summary, a majority of prospective and cross-sectional studies demonstrated
4 associations between higher blood Pb level and lower FSIQ in children ages 3-17 years
5 (e.g., Figure 5-2 and Table 5-3). While studies performed numerous tests, bias due to
6 increased probability of findings by chance was unlikely because most studies found a
7 consistent pattern of association across the ages of blood Pb level and FSIQ analyzed.
8 Across studies, FSIQ was measured with similar instruments scored on similar scales
9 with similar measurement error. The key supporting evidence is provided by the
10 prospective studies, which better indicated the temporal sequence between blood Pb
11 levels measured earlier in childhood or averaged over multiple years or tooth Pb levels
12 and FSIQ measured later in childhood. Prospective studies also had more extensive
13 consideration for potential confounding by maternal IQ and education, SES, birth weight,
14 smoking exposure, parental caregiving quality, and in a few cases, other birth outcomes
15 and nutritional factors. Further, the representativeness of findings was supported by
16 associations found in diverse populations (e.g., Boston, MA; Cincinnati, OH; Rochester,
17 NY; Cleveland, OH; Mexico City, Mexico; Port Pirie and Sydney, Australia; and
18 Kosovo, Yugoslavia) and in studies examining populations recruited from prenatal
19 clinics, hospital maternity departments, or schools with high follow-up participation and
20 lack of biased follow-up participation by blood Pb level and FSIQ. The few weak or null
21 associations found in Cleveland and Sydney cohorts were adjusted for similar potential
22 confounding factors and thus do not mitigate the otherwise strong evidence. The blood
23 Pb-FSIQ association in children was substantiated in a pooled analysis of seven
24 prospective studies by Lanphear et al. (2005) as well as multiple meta-analyses that
25 combined results across various prospective and cross-sectional studies (Pocock et al..
26 1994: Schwartz. 1994: Needleman and Gatsonis. 1990). with Schwartz (1994)
27 demonstrating the robustness of evidence to potential publication bias.
28 Across the prospective studies, blood Pb-associated FSIQ decrements were found with
29 concurrent, prenatal (maternal or cord), earlier childhood, multiple year average, or
30 lifetime average blood Pb levels. Associations also were found with tooth Pb levels.
31 There is no clear indication of a stronger association of FSIQ with blood Pb level
32 measured at a particular lifestage or time period. Concurrent blood Pb level in children
33 reflects recent and past Pb exposures. Thus, several observations point to an effect of
34 cumulative childhood Pb exposure. Blood Pb-associated FSIQ decrements were found in
35 populations with mean blood Pb levels 5-10 (ig/dL. A common finding was a supralinear
36 concentration-response relationship, i.e., a larger decrement in cognitive function per unit
37 increase in blood Pb level in children in the lower range of the population blood Pb level
38 distribution. In analyses restricted to children in the lower range of blood Pb levels
November 2012 5-73 Draft - Do Not Cite or Quote
-------
1 (e.g., peak <10 (ig/dL), associations were found in groups of children with mean blood
2 Pb levels 3-4 (ig/dL (Bellinger. 2008: Canfield. 2008: Hornung. 2008). Precision of effect
3 estimates was demonstrated in the pooled analysis by the narrow range of estimates,
4 -2.36 to -2.94 points per log increase in blood Pb level, obtained by excluding one study
5 at a time (Lanphear et al., 2005). Across individual studies, there was a wide range of
6 effect estimates reported for blood Pb-associated FSIQ decrements. However, there was
7 variability in model specification and blood Pb level ranges examined among studies
8 (Figure 5-2 and Table 5-3). Similarly larger effect estimates were found in the Boston
9 and Rochester cohorts, which differed in racial and SES distributions. Although these
10 studies had smaller sample sizes, they had at least as extensive consideration for potential
11 confounding as other studies (Canfield et al.. 2003a: Bellinger et al.. 1992). Each study
12 estimated larger effects for children whose peak blood Pb levels never exceeded
13 10 (ig/dL, -1.8 points (95% CI: -3.0, -0.60) per 1 (ig/dL increase in concurrent blood Pb
14 level in the Rochester cohort (Canfield et al.. 2003a) and -1.6 points (95% CI: -2.9, -0.2)
15 per 1 (ig/dL increase in age 2-year blood Pb level in the Boston cohort (Bellinger and
16 Needleman. 2003). These subsets of children had mean blood Pb levels of 3.3
17 (Rochester) and 3.8 (ig/dL (Boston), lower than those examined in other prospective
18 studies.
5.3.2.2 Bayley Scales of Infant Development
19 The Mental Development Index (MDI) of the Bayley Scales of Infant Development is a
20 widely used test of infant mental development. Black et al. [(2004) and Pollit (2005)]
21 asserted that the MDI is a reliable indicator of current development and cognitive
22 functioning of the infant, integrating cognitive skills such as sensory/perceptual acuities,
23 discriminations, and response; acquisition of object constancy; memory learning and
24 problem solving; vocalization and beginning of verbal communication; and basis of
25 abstract thinking. However, the MDI test is not an intelligence test, and MDI scores,
26 particularly before ages 2-3 years, are not necessarily correlated with later measurements
27 of FSIQ in children with normal development. In the review of the MDI evidence,
28 emphasis was placed on examinations at ages 2-3 years, which have test items more
29 similar to those in school-age IQ tests. Most of the prospective studies reviewed in the
30 2006 Pb AQCD (U.S. EPA. 2006b) found associations of prenatal, earlier infancy, and
31 concurrent blood Pb level with MDI score in children between ages 2 and 3 years, and
32 recent studies examined and found associations with cord blood Pb level (Table 5-4).
November 2012 5-74 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
The prospective studies found blood Pb-associated decrements in MDI in some large
(n = 146-592) populations with mean blood Pb levels 5-10 (ig/dL. Recruitment of
participants before or at birth without consideration of Pb exposure or maternal IQ, high
to moderate follow-up participation, and nonselective loss-to-follow-up in most studies
increase confidence that the observed associations are not due to selection bias.
Comparisons of blood Pb levels measured at various lifestages did not clearly indicate a
stronger effect on MDI of prenatal or postnatal childhood blood Pb levels. While the
prospective studies adjusted for birth outcomes and maternal IQ and education, most did
not adjust for other SES indicators or parental caregiving quality. Concurrent and cord
blood Pb levels were associated with MDI, with additional adjustment for SES and
HOME score in the Boston cohort and HOME score in the Yugoslavia cohort
(Wasserman et al., 1992; Bellinger etal.. 1987). In the Cleveland cohort, associations of
cord, age 6 month, and concurrent blood Pb levels with MDI at age 2 years became null
after adjusting for covariates including HOME score (Ernhart et al., 1988; Ernhart et al.,
1987). However, 50% of the cohort was born to alcoholic mothers and may be less
representative of the general U.S. population of children.
Table 5-4 Associations of blood Pb level with Bayley MDI in children ages
12 months to 3 years.
Study3
Study Population and
Methodological Details
(Presented in order of strength of study Blood Pb
design and consideration for potential Timing and
confounding)3 Levels (ug/dL)
MDI Assessment
Effect Estimate
(95% Cl)b
Bellinger et al. 249 children followed from birth
(1987) (1979-1981) to age 2 yr, Boston, MA.
Prospective. Recruitment from birth
hospital. High follow-up participation.
Participants had higher cord blood Pb
level, SES, HOME score, maternal
education and IQ, lower maternal age,
were nonwhite. Regression model
adjusted for maternal age, race, maternal
IQ, maternal education, years of smoking,
alcohol drinks/week in 3rd trimester, SES,
HOME score, sex, birth weight,
gestational age, birth order.
Prenatal (cord)
Low: <3
Medium: 6-7
High:> 10
Adjusted mean at age
2yr
High vs. low cord
blood
High vs. medium
cord blood
-4.8 (-7.3, -2.3)
-3.8 (-6.3, -1.3)
Jedrychowski et 444 children born 2001 -2004 followed
al. (2009b) prenatally to age 3 yr, Krakow, Poland.
Prospective cohort examining multiple
exposures and outcomes. Recruitment
from prenatal clinic. High follow-up
participation. Log linear regression model
adjusted for maternal education, birth
order, prenatal smoking, sex, and within-
subject MDI correlation. Did not consider
potential confounding by HOME score.
Prenatal (cord)
Geometric mean
(range): 1.29
(0.44-5)
Interval
analyzed:!.2-1.3 =
10th-90th
percentiles
Age 2 yr
Age 3 yr
-2.6 (-5.0,-0.21)
-2.3 (-4.3, -0.30)
November 2012
5-75
Draft - Do Not Cite or Quote
-------
Study3
Tellez-Rojo et
al. (2006)
Tellez-Rojo
(2008)
Claus Henn et
al. (2012)
Hu et al. (2006)
Pilsner et al.
(2010)
Surkan et al.
(2008)
Study Population and
Methodological Details
(Presented in order of strength of study
design and consideration for potential
confounding)3
1 93 children born 1 997-1 999 followed
prenatally to age 2 yr, Mexico City, Mexico
Cross-sectional. Recruitment from
prenatal clinic or birth hospital.
Participants had older, more educated
mothers, lower cord blood Pb level, and
slightly higher MDI. Log linear regression
model adjusted for sex, maternal age,
birth weight, maternal IQ, cohort.
Considered potential confounding by other
unspecified factors.
455 children born 1997-2000 followed
prenatally to age 3 yr, Mexico City, Mexico
Prospective, same cohort as above. No
selective participation of subjects. Linear
mixed effects regression adjusted for sex,
hemoglobin, gestational age, maternal IQ,
maternal education, blood Pb-blood Mn
interaction. Did not consider potential
confounding by HOME score.
1 46 children born 1 997-1 999 followed
prenatally to age 2 yr, Mexico City, Mexico
Prospective. Recruitment from prenatal
clinic. Moderate follow-up participation.
Eligible similar to non-eligible. Log linear
regression model adjusted for sex,
maternal age, current weight, height-for-
age Z score, maternal IQ, concurrent
blood Pb (in models examining blood Pb
at other lifestages). Considered potential
confounding by other unspecified factors.
255 children born 1994-1995 followed
prenatally to age 2 yr, Mexico City,
Mexico.
Prospective. Recruitment from birth
hospital. Low but not selective
participation. Linear regression model
adjusted for maternal age, maternal IQ,
marital status, parity, gestational age,
inadequate folate intake, MTHFR
genotype. Did not consider potential
confounding by HOME score.
309 children born 1991-2004 followed
from birth to age 3 yr, Mexico City,
Mexico.
Cross-sectional. Recruitment from birth
hospital. High participation rate. Linear
mixed effects model adjusted for sex,
maternal age, IQ, education, and self-
esteem, parity, grams/day alcohol,
smoking status, cohort. Did not consider
potential confounding by HOME score.
Blood Pb
Timing and
Levels (ug/dL)
Concurrent
Geometric mean:
2.9
Interval analyzed:
0.8-4.9 = range
Age 1 yr
Mean (SD): 5.1
(2.6)
Interval analyzed:
2.5-8.4 =10th-90t
h percentiles
Prenatal maternal
1st trimester
Mean (SD): 7.1
(5 1)
W- ' /
Prenatal maternal
3rd trimester
Mean (SD): 6.9
I A ^>\
(4.Z)
Prenatal cord
blood
Mean (SD): 6.2
(3.9)
Concurrent
Mean (SD): 4.8
(3.7)
Prenatal (cord)
Mean (SD):
6.7 (3.6)
Interval analyzed:
3.5-10.5 =
1 0th-90th
percentiles
Concurrent
Mean (SD): 6.4
(4.3)
Interval analyzed:
2.0 (10th
percentile)-10
MDI Assessment
Age 2 yr
Ages 1 to 3 yr
Blood Mn:
<2.0 ug/dL
Blood Mn:
2.0-2.8 ug/dL
Blood Mn >2.8 ug/dL
Age 2 yr
Prenatal 1st trimester
Interval analyzed: 2.5
(10thpercentile)-10)
Prenatal 3rd trimester
Interval analyzed: 2.8
(10thpercentile)- 10
Prenatal Cord blood
Interval analyzed: 2.5
(10th percentile)- 10
Concurrent
Interval analyzed:
1.6-9.1 = 10th-90th
percentiles
Age 2 yr
Ages 1 to 3 yr
All subjects
High maternal self-
esteem
Low maternal self-
esteem
Effect Estimate
/oco/ ^i\k
(95% Cl)
-1 .71 (-3.0, -0.42)
-3.0 (-5.22, -0.78)
-0.07 (-0.39, 0.25)
-2.2 (0, 4.44)
-0.91 (-1.8, -0.04)
-0.49 (-1.3, 0.31)
-0.07 (-0.93, 0.79)
-0.23 (-0.92, 0.45)
-0.73 (-1 .2, -0.23)
-0.1 8 (-0.45, 0.09)
0.36 (-0.50, 1.2)
-0.31 (-0.60, -0.02)
November 2012
5-76
Draft - Do Not Cite or Quote
-------
Study3
Vimpani et al.
(1985)
Wasserman et
al. (1992)
Solon et al.
(2008)
Ernhart et al.
(1988: 1987)
Study Population and
Methodological Details
(Presented in order of strength of study
design and consideration for potential
confounding)3
592 children followed prenatally to age 2
yr, Port Pirie, Australia.
Prospective. Residence near Pb smelter.
High baseline participation rate. Linear
regression model adjusted for maternal
age, education, IQ, workplace, and
prenatal marital status, paternal education
and workplace, parental relationship, child
birth rank, mouthing activity, oxygen use
at birth, apgar score, neonatal jaundice,
size for gestational age. Did not consider
potential confounding by HOME score.
392 children followed prenatally (from
1985) to age 2 yr, Kosovska Mitrovica and
Pristina, Yugoslavia.
Prospective. 53% live near Pb sources.
High follow-up participation, no selective
attrition. Log linear regression model
adjusted for sex, birth order, birth weight,
ethnic group, HOME, maternal education,
maternal age, maternal IQ.
502 children born 1997-2004, Visayas,
Philippines.
Cross-sectional. Census based
recruitment. No selective participation of
subjects. Two-stage linear regression
model to account for determinants of
blood Pb (sex, roof materials, water
source, breastfed for > 4 months) and
cognitive function (HOME score, maternal
education, maternal smoking, born
premature, region of residence).
359 children, followed prenatally to age 2
yr, Cleveland, OH
Prospective. Recruitment at birth
hospital. High follow-up participation, more
white, higher IQ, nonalcoholic mothers not
followed. 50% born to alcoholic mothers.
Linear regression adjusted for age, race,
sex, birth order, parental education,
maternal IQ, Authoritarian Family
Ideology, HOME.
Blood Pb
Timing and
Levels (ug/dL)
20% subjects had
age 2 yr blood Pb
levels >30
Concurrent
Means:
35.5 (K. Mitrovica)
8.4 (Pristina)
Concurrent
Mean (SD):
7.1 (7.7)
Interval analyzed:
1.6 (10th
percentile) -10
Means (SD):
Prenatal cord:
6.0(2.1)
Age 6 mo:
10.1 (3.3)
Concurrent:
16.7(6.5)
MDI Assessment
Age 2 yr
Maternal avg prenatal
blood Pb
Cord blood Pb
Age 6 mo blood Pb
Age 2 yr blood Pb
Lifetime (to age 2 yr)
ava
** • a
Age 2 yr
Cord blood Pb
6 mo blood Pb
12 mo blood Pb
18 mo blood Pb
24 mo blood Pb
Ages 6 mo to 3 yr
Age 2 yr
Prenatal cord
Age 6 mo blood Pb
Concurrent blood Pb
Effect Estimate
/oco/ ^i\k
(95% Cl)
-0.64
0.03
-0.40, p <0.05
-0.06
-0.31, p<0.05
Per three-fold
increase in
blood Pb
-1.7, p = 0.12
-1.1, p = 0.34
-1.7, p = 0.17
-1.8, p = 0.16
-2.5, p = 0.03
-1 .07 for
population mean
serum folate of
225 ug/dL, 95%
Cl: not available
Variance
estimates
0.0003, t = -0.21
0.00, p = 0.95
0.00, p = 0.95
MDI = Mental Development Index, MTHFR = methylenetetrahydrofolate reductase
aStudies are presented in order of strength of study design and consideration for potential confounding. All Mexico City studies
were kept together.
bExcept where noted, effect estimates are standardized to a 1 ug/dL increase in blood Pb level for the lowest blood Pb range
examined in the study or for blood Pb level up to 10 ug/dL.
1
2
o
J
4
5
6
7
A large study of 444 children in Krakow, Poland, found cord blood Pb-associated
decrements in MDI at ages 2 and 3 years with lower cord blood Pb levels (median
1.23 (ig/dL, 5th-95th: 1.24-1.34 (ig/dL) than examined in other studies (Jedrychowski et
al.. 2009b) (Table 5-4). However, cord blood Pb levels reflect the pregnancy blood Pb
levels of mothers. Evidence indicates increased mobilization of Pb from bone to blood in
pregnant women (Sections 4.2.2.4 and 4.3.5.2). Thus, there is uncertainty regarding the
Pb exposure scenarios that contribute to associations between cord blood Pb level and
MDI in children. Jedrychowski et al. (2009b) estimated a larger decrease in MDI per unit
November 2012
5-77
Draft - Do Not Cite or Quote
-------
1 increase in cord blood Pb level among the 233 males than among the 223 females. Other
2 observations have indicated increased susceptibility of the developing male central
3 nervous system (CNS) to environmental insults (Moffitt et al., 2001). Although median
4 cord blood Pb levels were similar in males (1.35 (ig/dL) and females (1.41 (ig/dL), the
5 mean age 3-year MDI score was slightly lower among males than among females (101
6 and 105, respectively).
7 Multiple studies in various Mexico City cohorts reported associations of prenatal
8 (maternal or cord) or child postnatal blood Pb levels with decrements in MDI in children
9 between ages 1 and 3 years (Claus Henn et al.. 2012; Pilsner et al. 2010; Surkan et al..
10 2008: Hu et al.. 2006: Tellez-Roio et al.. 2006). Hu et al. (2006) compared associations
11 among prenatal maternal blood Pb levels measured at different trimesters among 146
12 children at age 2 years. Increases in first trimester maternal blood Pb levels (whole blood
13 or plasma) were associated with larger decreases in MDI scores than increased in
14 maternal third trimester, cord, or child concurrent blood Pb levels (Table 5-4). These
15 results were adjusted for sex, 2-year blood Pb level, height-for-age Z score, weight,
16 maternal age, and maternal IQ. Model covariates did not include SES, maternal
17 education, or HOME score; however, a larger list of unspecified potential confounding
18 factors was considered.
19 Consistent with several findings for FSIQ, Tellez-Rojo et al. (2006) found larger effect
20 estimates in children with lower blood Pb levels. In linear models, a 1 (ig/dL increase in
21 concurrent blood Pb level was associated with a -1.71 point (95% CI: -3.0, -0.42)
22 decrease in MDI among 193 children with concurrent blood Pb levels <5 (ig/dL and a
23 -1.0 point (95% CI: -1.8, -0.26) decrease among 294 children with concurrent blood Pb
24 levels <10 (ig/dL. In a follow-up of the same cohort to age 3 years, Claus Henn et al.
25 (2012) found inconsistent interactions between blood Mn and Pb levels. Investigators
26 selected mid-range (2.0-2.8 (ig/dL) blood Mn levels as the reference group based on
27 previous observations that MDI scores were least affected by increases in blood Mn level
28 in this group. Larger blood Pb-associated MDI decrements were found in the 91 children
29 each with blood Mn levels <2.0 (ig/dL and >2.8 (ig/dL with age 1 year blood Pb level but
30 not age 2 year blood Pb level. Adjustment for sex, gestational age, hemoglobin, maternal
31 IQ, maternal education, and visit produced more negative effect estimates. Kim et al.
32 (2009b) also found effect modification by blood Mn levels for the association between
33 blood Pb level and FSIQ, but in older children ages 8-11 years (Section 5.3.2.1).
34 Other recent studies in Mexico City examined effect modification by maternal self-
35 esteem, genetic variants, and nutritional status. Surkan et al. (2008) stratified data by the
36 level of maternal self-esteem as reported by mothers. Higher age 2-year blood Pb level
37 was associated with lower MDI score among children with mothers in the lowest three
November 2012 5-78 Draft - Do Not Cite or Quote
-------
1 quartiles of self-esteem but not among children with mothers in the highest quartile of
2 self-esteem (Table 5-4). Model covariates included cohort, sex, parity, and maternal IQ,
3 age, education, smoking, alcohol consumption, and self-esteem. These findings indicated
4 that maternal psychosocial functioning may influence the effects of Pb on the mental
5 development of young children.
6 In another study in Mexico City, higher cord blood Pb level was associated with a lower
7 MDI score in children at age 2 years (-0.73 points [95% CI: -12, -0.23] in MDI per
8 1 (ig/dL increase in cord blood Pb level in a linear model) (Pilsner et al., 2010).
9 Investigators reported a lack of effect modification by genetic variants in the
10 methylenetetrahydrofolate reductase (MTHFR) enzyme, which is involved in folate
11 metabolism. The MTHFR 677 TT genotype produces an enzyme with lower metabolic
12 activity, is associated with lower serum folate levels (Kordas et al.. 2009). and in this
13 Mexico City cohort, was associated with lower MDI score at age 2 years. Results from
14 stratified analyses were not reported, thus differences in the magnitude of association
15 between genotypes could not be compared.
16 Consistent with the prospective evidence, a recent cross-sectional analysis indicated an
17 association between higher concurrent blood Pb level and lower MDI score children in
18 the Philippines, ages 6 months to 3 years (Solon et al., 2008). Although HOME score,
19 years of schooling of child, premature birth, region of residence, and maternal education
20 and smoking were examined as potential confounding factors, adjustment was made in
21 two stages: first, adjustment for blood Pb determinants and second, adjustment for MDI
22 determinants. This method may not adequately control for the variance shared by blood
23 Pb level and MDI. In this cohort, children with lower folate levels had larger
24 Pb-associated decreases in cognitive function. Among children with folate levels
25 < 230 (ig/mL, blood Pb level had an association with lower MDI scores in the range of
26 0.80 to 2.44 points. Among children with higher folate levels, blood Pb level was not
27 estimated to have a negative impact. The results from this study indicated a moderating
28 effect of folate on blood Pb levels as folate levels were not associated with MDI. Higher
29 folate level has been associated with lower blood Pb level due to the role of folate in
30 increasing Pb excretion by inhibiting the binding of Pb to blood elements.
31 In summary, evidence consistently indicates associations of higher blood Pb levels with
32 lower MDI scores in children ages 2-3 years (Table 5-4). Key evidence was provided by
33 prospective studies, in particular those that adjusted for maternal IQ and education, SES,
34 birth outcomes, and HOME score rWasserman et al.. 1992; Bellinger et al.. 1987).
35 Several large studies contributed to the evidence (n = 146-592), and several studies had
36 high to moderate follow-up participation, and nonselective loss-to-follow-up, which
37 reduces the likelihood of selection bias. Higher blood Pb level was associated with lower
November 2012 5-79 Draft - Do Not Cite or Quote
-------
1 MDI scores in a few different cohorts in Mexico City and other populations, and while
2 most adjusted for maternal education and IQ, they did not examine potential confounding
3 by parental caregiving quality. The lack of association observed in the Cleveland cohort
4 does not mitigate the otherwise compelling evidence, given the high prevalence of
5 prenatal alcohol exposure in this cohort and the evidence in the Boston and Yugoslavia
6 cohorts that adjusted for several potential confounding factors. MDI was associated with
7 prenatal (cord or maternal) and concurrent blood Pb levels. Comparison among blood Pb
8 levels at different lifestages did not consistently indicate a stronger effect on MDI of
9 prenatal or postnatal Pb levels. Pb-associated decrements in MDI were found in
10 populations with mean blood Pb levels 5-10 (ig/dL. However, a cross-sectional analysis
11 in children in Mexico City found a larger decrement in age 2 year MDI per unit increase
12 in concurrent blood Pb level among children with blood Pb levels <5 (ig/dL versus
13 5-10 (ig/dL or <10 (ig/dL (Tellez-Rojo et al.. 2006). An association was found in children
14 in Poland with lower cord blood Pb levels, median 1.23 (ig/dL (Jedrychowski et al.,
15 2009b). However, since cord blood Pb levels reflect blood Pb levels of mothers, which in
16 turn are influenced by Pb mobilized from bone to blood, the specific Pb exposure
17 scenario contributing to the observed associations is uncertain. Overall, while evidence
18 indicates associations of higher prenatal and postnatal child blood Pb levels with lower
19 MDI scores in young children (ages 2-3 years), the impact on later cognitive function is
20 not certain since MDI scores do not necessarily predict later IQ in children with normal
21 development.
5.3.2.3 Learning and Memory in Children
Epidemiologic Studies of Learning and Memory in Children
22 The small body of studies in the 2006 Pb AQCD did not clearly indicate associations
23 between higher blood Pb level and poorer memory or learning (i.e., acquisition of new
24 information) in children ages 5-17 years (Table 5-5). Studies used various tests to assess
25 learning and memory, which may account for some of the heterogeneity observed in
26 effect estimates. Evidence from prospective analyses in the Rochester, Boston, and
27 Cincinnati cohorts was mixed, with an association found in the Rochester cohort at age 5
28 years (Canfield et al.. 2004). associations in the positive and negative direction in the
29 Boston cohort across the various tests and ages of blood Pb examined at ages 5 and 10
30 years (Stiles and Bellinger. 1993; Bellinger et al.. 1991). and blood Pb-associated
31 improved memory found in the Cincinnati cohort at age 15-17 years (Ris et al.. 2004).
32 Previous cross-sectional studies found associations between higher concurrent blood Pb
33 levels and poorer learning and memory, including the large (n = 4,852) study of children
November 2012 5-80 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
ages 6-16 years participating in NHANES (Lanphear et al.. 2000). Several recent studies,
all cross-sectional, also found associations between higher concurrent blood Pb level and
poorer memory in children ages 6-16 years. Some were variants of previous studies
(Krieg et al.. 2010; Froehlich et al.. 2007): others had limited implications because of
little consideration for potential confounding (Counter et al., 2008; Min et al., 2007).
The prospective studies had smaller sample sizes (n = 148-195) than cross-sectional
studies (n = 246-4,852) but greater examination of potential confounding. Further,
recruitment of participants before or at birth, moderate to high follow-up participation,
and in most cases follow-up not biased to higher blood Pb levels and lower cognitive
function reduce the likelihood of selection bias (Table 5-5). Another strength was the
examination of earlier childhood or lifetime average blood Pb levels, which better
indicated the temporal sequence between Pb exposure and decrements in learning and
memory.
Table 5-5 Associations between blood Pb levels and performance on tests of
learning and memory in children.
Study
Canfield et
al. (2004)
Froehlich et
al. (2007)
Study Population and Methodological
Details
(Prospective studies first, then cross-sectional
studies. Within each category, presented in
order of strength of study design)
174 children born 1994-1995 followed from age
6 mo to 5 yr, Rochester, NY
Prospective. Recruitment from study of dust
control. 73% nonwhite. High follow-up participation,
no selective attrition. Linear regression model
adjusted for neonatal intensive care unit (NICU)
admission, sex, age, spatial span length. Also
considered potential confounding by prenatal
smoking, household income, maternal IQ and
education, ethnicity, HOME, breastfeeding
duration, 1st prenatal visit, spatial working memory
problem, birth weight, marital status, household
crowding.
174 children born 1994-1995 followed from age
6 mo to age 5 yr, Rochester, NY
Cross-sectional. Same cohort as above. High
follow-up participation, no selective attrition. Linear
regression model adjusted for income (spatial
working memory), HOME, maternal IQ, race
(spatial span). Also considered potential
confounding by transferrin saturation, prenatal
smoking exposure, maternal education, age, NICU,
sex.
Blood Pb
Levels
(ug/dL)
Lifetime (to age
5 yr) avg
Mean (SD): 7.2
(3.6)
Interval
analyzed: 3.5
(10th
percentile)-10
Concurrent
Mean (SD): 6.1
(4.9)
Interval
analyzed: 1.9
/•] r\*u
^ i uin
percentile)-10
.
Memory/ SI?fV!n^'ma*e
Learning Test (95/oCI)
Spatial span total -0.1 1 (-0.20, -0.02)b
errors CANTAB
Age 5 yr
Spatial working -0.51 (-1.2, 0.1 6)b
memory,
total errors
Spatial span -0.02 (-0.04, 0)
length
CANTAB
Age 5 yr
November 2012
5-81
Draft - Do Not Cite or Quote
-------
Study
Bellinger et
al. (1991)
Stiles and
Bellinger
(1993)
Riset al.
(2004)
Lanphear et
al. (2000)
Krieg et al.
(2010)
Study Population and Methodological
Details
(Prospective studies first, then cross-sectional
studies. Within each category, presented in
order of strength of study design)
1 70 children followed from birth (1 979-1 981 ) to age
5 yr, Boston, MA area
Prospective. Recruitment at birth hospital.
Moderate follow-up participation. More participants
were white, had higher age 2 yr HOME score,
higher postnatal blood Pb levels. Log linear
regression model adjusted for SES, maternal IQ
and marital status, preschool attendance, HOME,
out of home care, residence changes, medication
use in previous 12 mo, # adults in home, child sex,
race, birth weight, birth order.
1 48 children followed from birth (1 979-1 981 ) to age
10yr, Boston, MA area
Prospective. Same cohort as above. Moderate
follow-up participation, participants had higher SES
and HOME score. Linear regression model
adjusted for HOME, family stress, race, marital
status (earlier childhood blood Pb), HOME, family
stress, maternal age and race, birth weight, number
of daycare situations to age 57 mo (concurrent
blood Pb).
195 children followed prenatally (1979-1985) to age
15-1 7 yr, Cincinnati, OH
Prospective. Recruitment at prenatal clinic. High
follow-up participation, no selective attrition. Mostly
African-American. Linear regression model
adjusted for SES, maternal IQ, HOME, adolescent
marijuana use, and obstetrical complications. Also
considered potential confounding by birth
outcomes, maternal age, prenatal smoking,
alcohol, marijuana, and narcotics use, number of
previous abortions, stillbirths, gravidity, parity,
caregiver education, public assistance, child age,
sex, health, Fe status
4,852 children ages 6-1 6 yr (born 1972-1988), U.S.
NHANES III (1988-1994)
Cross-sectional. Large U.S. representative study
of multiple risk factors and outcomes. Linear
regression model adjusted for sex, race/ethnicity,
poverty index ratio, reference adult education,
serum ferritin and cotinine levels. Did not consider
potential confounding by parental cognitive
function, HOME score.
766-780 children ages 1 2-1 6 yr (born 1 975-1 982),
U.S. NHANES III (1991 -1994)
Cross-sectional. Large U.S. representative study
of multiple risk factors and outcomes. Log linear
regression model adjusted for sex, caregiver
education, family income, race/ethnicity, test
language. Did not consider potential confounding
by parental cognitive function, HOME score.
Blood Pb
Levels
(ug/dL)
Earlier
childhood (age
2yr)
Mean (SD):
7.0 (6.6)
Interval
analyzed: 1.8
(10th
percentile)-10
Concurrent
blood Pb levels
NR
Earlier
childhood
Exact levels NR
but concurrent
mean reported
to be <8
Memory/
Learning Test
Memory
Age 2 yr blood
Pb
Concurrent blood
Pb
McCarthy Scale
of Children's
Abilities,
Age 5 yr
Effect Estimate
(95% Clf
-0.1 4 (-0.52, 0.25)
0.10 (-0.41, 0.61)
Perseveration score, CVLT, Age 10 yr
Age 1 yr blood
Pb
Age 2 yr blood
Pb
0.02 (0, 0.04)b
-0.03 (-0.05, -0.01 )b
# trials to 1st category, WCST, Age 10 yr
Earlier
childhood (age
6.5 yr)
Mean (SD): NR
Concurrent
Geometric
mean: 1.9
(5th-95th:
1.70-2.10)
Interval
analyzed:
1 .74-2.06 =
1 0th-90th
percentiles
Concurrent
Mean
(5th-95th): 1 .95
(1 .63-2.27)
Interval
analyzed:
1.69-2.19 =
1 0th-90th
percentiles
Age 1 yr blood
Pb
Memory
composite of
SubtestsofCVLT
Ages 15-1 7 yr
Digit Span
WISC-R
Ages 6-1 6 yr
Digit span
WISC-R
Ages 12-1 6 yr
-0.44 (-0.93, 0.05)b
0.01 (-0.02, 0.05)
-0.05 (-0.89, -0.01)
-0.42 (-0.65, -0.18)
November 2012
5-82
Draft - Do Not Cite or Quote
-------
Study
Surkan et
al. (2007)
Kordas et
al. (2006)
Study Population and Methodological
Details
(Prospective studies first, then cross-sectional
studies. Within each category, presented in
order of strength of study design)
389 children ages 6-10 yr, Boston, MA,
Farmington, ME
Cross-sectional. Recruitment from trial of
amalgam fillings. High participation rate. Higher
participation by white children in Maine. Analysis of
covariance adjusted for caregiver IQ, child age,
SES, race, and birth weight, Also considered
potential confounding by caregiver education and
marital status, parenting stress, and maternal
utilization of prenatal or annual health care but not
HOME score.
293 children, age 7 yr, Torreon, Mexico.
Cross-sectional. Recruitment at prenatal clinic.
High participation rate. Residence near metal
Blood Pb
Levels
(ug/dL)
Concurrent
Mean (SD): 2.2
(1.6)
Concurrent
Interval
analyzed:
Memory/
Learning Test
General memory
index, WRAML
Ages 6-1 0 yr
Stern berg
memory
Age 7 yr
Effect Estimate
(95% Clf
-6.7 (-12.1, -1.2)°
Blood Pb group
5-10 ug/dL vs.
1-2 ug/dL
-0.1 6 (-0.37, 0.05)
Chiodo et
al. (2004)
Concurrent
Mean (SD):
5.4 (3.3)
Interval
analyzed:
2.3-9.5 =
10th-90th
percentiles
Verbal learning,
WRAML
Corsi Backward
Spatial Span
Age 7.5 yr
-0.20, p>0.05d
-0.22, p>0.05d
foundry. Linear regression model adjusted for child 2.1-10.0
sex, age, school, birth order, hemoglobin, forgetting
homework, household possessions and crowding,
house ownership, maternal education, family
structure, urinary As, tester. Did not consider
potential confounding by HOME score or parental
cognitive function.
246 children, age 7.5 yr, Detroit, Ml area
Cross-sectional. Recruitment at prenatal clinic. All
African-American. High prevalence of prenatal
alcohol exposure. High participation rate. Log linear
regression model adjusted for SES (all outcomes).
SES, caregiver vocabulary, disruption in caregiver
(verbal learning). HOME score, child age, child sex,
caregiver education, parity (spatial span). Also
considered potential confounding by maternal
prenatal marijuana, smoking, or cocaine use,
crowding, child life stress, caregiver age, life stress,
and psychology, conflict tactics, prenatal alcohol
exposure, family functioning, # children <18 years.
Note: Results are presented first for prospective studies then for cross-sectional studies. Results from the same cohort are kept
together. Within each category, results are presented in order of strength of study design.
CANTAB = Cambridge Neuropsychological Testing Automated Battery, CVLT = California Verbal Learning Test,
WCST = Wisconsin Card Sorting Test, WISC-R = Wechsler Intelligence Scale for Children Revised, WRAML = Wide Range
Assessment of Memory and Learning
"Effect estimates are standardized to a 1 ug/dL increase in blood Pb level in the lowest range of blood Pb levels examined in the
study or the interval from the 10th percentile to 10 ug/dL or the 90th percentile, whichever is lower.
bThe direction of the effect estimate was changed such that a negative estimate represents poorer performance and a positive
estimate represents better performance.
°Effect estimate compares test performance of children in higher blood Pb groups to children in lowest blood Pb group.
Sufficient data were not provided to calculate 95% Cl.
1
2
3
4
5
6
7
8
9
10
There were contrasting associations between blood Pb levels and memory in the
Rochester and Boston cohorts at age 5 years (Canfield et al.. 2004; Bellinger et al.. 1991).
Although different time periods of blood Pb level were examined, the population means
were similar, ~7 (ig/dL. In the Rochester cohort, a 1 (ig/dL increase in higher lifetime
average blood Pb level was associated with 0.11 (95% CI: 0.02, 0.20) more total errors
on the spatial span memory task (i.e., errors in replicating a sequence pattern) with
adjustment for neonatal intensive care unit (NICU) admission, sex, age, and spatial span
length and consideration for several other factors (Table 5-5) (Canfield et al.. 2004).
Recent evidence extended findings to associations between poorer performance on a
spatial working memory tasks and higher concurrent blood Pb levels (Froehlich et al..
November 2012
5-83
Draft - Do Not Cite or Quote
-------
1 2007). which also reflect represent past and recent Pb exposure. In the recent analysis,
2 associations were found with spatial span length (number of squares recalled correctly)
3 with adjustment for HOME score, maternal IQ, and race; and with spatial working
4 memory errors with adjustment for family income. In each analysis of the Rochester
5 cohort, multiple associations were examined; however, there were consistent patterns
6 blood Pb-associated decrements in cognitive function observed across the various indices
7 of memory, learning, and executive function examined. Coherence for associations with
8 performance on spatial span and spatial working memory tasks was found with evidence
9 in rodents for Pb-induced impaired performance on visual-spatial memory tasks in the
10 Morris water maze and working memory tasks in the radial arm maze, respectively
11 (discussed below). In contrast, in the Boston cohort at age 5 years, concurrent blood Pb
12 level was not associated with poorer memory, as assessed using the McCarthy Scale of
13 Children's Abilities (Bellinger et al.. 1991). These results were adjusted for more
14 potential confounding factors than results from other studies (Table 5-5). including SES,
15 maternal IQ, and HOME score. Higher age 2 year blood Pb level was associated with
16 poorer memory at age 5 years, but the association lacked precision (Table 5-5).
17 In the Boston cohort at age 10 years, associations were inconsistent across the multiple
18 tests of memory and learning and time periods of blood Pb levels (ages 1, 2, 5, 10 years)
19 examined (Stiles and Bellinger. 1993). For example, higher age 1 year blood Pb level was
20 associated with better memory (i.e., fewer errors in recalling word list) as assessed with
21 the California Verbal Learning Test. In the Cincinnati cohort at age 15-17 years, higher
22 earlier childhood (age 6.5-year) blood Pb level also was associated with better memory
23 (composite score of various subtests of the California Verbal Learning Test with
24 adjustment for similar potential confounding factors plus adolescent marijuana use (Ris et
25 al.. 2004). In two independent Boston-area cohorts examined at different ages, poorer
26 learning (number of trials to sort cards properly or number of categories achieved) as
27 assessed with the Wisconsin Card Sorting Test (WCST) was associated with higher age 1
28 year blood Pb level in children ages 10 years (Stiles and Bellinger. 1993) and with higher
29 childhood tooth Pb levels in young adults ages 19-20 years (-0.6 categories [95% CI:
30 -1.0, -0.21] per natural log unit increase in tooth Pb level [collected from age 5-8 years],
31 with adjustment for parental IQ, maternal age and education, SES, sex, birth order,
32 current smoking status, drug use, and alcohol use) (Bellinger et al.. 1994a). In the
33 younger cohort age 10 years, decrements in learning as assessed by performance on the
34 WCST were not consistently found across the various ages of blood Pb level examined
35 (Stiles and Bellinger. 1993).
36 The cross-sectional studies examined potential confounding by parental education and
37 SES, but a notable omission was consideration for HOME score. Chiodo et al. (2004)
38 found a concurrent blood Pb-associated decrement in spatial memory with adjustment for
November 2012 5-84 Draft - Do Not Cite or Quote
-------
1 HOME score. However, the results may lack generalizability because of the high
2 prevalence of prenatal alcohol exposure. HOME score was not associated with memory
3 in every study. For example, in the Rochester cohort, only household income remained
4 significantly associated with total errors in the spatial working memory task and was
5 included in the final model (Froehlich et al., 2007). Therefore, the confounding factors
6 may vary among populations. Most cross-sectional studies found Pb-associated
7 decrements in memory in populations with mean concurrent blood Pb levels 5-8 (ig/dL.
8 Despite the lack of information on HOME score, the cross-sectional analyses of children
9 participating in NHANES III had several strengths, including large sample sizes
10 (n = 760-4,852), high participation rates, lower likelihood of selection bias due to the
11 examination of multiple risk factors and outcomes, nationally-representative results, and
12 examination of the shape of the concentration-response relationship (Krieg etal.. 2010;
13 Lanphear et al.. 2000). An increase in concurrent blood Pb level was associated with a
14 larger decrement in digit span score in adolescents ages 12-16 years (Krieg et al., 2010)
15 than children 6-16 years (Lanphear et al.. 2000); however, the influence of higher past Pb
16 exposures is likely greater in older children. In the analysis of children ages 6-16 years,
17 Lanphear et al. (2000) estimated the largest decrement in memory score per unit increase
18 in blood Pb level in children with concurrent blood Pb levels <2.5 (ig/dL (-0.25 [95% CI:
19 -0.58, 0.08] points per 1 (ig/dL increase in concurrent blood Pb level versus -0.05 [95%
20 CI: -0.09, -0.01] among all subjects). A nonlinear concentration response relationship
21 also was found among children ages 7 years living near a metal foundry in Torreon,
22 Mexico, with a larger Pb-associated decrement in memory found among children with
23 concurrent blood Pb levels <10 (ig/dL (Kordas et al.. 2006).
lexicological Studies of Learning and Memory
24 As described in the preceding sections, blood Pb levels are consistently associated with
25 decrements in FSIQ in children but show more variable associations with performance on
26 neuropsychological tests of learning and memory. A relationship between Pb exposure
27 and cognitive function decrements is supported further by evidence for Pb-induced
28 impairments in memory and learning in animals. The 2006 Pb AQCD (U.S. EPA. 2006b)
29 reported Pb-induced impaired memory and learning primarily in animals with Pb
30 exposures that resulted in blood Pb levels 30-50 (ig/dL; however, some studies observed
31 impairments in rodents (pre- and/or post-natal Pb exposure) and monkeys (lifetime
32 postnatal Pb exposure) with blood Pb levels 14-25 (ig/dL (Altmann et al.. 1993; Rice and
33 Karpinski. 1988; Gilbert and Rice, 1987). Several recent studies added to the evidence for
34 impaired learning and memory in animals with lower blood Pb levels in the range
35 relevant to humans, 8-17 (ig/dL (Cory-Slechta et al., 2010; Niu et al., 2009; Virgolini et
36 al.. 2008a; Stangle et al.. 2007). Effects in animals with lower blood Pb levels generally
November 2012 5-85 Draft - Do Not Cite or Quote
-------
1 were found with gestational/lactational Pb exposures. Results from toxicological studies
2 on learning as well as other nervous system endpoints that provided concentration-
3 response information (i.e., those with multiple Pb exposure concentrations) are shown in
4 Figure 5-3 and associated Table 5-6. These results demonstrate the coherence among
5 inter-related CNS changes induced by Pb exposure in animals, including deficits in CNS
6 development and plasticity and alterations in neurotransmitters, which mediate cognitive
7 function.
November 2012 5-86 Draft - Do Not Cite or Quote
-------
Behavioral; Neonate; Rat; Fernale(l)
Beh.ivior.il; Neonale; Ral; Bolh (2)
BDhaviorat: Nconate; Rat; Male (i)
Beli.ivior.il; Neonatc; Rat; Bolh(<1)
Cognition; NeolhUD; Rat; Both (5)
Cognition; Adull; Rat; Male(G)
Cognition;Adull: Monkey; Bolh (7)
Cognition; Ncon.lte; Mouse; Both(8)
l_ortniticin;N<>onate: Rat; Both (9)
Cognition;Neonate; Rat; Both (1)
' .,,(,. ,. ,1, i ..,,.• (J. ",,.,!• Rat; Female (10)
• ,,ih, ,,'.1 ,',.,,„• !!.•..,,,,i. Ral; Male(ll)
i ..rii. ..-.I.-(..!!,• .-...lull Ral; M ,i, i i •,
Corticosterone;Adult; Ral; Male(13)
Morphology; Neonale; Rat;Both I M)
Morphology: Adult; Rat; Male (6)
Morphology; AduH: Montey; Female (15)
Morphology; Adult; Ral;Male (1C)
Morphology. Neonale; Mouse; Bolh (8)
Morphologv; Neonate; Rat; Both (9)
Motor lunction; Neon.ile; Mouse; M.ile(J7)
M..r,.r Inn. n,,i, ' I'lli II.,- M i, I i i
Motor (unction; Neonale; Rat; Both (5)
Mm or dinci inn; Adull: Rat; Male(13)
Molor (unction; Ncnn.ite; Ral; Both (1)
Mot or (unction; Neon ate; Rat; Male(3)
Ncurophysiology; Neonale; Rat; Both (1*1)
Neuro!ransmiller; Neonate; Mouse; Both(18)
Neurotransmilter, Neonate; Mouse: Both(17)
Neurotransmltter Adult; Rat; Female (13)
Ncurotr.insmillor; Adull; Ral; Male(13)
NeurotransmKten Neonale; Rat; Female (11)
Ond.Ttivc St ross; Adull; Monkey; Female (IS)
»...i'l.itivi;Sli<£.s; Neonale; Mouse; Bolh (8)
Physical Develop me nl; Adull; Ral; Female (14)
I'll"'.!, .il (>i,vi'li>|iMii'iil: H.,nii.ili-. R.it. (•i-ni.ili'tl)
F'livsii'-il ll.".'fli-.|im.Ti1; M.'on.il,-: fi.fl; Mal.-(i)
Physical [levebpmcnl; Adull; Ral; M.ile(C)
ProlKefMion/dlfl/survival; Neonate; Rat; Both (9)
Stress-Induced cort icosterone; Neonale; (tat: Fern ale (10)
Slrcss-induccd cofticosti;ronc;Adull; Ral; Female(13)
Slriiii-induced cort icosterone; Neonale; Ral; Fern ale (11)
Stress-induced motorfunction; Adull; Rat; Both(13)
Stress-induced neurotransmiltcr: Neonate; Rat; Female (10)
Stress-induced ncumtransmltter. Adult; Ral; Female (13)
Slress-inducedneurotranimlller; Adult; Rat; Male(13)
Siren-induced neu retransmit tec Neonale; Rat; Male (11)
• Highest Concentration
*Lowfl5tCooc. with Response
A Highest Cone with No Response
o LosvestConcentration
10
100
1,000
Blood Lead (ng/dl)
Note: This figure illustrates nervous system effects associated with Pb exposure in studies that examined multiple exposure
concentrations. Dosimetric representation reported by blood Pb level. (ID corresponds to studies described in Table 5-6)
Figure 5-3 Summary of Pb exposure-nervous system concentration-
response information from toxicological studies.
November 2012
5-87
Draft - Do Not Cite or Quote
-------
Table 5-6 Summary of findings from neurotoxicological concentration-
response array presented in Figure 5-3.
Study
ID in
Figure
5-3 Reference
Blood Pb
Level
(ug/dL)
Outcome
1
10
Beaudin et
al. (2007)
Grant et al.
(1980)
13 & 31 Behavior, neonate: Lactational Pb exposure, offspring deficient in Reward Omission testing.
Physical development; Postnatal Pb exposure (birth to 4 weeks of age): Pb-induced
development of over-reactivity to reward omission and errors is reversible with chelation
treatment.
57
Behavior, neonate: chronic Pb exposure (drinking water) to dams and pups, Changed behavior,
males.
Kishi et al. 59 & 186 Behavior, neonate: Pb exposure (oral gavage of pups) during lactational period, Changed
(1983) emotional behavior, males and females.
Motor function, neonate: Pb exposure (oral gavage of pups) during lactational period, motor
function (rotarod performance) impaired, both sexes.
Physical development; Pb exposure during lactation (oral gavage): Delayed development of
righting reflex in male rats.
Overmann 33, 174 & Behavior-Pb exposure (oral gavage of pups) during lactation: aversive conditioning affected by
(1977) 226 Pb exposure, male and females.
Cognition-Pb exposure (oral gavage of pups) during lactation: Response inhibition impaired,
both sexes.
Motor function- Pb exposure (oral gavage of pups) during lactation: Increased motor activity
and impaired motor coordination (rotarod), male and females.
Stangle et 13 & 31 Cognition; Developmental Pb exposure (PND1-PND30): Impaired learning with visual
al. (2007) discrimination task, heightened response to errors, both sexes.
Motor function; Developmental Pb exposure (PND1-PND30): Alcove latency and response
latency significantly affected by Pb exposure, both sexes.
Gong &
Evans
(1997)
38 & 85 Cognition-Adult male 21 day Pb exposure: Hyperactivity with Habituation to new cage
environment.
Morphology; 21 day Pb exposure to adult males: Marker of neuronal injury-elevated
hippocampal glial fibrillary acidic protein (GFAP).
Physical development; Adult male rats (21 day Pb exposure): Neurotoxicity measured with
brain glial fibrillary acidic protein (GFAP).
Rice (1990) 32 & 36
Cognition-Chronic Pb exposure from birth: Spatial discrimination reversal task impairment, both
sexes.
Lietal.
(2009C)
80 & 100 Cognition-Gestational & lactational Pb exposure: Morris water maze performance impaired.
Morphology; Gestational & lactational Pb exposure: Increased levels of inflammatory cytokines
& exocytosis related proteins in brains of pups at weaning, both sexes.
Oxidative stress-gestational and lactational Pb exposure: Elevated hippocampal TNF levels in
offspring, males and females.
80 & 102 Cognition- Gestational & lactational Pb exposure: Morris water maze performance impaired.
Morphology: Increased levels of Alzheimer disease-associated proteins in mice with gestational
and lactational Pb exposure, both sexes.
Proliferation/diff/survival, gestational & lactational Pb exposure: Increased hippocampal
expression of P-tau and amyloid beta in male and female pups.
10 & 13 Corticosterone: Lifetime Pb +/- stress: Correlation between 9-month old's corticosterone level
and frontal cortex dopamine levels in behaviorally tested female offspring.
Stress: Corticosterone-Lifetime Pb plus stress: Affects Fl performance, dopamine and serotonin
levels in female offspring.
Stress: Corticosterone-neurotransmitter-Lifetime Pb exposure in female rats plus stress:
Dopamine homeostasis affected.
11 Virgolini et 25 Corticosterone: Maternal Pb plus stress: Elevated corticosterone in male offspring with prenatal
al. (2008b) stress + offspring stress was further enhanced with Pb exposure.
Stress: Corticosterone-Maternal Pb plus stress: Affects Fl performance.
Lietal.
(201 Ob)
Cory-
Slechta et
al. (2010)
November 2012
5-8
Draft - Do Not Cite or Quote
-------
Study
ID in
Figure
5-3 Reference
16
18
Blood Pb
Level
(ug/dL)
Outcome
Neurotransmitter; Gestational and lactational Pb exposure: Induced dopamine and serotonin
changes in rat offspring.
Stress induced neurotransmitter effects, Maternal Pb plus stress: serotonin and 5-HIAA
(serotonin metabolite), and dopamine turnover were significantly affected in males.
12 Virgolini et 15&27 Corticosterone: Chronic Pb exposure from weaning: Pb exposure alone decreased basal
al. (2005) plasma corticosterone levels at 5 months of age, males.
Motor function: Chronic Pb exposure from weaning: Locomotor activity significantly decreased
Fl response rates & increased post-reinforcement pause period in a concentration-dependent
manner, males.
Stress: Corticosterone-Chronic Pb plus stress: Affects neurotransmitters & Fl performance
13 Virgolini et 31 Corticosterone: Maternal Pb exposure (gestation and lactation) +/- stress: Differential basal
al. (2008a) corticosterone levels between behavioral and non-behavioral tested rats, females.
11 &/or Stress: Corticosterone-Maternal Pb plus stress: Affects Fl performance, dopamine, serotonin,
31 and NE levels.
Motor function: Maternal Pb +/- stress: Increased locomotor activity (run rate) with Pb and
stress exposure.
31 Neurotransmitter; Gestational and lactational Pb exposure: Induced NE aberrations in adult rat
offspring (both sexes).
Stress induced motor function: Maternal Pb +/- stress: Increased locomotor activity (run rate)
with Pb and stress exposure.
Stress induced neurotransmitter; Gestational and lactational Pb exposure + stress: Induced
HVA (monoamine neurotransmitter metabolite) and NE aberrations in female adult rat offspring.
14 Hu et al. 4&12 Morphology; Gestational Pb exposure: Neurite outgrowth marker PSA-NCAM decreased in rat
(2008b) pups, both sexes.
Neurophysiology; gestation Pb exposure: decreased hippocampal sialyltransferase activity,
both sexes.
Physical development; t-Gestational Pb exposure: Early brain synapse development impaired
(hippocampal PSA-NCAM).
15 Wuetal. 19&26 Morphology: Elevated expression of Alzheimer's disease-related genes and Tc factors in aged
(2008a) brains of female monkeys (exposed to Pb as infants).
Oxidative stress: Elevated oxidative DNA damage in aged brains of female monkeys (exposed
to Pb as infants).
Tavakoli-
Nezhad et
al. (2001)
18, 29, & Morphology; 3 to 6 weeks of Postnatal (starting at PND22) Pb exposure in males: Decreased
54 number of spontaneously active midbrain dopamine neurons.
17 Leasure et 10 & 42 Motor function; Mouse maternal (dam) Pb exposure: Induced decreased rotarod performance
al. (2008) in offspring (1 year-old male offspring).
Neurotransmitter; Mouse maternal (dam) Pb exposure: Affects 1 -year old male offspring
dopamine homeostasis, both sexes.
Fortune &
Lurie (2009)
8&43
Neurotransmitter; Mouse maternal (dam) Pb exposure: Affects offspring superior olivary
complex (auditory) neurotransmitters, both sexes.
Learning and Memory - Morris Water Maze
November 2012
5-89
Draft - Do Not Cite or Quote
-------
1 In the Rochester cohort of children, blood Pb level was associated with poorer
2 performance on tests of spatial memory (Table 5-5). In animals, spatial memory has been
3 tested using the Morris water maze apparatus. The Morris water maze can be used to test
4 spatial memory and learning ability by assessing the time or distance taken for a rodent to
5 swim to a submerged platform using visual cues and in subsequent trials with the
6 platform removed, the time spent in the previous location of the platform. The
7 2006 Pb AQCD (U.S. EPA. 2006b) reported mixed effects of Pb on memory; some
8 studies found Pb-induced impaired memory in animals whereas others found improved
9 memory with Pb exposure (U.S. EPA. 2006b). Evidence was more consistent for
10 Pb-induced impairments in long-term memory, i.e., stored information, which was found
11 in animals with gestational, gestational/lactational, or gestation/lifetime Pb exposure
12 producing blood Pb levels 23-32 (ig/dL (Yang etal.. 2003; Jettetal.. 1997; Kuhlmann et
13 al.. 1997). Using the Morris water maze, Jett et al. (1997) found an effect of gestational
14 plus lactational Pb exposure of female rats (Pb acetate in maternal feed 10 days prior to
15 mating to PND21) on long-term memory but not working memory, which is memory for
16 information that changes frequently that is not stored permanently. Kuhlmann et al.
17 (1997) compared various lifestages of Pb exposure and found impaired learning and long-
18 term memory using the Morris water maze in Long Evans rats exposed to Pb during
19 gestation and lactation (via maternal diet) or over a lifetime from gestation through
20 adulthood each producing peak blood Pb levels of 59 (ig/dL, which are higher than those
21 relevant to humans. Pb exposure during only the post-weaning period, producing a more
22 relevant blood Pb level of 26 (ig/dL did not affect memory.
23 In contrast with Kuhlmann et al. (1997). recent evidence points to impairments in
24 memory and learning as assessed with the Morris water maze with postweaning Pb
25 exposure, albeit with higher blood Pb levels than relevant to humans. Impaired learning,
26 i.e., slower decrease in time to escape from the Morris water maze across training trials,
27 was found in weanling Sprague-Dawley male rats exposed to 400 or 800 mg/L Pb acetate
28 for 8 weeks in drinking water (Fan etal.. 2010; Fan et al.. 2009a). Further, various dietary
29 supplements (oral gavage of methionine, taurine, Zn, ascorbic acid or glycine) or
30 methionine-choline given before or concomitantly with Pb exposure in these weanling
31 males mitigated the effect of Pb on escape latency to resemble that of control pups at the
32 end of training (Fan et al.. 2010: Fan et al.. 2009a). In Fan et al. (2009a). Zn and
33 methionine given before Pb exposure attenuated Pb-induced impairments in spatial
34 memory and learning, whereas glycine, taurine, vitamin C, vitamin Bl, tyrosine had no
35 effect (blood Pb levels ranged from 7 to 70 (ig/dL depending on nutrient status and
36 recovery time). In Fan et al. (2010). Pb-exposed rat pups (blood Pb level 50.2 (ig/dL)
37 showed deficits in retaining information about the platform location after test day 2 and
38 3. Concurrent methionine-choline treatment mitigated these impairments in spatial
39 memory. The action of methionine, a source of sulfur, may be attributable to its function
November 2012 5-90 Draft - Do Not Cite or Quote
-------
1 as a chelator and/or free-radical scavenger. Choline is important for cell membranes and
2 neurotransmitter synthesis (Zeisel and Blusztajn. 1994). and unrelated to Pb exposure,
3 choline supplementation of rats PND16-PND30 was shown to attenuate normal age-
4 related declines in spatial memory (Meek et al.. 2007).
5 Consistent with Kuhlmann et al. (1997). other recent studies found impairments in
6 learning and memory in animals as assessed using the Morris water maze with
7 gestational/lactational Pb exposures, albeit at higher concentrations than those relevant to
8 humans. In Li et al. (2009c). rodents were exposed GD1-PND21 to Pb acetate via the
9 drinking water of dams (1,000-10,000 ppm) and had corresponding blood Pb levels of
10 40-100 (ig/dL at PND21. Beginning at weaning, learning and spatial memory were
11 assessed in pups with a reversal procedure in the Morris water maze. Pb-exposed pups
12 with blood Pb levels of 80 and 100 (ig/dL had statistically significant increases in escape
13 latency, indicating impaired spatial memory and learning (Li et al.. 2009c). The pups in
14 Li et al. (2009c) were not separated by sex. Cao et al. (2009) found that long-term
15 postnatal Pb exposure from birth (2,000 ppm Pb acetate) impaired spatial memory in
16 male Wistar rats as adults (PND81-90), and these effects were exacerbated by long-term
17 administration of melatonin (3 mg/kg by gastric gavage, 60 days from weaning).
18 Mechanistic support for effects on learning and memory was provided in this study by
19 observations in the hippocampal dentate gyrus that Pb exposure also impaired long-term
20 potentiation (LTP), a major cellular mechanism underlying learning and memory.
Working Memory - Delayed Spatial Alternation
21 Working memory also can be measured by testing delayed spatial alternation (DSA). In
22 DSA, an animal receives rewards by alternating responses between two locations or
23 levers. This test requires working memory because the correct response changes between
24 trials, and the animal must determine which response is correct based on memory of its
25 previous response. Impaired working memory is indicated by increased response errors,
26 decreased percent of correct responses, and increased response perseverative errors
27 (i.e., repeatedly pressing the same lever without moving to the other lever when the
28 reward is moved. Studies detailed in earlier Pb AQCDs showed that Pb-exposed animals
29 had deficits in working memory as assessed with DSA (Alberand Strupp. 1996; Rice and
30 Gilbert. 1990b; Rice and Karpinski. 1988; Levin etal. 1987). Studies in nonhuman
31 primates showed that there were multiple lifestages and durations of Pb exposure that
32 induced poorer performance on DSA tasks, including lifetime from birth or later
33 postnatal (i.e., post-weaning to time of testing) (Rice and Gilbert. 1990b: Rice and
34 Karpinski. 1988; Levin etal.. 1987). Pb-induced impairments in DSA task performance
35 have been observed less consistently in rats with juvenile only or juvenile to adult
November 2012 5-91 Draft - Do Not Cite or Quote
-------
1 exposure. In fact, postweaning Pb exposure was shown to increase accuracy in
2 performance on DSA tasks in rodents (Cory-Slechta et al.. 1991).
Learning and Memory - Y Maze
3 The three-branch radial Y-maze test evaluates learning as the number of days required to
4 learn the maze (90% correctly). The Y-maze has a light at the end of one of the branches.
5 The branch with the illuminated light is the safe area whereas the other two branches are
6 electrified and cause a mild electric shock when entered. The spatial memory test
7 assessed by the Y-maze test shares homology with the spatial working memory test
8 conducted by Froehlich et al. (2007) in the Rochester cohort, which require children to
9 search boxes for a reward and avoid returning to boxes where the reward was previously
10 found. A recent study using the Y-maze showed impaired learning in Wistar albino rat
11 offspring exposed to Pb from lactation to adulthood up to 12 weeks of age (300 mg/dL
12 Pb acetate in dam drinking water and then in offspring drinking water postweaning) (Niu
13 et al.. 2009). Pb induced statistically significant impairments in learning at 8, 10 and
14 12 weeks of age but not age 6 weeks. These effects on learning were found with blood Pb
15 levels relevant to humans, 17 (ig/dL, as measured at age 6 weeks. Mechanistic support for
16 Pb-induced learning impairments was provided by concomitant observations of
17 Pb-induced attenuation in levels of hippocampal glutamate (Section 5.3.11.4). which
18 mediates signaling pathways involved in LTP.
Learning - Schedule-Controlled Behavior Testing
19 The 2006 Pb AQCD described the effects of Pb exposure on learning in animals as
20 measured with schedule-controlled behaviors using Fixed Interval (FI) or Fixed Ratio
21 (FR) operant conditioning reinforcement schedules and indicated differential effects by
22 Pb exposure concentration, with low-level (e.g., 11 (ig/dL) and high-level Pb (peak levels
23 of 115 (ig/dL) exposures increasing and decreasing FI response rate, respectively. This
24 nonlinear response has been examined further in recent work by the Cory-Slechta
25 laboratory, much of which also examined the interaction between stress and Pb exposure.
26 Impaired performance in FI testing with Pb exposure also supports the effects of Pb on
27 attention-related behavioral problems (Section 5.3.3.1).
28 Recent evidence indicated that certain learning impairments induced by certain levels of
29 Pb exposure were modifiable. Female rats were exposed to 300 ppm Pb acetate via dam
30 drinking water from birth through lactation PND1-PND21 and 30 ppm via their own
31 drinking water to PND30. Rats subsequently administered succimer PND31-PND52
32 (twice daily by oral gavage, resulting in blood Pb level 2.8 (ig/dL) performed better on
33 visual discrimination tasks than did the rats exposed to Pb alone (blood Pb level at
November 2012 5-92 Draft - Do Not Cite or Quote
-------
1 PND52: 12.6 (ig/dL) (Stangle et al.. 2007). Brain Pb levels also were lower in the
2 Pb+succimer group (196 ng/g) than Pb-only group (1,040 ng/g). Succimer alone in the
3 absence of Pb exposure resulted in some cognitive impairment in this study, and rats
4 given succimer after a higher Pb concentration (300 ppm, blood Pb level 8.5 (ig/dL) did
5 not have better learning ability than rats exposed to 300 ppm Pb alone (blood Pb level:
6 31 (ig/dL). Therefore, succimer administration may not completely alleviate the effects of
7 Pb exposure on learning impairments.
Learning Ability with Stress
8 The combined paradigm of Pb exposure and stress experienced by a laboratory animal
9 has been examined by the Cory-Slechta laboratory, which has focused on the common
10 pathway of altered HPA axis and brain neurotransmitter levels. Depending on the timing
11 of stress and Pb exposure concentration, greater impairments in learning were found in
12 animals with dietary Pb exposure that resulted in blood Pb levels relevant to humans
13 when combined with stress. Evidence additionally indicates that associations of Pb
14 exposure and stress with learning deficits (multiple schedule of repeated learning and
15 performance in females) may be related to aberrations in corticosterone and dopamine.
16 As indicated in Figure 5-3 and Table 5-6. Pb exposure with stress has been shown to
17 increase corticosterone levels and exacerbate Pb-induced dopamine release and learning
18 impairments. For example, learning deficits in female rat offspring at age 2 months were
19 enhanced following lifetime Pb exposure combined with prenatal stress, i.e., maternal
20 restraint (Cory-Slechta et al.. 2010). This exposure paradigm involved exposure of dams
21 to Pb acetate from 2 months prior to mating through lactation and exposure of their pups
22 from a mixed sex litter via drinking water Pb (50 ppm) through the remainder of their
23 lifetime (2 months). The peak blood Pb levels of pups (age 5-6 days) ranged from 10 to
24 13 (ig/dL. Learning impairments were found in repeated learning assessments but not
25 performance assessments. Pb/stress was found to increase the total number of responses
26 required to learn a sequence. Pb/stress exposure also affected dopamine from the frontal
27 cortex and dopamine turnover in the nucleus accumbens, which are processes underlying
28 cognition. Also, Pb-exposed offspring with and without maternal stress exposure had
29 statistically significant decreases in hippocampal nerve growth factor versus controls.
30 Another study of lifetime Pb exposure (50 or 150 ppm Pb acetate in drinking water to
31 dams from 2 weeks before pregnancy though lactation and to offspring thereafter)
32 indicated a potentiation of effects on learning with Pb and stress co-exposure, with stress
33 given prenatally via dams or postnatally to offspring. Lifetime 50 ppm Pb exposure plus
34 prenatal or postnatal stress resulting in blood Pb levels 11-16 (ig/dL decreased the post-
35 reinforcement pause (PRP) period in female offspring when examined starting at age
36 2 months (Rossi-George et al.. 2011) (Table 5-7. rightmost column). Animals with
November 2012 5-93 Draft - Do Not Cite or Quote
-------
1 150 ppm Pb exposure and blood Pb levels 25-33 (ig/dL had decreased PRP only with
2 prenatal stress. Within the FI schedule, the PRP represents timing capacity or proper
3 temporal discrimination and refers to the period during which the animal waits or pauses
4 before depressing the lever for a reward. In this case, decreased pause or PRP interval in
5 Pb plus stress-exposed animals indicates that they started responding earlier than did
6 controls. These results also point to an effect of Pb on increasing impulsivity
7 (Section 5.3.3.1). Separately, the overall FI response rate, which also indicates
8 impulsivity (i.e., rate of not withholding responses), was significantly increased with
9 50 ppm Pb exposure alone and with co-exposure to maternal or offspring stress. At
10 150 ppm, Pb increased FI response rate only with co-exposure to stress (maternal or
11 offspring). Biochemical analysis revealed alterations in frontal cortex norepinephrine,
12 reductions in dopamine homeostasis in the nucleus accumbens, and enhancement of the
13 striatal monoamine system as possible mechanistic contributions to Pb-induced
14 impairments in learning.
November 2012 5-94 Draft - Do Not Cite or Quote
-------
Table 5-7 Summary of effects of maternal and lifetime Pb exposure on
Fl performance observed by Cory-Slechta and colleagues.
Maternal Pbb
Pb (ppm)
Overall rate3
PRPa
Lifetime Pb"
Overall rate3
PRP"
0 ppm:
0-PS
0-OS
No Significant Effect
No Significant Effect
No Significant Effect
*i -23%
No Significant Effect
No Significant Effect
No Significant Effect
No Significant Effect
50 ppm:
50-NS
50-PS
50-OS
No Significant Effect
No Significant Effect
*t 64.9%
No Significant Effect
No Significant Effect
No Significant Effect
*t 95%
*t 79.2%
*t 74.7%
No Significant Effect
*i -42%
* |-39.3%
150 ppm:
1 50-NS
1 50-PS
1 50-OS
*t 42.4%
No Significant Effect
*t 59.2%
*| -30.3%
*i -25.7%
No Significant Effect
No Significant Effect
*t 90.7%
*t 78.5%
No Significant Effect
*i -44.7%
No Significant Effect
"Based on calculation of group mean values across session block post-stress challenge for both maternal and lifetime Pb exposure
studies. All calculations represent percent of 0-NS control values; f represents increase; J, represents decrease.
bData from Virgolini et al. (2005). 'Denotes significant effect versus 0 ppm control (p <0.05).
°Data from current study, Rossi-George et al. (2011)
'Denotes significant effect vs. 0 ppm control (p <0.05).
Source: Reprinted with permission of Elsevier Science, Table 1 of Rossi-George et al. (2011).
Notes:
PRP = Post-reinforcement pause; PS = Prenatal (maternal stress); OS= Offspring stress.
Overall results demonstrated that lifetime Pb exposure (right column) with or without prenatal stress induced learning deficits in
female rats as demonstrated by an increase in overall rate and decreased PRP. Mechanistically, these authors proposed that
associations of Pb and stress with learning deficits may be related to aberrations in corticosterone and dopamine. Prenatal Pb
exposure alone (left column) induced similar responses during testing at age 2 months.
November 2012
5-95
Draft - Do Not Cite or Quote
-------
1 A separate investigation from the same laboratory similarly indicated a potentiation of
2 effects with Pb and stress co-exposures but with developmental Pb exposure from
3 two months prior to mating through lactation (50 or 150 ppm Pb acetate in drinking
4 water) (Virgolini et al.. 2008a). Dams were subject to restraint stress at GD16-GD17.
5 Prenatal stress or Pb exposure alone did not affect FI performance in offspring.
6 Compared with controls, marked increases in response rates on FI performance were
7 found in the 50 ppm Pb plus prenatal stress female offspring at age 2-3 months, whose
8 mean blood Pb level was 19 (ig/dL at weaning. Using the same Pb exposure protocol,
9 Virgolini et al. (2008b) expanded evidence for Pb-stress interactions through the
10 examination of the effects of additional adult intermittent stress (cold, novelty or
11 restraint) on FI performance, corticosterone, and dopamine. Compared with females with
12 stress but no Pb, female offspring with adult intermittent restraint and cold stress and
13 higher dose Pb (150 ppm) had statistically significant increases in FI response rate and
14 decreased PRP, i.e., increased impulsivity (Figure 5-4, A panel). Male offspring showed
15 decreased FI response rates due to decreased run rate with restraint stress at the lower Pb
16 dose (50 ppm) (Figure 5-4. B panel). At the higher dose of Pb, males showed increased
17 FI response rates and increased run rates with cold stress.
18 Pb exposure over various developmental windows in rodents has been shown to affect the
19 HPA axis, as measured by the levels of corticosterone, the major glucocorticoid involved
20 in stress responses in rodents. Thus, modulation of corticosterone may provide a
21 mechanistic explanation for learning deficits (FI testing in females) found with Pb and
22 stress co-exposure in rodents. As examined by the Cory-Slechta laboratory, exposure to
23 Pb induced differential changes in corticosterone levels in each sex, depending on the age
24 of the animal and the timing of exposure, developmental (gestational and lactational),
25 post-weaning, or lifetime (Rossi-George et al.. 2011; Cory-Slechta et al.. 2010; Virgolini
26 et al.. 2008aV
November 2012 5-96 Draft - Do Not Cite or Quote
-------
A.
B.
Novelty
Novelty
C 60 164
0 SO 150
o 50 1SQ
0 50 160
Pastreinforcement Pause Time
0 SO 150
0 60 1SO
1ED * 120-i
III III
0 50 150 0 SO 150
iJtete
150 + 120-j
iJin Hi
0 50 150 0 50 150 0 50 150
Postreinforcemenl Pause Time
SOO -I MO -| 12S -i 150 -, 180-1 150 -, _!_
I L III ll III ill
0 60 ISO 0 40 150 0 SO 150
° M ,!" ° " "° ° » 1M Pb(ppm)] [Pbfepn,)] [Pblppm,]
[Pb(ppm)] [Pb(ppm)] [Pb (ppm)]
Note: * denotes significantly different from 0 ppm Pb. ~denot.es p = 0.07. + denotes significantly different from the 50 ppm Pb group.
Each column presents results for a particular stressor (restraint, cold, novelty). In females (panel A), gestational/lactational 50 and
150 ppm Pb exposure increased overall rate and decreased post-reinforcement-pause time over that of cold or restraint stress given
in adulthood. In males (panel B), gestational/lactational Pb exposure did not alter the effects of the stressors.
Source: Reprinted with permission of Elsevier Science, Virgolini et al. (2008b).
Figure 5-4 Changes in Fixed Interval performance in (A) female and (B) male
offspring with gestational/lactational Pb exposure plus various
stressors given in adulthood.
November 2012
5-97
Draft - Do Not Cite or Quote
-------
1 Because animals that are used for FI testing are regularly handled by laboratory personnel
2 and often participate in other tests of cognition, their baseline level of stress may be
3 skewed from that of a laboratory animal that constantly remains in a cage without daily
4 handling. Because effects on the HPA axis are of interest to Pb researchers, the baseline
5 corticosterone levels of animals that have participated in behavior testing (FI) and those
6 who have not (NFI) have been compared after gestational/lactational Pb exposure.
7 Virgolini et al. (2008b) found that baseline corticosterone levels were significantly
8 different between FI and NFI animals. Also, the effect of combined
9 gestational/lactational Pb exposure plus maternal stress on corticosterone was compared
10 in FI and NFI animals. At the baseline age of 4-5 months, Pb exposure with or without
11 stress did not induce differences in corticosterone levels in FI females but did in males
12 (Virgolini et al., 2008b). In the FI males, 50 ppm Pb exposure decreased corticosterone
13 versus control (no Pb exposure), and 150 ppm Pb exposure elevated corticosterone versus
14 control. In male NFI animals, a U shaped concentration-response was found, with 50 ppm
15 Pb exposure reducing corticosterone over than in the controls or with 150 ppm Pb
16 exposure. In the NFI males, stress did not affect corticosterone levels or interact with the
17 effect of Pb exposure. NFI females exposed to 150 ppm Pb had significantly elevated
18 corticosterone versus control (no Pb exposure). These data demonstrate that behaviorally
19 tested animals have altered HPA axis and altered responses to Pb exposure versus
20 animals that are housed under conditions without daily handling by caregivers.
21 Lifetime Pb exposure beginning in gestation (150 ppm drinking water of dams from
22 2 months prior to mating through lactation, then continuing in offspring water) induced
23 increases in basal (age 2 months, before behavioral testing) corticosterone only in female
24 offspring but not final (age 10 months, after testing) corticosterone in either female or
25 male offspring (Rossi-George et al., 2011) (Figure 5-5). The Pb-related increase in
26 corticosterone was found in animals with blood Pb levels of 19 and 31 (ig/dL measured at
27 PND21. Pb-stress interactions were observed at age 2 months but not 10 months (final,
28 after behavioral testing). At age 2 months, Pb plus stress attenuated the Pb-induced
29 elevations in corticosterone to baseline levels (Figure 5-5). By 10 months of age, these
30 offspring had lower corticosterone concentrations versus control animals. In males,
31 corticosterone levels were not affected significantly by Pb and/or stress at 2 (basal) or
32 10 (final) months of age (Figure 5-5) (Rossi-George et al.. 2011).
November 2012 5-98 Draft - Do Not Cite or Quote
-------
Female
900n
Basal
Final
PbxS
600-
O)
300-
900
Final
600
300
0 50 150 0 50 150
Pb Exposure (ppm)
so iso o so iso
Pb Exposure (ppm)
Note: Corticosterone levels are noted on the y-axis. 'denotes significantly different from NS control (Black bars); # denotes
significantly different from corresponding Pb-NS value; + differs from 50-NS. (0, 50, 150 ppm) and/or stress (PS [dam stress, white
bar with black dots] or OS [PS followed by offspring stress, black bar with white spots]). Basal measurements were taken at
2 months of age, prior to the initiation of behavioral testing and final measurements were taken at 10 months, after behavioral
testing.
Source: Rossi-George et al. (2011)
Figure 5-5 Mean basal and final corticosterone levels of female and male
offspring exposed to lifetime Pb.
1 Pb-stress effects on corticosterone have not been consistent. In another study, gestational
2 plus lactational Pb (50, 100 ppm) did not affect baseline corticosterone levels in females
3 and there was no interaction with stress (Virgolini et al., 2008a). In males, stress
4 increased baseline corticosterone in the 150 ppm Pb group. In animals given intermittent
5 stress as adults and not behaviorally tested, Pb decreased corticosterone levels in females
6 but not males, which may explain the observations of Pb plus stress increases in
7 decreases in FI response rates in females (Virgolini et al., 2008b). Post-weaning exposure
8 of male rodents to Pb (PND21-age 5 months) produced a U-shaped concentration-
9 response for corticosterone prior to FI testing, with a significant decrement in basal
10 corticosterone levels in the 50 ppm exposure group versus control and the 150 ppm Pb
11 group (Virgolini et al., 2005). Lifetime Pb exposure and prenatal stress both reduced
12 corticosterone in animals behaviorally tested at age 4 months but not in animals at age
13 11 months not behaviorally tested (Cory-Slechta et al., 2010).
14 Another study examined the effects of Pb acetate on the HPA axis but examined the
15 interaction with an outside stress administered using control vehicle injections (Rossi-
16 George et al., 2009). The corticosterone response to this vehicle injection stress was
November 2012
5-99
Draft - Do Not Cite or Quote
-------
1 prolonged in a nonlinear concentration-dependent manner in both sexes with the most
2 profound effects observed at the lower 50 ppm Pb dose. Maternal stress also prolonged
3 the corticosterone stress response to vehicle injection and enhanced the Pb effect in
4 males. To test the negative feedback of the HPA axis, exogenous dexamethasone (DEX)
5 was administered to suppress endogenous corticosterone. The DEX test revealed HPA
6 axis hypofunction. Specifically, Pb and Pb plus maternal stress initially reduced the
7 ability of DEX to suppress corticosterone. With time, the effect of DEX in males induced
8 prolonged corticosterone suppression or failure to return to baseline as was observed in
9 control animals. Rossi-George et al. (2011) additionally found that Pb and/or maternal
10 stress significantly impacted the negative feedback by increasing nuclear glucocorticoid
11 receptor levels. In summary, prenatal Pb exposure induced HPA negative feedback
12 hypofunction in both male and female offspring with an inverse U concentration-
13 response relationship. This negative feedback loop was impacted more at the lower Pb
14 dose (50 ppm) versus the higher dose (150 ppm) (Rossi-George et al., 2011).
15 To summarize the results for Pb-stress interactions in animals (Table 5-6). lifetime Pb
16 exposure when combined with stress was found to exacerbate learning impairments
17 compared with Pb exposure or stress alone, although not across all tests or Pb doses. The
18 interaction between Pb and stress may be mediated via effects on corticosterone and
19 dopamine. Lifetime Pb exposure was found to increase basal (at age 2 months)
20 corticosterone levels in females, and co-exposure to stress attenuated the response (Rossi-
21 George etal.. 2011; 2009). Males given lifetime Pb exposure had no statistically
22 significant corticosterone response to Pb exposure; whereas males with
23 gestational/lactational Pb exposure had statistically significant decreases in corticosterone
24 at 5 months of age in the 50 ppm exposure group only (but not in 150 ppm Pb exposure
25 group). On the other hand, females had concentration-dependent corticosterone responses
26 to Pb exposure in both exposure models (lifetime Pb exposure and gestational/lactational
27 Pb exposure). Maternal stress alone also led to HPA axis negative feedback hypofunction
28 in offspring. Pb exposure plus maternal stress enhanced negative feedback in males and
29 attenuated this effect in females. Pb exposure with or without maternal stress prolonged
30 the effect of DEX-dependent corticosterone suppression in males. These data together
31 show that HPA axis alterations could provide a link for interactions found between Pb
32 and stress in impairing learning.
November 2012 5-100 Draft - Do Not Cite or Quote
-------
5.3.2.4 Executive Function in Children
1 Epidemiologic evidence presented in the 2006 Pb AQCD (U.S. EPA. 2006b) indicated
2 associations between higher blood or tooth Pb levels and poorer performance on tests of
3 executive function in children and young adults. Associations were found with indices of
4 executive function such as strategic planning, organized search, flexibility of thought and
5 action to a change in situation, and control of impulses (described in greater detail in
6 Section 5.3.3.1). Prospective analyses in two Boston area and the Rochester cohorts
7 provided key evidence with examination of blood Pb levels preceding executive function
8 testing and adjustment for several potential confounding factors (Canfield et al.. 2004;
9 2003b: Bellinger et al.. 1994a: Stiles and Bellinger. 1993). Further, recruitment of
10 participants before or at birth, moderate to high follow-up participation, and in most cases
11 follow-up not biased to higher blood Pb levels and lower cognitive function increase
12 confidence that the observed associations are not due to selection bias (Table 5-8).
13 Among the few recent cross-sectional studies, most found concurrent blood Pb-associated
14 decrements in executive function, including an analysis of the Rochester cohort
15 (Froehlich et al.. 2007). Evidence from other recent studies had weaker implications due
16 to the limited consideration of potential confounding (Nelson and Espy. 2009; Vega-
17 Dienstmaier et al.. 2006). Several of the prospective and cross-sectional studies
18 performed multiple tests of cognitive function, including executive function. However,
19 except for Stiles and Bellinger (1993). a consistent pattern of association was found
20 across the various tests performed. Thus, the evidence does not appear to be biased by
21 associations found by chance alone.
22 Studies in children found Pb-associated decreases in executive function using various
23 tests including the Intra-Extra Dimensional Set Shift, WCST, and Stoop test (Table 5-8).
24 As discussed below, studies in animals also demonstrated Pb-induced decrements in
25 executive function, including rule learning and reversal, which also were associated with
26 blood Pb levels in children. This coherence between findings in animals and humans for
27 analogous domains further supports a relationship between Pb exposure and decrements
28 in cognitive function. Additional biological plausibility for Pb-associated decrements in
29 executive function is provided by toxicological evidence for Pb-induced changes in the
30 availability of dopamine (Section 5.3.11). a neurotransmitter that affects executive
31 functions mediated by the prefrontal cortex. Recent work shows that executive function
32 in animals is affected by N-Methyl-D-aspartic acid or N-Methyl-D-aspartate (NMDA)
33 receptors and dopamine-like receptors (Herold. 2010). which are two well-characterized
34 targets of Pb.
November 2012 5-101 Draft - Do Not Cite or Quote
-------
1 In the Boston cohort, children tested at a relatively older age (10 years), when testing is
2 more reliable, a 1 (ig/dL increase in age 5 year blood Pb level was associated with 0.05
3 (95% CI: 0.01, 0.09) more perseverative errors on the WCST (errors in sorting cards
4 according to a change in rule) (Stiles and Bellinger. 1993). In this cohort, results were
5 inconsistent across the various cognitive tests. However, associations were more
6 consistent for executive functions assessed by the WCST. In another cohort ages
7 19-20 years from towns around Boston, higher tooth Pb levels (from ages 5-8 years) were
8 associated with more errors on the WCST in sorting by the set rules and poorer
9 performance on the Stroop Color and Color-word tests, which test the ability of subjects
10 to shift focus to another dimension of stimulus that defines correct responding (Bellinger
11 etal.. 1994a).
November 2012 5-102 Draft - Do Not Cite or Quote
-------
Table 5-8 Associations between blood or tooth Pb levels and performance of
tests of executive function in children and young adults.
Study
Stiles
and
Bellinger
I A QQO\
(1 b)b)O)
Bellinger
etal.
(1994a)
Canfield
etal.
(2004)
Froehlich
etal.
(2007)
Canfield
etal.
(2003b)
Study Population and Methodological
Details
(Presented first for prospective studies then for
cross-sectional studies. Within each category,
results are presented in order of strength of study
design)
148 children followed from birth (1979-1981) to age
1 0 yr, Boston, MA area
Prospective. Recruitment at birth hospital.
Moderate follow-up participation, participants had
higher SES and HOME score. Linear regression
model adjusted for HOME score, family stress,
race, marital status (5 yr blood Pb), HOME score,
family stress, maternal age and race, birth weight,
# daycare situations to age 57 mo (concurrent).
79 young adults, born 1970, followed from 1st grade
to age 19-20 yr, Boston, MA area
Prospective. Moderate follow-up participation.
Participation from higher SES, females, higher initial
IQ but no affect on association with tooth Pb level.
Regression model adjusted for parental IQ, sex,
SES, current drug, alcohol and illicit drug use,
maternal education and age, birth order. Also
considered potential confounding by other
unspecified factors.
174 children born 1994-1995 followed from age
6 mo to 5 yr, Rochester, NY
Prospective. Recruitment from study of dust
control. 73% nonwhite. High follow-up participation,
no selective attrition. Linear regression model
adjusted for NICU admission, HOME, prenatal
maternal smoking, household income, child sex,
average crowding in home (I ED); maternal IQ,
HOME, prenatal smoking, household income, child
sex (Stockings of Cambridge). Also considered
potential confounding by breastfeeding duration,
maternal ethnicity, first prenatal visit, spatial working
memory problem, age at testing, birth weight,
marital status, maternal education and spatial span
length
174 children born 1994-1995 followed from age
6 mo to age 5 yr, Rochester, NY
Cross-sectional. Same cohort as above. High
follow-up participation, no selective attrition. Linear
regression model adjusted for NICU, sex (rule
learning). Also considered potential confounding by
income, HOME score, maternal IQ and education,
prenatal smoking exposure, race, and age,
transferrin saturation.
150 children born 1994-1995 followed from age
6 mo to age 4.5 yr, Rochester, NY
Cross-sectional. Same cohort as above. High
follow-up participation, no comparison of
nonparticipants. Linear mixed effects model
adjusted for: child sex, maternal IQ, education, and
prenatal smoking, household income, marital status,
HOME score. Also considered potential
confounding by age, birth order, attention rating,
race, gestational age, color/shape knowledge, child
IQ
Blood Pb
Levels
(ug/dL)
Earlier
childhood
(age 5 yr)
Concurrent
mean <8
Exact levels
NR mean
reported to
be<8
Deciduous
tooth (age
5-8 yr)
Mean (SD):
13.7
(11.1)ug/g
10th-90th:
4.3-26.4
Lifetime (to
age 5 yr) avg
Mean (SD):
7.2 (3.6)
10th-90th:
3.5-11.8
Concurrent
Mean (SD):
6.1 (4.9)
10th-90th:
1.9-11 .7
Concurrent
Mean: 6.5
10th-90th:
data not
available
Executive
Function Test
Perseverative errors,
Age 5 yr blood Pb
Concurrent blood Pb
Mean time to
complete color-word
test, Stroop test
Perseverative
responses, WCST
Ages 1 9-20 yr
Stages Completed -
Intra-Extra
Dimensional (IED)
Ortt OKift
bet bhitt
Stockings of
Cambridge problems
solved in minimum
moves
CANTAB
Age 5 yr
Stages Completed -
Intra-Extra
Dimensional Set
C Kift
onlTt
CANTAB
Aae 5 vr
nyc \j yi
Inhibit Efficiency
(# correct-
incorrect)/phase
duration
Shape School Task
Repeated measures
at ages 4 and 4.5 yr
Effect Estimate
/nco/ /^i\3
(95% Cl)
WCST, Age 10yr
-0.05 (-0.09, -0.01 )b
-0.05 (-0.11,0.01)"
-0.68 (0.28, 1 .08)b
-0.37(0.10,0.64)"
-0.11 (-0.21, -0.01)
-0.08 (-0.1 7, 0.001)
-0.06 (-0.1 2,0)°
-0.01 9 (-0.03, -0.007)
November 2012
5-103
Draft - Do Not Cite or Quote
-------
Study
Surkan
etal.
(2007)
Study Population and Methodological
Details
(Presented first for prospective studies then for
cross-sectional studies. Within each category,
results are presented in order of strength of study
design)
389 children ages 6-10 yr, Boston, MA ,Farmington,
ME
Cross-sectional. Recruitment from trial of
Blood Pb
Levels Executive
(ug/dL) Function Test
Concurrent
Mean (SD):
2.2(1.6)
Effect Estimate
(95% Cl)a
Blood Pb level 5-10
vs. 1-2 ug/dLd
amalgam fillings. High participation rate. Higher
participation of white children in Maine. Analysis of
covariance adjusted forcaregiver IQ, child age,
SES, race, and birth weight, Also considered
potential confounding by caregiver education and
marital status, parenting stress, and maternal
utilization of prenatal or annual health care but not
parental caregiving quality.
rciacvciauuii ciiuia, a 1 a / 1/I c -3 7\
WCST -9.19 (-14.6,-3.7)
Stroop color-word
interference
0.75 (-1.6, 3.1)
Chiodo
etal.
(2004)
Choet
al.
(2010)
246 children, age 7.5 yr, Detroit, Ml area
Cross-sectional. Recruitment at prenatal clinic. All
African American High prevalence of prenatal
alcohol exposure. High participation rate. Log linear
regression model adjusted for SES, family
functioning, # children <1 8 yr, caregiver vocabulary,
prenatal alcohol use, caregiver education, child sex.
Also considered potential confounding by HOME,
maternal prenatal marijuana, smoking, or cocaine
use, crowding, child life stress, caregiver age, life
stress, and psychology, conflict tactics, disruption in
caregiver, parity, child age.
667 children ages 8-1 1 yr, born 1 997-2000, 5
Korean cities
Cross-sectional. School-based recruitment,
moderate participation rate. Log linear regression
model adjusted for age, sex, parental education,
maternal IQ, child IQ, birth weight, urinary cotinine.
Did not consider potential confounding by parental
caregiving quality.
Concurrent
Mean (SD):
5.4 (3.3)
Interval
analyzed:
2.3-9.5= 10t
h-90th
percentiles
Concurrent
Mean (SD):
1.9(0.67)
Interval
analyzed:
1.2-2.8=101
h-90th
percentiles
Perseverative errors,
WCST
Age 7.5 yr
Color-word score
Stroop test
Ages 8-1 1 yr
-0.49, p >0.05e
0 (-0.09, 0.08)
Note: Results are presented first for prospective studies then for cross-sectional studies. Within each category, results are
presented in order to strength of study design.
WCST = Wisconsin Card Sorting Test, CANTAB = Cambridge Neuropsychological Testing Automated Battery.
"Effect estimates are standardized to a 1 ug/dL increase in blood Pb level or 1 ug/g in tooth Pb level in the 10th-90th percentile
interval.
bThe direction of the effect estimate was changed such that a negative estimate represents poorer performance.
°95% Cl: was constructed using a standard error that was estimated from the reported p-value.
dEffect estimates compare test performance of children in higher blood Pb groups to children in lowest blood Pb group.
Sufficient data were not provided to calculate 95% Cl.
1
2
3
4
5
6
7
8
9
10
11
12
Results from the Rochester cohort at ages 4 and 5 years indicated associations of
concurrent and lifetime average blood Pb level with lower inhibition efficiency in the
Shape School task (i.e., giving correct responses and withholding incorrect responses)
(Canfield et al.. 2003b). poorer problem solving on a spatial planning task (Canfield et
al.. 2004). and poorer rule learning and reversal (Froehlich et al.. 2007). Associations
with Shape School tasks were attenuated and lost precision with adjustment for attention
ratings, color/shape knowledge, and child IQ. These results suggest that the effect of Pb
exposure on executive function may be mediated through effects on knowledge.
Froehlich et al. (2007) found a larger concurrent blood Pb-associated decrement in a rule
learning and reversal task in the Intra-Extra Dimensional Set Shift in children with the
DRD4 exon III 7-repeat microsatellite (assessed using a blood Pb-DRD4-7 interaction
term, p = 0.042). While this evidence for effect modification is based on a smaller subset
November 2012
5-104
Draft - Do Not Cite or Quote
-------
1 of subjects (n = 34/174), they add support for Pb-associated decreases in executive
2 function because dopamine is a key neurotransmitter that regulates executive function,
3 the DRD4-7 variant is associated with reduced dopamine-induced signaling in
4 downstream pathways (e.g., cyclic AMP), and the DRD4-7 variant was associated with
5 poorer executive function in this cohort. The association of concurrent blood Pb level
6 with impaired rule learning and reversal also was greater in boys, who had lower mean
7 scores than girls.
8 In addition to assessment of earlier or cumulative Pb biomarkers, a strength of the
9 prospective studies was the consideration for numerous potential confounding factors.
10 The potential confounding factors varied among studies based on their association with
11 executive function and/or influence on the Pb-executive function relationship. Some
12 prospective studies demonstrated Pb-associated decrements in executive function with
13 adjustment for SES, maternal IQ, and HOME score (Canfield et al., 2004; Stiles and
14 Bellinger. 1993). Others considered and excluded potential confounding by HOME score,
15 parental smoking, maternal education, or birth outcomes (Table 5-8).
16 The prospective studies indicated blood Pb-associated decrements in executive function
17 in populations with a mean lifetime average blood Pb level of 7.2 (ig/dL and a mean
18 concurrent blood Pb level of 6.5 (ig/dL. Associations in populations with lower mean
19 blood Pb levels (2-5 (ig/dL), as assessed in cross-sectional studies with concurrent blood
20 Pb level, were not as clearly demonstrated. While these studies adjusted for SES and
21 parental cognitive function, most did not examine potential confounding by parental
22 caregiving quality, i.e., HOME score. Among children in New England ages 6-10 years
23 with mean concurrent blood Pb level 2.2 (SD: 1.6) (ig/dL, Pb-associated decrements in
24 executive function assessed by WCST, Trail-making, and Verbal cancellation tests were
25 observed primarily in the group with blood Pb levels 5-10 (ig/dL. In this study, higher
26 blood Pb level was not associated with poorer color-word score in the Stroop test. Cho
27 et al. (2010) did not find a Pb-associated lower color-word score among children in five
28 Korean cities ages 8-11 years with mean concurrent blood Pb level 1.9 (SD: 0.7) (ig/dL.
29 Other studies found associations in children with mean blood Pb levels 2-5 (ig/dL but had
30 limited implications because of lack of representativeness of a population with high
31 prevalence of prenatal alcohol exposure (Chiodo et al.. 2004) or lack of consideration for
32 potential confounding (Nelson and Espy. 2009).
33 The associations observed in children between blood or tooth Pb levels and poorer
34 executive function as assessed by the rule learning and reversal components of the Intra-
35 Extra Dimensional Set Shift, Stroop Test, and WCST are supported by observations in
36 animals of Pb-induced impairment in analogous measures of cognitive flexibility, tested
37 with discrimination reversal learning and concurrent random interval (RI-RI) scheduling.
November 2012 5-105 Draft - Do Not Cite or Quote
-------
1 In animals, some of these observations were made with Pb exposures relevant to humans.
2 These tests of cognitive flexibility measure the ability of humans and animals to adjust
3 their responses in reaction to changes in reinforcement. Poorer performance in both
4 children and animals is indicated by increased response errors, decreased percent correct
5 responses, and perseverative responding (e.g., persistence in making a previously-
6 rewarded response after a new shift in reinforcement). As reviewed in the
7 2006 Pb AQCD (U.S. EPA. 2006b). several lines of evidence indicated Pb-induced
8 impairments in executive function in animals. Lifetime dietary Pb exposures beginning at
9 birth or after weaning that produced peak blood Pb levels of 19-36 (ig/dL were found to
10 induce poorer performance on discrimination reversal learning tasks in monkeys ages
11 5-10 years (Rice and Gilbert. 1990b: Gilbert and Rice. 1987). Recent work has shown
12 that discrimination reversal learning involves NMDA receptors and dopamine-like
13 receptors (Herold. 2010). which are two well-characterized targets of Pb. Gestational Pb
14 exposure (blood Pb of dams >40 (ig/dL) was found to impair cognitive flexibility in
15 squirrel monkeys, ages 5-6 years, as indicated by a slower shift or lack of a shift to the
16 lever reinforced more frequently under RI-RI scheduling (Newland et al., 1994). Rats
17 also showed Pb-induced impairments on discrimination reversal tasks, but the authors
18 attributed the changes to learning-related problems instead of impaired executive function
19 (Garavan et al.. 2000: Hilson and Strupp. 1997V
5.3.2.5 Academic Performance and Achievement in Children
20 As described in preceding sections, a large body of evidence demonstrates Pb-associated
21 decrements in FSIQ, with more variable findings for performance on tests of learning and
22 memory. Lower FSIQ and learning are linked with poorer academic performance and
23 achievement, which may have important implications for success later in life. Further,
24 academic performance may better assess the knowledge of an individual in the actual
25 subject areas studied, whereas aptitude tests are used to predict future performance. In
26 addition to FSIQ, the 2006 Pb AQCD described associations between blood Pb levels in
27 children ages 5-18 years and poorer performance on tests of math and reading skills,
28 vocabulary, and spelling, objective measures such as high school completion and class
29 rank, and teacher ratings of academic functioning. Associations continued to be reported
30 in recent studies, including prospective studies examining performance on academic
31 achievement tests and an additional analysis of adolescents participating in NHANES
32 (Table 5-9). Findings from other recent studies had weaker implications because of the
33 lack of representativeness of populations with high prevalence of prenatal alcohol or drug
34 exposure (Min et al.. 2009; Chiodo et al.. 2007). Multiple testing was common in studies;
35 however, the consistent pattern of blood Pb-associated decrements in academic
November 2012 5-106 Draft - Do Not Cite or Quote
-------
1 performance across the various tests conducted increases confidence that the evidence is
2 not unduly biased by a higher probability of associations found by chance alone.
3 Key evidence supporting associations between blood Pb level and performance on tests
4 of quantitative, reading, vocabulary, and spelling skills was provided by previous
5 prospective studies in the Boston and Cincinnati cohorts (Ris et al.. 2004; Bellinger etal..
6 1991). Associations with earlier childhood blood Pb levels better characterized the
7 temporal sequence between Pb exposure and poorer academic performance. Evidence
8 from prospective studies did not strongly indicate selection bias with recruitment of
9 participants before or at birth, moderate to high follow-up participation, and in most cases
10 follow-up not biased to higher blood Pb levels and lower cognitive function (Table 5-9).
11 An additional strength of the prospective studies was the consideration of several
12 potential confounding factors (Table 5-9). including birth outcomes, exposure to smoking
13 and drugs, and nutritional status and the adjustment for SES, parental education and IQ,
14 and HOME score. Evidence for associations between blood Pb levels and reading, math,
15 and vocabulary skills provides coherence for the associations observed between blood Pb
16 levels and FSIQ, which includes components of quantitative reasoning and language
17 ability.
November 2012 5-107 Draft - Do Not Cite or Quote
-------
Table 5-9 Associations between blood or tooth Pb levels and measures of
academic performance and achievement in children and young
adults.
Study
Study Population and
Methodological Details
(Presented first for prospective studies then
for cross-sectional studies. Within each
category, results are presented in order to
strength of study design)
Blood or
Tooth Pb
Levels
(ug/dL)
Indicator of
Academic
Performance/
Achievement
Effect Estimate
(95% Cl)a
Studies of neuropsychological testing of academic performance
Bellinger et al.
(1991)
170 children followed from birth (1979-1981)
to age 5 yr, Boston, MA area
Prospective. Recruitment at birth hospital.
Moderate follow-up participation. More
participants were white, had higher age 2 yr
HOME score, and higher postnatal blood Pb
levels. Log linear regression model adjusted
for SES, maternal IQ and marital status,
preschool attendance, HOME, out of home
care, residence changes, medication use in
previous 12 mo, number of adults in home,
child sex, race, birth weight, birth order.
Earlier
childhood
(age 2 yr)
Mean (SD):
7.0 (6.6)
Interval
analyzed:
1.8 (1 Oth
percentile)-
10
Verbal
-0.09 (-0.51, 0.34)
Quantitative
McCarthy Scale of
Children's Abilities
Age 5 yr
-0.30 (-0.65, 0.05)
Dietrich et al. 258 children followed prenatally (1979-1985)
(1991) to age 4 yr, Cincinnati, OH
Prospective. Recruitment at prenatal clinic.
High follow-up participation, no selective
attrition. Mostly African American. Linear
regression adjusted for SES, birth weight,
maternal IQ, prenatal marijuana use, HOME,
child race, preschool attendance. Also
considered potential confounding by birth
outcomes, maternal age, prenatal smoking,
alcohol use and narcotics use, # previous
abortions, stillbirths, gravidity, parity,
caregiver education, public assistance, child
age, sex, health, Fe status
Earlier
childhood
(Age 2 yr):
NR
Achievement score,
KABC
0.06, p >0.05b
Concurrent:
NR
0.01,p>0.05b
Lifetime
avg: NR
0.07, p >0.05b
Ris et al. (2004) 195 children followed prenatally (1979-1985)
to age 15-17 yr, Cincinnati, OH
Prospective. Same cohort as above. High
follow-up participation, no selective attrition.
Linear regression adjusted for SES, maternal
IQ, HOME, adolescent marijuana use, and
obstetrical complications. Also considered
potential confounding by birth outcomes,
maternal age, prenatal smoking, alcohol,
marijuana, and narcotics use, # previous
abortions, stillbirths, gravidity, parity,
caregiver education, public assistance, child
age, sex, health, Fe status
Earlier
childhood
(age 6.5 yr)
Mean (SD):
NR
Reading, spelling,
math,
vocabulary
composite
WRAT-III and
WISC-III
Ages 15-17 yr
-0.081 (-0.17, 0.003)
Chandramouli 488 children followed from age 30 mo (born
et al. (2009) 1991 -1992) to 7-8 yr, Avon, U. K.
Prospective. All births in area eligible.
Similar characteristics as U.K. census, high
participation at baseline and follow-up.
Participants had better educated mothers,
who smoked less, better home environment.
Regression model adjusted for maternal
education and smoking, home ownership,
home facilities score, family adversity index,
paternal SES, parenting attitudes at 6 mo,
child sex. Also considered potential
confounding by child IQ.
Age 30 mo
Mean (SD):
NR
Group 1:
0-2
Group2: 2-5
Group 3:
5-10
Group 4:
Standardized
Achievement Test
Ages 7-8 yr
Per doubling blood Pb
-0.3 (-0.5,-0.1)°
November 2012
5-108
Draft - Do Not Cite or Quote
-------
Study
Miranda et al.
(2009)
Lanphear et al.
(2000)
Krieg et al.
(2010)
Surkan et al.
(2007)
Kordas et al.
(2006)
Study Population and
Methodological Details
(Presented first for prospective studies then
for cross-sectional studies. Within each
category, results are presented in order to
strength of study design)
57,568 children, 4th grade, all counties, NC
Prospective: based on data from
surveillance databases. Quantile regression
adjusted for sex, age of blood Pb
measurement, race, enrollment in free lunch
program, parental education, charter school.
Did not consider potential confounding by
parental caregiving quality or cognitive
function.
4,852 children ages 6-16 yr (born
1972-1988), U.S. NHANES III (1988-1994)
Cross-sectional. Large U.S. representative
study of multiple risk factors and outcomes.
High, non-selective participation. Linear
regression model adjusted for sex,
race/ethnicity, poverty index ratio, reference
adult education, serum ferritin and cotinine
levels, Did not consider potential
confounding by parental cognitive function or
parental caregiving quality.
766-780 children ages 12-16 yr (born
1975-1982), U.S. NHANES III (1991-1994)
Cross-sectional. Large U.S. representative
study of multiple risk factors and outcomes.
Log linear regression model adjusted for sex,
caregiver education, family income, race-
ethnicity, test language. Did not consider
potential confounding by parental cognitive
function or caregiving quality.
389 children ages 6-10 yr, Boston, MA,
Farmington, ME
Cross-sectional. Recruitment from trial of
amalgam fillings. High participation rate.
Higher participation of white children in
Maine. Analysis of covariance adjusted for
caregiver IQ, child age, SES, race, and birth
weight, Also considered potential
confounding by caregiver education and
marital status, parenting stress, and
maternal utilization of prenatal or annual
health care but not parental caregiving
quality.
294 children, age 7 yr, Torreon, Mexico.
Cross-sectional. Recruitment at prenatal
clinic. High participation rate. Residence
near metal foundry. Linear regression model
adjusted for child sex, age, school, birth
order, hemoglobin, forgetting homework,
household possessions and crowding, house
ownership, maternal education, family
structure, urinary As, tester. Did not consider
potential confounding by parental cognitive
function or caregiving quality.
Blood or
Tooth Pb
Levels
(ug/dL)
Earlier
childhood
(ages 9-36
mo)
Median
(25th-75th):
4.8 (3-6)
Concurrent
Geometric
mean: 1.9
(5th-95th:
1.70, 2.10)
Interval
analyzed:
1 .74-2.06
= 1 0th-90th
percentile
Concurrent
Mean
(5th-95th):
1.95
(1 .63-2.27)
Interval
analyzed:
1.69-2.19 =
10th-90th
percentile
Concurrent
Mean (SD):
2.2(1.6)
Concurrent
Geometric
mean
(range):
10.2
(2-43.8)
Interval
analyzed:
2.1-10.0
Indicator of
Academic
Performance/
Achievement
Reading
4th grade end-of-
grade test score
Math score
Reading score
WRAT-R
Ages 6-1 6 yr
Math score
Reading score
WRAT-R
Ages 1 2-1 6 yr
Reading score
Math score
WIAT
Ages 6-1 0 yr
Math achievement
test
PPVT
Age 7 yr
Effect Estimate
(95% Cl)a
Score vs. blood Pb
1 ug/dL
2 ug/dL:
-0.30 (-0.58, -0.01 )d
3 ug/dL:
-0.46 (-0.73, -0.1 9)d
4 ug/dL:
-0.52 (-0.79, -0.24)d
5 ug/dL:
-0.80 (-1.08, -0.51 )d
-0.70 (-1.0, -0.37)
-0.99 (-1 .4, -0.62)
-2.5 (-4.5, -0.50)
-2.9 (-4.3, -1.5)
Blood Pb5-10 ug/dL
vs. 1-2:
-5.20 (-9.45, -0.95)d
-4.02 (-7.6, -0.43)
-0.42 (-0.92, 0.08)
-0.71 (-1 .4, 0.02)
November 2012
5-109
Draft - Do Not Cite or Quote
-------
Study
Study Population and
Methodological Details
(Presented first for prospective studies then Blood or Indicator of
for cross-sectional studies. Within each Tooth Pb Academic
category, results are presented in order to Levels Performance/
strength of study design) (ug/dL) Achievement
Effect Estimate
(95% Cl)a
Chiodo et al. 506 children, age 7 yr (born 1982-1984),
(2007) Detroit, Ml area.
Cross-sectional. All African American. High
prevalence of prenatal drug exposure. High
follow-up participation. Linear regression
model adjusted for caregiver education,
SES, HOME, maternal IQ, child sex, prenatal
marijuana use (all outcomes), Caregiver
concurrent psychological symptoms (Math),
Child age, maternal custody (Reading). Also
considered potential confounding by prenatal
cigarettes/day, alcohol use, cocaine use, #
children in home, caretaker marital status,
concurrent alcohol/week, current maternal
cigarettes/day, and current marijuana use.
Concurrent
Mean (SD):
5.0 (3.0)
Interval
analyzed:
2.1-8.7=10
th-90th
percentiles
Math
-0.17 (-0.27, -0.07)°
Reading
Metropolitan
Aptitude Test
Age 7 yr
-0.06, p>0.05b
Chiodo et al.
(2004)
246 children, age 7.5 yr, Detroit, Ml area Concurrent
Cross-sectional. Recruitment at prenatal Mean (SD):
clinic. All African American. High prevalence 5.4 (3.3)
of prenatal alcohol exposure. High Interval
participation rate. Log linear regression analyzed'
model adjusted for SES (all outcomes). 23-95 =10
HOME, caregiver vocabulary, prenatal th-90th
alcohol use (arithmetic). Caregiver percentiles
vocabulary, disruption in caregiver (verbal H
learning). Also considered potential
confounding by maternal prenatal marijuana,
smoking, or cocaine use, crowding, child life
stress, caregiver age, life stress, and
psychology, conflict tactics, family
functioning, # children <18 years, caregiver
education, child sex and age, parity.
Verbal learning
(WRAML)
-0.20, p>0.05b
Arithmetic (WISC-
III)
Age 7.5 yr
-0.1 7, p>0.05b
Min et al. (2009) 267 children, age 11 yr (born 1994-1996),
Cleveland, OH
Prospective, Recruitment at birth hospital.
86% African American with high prevalence
of prenatal drug and alcohol exposure.
Moderate follow-up participation to age 4 yr,
high retention to age 11 yr. Higher
participation from African American and
married mothers. Linear regression model
adjusted for HOME score, maternal birth
vocabulary score, head circumference at
birth (both outcomes), prenatal cocaine use
(math), child sex, prenatal cocaine and
alcohol use, current caregiver alcohol use
(reading age 11 yr). Also considered
potential confounding by maternal education,
Fe deficiency, maternal psychological
distress, race.
Earlier
childhood
(Age 4 yr)
Mean
(range): 7.0
(1.3-23.8)
Interval
analyzed:
3.0 (1 Oth
percentile)-
10
Math
-0.45 (-0.84, -0.06)
Reading
WJTA
Age 11 yr
-0.58 (-1.0, -0.13)
Studies of School Performance
Fergusson et al.
(1997)
881 children followed from birth to age 16-18
yr, Christchurch, New Zealand
Prospective. Moderate follow-up
participation, attrition did not affect results.
Regression model adjusted for maternal age,
punitiveness, standard of living,
breastfeeding duration, parental conflict,
class level, residence on busy roads. Also
considered potential confounding by sex,
ethnicity, maternal education, family size,
HOME, SES, # schools attended, ethnicity,
paternal education, parental smoking, child
birth outcomes, weatherboard housing.
Tooth Pb
(age 6-8 yr)
Mean (SD):
6.2
(3.7) ug/g
Percent leaving
school with no
qualifications
Age 16-18 yr
0-2 ug/g: 15.6
3-5 ug/g: 16.7
6-8 ug/g: 18.1
9-11 ug/g: 19.7
12+ug/g: 24.1
November 2012
5-110
Draft - Do Not Cite or Quote
-------
Study
Needleman et
al. (1990)
Study Population and
Methodological Details
(Presented first for prospective studies then
for cross-sectional studies. Within each
category, results are presented in order to
strength of study design)
132 young adults followed from 1st/2nd
grade to age 18 yr, Chelsea, Sommerville,
MA
Prospective. Recruitment at schools. Low
follow-up participation. Participants had
lower tooth Pb, higher parental education,
SES, maternal IQ. Participation status did
not alter tooth Pb-childhood IQ association.
Logistic regression adjusted for maternal age
at birth, education, and IQ, family size, sex,
age at testing, birth order, alcohol use,
mother and child left hospital together. Did
not examine potential confounding by
parental caregiving quality.
Blood or
Tooth Pb
Levels
(ug/dL)
Tooth Pb
(1st/2nd
grade)
distribution
<10 ppm,
50%
10-1 9.9 pp
m: 22.7%
>20 ppm:
27.3%
Indicator of
Academic
Performance/
Achievement
Failure to graduate
high school
Highest grade
achieved
Effect Estimate
(95% Cl)a
OR >10 ppm vs.
<10 ppm
7.4 (1 .4, 40.8)d
-0.03 (-0.05, 0) per
natural log increase in
tooth Pb
Study of teacher ratings of academic performance
Leviton et al.
(1993)
1923 children followed from birth
(1979-1980) to age 8 yr, Boston, MA
Prospective. Recruitment from birth
hospital. High participation at baseline and
follow-up. Regression model adjusted for
single parent family, gestational age,
maternal education, ethnicity, #
children, daycare in first 3 years. Also
considered potential confounding by other
unspecified factors.
Prenatal
(cord)
Mean: 6.8
Tooth Pb
(Age 6 yr)
Mean: 3.3
Reading, BTQ, Age
Syr
Prenatal (cord)
Tooth Pb
RR (yes/no) per loge
increase
Girls: 1.7(0.9, 3.3)
Boys: 1 .3 (0.8, 2.2)
Girls: 2.2 (1.1, 4.2)
Boys: 1 .2 (0.7, 2.2)
Note: Results are organized by method of outcome assessment then by prospective or cross-sectional design. Within each
category, results are presented in order to strength of study design.
KABC = Kaufman Assessment Battery for Children, WRAT = Wide Range Achievement Test, WISC = Wechsler Intelligence Scale
for Children, BTQ = Boston Teacher's Questionnaire, WIAT = Wechsler Individual Achievement Test, PPVT = Peabody Picture
Vocabulary Test, WRAML = Wide Range Assessment of Memory and Learning, WJTA = Woodcock Johnson-Ill Tests of
Achievement.
aEffect estimates are standardized to a 1 ug/dL increase in blood Pb level in the interval from the 10th percentile of blood Pb level
to the 90th percentile or 10 ug/dL, whichever is lower.
bSufficient data were not provided to calculate 95% Cl.
°95% Cl: was constructed using a standard error that was estimated from the reported p-value.
dEffect estimates compare test performance of children in higher blood Pb groups to children in lowest blood Pb group.
1
2
3
4
5
6
7
8
9
10
11
12
13
The ages at which associations between blood Pb level and performance on academic
achievement tests were found varied between prospective studies. In the Boston cohort
with lower blood Pb levels (mean: 7.0 (ig/dL), a 1 (ig/dL increase in age 2 year blood Pb
level was associated with a -0.30-point (95% Cl: -0.65, 0.05) change in quantitative skills
score age at 5 years in the blood Pb interval 1.8-10 (ig/dL (Bellinger et al.. 1991) with
adjustment for SES, maternal IQ and marital status, preschool attendance, HOME, out of
home care, residence changes, medication use in previous 12 months, number of adults in
home, child sex, race, birth weight, and birth order. Evidence did not strongly indicate an
association with verbal skills. In the Cincinnati cohort with higher blood Pb levels, age
6.5 year blood Pb level was associated with decrements in academic performance at ages
15-17 years and 5 years (Ris et al.. 2004; Dietrich et al.. 1992) but not 4 years (Dietrich et
al.. 1991). These differences within the Cincinnati cohort could be attributed to changes
in blood Pb levels overtime or age-related differences in reliability of tests of learning.
November 2012
5-111
Draft - Do Not Cite or Quote
-------
1 Recent prospective studies found associations for earlier childhood blood Pb levels but
2 did not have blood Pb measurements available at other time periods for comparison. The
3 records-based analysis (Miranda et al., 2009; 2007a), multi-factorial nature, or high
4 participation rate of recent studies (Chandramouli et al.. 2009) do not indicate a strong
5 influence of selection bias. Miranda et al. (2009; 2007a) linked higher blood Pb levels
6 measured at ages 0-5 years, as ascertained from a surveillance database, with lower end-
7 of-grade (EOG) test scores in 8,600 fourth grade children in seven of the largest counties
8 in North Carolina and then in 57,678 children in the entire state. A strength of the
9 analyses was the availability of individual-level data on a large number of children
10 representative of the North Carolina fourth grade population. The large numbers of
11 children with blood Pb levels 2-5 (ig/dL provided greater power to estimate the effects of
12 Pb in the lower range of blood Pb levels. In each analysis, children with an earlier blood
13 Pb level of 2 (ig/dL had lower EOG scores (p < 0.05) compared with children with a
14 blood Pb level of 1 (ig/dL. Further, across deciles of blood Pb level, the decrease in EOG
15 score generally was monotonic (Figure 5-6). Because these children were born in the
16 early- to mid-1990s and blood Pb levels were measured earlier in childhood, it is less
17 likely that associations were influenced by higher past Pb exposures.
18 Due to the records-based study design, investigators had a smaller set of potential
19 confounding factors available than those considered in the prospective studies described
20 above. Results were adjusted for sex, race, school-type, school district, age of blood Pb
21 measurement, parental education, participation in a free or reduced lunch program as a
22 measure of SES, and in the analysis of seven North Carolina counties, daily use of a
23 computer as a measure of a stimulating home environment (Miranda et al., 2007a). While
24 there may be no complete single measure of SES and parental caregiving quality, the
25 covariates examined in these analyses are not as well characterized, and the results may
26 be subject to residual confounding.
November 2012 5-112 Draft - Do Not Cite or Quote
-------
£-0,
u
-------
1
2
3
risk (lower parental education, enrollment in a school lunch program) had a greater
magnitude of negative association with EOG score, blood Pb level was independently
associated with EOG score decrements that were as large as 1 to 2 points.
« o
-2
I
8
W
n
UJ
c
• -8
I
8-10
o
-12
Cummulative Deficit: Decrease in EOG scores by multiple risk factors
5% 10% 15% 20% 25% 30% 35% 40% 45% 50% 55% 60% 65% 70% 75% 80% 85% 90% 95%
t
Baseline (BLL=lMg/dL, no school
lunch program, parents completed college)
Effect of reduction in parental education
> from completed college
to only completed high school
Income effect as indicated by
enrollment in school lunch program
} Effect of increased BLL from 1 to 5 pg/dL
Quantile
Note: Baseline score calculated for a hypothetical referent individual with a blood Pb level of 1 ug/dL, parents completed college,
and not enrolled in the school lunch program (i.e., model covariates = zero). Vertical bars indicate the decrease in EOG score
associated with blood Pb level and covariates in various percentiles of EOG score (lowest to highest, left to right). An increase in
earlier childhood blood Pb level is associated with a larger decrease in EOG score (larger black bars on left) among children in the
5th and 10th percentiles of EOG score than children in the 90th and 95th percentiles (smaller black bars on right).
Source: Reprinted with permission of Elsevier Science, Miranda et al. (2009)
Figure 5-7 Greater reduction in End-of-Grade (EOG) scores with increasing
blood Pb level in lower percentiles of the test score distribution.
4
5
6
1
8
9
10
11
12
Similar to Miranda et al. (2009). Chandramouli et al. (2009) found associations between
earlier childhood blood Pb levels (age 30 months) and later academic performance
(Standard Assessment Test [SAT] at age 7 years). In this study of 488 children in the
U.K., who had similar sociodemographic characteristics as those found in the U.K.
census, a doubling of age 30 month blood Pb level was associated with a 0.3-point (95%
CI: 0.1, 0.5) decline in SAT grade. Results were adjusted for maternal education and
smoking, home ownership, parental SES and several factors related to caregiving quality
including home facilities score, family adversity index, and parenting attitudes. In
analyses of blood Pb level categories, lower SAT scores were most clearly indicated in
November 2012
5-114
Draft - Do Not Cite or Quote
-------
1 children with age 30 month blood Pb levels >5 (ig/dL. Children with blood Pb levels
2 2-5 (ig/dL generally did not have lower SAT scores than children with blood Pb levels
3 0-2 ng/dL.
4 Consistent with prospective studies, cross-sectional studies found associations between
5 higher concurrent blood Pb level and lower scores on tests of math and reading, including
6 large studies of children participating in NHANES. While cross-sectional studies
7 considered potential confounding by SES and caregiver education, few considered
8 parental cognitive function, and none considered parental caregiving quality. Lanphear et
9 al (2000) and Krieg et al. (2010) found concurrent blood Pb-associated decrements in
10 math and reading score among 4,852 children ages 6-16 years and 766-780 children ages
11 12-16, respectively, participating in NHANES. The examination of multiple exposures
12 and outcomes in NHANES increases confidence that associations are not unduly
13 influenced by selection bias. While the mean blood Pb levels were low in these study
14 populations, ~2 (ig/dL, the influence of higher past Pb exposures on findings cannot be
15 excluded. Consistent with studies of FSIQ, Lanphear et al. (2000) found a supralinear
16 concentration-response relationship. A 1 (ig/dL increase in concurrent blood Pb level was
17 associated with a change in reading score of-0.70-points (95% CI: -1.0, -0.37) among all
18 subjects and -1.1-points (95% CI: -1.54, -0.58) among the 4, 043 children with blood Pb
19 levels <5 (ig/dL. A supralinear concentration-response relationship also was found in
20 children ages 7 years in Mexico living near a metal foundry as indicated by larger blood
21 Pb-associated decrements in math and vocabulary scores among children with concurrent
22 blood Pb levels <10 (ig/dL (Kordas et al.. 2006). In contrast with these findings, among
23 children ages 6-10 years in New England decrements in reading and math scores were
24 found in association with higher blood Pb levels, i.e., blood Pb levels 5-10 (ig/dL
25 compared with blood Pb levels 0-2 (ig/dL (Surkan et al.. 2007).
26 Prospective studies in a Boston, MA area cohort and New Zealand cohort found
27 associations of tooth Pb levels measured at an earlier age (ages 6-8 years) with school
28 performance ascertained at ages 16-18 years from school records (Fergusson et al.. 1997,
29 1993; Needleman et al.. 1990). suggesting the effect of early exposure to Pb may be
30 persistent. In the New Zealand cohort at ages 12-13 and 18 years, recruitment rate and
31 follow-up participation were high, and model correction for nonrandom sample attrition
32 produced robust results, indicating lack of undue selection bias (Fergusson et al.. 1997.
33 1993). Further, associations observed at ages 12-13 years between higher tooth Pb level
34 and lower teacher ratings of math, reading, and writing abilities (Fergusson et al.. 1993).
35 which are subject to greater measurement error, were supported by associations observed
36 at age 18 years with more objective measures such as lower probability of completion of
37 high school and lower scores on school exams (Fergusson et al.. 1997). In this cohort,
38 Pb-associated decrements in school performance were found with consideration for
November 2012 5-115 Draft - Do Not Cite or Quote
-------
1 potential confounding by several factors including SES, parental education, HOME
2 score, sex, ethnicity, number of school changes, perinatal history, breastfeeding, maternal
3 age, and residence in weatherboard housing and near busy roads.
4 In one Boston-area cohort, age 6-8 years tooth Pb level >20 (ig/g was associated with
5 dropping out of high school at age 18 years with an odds ratio of 7.4 (95% CI: 1.4, 40.7)
6 (Needleman et al.. 1990). The relatively small sample size (n = 132) and adjustment for
7 several potential confounding factors, including maternal education, IQ, and age, SES,
8 and subject alcohol use may have contributed to the imprecision of the effect estimate.
9 Parental caregiving quality was not considered. Participation was biased to children with
10 lower tooth Pb levels and higher SES. This selection bias likely did not produce a
11 spurious association; however, the results may be less generalizable to the original study
12 population. In another Boston-area cohort, higher tooth Pb level at age 6 years was
13 associated with higher teacher ratings of age 8 year spelling and reading difficulties in
14 girls but not boys (Leviton et al.. 1993). Despite the large sample size (n = 1923) and
15 high follow-up participation, the study did adjust for SES or parental caregiving quality.
16 However, other unspecified potential confounding factors were considered.
5.3.2.6 Integrated Summary of Cognitive Function in Children
17 Results from recent epidemiologic and animal studies add to the strong evidence base
18 reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b) demonstrating that Pb exposure is
19 associated with decrements in cognitive function in children, based on associations
20 observed with FSIQ, and also executive function, and academic performance.
21 Associations with performance on tests of learning and memory were less consistently
22 found (Table 5-5). A large epidemiologic evidence base demonstrates associations of
23 higher blood Pb level with lower FSIQ in school-aged children (Figure 5-2 and Table
24 5-3). with smaller bodies of evidence indicating associations with lower scores on tests of
25 executive function, and academic performance in children ages 4 to 18 years (Table 5-8
26 and Table 5-9). There was no clear indication that blood Pb level was more strongly
27 associated with performance in a particular domain of cognitive function. The
28 Pb-associated decrements in cognitive function observed in children were strongly
29 supported by observations in animals of decrements in learning, memory, and executive
30 function with relevant dietary Pb exposures. In particular, coherence was found between
31 observations of Pb-associated decrements in performance on spatial span tasks in
32 children and Morris water maze in animals both of which test visual spatial memory on
33 spatial working memory tasks in children and the radial arm maze in animals
34 (Section 5.3.2.3). Coherence also was found with Pb-associated changes in performance
35 on tests of rule learning and reversal in humans and animals (Section 5.3.2.4) both of
November 2012 5-116 Draft - Do Not Cite or Quote
-------
1 which assess cognitive flexibility. Additional biological plausibility for Pb-associated
2 cognitive function decrements was provided by toxicological evidence for the effects of
3 Pb on neurophysiological and neurochemical processes that mediate cognition.
4 Compelling epidemiologic evidence for Pb-associated cognitive function decrements was
5 described in the 2006 Pb AQCD (U.S. EPA. 2006b) for FSIQ (see Section 5.3.2.1 of this
6 ISA also). Across studies, FSIQ was measured with similar instruments (i.e., WISC-R,
7 WISC-III, WPPSI, Stanford-Binet) scored on a similar scale with similar measurement
8 error. Associations were found in most of the prospective studies, conducted in the U.S.,
9 Mexico, Europe, and Australia, in representative populations with high follow-up
10 participation without indication of selective participation among children with higher
11 blood Pb levels and lower cognitive function (Figure 5-2 and Table 5-3) that could
12 produce spurious associations. The prospective studies found associations of blood Pb
13 levels measured concurrently with FSIQ (ages 4-17 years) and earlier in life (i.e., prenatal
14 cord or maternal, age 2 year) or averaged over multiple years, better establishing the
15 temporal sequence between Pb exposure and cognitive function decrements than cross-
16 sectional studies, multiple testing was common; however, the consistent pattern of
17 association observed across the ages of blood Pb level and/or cognitive test examined in
18 most previous and recent studies increases confidence that the evidence is not unduly
19 biased by a higher probability of associations found by chance alone. Another strength of
20 the prospective evidence was the consideration of several potential confounding factors.
21 As indicated in Table 5-3. results from most cohorts were adjusted for maternal IQ and
22 education, child sex and birth weight, SES, and HOME score. Although not considered as
23 frequently, some studies also indicated lack of confounding by parental smoking, birth
24 order, and nutritional status. The robustness of the blood Pb-FSIQ association in children
25 was substantiated in a pooled analysis of seven prospective studies by Lanphear et al.
26 (2005) as well as multiple meta-analyses that combined results across various prospective
27 and cross-sectional studies (Pocock et al., 1994; Schwartz, 1994; Needleman and
28 Gatsonis. 1990). with Schwartz (1994) demonstrating the robustness of evidence to
29 potential publication bias.
30 Comparisons of effect estimates across studies are difficult because of the variability in
31 population blood Pb distributions, lifestage of blood Pb level examined, type of model
32 examined (linear versus nonlinear), and tests conducted. The pooled analysis of seven
33 prospective cohorts demonstrated precision of effect estimates by finding a relatively
34 narrow range of effect estimates, -2.36 to -2.94 points per natural log increase in blood Pb
35 level, excluding one study at a time (Lanphear et al.. 2005). In a linear model, a greater
36 decrease in FSIQ estimated for a 1 (ig/dL increase in concurrent blood Pb for the 244
37 children who had peak blood Pb levels <10 (ig/dL (-0.80 points [95% CI: -1.74, -0.14])
38 and the 103 children with peak blood Pb levels <7.5 (ig/dL (-2.9 points [95% CI: -5.2,
November 2012 5-117 Draft - Do Not Cite or Quote
-------
1 -0.71]). Among children with peak blood Pb levels <10 (ig/dL and <7.5 (ig/dL, the
2 median concurrent blood Pb levels were 4.2 (ig/dL and 3.2 (ig/dL, respectively (Hornung.
3 2008). Among individual studies, a wide range of effect estimates was reported.
4 However, studies varied in model specification and the blood Pb level range examined.
5 Similarly large effects were estimated in the Boston and Rochester cohorts, which
6 differed widely in racial and SES distributions (Canfield et al.. 2003a: Bellinger et al..
7 1992). While the sample sizes were smaller, these studies had least as extensive
8 consideration for potential confounding as other studies. Further, each study estimated
9 larger effects for children whose peak blood Pb levels never exceeded 10 (ig/dL, -1.8
10 points (95% CI: -3.0, -0.60) per 1 (ig/dL increase in concurrent blood Pb level in the
11 Rochester cohort (n = 101, 59%) (Canfield et al.. 2003a) and -1.6 points (95% CI: -2.9,
12 -0.2) per 1 (ig/dL in age 2-year blood Pb level in the Boston cohort (n = 48, 32%)
13 (Bellinger and Needleman. 2003). These subsets of children had mean blood Pb levels of
14 3.3 (Rochester) and 3.8 (ig/dL (Boston). Some recent cross-sectional studies estimated
15 smaller effects but with examination of populations with higher concurrent blood Pb
16 levels (means: 7, 8.7 (ig/dL) using a linear model (Kordas et al.. 2011; Min et al.. 2009).
17 Other recent studies estimated similar effects as previous studies although the log-linear
18 models make comparisons difficult. Among children ages 3-7 years in India, a 1 (ig/dL
19 increase in concurrent blood Pb level was associated with a 1.2-point decrease (95% CI:
20 -1.9, -0.37) in FSIQ from the 10th percentile of blood Pb level 5.8 to 10 (ig/dL (Roy et
21 al.. 2011). Kim et al. (2009b) found that a 1 (ig/dL increase in concurrent blood Pb level
22 was associated with a 3.2-point decrease (95% CI: -6.1, -0.23) in FSIQ among children
23 ages 8-11 years in Korea with blood Mn levels >1.4 (ig/dL in the 10th-90th percentile
24 interval of blood Pb level (0.9-2.8 (ig/dL). In this study, the potential influence of higher
25 past Pb exposures cannot be excluded. Further, while these recent studies adjusted for
26 parental education and SES, parental caregiving quality was not examined. The relatively
27 low blood Pb levels in the Rochester and Boston cohorts, consideration of peak blood Pb
28 levels, and the adjustment for several potential confounding factors indicate that their
29 results may be more representative.
30 Previous prospective studies, several of which contributed to the FSIQ evidence,
31 provided key evidence for associations of blood or tooth Pb level with decrements in
32 executive function and academic performance for the reasons described for FSIQ.
33 Endpoints associated with blood or tooth Pb level included rule learning and reversal,
34 reading and math skills assessed using neuropsychological tests and school performance
35 assessed from school records. Higher concurrent blood Pb level was associated with
36 lower scores on tests of math and reading in the large study of children participating in
37 NHANES (Lanphear et al.. 2000). Recent studies conducted in the U.S., Mexico, Europe,
38 and Asia, most of which were cross-sectional, also found associations between higher
39 blood Pb level and lower cognitive function. The few recent prospective studies indicated
November 2012 5-118 Draft - Do Not Cite or Quote
-------
1 associations between higher earlier childhood blood Pb level, ages 9-36 month and age
2 30 months, respectively, with poorer academic performance in children in North Carolina
3 at age 9 years (Miranda et al.. 2009) and in children ages 7 years in the U.K.
4 (Chandramouli et al.. 2009).
5 In most studies that provided unadjusted and adjusted effect estimates, blood Pb level
6 was associated with a smaller but statistically significant decrement in FSIQ after
7 adjusting for potential confounding factors (Palaniappan et al.. 2011; Kim et al.. 2009b:
8 Lanphear et al., 2005; Canfield et al., 2003a). The consideration for potential
9 confounding varied among studies. Most studies adjusted for SES-related variables such
10 as the Hollingshead Index, household income, and/or parental education. Several, in
11 particular the prospective studies, adjusted for parental cognitive function and parental
12 caregiving quality commonly evaluated as HOME score. Overall, recent studies
13 considered potential confounding by SES and parental IQ or education but not parental
14 caregiving quality. Analyses of associations between potential confounding factors and
15 blood Pb level and cognitive function indicated that the confounding factors may vary
16 across populations and endpoints. In the Cleveland cohort, adjustment for HOME score
17 attenuated the blood or tooth Pb level-cognitive function relationships (Greene and
18 Ernhart. 1993: Greene et al.. 1992: Ernhart et al.. 1988). In the Rochester cohort, HOME
19 score met the criteria for adjustment in models for FSIQ (Canfield et al., 2003a) but not
20 all measures of memory and executive function (Froehlich et al.. 2007: Canfield et al..
21 2004: 2003b). Adjustment for SES is difficult as it is highly correlated with Pb exposure
22 and there is no single measure that represents SES. Residual confounding also is likely by
23 factors not considered. The combination of evidence from prospective studies that
24 considered several well-characterized potential confounding factors plus evidence that Pb
25 exposure induces impairments in cognitive function in animals, in particular, spatial
26 memory and executive function, which are associated with blood or tooth Pb levels in
27 children increases confidence that the associations between blood and tooth Pb levels and
28 cognitive function observed in children represent a relationship with Pb exposure.
29 With regard to important lifestages and durations of Pb exposure, toxicological evidence
30 clearly demonstrates impaired learning and memory in animals exposed to Pb
31 gestationally with or without early postnatal exposure. Impairments in learning and
32 memory observed with lower blood Pb levels (8-17 (ig/dL) were found with Pb exposures
33 that began during the gestational or lactation period. The effect of early life Pb exposures
34 is supported by evidence that processes such as neurogenesis and synaptic pruning are
35 highly active during the first few years of life (Rice and Barone. 2000: Landrigan et al..
36 1999). However, evidence in a group of monkeys also indicates impaired learning with
37 Pb exposure beginning later during the juvenile period, indicating that Pb exposure in
38 infancy is not necessary to induce impairments in cognitive function (Rice. 1992b. 1990:
November 2012 5-119 Draft - Do Not Cite or Quote
-------
1 Rice and Gilbert. 1990b). Epidemiologic studies also point to cognitive function
2 decrements associated with blood Pb levels measured at various lifestages and time
3 periods. Among studies of young children <3 years, several found stronger associations
4 of MDI with prenatal (maternal or cord) blood Pb than with postnatal child blood Pb (Hu
5 et al., 2006; Bellinger et al.. 1987; Dietrich et al.. 1987a; Vimpani et al.. 1985). However,
6 in older children, ages 4-17 years, in whom cognitive function is more stable and reliably
7 measured, decrements in cognitive function were associated with more strongly with
8 indicators of postnatal Pb exposure, i.e., concurrent, early childhood, and cumulative
9 average blood Pb levels as well with tooth Pb levels. Evidence did not clearly identify an
10 individual critical postnatal time period or duration of Pb exposure in terms of risk of
11 developing cognitive function decrements. Because of the contribution of bone Pb levels
12 to concurrent blood Pb levels in children, associations with concurrent blood Pb levels
13 may reflect an effect of past and recent Pb exposures.
14 Previous prospective studies found blood Pb-associated decrements in cognitive function
15 in populations with mean blood Pb levels 5-10 (ig/dL (Table 5-3). In analyses restricted
16 to children in the lower range of the blood Pb distribution (e.g., <10 (ig/dL), associations
17 with FSIQ were found in groups of children with mean age 2 year or concurrent blood Pb
18 levels 3-4 (ig/dL with consideration of peak blood Pb levels (Bellinger. 2008; Canfield.
19 2008; Hornung. 2008). Several recent studies found associations of FSIQ with lower
20 blood Pb levels (primarily concurrent), population means 2-5 (ig/dL, (Kim et al.. 2009b;
21 Jusko et al.. 2008; Zailinaet al., 2008; Chiodo et al., 2007) for FSIQ but not consistently
22 for other indices of cognitive function (Cho et al.. 2010; Miranda et al.. 2010;
23 Chandramouli et al., 2009; Surkan et al., 2007). Many of these recent studies had
24 uncertainties related to the influence of higher past Pb exposures, high prevalence of
25 prenatal drug exposure, or potential confounding. Several recent toxicological studies
26 added to the evidence for impaired learning and memory in animals with lower blood Pb
27 levels, 8-17 (ig/dL (Cory-Slechta et al.. 2010; Niu et al.. 2009; Virgolini et al.. 2008a;
28 Stangle et al.. 2007). Recent evidence from the Cory-Slechta laboratory found learning
29 impairments with lower lifetime Pb exposures when combined with stress, which
30 potentially may be mediated via effects on corticosterone and dopamine (Rossi-George et
31 al.. 2011; Cory-Slechta et al.. 2010; Rossi-George et al.. 2009; Virgolini et al.. 2008a).
32 The biological plausibility for epidemiologic and toxicological evidence linking Pb
33 exposure to decrements in cognitive function is provided by the well-characterized
34 toxicological evidence for Pb exposure interfering with development of the brain and
35 activity of neurochemical processes that mediate cognitive function (Section 5.3.11). Pb
36 has been shown to increase the permeability of the blood-brain barrier and deposit in the
37 CNS. Pb has been shown to impair neurogenesis, synaptic architecture, and neurite
38 outgrowth. Cognitive function is mediated by the cortical and subcortical structures of the
November 2012 5-120 Draft - Do Not Cite or Quote
-------
1 brain that integrate function in the hippocampus, prefrontal cortex, and nucleus
2 accumbens using dopamine and glutamate as primary neurotransmitters. Experimental
3 studies have shown that Pb induces changes in dopamine and glutamate release in these
4 regions and decreases long-term potentiation, which is a major cellular mechanism
5 underlying synaptic plasticity and learning and memory.
5.3.2.7 Epidemiologic Studies of Cognitive Function in Adults
Adults without Occupational Pb Exposures
6 As described in the preceding section, Pb exposure that begins in gestation and lasts
7 through the early postnatal period or for a lifetime or begins after infancy has been shown
8 to induce learning impairments in adult animals. Less well characterized are learning
9 impairments in adult animals due to adult-only Pb exposures. As reported in the
10 2006 Pb AQCD, epidemiologic studies have examined cognitive performance in adults
11 without occupational Pb exposure primarily in association with concurrently measured
12 blood and bone Pb levels and have found associations with bone Pb levels but not blood
13 Pb levels (U.S. EPA. 2006b). Recent studies produced similar findings and provided new
14 evidence from prospective analyses (Table 5-10).
15 Evidence was provided by large cohorts examining multiple exposures and outcomes,
16 reducing the likelihood of selective participation of subjects with higher Pb exposures
17 and cognitive deficits. Most studies performed multiple tests of cognitive function.
18 However, associations with bone Pb level were not isolated to a few tests. Several
19 publications are available; however, many are variant analyses in the same population
20 (e-g-, Normative Aging Study [NAS], NHANES) and are not considered as all
21 independent assessments of the Pb-cognitive function relationship. Further, although
22 evidence is available from longitudinal cohorts, most analyses are cross-sectional
23 examining the association between one measurement of cognitive function and a
24 concurrent measure of blood or bone Pb level. Because temporality cannot be
25 determined, causal inference regarding the effects of Pb exposure is limited. In analyses
26 of bone Pb level, this limitation is mitigated somewhat because bone Pb level reflects
27 several years of exposure. Additionally, with blood and bone Pb level, it is difficult to
28 characterize the specific timing, duration, frequency, and level of Pb exposure that
29 contributed to associations observed with cognitive function. This uncertainty may apply
30 particularly to assessments of blood Pb levels, which in nonoccupationally-exposed
31 adults, reflect both current exposures and cumulative Pb stores in bone that are mobilized
32 during bone remodeling (Sections 4.3 and 4.7.3). Although studies adjusted for age, a
33 common limitation is the potential for residual confounding by age because of the strong
November 2012 5-121 Draft - Do Not Cite or Quote
-------
1 correlation between increasing bone Pb levels and increasing age. However, the
2 coherence with evidence for cognitive function decrements associated with long-term Pb
3 exposure in animals provides support for associations observed in adults. Because of the
4 difficulty in establishing the temporal sequence between Pb exposure and cognitive
5 function in cross-sectional studies, in the review of evidence, emphasis was placed on
6 prospective analyses. Emphasis also was placed on studies that considered several
7 potential confounding factors such as age, education, SES, smoking, and alcohol use.
Evidence from Prospective Studies
8 Key evidence for the effects of Pb exposure on cognitive function of adults has been
9 provided by recent prospective analyses of the large Baltimore Memory Study (BMS)
10 and NAS. Strengths of these studies include comparisons of associations between bone
11 and blood Pb levels, the repeated assessment of Pb biomarker levels and cognitive
12 performance, the high follow-up participation of subjects, and lack of selective attrition
13 by Pb biomarker levels and demographic characteristics. In particular, the repeated
14 assessments permitted the examination of associations of bone Pb levels with changes in
15 cognitive function over time, which better established the temporal sequence between Pb
16 exposure and subsequent changes in cognitive function. The BMS and NAS differed in
17 many respects, including sex and race of subjects, the test instruments used, and potential
18 confounding factors considered. The BMS included men and women, 50-70 years of age,
19 residing in Baltimore, MD. A total of 1,140 out of 2,351 (48.5%) subjects participated
20 from neighborhoods that represented a diversity of race and SES. This study was unique
21 in that it included a large proportion of African-Americans (n=395). In comparison, the
22 NAS involved only men (original n = 2,280) residing in the Greater Boston area. Subjects
23 primarily were white and at enrollment were ages 21 to 80 years and had no current or
24 past chronic medical conditions. Both studies adjusted for age and education. The BMS
25 additionally adjusted for household wealth, and the NAS additionally adjusted for
26 smoking and alcohol intake. Results from both of these cohorts with different
27 demographics and methodology indicated Pb-associated cognitive function decrements.
28 In the BMS, longitudinal analyses involved repeat cognitive testing at 14-month
29 intervals. Most subjects completed follow-up; 91% at the second round of testing and
30 83% at the third round (Bandeen-Roche et al., 2009). An interquartile range higher
31 baseline tibia Pb level (12.7 ug/g) was associated with a 0.019 unit (95% CI: -0.031,
32 -0.007) per year decrease in eye-hand coordination z-score, with adjustment for age, sex,
33 race, SES, and interviewer, with a larger decrease estimated for African Americans than
34 for whites (Table 5-10). Results were not homogeneous across the various tests
35 performed. Tibia Pb levels were more weakly associated with time-related decreases in
36 language, processing speed, and executive function; however, most effect estimates were
November 2012 5-122 Draft - Do Not Cite or Quote
-------
negative in direction. Further, for language and executive function, tibia Pb level was
associated with greater decreases in scores among whites than African Americans.
Table 5-10 Associations of blood and bone Pb levels with cognitive function in
adults.
Study Population and
Methodological Details
(Presented first for
prospective analyses then
for cross sectional analyses.
Within categories, studies
are presented in order of
strength of methodology)
Subgroup
(where
Cognitive Test examined)
Blood Pb Effect
Estimate
(95% Cl)
Bone Pb Effect Estimate
(95% Cl)
Prospective Studies:
Bandeen-Roche et al. (2009)
943, ages 50-70 yr at baseline,
BMS, Baltimore, MD
Large sample of men and
women of various
races/ethnicities with repeated
measures of cognitive function
and tibia Pb. High follow-up
participation over 28 mo.
Marginal linear regression
models adjusted for age, sex,
household wealth, education,
race/ethnicity, interviewer. Did
not consider potential
confounding by history of
smoking or alcohol use.
Tibia Pb Mean (SD):
19 (12.7) ug/g
NOT EXAMINED
Longitudinal associations
Eye/hand African-
coordination - Americans
Purdue pegboard
Change in Z-scores per
IQR increase:
-0.032 (-0.052, -0.012)/yr
-0.009 (-0.024, 0.006)/yr
Cross-sectional associations
Verbal
memory/learning
- Rey auditory
verbal learning
test
Language -
Boston naming
test
African-
Americans
White
African-
Americans
White
0.006 (-0.09, 0.10)
-0.076 (-0.15, 0.001)
0.065 (-0.010,0.14)
-0.024 (-0.12, 0.07)
Weisskopfetal. (2007b)
405-749 males, mean age 68.7
yr at baseline, NAS, Boston,
MA area.
Large sample, only men,
primarily white. Repeated
measures of cognitive function
and tibia Pb. High follow-up
participation over 3.5 yr.
Linear repeated measures
analysis adjusted for age, age2,
education, smoking status,
current alcohol intake, yr
between bone Pb
measurement and first
cognitive test, yr between
cognitive tests. Also evaluated
language, computer
experience, physical activity.
Mean (IQR)
Tibia Pb:20(15) ug/g
Patella Pb: 25 (20) ug/g
NOT EXAMINED
Longitudinal associations
Visuospatial skills - pattern
comparison (+ = poorer
performance), NES2
Executive function - verbal fluency,
WISC-R
Short-term memory - word list,
CERAD
Change in score over 3.5 yr
per IQR increase:
Tibia: 0.079 (0.04, 0.12)
Patella: 0.073 (0.04, 0.12)
Tibia:-0.04 (-0.16, 0.08)
Patella: -0.086 (-0.20, 0.03)
Tibia:-0.028 (-0.12, 0.06)
Patella:-0.081 (-0.17, 0.005)
Cross-sectional associations
Visuospatial skills - pattern
comparison latency
(+ = poorer performance), NES2
Executive function - verbal fluency,
WISC-R
Short-term memory - word list,
CERAD
Tibia:-0.03 (-0.17, 0.11)
Patella:-0.02 (-0.14, 0.11)
Tibia:-0.27 (-0.70, 0.16)
Patella:-0.22 (-0.62, 0.17)
Tibia: 0.12 (-0.20, 0.32)
Patella: 0.012 (-0.18, 0.41)
November 2012
5-123
Draft - Do Not Cite or Quote
-------
Study3 Population and
Methodological Details
(Presented first for
prospective analyses then
for cross sectional analyses.
Within categories, studies
are presented in order of
strength of methodology)
Cognitive Test
Subgroup
(where
examined)
Blood Pb Effect
Estimate
(95% Cl)
Bone Pb Effect Estimate
(95% Cl)
Wang et al. (2007a)
358 males, median age: 67 yr,
MAS, Boston, MA area
Same cohort as above. Subset
representative of full cohort.
Linear regression adjusted for
age, years of education,
smoking status, pack-years
smoking, nondrinker,
grams/day alcohol
consumption, English as first
language, computer
experience, diabetes.
Tibia Pb: Median (IQR) 19
(15)ug/g
NOT EXAMINED
Mini Mental State
Exam Score
HFE wildtype
One HFE
variant
Two HFE
variants
Change in Score per
IQR increase:
-0.02 (-0.10, 0.07)/yr
-0.14 (-0.33, 0.04)/yr
-0.63 (-1.04,-0.21 )/yr
Cross-sectional Studies:
Shih et al. (2006)
985 adults, mean age: 59 yr,
BMS, Baltimore, MD
Large sample. Subjects with
tibia Pb measured were more
educated and white. Compared
blood/bone associations.
Linear regression adjusted for:
Model A: age, sex, technician,
presence of APOE-E4 allele
Model B: Model I, years of
education, race/ethnicity, wealth
Did not consider potential
confounding by history of
smoking or alcohol use.
Mean (SD)
Concurrent blood Pb: 3.5
(2.2) ug/dL
TibiaPb:18.7(11.2)ug/g
Language -
Boston naming
test
Eye-hand
coordination -
Purdue Pegboard,
trail making
Executive
functioning -
Purdue Pegboard,
Stroop and trail
making test
Visuoconstruction
- Rey complex
figure copy
Model A
Model B
Model A
Model B
Model A
Model B
Model A
Model B
Score per 1 |jg/dL
increase:
-0.006 (-0.03, 0.017)
-0.002 (-0.02, 0.016)
-0.011 (-0.03, 0.01)
-0.008 (-0.02, 0.002)
-0.01 4 (-0.03, 0.005)
-0.010 (-0.03, 0.007)
-0.01 9 (-0.05, 0.008)
-0.01 4 (-0.04, 0.01)
Score per 1 |jg/g increase:
-0.008 (-0.01 , -0.004)
0.0006 (-0.003, 0.004)
-0.008 (-0.01 , -0.004)
-0.008 (-0.02, 0.002)
-0.008 (-0.01 , -0.004)
-0.003 (-0.006, 0.0008)
-0.01 2 (-0.02, -0.007)
-0.004 (-0.01 , 0.0003)
Glass etal. (2009)
1,001 adults, mean age 59 yr,
BMS, Baltimore, MD
Large sample. High participation
rate: 91 %.
Multilevel hierarchical regression
model adjusted for age, sex,
race/ethnicity, education, testing
technician, time of day.
Investigator assessed NPH.
Did not consider potential
confounding by history of
smoking or alcohol use.
Tibia PbMean (SD): 18.8
NOT EXAMINED
Language -
Boston naming
test
Eye-hand
coordination -
Purdue Pegboard,
trail making test
Executive
functioning -
Purdue Pegboard,
Stroop, trail
making test
Visuoconstruction
- Rey complex
figure copy
2nd tertile NPH
SrdtertileNPH
2nd tertile NPH
3rd tertile NPH
2nd tertile NPH
3rd tertile NPH
2nd tertile NPH
3rd tertile NPH
Score per 1 |jg/g increase:
0.001 (-0.008, 0.009)b
-0.009 (-0.017,-0.0001 )b
-0.004 (-0.012, 0.005)b
-0.006 (-0.015, 0.002)b
-0.002 (-0.010,0.006)"
-0.010 (-0.018, -0.002)b
-0.003 (-0.014, 0.008)b
-0.006 (-0.017, 0.005)b
November 2012
5-124
Draft - Do Not Cite or Quote
-------
Study3 Population and
Methodological Details
(Presented first for
prospective analyses then
for cross sectional analyses.
Within categories, studies
are presented in order of
strength of methodology)
Subgroup
(where
Cognitive Test examined)
Blood Pb Effect
Estimate
(95% Cl)
Bone Pb Effect Estimate
(95% Cl)
Weuve et al. (2006)
720 males, ages > 45 yr, MAS,
Boston, MA area
Large sample. High follow-up
participation. Compared
blood/bone associations.
Linear mixed effects regression
adjusted for smoking status,
grams/day alcohol consumption,
calorie adjusted calcium intake,
regular energy expenditure on
leisure time physical activity,
diabetes.
Additional adjustment for dietary
factors.
Median (IQR)
Concurrent blood Pb: 5.2
(3) ug/dL
Tibia: 19(15) ug/g
Patella: 27 (21) ug/g
Mini Mental State
Exam Score
ALAD wildtype
ALAD-2 carrier
ALAD wildtype
ALAD-2 carrier
Score per IQR
increase:
-0.05 (-0.16, 0.06)
-0.29 (-0.56, -0.02)
Score per IQR increase:
Tibia:-0.05 (-0.21, 0.12)
-0.16 (-0.58, 0.27)
Patella: -0.07 (-0.23, 0.09)
-0.26 (-0.64, 0.12)
Rajanetal. (2008)
486-959 males, ages > 45 yr,
MAS, Boston, MA area
Large sample. Compared
blood/bone associations. Linear
regression adjusted for blood Pb
main effect, ALAD genotype,
age at cognitive test, education,
grams/day alcohol consumption,
pack-years smoking, English as
first language. Also considered
smoking status, income,
physical activity, diabetes,
coronary heart disease.
Concurrent blood Pb Mean
(SD): 5.3(2.9) ug/dL(ALAD
wildtype), 4.8 (2.7) ug/dL
(ALAD2 carriers)
Tibia Mean (SD): 21.9
(13.8) ug/g (ALAD wildtype),
21.2(11.6) ug/g (ALAD2
carriers)
Patella Mean (SD): 29.3
(19.1) ug/g (ALAD wildtype),
27.9 (17.3) ug/g (ALAD2
carriers)
Visuospatial - constructional Praxis,
CERAD
Executive function - verbal fluency,
CERAD
Verbal memory - word list memory,
CERAD
Perceptual speed - mean latency
continuous performance, NES
Score*ALAD2 per
IQR increase:
-0.05 (-0.23, 0.13)°
-0.03 (-0.22, 0.16)°
0.003 (-0.18, 0.19)°
-0.18 (-0.42, 0.06)°
Score*ALAD2 per IQR
increase:
Tibia: -0.25 (-0.49, -0.02)°
Patella: 0.02 (-0.19, 0.23)°
Tibia:-0.11 (-0.34,0.13)°
Patella:-0.03 (-0.24, 0.19)°
Tibia: 0.08 (-0.15, 0.31)°
Patella: 0.14 (-0.07, 0.34)°
Tibia: -0.25 (-0.59, 0.08)°
Patella:-0.16 (-0.44, 0.12)°
November 2012
5-125
Draft - Do Not Cite or Quote
-------
Study3 Population and
Methodological Details
(Presented first for
prospective analyses then
for cross sectional analyses.
Within categories, studies
are presented in order of
strength of methodology)
Subgroup
(where
Cognitive Test examined)
Blood Pb Effect
Estimate
(95% Cl)
Bone Pb Effect Estimate
(95% Cl)
Weuve et al. (2009)
587 females, ages 47-74 yr,
Nurses' Health Study, Boston,
MA area
Large sample of only females.
Compared blood/bone
associations. Generalized
estimating equations adjusted
forage and age2 at Pb
assessment, age at cognitive
assessment, education,
husband's education, alcoholic
drinks/week, smoking status,
physical activity, use of aspirin,
ibuprofen, Vitamin E
supplements, menopausal
status, and postmenopausal
hormone use. Additional
adjustment for nutrition factors,
medication use, mental health.
Assessed outcomes over
telephone but a mean 5 years
after Pb measured.
Mean (SD)
Concurrent blood Pb 2.9
(1.9)ug/dL
Tibia Pb 10.5(9.7) ug/g
Patella Pb 12.6(11.6) ug/g
Orientation, registration, immediate
verbal memory with TICS. Immediate
and delayed paragraph recall,
category fluency, digit span
backwards (working memory,
attention) with EBMT
Composite Z-score
Composite except letter fluency
Z-score per SD
increase:
Z-score per SD increase:
-0.015
(-0.069, 0.039)
0.016
(-0.071, 0.039)
Tibia: -0.040 (-0.09, 0.004)
Patella:-0.012 (-0.06, 0.03)
Tibia:-0.05 (-0.10,-0.003)
Patella:-0.033 (-0.08, 0.014)
Krieg and Butler (2009)
2,823 adults, ages 20-59 yr,
Large U.S. representative
NHANES III (1991-1994).
Log-linear regression model
adjusted forage, sex, education,
family income, race-ethnicity,
computer or video-game
familiarity, alcohol use within the
last 3 h, test language, sampling
unit and stratum. Did not
consider potential confounding
by smoking.
Concurrent Blood mean (SD):
2.88(6.91) ug/dL
Symbol Digit
Substitution
(mean total
latency, sec)
Serial digit
learning total
score, NES
Ages 20-39 yr
Ages 40-59 yr
Ages 20-39 yr
Ages 40-59 yr
Z-score per 1
increase:
-0.097 (-0.42, 0.23)d
-0.290 (-0.60, 0.02)d
-0.1 17 (-0.46, 0.23)d
0.401 (-0.19, 1.0)d
NOT EXAMINED
November 2012
5-126
Draft - Do Not Cite or Quote
-------
Study3 Population and
Methodological Details
(Presented first for
prospective analyses then
for cross sectional analyses.
Within categories, studies
are presented in order of
strength of methodology)
Krieg et al. (2009)
2,090 adults, ages 20-59 yr,
1 976 adults, ages > 60 yr. Large
U.S. representative NHANES III
(1991-1994).
Log linear regression model
adjusted for sex, age, education,
family income, race-ethnicity,
computer or video game
familiarity, alcohol use in the last
3 h, test language (20-59 yr) and
sex, age, education, family
income, race-ethnicity, test
language (> 60 yr), sampling
unit and stratum. Did not
consider potential confounding
by smoking.
Concurrent Blood Pb Mean (SD)
Age 20-59 yr: 2.85 (7.31) ug/dL
Age > 60 yr: 4.02 (3.56) ug/dL
Krieg et al. (201 0)
2,093 adults, ages 20-59 yr,
1 ,799 adults, ages > 60 yr.
Large U.S. representative
NHANES III (1991-1994).
Log linear regression model
adjusted for sex, age, education,
family income, race-ethnicity,
computer or video game
familiarity, alcohol use in the last
3h, test language, sampling unit,
stratum (20-59 yr) and sex, age,
education, family income, race-
ethnicity, test language,
sampling unit, and stratum (s 60
yr). Did not consider potential
confounding by smoking.
Concurrent blood Pb Mean (SD)
Age 20-59 yr: 2.85 (7.32) ug/dL
Age > 60 yr: 4.02 (3.39) ug/dL
Cognitive Test
Symbol Digit
Substitution
(mean total
latency)
Serial digit
learning total
score
Word recall,
number correct
Story recall,
number correct,
Neurobehavioral
Evaluation
System
Symbol Digit
Substitution
(mean total
latency, sec)
Serial digit
learning total
score
Word recall,
number correct
Story recall,
number correct
Neurobehavioral
Pvsli istinn
QVCtlUCt UUI 1
System
Subgroup
(where
examined)
Ages 20-59 yr
ALAD wildtype
ALAD-2 carrier
Ages 20-59 yr
ALAD wildtype
ALAD-2 carrier
Ages > 60 yr
ALAD wildtype
ALAD-2 carrier
Ages > 60 yr
ALAD wildtype
ALAD-2 carrier
Age group and
VDR haplotype
Ages 20-59 yr
CC haplotype
CT haplotype
TC haplotype
TT haplotype
Ages 20-59 yr
CC haplotype
CT haplotype
TC haplotype
TT haplotype
Ages > 60 yr
CC haplotype
CT haplotype
TC haplotype
TT haplotype
Ages > 60 yr
CC haplotype
CT haplotype
TC haplotype
TT haplotype
Blood Pb Effect
Estimate Bone Pb Effect Estimate
(95% Cl) (95% Cl)
Z-score per 1 [tgldL
increase: NOT EXAMINED
-0.1 32 (-0.358, 0.095)d
-0.526 (-1.11 8, 0.066)d
-0.022 (-0.526, 0.482)d
0.025 (-0.406, 0.456)d
-0.075 (-0.285, 0.135)
0.025 (-0.406, 0.456)
0.085 (-0.0997, 0.271)
-0.466 (-1 .072, 0.139)
Score per log
increase: NOT EXAMINED
-20 (-44, 4.0)d
0.73 (-1 .4, 2.9)d
-2.6 (-5.3, 0.07)d
-3.6 (-7.2, 0.05)d
8.0(0.61, 15.4)
1 .0 (-0.89, 2.9)
-1.4 (-3.1, 0.29)
-0.01 4 (-2.8, 2.5)
-0.65 (-1.5, 0.25)
-0.08 (-0.34, 0.19)
-0.03 (-0.40, 0.33)
-0.08 (-0.76, 0.60)
0.29 (-3.3, 3.9)
0.01 (-0.38, 0.40)
0.07 (-0.64, 0.78)
-0.22 (-0.86, 0.43)
November 2012
5-127
Draft - Do Not Cite or Quote
-------
Study3 Population and
Methodological Details
(Presented first for
prospective analyses then
for cross sectional analyses.
Within categories, studies
are presented in order of
strength of methodology)
Cognitive Test
Subgroup
(where
examined)
Blood Pb Effect
Estimate
(95% Cl)
Bone Pb Effect Estimate
(95% Cl)
Van Wijngaarden et al. (2009)
47 adults, mean age 61.5 yr,
Rochester, NY
Small sample size, without
consideration for potential
confounding by smoking or
alcohol use. Linear regression
adjusted forage, sex,
educational level, history of
hypertension. Excluded subjects
with BMI >32 kg/m2.
Mean (SD)
Tibia: 2.0 (5.2) ug/g
Calcaneus: 6.1 (8.5) ug/g
Delayed matching
to sample, %
correct
CANTAB
NOT EXAMINED
Paired Associate
Learning, total
trials adjusted
(increase = poorer
performance)
CANTAB
Calcaneus
Lowest fertile: 87.56e
Medium tertile: 86.67
Highest Tertile: 80.67, p=0.03
Tibia
Lowest tertile: 85.42e
Medium tertile: 87.08
Highest tertile: 82.44, p=0.25
Calcaneus
Lowest tertile: 2.54e
Medium tertile: 2.61
Highest tertile: 2.72, p = 0.21
Tibia
Lowest tertile: 2.62e
Medium tertile: 2.59
Highest tertile: 2.66, p = 0.79
Gao et al. i
188 adults, mean age 69.2 yr,
Rural Sichuan and Shandong
Provinces, China.
Small sample size. Subset was
younger, had more education,
and higher BMI than full cohort.
Separate ANCOVA adjusted for
age, sex, education, BMI, or
APOE £4. History of smoking
and alcohol consumption not
associated with cognitive score.
Concurrent plasma Pb Mean
(SD): 0.39 (0.63) ug/dL
Composite
cognitive Z-score
Word list learning,
word recall
(CERAD), CSID,
IU story recall,
Animal naming
fluency test, IU
token test of
language and
working memory
Z-score per 1 |jg/dL
plasma Pb
increase:
42.8(21.4, 64.2)
NOT EXAMINED
Note: Effect estimates in bold indicate the stronger association between blood Pb and bone Pb level. IQR = Interquartile range,
BMS = Baltimore Memory Study, NAS = Normative Aging Study, NES2 = Neurobehavioral Evaluation System 2, WISC-R =
Wechsler Adult Intelligence Scale-Revised, CERAD = Consortium to Establish Registry for Alzheimer's disease, HFE = Human
Hemochromatosis protein, NPH = Neighborhood Psychosocial Hazard, TICS = Telephone Interview for Cognitive Status,
EBMT = East Boston Memory Test, CANTAB = Cambridge Neuropsychological Test Automated Battery, CSID = Community
Screening Instrument for Dementia, IU = Indiana University.
"Studies are presented first for prospective analyses then for cross sectional analyses. Within categories, studies are presented in
order of strength of methodology.
bEffect estimates indicate interactions between Pb and category of NPH, with the lowest tertile of NPH serving as the reference
group.
°Effect estimates indicate interactions between Pb and ALAD genotype.
dThe directions of effect estimates were changed to indicate a negative slope as a decrease in cognitive performance.
eResults refer to mean cognitive function scores among tertiles of bone Pb. Tertile concentrations not reported.
November 2012
5-128
Draft - Do Not Cite or Quote
-------
1 Similar to the BMS, among NAS men, higher baseline tibia Pb levels were associated
2 with decreases in cognitive performance overtime in longitudinal analyses with repeated
3 measures of cognitive function plus a bone Pb-time interaction term in order to estimate
4 the association between baseline bone Pb level and decline in cognitive test score over
5 time (Weisskopf et al. 2007b). This NAS analysis expanded the evidence base by also
6 finding associations with patella Pb levels. Two measurements of cognitive function,
7 collected approximately 3.5 years apart were available for 60-70% of participants. Both
8 tibia and patella Pb levels were associated with decrements in executive function, short-
9 term memory, and visuospatial skills (as indicated by increased response latency on a
10 pattern comparison test). The strongest effect was estimated for the latter. Weisskopf et
11 al. (2007b) also found a nonlinear association with patella Pb, with latency times
12 becoming worse over time (i.e., larger values indicating slower response time) up to
13 approximately 60 ug/g patella Pb then leveling off at higher levels (Figure 5-8). A
14 20 ug/g difference in patella Pb level was associated with an increase in latency of 0.073
15 ms (95% CI: 0.04, 0.12) among all men and a 0.15 ms increase among men with patella
16 Pb level <60 ug/g. Both patella and tibia Pb were associated with fewer errors on the
17 pattern comparison test. The authors proposed that this may be related to slowing reaction
18 time to improve accuracy. When the nine men with the highest bone Pb levels were
19 removed, the association with fewer errors was no longer statistically significant.
20 However, the authors did not indicate whether the point estimate changed.
November 2012 5-129 Draft - Do Not Cite or Quote
-------
.™ CSj
E o
|
I P
o o
cc °
1
c 2
I
P CO
8 ? J
20 40 60
Patella lead (M9/g)
80
Note: Models are adjusted for age, age squared, education, smoking, alcohol intake, years between bone Pb measurement and first
cognitive test, and years between the cognitive tests. The 9 subjects with the highest patella Pb levels (>89 ug/g bone mineral) were
removed. The estimated concentration-response is indicated by the solid line and the 95% confidence interval by the dashed lines.
The patella Pb level-associated increase in response latency is larger among men with patella Pb levels <60 ug/g. Patella Pb levels
of all individual subjects are indicated by short vertical lines on the abscissa, (reference = 0 at mean of patella Pb level).
Source: Reprinted with permission of Williams & Wilkins, Weisskopf et al. (2007b).
Figure 5-8 Nonlinear association between patella Pb level and the relative
change over 3.5 years in response latency on the pattern
comparison test in men from the Normative Aging Study.
1 Longitudinal analysis of the NAS cohort also indicated that hemochromatosis (HFE)
2 gene variants modified the blood Pb-cognition association (Wang et al.. 2007a). In
3 models adjusted for age, years of education, smoking status, pack-years smoking,
4 nondrinker, grams/day alcohol consumption, English as first language, computer
5 experience, and diabetes, an interquartile range higher tibia Pb level (15 ug/g) was
6 associated with a 0.22 point steeper annual decline (95% CI: -0.39, -0.05) in Mini-Mental
7 State Examination score (MMSE, which assesses cognitive impairment in a number of
8 domains) among the 130 (36%) men with either the H63D or C282Y variant. The
9 association was found to be nonlinear, with larger Pb-associated declines observed at
10 higher tibia Pb levels (Figure 5-9. solid line). The change in MMSE score associated with
11 15 ug/g higher tibia Pb levels was comparable to that found between NAS men who were
12 4 years apart in age. Tibia Pb level was not associated with a decline in MMSE score in
13 men with the HFE wildtype genotype (Figure 5-9. dashed line). Bone Pb levels did not
November 2012
5-130
Draft - Do Not Cite or Quote
-------
1
2
3
4
differ widely by HFE variant. HFE variants, H63D and C282Y, are associated with
hemochromatosis, a disease characterized by higher iron body burden. Iron metabolism
has been hypothesized to affect neurodegenerative diseases, which may explain the
observed effect modification. However, firm conclusions are not warranted.
o
o
uj _n?
GO U'^
•- -0.4
o
O)
c
CD
"§ -0.6
< -0.8
WFfwildtype
HFE variant allele
0 10 20 30 40 50
Tibia lead biomarker (ug/g)
Note: The lines indicate curvilinear trends estimated from the penalized spline method. Among hemochromatosis (HFE) wild-types,
the association between tibia Pb and annual cognitive decline was nearly null (dashed line). Among variant allele carriers, the
association tended to deviate from linearity (solid line, p = 0.08), with a greater tibia Pb-associated decline in MMSE observed
among men with higher tibia Pb levels. The model was adjusted for age, years of education, smoking status, pack-years smoking,
nondrinker, grams/day alcohol consumption, English as first language, computer experience, and diabetes.
Source: Wang et al. (2007a).
Figure 5-9 Nonlinear association of tibia Pb level with annual rate of
cognitive decline, by hemochromatosis genotype in men from the
Normative Aging Study.
5
6
7
Evidence from Cross-sectional Studies
Associations between bone Pb levels and decrements in cognitive function in adults also
are supported by evidence from several cross-sectional studies conducted in the BMS and
NAS cohorts and other populations. The cross-sectional studies have contributed
evidence for stronger associations of cognitive function decrements with bone Pb levels
than blood Pb level and for associations with adjustment for additional potential
November 2012
5-131
Draft - Do Not Cite or Quote
-------
1 confounding factors such as diet and medication use. While cross-sectional studies
2 examined factors that may potentially increase risk of Pb-associated cognitive function
3 decrements in adults, they each examined different factors and did not produce
4 conclusive evidence. These subgroup analyses also are subject to higher probability of
5 finding an association by chance.
6 In addition to comparisons of blood and bone Pb levels, cross-sectional analyses of the
7 BMS included detailed analysis of potential confounding, although smoking and alcohol
8 use were not examined. Among 991 adults, both higher concurrent blood and bone Pb
9 level were associated with poorer performance in tests of language, processing speed,
10 eye-hand coordination, executive function, verbal memory and learning, visual memory,
11 and visuoconstruction; however, associations with tibia Pb level tended to be larger in
12 magnitude (Table 5-10) (Shih et al. 2006). Mean (SD) blood and tibia Pb levels were
13 3.46 (2.23) ug/dL and 18.7 (11.2) ug/g, respectively. Tibia Pb levels were associated with
14 worse performance on tests in all domains with adjustment for age, sex, testing
15 technician, and presence of the apolipoprotein (APO)E-e4 allele (potential risk factor for
16 Alzheimer's Disease). The magnitudes of associations were attenuated with additional
17 adjustment for education, race, and household wealth; however, in these more fully-
18 adjusted models, higher tibia Pb levels remained associated with poorer performance in
19 all domains except language and processing speed. The strongest association was found
20 for visuoconstruction, which assesses visuospatial skills and motor skills. A 1 ug/g bone
21 higher tibia Pb level was associated with a 0.0044 SD (95% CI: -0.0091, 0.0003) lower
22 visuoconstruction score. Analysis of tibia Pb as a quadratic term did not indicate a
23 nonlinear relationship with visuoconstruction.
24 In contrast with longitudinal results in BMS, race-stratified analyses of persistent effects
25 in cross-sectional analyses indicated that tibia Pb levels were associated with greater
26 decreases in performance on tests of eye-hand coordination, executive function, and
27 verbal memory and learning among whites than among African Americans (Bandeen-
28 Roche et al., 2009). Among all subjects, tibia Pb-associated decrements in cognitive
29 performance were modified by neighborhood level psychosocial stress. Specifically,
30 higher tibia Pb levels were associated with larger decrements, particularly in language,
31 eye-hand coordination, and executive function, among subjects living in neighborhoods
32 with a greater number psychosocial hazards (e.g., number of violent crimes, emergency
33 calls, off-site liquor licenses as assessed by investigators) (Glass et al.. 2009) (Table
34 5-10). Results were adjusted for age, sex, race/ethnicity, education, testing technician,
35 and time of testing. Subjects living near more psychosocial hazards had slightly higher
36 tibia Pb levels. In support of these results, several studies have found Pb-stress
37 interactions in impaired learning and memory of adult animals with Pb exposures
November 2012 5-132 Draft - Do Not Cite or Quote
-------
1 beginning in gestation and lasting through post-weaning or to the time of testing
2 (Section 5.3.2.3).
3 The 2006 Pb AQCD (U.S. EPA. 2006b) described cross-sectional associations of both
4 blood and tibia Pb levels with poorer cognitive performance among 141 NAS men
5 (Pavton et al.. 1998). Several recent, larger cross-sectional NAS analyses corroborated
6 previous findings for bone Pb but generally indicated weak associations with concurrent
7 blood Pb levels and only in groups with specific genetic variants. In contrast with the
8 longitudinal analyses, Weisskopf et al. (2007b) found that repeat measures of bone Pb
9 levels were inconsistently associated with cognitive function (improved and poorer
10 performance) in cross-sectional analyses. Among 720 NAS men 45 years of age and
11 older, higher concurrent blood and bone Pb levels were associated with lower MMSE
12 scores among 149 ALAD-2 carriers (Weuve et al.. 2006). with a larger decrease found
13 for an increase in blood Pb level. A 3 (ig/dL higher concurrent blood Pb level (the
14 interquartile range) was associated with a 0.26 point lower mean MMSE score (95% CI:
15 -0.54, -0.01) among ALAD-2 carriers and a 0.04 point lower score (95% CI: -0.16, -0.07)
16 among noncarriers. A subsequent NAS analysis (n = 486-959) did not find a consistent
17 direction of modification of the association of blood or bone Pb levels with tests of
18 cognitive function in various domains by ALAD genotype (Rajan et al.. 2008). An
19 interaction between higher tibia Pb level and ALAD-2 genotype was found only for
20 visuospatial skills (constructional praxis test), and between patella Pb level and ALAD-2
21 genotype for perceptual speed (pattern comparison test). The potential direction of effect
22 modification by the ALAD-2 genotype is not clear as the greater affinity of the ALAD-2
23 enzyme subunit for Pb may increase risk of Pb-associated health effects by increasing
24 blood Pb levels, or it may diminish Pb-associated health effects by sequestering Pb in the
25 bloodstream and decreasing its bioavailability.
26 Cross-sectional studies examined a larger number of potential confounding factors than
27 did longitudinal analyses. The NAS found blood and bone Pb-associated decrements in
28 cognitive function with adjustment for dietary factors, physical activity, medication use,
29 and comorbid conditions (Raian et al.. 2008; Weuve et al.. 2006) (Table 5-10). As in the
30 BMS and NAS, tibia and patella Pb levels were more consistently associated with
31 cognitive performance than was blood Pb levels in 587 healthy women in the Boston,
32 MA area participating in the Nurses' Health Study (Table 5-10) (Weuve et al.. 2009).
33 Additional potential confounding factors examined in this group included use of aspirin,
34 ibuprofen, or vitamin E, mental health, and antidepressant use. Blood, patella, and tibia
35 Pb levels were measured between ages 47 and 74 years and an average of 5 years before
36 cognitive testing. Contrary to expectation, higher patella and tibia Pb levels were
37 associated with higher scores on the "f" naming test (naming words that begin with f). In
38 separate models, the "f' naming test was omitted from a composite index of all cognitive
November 2012 5-133 Draft - Do Not Cite or Quote
-------
1 tests performed by phone, and a one SD higher tibia Pb level was associated with
2 0.051-point lower (95% CI: -0.10, -0.003) composite cognitive function z-score (Table
3 5-10). A similar magnitude of decrease was estimated for an increase in age of 3 years in
4 these women. The magnitude of association was smaller for an SD increase in patella Pb
5 level (-0.033 [95% CI: -0.080, 0.014]), and a weak association was found for an SD unit
6 increase in blood Pb level (-0.016 [95% CI: -0.071, 0.039]).
7 Several analyses of the large, U.S.-representative NHANES III (1991-1994) population
8 of men and women investigated effect modification by age and genetic variants. Only
9 blood Pb levels were available and were measured in samples collected concurrently with
10 cognitive testing. These analyses adjusted for several of the same potential confounding
11 factors as other studies, with the exception of smoking. Krieg and Butler (2009) found
12 blood Pb level to be associated weakly with poorer performance on tests of learning and
13 visuospatial skills among adults ages 20-39 years and inconsistently in adults ages 40-59
14 years. Krieg et al. (2009) further found inconsistent associations with word and story
15 recall in adults ages > 60 years. Because of the different types and numbers of tests
16 administered, it is difficult to compare findings between adults less than and greater than
17 age 60 years. In the subset of the population with genetic analysis, blood Pb-cognitive
18 function associations were not found to be modified by ALAD genetic variants in a
19 consistent direction (Krieg et al.. 2009). Among adults ages 20-59 years and > 60 years,
20 higher concurrent blood Pb level was associated with a larger decrement in performance
21 on some tests in ALAD-2 carriers and other tests in ALAD -1 subjects (Table 5-10).
22 Krieg et al. (2010) found differences in the association between concurrent blood Pb level
23 and scores on a symbol-digit substitution test by the VDR variants, rs731236 and VDR
24 rs2239185, and by the VDR haplotype, which have unclear functional relevance. Similar
25 to observations in adolescent NHANES participants (Section 5.3.2.5). results were not
26 uniform across the various tests. However, for several tests, blood Pb level was
27 associated with greater decrements in cognitive performance among adults with the CC
28 genotypes of VDR variants.
29 Other cross-sectional studies with fewer subjects generally produced results consistent
30 with those from the larger studies described above. A study of 188 rural Chinese men and
31 women found a weak association between higher plasma Pb levels and a lower composite
32 cognitive score based on a battery of in-person administered tests (Gao et al.. 2008).
33 Results were adjusted for age, sex, education, BMI, or APOE-e4 in individual ANCOVA
34 analyses. Smoking and alcohol use were not associated with cognitive performance in
35 this group. Pb in plasma is not bound to erythrocytes, as is about 99% of blood Pb, and is
36 the fraction delivered directly to soft tissue (Chuang et al.. 2001; Hernandez-Avila et al..
37 1998). The results of Gao et al. (2008) may provide information on the cognitive effects
38 of a more bioavailable fraction of Pb dose; however, because there is little investigation
November 2012 5-134 Draft - Do Not Cite or Quote
-------
1 of plasma Pb, firm conclusions are not warranted. Among 47 men and women in
2 Rochester, NY (age 55-67 years), subjects in the highest two tertiles of calcaneal bone
3 (heel bone with higher turnover rate than tibia) Pb level performed worse on delayed
4 matching-to-sample and paired associated learning tasks than subjects in the lowest tertile
5 (exact Pb levels in tertiles not reported) (van Wijngaarden et al. 2009). In analyses of
6 tibia Pb levels, subjects in the highest tertile of tibia Pb level did not consistently perform
7 worse on the various cognitive tests (Table 5-10).
Adults with Occupational Pb Exposures
8 The 2006 Pb AQCD (U.S. EPA. 2006b) concluded that in adults, blood Pb levels were
9 associated with cognitive function more consistently among those with occupational Pb
10 exposures. These findings were supported by results from a few recent studies of
11 occupationally-exposed adults. Several of these associations were found with adjustment
12 for fewer but a similar set of potential confounding factors as in nonoccupational studies;
13 however, other occupational exposures were not considered. A prospective analysis was
14 conducted in former male Pb battery workers whose occupational exposure had ceased
15 0.02 to 16 years (median: 6) before follow-up testing in 2001-2004 (Khalil et al.. 2009a).
16 Subjects included 83 of 288 workers (in 2004 mean age: 54 years, median tibia Pb level:
17 57 ug/g) and 51 of 181 controls (mean age: 55 years, median tibia Pb level: 12 ug/g)
18 from the 1982 Lead Occupational Study in Pennsylvania. While the follow-up
19 participation was low, participation was not biased to poor performers on cognitive tests
20 at baseline. In former Pb-exposed workers, a 10 ug/g higher peak tibia Pb levels was
21 associated with a -0.352 change in total cognitive function score (e.g., learning, memory,
22 executive function, general intelligence, spatial function, psychomotor speed) between
23 1982 and 2004. In controls, higher tibia Pb levels were associated with improved
24 performance on several tests. Results were adjusted for age, education, income, blood
25 pressure, years of employment, years since last worked, smoking, alcoholic drinks/week,
26 and baseline score. Cross-sectional associations indicated stronger associations of
27 concurrent tibia Pb level than concurrent blood Pb level (median: 12 ug/dL) with poorer
28 cognitive performance in former Pb-exposed workers. In controls, higher concurrent
29 blood Pb levels were associated with larger decrements in cognitive performance. As in
30 nonoccupationally-exposed adults, the stronger findings for tibia Pb levels in former
31 Pb-exposed workers indicate stronger effects of long-term cumulative Pb exposures than
32 recent exposures on cognitive function. The associations for concurrent blood Pb levels
33 in controls also may reflect effects of past exposures.
November 2012 5-135 Draft - Do Not Cite or Quote
-------
1 Blood and tibia Pb levels also were associated with cognitive performance in a follow-up
2 of 652 Pb-exposed workers (mean age: 43.4 years, mean blood Pb level: 30.9 ug/dL) in
3 Korea, whose patella Pb levels were measured (Dorsey et al., 2006). Higher patella Pb
4 levels were associated with poorer manual dexterity, executive function, and verbal
5 memory with adjustment for age, sex, education, and job duration. The associations for
6 patella Pb level were not as strong as those previously found for either blood or tibia Pb
7 levels in these workers (Schwartz et al., 2005; Schwartz et al., 2001).
8 Other occupational studies aimed to characterize factors that either mediate or modify the
9 association between Pb biomarkers and cognitive function. Both a working lifetime time-
10 weighted integrated blood Pb level (an index of cumulative exposure) (p = 0.09) and tibia
11 Pb level (p = 0.08) were associated with longer times to complete the grooved pegboard
12 test among current Pb smelter workers (Bleecker et al.. 2007b). In the same workers
13 (n = 112, mean age: 38 years), higher time-weighted integrated blood Pb level was
14 associated with decrements in executive function, learning, and memory among those
15 with lower cognitive reserve (i.e., > 12th grade reading level by Wide Range
16 Achievement Test-R) (Bleecker et al.. 2007a). Subjects with lower and higher cognitive
17 reserve were matched by blood Pb level (mean: 26 ug/dL), and results were adjusted for
18 age, depression scale, and current alcohol use.
19 Apolipoprotein E is a transport protein for cholesterol and lipoproteins and has been
20 found to regulate synapse formation (connections between neurons). A genetic variant,
21 called the ApoE-e4 allele is a haplotype between 2 exonic SNPs and has been associated
22 with a two-fold increased risk of developing Alzheimer's disease, although the majority
23 of such individuals still do not develop the disease. Thus, it is biologically plausible that
24 ApoE-e4 carriers may be biologically susceptible to cognitive dysfunction. A study of
25 529 U.S. male, former tetra-ethyl Pb workers found that higher peak tibia Pb levels were
26 associated with lower scores on tests of executive function, vocabulary, and memory
27 (Stewart et al.. 2002). and for several tests, larger decrements among the 118 men with at
28 least one ApoE-e4 allele. Results were adjusted for age, race, education, depression,
29 testing technician, and visit number. The group with at least one ApoE-e4 allele had
30 slightly higher peak tibia Pb levels (mean: 26.2 versus 23.1 ug/g) and a larger percentage
31 of non-white subjects but were similar in age, education, and time since employment.
Summary of Cognitive Function in Adults
32 In summary, consistent with evidence described in the 2006 Pb AQCD, recent studies
33 found that higher bone Pb levels were associated decrements in cognitive function in
34 adults without occupational Pb exposure (Table 5-10). Much of this evidence was
35 provided by analyses of the BMS and NAS, with additional findings reported in the
November 2012 5-136 Draft - Do Not Cite or Quote
-------
1 Nurses' Health Study and smaller populations. Nonetheless, the multiple risk factors and
2 health outcomes examined in most of these cohorts reduces the likelihood of selection
3 bias by Pb exposure or cognitive function. While the NAS and Nurses' Health Study
4 included primarily white men and white women, respectively, the BMS examined a more
5 diverse population of men and women of several different race/ethnicities. There was
6 variability in associations across the various domains of cognitive function tested within
7 studies; however, higher bone Pb levels were associated with decrements in most of the
8 tests performed. In several populations, higher bone Pb levels were associated with
9 decrements in executive function, visuospatial skills, learning, and memory.
10 Key evidence for bone Pb-associated cognitive decrements was provided by recent
11 prospective analyses that demonstrated that higher tibia (means 18.8, 20 (ig/g) and patella
12 (mean 25 (ig/g) bone Pb levels measured at baseline were associated with subsequent
13 declines in cognitive function over 2- to 4-year periods (Bandeen-Roche et al., 2009;
14 Weisskopf et al.. 2007b). These findings indicate that long-term Pb exposure may
15 contribute to ongoing declines in cognitive function in adults. These associations were
16 found with adjustment for potential confounding by age, education, smoking, and alcohol
17 use in the NAS and age, sex, race, household wealth, and education in the BMS.
18 Supporting evidence was provided by most cross-sectional analyses that adjusted for
19 several of the potential confounding factors described above plus dietary factors, physical
20 activity, medication use, and comorbid conditions (Rajan et al., 2008; Weuve et al..
21 2006). Cross-sectional studies generally demonstrated larger decrements in cognitive
22 function in adults in association with tibia or patella Pb levels than with concurrent blood
23 Pb levels. In comparisons of associations with patella and tibia Pb levels in the NAS and
24 Nurses' Health Study, tibia Pb levels were not consistently associated with larger
25 decreases in cognitive performance (Weuve et al.. 2009; Weisskopf et al.. 2007b). In
26 NHANES analyses, concurrent blood Pb levels were associated with lower cognitive
27 function in particular age and genetic variant subgroups but not consistently across the
28 various cognitive tests evaluated (Krieg et al.. 2010; Krieg and Butler. 2009; Krieg et al.,
29 2009). NHANES did not have bone Pb measures for comparison.
30 Because bone Pb is a major contributor to blood Pb levels, blood Pb level also can reflect
31 longer term exposures, including higher past exposures, especially in adults without
32 occupational exposures. Thus, in the NHANES results, it is difficult to characterize the
33 relative contributions of recent and past Pb exposures to the associations observed
34 between concurrent blood Pb level and cognitive function. In other cohorts, the
35 discrepant findings for blood and bone Pb levels indicate that cumulative Pb exposure
36 that likely included higher past exposures, may be a better predictor of cognitive function
37 in adults than is blood Pb level. Additional support for the effects of cumulative or past
November 2012 5-137 Draft - Do Not Cite or Quote
-------
1 Pb exposure is provided by analyses of a few child cohort as adults, which indicated that
2 childhood tooth (from ages 5-8 years) and blood Pb levels (e.g., age 10 years) were
3 associated with decrements in cognitive function in adults ages 19-30 years) (Mazumdar
4 et al.. 2011; Bellinger et al.. 1994a). An uncertainty related to the evidence for cognitive
5 function decrements associated with bone Pb levels is the potential residual confounding
6 by age. Although studies adjusted for age, the high correlation between increasing age
7 and bone Pb levels (Section 4.3.5) makes it difficult to distinguish the independent effect
8 of Pb exposure. However, the coherence with evidence for cognitive function decrements
9 associated with long-term Pb exposure in animals provides support for associations
10 observed in human adults.
11 Cross-sectional analyses provided information on potential effect modification of bone
12 Pb- and blood Pb-associated decrements in cognitive function in adults by race,
13 psychosocial stress, and genetic variants. Inconsistencies were found for effect
14 modification by race in the BMS, ALAD-2 genotype in the NAS and NHANES, and
15 VDR genotype in NHANES. Larger tibia Pb-associated decrements in cognitive function
16 was found in NAS men with HFE variants and in BMS subjects living near more
17 psychosocial hazards. Evidence does not clearly indicate whether the observed effect
18 modification reflects chance, a change in the toxicokinetics of Pb that alters Pb dose at
19 the biological site of action, or a direct biological interaction that increases the toxicity of
20 Pb in the target tissue. However, such effect modification serves to strengthen inferences
21 about associations between Pb biomarkers and cognitive function since it is unlikely that
22 potential confounding factors vary by levels of the modifying factor, particularly genetic
23 variants. However, because there is little available evidence and inconsistent evidence for
24 some factors, firm conclusions regarding effect modification are not warranted.
25 In contrast with nonoccupationally-exposed adults, in adults with former and current
26 occupational Pb exposures, cognitive function decrements were associated with both
27 blood (means: 12 [former workers]-31 ug/dL) and bone Pb levels. Thus, among
28 Pb-exposed workers, both current and cumulative Pb exposures may affect cognitive
29 function. Several of these studies considered confounding by a similar set of potential
30 confounding factors as studies of adults without occupational Pb exposures but did not
31 consider other occupational exposures. In the prospective study of former Pb workers,
32 peak tibia Pb levels were associated more strongly with cognitive performance than were
33 blood Pb levels (Khalil et al.. 2009a). Thus, in the absence of higher current Pb
34 exposures, cumulative Pb exposures may have a greater effect on cognitive function in
35 adults.
November 2012 5-138 Draft - Do Not Cite or Quote
-------
5.3.3 Attention-related Behavioral Problems in Children
1 The effects of Pb exposure on attention-related behavioral problems such as inattention,
2 impulsivity, hyperactivity, and ADHD have not been examined as extensively as effects
3 on cognition. Behavioral effects are more complex to study than are cognitive effects,
4 particularly FSIQ. There are fewer objective tests of attention-related behavioral
5 problems with as strong psychometric properties or as rigorous validation as IQ tests. In
6 several studies, attention-related behavioral problems were assessed using teacher and/or
7 parent ratings which are subject to greater measurement error. However, domain-specific
8 neuropsychological assessments are advantageous as they may provide greater insight
9 into whether there is a particular domain more susceptible to the effects of Pb exposure.
10 As with cognitive function, in the evaluation of epidemiologic evidence for attention-
11 related behavioral problems, greater emphasis was placed on evidence from
12 neuropsychological tests than from parent or teacher ratings and prospective studies with
13 repeated assessments of blood Pb levels and behavior, studies of older children in whom
14 outcomes are more reliably measured, and studies of children whose blood Pb levels are
15 less influenced by higher past Pb exposures. Similar to cognitive function, associations
16 between blood Pb levels and attention-related behavioral problems potentially may be
17 confounded by factors such as parental SES, education, and IQ, quality and stability of
18 the caregiving environment, and nutritional status. Accordingly, greater weight was given
19 to studies with greater consideration for potential confounding in the study design or in
20 statistical analyses. Consideration also was given to studies assessing effects relevant to
21 blood Pb levels in contemporary U.S. children (i.e., <5 (ig/dL).
22 Some studies that found associations between concurrent blood Pb levels and attention-
23 related behavioral problems were not major considerations in drawing conclusions and
24 are not discussed in detail in the following sections. These studies examined populations
25 that have high prevalence of prenatal alcohol or drug exposure (Chiodo et al.. 2007;
26 2004). or had earlier childhood chelation (Chen et al.. 2007). and thus may not be
27 representative of current children in the U.S. population. Others had limited consideration
28 for potential confounding (Liu et al.. 20 lib), or examined infants in whom inattention
29 ratings may not predict behavior in later childhood (Plusquellec et al.. 2007).
November 2012 5-139 Draft - Do Not Cite or Quote
-------
5.3.3.1 Inattention and Impulsivity
Epidemiologic studies of Inattention and Impulsivity in Children
1 Attention is the ability to maintain a consistent focus on an activity or relevant stimuli
2 and can be assessed by examining sustained attention, impulsivity, or distractibility.
3 Several epidemiologic studies reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b)
4 reported associations between blood, tooth, or bone Pb levels with inattention in children
5 ages 8-17 years, including prospective studies described in previous sections for
6 cognitive function (Ris et al., 2004; Fergusson et al., 1993; Leviton et al., 1993). As
7 described in this section, recent studies also found associations of blood Pb levels with
8 inattention in children ages 8-17 years. Many previous studies of inattention included
9 children with higher blood Pb levels than those of most current U.S. children. Recent
10 studies, most of which were cross-sectional, provided evidence of blood Pb-associated
11 inattention in populations of children with mean concurrent blood Pb levels 2 to 5 (ig/dL
12 (Cho et al., 2010; Nicolescu et al.. 2010; Plusquellec et al., 2010); however, limitations
13 include the cross-sectional design of studies and potential influence of higher past Pb
14 exposures. In the collective body of literature, most evidence was for inattention rated by
15 teachers, parents, or blinded examiners; however, associations were consistently found
16 for more objective measures such as the continuous performance test (CPT) (Figure 5-10
17 and Table 5-11). Thus, evidence does not indicate undue influence by biased reporting of
18 inattention by parents of children with high Pb exposures. Epidemiologic findings for
19 inattention and impulsivity are supported by the coherence of findings in Pb-exposed
20 animals of poorer response inhibition in Schedule Controlled Behavior Tests and poorer
21 performance on signal detection tests with distracting stimuli (discussed below). In
22 particular, both evidence in children and animals indicates Pb-associated poorer
23 performance on test of response inhibition, i.e., continued responses to stop signals.
24 Most studies that assessed inattention with the objective CPT found associations with
25 blood Pb level (Figure 5-10 and Table 5-11). including prospective studies in Cincinnati
26 and in Chelsea/Sommerville, MA, which indicated associations of higher prenatal or
27 earlier childhood blood Pb levels or tooth Pb levels with increases in commission and
28 omission errors or reaction time in adolescents and young adults (Ris et al.. 2004;
29 Bellinger et al.. 1994a). In the CPT, subjects are assessed for their ability to maintain
30 focus during a repetitive task and respond to targets or inhibit responses. These findings
31 from prospective studies characterized the temporal sequence between Pb exposure and
32 inattention better than cross-sectional studies and made reverse causation a less likely
33 explanation for observed associations. These studies recruited cohorts from schools or
34 prenatal clinics and had moderate to high follow-up participation that was not conditional
35 on blood or tooth Pb levels, which reduces the likelihood of selection bias. Among
November 2012 5-140 Draft - Do Not Cite or Quote
-------
1 primarily white, higher SES young adults ages 19-20 years, Bellinger et al. (1994a) found
2 that compared with the group with age 5-8 year tooth Pb levels 2.9-5.9 ppm, the group
3 with tooth Pb levels >19.9 ppm had fewer correct response on the CPT and had longer
4 reaction times for correct responses but did not commit more commission errors
5 (responding to a nontarget). In the mostly African-American, lower SES Cincinnati
6 cohort, Ris et al. (2004) found increased inattention (composite of CPT outcomes) in
7 association with prenatal maternal, age 3-60 month average, and age 78 month blood Pb
8 levels in adolescents ages 15-17 years, particularly among males. Although blood Pb
9 levels at older ages were not examined and results do not exclude an effect of more recent
10 Pb exposures, the combined evidence from these prospective studies points to an effect
11 on inattention of cumulative earlier childhood Pb exposures. Both studies considered
12 several potential confounding factors, including SES, parental IQ, maternal education,
13 and self drug use. HOME score was considered only in the Cincinnati cohort. Ris et al.
14 (2004) also considered potential confounding by prenatal drug and alcohol exposure,
15 birth outcomes, and iron status.
16 Recent studies that assessed inattention with neuropsychological tests, primarily in non-
17 U.S. populations, found Pb-associated increases in inattention, although all were cross-
18 sectional design and did not consider potential confounding by parental caregiving
19 quality. Further, sufficient data were not provided to assess whether participation was
20 biased to those with higher Pb exposure and inattention. A study in children ages 8-11
21 years in Korea demonstrated poorer performance on some indices of the CPT with
22 relatively low blood Pb levels (mean 1.9 (ig/dL) (Cho et al.. 2010): however, the
23 contribution of higher earlier Pb exposures cannot be excluded. Specifically, higher
24 concurrent blood Pb levels were associated with more commission errors, but weakly
25 with other parameters of the CPT (Figure 5-10 and Table 5-11). Results were adjusted for
26 age, sex, paternal education, maternal IQ, child IQ, city of residence, birth weight, and
27 urinary cotinine, the latter of which was more strongly associated with CPT performance
28 than blood Pb level and the primary cause of attenuation of blood Pb effect estimates.
29 Cho et al. (2010) found that mean blood Pb levels were similar in children with and
30 without (1.80 and 1.93 (ig/dL, respectively, p = 0.32) parental report of history of
31 neuropsychiatric disease (e.g., ADHD, learning disability, depression, obsessive-
32 compulsive disorder); however, history may not accurately represent current parental
33 caregiving quality.
November 2012 5-141 Draft - Do Not Cite or Quote
-------
Study
Risetal. (2004)
Choetal. (2010)
Nicolescuetal. (2010)
Niggetal.(2008)
Chiodoetal.(2007)a
Chiodoetal.(2004)a
Bumsetal. (1999)
Wasserman etal. (2001)
Canfieldetal. (2003b)
Nicolescuetal. (2010)
Silvaetal. (1988)
Plusquellecetal.(2010)
Kordasetal.(2007)
Royetal. (2009a)
Chiodoetal.(2007)a
Chiodo etal. (2004)=
Nicolescuetal. (2010)
Silvaetal. (1988)
Plusquellecetal.(2010)
Royetal. (2009a)
Niggetal. (2008)=
Chiodoetal.(2007)a
Chen etal. (2007)
Choetal. (2010)
Chen etal. (2007)
Mean (SD) blood Blood Pb
Pb(ug/dL) interval analyzed
NR NR
1.9(0.67)
3.7 (2.6)
1.04(0.53)
5.0 (3.0)
5.4 (3.3)
5.4 (3.3)
GM: 14.3(13.5-15.1)
GM: 13.9(13.2-14.6)
6.5(1.5)
6.5
3.7 (2.6)
11.1 (4.9)
5.4 (5.0)
11.5(6.1)
11.4(5.3)
5.0 (3.0)
5.4 (3.3)
3.7 (2.6)
11.1 (4.9)
5.4 (5.0)
11.4(5.3)
1.04(0.53)
5.0 (3.0)
12.0(5.2)
1.9(0.67)
12.0(5.2)
1.2-2.8
2.0-8.5
0.5-1.7
2.1-8.7
2.3-9.5
2.3-9.5
13.7-14.9
13.3-14.4
4.7-8.4
Data N/A
2.0-8.5
5.9-10
1.4-10.8
5.4-10
5.8-10
2.1-8.7
2.3-9.5
2.0-8.5
5.9-10
1.4-10.8
5.8-18.3
0.5-1.7
2.1-8.7
6.5-10
1.2-2.8
6.5-10
Outcome
Inattention composite, Continuous Performance Test
Comissions errors, Continuous Performance Test
Response time — <
Go/no go, Test of Attentional Performance
Stoptask
Comission errors, Continuous Performance Test ^
Omission errors
Number of errors, Continuous Performance Test
Comissionserrors
Reaction Time
Attention problems, boys, Child BehaviorChecklist —
Attention problems, girls
Attention problems, Child BehaviorChecklist H
Inattention, control phase, Shape School
Inattention , com pleted phases
Inattention, parent, Conners —
Inattention, teacher
Inattention, parent, Rutter —
Inattention, teacher
Off task duration, InfantBehavioral Rating Scale
Impulsivity
Off task passive
Inattention, Conners
Inattention, AchenbachTeacher Report Form
Inattention, Child Behavior Checklist
Hyperactivity, parent, Conners
Hyperactivity, teacher
Hyperactivity, parent, Rutter
Hyperactivity, teacher
Global activity scale, InfantBehavioral Rating Scale
Hyperactivity, Conners
Hyperactivity/impulsivity, Child BehaviorChecklist
Hyperactivity, teacher, PROBS-14
Hyperactivity,' indirect —
Total ADHD rating, teacher, Korean ADHD Rating Scale
Total ADHD rating, parent
ADHD index, indirect
Inattention/
» * Tmpulsivity
-*-
— »
*fc
w&
A It
* . '
a —
i
•
•—
-•—
— •
•
•
-•-
-m-
•
•
« Hyperactivity
:
•
•
0
t* Total ADHD
^ Rating
-0.5
-0.3
-0.1
0.1
0.3
0.5
Standard deviation change perl |jg/dL increase in
blood Pb level in various intervals of blood Pb level
aStandard errors were estimated from p-values or sufficient data were not provided to calculate 95% CIs.
Note: Regression coefficients were scaled to their standard deviation to facilitate comparisons among tests with different scales.
Small effect estimates should not necessarily indicate lack of effect or weak effect. Results are categorized by outcome category:
inattention/impulsivity (with objective tests presented first), hyperactivity ratings, total ADHD ratings. Within categories, results
generally are presented in order of strength of study design. Effect estimates are standardized to a 1 ug/dL increase in blood Pb
level in the interval from the 10th percentile to the 90th percentile or 10 ug/dL, whichever was lower. For studies with 10th
percentiles of blood Pb level > 10 ug/dL, effect estimates are standardized to a 1 ug/dL increase in blood Pb level in the interval
from the 10th to 90th percentile of blood Pb level. The percentiles are estimated using various methods and are approximate values.
Effect estimates are assumed to be linear within the 10th to 90th percentile interval of blood Pb level. Gray, orange, blue, and black
symbols represent associations with lifetime average, prenatal (maternal), earlier childhood, and concurrent blood Pb levels,
respectively.
Figure 5-10 Associations of blood Pb levels with attention-related behavioral
problems in children.
November 2012
5-142
Draft - Do Not Cite or Quote
-------
Table 5-11 Additional characteristics and quantitative results for studies
presented in Figure 5-10.
Study
Bellinger et al.
(1994a)
Risetal.
(2004)
Cho et al.
(2010)
Nicolescu et al.
(2010)
Study Population and
Methodological Details
79 young adults, born 1970, followed
from first grade to age 1 9-20 yr, Boston,
MA area
Prospective. Moderate follow-up
participation. Participation from higher
SES, females, higher initial IQ but no
affect on association with tooth Pb level.
Regression model adjusted for parental
IQ, sex, SES, current drug, alcohol and
illicit drug use, maternal education and
age, birth order. Also considered
potential confounding by other
unspecified factors.
195 children followed prenatally
(1 979-1 985) to age 1 5-1 7 yr, Cincinnati,
OH
Prospective. Recruitment at prenatal
clinic. High follow-up participation, no
selective attrition. Mostly African
American. Linear regression model
adjusted for SES, maternal IQ, HOME,
adolescent marijuana use, obstetrical
complications. Also considered potential
confounding by birth outcomes, maternal
age, prenatal smoking, alcohol,
marijuana, and narcotics use, gravidity,
# previous abortions, stillbirths, parity,
caregiver education, public assistance,
child age, sex, health, Fe status.
667 children ages 8-1 1 yr, born
1 997-2000, 5 Korean cities
Cross-sectional. School-based
recruitment, moderate participation rate.
Log linear regression model adjusted for
age, sex, parental education, maternal
IQ, child IQ, birth weight, urinary
cotinine. Did not consider potential
confounding by parental caregiving
quality.
83 children ages 8-1 2 yr (born
1995-1999), Bucharest and Pantelimon,
Romania
Cross-sectional. Pantelimon near
former metal processing plant. Low
correlations blood Pb with blood Al, Hg.
No information on participation rate. Log
linear regression model adjusted for city,
sex, age, computer experience,
handedness, eye problems, # siblings,
parental education, prenatal smoking
and alcohol use, parental ever having
psychological/psychiatric problem.
Blood /
tooth Pb
Levels
(ug/dL
or ug/g)
Deciduous
tooth (age
5-8 yr)
Q1:
2.9-5.9 ppm
Q2:
6.0-8.7 ppm
Q3:
8.8-19.8 ppm
Q4: 19.9-51.8
ppm
Multiple time
periods
Mean (SD):
Not Reported
Concurrent
|\yia~,n /onV
IvIcaM ^OU^.
1.9(0.67)
Interval
analyzed:
1.2-2.8 =
1 Oth-90th
percentiles
Concurrent
Median
(IQR): 3.7
(2.6)
Interval
analyzed:
2.0-8.5 =
1 0th-90th
percentiles
Outcome
Mean (SE) Correct
Responses per quartile
Mean (SE) Reaction time
errors per quartile
Continuous Performance
Test (CPT)
Ages 19-20yr
Inattention composite,
CPT
Prenatal (maternal)
3-60 mo avg
78 mo
Ages 15-17 yr
Commission errors, CPT
Response time, CPT
Total ADHD rating,
teacher
Total ADHD rating,
parent
Korean ADHD Rating
Scale IV
Ages 8-1 1 yr
Go/no go, KITAP
Inattention parent rating
Inattention teacher rating
Hyperactivity, parent
rating
Hyperactivity, teacher
rating
German Conners Rating
Ages 8-1 2 yr
Effect Estimate
/OCO/ ^l\^
(95% Cl)
Q1: 98.0(1.0)b
Q2: 97.6(1.1)
Q3: 96.9(1.1)
Q4: 94.6 (1.1)
Q1: 361.2(16.5)
Q2: 374.2(17.3)
Q3: 370.7(17.9)
Q4: 385.0(17.1)
0.16(0.04,0.27)
0.11 (0.04,0.19)
0.12(0.02,0.22)
0.03 (-0.01 , 0.07)°
-0.01 (-0.05, 0.03)°
0.042(0.017,0.067)°
0.010 (-0.01 3, 0.033)°
8.9% (-1.3, 19.3)
1 .3% (-3.3, 5.9)d
4.5% (-1.3, 10.3)d
6.3% (-0.60, 13.2)d
5.2% (-2.4, 12.8)d
November 2012
5-143
Draft - Do Not Cite or Quote
-------
Study
Nigg et al.
(2008)
Needleman et
al. (1979)
Chiodo et al.
(2007)
Chiodo et al.
(2004)
Study Population and
Methodological Details
1 50 children ages 8-1 7 yr, Birth yr and
location NR
Case-control study of ADHD.
Recruitment with advertisements. Could
have biased participation by Pb
exposure. Regression-based path
analysis adjusted for sex and income.
Did not consider potential confounding
by parental education or caregiving
quality.
158 children in 1st/2nd grade (born
1968-1971), Chelsea, Sommerville, MA
Cross-sectional. Recruitment from
schools. Only 6.7% selected based on
low and high tooth Pb levels. Moderate
participation rate but no selective
participation based on tooth Pb or
teacher ratings. Analysis of covariance
adjusted for paternal SES, maternal age,
# pregnancies, maternal education and
parental IQ. Did not consider potential
confounding by parental caregiving
quality.
506 children, age 7 yr (born 1982-1984),
Detroit, Ml area.
Cross-sectional. Recruitment at
prenatal clinic. All African American.
High prevalence prenatal drug exposure.
High follow-up participation. Linear
regression model adjusted for child sex,
prenatal marijuana use (commission
errors), caregiver education, HOME,
maternal IQ, cocaine use, prenatal
alcohol use and cigarettes/day (omission
errors), child age and sex (hyperactivity),
child age, caretaker education, SES,
HOME, maternal age and IQ, prenatal
alcohol use, current marijuana use
(inattention). Also considered potential
confounding by # children in home,
caretaker marital status, concurrent
alcohol/week, current maternal
cigarettes/day, caregiver concurrent
psychological symptoms, maternal
custody.
246 children, age 7.5 yr, Detroit, Ml area
Cross-sectional. Recruitment at
prenatal clinic. All African American.
High prevalence prenatal alcohol
exposure. High participation rate. Log
linear regression model adjusted for
SES (all outcomes). Prenatal smoking
exposure, caregiver vocabulary (# errors
CPT), caregiver vocabulary, child age
(commission errors, CPT). HOME,
prenatal alcohol and smoking exposure,
disruption in caregiver (Inattention
rating). Also considered potential
confounding by caregiver education,
family functioning, # children <1 8 years,
maternal prenatal marijuana, smoking,
or cocaine use, parity, crowding, child
sex, child life stress, caregiver age, life
stress, and psychology, conflict tactics.
Blood /
tooth Pb
Levels
(ug/dL
or ug/g)
Concurrent
|\yia~,n /onV
IvIcaM ^OU^.
8-11 yr: 1.04
(0.53)
1 2-1 7 yr:
1 .03 (0.54)
Interval
analyzed:
0.5-1 .7 =
1 0th-90th
percentiles
Tooth Pb
(1st/2nd
grade)
High
>27.0 ppm
(n = 58)
Low
<5.1 ppm
(n = 1 00)
Concurrent
Mean (SD):
5.0 (3.0)
Interval
analyzed:
2.1-8.7 =
1 Oth-90th
percentiles
Concurrent
Mean (SD):
5.4 (3.3)
Interval
analyzed*
2.3-9.5 =
1 0th-90th
percentiles
Outcome
Stop task
Hyperactivity/impulsivity
Teacher, parent rating,
Child Behavioral
Checklist, ADHD Rating
Scale
Ages 8-1 7 yr.
Reaction time, 12 sec
delay
% negative response,
teacher rating
Impulsive
Hyperactive
Commission Errors (%),
CPT
Omission Errors (%),
CPT
Inattention, PROBS-14
Hyperactivity, Achenbach
Teacher Report Form
Anp 7 vr
y y
Number of errors, CPT
Commission errors, CPT
Reaction time, CPT
Inattention, Examiner
rating
Child Behavior Checklist
Age 7.5 yr
Effect Estimate
/OCO/ ^l\^
(95% Cl)
0.38(0.16,0.60)°
0.21 (0, 0.42)°
Mean (SD)
Low tooth Pb: 0.35
(0.08)
High tooth Pb: 0.37
CO 09)
\\j.\j*jf
High vs. low tooth Pb
25 vs. 9%, p = 0.01
16 vs. 6%, p = 0.08
-0.08, p >0.05e
0.18(0.07,0.29)°
0.13(0.03,0.23)°
0.13(0.03,0.23)°
0.35 (0, 0.69)°
0.05, p >0.05e
0.25, p >0.05e
0.15(0.04,0.26)°
November 2012
5-144
Draft - Do Not Cite or Quote
-------
Study
Fergusson et
al. (1993)
Chandramouli
et al. (2009)
Leviton et al.
(1993)
Burns et al.
(1999)
Study Population and
Methodological Details
878 children followed from birth to age
13yr, Christchurch, New Zealand
Prospective. Moderate follow-up
participation, attrition did not affect
results. Log linear regression model
adjusted for sex, ethnicity, maternal
education, family size, HOME, SES, #
schools attended. Also considered
potential confounding by ethnicity,
maternal age, paternal education,
breastfeeding duration, parental
smoking, child birth outcomes, residence
on busy roads, weatherboard housing.
488 children followed from age 30 mo
(born 1 991 -1992) to 7-8 yr, Avon, U.K.
Prospective. All births in area eligible.
Similar characteristics as U.K. census,
high participation at baseline and follow-
up. Participants had better educated
mothers, who smoked less, better home
environment. Regression model
adjusted for maternal education and
smoking, home ownership, home
facilities score, family adversity index,
paternal SES, parenting attitudes at 6
mo, child sex. Also considered potential
confounding by child IQ.
1 ,923 children followed from birth
(1979-1980) to age 8 yr, Boston, MA
Prospective. Recruitment from birth
hospital. High participation at baseline
and follow-up. Log linear regression
model adjusted for single parent family,
gestational age, maternal education,
ethnicity, # children, daycare in first
3 years. Also considered potential
confounding by other unspecified
factors.
322 children followed from birth
(1979-1 982) to age 11-1 Syr, Port Pirie,
Australia.
Prospective. Moderate follow-up
participation. Participants had higher
birth weight, older mothers, less
educated fathers. Log linear regression
model adjusted for maternal age,
prenatal smoking status, birth weight,
type of feeding, length of breastfeeding,
maternal education, maternal IQ,
paternal education, concurrent maternal
psychopathology, birth order, family
functioning, paternal occupation, parent
smoking, marital status, HOME, child IQ.
Blood /
tooth Pb
Levels
(ug/dL
or ug/g)
Tooth Pb
(age 6-8 yr)
Mean (SD):
6.2 (3.7) ug/g
Age 30 mo
Mean (SD):
Not Reported
Group 1 : 0-2
Group2: 2-5
Group 3:
5-10
Group 4: >10
Prenatal
(cord) Mean:
6.8
Tooth Pb
(Age 6 yr)
Mean: 3.3
Lifetime avg
(to age 11-13
yr) blood
Boys:
GM: 14.3
(5th-95th)
(13.5-15.1)
10th-90th:
13.7-14.9
Girls:
GM13.9
(5th-95th)
13.2-14.6)
10th-90th:
13.3-14.4
Outcome
Inattention/restlessness
Rutterand Conners'
ratings
Age 13yr
Selective inattention
Test of Everyday
Attention for Children
Ages 7-8 yr
Hyperactivity, teacher
Strengths and Difficulties
Questionnaire,
Ages 7-8 yr
Daydreaming
Prenatal (cord)
Tooth Pb
Boston Teacher
Questionnaire, Age 8 yr
Attention problems, boys
Attention problems, girls
Maternal rating by Child
Behavior Checklist
Age 11-1 Syr
Effect Estimate
(95% Cl)a
0.06(0, 0.12)b'°
OR vs. 0-2 ug/dLas
reference
2-5 ug/dL:
0.97 (0.62, 1 .52)'
5-10 ug/dL:
1.01 (0.64, 1.61)f
>10 ug/dL:
0.88(0.42,1.85)''
2-5 ug/dL:
0.84 (0.47, 1 .52)'
5-10 ug/dL:
1.25(0.67,2.33)'
>10ug/dL:
2.82(1.08,7.35)'
RR (yes/no) per loge
increase
Girls: 1.3 (0.8, 2.2)'
Boys: 1.0(0.6, 1.5)'
Girls: 1 .5 (0.9, 2.6)b'f
Boys: 1.1 (0.7, 1.7)"''
0.02 (-0.04, 0.09)
0.07(0.02,0.12)
November 2012
5-145
Draft - Do Not Cite or Quote
-------
Study
Wasserman et
al. (2001)
Canfield et al.
(2003b)
Silva et al.
(1988)
Plusquellec et
al. (201 0)
Study Population and
Methodological Details
191 children followed from birth to age
5 yr, Pristina, Yugoslavia.
Cross-sectional. High follow-up
participation. Participants had lower
maternal education, were Albanian, had
higher age 4 blood Pb. Generalized
estimating equations with log blood Pb
adjusted for child sex and age, ethnicity,
HOME, maternal education, birth weight,
maternal smoking. Did not consider
potential confounding by maternal IQ.
150 children born 1994-1995 followed
from age 6 mo to age 4.5 yr, Rochester,
NY
Cross-sectional. Recruitment from
study of dust control. 73% nonwhite.
High follow-up participation, no
comparison of nonparticipants. Linear
mixed effects model adjusted for age,
gestational age, maternal IQ and
education, HOME, race, color/shape
knowledge, child IQ (control phase),
birth order, marital status, race
(completed phases). Also considered
potential confounding by child sex, birth
weight, household income, prenatal
smoking exposure.
535 children age 1 1 yr (born
1972-1973), Dunedin, New Zealand
Cross-sectional. Moderate participation
rate. Participants were of higher SES
and non-Maori. Log linear regression
adjusted for SES, maternal verbal skills,
change in residence and school, solo
parenting, child/parent separation,
maternal age at first birth, family
relations, marriage guidance sought,
maternal mental health symptoms, child
sex, IQ. Did not consider potential
confounding by parental caregiving
quality.
90-98 children, ages 5-6 years (born
1993-1996), Inuit communities, Quebec,
Canada
Cross-sectional. Study of multiple
exposures. Low but no selective
participation by Pb, PCBs, Hg. Log linear
regression model adjusted for birth
weight, sex, parity, caregiver education
(impulsivity) and birth weight, SES, child
blood hemoglobin (off task duration).
Also examined potential confounding by
# children in home, residents per room,
caretaker psychological distress,
nonverbal reasoning, and linguistic
acculturation, HOME, prenatal alcohol
and illicit drug use and cigarettes/day,
serum Se, fatty acids.
Blood /
tooth Pb
Levels
(ug/dL
or ug/g)
Concurrent
Mean (SD):
6.5 (1 .48)
Interval
analyzed:
4.7-8.4 =
1 Oth-90th
percentiles
Concurrent
Mean: 6.5
10th-90th:
data not
available
Concurrent
Mean (SD):
11.1 (4.91)
Interval
analyzed: 5.9
(10th
percentile)-
10
Concurrent
Mean (SD):
5.4 (5.0)
Interval
analyzed:
1.4-10.8 =
1 0th-90th
percentiles
Outcome
Attention problems
Maternal rating, Child
Behavior Checklist
Repeated measures
ages 4-5 yr.
Inattention, control phase
Inattention, inhibit phase
Examiner rating during
Shape School Task
Repeated measures at
ages 4 and 4.5 yr
Inattention, parent
Inattention, teacher
Hyperactivity, parent
Hyperactivity, teacher
Rutter Behavior
Questionnaire
Off task duration
Impulsivity
Global activity rate
Examiner ratings
modified Infant
Behavioral Rating Scale
at ages 5-6 yr.
Effect Estimate
/OCO/ ^l\^
(95% Cl)
0 (-0.02, 0.02)
0.01 (-0.01,0.04)
0.008 (-0.02, 0.04)
0.06 (-0.03, 0.16)
0.15(0.06,0.25)
0.13(0.03,0.23)
0.12(0.01,0.22)
0.02 (0, 0.039)
0.019(0.001,0.036)
0.014(0.006,0.022)
November 2012
5-146
Draft - Do Not Cite or Quote
-------
Study
Kordas et al.
(2007)
Roy et al.
(2QQ9a)
Rabinowitz et
al. (1992)
Chen et al.
(2007)
Study Population and
Methodological Details
1 57 children ages 6-8 yr (born
1993-1995), Torreon, Mexico.
Cross-sectional. 26% of larger study
selected for classroom observation.
Residence near metal foundry. Linear
regression model adjusted forage, sex,
SES, home ownership, crowding in
home, maternal education, family
structure, forgetting homework. Also
considered potential confounding by
micronutrients but not parental
caregiving quality.
756 children ages 3-7 yr (born
1998-2003), Chennai, India
Cross-sectional. Recruitment at
schools. No information provided on
participation. Log linear regression
model adjusted for age, sex,
hemoglobin, average monthly income,
parental education, number of other
children, clustering in school and
classroom. Did not consider potential
confounding by parental caregiving
quality.
493 children, grades 1-3, Taiwan
Cross-sectional. Some reside near
smelter. High participation rate. Logistic
regression model adjusted for sex,
grade, # adults at home, child longest
hospital stay. Also considered potential
confounding by parental education, SES,
birth outcomes, handedness, language
at home, prenatal medicine, alcohol,
smoking but not parental caregiving
quality.
780 children in TLC trial followed ages
2-7 yr, Baltimore, MD; Cincinnati, OH;
Newark, NJ; Philadelphia, PA
Cross-sectional. Mostly African
American. 50% given chelation at ages
12-33 mo, blood Pb levels 20-44 ug/dL
No information on participation rate.
Regression-based path analysis
adjusted for city, race, sex, language,
parental education, parental
employment, single parent, age at blood
Pb measurement, caregiver IQ.
Considered potential confounding by
chelation but not parental caregiving
quality.
Blood /
tooth Pb
Levels
(ug/dL
or ug/g)
Concurrent
Mean (SD):
11.5(6.1)
Interval
analyzed: 5.4
(10th
percentile)-
10
Concurrent
Mean (SD):
11.4(5.3)
Interval
analyzed: 5.8
(10th
percentile)-10
Tooth Pb
(grades 1 -3)
Mean (SD):
4.6 (3.5)
Concurrent
Mean (SD)'
12.0(5.2) '
Interval
analyzed: 6.5
(10th
percentile)-
10
Outcome
Off task passive behavior
Examiner rating,
instrument developed by
investigator
Ages 6-8 yr
Inattention z-score
Hyperactivity z-score
Teacher ratings,
Conners' ADHD/DSM-IV
Scales Ages 3-7 yr.
Hyperactivity Syndrome
Boston Teacher
Questionnaire
Grades 1-3
Hyperactivity Index
ADHD index
Parent ratings, Conners
Scale-Revised, Age 7 yr
Effect Estimate
/OCO/ ^l\^
(95% Cl)
0.034 (0.005, 0.063)
0.031 (0.006, 0.056)
0.01 7 (-0.005, 0.039)
OR vs. <2.3 ug/g
reference
9 "V7 Lin/rr
^.o / HU'U- k
1.9(0.53,7.8)"
>7 ug/g:
2.8 (0.68, 2.8)b
Direct:
0.08 (-0.06, 0.22)
Indirsct'
0.04 (-0.06, 0.13)
Direct:
0.04 (-0.1 0,0.18)
Indirect:
0.07(0.03,0.11)
Direct = independent
of IQ.
Indirect = mediated
through IQ
November 2012
5-147
Draft - Do Not Cite or Quote
-------
Study
Froehlich et al.
(2009)
Study Population and
Methodological Details
2,588 children, ages 8-15 yr (born
1986-1996), U.S. NHANES 2001-2004
Cross-sectional. U.S. representative
results, study of multiple risk factors and
outcomes, high participation rate.
Logistic regression adjusted for current
household ETS exposure, sex, age,
race/ethnicity, income, preschool
attendance, maternal age, birth weight,
and interaction terms for Pb and prenatal
ETS interaction. Did not consider
potential confounding by parental
education orcaregiving quality.
Blood /
tooth Pb
Levels
(ug/dL
or ug/g)
Concurrent
Tertile 1: <0.8
Tertile 2:
0.9-1.3
Tertile 3: >1.3
Outcome
ADHD DSM-IV criteria
met
Parental rating, ADHD
DISC module
Age 8-1 5 yr
Effect Estimate
/OCO/ ^l\^
(95% Cl)
OR vs. <0.8 ug/dL
0.9-1 .3 ug/dL:
1.7(0.97,2.9)'
>1.3 ug/dL:
2.3(1.5,3.8)'
"Effect estimates are standardized to a 1 ug/dL increase in blood Pb level in the interval from the 10th percentile of blood Pb level
to 10 ug/dLorthe 90th percentile, whichever is lower and scaled to the standard deviation of the test score to facilitate
comparisons among tests that are scored on different scales. For studies with 10th percentiles of blood Pb level > 10 ug/dL, effect
estimates are standardized to a 1 ug/dL increase in blood Pb level in the interval from the 10th to 90th percentile of blood Pb level.
""Results for tooth Pb not presented in Figure 5-10.
""Standard error was estimated from the reported p-value.
dResults represent the change in false alarm rate.
Sufficient data were not provided to calculate 95% CIs.
'Results not presented in Figure 5-10 because OR or RR reported in papers.
1
2
o
J
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Response inhibition is a measure of impulsivity and in children has been assessed with
stop signal tasks, which measures the execution of action and the inhibition of that action
when given a stop signal. Recent cross-sectional studies found that children with higher
concurrent blood Pb levels had increased responses with stop signals. Among children
ages 8-12 years in Romania, a 1 (ig/dL increase in concurrent blood Pb level was
associated with a 8.9% (95% CI: -1.3, 19.3) increased false-alarm rate in responses to
stop signals with adjustment for city, sex, age, computer experience, handedness, eye
problems, number of siblings, parental education, prenatal alcohol and smoking exposure,
and parental report of parental psychopathology (Nicolescu et al.. 2010). It is uncertain
how history of parental psychopathology may be related to current caregiving quality.
Children in one town lived near a metal processing plant; however, blood Pb levels of Al
and Hg (other neurotoxic metals), were not associated with the stop signal task. In a case-
control study of children with ADHD, Nigg et al. (2008) found that higher concurrent
blood Pb level was associated poorer response inhibition on a stop task, which in turn
was associated with higher hyperactivity/impulsivity ratings. Path analysis showed that
the association between blood Pb level and hyperactivity/impulsivity ratings was
mediated by poorer performance on the stop task.
In addition to objective tests of attention, studies found Pb-associated increases in
inattention as rated by teachers, parents, and independent examiners (Figure 5-10 and
Table 5-11). Results from prospective studies indicated associations in children ages 7-13
years in Australia, New Zealand, and Boston, MA (Burns etal., 1999; Fergusson et al..
1993; Leviton et al.. 1993) but less so in younger children ages 4-5 years in Rochester
November 2012
5-148
Draft - Do Not Cite or Quote
-------
1 and Yugoslavia (Canfield et al.. 2003b; Wasserman et al.. 2001). These studies had
2 population-based recruitment and moderate to high follow-up, without indication of
3 biased participation by children with higher blood Pb levels and attention problems.
4 Among older children, inattention was associated with higher lifetime (to age 11-13
5 years) average (Burns etal.. 1999). and tooth (collected at ages 6-8 years) Pb levels
6 (Fergusson et al.. 1993; Leviton et al.. 1993). In separate cohorts, lifetime average blood
7 Pb level (Burns et al.. 1999) and tooth Pb level (Leviton et al.. 1993) were associated
8 with higher inattention ratings among girls than boys. With the exception of the Boston,
9 MA analysis, consideration of potential confounding did not differ widely among the
10 prospective studies, with most adjusting for maternal education, SES, and parental
11 caregiving quality (e.g. HOME) (Table 5-11). Prospective studies that found associations
12 examined higher blood Pb levels. The mean lifetime (to age 11-13 years) average blood
13 Pb level in the Port Pirie, Australia cohort was ~14 (ig/dL (Burns et al.. 1999).
14 Chandramouli et al. (2009) did not find an association between higher age 30 month
15 blood Pb levels and higher ratings of inattention in U.K. children ages 7-8 years.
16 Associations with inattention were not found in the Rochester and Yugoslavia cohorts
17 with lower concurrent blood Pb levels, means 6-7 (ig/dL (Canfield et al.. 2003b:
18 Wasserman et al., 2001). The studies in the Rochester and Yugoslavia cohorts examined
19 lower blood Pb levels but also had smaller sample sizes and examined younger children
20 ages 4-5 years, in whom patterns of behavior are less well established and in whom
21 inattention ratings may be less reliably measured. Nonetheless, these few weak findings
22 do not mitigate the otherwise compelling evidence, including that for attention-related
23 behavioral problems assessed with neuropsychological tests and that observed in animals.
24 Canfield et al. (2003b) found associations between higher concurrent blood Pb level and
25 higher ratings of inattention, but they were attenuated with adjustment for child color and
26 shape knowledge and FSIQ, which suggested that poorer knowledge of the task
27 parameters may increase distraction. However, other studies found associations between
28 blood Pb level and inattention with adjustment for child IQ (Cho etal.. 2010; Nigg et al..
29 2008). indicating the relationship between inattention and cognitive function may vary
30 across populations.
31 Several cross-sectional studies found associations between higher concurrent blood Pb
32 level and inattention among children, with several studies examining older children, 8-17
33 years (Nicolescu et al.. 2010; Nigg et al.. 2008; Silvaetal.. 1988). These studies had
34 population-based recruitment but did not provide sufficient information to assess
35 potential selection bias. While most of these studies examined potential confounding by
36 parental education or cognition, and parental history of psychopathology, none
37 considered parental caregiving quality. Whereas a previous study in New Zealand
38 examined children with relatively high blood Pb levels (mean 11 (ig/dL) (Silva et al..
39 1988). some recent studies provided evidence of association between blood Pb level and
November 2012 5-149 Draft - Do Not Cite or Quote
-------
1 higher ratings of inattention in populations with relatively low concurrent blood Pb levels
2 (means: 1, 3.7 (ig/dL) (Nicolescu et al.. 2010; Nigg et al.. 2008). However, contributions
3 from higher past Pb exposures cannot be excluded. Past Pb exposures especially may
4 have an influence in the study of children ages 8-12 years in Romania, 55% (n = 46/83)
5 of whom lived near a former metal processing plant and had higher concurrent blood Pb
6 levels (mean 5.1 versus 3.2 (ig/dL) (Nicolescu et al.. 2010). Among all subjects, a
7 1 (ig/dL increase in concurrent blood Pb level was associated with a 4.5% higher (95%
8 CI: -1.3, 10.3%) teacher rating of inattention and a weaker, imprecise 1.3% (95% CI:
9 -3.3,5.9) higher parent rating. Blood Pb levels were not correlated with blood levels of
10 Al or Hg, and neither of these other metals were associated with inattention ratings. The
11 association did not change substantially in an analysis that excluded the 5 children with
12 blood Pb levels > 10 (ig/dL. Adjustment for city, sex, age, computer experience,
13 handedness, eye problems, number of siblings, parental education, prenatal smoking
14 exposure, prenatal alcohol exposure, and parental history of psychological or psychiatric
15 problems resulted in less precise blood Pb level effect estimates, although investigators
16 did not report the magnitude of change in the effect estimate.
17 Consistent with Nicolescu et al. (2010). Nigg et al. (2008) found associations of
18 concurrent blood Pb levels with parent and teacher ratings of a composite
19 hyperactivity/impulsivity index in a group of children (location not reported) with and
20 without ADHD (ages 8-17 years) with a mean blood Pb level of ~1 (ig/dL. The case-
21 control design of the study could have resulted in biased participation of ADHD children
22 with higher blood Pb levels. Nigg et al. (2008) also found the Pb-associated increase in
23 hyperactivity/impulsivity to be independent of the association with IQ using regression-
24 based path analysis, a more rigorous method to characterize the impact of one variable on
25 the association of another in the model after controlling for other previous variables. With
26 adjustment for sex and income, concurrent blood Pb level was directly associated with
27 hyperactivity/impulsivity, and the association was not completely mediated by the blood
28 Pb-IQ association. Instead, the association between blood Pb level and IQ was found to
29 be mediated by the association with hyperactivity/impulsivity. Other potential
30 confounders including parental IQ and caregiving quality were not examined. Other
31 recent cross-sectional studies of older children found associations between higher
32 concurrent blood or hair Pb level and higher ratings of inattention (independent examiner
33 or parent) in children in Mexico and China living near Pb sources (Bao et al., 2009;
34 Kordas et al.. 2007). As in most other cross-sectional studies, these results were adjusted
35 for SES and parental education.
36 Cross-sectional studies that included younger children (ages 3-5 years) also found
37 associations between concurrent blood Pb level and higher inattention as rated by
38 teachers or study examiners (Plusquellec et al., 2010; Roy et al.. 2009a). In these younger
November 2012 5-150 Draft - Do Not Cite or Quote
-------
1 children, attention-related behaviors may be less reliably measured and may not predict
2 later childhood behavior. A study conducted in Inuit children, ages 5-6 years, living in
3 Quebec, Canada reported associations between blood Pb levels and measures of
4 inattention with consideration of several potential confounding factors (Plusquellec et al.
5 2010). Concurrent blood Pb level but not cord blood Pb level was associated with
6 impulsivity and duration of off task behavior as rated by study examiners (Plusquellec et
7 al., 2010). Fraser et al. (2006) additionally indicated that at ages 5-6 years, the
8 relationship between concurrent blood Pb level and motor function (i.e., transversal sway,
9 reaction time) may be mediated by the association between blood Pb level and
10 inattention/impulsivity. The various associations were adjusted for different factors but
11 included SES, caregiver education, birth weight, and blood hemoglobin. HOME score
12 and micronutrient levels were not associated with inattention or impulsivity and thus
13 were not included in models. In this population that has high consumption offish, blood
14 levels of poly chlorinated biphenyls and Hg were not associated with inattention or
15 impulsivity ratings. Other recent cross-sectional studies that included younger children
16 (ages 3-7 years) found associations between higher concurrent blood Pb level and higher
17 parent or teacher ratings of inattention in populations with higher blood Pb levels (mean
18 11.4 (ig/dL or median 13.2 (ig/dL) (Liu et al.. 20 lib: Roy et al.. 2009a). While these
19 studies adjusted for or considered potential confounding by SES, child age, and parental
20 education, they did not examine potential confounding by parental caregiving quality.
lexicological Studies of Inattention and Impulsivity
21 The associations described in the preceding section between blood Pb level and
22 inattention and impulsivity in children are supported by findings in animals for
23 Pb-induced impaired ability to inhibit inappropriate responses and increased
24 perseveration. In animals, tests of response inhibition include Signal Detection with
25 Distraction, Differential Reinforcement of Low Rates of Responding (DRL), Fixed
26 Interval (FI) testing, FI with Extinction, or Fixed Ratio (FR)/waiting-for-reward, with
27 impulsivity indicated by premature responses, decreased pause time between two
28 scheduled events, and increased perse veration. Some of these tests also have been used to
29 assess learning (Section 5.3.2.3). and the interactions observed between Pb exposure and
30 maternal or offspring stress also may apply to effects on impulsivity. Multiple earlier
31 studies and those included in the 2006 Pb AQCD showed that early life Pb exposure
32 impaired response inhibition as assessed with these aforementioned tests, and recent
33 studies provide supporting evidence. Discrimination reversal, which also measures
34 response inhibition by rewarding the withholding of responses, has been shown to be
35 affected by Pb exposure. Spatial and non-spatial discrimination reversal (i.e., reversal of a
36 previously learned habit) was significantly affected after developmental Pb exposure and
November 2012 5-151 Draft - Do Not Cite or Quote
-------
1 was exacerbated with distracting stimuli. The collective evidence indicates that
2 impulsivity in rodents and nonhuman primates is significantly affected by Pb exposure
3 that results in blood Pb levels in the range relevant to humans, i.e., 11-31 (ig/dL.
4 Toxicological studies provide more consistent evidence for the effects of Pb exposure on
5 impulsivity in animals than on sustained attention. The 2006 Pb AQCD (U.S. EPA.
6 2006b) reported inconsistent findings for the effects of Pb exposure on sustained attention
7 deficits in animals as assessed by a signal detection test with distracting stimuli, a test
8 recording omissions after exposure to an external distraction. In this test, animals earn
9 food rewards by discriminating correctly between a target and distracter light.
10 Postweaning Pb exposure that produced blood Pb levels of <5, 16, or 28 ug/dL did not
11 affect performance in the signal detection test with distracting stimuli in adult rats
12 (Brockel and Cory-Slechta. 1999a). A similar lack of effect was reported in a recent study
13 of female rats exposed to 20 or 300 ppm Pb acetate in drinking water during lactation
14 (PND1-PND30) with resultant blood Pb levels on PND52 of 13 or 31 (ig/dL, respectively
15 (Stangle et al., 2007). However, in this study, Pb exposure induced impulsivity as
16 indicated by premature responses in a discrimination reversal task. Impulsivity was not
17 improved with the chelator succimer, indicating persistence of effects. Impulsivity was
18 found in monkeys exposed to Pb from birth to time of testing at age 3-4 years with blood
19 Pb levels 15 and 25 (ig/dL; however, effects were reversible, as Pb-exposed monkeys did
20 not improve performance as quickly but eventually acquired reinforcement rates equal to
21 that in controls (Rice and Gilbert. 1985). Previous evidence indicated that Pb exposure of
22 laboratory animals induces distractibility. Spatial and non-spatial discrimination reversal
23 was significantly affected after lifetime Pb exposure in monkeys ages 9-10 years (blood
24 Pb levels 15, 25 (ig/dL) and was exacerbated with distracting stimuli. Repeated learning
25 testing revealed that these deficits likely were not due to sensory or motor impairment
26 (Gilbert and Rice. 1987).
27 The effects of Pb exposure on inattention and impulsivity in the 1986 Pb AQCD were
28 indicated by aberrant performance on operant conditioning tasks in rodents and
29 non-human primates (U.S. EPA. 1986b). The 2006 Pb AQCD reported consistent
30 findings for Pb exposure (producing blood Pb levels: 58-94 (ig/dL) affecting FI response
31 rates, by means of decreased interresponse times. Some studies indicated decreased
32 interresponse times in animals with blood Pb levels 11-15 (ig/dL (U.S. EPA, 2006b). The
33 effects of Pb exposure on impulsivity also have been demonstrated as shorter time
34 Pb-exposed animals will wait for reward in FR/waiting for reward testing. In this test,
35 animals can obtain food by pressing a lever for a set number of times. Free food is
36 delivered with increasingly long time intervals so long as animals inhibit additional lever
37 presses. Animals can reset the schedule to return to the FR component at any time.
38 Brockel and Cory-Slechta (1998) exposed male Long-Evans rats to 0, 50, or 150 ppm
November 2012 5-152 Draft - Do Not Cite or Quote
-------
1 Pb acetate in drinking water from weaning, which produced respective blood Pb levels of
2 <5, 11, and 29 ug/dL after 3 months of exposure. After 40 days of exposure, the 150 ppm
3 Pb-exposed rats responded more quickly in the FR component and reset the schedule
4 (thus shortening the waiting period) more frequently than did the 50 ppm Pb-exposed rats
5 and controls. In the waiting component, wait time was significantly lower in both Pb
6 exposure groups compared to controls. The behavior of the 150 ppm Pb-exposed rats
7 suggested a low tolerance for waiting, but 150 ppm Pb exposure also yielded more
8 reinforcers per session and a higher response-to reinforcement-ratio than achieved by the
9 50 ppm Pb group and controls. Mechanistic understanding of the aforementioned
10 Pb-induced impulsivity was provided by a study with similar postweaning dosing of 0,
11 50, and 150 ppm Pb that yielded respective blood Pb levels of <5, 10, and 26 ug/dL after
12 3 and 7 months of exposure. Administration of a D2 receptor agonist reversed the
13 Pb-induced parameters assessed by FR schedule testing, suggesting a role for dopamine-
14 like receptors in Pb-induced impulsivity (Brockel and Cory-Slechta. 1999b).
15 Pb-induced impulsivity appears to be related to emotionality, which was found in
16 Pb-exposed rats trained to perform an olfactory discrimination task, albeit at higher Pb
17 exposures than those relevant to humans. In this study, rats were given early postnatal Pb
18 exposure (300 ppm Pb acetate via dam drinking water PND1-PND17 then either 20 or
19 300 ppm PND18-PND30 in their own drinking water) which produced blood Pb levels of
20 40-60 and 100-140 (ig/dL). The offspring were tested as young adults on a food-
21 motivated olfactory discrimination task in which rewards for correct responses were
22 occasionally and unpredictably omitted. Pb-exposed animals were more sensitive both to
23 their own errors and to reward omission than controls, suggesting a lowered capacity for
24 regulating arousal and emotion. Administration of succimer, a chelating agent, after the
25 Pb exposure period (PND31-PND52) normalized reactivity to reward omission and errors
26 in the Pb-treated rats, but increased the reactivity in the control animals (Beaudin et al..
27 2007). Similar observations were made by the same laboratory for heightened reactivity
28 to errors in tests of visual discrimination and visual sustained attention in rats exposed to
29 20 or 300 ppm Pb acetate in drinking water PND1-PND30 (Stangle et al.. 2007)
30 In animals, Pb-induced increases in inattention have been indicated using tests that are
31 not direct assessments of inattention but that examine behaviors linked to inattention. For
32 example, a study reported that impaired performance on auditory threshold tasks in
33 Pb-exposed monkeys was likely due to inattention (Laughlin et al.. 2009). Rhesus
34 monkeys were exposed to Pb acetate from gestation (drinking water of mothers, 3 months
35 prior to mating) to birth or postnatally from birth to age 5.5 months at weaning and had
36 resultant bone Pb levels at 11 years of 7 and 13 (ig/g for prenatal and postnatal groups,
37 respectively, and blood Pb levels during Pb exposure of 35 and 46 (ig/dL, respectively.
38 Animals were tested at age 13 years when blood Pb levels had returned to baseline levels.
November 2012 5-153 Draft - Do Not Cite or Quote
-------
1 The inability of some of the monkeys to engage or focus attention on the task at hand
2 yielded fewer available measurements in Pb-exposed animals versus control rhesus
3 monkeys. These observations were made in monkeys with higher peak blood Pb levels
4 than those relevant to humans.
5 In summary, several studies in animals indicate that early developmental Pb exposure of
6 rodents and non-human primates, some producing blood Pb levels relevant to humans,
7 increases impulsivity as indicated by impaired response inhibition. Evidence for
8 Pb-induced decrements in sustained attention is less consistent. The observations for
9 Pb-induced increases in impulsivity in animals provide support for associations observed
10 in children between blood and tooth Pb levels and impaired response inhibition and also
11 higher ratings of inattention and impulsivity.
5.3.3.2 Hyperactivity
12 Studies reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b) indicated associations
13 between higher concurrent blood Pb level or tooth Pb level and higher parent and teacher
14 ratings of hyperactivity in children ages 6-11 years in the U.S. and Asia (Rabinowitz et
15 al.. 1992: Silvaetal.. 1988: Gittleman and Eskenazi. 1983: Needleman et al.. 1979:
16 David et al.. 1976). The case-control or cross-sectional design of studies limited
17 understanding of the temporal sequence between Pb exposure and hyperactivity. Several
18 recent studies, including a prospective study (Chandramouli et al., 2009). also found
19 associations between blood Pb level and hyperactivity as rated by teachers and parents
20 (Figure 5-10 and Table 5-11). Overall studies indicated associations with mean or group
21 blood Pb levels >10 (ig/dL.
22 The recent prospective study of children in the U.K. addressed some of the limitations of
23 previous cross-sectional studies by demonstrating an association between higher earlier
24 childhood (age 30 months) blood Pb level and higher teacher ratings of hyperactivity
25 later in childhood at age 7-8 years with adjustment for several potential confounding
26 factors, including home facilities score and family adversity index (Chandramouli et al..
27 2009). In addition to the prospective design, the study had a high participation rate at
28 baseline and follow-up from a population with similar characteristics as reported in the
29 U.K. census. Increases in hyperactivity were found primarily in the group of children
30 with blood Pb levels >10 (ig/dL and were independent of associations with IQ.
31 Among cross-sectional studies, adjustment for SES, maternal education and IQ was
32 common; however, few adjusted for parental caregiving quality. Silva et al. (1988) and
33 Nicolescu et al. (2010) respectively, adjusted for current and history of maternal
34 psychopathology, whose relationships with parental caregiving quality are not well
November 2012 5-154 Draft - Do Not Cite or Quote
-------
1 characterized. Both studies found associations of concurrent blood Pb level with
2 hyperactivity as rated by teachers and parents, and Silva et al. (1988) found the
3 association in children age 11 years in New Zealand to persist with adjustment for child
4 IQ. The group of children in New Zealand (Silva et al.. 1988) had a higher mean
5 concurrent blood Pb level than the group in Romania (Nicolescu et al.. 2010) (11.1 versus
6 3.7 (ig/dL). Other cross-sectional studies found associations with hyperactivity in
7 younger children, in whom behavior may be rated less reliably. Plusquellec et al. (2010)
8 found an association in children ages 5-6 years with relatively low concurrent blood Pb
9 levels, mean 5.4 (ig/dL, and found that HOME score and caretaker distress were not
10 associated with hyperactivity. Roy et al. (2009a) found a Pb-associated increase in
11 hyperactivity in children ages 3-7 years in India with a mean concurrent blood Pb level of
12 11.4(ig/dL.
13 Pb also has been associated with hyperactivity in animals, but the relevance to
14 observations in children is not clear. In a recent study, Pb exposure from gestation to the
15 early postnatal period (PND10) (low and high dose Pb: 10 and 42 (ig/dL blood Pb level at
16 PND10, respectively) increased activity of male mice at age 1 year with co-treatment
17 with amphetamines but not female mice (Leasure et al.. 2008). Without amphetamines,
18 Pb induced less activity of mice, and the low Pb dose inhibited activity more than the
19 high Pb dose did. In addition to the effects of Pb on impulsivity, Stangle et al. (2007)
20 found Pb-induced decreases in arousal. In one theory of ADHD, low arousal levels
21 contribute to excessive self-stimulation or hyperactivity such that an optimal level of
22 arousal can be attained (Swanson et al.. 2011).
5.3.3.3 Ratings of Attention Deficit Hyperactivity Disorder-related Behaviors
23 The 2006 Pb AQCD (U.S. EPA. 2006b) did not examine ADHD specifically. However,
24 in addition to finding associations with inattention, impulsivity, and hyperactivity, some
25 of the recent epidemiologic studies described in the preceding sections found associations
26 between higher concurrent blood Pb level and higher parental and teacher ratings of
27 ADHD-related behaviors (Cho et al.. 2010: Nicolescu etal.. 2010: Roy et al.. 2009a).
28 which are a composite of the various behaviors that are evaluated in the diagnosis of
29 ADHD. The strengths and limitations of these studies have been described in the
30 preceding sections. Main limitations were the cross-sectional design, lack of
31 consideration for potential confounding by parental caregiving quality, and lack of
32 validation of ADHD ratings with a clinical diagnosis. Thus, the evidence specifically for
33 these total ADHD index ratings were emphasized less than evidence for inattention and
34 impulsivity in drawing conclusions about the effects of Pb exposure on attention-related
35 behavioral problems.
November 2012 5-155 Draft - Do Not Cite or Quote
-------
1 The large, U.S. representative analysis of children participating in NHANES 2001-2004
2 found an association between concurrent blood Pb level in children ages 8-15 years and
3 parental assessment of child ADHD-related behaviors using the Diagnostic Interview
4 Schedule for Children which uses DSM-IV criteria to identify children at increased risk
5 of meeting diagnostic criteria for ADHD (Froehlich et al.. 2009). Compared with children
6 with concurrent blood Pb levels <0.8 (ig/dL, children with concurrent blood Pb levels
7 >1.3 (ig/dL had elevated odds of parentally-rated ADHD-related behaviors with an OR of
8 2.3 (95% CI: 1.5, 3.8). These results were adjusted for current household smoking
9 exposure, sex, age, race/ethnicity, income, preschool attendance, maternal age, and birth
10 weight. A similar OR was estimated when children with concurrent blood Pb levels
11 >5.0 (ig/dL were excluded from the highest tertile. The strongest association was
12 observed in children with both high blood Pb level and prenatal tobacco smoke exposure.
13 Compared to children with blood Pb levels <0.8 (ig/dL with no exposure to prenatal
14 tobacco smoke, children with blood Pb levels >1.3 (ig/dL and exposure to prenatal
15 tobacco smoking had the highest odds of parentally-rated ADHD-related behavior (OR:
16 8.1 [95% CI: 3.5, 18.7]). Although ADHD-related behavior was associated with low
17 concurrent blood Pb levels (1.3-5 (ig/dL), the contribution of higher past Pb exposures of
18 adolescents born in the late 1980s cannot be excluded. Roy et al. (2009a) also found an
19 association with teacher ratings of ADHD-related behaviors using DSM criteria in
20 children in Chennai, India; however, the study population included some very young
21 children (i.e., age 3 years) and had relatively high blood Pb levels (mean: 11.4 (ig/dL).
22 Other recent cross-sectional studies found Pb-associated higher ratings of ADHD-related
23 behaviors using instruments that do not follow DSM criteria. Among children ages
24 8-11 years in Korea, Cho et al. (2010) found a stronger relationship with a total ADHD-
25 related behaviors index as rated by teachers than parents. Mean ADHD ratings by teacher
26 and parents were similar (both 9.1); however, parental ratings had greater variability (SD:
27 11.5 for parents and 8.6 for teachers), which may have contributed to differences in
28 association. Among children in Romania, concurrent blood Pb level was associated
29 similarly with parent and teacher ratings of ADHD-related behaviors (Nicolescu et al..
30 2010). As with individual attention-related behaviors described in preceding sections,
31 blood Al and Hg levels were not associated with ratings of ADHD-related behaviors.
32 Based on a log-linear model, a 1 (ig/dL increase in concurrent blood Pb level within the
33 10th-90th percentile interval (1.8-7.1 (ig/dL) was associated with a 4% increase (95% CI:
34 0, 10) in rating of ADHD-related behavior. The association did not change substantially
35 in an analysis that excluded the 5 children with blood Pb levels > 10 (ig/dL. Both studies
36 considered potential confounding by parental history of psychopathology. In Cho et al.
37 (2010). children of parents with history of psychiatric disease had lower blood Pb levels.
38 In Nicolescu et al. (2010). parental history of psychological or psychiatric problems was
39 weakly correlated with parental (r = 0.24, p < 0.05) and teacher (p = 0.12) rating of
November 2012 5-156 Draft - Do Not Cite or Quote
-------
1 ADHD-related behavior, and ORs were fairly similar for ADHD score rated by teachers
2 and parents. Although parental history of psychopathology was examined in a few
3 studies, its relationship with current parental caregiving quality is not well characterized.
5.3.3.4 Attention Deficit Hyperactivity Disorder in Children
4 The 2006 Pb AQCD (U.S. EPA, 2006b) did not review studies of prevalence or incidence
5 of ADHD diagnosis but noted lack of conclusive evidence for the effect of Pb exposure
6 on ADHD based on a few small studies comparing blood Pb levels between children with
7 and without hyperactivity as identified by parents, teachers, or schools (Gittleman and
8 Eskenazi. 1983; David et al., 1972). As described in the previous section, several recent
9 cross-sectional studies found associations of concurrent blood Pb level with parent and
10 teacher ratings of a total ADHD index, a composite index of inattention, impulsivity, and
11 hyperactivity. Results from a small body of recent studies also indicate associations of
12 higher concurrent blood Pb level with prevalence of diagnosed ADHD in children ages
13 4-17 years (Nigg et al.. 2010: 2008: Wang et al. 2008d: Braun et al.. 2006). All of the
14 studies were cross-sectional; thus, the temporal sequence between Pb exposure and
15 ADHD incidence cannot be established. While there is coherence with evidence from
16 prospective studies in other populations for associations of blood Pb levels with
17 inattention, hyperactivity, and impulsivity, evidence specifically for ADHD prevalence
18 was emphasized less than evidence for inattention and impulsivity in drawing
19 conclusions about the effects of Pb exposure on attention-related behavioral problems.
20 Associations between concurrent blood Pb level and ADHD prevalence were found in
21 case-control studies conducted in different populations of children. While a potential
22 limitation of these studies is selection bias arising from the nonrandom population
23 sample, a common strength is their independent diagnosis of ADHD in a structured
24 manner using parental and teacher ratings of behavior followed by independent
25 assessment by multiple clinicians using DSM-IV criteria (Nigg et al.. 2010; 2008; Wang
26 et al., 2008d). Nigg et al. (2010; 2008) found an association between concurrent blood Pb
27 level and ADHD diagnosis in relatively small (n = 150, 236) groups of children ages
28 6-17 years from the same community, with controls selected from healthy children who
29 responded to community advertisements. Wang et al. (2008d_) found an association in a
30 larger (n = 1,260) group of children in China, with controls selected from children
31 attending the same pediatric clinic for respiratory infections.
32 Braun et al. (2006) found an association in children ages 4-15 years participating in
33 NHANES 1999-2002. ADHD was ascertained by parent-report of ADHD diagnosis or
34 use of stimulant medication, which is subject to reporting bias; however, the examination
November 2012 5-157 Draft - Do Not Cite or Quote
-------
1 of multiple risk factors and outcomes in NHANES reduces the likelihood of biased
2 participation and reporting of ADHD by parents of children with higher Pb exposure.
3 NHANES is not a random sample, but a strength over other studies that examined the
4 prevalence of ADHD diagnosis is the large (n = 4,704) sample size and the nationally-
5 representative results produced with adjustment for sampling weights in models.
6 Surveillance data indicate that states with a higher percentage of children with blood Pb
7 levels > 10 (ig/dL have lower prevalence of diagnosed ADHD (CDC. 2012, 201 Ib).
8 These data reduce the potential for confounding of associations observed in the NHANES
9 population by regional differences in blood Pb levels and ADHD prevalence.
10 With respect to blood Pb levels associated with ADHD diagnosis, analyses of the
11 concentration-response relationship indicated monotonic increases in ORs across blood
12 Pb level groups (Wang et al.. 2008d: Braun et al.. 2006). In the analysis of children in
13 NHANES, compared to children with concurrent blood Pb level <0.8 (ig/dL, children
14 with concurrent blood Pb level >2.0 (ig/dL (maximum not reported) had higher
15 prevalence of ADHD with an OR of 4.1 (95% CI: 1.2, 14.0). A similar OR was estimated
16 for children with blood Pb levels 2.0-5.0 (ig/dL (Braun et al.. 2006V In the study of
17 children in China, the highest OR was found in children with concurrent blood Pb levels
18 > 10 (ig/dL but also was elevated in the group with blood Pb levels 5-10 (ig/dL (OR: 4.92
19 [95% CI: 3.47, 6.98] compared with children with blood Pb level <5 (ig/dL) (Wang et al..
20 2008d). Other evidence indicated associations at lower blood Pb levels, i.e., population
21 means ~1 (ig/dL or group with levels >0.8 (ig/dL (Nigg etal.. 2010; 2008; Braun et al..
22 2006). However, the examination of adolescents adds uncertainty regarding the relative
23 contributions of higher past Pb exposures and current exposures to the observed
24 associations. Blood Pb levels are higher in early childhood, and among children
25 participating in NHANES who were born 1984-1998, some likely had higher early-life
26 Pb exposures from the use of leaded gasoline in the U.S. (Braun et al.. 2006).
27 Consideration for potential confounding varied among studies. In three-way analyses of
28 covariance, Nigg et al. (2008) adjusted for sex and household income, and Nigg et al.
29 (2010) adjusted for maternal IQ and prenatal smoking exposure. However, in preliminary
30 analyses, Nigg et al. (2010) considered blood hemoglobin, household income, age, sex,
31 and race/ethnicity as potential confounding factors. The analysis of children participating
32 in NHANES adjusted for age, race, prenatal smoking exposure, postnatal smoker in the
33 home, preschool/child care attendance, health insurance coverage, and ferritin levels but
34 initially considered poverty to income ratio, birth weight, and admission to the neonatal
35 intensive care unit (Braun et al.. 2006). The results for children in China were adjusted
36 for similar covariates and also family (parent and sibling) history of ADHD diagnosis,
37 ascertained from clinical records (Wang et al.. 2008d). Family history of ADHD was
38 selected as a covariate based on its association with child ADHD; no information was
November 2012 5-158 Draft - Do Not Cite or Quote
-------
1 provided on its association with child blood Pb level. None of the studies of ADHD
2 prevalence considered potential confounding by current parental caregiving quality.
3 In recent commentaries to studies reporting associations between blood Pb level and
4 ADHD in children, Brondum (2011, 2007) asserted the need for studies to consider
5 confounding by parental history of ADHD. Given the highly heritable nature of ADHD,
6 parental ADHD is a strong risk factor for ADHD in children (Faraone and Doyle. 2001);
7 however, data have not characterized well associations of parental history of ADHD and
8 blood Pb level in the child. Therefore, it is uncertain whether the lack of adjustment for
9 parental history of ADHD produces spurious associations between blood Pb level and
10 ADHD in children. Further, because parental history of ADHD likely explains a large
11 portion of variance in child ADHD, not removing that variance with statistical adjustment
12 may mask the smaller magnitude of risk due to other factors, including Pb, not produce
13 spurious associations. Studies that examined parenting behaviors in parents with current
14 ADHD have indicated that parents with ADHD show negative parenting control,
15 i.e., over-reactive disciplining, lack of planning, and disorganization but have not
16 consistently indicated that parents with ADHD have poorer emotional responsiveness,
17 i.e., involvement with the child (as reviewed in Johnston et al.. 2012). Thus, the potential
18 for parental ADHD to produce spurious associations between child blood Pb level and
19 child ADHD is not well characterized.
5.3.3.5 Integrated Summary of Attention-related Behavioral Problems
20 Although not examined as extensively as cognitive function, epidemiologic studies have
21 found associations of childhood blood and tooth Pb levels with attention-related
22 behavioral problems in children and young adults, with more compelling evidence for
23 increases in inattention and impulsivity than hyperactivity, ratings of ADHD-related
24 behaviors, or ADHD diagnosis. The evidence for inattention and impulsivity is provided
25 by both prospective and cross-sectional studies, whereas evidence for hyperactivity,
26 ratings of ADHD-related behaviors, and ADHD diagnosis is provided primarily by cross-
27 sectional studies. With analysis of earlier childhood blood or tooth Pb levels and later
28 childhood behavioral problems, the prospective studies better characterized the temporal
29 sequence between exposure and outcome. In cross-sectional associations with concurrent
30 blood Pb levels, there is greater uncertainty regarding the potential for reverse causation.
31 Associations between blood or tooth Pb levels and attention-related behavioral problems
32 were found in diverse populations in North America, Europe, Asia, Australia, and New
33 Zealand. Most studies had population-based recruitment from prenatal clinics, hospitals
34 at birth, or schools and had moderate to high participation. A few prospective studies had
35 increased loss-to-follow-up in certain groups, for example, lower SES, lower earlier
November 2012 5-159 Draft - Do Not Cite or Quote
-------
1 FSIQ, lower HOME score. This potential selection bias can reduce the generalizability of
2 findings to the original study population, but there was not strong indication that
3 participation was biased to those with higher blood Pb levels and greater attention-related
4 behavioral problems. Multiple testing was common; however, in most studies, the
5 consistent pattern of association observed across the ages of blood Pb level and/or
6 behavior examined increases confidence that the evidence is not unduly biased by the
7 increased probability of finding associations by chance alone.
8 A large evidence base indicates associations of blood and tooth Pb levels with inattention
9 and impulsivity as assessed using neuropsychological tests or ratings by parents or
10 teachers (Section 5.3.3.1). Observations of associations across the various methods of
11 assessment increase confidence that the collective evidence is not unduly influenced by
12 biased reporting of inattention by parents of children with higher blood Pb levels. Most
13 studies that examined inattention with the CPT found associations with blood Pb level
14 (Figure 5-10 and Table 5-11). including prospective studies, which indicated increases in
15 commission and omission errors or reaction time in association with higher prenatal
16 (maternal) and earlier childhood (age 3-60 month average, age 78 month) blood Pb levels
17 in the Cincinnati cohort at ages 15-17 years (Ris et al., 2004) and with higher tooth Pb
18 (from ages 5-8 years) levels in Boston-area young adults at ages 19-20 years (Bellinger et
19 al., 1994a). Results from prospective studies also indicated higher parental and teacher
20 ratings of inattention in association with higher lifetime average blood Pb levels in
21 children ages 11-3 years in Port Pirie, Australia (Burns et al., 1999) and with tooth Pb
22 levels (from ages 6-8 years) in children ages 8-13 years in New Zealand, and Boston, MA
23 (Fergusson et al., 1993; Leviton et al., 1993). The mean blood Pb levels (prenatal cord,
24 early childhood, lifetime average) in these populations were 7-14 (ig/dL. In children,
25 inattention was associated with biomarkers of Pb exposure representing several different
26 lifestages and time periods. Prospective studies did not examined a detailed blood Pb
27 history, and results do not identify an individual critical lifestage, time period, or duration
28 of Pb exposure associated with inattention in children. Associations in prospective studies
29 with tooth Pb level, earlier childhood average and lifetime average blood Pb levels point
30 to an effect on inattention of cumulative childhood Pb exposure. Indicators of more
31 recent Pb exposures were not examined. Evidence did not strongly indicate associations
32 between concurrent blood Pb levels and inattention ratings in the Rochester and
33 Yugoslavia cohorts (Canfield et al., 2003b; Wasserman et al., 2001). This latter group of
34 studies examined lower blood Pb levels, means 6.5 (ig/dL, but younger children ages 4-5
35 years, in whom behaviors may be less reliably measured.
36 An additional strength of the prospective studies was their more extensive consideration
37 for potential confounding. Although the specific factors varied by study, prospective
38 studies of inattention adjusted for factors such as SES, parental IQ, maternal education,
November 2012 5-160 Draft - Do Not Cite or Quote
-------
1 HOME score, self drug use, prenatal drug and alcohol exposure, and birth outcomes.
2 Adjustment for SES is difficult as it is highly correlated with Pb exposure and there is no
3 single measure that represents SES. Residual confounding also is likely by factors not
4 considered. The combination of evidence from prospective studies that considered
5 several well-characterized potential confounding factors plus coherence with evidence
6 that Pb exposure induces impulsivity in animals increase confidence that the associations
7 between blood and tooth Pb levels and inattention and impulsivity observed in children
8 represent a relationship with Pb exposure.
9 Several recent cross-sectional studies provided evidence of associations of higher
10 concurrent blood Pb level with increases in impulsivity using response inhibition tests
11 and higher inattention ratings in children ages 8-17 years. These associations were found
12 in populations with mean concurrent blood Pb levels 1 and 4 (ig/dL (Nicolescu et al..
13 2010; Nigg etal.. 2008). However, the contribution of higher Pb exposures earlier in
14 childhood cannot be excluded. Further, while these recent studies considered potential
15 confounding by parental education, they had less consistent consideration for other SES-
16 related factors or parental caregiving quality than prospective studies. Some considered
17 parental history of psychopathology (Cho etal.. 2010; Nicolescu et al.. 2010); however,
18 its relationship with parental caregiving quality is not well characterized. Recent cross-
19 sectional studies that included younger children (ages 3-5 years) also found associations
20 between concurrent blood Pb level and higher inattention as rated by teachers or study
21 examiners (Plusquellec et al., 2010; Roy et al., 2009a); however, ratings in young
22 children may be less reliably measured. With the exception of the Rochester cohort study
23 (Canfield et al., 2003b). several studies found associations between blood Pb level and
24 inattention with adjustment for child IQ (Cho et al.. 2010; Nigg et al.. 2008; Silva et al..
25 1988). supporting an effect of Pb exposure on inattention independent of effects on
26 cognitive function.
27 The epidemiologic findings for impulsivity are supported by observations in rats and
28 monkeys of Pb-induced impaired response inhibition in tests of discrimination reversal
29 learning and FR/waiting for reward (Stangle et al.. 2007; Brockel and Cory-Slechta.
30 1998; Rice and Gilbert. 1985). Coherence is found particularly with associations found
31 between blood Pb levels and impaired response inhibition in children as assessed using
32 the stop signal task. Impulsivity in animals was found with early postnatal (lactation) and
33 lifetime dietary Pb exposures relevant to humans, i.e., resulting in blood Pb levels
34 11-31 (ig/dL. The effects of early postnatal Pb exposure are consistent with the
35 continuing development of the nervous system and greater Pb absorption and retention
36 during early life. The findings in children and animals for Pb-associated impulsivity are
37 supported by observations that Pb affects dopaminergic neurons of the frontal cortex and
38 striatum of the brain by altering dopamine release and receptor density. The circuitry in
November 2012 5-161 Draft - Do Not Cite or Quote
-------
1 these regions is thought to mediate response inhibition. In animals, the effects of Pb on
2 sustained attention have been inconsistent; however, studies find Pb-induced increases in
3 distractibility. Attention-related behavioral problems also have been linked with changes
4 in the hippocampus, and evidence describing the effects of Pb on hippocampal functions
5 further supports the mode of action for Pb-associated increases in attention-related
6 behavioral problems.
7 Although examined in fewer studies, hyperactivity has been linked with higher blood and
8 tooth Pb levels in children (Section 5.3.3.2). Previous findings were limited to cross-
9 sectional and case-control studies. However, a recent prospective study found higher
10 teacher ratings of hyperactivity in children in the U.K. ages 7-8 years with age 30 month
11 blood Pb levels >10 (ig/dL, with adjustment for maternal education and smoking, SES,
12 home facilities score, family adversity index, plus other factors (Chandramouli et al..
13 2009). Among cross-sectional studies, associations were found with adjustment for SES
14 and maternal education; however, parental caregiving quality was examined infrequently.
15 Recent studies also found associations with higher parental ratings of a composite index
16 of ADHD-related behaviors, including a large NHANES analysis that used DSM-IV
17 criteria (Froehlich et al.. 2009). In the few available studies, concurrent blood Pb levels
18 were associated with prevalence of diagnosed ADHD in children (Section 5.3.3.4). There
19 is coherence with evidence from prospective studies for associations of blood and tooth
20 Pb levels with inattention, hyperactivity, and impulsivity, which comprise ADHD.
21 However, the small number of studies, their cross-sectional or case-control design, and
22 lack of consideration for potential confounding by parental caregiving quality preclude
23 conclusions regarding the relationship between Pb exposure and ADHD specifically.
5.3.4 Conduct Problems
5.3.4.1 Epidemiologic Studies of Conduct Problems in Children
24 The 2006 Pb AQCD (U.S. EPA. 2006b) described several prospective studies that
25 demonstrated associations of higher blood, tooth, and bone Pb levels with conduct
26 problems in children as rated by parents or teachers [(U.S. EPA. 2006b). and see Table
27 5-12 from this ISA]. A few previous studies found associations with criminal offenses in
28 adolescents or young adults. Supporting evidence from recent prospective studies
29 included follow-up of previous cohorts to older ages (Table 5-12). Recent cross-sectional
30 studies found associations between concurrent blood Pb level and ratings of misconduct,
31 but several had limitations aside from establishing temporality, including prenatal drug
32 and alcohol exposure, treatment with chelators earlier in childhood, and less extensive
November 2012 5-162 Draft - Do Not Cite or Quote
-------
1 consideration for potential confounding. Previous studies found associations with mean
2 blood Pb levels >10 (ig/dL. Recent evidence indicated associations with lower blood Pb
3 levels, means 1-8 (ig/dL. However, in these children and young adults, the influence of
4 higher Pb exposures earlier in childhood cannot be excluded. In the evaluation of
5 epidemiologic evidence for conduct problems, greater emphasis was placed on evidence
6 from prospective studies and studies with greater consideration for potential confounding.
November 2012 5-163 Draft - Do Not Cite or Quote
-------
Table 5-12 Associations between blood Pb level and misconduct in children and
young adults.
Study3
Prospective
Dietrich et al.
(2001 )
Burns et al.
(1999)
Chandramouli
et al. (2009)
Needleman et
al. (1996)
Study Population and
Methodological Details
Studies of Ratings of Misconduct
195 children followed from birth (1979-1985)
to age 15-17 yr, Cincinnati, OH
Recruitment at prenatal clinic. High follow-up
participation, no selective attrition. Primarily
African American. Linear regression model
adjusted for HOME score, parental IQ,
current SES, birth weight. Also considered
potential confounding by maternal age, other
birth outcomes, prenatal smoking, alcohol
use, and marijuana use, Fe status, ear
infections, sex, age, caregiver education,
public assistance, attendance at preschool
program, # children and adults in home.
322 children followed from birth (1979-1982)
to age 11-1 Syr, Port Pirie, Australia.
Moderate follow-up participation. Participants
had higher birth weight, older mothers, less
educated fathers. Log linear regression model
adjusted for maternal age, prenatal smoking
status, birth weight, type of feeding, length of
breastfeeding, maternal education, IQ, and
concurrent psychopathology, paternal
education, birth order, family functioning,
paternal occupation, parent smoking, marital
status, HOME, child IQ.
488 children followed from birth (1991-1992)
to age 7-8 yr, Avon, U.K.
All births in area eligible. Similar
characteristics as U.K. census, high
participation at baseline and follow-up.
Participants had better educated mothers,
who smoked less, better home environment.
Regression model adjusted for maternal
education and smoking, home ownership,
home facilities score, paternal occupation,
family adversity index, parenting attitudes at 6
mo. Also considered potential confounding by
child IQ.
301 boys selected from prospective cohort
followed from first grade to age 11 yr,
Pittsburgh, PA
Nested case-control. Moderate participation
rate. Participants had higher SES, lower
maternal IQ, smaller family size, higher IQ,
were nonwhite. ANCOVA adjusted for
maternal age, IQ, occupation, and education,
presence of both parents in home, Number
(#) of children in home, race, history of
medical problems, age, score at age 7 yr. Did
not consider potential confounding by
parental caregiving quality.
Blood
(ug/dL),
Tooth or
Bone (ug/g)
Pb Levels
0-6 yr avg
blood: NR
Lifetime (age
11-13 yr) avg
blood
GM (5th-95th)
Boys 1 4.3
(13.5-15.1)
Girls: 13.9
(13.2-14.6)
Intervals
analyzed:
Boys
13.7-14.9,
Girls
13.3-14.4 =
10th-90th
percentiles
Age 30 mo
blood
Mean (SD): NR
Reference: 0-2
2nd group: 2-5
3rd group: 5-10
4th group: >10
Bone at 10.2yr:
NR
Bone Pb levels
in high and low
groups NR
Outcome
Self-Report of
Delinquent
Behavior Score
Parental Report
of Predelinquent
and Delinquent
Bshsvior SCOTS
at ages 15-1 7 yr
Aggressive
Score, boys
Aggressive
Score qirls
Destructive
score, boys
Destructive
score, girls
Maternal rating
by Child
Behavior
Checklist at
ages 11-1 Syr
Antisocial
sctivitiss
Parent or
teacher rating by
Antisocial
Behavior
Interview at age
Syr
Delinquency
score (square
root)
Aggression
score (square
root)
Parent rating by
Child Behavior
Checklist at age
11 yr
Effect Estimate
/oco/ ^i\k
(95% Cl)
0.10(0.01,0.193)
0.09 (-0.02, 0.20)
0.17(0.08, 0.26)
0.10(0, 0.21)
0.06 (0.02, 0.09)
0.01 (-0.01 , 0.04)
ORs for increase in
score vs. reference
2nd: 0.93 (0.47, 1 .83)°
3rd: 1.44(0.73, 2.84)°
4th: 2.90 (1.05, 8.03)°
Low Pb group: 1.18
High Pb: 1.45, p=0.04
Low Pb: 2.43
High Pb: 2.98, p=0.009
November 2012
5-164
Draft - Do Not Cite or Quote
-------
Study3
Wasserman
et al. (2001)
Bellinger et
al. (|994b)
Study Population and
Methodological Details
191 children followed prenatally (from 1985)
to age 4-5 yr, Pristina, Yugoslavia.
Recruitment from prenatal clinics. High follow-
up participation, participants had less
educated mothers, higher concurrent blood
Pb, were Albanian. Log linear regression
model adjusted for sex, ethnicity, age,
maternal education, HOME, birth weight,
maternal smoking history.
1 ,782 children followed from birth
(1 979-1 980) to age 8 yr, Boston, MA area
Recruitment at birth hospital. High follow-up
participation. More participants were white,
had lower cord blood Pb levels, better birth
outcomes. Log linear regression model
adjusted for prepregnant weight, race,
Cesarean section, maternal marital status,
prenatal care, paternal education, colic, child
current medication use, sibship size, sex,
birth weight. Also considered potential
confounding by public assistance, prenatal
smoking, maternal education but not parental
caregiving quality.
Blood
(ug/dL),
Tooth or
Bone (ug/g)
Pb Levels
Lifetime (to age
4-5 yr) avg
blood
Mean (SD):
9.6(1.5)
Interval
analyzed: 7.8
(A r\iu
(i utn
percentile)-10
Tooth (age 6 yr)
Mean (SD):
3.4 (2.4) ug/g
10th-90th:
•1 o c o
1 .Z-D.O
Outcome
Aggressive
Score
Delinquent
Score
Maternal rating
by Child
Behavior
Checklist at
ages 4-5 yr
Total
externalizing
score T score,
inattentive,
nervous/
overact ive,
aggressive)
Teacher rating
by Child
Behavior Profile
at age 8 yr
Effect Estimate
/oco/ ^i\k
(95% Cl)
0.0 (-0.01, 0.01 7)
0.016(0.001,0.03)
0.51 (0.19,0.83)
Cross-sectional studies of Ratings of Misconduct
Braun et al.
(2008)
Chiodo et al.
(2007)
Sciarillo et al.
(1992)
2,867 children ages 8-15 yr (born
1986-1996), U.S. NHANES 2001-2004
Large multi-location study of multiple risk
factors and outcomes. Subjects with available
data were older, white, higher SES, with lower
blood Pb levels, had higher birth weight, and
fewer had household smokers. Logistic
regression adjusted for child age, poverty
income ratio, maternal age, sex, race, and
prenatal smoke exposure, cotinine levels. Did
not consider potential confounding by
parental caregiving quality.
451-460 African-American children, age 7 yr
(born 1989-1 991), Detroit, Ml
Recruitment at prenatal clinic. High
prevalence prenatal drug and alcohol
exposure. High participation rate. Linear
regression model adjusted for sex (both
outcomes), caretaker education, HOME,
maternal prenatal alcohol use, current
marijuana use (delinquent behavior),
maternal age, # children in home (social
problems). Also considered potential
confounding by SES, child age, maternal
prenatal and current maternal drug and
alcohol use, and IQ, current caretaker
psychopathology.
201 children (born 1984-1987) ages 2-5 yr,
Baltimore, MD.
High participation rate. Linear regression
adjusted for maternal education, employment
status, marital status, current depressive
symptom score, preschool children in the
home, child age, sex, Fe deficiency.
Concurrent
blood
Q1:<0.8
Q2: 0.8-1.0
Q3: 1.1-1.4
Q4: 1.5-10
Concurrent
blood
Mean (SD):
5.0 (3.0)
Interval
analyzed:
2.1-8.7 =
1 0th-90th
percentiles
Concurrent
blood
Mean (SD)
Low: 9.2 (2.9)
High: 27.8
(10.4)
Interval
analyzed: 5.9
(10th percentile
of lowgroup)-10
Conduct
disorder;
Parental report
using Diagnostic
Interview
Schedule for
Children-
Caregiver
Module at ages
8-1 5 yr
Delinquent
behavior
Social problems
Teacher rating
byAchenbach
Teacher Report
Form at age 7 yr
Total Behavioral
Problem score
(aggressive,
destructive,
somatic
problems, sleep
problems,
depressed,
social
withdrawal, etc)
Maternal rating
by Child
Behavior
Checklist at
ages 2-5 yr
ORs (yes/no) vs.
Q1 as reference
Q2: 7.24 (1.06, 49.5)°
Q3: 12.4(2.37, 64.6)°
Q4: 8.64 (1.87, 40.0)°
0.09(0, 0.1 8)d
0.10(0, 0.20)d
0.18(0.04,0.32)
November 2012
5-165
Draft - Do Not Cite or Quote
-------
Study3
Chen et al.
(2007)
Nigg et al.
(2008)
Study Population and
Methodological Details
622 children participating in TLC trial, age 7
yr, Baltimore, MD; Cincinnati, OH; Newark,
NJ; Philadelphia, PA.
Multi-city, high participation rate. High age 1-3
yr blood Pb levels 20-44 ug/dL that resulted in
chelation. Regression-based path analysis
adjusted for city, race, sex, language,
parental education and employment, single
parent, age at blood Pb measurement,
caregiver IQ. Considered potential
confounding by chelation treatment but not
parental caregiving quality. Direct=
independent of IQ. lndirect= mediated
through IQ
150 children, ages 8-1 7 yr, location NR.
Case-control study of ADHD. Recruitment by
community advertisements. High participation
rate. Did not consider potential confounding
factors.
Blood
(ug/dL),
Tooth or
Bone (ug/g)
Pb Levels
Concurrent
blood Mean
(SD): 8.0 (4.0)
Interval
analyzed: 3.9
(10th
percentile)-10
Concurrent
blood
Mean (SE):
Age 8-1 1 yr'
1 .04 (0.09) '
Age 1 2-1 7 yr:
Outcome
Externalizing
behavior
Direct, parent
Direct, Teacher
Indirect, parent
Indirect, Teacher
Behavior
Assessment
System for
Children
Oppositional
defiant disorder
Index
Parent, teacher
rating
Conners Rating
Scale-Revised
Effect Estimate
/oco/ ^i\k
(95% Cl)
OR for score > 60:
1 .024 (0.996, 1 .053)
1 .036 (1 .003, 1 .069)
1.008(1.002,1.014)
1.004(0.998,1.010)
r = 0.18, p O.05
Studies of Criminal Offenses
Wright et al.
(2008)
Fergusson et
al. (2008)
250 adults followed from birth (1979-1985) to
age 19-24yr, Cincinnati, OH
Prospective. Recruitment at prenatal clinic.
High follow-up participation. No selective
attrition. Negative binomial regression model
adjusted for maternal IQ and education, sex,
SES. Also considered potential confounding
by maternal prenatal smoking and prior
arrests, marijuana use, narcotic use, HOME,
birth weight, # children in the home, public
assistance in childhood.
911 children followed from birth (1977) to age
21 yr, Christchurch, New Zealand
Prospective. High follow-up participation.
Participants had lower SES. Negative
binomial regression model adjusted for
maternal education and prenatal
cigarettes/day, SES, ethnicity, family conflict,
physical abuse in childhood, parental alcohol
problems. Also considered potential
confounding by traffic density in childhood,
maternal age, paternal education, average
family income, maternal use of punishment,
parental drug use, parental bonding, child
marijuana use.
Age 6 yr
blood Pb
Median
(5th-95th):
6.8(3.4-18.3)
Age 0-6 yr avg
Median
(5th-95th):
12.3(6.0-26.3)
Tooth
(age 6-8 yr)
Mean:
6.2 ug/g
Criminal arrests
From county
records at ages
1 9-24 yr
Age 6 yr blood
Age 0-6 avg
blood
Convictions for
property and
violent offenses
From police
records at age
21 yr
RRs (yes/no):
1.05(1.01, 1.09)
1.01 (0.97, 1.05)
0.49(0.16,0.82)
November 2012
5-166
Draft - Do Not Cite or Quote
-------
Study3
Needleman et
al. (2002)
Study Population and
Methodological Details
340 adolescents, ages 12-18 yr (born
1974-1986), Pittsburgh, PA area.
Cross-sectional. 194 cases (county) and 146
controls (Pittsburgh high schools) from
different sources. Low participation from
cases. Logistic regression adjusted for race,
parental education and occupation, both
parents in home, # children in home,
neighborhood crime rate. Did not consider
potential confounding by parental caregiving
quality.
Blood
(ug/dL),
Tooth or
Bone (ug/g)
Pb Levels
Concurrent
Bone
Mean (SD)
in ppm
Cases:
11.0(32.7)
Controls:
1.5(32.1)
Outcome
Delinquent
status
From Juvenile
Court records at
ages 12-1 Syr
Effect Estimate
(95% Cl)b
ORs (yes/no) for bone
Pb level >25 ppm
White:
3.7(1.3, 10.5)°
African-American:
2.2(0.5, 10.0)°
aResults are organized according to outcome category, behavior ratings then documented criminal offenses. Within each category,
studies are organized by strength of design and analysis.
bUnless otherwise specified, effect estimates are standardized to a 1 ug/dL increase in blood Pb level or 1 ug/g increase in bone
or tooth Pb level in the interval from the 10th percentile to 10 ug/dL blood Pb or bone Pb or the 90th percentile, whichever is
lower.
°Odds in higher quantile of blood or bone Pb level compared to that in lowest quantile of blood or bone Pb level (reference).
d95% Cl was estimated from a reported p-value of 0.05.
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
Prospective studies provided key evidence for associations of earlier childhood and
lifetime average blood Pb levels and tooth Pb levels (from age 6-8 years) with conduct
problems such as delinquent behavior, aggression, antisocial activities, and destructive
behavior as rated by parents and/or teachers in children ages 8-17 years (Table 5-12).
These studies had moderate to high follow-up participation that was not conditional on
blood Pb levels, reducing the likelihood of selection bias. Across studies, behaviors were
assessed using various tests, but several studies used the Child Behavior Checklist.
Associations were found with both parent and teacher ratings, increasing confidence that
biased reporting of conduct problems by parents of children with higher Pb exposures did
not unduly influence the collective body of evidence. The evidence from prospective
studies is substantiated by associations observed in diverse populations (i.e., U.K.,
Cincinnati, Port Pirie, Australia) that considered several potential confounding factors
including multiple SES-related factors, parental caregiving quality, smoking exposure,
and birth outcomes (Chandramouli et al.. 2009; Dietrich et al.. 2001; Burns etal. 1999).
In the Yugoslavia cohort, lifetime average blood Pb level was associated with parent
ratings of aggressive and delinquent behavior with consideration for similar potential
confounding factors but in children from ages 4-5 years, in whom behaviors may be less
reliably measured or predictive of behavior at older ages (Wasserman et al.. 2001).
November 2012
5-167
Draft - Do Not Cite or Quote
-------
1 Collectively, the evidence from prospective studies indicated associations of teacher and
2 parental ratings of aggressive, destructive, antisocial, and delinquent behavior with
3 measures of cumulative Pb exposure, i.e., age 0-6 year average blood, lifetime average
4 blood, tooth, and bone Pb level. Associations with tooth (Bellinger et al.. 1994b) and
5 bone (Needleman et al., 1996) Pb level, collected prior to or at the same time as behavior
6 assessment, respectively, were found with adjustment for several potential confounding
7 factors as noted above, with the exception of parental caregiving quality. Collective
8 evidence from prospective studies most clearly indicated associations between blood Pb
9 level and ratings of conduct problems with population mean or group blood Pb levels
10 > 10 ug/dL (Chandramouli et al.. 2009; Dietrich etal.. 2001; Wasserman et al.. 2001;
11 Burns et al.. 1999). Among children ages 7-8 years in the U.K. born in the 1990s,
12 Chandramouli et al. (2009) recently found that compared with children with age
13 30 month blood Pb levels 0-2 ug/dL, children with blood Pb levels >10 ug/dL had
14 increased odds of greater antisocial activities as rated by parents or teachers with an OR
15 of 2.9 (95% CI: 1.05, 8.0). The Boston cohort was found to have lower childhood blood
16 Pb levels (Table 5-3); however, tooth Pb level was associated with a higher total
17 externalizing behavior score, which also included inattention (Bellinger et al.. 1994b).
18 As described above, cross-sectional studies also indicated blood Pb-associated higher
19 ratings of conduct problems; however, because of their various limitations discussed
20 below, their results were less of a consideration in drawing conclusions about the effects
21 of Pb exposure on conduct problems. In the recent analysis of 2,867 children ages 8-15
22 years participating in NHANES 2001 -2004, Braun et al. (2008) analyzed blood Pb level
23 as a categorical variable and found higher prevalence of conduct disorder as ascertained
24 by parental questionnaire with concurrent blood Pb levels in the range of 0.8 to
25 1.0 ug/dL. Compared with children with blood Pb levels <0.8 ug/dL, the OR in children
26 with blood Pb levels 0.8-1.0 ug/dL was 7.24 (95% CI: 1.06, 49.5). The wide 95% CIs
27 likely were due to the small numbers of cases of conduct disorder. For example, there
28 were 22 children rated as having conduct disorder in the group with blood Pb levels
29 0.8-1.0 ug/dL. Nigg et al. (2008) found a blood Pb-associated higher rating (parent or
30 teacher) of oppositional defiant disorder in a population with similarly low concurrent
31 blood Pb levels, means ~1 ug/dL. However, the implications are limited because of the
32 case-control design (Section 5.3.3.4) and lack of consideration of potential confounding
33 factors. Further, because these studies examined adolescents who likely had higher earlier
34 childhood Pb exposures, there is uncertainty regarding the level, timing, frequency, and
35 duration of Pb exposure that contributed to the observed associations.
November 2012 5-168 Draft - Do Not Cite or Quote
-------
1 Consideration for potential confounding varied among the cross-sectional studies of
2 conduct problems. With the exception of Nigg et al. (2008). most considered SES.
3 However, only a few considered parental caregiving quality (i.e.,. HOME score) or
4 current maternal psychopathology. An analysis of children in Baltimore, MD adjusted
5 results for current maternal depressive symptoms but examined children less than age 5
6 years and analyzed misconduct as a component of total behavior problem score, which
7 included internalizing behaviors (Sciarillo et al., 1992). Chiodo et al. (2007) found that
8 higher concurrent blood Pb level was associated with higher teacher ratings of social
9 problems and delinquent behavior in children ages 7 years in Detroit, MI with adjustment
10 for HOME score and initial consideration of current maternal psychopathology; however,
11 the study population had high prevalence of prenatal drug and alcohol exposure. Neither
12 of these exposures met the criteria for inclusion in the model, indicating lack of
13 confounding by these factors. Nonetheless, the results may not be representative of the
14 general U.S. population of children. Lack of representativeness also may pertain to the
15 results of Chen et al. (2007). who examined children who were given chelators at ages
16 1-3 years because of high earlier childhood blood Pb levels (20-44 ug/dL).
17 While few in number, evidence from prospective studies also indicated associations of
18 biomarkers of earlier childhood Pb exposure with delinquent and criminal acts as
19 objectively assessed from government records. These studies of delinquent and criminal
20 acts examined Pb levels in blood or tooth samples collected in the 1980s when Pb
21 exposures were much higher than those of the current U.S. population (Fergusson et al..
22 2008; Wright et al.. 2008). However, the prospective study design and consideration for
23 several potential confounding factors increase confidence that the observed associations
24 represent a relationship with Pb exposure.
25 In the Cincinnati cohort, prenatal cord and age 0-6 year average blood Pb levels were
26 associated with self- and parent-reported delinquent and social acts at ages 16-17 years
27 (Dietrich et al.. 2001). Wright et al. (2008) recently extended these findings to include
28 associations of prenatal cord and age 6 year blood Pb levels with criminal and violent
29 criminal arrests at ages 19-24 years. In models that adjusted for maternal IQ, sex, SES
30 score, and maternal education, the relative risks (RRs) for total arrests per 1 ug/dL
31 increment in blood Pb level were 1.07 (95% CI: 1.01, 1.13) for prenatal blood Pb level,
32 1.01 (95% CI: 0.97, 1.05) for age 0-6 year average blood Pb level, and 1.05 (95% CI:
33 1.01, 1.09) for blood Pb level at age 6 years. Interactions terms for blood Pb by sex were
34 not statistically significant; however, the attributable risk was considerably higher for
35 males (0.85 arrests/year [95% CI: 0.48, 1.47]) than for females (0.18 [95% CI: 0.09,
36 0.33]). A strength of Wright et al. (2008) was the detailed examination of potential
37 confounding by a large number of variables (Table 5-12). All of the examined covariates
38 were weakly correlated with blood Pb levels (r = 0.24-0.35), thereby reducing the
November 2012 5-169 Draft - Do Not Cite or Quote
-------
1 potential for confounding by the examined factors. Nonetheless, variables such as
2 maternal IQ, maternal education, and SES were included in the model because they were
3 associated with arrests in the full multivariate model or changed the blood Pb level
4 estimate by more than 10%. HOME score was similar between subjects with and without
5 criminal arrest records and did not meet the criteria for inclusion in final models.
6 The study of the New Zealand cohort also considered several potential confounding
7 factors such as family functioning and parental bonding (Table 5-12) (Fergusson et al..
8 2008). Per 1 ug/g Pb in teeth obtained between ages 6 and 8 years, there was a 0.49 (95%
9 CI: 0.16, 0.82) increase in the number of documented violent or property convictions at
10 ages 14-21 years. Results were adjusted for SES, ethnicity, maternal education, family
11 conflict, prenatal smoking exposure, physical abuse in childhood, and parental
12 alcoholism. The effect estimate for tooth Pb level decreased in adjusted models and was
13 found to account for <1% in the variance of criminal convictions; however, the
14 association remained statistically significant.
15 The epidemiologic studies described above employed different designs and assessed
16 conduct problems using different behaviors and methods. The consistency of association
17 of Pb biomarker levels with conduct problems was corroborated in a recent meta-analysis
18 (Marcus et al.. 2010) that included 19 studies (several of which are described above) with
19 a total of 8,561 children and adolescents (mean ages ranging from 3.5 years to
20 18.4 years). Effect estimates were converted to Pearson correlation coefficients, and the
21 combined effect estimate was r = 0.19 (95% CI: 0.14, 0.23). The key finding of this study
22 was the robustness of associations to between-study sources of heterogeneity. In the
23 meta-analysis, effect sizes did not differ significantly between prospective and cross-
24 sectional studies, among studies that examined different conduct problems
25 (i.e., opposition defiance, delinquency, externalizing problems), or among studies that
26 assessed conduct disorders using self-report, teachers report, or criminal records.
27 Adjustment for covariates such as SES, birth weight, parental IQ, and home environment
28 did not attenuate the relationship between blood Pb level and conduct problems. In
29 addition to strengthening the evidence for the independent associations of Pb biomarker
30 levels with conduct problems, the results indicated that the lack of adjustment for any
31 particular covariate, including HOME score, does not warrant limiting inferences from a
32 particular study. The major source of heterogeneity in effect estimates was the biomarker
33 of Pb examined. A larger magnitude of effect was estimated for hair Pb levels compared
34 with bone or blood Pb levels, which had similar effect sizes. The authors suggested that
35 hair Pb may be a better indicator of cumulative Pb exposure compared to bone Pb levels
36 due to the high turnover of bone throughout childhood and into adolescence; however, an
37 empirical basis for interpreting hair Pb measurements in terms of body burden or
38 exposure has not been firmly established (Section 4.3.4.2).
November 2012 5-170 Draft - Do Not Cite or Quote
-------
1 Several studies of misconduct aimed to characterize whether associations with
2 biomarkers of Pb exposure were independent of effects on IQ and educational attainment.
3 Most studies found that associations of Pb biomarkers with conduct problems remained
4 statistically significant in a model that additionally adjusted for child IQ or educational
5 attainment, indicating that Pb exposure may have a direct effect on misconduct
6 independent of its effect on cognitive function (Chandramouli et al.. 2009; Fergusson et
7 al., 2008; Burns et al.. 1999). However, simple statistical adjustment for cognitive
8 function indices may result in an underestimate of the effect on misconduct because a
9 decrement in cognitive function may lie on the causal pathway to behavioral problems.
10 Chen et al. (2007) used path analysis to characterize the direct effects and indirect effects
11 (mediated through child IQ) of blood Pb level on total externalizing problem ratings at
12 age 7 years; however, results were inconsistent. A direct effect was estimated for
13 externalizing problems rated by teachers, and an indirect effect was estimated for
14 problems rated by parents (Table 5-12). These findings may have limited applicability to
15 the general U.S. population given that some children in the study had been treated with
16 chelators at ages 1-3 years because of high blood levels, and it is uncertain whether the
17 observed associations were due to the residual effect of high earlier blood Pb levels
18 (20-44 jig/dL).
5.3.4.2 lexicological Studies of Aggression
19 While recent studies were not identified, evidence available in the 2006 Pb AQCD (U.S.
20 EPA. 2006b) pointed to the effects of Pb on changes in social behavior in rodents and
21 nonhuman primates. Most observations comprised Pb-induced increases in social and
22 sexual investigation, as indicated by sniffing, grooming, following, mounting, and
23 lordosis behavior. In animals, the social behavior most comparable to epidemiologic
24 findings in children is aggression; however, the effects of Pb on aggression in animals
25 were inconsistent. In animals, aggression was assessed as threats, attacks, bites, chases,
26 and offensive posture in encounters with other animals, and Pb exposure was found to not
27 affect, decrease, and increase aggression. Pb exposure generally was not found to affect
28 aggression in juvenile animals; however, increased aggression was found in adult animals
29 with high concentration (>2,500 ppm) gestational plus postnatal dietary Pb exposure.
30 Delville (1999) found Pb-induced increases in aggression with the lowest concentration
31 Pb exposure examined among all animal studies. Golden hamsters exposed to 100 ppm
32 Pb acetate GD8-PND42 in drinking water had blood Pb levels of 10 to 15 ug/dL at
33 PND42. As adults at PND45, Pb-exposed animals displayed more aggression as
34 measured by attacking and biting an intruder put in the cage. In mice, higher Pb exposure
35 produced mixed findings. BK:W mice exposed to 1,300 ppm Pb acetate in drinking water
November 2012 5-171 Draft - Do Not Cite or Quote
-------
1 from gestation through age 18 weeks displayed increased social and sexual investigation
2 but not aggression in males (femur Pb level at 18 weeks: 5,364 uM Pb/g ash) or females
3 (femur Pb level at 18 weeks: 4,026 uM Pb/g ash) (Donald et al.. 1986). Additional
4 investigation from the same laboratory exposed BK:W mice to 2,500 ppm Pb acetate in
5 drinking water from gestation through age 17-18 weeks, and found shorter latencies to
6 aggression in Pb-exposed mice than in controls (Donald et al.. 1987). In juvenile Long
7 Evans hooded rats, lactation-only (PND1-PND21) exposure to 670 ppm Pb chloride in
8 drinking water increased "rough and tumble" play behavior at PND36 which was not
9 characterized as aggression because of the lack of injury, submissive posturing, or escape
10 attempts in encounters with other animals (Holloway and Thor. 1987).
5.3.4.3 Summary of Conduct Problems
11 Although not examined as extensively as cognitive function, previous and recent
12 prospective studies consistently demonstrate Pb-associated increases in delinquent
13 behavior, aggression, antisocial activities, and destructive behavior as rated by parents
14 and teachers in children and as assessed with government records of criminal offenses in
15 adolescents and young adults (Table 5-12). Most studies examined multiple behaviors;
16 however, the consistent pattern of association observed across the ages of blood Pb level
17 and/or behaviors examined increases confidence that the evidence is not unduly biased by
18 an increased probability of associations by chance alone. Recent cross-sectional studies
19 found associations between concurrent blood Pb level and ratings of misconduct, but
20 several had additional limitations aside from study design, including prenatal drug and
21 alcohol exposure, treatment with chelators, or limited consideration for potential
22 confounding (Nigg et al.. 2008: Chen et al.. 2007: Chiodo et al.. 2007). The most
23 informative cross-sectional study was that finding a 7.24 (1.06, 49.47) higher odds of
24 conduct disorder in adolescents ages 8-15 years participating in NHANES with
25 concurrent blood Pb levels 0.8-1.0 (ig/dL compared with blood Pb levels <0.8 (ig/dL with
26 adjustment for age, sex, race, poverty to income ratio, and smoking exposure (Braun et
27 al.. 2008). However, the association was imprecise and could have been influenced by
28 higher past Pb exposures of the adolescents. Further, potential confounding by parental
29 caregiving quality was not examined. Evidence of Pb-induced aggression in animals was
30 mixed in adult animals with lifetime Pb exposure beginning in gestation and not indicated
31 in juvenile animals.
32 The evidence in children from prospective studies is substantiated by analyses of school-
33 aged children (ages 8-17 years) in populations from various locations and SES (i.e., U.K.,
34 Cincinnati, Port Pirie, Australia) with high participation rates, lack of indication of
35 substantial selection bias, and consideration of several potential confounding factors
November 2012 5-172 Draft - Do Not Cite or Quote
-------
1 including multiple SES-related factors, parental caregiving quality, smoking exposure,
2 and birth outcomes (Chandramouli et al.. 2009; Dietrich et al.. 2001; Burns etal. 1999).
3 Pb biomarker levels were associated with both parent and teacher ratings of conduct
4 problems, reducing the likelihood of biased reporting of conduct problems by parents of
5 children with higher Pb biomarker levels. Recent prospective studies of criminal offenses
6 in young adults, ages 19-24 years, strengthened previous evidence with consideration for
7 potential confounding by factors such as SES, smoking, drug, and alcohol exposure, and
8 parental caregiving quality (Fergusson et al.. 2008; Wright et al.. 2008). In the Cincinnati
9 cohort, a 1 ug/dL increase in age 6 year blood Pb level was associated with an increased
10 risk of total arrests with an RR of 1.05 (95% CI: 1.01, 1.09) with adjustment for maternal
11 IQ, sex, SES, and maternal education (Wright et al.. 2008). In the New Zealand cohort, a
12 1 ug/g Pb in teeth obtained between ages 6 and 8 years was associated with a 0.49 (95%
13 CI: 0.16, 0.82) increase in the number of documented violent or property convictions at
14 ages 14-21 years (Fergusson et al., 2008) with adjustment for SES, ethnicity, maternal
15 education, family conflict, prenatal smoking exposure, physical abuse in childhood, and
16 parental alcoholism. Further support for Pb-associated increases in conduct problems was
17 provided by a recent meta-analysis that found that evidence was robust to heterogeneity
18 in study design, definition and assessment method of conduct problems, potential
19 confounding variables examined, and population mean blood Pb levels (Marcus et al..
20 2010).
21 Associations of conduct problems (parent/teacher ratings and criminal offenses) with
22 earlier childhood blood (e.g., age 30 month, age 6 year), early childhood average blood
23 (e-g-, age 0-6 year), lifetime average blood (to age 11-13 years), tooth, and bone Pb levels
24 pointed to the effects of early childhood or cumulative Pb exposures. Associations were
25 found with a mean lifetime (to age 11-13 years) average blood Pb level of 14 (ig/dL,
26 mean age 6 year blood Pb level of 6.8 (ig/dL, and age 30 month blood Pb levels
27 >10 (ig/dL. Recent cross-sectional studies found associations with concurrent blood Pb
28 level and lower blood Pb levels, means 1-8 (ig/dL, but the study limitations detailed
29 above limit inferences regarding the effects of Pb exposure on conduct problems in these
30 populations. Most prospective studies did not analyze Pb biomarker levels at multiple
31 lifestages and time periods, including later childhood and more recent adult exposures, or
32 examine differences in association between Pb exposures at various lifestages and time
33 periods. The evidence does not identify an individual critical lifestage, time period, or
34 duration of Pb exposure associated with conduct problems in children or exclude an
35 effect of more recent Pb exposures.
November 2012 5-173 Draft - Do Not Cite or Quote
-------
5.3.5 Internalizing Behaviors in Children
5.3.5.1 Epidemiologic Studies of Internalizing Behaviors in Children
1 A majority of investigations of the effects of Pb on behavior in children has focused on
2 externalizing behaviors such as inattention, hyperactivity, aggression, and delinquency.
3 However, several studies also have linked biomarkers of Pb exposure in children with
4 internalizing behaviors characterized by directing feelings and emotions inward,
5 i.e., withdrawn behavior, symptoms of depression, fearfulness, and anxiety. Whereas
6 some studies found stronger associations for externalizing behaviors than for
7 internalizing behaviors (Plusquellec et al.. 2010; Wasserman et al.. 2001; Bellinger et al..
8 1994a; Sciarillo et al.. 1992). others did not find a clear difference in the strength of
9 association (Roy et al.. 2009b; Chiodo et al.. 2004; Bellinger et al.. 1994b). Internalizing
10 behaviors were assessed frequently using the Child Behavior Checklist, and as with
11 attention-related behavioral problems and conduct problems, were rated by parents and/or
12 teachers. Associations with both parent and teacher ratings increase confidence that
13 biased reporting of internalizing behaviors by parents of children with higher blood Pb
14 levels did not unduly influence the collective body of evidence. Most studies had
15 moderate to high follow-up participation. With the exception of the Yugoslavia cohort,
16 participation was not biased to higher blood Pb levels. Additionally, in most studies, a
17 consistent pattern of association was observed across the ages of blood Pb level and/or
18 multiple behaviors examined, which increases confidence that the evidence is not
19 strongly biased by an increased probability of associations found by chance alone.
20 Key evidence was provided by prospective studies in various populations, i.e., Boston,
21 Port Pirie, Australia, and Yugoslavia. Collectively, these studies found higher ratings for
22 internalizing behaviors in children (n = 322-1,511, ages 3-13 years) in association with
23 concurrent blood Pb level, lifetime average blood, and tooth Pb levels (Wasserman et al..
24 2001; Burns etal. 1999; Wasserman et al.. 1998; Bellinger et al.. 1994b). In the Port
25 Pirie cohort, Burns et al. (1999) found that higher lifetime average blood Pb levels (mean:
26 -14 (ig/dL) were associated with parental ratings of externalizing behaviors more
27 strongly in boys and with internalizing behaviors (i.e., withdrawn, anxious/depressed)
28 more strongly in girls ages 11-13 years, which may indicate sex differences in the effect
29 of Pb or differences in the types of behaviors that are observed and reported in girls
30 versus boys. Based on a log-linear model, a 1 (ig/dL increase in lifetime average blood Pb
31 level was associated with increased odds of an anxious/depressed rating above the
32 median of 1.04 (95% CI: 1.0, 1.09) among 159 boys ages 11-13 years (in the 10th-90th
33 percentile interval of blood Pb level 13.7-14.9 (ig/dL) and 1.08 (95% CI: 1.01, 1.15)
34 among 163 girls ages 11-13 years (in the 10th-90th percentile interval of blood Pb level).
November 2012 5-174 Draft - Do Not Cite or Quote
-------
1 These associations were found with the adjustment for factors related to SES and parental
2 caregiving including HOME score, family functioning score, and current maternal
3 psychopathology (General Health Questionnaire).
4 Differences between externalizing and internalizing behaviors also were found in the
5 Yugoslavia cohort according to age of assessment and blood Pb levels. This cohort was
6 examined between ages 3 and 5 years, ages at which behaviors may be less reliably
7 assessed. Among 379 children ages 3 years from the higher and lower Pb exposure
8 towns, higher cord and concurrent blood Pb levels were associated with higher maternal
9 ratings of anxious-depressed, withdrawn, and externalizing behaviors, with stronger
10 associations found for concurrent blood Pb level (mean: 25.8 (ig/dL) (Wasserman et al..
11 1998). Among 191 children ages 4-5 years from the lower Pb exposure town, higher
12 lifetime average blood Pb level (mean: 9.6 (ig/dL), was associated with higher ratings of
13 delinquent behavior and internalizing behaviors, with stronger associations found for
14 delinquent behavior (Wasserman et al.. 2001). A log unit increase in higher lifetime
15 average blood Pb level was associated with a 0.22 log (95% CI: -0.04, 0.47) higher rating
16 of withdrawn behavior and 0.19 log (95% CI: -0.05, 0.43) higher rating of anxious-
17 depressed behavior in children at ages 4-5 years. Results at each age were adjusted for
18 age, sex, HOME score, and maternal education. Results at age 3 years and 4-5 years were
19 additionally adjusted for residence type and maternal history of smoking, respectively.
20 With regard to important lifestages or durations of Pb exposure, results from prospective
21 studies did not clearly demonstrate differences in association among biomarkers
22 measured at various lifestages or time periods. The importance of childhood cumulative
23 exposure was indicated by associations with lifetime average blood Pb levels in the Port
24 Pirie cohort (to age 11-13 years) and in the Yugoslavia cohort (to age 5 years) and with
25 tooth Pb (from age 6 years) levels in the Boston cohort at age 8 years. In the Boston
26 cohort, Pb levels measured in teeth (mean: 3.4 (ig/g) but not cord blood were associated
27 with a higher teacher rating of internalizing behaviors at age 8 years (Bellinger et al..
28 1994b). In another Boston-area cohort, tooth (collected at first or second grade) Pb levels
29 were not associated with self-rated symptoms of depression (Profile of Mood States
30 questionnaire) at ages 19-20 years (Bellinger etal., 1994a). Prospective studies did not
31 analyze a detailed history of Pb biomarker levels to evaluate persistence of effects of
32 early exposure or to identify an individual critical lifestage or time period of Pb exposure
33 associated with increases in internalizing behaviors. The available evidence does not
34 preclude an effect of later childhood or more recent Pb exposure.
35 In the Cincinnati cohort, using structural equations, Dietrich et al. (1987b) found that the
36 associations of prenatal maternal and infant age 10 day blood Pb level (respective means:
37 8.3 and 4.9 (ig/dL) with poorer mood in infants (n = 185) ages 6 months were indirect,
November 2012 5-175 Draft - Do Not Cite or Quote
-------
1 meaning they were mediated through lower birth weight and/or shorter gestation. These
2 results suggested that Pb may exert its effects by impairing nervous system development.
3 The fetal period is an active period for neuronal differentiation, dendritic branching, and
4 synaptogenesis, which if impaired by Pb exposure, could have broad implications on a
5 wide range of subsequent neurodevelopmental effects. There are few such analyses, and
6 the findings are limited by the lower reliability of mood assessed in infancy.
7 Cross-sectional studies found associations between concurrent blood or hair Pb levels and
8 teacher and parent ratings of internalizing behaviors in children. Associations were found
9 in children ages 3-16 years (n = 303-756) in China and India with mean concurrent blood
10 Pb levels 9-14 (ig/dL (Liuetal.. 2011b: Bao et al.. 2009: Roy et al.. 2009a). Results were
11 adjusted for family income and parental education but not caregiving quality. In the few
12 studies of populations with mean blood Pb levels ~5 (ig/dL, results were inconsistent.
13 Chiodo et al. (2004) found an association with internalizing behaviors with adjustment
14 for child and caregiver life stress and marital status in 246 children age 7 years in Detroit,
15 MI who had high prevalence of prenatal alcohol or drug exposure. HOME score, SES,
16 maternal education, prenatal alcohol exposure, drug exposure were not found to influence
17 associations with blood Pb level; however, the results may lack generalizability to the
18 general population of U.S. children. A study that examined 79-91 Inuit children (age
19 5 years) in Quebec, Canada, did not find associations between concurrent blood Pb level
20 and internalizing behaviors with consideration of potential confounding by HOME score,
21 caregiver education and IQ, blood Hg levels, and prenatal smoking and alcohol exposure
22 (Plusquellecetal.. 2010).
23 Associations of Pb biomarkers with internalizing behaviors in children were observed
24 with consideration for a wide range of potential confounding factors, most commonly,
25 age, birth outcomes, parental education, and other SES-related factors. Parental
26 caregiving quality was evaluated in few studies. Blood Pb-associated higher ratings of
27 internalizing behaviors were found with adjustment for HOME score in the Yugoslavia
28 cohort (Wasserman et al., 2001; 1998). and HOME, family functioning, and current
29 maternal psychopathology (General Health Questionnaire) in the Port Pirie cohort (Burns
30 et al.. 1999). Several studies, each of which adjusted for a different set of covariates,
31 found similar or slightly attenuated effect estimates in univariate and multivariate models
32 (Wasserman etal.. 2001; Burns etal.. 1999: Bellinger et al.. 1994b). Collectively, these
33 observations increase confidence that the observed associations with Pb biomarkers
34 reflect a relationship with Pb exposure.
November 2012 5-176 Draft - Do Not Cite or Quote
-------
5.3.5.2 Toxicological Studies of Internalizing Behaviors
1 As in epidemiologic studies, toxicological studies have focused more on cognitive
2 function and attention-related behavioral problems and less on emotional- and mood-
3 related behaviors. Pb biomarker levels have been associated with ratings of withdrawn
4 behavior, depression, and anxiety in children, and this evidence is supported by findings
5 of Pb-induced anxiety, emotionality and depression-like behaviors in animals.
6 Emotionality has been indicated by loss of motivation and increased frustration in
7 response to errors and reward emission in visual or olfactory discrimination task trials in
8 Pb-exposed rats (Beaudin et al.. 2007; Stangle et al.. 2007). In each study, Long-Evans
9 rats were exposed to Pb during and after lactation (300 ppm Pb acetate via dam drinking
10 water PND1-PND17 then either 20 or 300 ppm PND17-PND30 in own water, with
11 respective blood Pb levels of 13 and 31 (ig/dL on PND52). In Beaudin et al. (2007).
12 greater disruption in performance (i.e., failure to enter testing alcove) after committing
13 errors and having rewards omitted was found in rats with blood Pb levels 13 and
14 31 (ig/dL tested at age 9-15 weeks. Pb-exposed rats also had greater response latency
15 with reward omission as indicated by the entrance into the testing alcove but failure to
16 respond within a set period of time. In Stangle et al. (2007). increased reactivity to errors
17 and reward omission was found in rats with blood Pb levels 31 (ig/dL. Blood Pb levels
18 were measured after a lag period, and peak blood Pb levels in these animals may have
19 been higher than those reported. In rhesus monkeys, emotional dysregulation was
20 indicated by tactile defensiveness after exposure to Pb acetate/50% glucose in 4 cc daily
21 milk formula from PND8 to ages 1-2 years (producing blood Pb 35-40 (ig/dL) (Moore et
22 al.. 2008).
23 In other studies, Wistar rats showed emotionality and depression-like behavior in the
24 open field test and forced swim test (i.e., Porsolt's Test) with gestational/lactational Pb
25 exposure (de Souza Lisboa et al.. 2005). The open field test monitors activity levels and
26 movements of animals in three dimensions. Depression-like behavior is indicated by
27 freezing behavior and low levels of activity. Emotionality is indicated by grooming or
28 freezing. In the forced swim test, animals are placed in a vertical cylinder of water from
29 which there is no escape and monitored for duration of struggling or attempt to escape.
30 Animals that stop quickly are ascribed a depression-like phenotype. As in many other
31 neurobehavioral tests, sex-specific differences in responses were found. Pb-exposed
32 males showed increased emotionality in the open field test as indicated by fewer counts
33 of rearing. Pb-exposed females showed a depression-like phenotype in the forced swim
34 test as indicated by longer time of immobility (de Souza Lisboa et al.. 2005). While
35 measured blood Pb levels of rats were low, 5-7 (ig/dL, they were measured after a lag in
36 exposure (PND70) and produced by oral gavage (10 mg/day) of mothers, a route that
November 2012 5-177 Draft - Do Not Cite or Quote
-------
1 may have uncertain relevance to human routes of Pb exposure. Pb-induced immobility in
2 the forced swim test also was found with 6-weeek postnatal Pb exposure via drinking
3 water but producing blood Pb levels 40 (ig/dL. Reducing internal Pb dose with the
4 chelator succimer reversed Pb-induced immobility (Stewart et al. 1996).
5 Depression initially may seem like an unexpected effect of immune modulation, but it has
6 been linked to an interaction between the CNS and the immune system via alterations in
7 cytokines such as IL-6 (Section 5.6.6.1). Dyatlov and Lawrence (2002) found that dietary
8 Pb exposure through lactation and a brief period after weaning (500 (JVI, PND1-PND22,
9 resultant blood Pb level: 17 (ig/dL) potentiated sickness behavior in mice in response to
10 bacterial infection. Sickness behavior was evidenced by an increase in serum IL-6 levels
11 with an accompanying decrease in food and water intake and increase in body weight.
12 This phenotype was correlated with decreases in the populations of several T cell
13 subtypes. Pb exposure also potentiated release of IL1(3, which plays an important role in
14 inflammatory responses to infection and has been shown to inhibit hippocampal
15 glutamate release in young but not aged animals. Sickness behavior also was induced in
16 Pb-exposed animals with IL-6 and IL-1 administration without infection, further
17 supporting a role for immunomodulation in mediating sickness behavior.
18 Pb exposure had mixed effects on anxiety-related responses as measured by the elevated
19 plus maze, which assesses behavior of rodents in an unfamiliar environment. The maze is
20 elevated above the floor and consists of two arms that are enclosed with walls intersected
21 with two arms that have no walls. The animal is placed in the center of the maze, and
22 longer latency to enter an open arm, and lower frequency and duration of entries into an
23 open arm are indicative of anxiety. In a study of gestational/lactational (GD1-PND24)
24 exposure to 2.84 mg/mL Pb acetate trihydrate in drinking water, Sprague-Dawley rats did
25 not differ in anxiety-related responses from controls. Blood Pb levels of Pb-exposed rats
26 were higher than those relevant to humans, 65.8 (ig/dL at PND25 (Molina et al.. 2011).
27 Another study exposed female rats postnatally (PND1-PND30) to 2,000 ppm Pb acetate
28 in drinking water, which yielded lower blood Pb levels, 34 (ig/dL. Pb-exposed rats had an
29 increase in anxiety-related behavior at PND60, as indicated by a lower percentage of
30 open arm entries and less time spent in the open arms (Foxetal. 2010).
November 2012 5-178 Draft - Do Not Cite or Quote
-------
5.3.5.3 Summary of Internalizing Behaviors
1 Internalizing behaviors, i.e., withdrawn behavior, symptoms of depression, anxiety, have
2 been examined less than externalizing behaviors (i.e., attention-related behavioral
3 problems, misconduct). However, several prospective studies found associations of
4 higher parental and teacher ratings of internalizing behaviors with higher tooth or lifetime
5 average blood Pb levels in children ages 8-13 years (Burns et al.. 1999; Bellinger etal..
6 1994b) and higher concurrent and lifetime average blood Pb levels in children ages 3-5
7 years (Wasserman et al.. 2001; 1998). Collectively, the lack of biased participation by
8 subjects with higher blood Pb levels and associations found with both parent and teacher
9 ratings increase confidence that the evidence is not unduly influenced by biased reporting
10 of behaviors by parents of children with higher blood Pb levels. The prospective studies
11 found associations with adjustment for several potential confounding factors, including
12 SES and parental education and caregiving quality. In prospective studies, associations
13 were found with tooth Pb levels and lifetime average and concurrent blood Pb levels.
14 While, there is not a clear indication of an individual critical lifestage or time period of
15 Pb exposure associated with internalizing behaviors, several observations point to an
16 effect of cumulative childhood Pb exposure. These results do not preclude an effect of
17 later childhood or more recent Pb exposures. In prospective studies, associations with
18 internalizing behaviors were found with a mean lifetime average blood Pb levels of
19 14 (ig/dLto age 11-13 years (Burns etal.. 1999) and 9.6 (ig/dLto age 4.5-5 years
20 (Wasserman et al.. 2001). In children ages 4.5-5 years (Yugoslavia cohort), in whom
21 behavioral ratings may be less reliably measured, lifetime average blood Pb level was
22 associated more strongly associated with the rating of delinquent behavior than ratings of
23 internalizing behaviors (Wasserman et al.. 2001). Cross-sectional studies provided
24 supporting evidence of concurrent blood Pb-associated increases in internalizing
25 behaviors in children ages 3-16 years, and several considered potential confounding by a
26 similar set of factors as did the prospective studies. However, associations in populations
27 with mean concurrent blood Pb levels ~5 (ig/dL were inconsistent.
28 Evidence in children is supported by observations that dietary Pb exposure (early
29 postnatal to day 22 or 30) resulted in depression-like behavior and emotionality in
30 rodents, (Beaudin et al.. 2007; Stangle et al.. 2007; Dyatlov and Lawrence. 2002) and
31 rhesus monkeys (postnatal to age 1-2 years) (Moore et al.. 2008). with some evidence in
32 rodents at blood Pb levels relevant to humans (13-31 (ig/dL). Evidence for Pb-induced
33 anxiety in animals is mixed. Mode of action support is provided by well-documented
34 evidence for Pb-induced changes in the HPA axis (Section 5.3.2.3) and dopaminergic and
35 GABAergic systems (Sections 5.3.2.2 and 5.3.11.4). which are involved in regulating
36 mood and emotional state.
November 2012 5-179 Draft - Do Not Cite or Quote
-------
5.3.6 Psychopathological Effects in Adults
5.3.6.1 Epidemiologic Studies of Psychopathological Effects in Adults
1 The potential effects of Pb exposure on mood and psychopathological effects
2 (e.g., anxiety, depression, schizophrenia) in adults have been examined less than that in
3 children or cognitive function in adults. However, evaluation of mood states is an integral
4 part of the neurocognitive test battery of the World Health Organization (WHO), and it
5 has been suggested that indices of the Profile Of Mood States may be particularly
6 sensitive to toxicant exposures (Johnson et al.. 1987). As with other nervous system
7 endpoints in adults, several studies reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b)
8 found higher prevalence of self-reported mood disorders and anxiety among Pb-exposed
9 workers (n = 43-576, mean concurrent or peak blood Pb levels: 31-79 (ig/dL) in
10 association with higher blood Pb levels or compared with unexposed controls
11 (n = 24-181, mean blood Pb levels: 15-38 (ig/dL) (Schwartz etal.. 2005; Maizlish et al..
12 1995; Parkinson et al.. 1986; Baker etal.. 1985; Baker etal.. 1984; Lilisetal.. 1977).
13 Several studies considered potential confounding by age, sex, education, medical
14 conditions, smoking, and alcohol use, but only Maizlish et al. (1995) examined other
15 occupational exposures. Most studies were cross-sectional, which makes uncertain the
16 temporal sequence between Pb exposure and development of psychopathological effects.
17 The few studies of adults without occupational Pb exposures participating in NAS and
18 NHANES demonstrated associations of concurrent blood and bone Pb level with
19 psychopathological effects. As bone Pb is a major contributor to blood Pb levels in adults
20 without current occupational Pb exposure, cross-sectional associations with each
21 biomarker may indicate effects of cumulative Pb exposure. These previous and recent
22 cross-sectional studies found associations with adjustment for several potential
23 confounding factors, including age, education, employment status, and alcohol use.
24 Further, the examination of multiple exposures and outcomes in these studies reduces the
25 likelihood of participation conditional on Pb exposure and psychopathological effects.
26 Analyses of 526 men ages 48-70 years in the NAS indicated associations of both higher
27 concurrent blood (mean: 6.3 (ig/dL [SD: 4.16]) and tibia (mean: 21.9 ng/g [SD: 13.5]) Pb
28 levels with higher prevalence of self-reported depression and anxiety (Rhodes et al..
29 2003). In a recent analysis of 744 NAS men ages 48-94 years, Rajan et al. (2007) found
30 associations of symptoms assessed using the Brief Symptom Inventory with patella and
31 tibia Pb levels. A 14 (ig/g increase in tibia Pb level was associated with an increased odds
32 of an anxiety score above the median of 1.18 (95% CI: 0.98, 1.42) and of depression
33 score above the median of 1.16 (95% CI: 0.97, 1.38). Similar effect estimates were found
November 2012 5-180 Draft - Do Not Cite or Quote
-------
1 for patella Pb level. Effect modification by ALAD genotype was not in a consistent
2 direction. For most mood symptoms, tibia bone Pb levels were associated with larger
3 ORs among the 587 men with the ALAD 1-1 genotype. In contrast, ORs for associations
4 between patella Pb levels and symptoms such as depression and positive symptom
5 distress index were larger among the 121 ALAD-2 carriers. In the NAS, effect
6 modification by ALAD genotype also was inconsistent for associations between tibia Pb
7 levels and cognitive performance (Rajan et al., 2008) (Section 5.3.2.7). The relationship
8 between ALAD-2 genotype and Pb bioavailability is not clear (Section 5.2.3.3).
9 A recent analysis of 1,987 adults ages 20-39 years participating in NHANES 1999-2004
10 was the largest study of psychopathological effects in adults and included both men and
11 women of multiple races and ethnicities (Bouchard et al., 2009). However, only
12 concurrent blood Pb levels were available for analysis. Various symptoms were examined
13 using the WHO Composite International Diagnostic Interview, which follows DSM
14 criteria. Adults with concurrent blood Pb levels >0.7 (ig/dL had higher prevalence of all
15 three self-reported disorders. Adults in the highest quintile of concurrent blood Pb level
16 (> 2.11 (ig/dL) had the highest OR for major depressive disorder (OR: 2.32 [95% CI:
17 1.13, 4.75]) and panic disorder (OR: 4.94 [95% CI: 1.32, 18.48]) compared with adults
18 with blood Pb levels <0.7 (ig/dL with adjustment for age, sex, race, education, and
19 poverty to income ratio. A monotonic increase in ORs was not found across the quintiles
20 of blood Pb levels. For all endpoints, ORs were larger in analyses excluding current
21 smokers. While associations were found with relatively low concurrent blood Pb levels,
22 there is uncertainty regarding the magnitude, timing, frequency, and duration of Pb
23 exposure that contributed to the observed associations.
24 In analyses of cohorts in California and New England born in the 1950s and 1960s, Opler
25 et al. (2008; 2004) reported associations between higher levels of cord plasma 5-ALA
26 and subsequent diagnosis of schizophrenia spectrum disorder (ascertained using DSM-IV
27 criteria) in adolescence and adulthood. Because of the lack of direct measurements of Pb
28 biomarker levels, post hoc analysis, and limited consideration for potential confounding,
29 firm conclusions are not warranted. Investigators measured 5-ALA levels in stored serum
30 samples as surrogates for Pb exposure only citing previous observations of a high
31 correlation (0.90) between categorized 5-ALA levels (cutpoint 9.05 ng/mL) and blood Pb
32 levels (cutpoint 15 (ig/dL). In the California cohort, 5-ALA level > 9.05 ng/mL was
33 associated with schizophrenia spectrum disorder with an OR of 2.43 (95% CI: 0.99,
34 5.96), with adjustment for maternal age at delivery. In pooled analyses of the California
35 and New England cohorts, 5-ALA level > 9.05 ng/mL was associated with schizophrenia
36 spectrum disorder with an OR of 1.92 (95% CI: 1.05, 3.52), with adjustment for maternal
37 age and education. An adjusted OR was not presented for the New England cohort alone,
November 2012 5-181 Draft - Do Not Cite or Quote
-------
and it appeared that the association in the pooled dataset was influenced by that found in
the California cohort.
5.3.6.2 Toxicological Studies of Mechanisms of Psychopathological Effects
3 An environmental origin of schizophrenia was proposed years ago (Tsuang. 2000). and
4 while epidemiologic evidence is inconclusive, toxicological studies have provided
5 indirect evidence to explain how Pb exposure may contribute to schizophrenia
6 development (Figure 5-11). Pb exposure has been shown to reduce function in the
7 NMDA receptor (NMDAR) and decrease hippocampal neurogenesis, which have been
8 associated with schizophrenia-related endpoints. Pb may bind a divalent cation site in the
9 NMDAR and allosterically inhibit glycine binding (Hashemzadeh-Gargari and Guilarte.
10 1999). NMDAR antagonists have been shown to exacerbate schizophrenia symptoms in
11 affected individuals and induce a schizophrenic phenotype in unaffected subjects (Coyle
12 and Tsai. 2004). Evidence supports a decrease in hippocampal degenerate gyrus (DG)
13 neurogenesis as a mode of action for Pb-associated schizophrenia induction.
14 Developmental Pb exposure inhibits neurogenesis in animal models (Section 5.3.11.9).
15 Decreased neurogenesis is seen in patients with schizophrenia (Kempermann et al.. 2008;
16 Reif etal.. 2006) and animal models of schizophrenia (Maedaetal.. 2007). and
17 clozapine, a treatment for schizophrenia, restores hippocampal DG neurogenesis in
18 animal models of schizophrenia (TVIaeda et al.. 2007) (Figure 5-11). These DG pathways
19 are also NMDAR-dependent.
Human Populations or Cohorts
ALAD and Schizophrenia
Associated in Human Cohort
(Epidemiologic)
Rodent
, Studies
Pb exposure
NMDAR
Antagonism
Pharmacologically-
induced Schizophrenia in
Animal models
Figure 5-11 Schematic representation of the contribution of Pb exposure to
the development of a phenotype consistent with schizophrenia.
November 2012
5-182
Draft - Do Not Cite or Quote
-------
5.3.6.3 Summary of Psychopathological Effects in Adults
1 Studies of Pb exposure and behavior in adults have focused on psychopathological effects
2 rather than aggression and criminal behavior. Evidence links occupational Pb exposure
3 with self-reported mood disorders and anxiety, although the cross-sectional design and
4 potential confounding by other occupational exposures limits the implications. However,
5 supporting evidence is provided by a few but large (n = 744 and 1,787) cross-sectional
6 studies in nonoccupationally-exposed adults that found associations of concurrent blood
7 (Bouchard et al.. 2009) and tibia (Raian et al.. 2008) Pb levels with depression and
8 anxiety as assessed with widely-used questionnaires such as the Brief Symptom
9 Inventory and the WHO Composite International Diagnostic Interview. Evidence was
10 provided by the NAS study of men (primarily white) and study of men and women
11 (various races/ethnicities) participating in NHANES, both of which involve the
12 examination of multiple exposures and outcomes. Studies in adults with and without
13 occupational Pb exposure found associations with adjustment for several confounding
14 factors, including age, education, employment status, and alcohol use. The cross-
15 sectional nature of these studies makes uncertain the temporal sequence between Pb
16 exposure and development of psychopathological effects and the critical level, timing,
17 frequency, and duration of Pb exposure. Both blood and bone Pb levels in adults reflect
18 cumulative exposure, and it is uncertain what are the relative contributions of past versus
19 recent Pb exposures to the observed associations.
20 The epidemiologic evidence is supported by observations that early postnatal (to just after
21 lactation) Pb exposure induces depressive- and anxiety-related phenotypes in animals
22 (Section 5.3.5.2). The mode of action is supported by evidence for Pb-induced changes in
23 the HPA axis and dopaminergic and GABAergic CNS processes, which mediate anxiety
24 and depression. While epidemiologic evidence for Pb-associated schizophrenia is
25 inconclusive, a few toxicological studies have shown that Pb exposure decreases
26 NMDAR function and hippocampal DG neurogenesis, which are found in animal models
27 of schizophrenia (agitation, trouble finding food, reduced swimming behavior).
28 Epidemiologic evidence indicates associations of Pb biomarker levels with depression
29 and anxiety in children and adults as rated by self, parents, or teachers. Differences in
30 associations for other behaviors may be related to what endpoints are examined. Studies
31 in children and young adults have focused on attention-related behavioral problems and
32 misconduct; studies of older adults did not examine such behaviors. Differential effects in
33 children and adults also may be expected given the predominance of different
34 neurophysiological processes operating at different ages, for example, neurogenesis and
35 brain development in children and neurodegeneration in adults. Differences in the effects
November 2012 5-183 Draft - Do Not Cite or Quote
-------
1 of Pb exposure between children and adults also may be related to differences in Pb
2 exposure profiles by age.
5.3.7 Sensory Organ Function
5.3.7.1 Epidemiologic Studies of Sensory Organ Function in Children
3 Although not as widely examined as cognitive and behavioral outcomes, several studies
4 found associations of higher blood Pb level with higher hearing thresholds or poorer
5 auditory processing in children, with evidence limited largely to that described in the
6 2006 Pb AQCD (U.S. EPA. 2006b). The prospective Cincinnati study with repeat
7 measures of blood Pb prenatally to age 5 years provided information on the temporal
8 sequence between Pb exposure and hearing effects and potential critical lifestages of
9 exposure and had extensive consideration for potential confounding. In this cohort,
10 poorer auditory processing in 215 children at age 57 months was associated with higher
11 prenatal maternal, neonatal (10-day), yearly age 1 to 5 year (means: 10.6-18.4 (ig/dL),
12 and lifetime average blood Pb levels, with the strongest associations found for neonatal
13 blood Pb level (mean: 4.8 [SD: 3.3] (ig/dL). A 1 (ig/dL higher neonatal blood Pb level
14 was associated with a 0.20-point (p < 0.01) and 0.26-point (p < 0.10) lower score on the
15 total and left ear Filtered Word test (indicating incorrectly identified, filtered, or muffled
16 words), with adjustment for hearing screen, social class, HOME score, birth weight,
17 gestational age, obstetrical complications, and maternal alcohol consumption (Dietrich et
18 al., 1992). Overall, the findings pointed to a stronger effect of Pb exposure in infancy.
19 Additional support was provided by the large U.S. cross-sectional NHANES II
20 (n = 4,519) (Schwartz and Otto. 1987) and Hispanic Health and Nutrition Examination
21 Survey (HHANES, n = 3,262) (Schwartz and Otto. 1991) studies. The examination of
22 multiple exposures and outcomes in these studies reduces the likelihood of participation
23 conditional on Pb exposure and hearing function. Each study found an association
24 between higher concurrent blood Pb level and higher hearing thresholds in children (ages
25 4-19 years). In HHNANES, an increase in concurrent blood Pb level (median: 8 (ig/dL)
26 from 6 to 18 (ig/dL also was associated with a 15% increase in the percentage of children
27 with a substandard hearing threshold (2,000 Hz). Higher concurrent blood Pb level also
28 was associated with higher hearing thresholds across several frequencies in a smaller
29 (n = 155) study of similarly aged (4-14 years) children in Poland with similar blood Pb
30 levels (median: 7.2 (ig/dL [range: 1.9-28]) (Osman et al.. 1999). In each of the studies in
31 children, associations persisted in analyses restricted to subjects with concurrent blood Pb
32 levels <10 (ig/dL. Each of these studies adjusted for different potential confounding
November 2012 5-184 Draft - Do Not Cite or Quote
-------
1 factors, but in stepwise regression analyses, each considered parental education, maternal
2 smoking, nutritional factors, and environmental noise. Across studies, associations were
3 found with adjustment for factors such as age, sex, ethnicity, family income, concurrent
4 or past colds, antibiotic use, degree of urbanization, and Apgar score.
5 Mechanistic support for associations with higher hearing thresholds in children was
6 provided by a few studies that found associations of blood Pb level with lower brainstem
7 auditory evoked potentials in children. Brainstem auditory evoked potentials measure
8 nerve electrical activity and are used to assess neurological auditory function. In
9 prospective analyses of the Mexico City cohort (n = 100, 113), Rothenberg et al. (2000;
10 1994b) reported associations with prenatal and postnatal blood Pb levels. At age 5-7
11 years, the shape of the concentration-response relationship differed between prenatal
12 maternal and postnatal (ages 1 and 4 years) blood Pb level. Higher blood Pb level at age 1
13 year and at age 4 years (mean reported for age 28 months: 10.8 (ig/dL) was associated
14 with lower interpeak intervals in auditory evoked potentials. Prenatal maternal blood Pb
15 level showed a biphasic relationship, with lower evoked potentials found with blood Pb
16 levels 1-8 (ig/dL and higher evoked potentials found with blood Pb levels 8-30 (ig/dL.
17 Results were adjusted for age, sex, and head circumference. In this cohort, maternal first
18 trimester blood Pb levels 10.5-32 ug/dL were associated with supernormal retinal ERG
19 (Rothenberg et al.. 2002b), the impact of which on visual impairment is not clear.
20 Associations with lower auditory evoked potentials also were found in small studies
21 (n = 13, 29) of children with higher concurrent blood Pb levels (i.e., range 6-84 (ig/dL)
22 than most of the current U.S. population (Holdstein et al.. 1986; Robinson et al.. 1985).
23 Recent cross-sectional studies aimed to identify the locus in the auditory system affected
24 by Pb exposure in the examination of a population of children (n = 53, 117, ages 2-18
25 years) living in Pb glazing communities in Ecuador with higher blood Pb levels than
26 those relevant to current U.S. children (means 33 and 37 (ig/dL) (Buchanan et al.. 2011;
27 Counter et al.. 2011). Concurrent blood Pb level was not correlated with the acoustic
28 stapedius reflex (Counter et al.. 2011) or distortion product otoacoustic emissions
29 (Buchanan et al.. 2011). indicating lack of effect on the auditory brainstem or inner ear,
30 respectively. Other loci were not examined, and potential confounding was not
31 considered.
November 2012 5-185 Draft - Do Not Cite or Quote
-------
5.3.7.2 Epidemiologic Studies of Sensory Organ Function in Adults
1 Studies of auditory function reviewed in the 2006 Pb AQCD consistently indicated
2 associations between blood Pb levels and changes in auditory evoked brainstem
3 potentials in occupationally-exposed adults but found less consistent associations with
4 hearing thresholds (U.S. EPA. 2006b). A few recent studies found increases in hearing
5 thresholds in Pb-exposed workers. A recent analysis of the NAS provided evidence in
6 nonoccupationally-exposed men for associations of tibia Pb levels with hearing loss.
7 Among 448 men in the NAS, higher tibia Pb level (mean: 22.5 ug/g) at mean age 64.9
8 years, measured up to 20 years after initial hearing testing, was associated with a faster
9 rate of increase in hearing threshold for frequencies of 1, 2, and 8 kHz and a pure tone
10 average. Men were free of hearing loss at baseline and had hearing tested repeatedly
11 (median 5 times per subject) over a median of 23 years (Park et al., 2010). Blood Pb was
12 not examined. In cross-sectional analyses adjusted for age, race, education, body mass
13 index, pack-years of cigarettes, diabetes, hypertension, occupational noise (based on a
14 job-exposure estimate), and presence of a noise notch (indicative of noise-induced
15 hearing loss), higher patella Pb level (mean 32.5 ug/g, measured within 5 years of hearing
16 test) was associated with a higher hearing threshold for frequencies greater than 1 kHz. A
17 21 ug/g (interquartile range) increase in patella Pb level was associated with pure tone
18 average hearing loss with an OR of 1.48 (95% CI: 1.14, 1.91) in adjusted analyses.
19 Similar, but slightly weaker associations were found for tibia bone Pb levels. In the NAS,
20 bone Pb levels were measured after the initial hearing measurement but reverse causation
21 is unlikely since bone Pb is an indicator of cumulative Pb exposure, and tibia Pb has a
22 half-life on the order of decades (Section 4.3). Bone Pb levels increase with age, and
23 although age was included as a model covariate, residual confounding by age is possible.
24 Recent cross-sectional studies added evidence for associations between higher concurrent
25 blood Pb levels and higher hearing thresholds in adults with occupational Pb exposures.
26 A hospital-based case-control study examined workers from diverse occupations and
27 examined potential confounding by other occupational exposures. Cases included
28 workers referred for hearing testing (average hearing thresholds above 25 dB), and
29 controls comprised workers with normal hearing thresholds who were having routine
30 occupational health examinations (Chuang et al.. 2007). Geometric mean blood Pb levels
31 were 10.7 ug/dL for the 121 cases and 3.9 ug/dL for the 173 controls. In models that
32 adjusted for age, smoking, alcohol consumption, years of noise exposure, as well as Mn,
33 As, and Se levels in blood, higher blood Pb levels were associated with higher hearing
34 threshold (0.5-6 kHz). The potential selection bias arising from the nonrandom
35 population sample may limit implications of these findings. Other studies found
36 associations of higher concurrent blood Pb level with increased hearing thresholds or
November 2012 5-186 Draft - Do Not Cite or Quote
-------
1 hearing loss in Pb-exposed workers (n = 183-259) but had limited or no consideration for
2 potential confounding (Forst et al.. 1997) and/or examined workers with mean blood Pb
3 levels >50 ug/dL (Hwang et al.. 2009; Wu et al.. 2000).
5.3.7.3 lexicological Studies of Sensory Organ Function
Effects on Auditory Function
4 The 2006 Pb AQCD (U.S. EPA. 2006b) described impaired auditory function in
5 nonhuman primates exposed to lifetime Pb beginning in gestation or birth to ages
6 8-13 years (resulting in blood Pb levels 33-170 (ig/dL during or just after Pb exposure)
7 (Rice. 1997: Lilienthal and Winneke. 1996). Pb-related effects persisted after Pb
8 exposure was terminated, and blood Pb levels had returned to baseline. Recent studies
9 provided similar evidence with lower Pb exposures and blood Pb levels. These
10 observations in animals are consistent with the epidemiologic associations described
11 above (Sections 5.3.7.1 and 5.3.7.2) but were related to higher Pb exposures than those
12 relevant for humans. Monkeys with lifetime Pb exposure from birth to age 13 years were
13 found to have decrements in auditory function, as evidenced by elevated thresholds and
14 increased latencies in brainstem auditory evoked potentials. Further, half of the pure tone
15 detection thresholds were above the control range at certain frequencies (Rice. 1997). In
16 addition to indicating hearing loss, brainstem auditory evoked potentials can indicate
17 impaired synaptic maturation and incomplete neuron axon myelination leading to
18 impaired neuronal conduction (Gozdzik-Zolnierkiewicz and Moszynski. 1969). Thus, the
19 findings from Rice (1997) and those described in the preceding sections for children may
20 indicate that Pb exposure impairs auditory nerve conduction. Studies in animals with
21 blood Pb levels >300 (ig/dL found that the cochlear nerve was especially sensitive to Pb
22 exposure (Gozdzik-Zolnierkiewicz and Moszvnski. 1969).
23 In a recent study, Laughlin et al. (2009) studied rhesus monkeys exposed to Pb acetate
24 prenatally to birth or postnatally from birth through weaning at age 5.5 months (maternal
25 drinking water, 3 months prior to mating until weaning, resulting in bone Pb levels at age
26 11 years of 7 and 13 (ig/g for prenatal and postnatal groups, respectively, and blood Pb
27 levels during Pb exposure of 35 and 46 (ig/dL, respectively). Auditory threshold testing
28 and threshold task testing was conducted at 13 years of age after blood Pb levels had
29 returned to those found in controls. At birth, animals were cross fostered, creating a
30 control group, a prenatal Pb group, and a postnatal Pb group; however, Pb-exposed
31 animals were analyzed as a single group. Pb exposure induced small, statistically
32 nonsignificant elevations in auditory thresholds in animals. Auditory threshold task-
33 related behavioral testing was also impaired in Pb-exposed animals. This study has
November 2012 5-187 Draft - Do Not Cite or Quote
-------
1 multiple limitations that could have contributed to that lack of statistically significant
2 aberrations, including limited power with the examination of 5 animals per group, the
3 inability of some of the monkeys to engage or focus on the task at hand which resulted in
4 fewer available measurements, differences between the sexes in inattention, and mixing
5 of the postnatal Pb and prenatal Pb-exposed animals into one group.
Effects on Vision
6 The 1986 and 2006 Pb AQCDs (U.S. EPA. 2006b) detailed the effects of Pb exposure
7 during perinatal development and adulthood on the visual system of animals, including
8 reduced visual acuity and supporting mechanisms of action such as alterations in the
9 retina (Fox et al.. 1997; Fox and Sillman. 1979). CNS visual processing areas (Costa and
10 Fox. 1983). and subcortical neurons involved in vision (Cline et al.. 1996). For example,
11 environmentally-relevant doses of Pb (10~3 (iM) in tadpoles inhibited the growth of
12 neurons in the subcortical retinotectal pathway, the main efferent from the retina (Cline et
13 al.. 1996). Pb-related aberrations in electrical responses in retinal cells, as measured by
14 electroretinograms (ERGs), have been found in rodents, nonhuman primates, and
15 children. Recent research expands upon the extant evidence by examining effects in
16 animals with lower Pb exposures or blood Pb levels.
17 Extensive work in nonhuman primates with Pb exposure during development or over a
18 lifetime (peak blood Pb levels 50-115 (ig/dL) showed dysfunction in temporal visual
19 function (responses to different frequencies of light flicker) under high luminance but no
20 change in spatial function (Rice. 1998). A recent study found no effects of Pb exposure
21 on spatial acuity as assessed with the modified Teller preferential looking paradigm
22 (Laughlin et al.. 2008) in rhesus monkeys exposed to Pb acetate postnatally (PND8-age
23 26 weeks via commercial milk formula, producing blood Pb levels of 35-40 ug/dL). In
24 monkeys, effects on vision were tested with higher Pb exposures than those relevant to
25 humans. Low-level developmental Pb exposure was found to result in sensorimotor
26 deficits in adult zebrafish (Rice et al.. 2011). Fish that were exposed as embryos (2 to 24
27 hours post-fertilization) to water containing 0.03 uM PbCl2 had impaired response to
28 visual stimulation (a rotating bar) under low light conditions. These zebrafish also failed
29 to respond normally to mechanosensory stimulation (0.01 and 0.03 uM PbCl2), showing a
30 significantly impaired startle response.
31 Animal toxicological evidence also shows that the lifestage of exposure and the dose of
32 Pb contribute to the complex and variable effects of Pb on the retina, as assessed by ERG
33 (summarized in Table 5-13). The biological relevance of these variable findings is
34 uncertain. Female rats exposed postnatally to 200 or 2,000 ppm Pb acetate exposure via
35 dam drinking water from birth through lactation, resulting in blood Pb levels of 19 and
November 2012 5-188 Draft - Do Not Cite or Quote
-------
1 59 ug/dL at weaning, respectively, had subnormal scotopic ERGs (decreased A- and B-
2 wave amplitudes) with decreased sensitivity and temporal resolution when assessed at
3 90 days of age (Fox et al., 1991). Similar results were obtained in multiple studies
4 conducted in in vitro models (Otto and Fox. 1993; Fox and Farber. 1988; Fox and Chu.
5 1988). Monkeys exposed to relatively high levels of Pb continuously from the prenatal
6 period to age 7 years (350 or 600 ppm Pb acetate, resulting in blood Pb levels of 40 and
7 50 ug/dL, respectively) had persistently increased maximal retinal ERG amplitude (B-
8 wave only, supernormality) and increased mean ERG latency when assessed 2 years after
9 Pb exposure was terminated when blood Pb levels were <10 ug/dL (Lilienthal et al.
10 1988).
November 2012 5-189 Draft - Do Not Cite or Quote
-------
Table 5-13 Summary of Pb-related effects observed on the visual system.
Study
Foxet al.
(2008)
Lilienthal et
al. (1988)
Fox et al.
(1997)
Rothenberg
etal.
(2002b)
F = Females,
" — " Denotes
Species
Long-
Evans
Rat
Rhesus
Monkey
Long-
Evans
Rat
Human
children
Pb Exposure Maximal Blood
Sex Protocol/Dose Pb Level (ug/dL)
F Prenatal-PNDIODW
Low, 27 ppm 1 2
Moderate, 55 ppm 24
High, 109 ppm 46
M Prenatal-lifetime, DW
& F 350 ppm -50
600 ppm -115
F PND1-PND21, DW
200 ppm DW 1 9
2,000 ppm DW 59
M Prenatal maternal a 10. 5
^ p 1st trimester
ERG
Abnormality
Supernormal
Supernormal
Subnormal
Supernormal
Supernormal
Subnormal
Subnormal
Supernormal
Retinal Retinal Retinal Cell
Progenitor cell Cellular Dopamine Layer
proliferation Apoptosis Levels Thickness
Yes Not affected Dose-dependent J, f
Yes Not affected Dose-dependent J, f
No Yes Dose-dependent J, J,
— — — —
— Yes — I
— Yes — I
— — — —
M = Males, PND = postnatal day, DW = Drinking water
not measured.
November 2012
5-190
Draft - Do Not Cite or Quote
-------
1 Pb exposure beginning in the gestational period (Pb acetate in drinking water from
2 2 weeks before mating to PND10) also induced supernormal ERGs in adult Long-Evans
3 rats, but only with low (27 ppm) and moderate (55 ppm) doses that produced blood Pb
4 levels 10-12 (ig/dL and 21-24 (ig/dL (Fox et al.. 2008) (Figure 5-12 and Table 5-13). This
5 exposure window represents the developmental period for the retina of the rat and is
6 analogous to gestational human retinal development. Subnormal ERGs were induced by
7 the high 109 ppm dose (Figure 5-12). which produced blood Pb levels 40-46 (ig/dL.
8 Results of this rodent study demonstrated persistent supernormal scotopic rod
9 photoreceptor-mediated ERGs in animals with blood Pb levels relevant to humans. These
10 findings were consistent with the associations observed between supernormal ERG and
11 prenatal maternal blood Pb levels > 10.5 ug/dL in male and female children (Rothenberg
12 et al., 2002b). The functional relevance of findings is uncertain as supernormal scotopic
13 ERGs may be recorded without other overt ophthamalogic changes and are rarely seen in
14 the clinical setting (Terziivanov et al., 1982).
15 Animal studies indicate that the dose of Pb and the exposure lifestage not only
16 differentially affect functional tests, i.e., ERG but also differentially affect retinal cell
17 numbers and morphology. Concomitant with Pb-induced supernormal ERGs, Fox et al.
18 (2008) found that 27 and 55 ppm gestational plus early postnatal Pb exposure increased
19 neurogenesis of rod photoreceptors and rod bipolar cells without affecting apoptosis of
20 Miiller glial cells and increased the number of rods in central and peripheral retina (Table
21 5-13). Concomitant with subnormal ERGs, higher-level gestational plus early postnatal
22 Pb exposure (109 ppm, blood Pb level 40-46 (ig/dL) decreased the number of rods in the
23 central and peripheral retina (Fox et al.. 2008). Early postnatal (PND1-PND21) Pb
24 exposure (200 or 2,000 ppm, producing blood Pb levels 19 and 59 (ig/dL) induced
25 scotopic ERG subnormality in adult rats, decreased the number of rods in the central and
26 peripheral retina, and decreased the retinal Zn concentration (Fox et al.. 1997) (Table
27 5-13). Similar observations were made in separate work in mice. Low and moderate
28 doses of Pb from gestation to PND10 (27 or 55 ppm Pb acetate in dam drinking water,
29 producing blood Pb levels of 12 and 25 ug/dL, respectively) induced greater and
30 prolonged rod bipolar cell neurogenesis and greater thickness and cell number of the
31 outer and inner neuroblastic layers of the retina (Giddabasappa et al., 2011). As in rats, at
32 higher doses of Pb (109 ppm Pb acetate, resulting in blood Pb levels of 56 ug/dL), there
33 was no increased rod neurogenesis in mice. Nitric oxide has been shown to regulate
34 retinal progenitor cell proliferation in chick embryos (Magalhaes et al.. 2006). Thus,
35 these authors postulated that impaired NO production may contribute to aberrant retinal
36 cell proliferation (Giddabasappa et al.. 2011).
November 2012 5-191 Draft - Do Not Cite or Quote
-------
•§ ; 0:0
I 1J»
I?
?-i I.WO
High iino
Note: *p <0.05. Low Pb = 27 ppm, blood Pb level 10-12 |jg/dL, Moderate Pb = 55 ppm, blood Pb level 21-24 |jg/dL, High
Pb = 109 ppm, blood Pb level 40-46 |jg/dL. Relative to controls (gray bars), low (white bars) or moderate (blue bars) Pb exposure
from gestation through postnatal day 21 induced supernormal electroretinograms (ERGs) whereas high Pb exposure (black bars)
induced subnormal ERGs.
Source: Fox et al. (2008)
Figure 5-12 Retinal a-wave and b-wave ERG amplitude in adult rats after
prenatal plus early postnatal Pb exposure.
1 Mechanistic understanding of the effect of Pb on the visual system includes the capability
2 of Pb to displace divalent cations, inhibit physiological enzymes, regulate cell
3 proliferation and apoptosis, perturb normal neuroanatomical formation (cytoarchitecture
4 in the brain), and affect neurotransmitters. The effects of Pb on the retina were shown to
5 be mediated by its inhibition of cGMP phosphodiesterase (PDE) (Srivastava et al.. 1995;
6 Fox and Farber. 1988). Independent of Pb exposure, pharmacological inhibition of cGMP
7 PDE has been linked with visual problems including alterations in scotopic ERGs (Laties
8 and Zrenner. 2002). Postnatal Pb exposure of animals (peak blood Pb levels: 19,
9 59 (ig/dL) or in vitro Pb exposure of rods isolated from these animals elevated cGMP
10 which contributed to elevated rod calcium concentration (Fox and Katz. 1992) and
11 subsequently induced apoptotic cell death in a concentration-dependent manner.
12 Pb has been shown to affect a plethora of neurotransmitters in the brain, and it has
13 recently been shown to affect neurotransmitters in the retina. With the aforementioned
14 gestational to PND10 exposure, Pb induced concentration-dependent decreases in adult
15 rat retinal synthesis of dopamine, which has functions in retinal growth and development
16 and signal transduction in rods and cones (Fox et al.. 2008) (Figure 5-13). As discussed in
17 the 2006 Pb AQCD (U.S. EPA. 2006b). other mode of action support for the effects of Pb
18 on the visual system is provided by observations of Pb-induced decreased Na+/K+ATPase
19 activity which have been reported in vitro and in vivo. Also, structural changes from
20 chronic Pb exposure (birth to age 6 years) included cytoarchitectural changes in visual
21 projection areas of the brain of monkeys; maximum blood Pb level in the low and high
22 dose group reached 20 (ig/dL and 220 (ig/dL, respectively (Reuhl etal.. 1989).
November 2012
5-192
Draft - Do Not Cite or Quote
-------
6
u e 4
2.0
I 1.6
si
>'! u
1— 0.8
0.4
0
I Dark-adapted
I Light-adapted
Control Low Moderate High
GLE
Control Low Moderate High
GLE
Control Low Moderate High
GLE
Control Low Moderate High
GLE
Note: *p O.05. GLE = Gestational Pb exposure to postnatal day 10. Low Pb = 21 ppm, blood Pb levels 10-12 ug/dL, Moderate
Pb = 52 ppm, blood Pb levels 21-24 ug/dL, High Pb = 109 ppm, blood Pb levels 40-46 ug/dL. A. DA = dopamine,
B. DOPAC = dopamine metabolite, C. HVA = dopamine metabolite, D. DOPAC/DA = ratio of dopamine metabolite to dopamine. Pb
exposure decreased dopamine, DOPAC, HVA, and DOPAC/DA in a concentration-dependent manner in light-adapted animals (blue
bars). In dark adapted animals (black bars), Pb exposure decreased dopamine, DOPAC, and DOPAC/DA but not always in
concentration-dependent manner.
Source: Fox et al. (2008)
Figure 5-13 Retinal dopamine metabolism in adult control and gestationally
Pb-exposed (GLE) rats.
5.3.7.4 Summary of Sensory Function
i
2
3
4
5
6
7
8
9
Children
Several studies indicated that higher blood Pb levels are associated with decrements in
auditory function in children ages 3-19 years, as evidenced by increases in hearing
thresholds. Results from the prospective Cincinnati cohort study (n = 215) provide key
evidence for associations of neonatal, yearly age 1 to 4 year, and lifetime average blood
Pb levels with increased hearing thresholds at age 57 months (Dietrich et al.. 1992). and
large (n = 3,000-4,000) cross-sectional analyses of children participating in NHANES
and HHANES provide supporting evidence for concurrent blood Pb levels (Schwartz and
Otto. 1991. 1987). The examination of multiple exposures and outcomes in NHANES
and HHANES reduces the likelihood of participation conditional on Pb exposure and
November 2012
5-193
Draft - Do Not Cite or Quote
-------
1 hearing function. In the Cincinnati cohort, mean blood Pb levels were 4.8 (ig/dL for
2 neonatal and 17.4 (ig/dL for lifetime average. In HHANES, the median concurrent blood
3 Pb levels was ~8 (ig/dL. Across studies, associations were found with adjustment for
4 factors such as age, sex, ethnicity, family income, concurrent or past colds, antibiotic use,
5 degree of urbanization, and Apgar score. Potential confounding by parental education,
6 nutritional factors, environmental noise, and maternal smoking also was considered.
7 Mechanistic evidence was provided by observations of associations between blood Pb
8 level with lower auditory evoked potentials in children, particularly associations found in
9 the prospective analysis of children in Mexico City with prenatal, age 1 year, and age
10 4-year blood Pb levels (Rothenberg et al.. 2000). Biological plausibility is provided by
11 evidence in animals indicating increased thresholds and increased latencies in brainstem
12 auditory evoked potentials in nonhumans primates with multi-year postnatal Pb exposure
13 beginning at birth (Rice. 1997; Lilienthal and Winneke. 1996). although auditory
14 assessment were made in adult animals ages 8-13 years. Pb exposure limited to the
15 gestational period or to the postnatal period to age 5 months was found to have weaker
16 effects (Laughlin et al.. 2009). In animals, auditory effects were found with higher blood
17 Pb levels (i.e., 33-170 (ig/dL) than those relevant to humans; thus, it is difficult to
18 ascertain support for observations in children.
19 Maternal first trimester blood Pb levels 10.5-32 ug/dL were associated with supernormal
20 retinal ERGs in children in Mexico City at ages 5-7 years (Rothenberg et al., 2002b). The
21 animal evidence showed ERGs in different directions depending on lifestage of Pb
22 exposure and blood Pb level. Supernormal ERGs were found in adult rats with prenatal
23 plus early postnatal (PND10) Pb exposure that produced blood Pb levels of 12 and
24 24 ug/dL (Fox et al., 2008). The implications of supernormal ERGs on visual impairment
25 is not clear, and the biological relevance of the nonlinear concentration-response is not
26 clear. For these reasons, the evidence for the effects of Pb on retinal ERGs was not a
27 major consideration in drawing conclusions about the effects of Pb exposure on sensory
28 function. In these animals, Pb exposure also increased rod cell neurogenesis and
29 decreased dopamine. Toxicological studies demonstrated a range of other effects on the
30 visual system including impaired visual function, and potential mechanisms such as
31 alterations in morphology and cell architecture, signaling, enzyme inhibition,
32 neurotransmitter levels, neuroanatomical development, cell proliferation, and retinal cell
33 apoptosis.
November 2012 5-194 Draft - Do Not Cite or Quote
-------
Adults
1 In adults, increased hearing thresholds or hearing loss were associated with bone Pb
2 levels in 448 NAS men who were unlikely to have had occupational Pb exposures (Park
3 et al., 2010) and with concurrent blood Pb levels in adults with current occupational Pb
4 exposure. In the NAS, the examination of multiple exposures and outcomes reduces the
5 likelihood of participation conditional on Pb exposure and hearing function. Among NAS
6 men, higher tibia Pb levels were associated with a faster rate of increase in hearing
7 thresholds over a 23 year follow-up with adjustment for age, race, education, body mass
8 index, pack-years of cigarettes, diabetes, hypertension, occupational noise, and presence
9 of a noise notch. Tibia Pb levels were measured up to 20 years after initial hearing
10 testing, and while Pb in tibia has a half-life on the order of decades, there is uncertainty
11 regarding the temporal sequence with changes in hearing thresholds. Temporality also is
12 difficult to establish in the cross-sectional occupational studies. A hospital-based case
13 control study found an association between higher concurrent blood Pb levels and higher
14 hearing thresholds among workers with relevant blood Pb levels (means 10.7 and
15 3.9 ug/dL in workers with and without hearing problems, respectively) (Chuang et al.,
16 2007). Among other factors, results were adjusted for other occupational exposures.
17 Biological plausibility is provided by evidence in animals with lifetime Pb exposure but
18 with higher blood Pb levels (i.e., 33-107 (ig/dL) than those relevant to humans. Adult
19 monkeys were found to have supernormal ERGs with developmental or lifetime Pb
20 exposure that produced blood Pb levels of 50 and 115 (ig/dL (Rice. 1998): however,
21 Pb-associated visual system effects in human adults are not well characterized.
5.3.8 Motor Function
22 Some studies in children have assessed fine motor function, i.e., response speed,
23 dexterity, as part of a battery of neurodevelopmental testing, and most have found
24 associations with blood Pb level. Fewer studies have examined gross motor function,
25 i.e., postural balance, action tremor, agility, but also have found associations with blood
26 Pb level in children. Poorer motor function also was found in Pb-exposed workers.
27 Key evidence from the prospective studies of the Cincinnati and Yugoslavia cohorts
28 demonstrated associations of blood Pb levels with poorer motor function with either
29 adjustment for or consideration for several potential confounding factors related to SES,
30 parental caregiving quality and education, smoking exposure, birth outcomes, sex, and
31 child health. In the Cincinnati cohort, higher earlier childhood average blood Pb levels
32 (0-5 year average or 78 month, exact levels not reported) were associated with poorer
33 fine (n = 195) (Ris et al., 2004) and gross motor function (n = 91) (Bhattacharya et al..
November 2012 5-195 Draft - Do Not Cite or Quote
-------
1 2006) assessed in adolescence (ages 12, 15-17 years). In this cohort recruited from birth,
2 follow-up participation was high and not conditional on blood Pb levels. The fine motor
3 function results were adjusted for maternal IQ, SES, HOME score, and adolescent
4 marijuana consumption. Collectively, these findings suggest the persistence of effects of
5 earlier childhood Pb exposure to later childhood; however, later childhood or concurrent
6 blood Pb levels were not examined. Assessments in Cincinnati cohort children at age 6
7 years indicated associations of concurrent (mean: 11.66 (ig/dL), lifetime average (mean:
8 12.3 (ig/dL), and neonatal (mean: 4.8 (ig/dL) but not prenatal maternal (mean: 8.4 (ig/dL)
9 blood Pb levels with poorer upper limb dexterity, fine motor composite score (n = 245)
10 (Dietrich et al., 1993a). and poorer postural balance (n = 202) (Bhattacharya et al., 1995).
11 These results were adjusted for HOME score and race. Additional covariates included
12 maternal IQ, SES, and sex for fine motor functions (Dietrich et al., 1993a) and height,
13 BMI, birth weight, bilateral ear infection, and foot area for postural balance
14 (Bhattacharya et al., 1995). Blood Pb levels were associated with fine and gross motor
15 function in unadjusted and adjusted analyses, increasing confidence that confounding by
16 the examined covariates did not unduly bias the observed associations. Prospective
17 analysis of the Yugoslavian cohort indicated an association of lifetime average blood Pb
18 level (exact levels not reported) with decrements in fine but not gross motor function at
19 age 4.5 years in 283 children (Wasserman et al.. 2000). Although only 50% of the cohort
20 was examined, and participation was greater among children with lower SES and HOME
21 score, participation was not conditional on higher blood Pb levels.
22 Supporting evidence was provided by most cross-sectional studies of motor function,
23 which found that concurrent blood Pb level was associated with poorer fine motor
24 function in children in Asia and Canada ages 3-16 years (n = 61-814) (Palaniappan et al..
25 2011; Min et al., 2007; Despres et al., 2005). In exception, Surkan et al. (2007) found that
26 higher concurrent blood Pb level was associated with better fine motor function as
27 indicated by faster finger tapping speed among 534 children in New England. This study
28 examined lower blood Pb levels than other studies (mean: 2.2 (ig/dL) but a similar age
29 range (6-10 years) and set of potential confounding factors (age, sex, caregiver IQ, SES,
30 race). Concurrent blood Pb level (mean: 5.0 (ig/dL) was associated with greater sway
31 oscillation, alternating arm movements, and action tremor in 110 Inuit children (ages 4-6
32 years) in Quebec, Canada (Despres et al.. 2005) with consideration for potential
33 confounding by factors such as HOME score, maternal education and several nutrient
34 levels. The population of Inuit children was selected from subsistence fishing
35 communities, who have higher exposure to Hg and polychlorinated biphenyls. Several
36 indices of fine and gross motor function were associated with blood Pb level, with
37 adjustment for these other exposures. Min et al. (2007) found impaired fine motor
38 function in 61 children in Korea with a mean concurrent blood Pb level of 2.9 (ig/dL;
39 however, the results were not adjusted by SES-related variables.
November 2012 5-196 Draft - Do Not Cite or Quote
-------
1 An association of Pb exposure with poorer motor function in adults was found in
2 Pb-exposed workers (Iwata et al.. 2005). although implications are limited by the cross-
3 sectional design, high concurrent blood Pb levels (mean: 40 ug/dL), and lack of
4 consideration for potential confounding by other occupational exposures. Among 121 Pb
5 smelter workers in Japan, higher blood Pb level was associated with greater sagittal sway
6 with eyes open (p <0.05) and eyes closed (p <0.01) and transversal sway with eyes closed
7 (p <0.05) with adjustment for age, height, smoking status, and drinking status. The
8 authors calculated a benchmark dose level (Budtz-Jorgensen et al.. 2001; NRC. 2000) of
9 14.3 ug/dL from a linear concentration-response model. A supralinear concentration-
10 response function was found to fit the data slightly better than was a linear function.
11 Pb exposure has shown mixed effects on endurance, balance, and coordination in animals
12 as measured by rotarod performance and treadmill testing. Lower concentration
13 gestational plus early postnatal (to PND10) Pb exposure (27 ppm, producing peak blood
14 Pb level < 10 (ig/dL at PNDO-PND10) resulted in significantly poorer rotarod
15 performance (i.e., falling off more quickly) than did higher exposure (109 ppm, blood Pb
16 level: 33-42 (ig/dL) in male (but not female) adult mice, indicative of a nonlinear
17 concentration-response relationship (Leasure et al.. 2008). Other rotarod experiments
18 examining various speeds of rotarod rotation and higher Pb exposures producing blood
19 Pb levels >60 (ig/dL, some administered by routes with uncertain relevance to humans,
20 yielded mixed results (Kishietal. 1983; Grant etal.. 1980; Overmann. 1977). Herring
21 gull chicks injected with a single i.p. bolus dose of Pb (100 mg/kg Pb acetate, a dose
22 selected to represent exposure in the wild) had slower development of motor skills versus
23 control birds, as assessed by the treadmill test (Burger and Gochfeld. 2005).
24 In summary, epidemiologic evidence demonstrates associations of higher blood Pb levels
25 with poorer fine and gross motor function in children ages 3-17 years. Little evidence is
26 available in adults. Prospective analyses of the Cincinnati and Yugoslavia cohorts
27 (n = 91-283) that considered several potential confounding factors such as SES and child
28 health found associations with earlier childhood blood Pb levels (i.e., age 78 month, 0-5
29 year average) in adolescents (Bhattacharya et al.. 2006; Ris et al.. 2004) and neonatal,
30 lifetime average, and concurrent blood Pb levels in children ages 4-6 years (Wasserman
31 et al.. 2000; Bhattacharya et al.. 1995; Dietrich et al.. 1993a). In the Cincinnati cohort,
32 neonatal blood Pb levels were lower than concurrent or lifetime average blood Pb levels
33 at age 6 years (means 4.8, 11.7, 12 (ig/dL, respectively). In cross-sectional studies that
34 examined similar potential confounding factors, results were inconsistent in populations
35 with lower blood Pb levels (means <5 (ig/dL) (Surkan et al.. 2007; Despres et al.. 2005).
November 2012 5-197 Draft - Do Not Cite or Quote
-------
5.3.9 Seizures in Animals
1 Previous studies did not consistently show that Pb exposure induced seizures in animals.
2 Pb-induced seizures were found in male Wistar rats exposed to Pb acetate postnatally
3 (250-1,000 ppm in drinking water PND30-PND60, resulting in blood Pb levels of
4 -20-42 (ig/dL), as indicated by a decrease in the elapsed time required to develop the first
5 myoclonic jerk and tonic-clonic seizure (Arrieta et al.. 2005). Also, the dose of the
6 seizure-inducing agent pentylenetetrazol (PTZ) required to induce seizures significantly
7 decreased in all Pb dose groups. Other studies showed no effect of Pb exposure on
8 seizures (Schwark et al.. 1985; Alfano and Petit 1981). In a study of early postnatal
9 Pb acetate exposure (2,000 ppm in drinking water PND1-PND25), Pb had variable effects
10 on induction of seizures in Sprague Dawley rats examined at PND25 or PND50,
11 depending on the convulsant-inducing agent administered (Chen and Chan. 2002). Chen
12 and Chan (2002) hypothesized that the variable effects may be due to the selective effects
13 on inhibitory and excitatory neurotransmission by age and blood Pb level, which were
14 47 (ig/dL and 11 (ig/dL at PND25 and PND50, respectively.
15 Recent investigation expanded on the work by Arrieta et al. (2005) by showing that Pb
16 exposure may induce seizure activity in another rodent species, BALB/c mice. Adult
17 (ages 2-3 months) male BALB/c mice were exposed to Pb acetate for 30 days via
18 drinking water (range of blood Pb levels 50-400 ppm Pb groups: 6.4-18 (ig/dL)
19 (Mesdaghinia et al.. 2010). Except for 50 ppm Pb exposure, all other Pb concentrations
20 significantly reduced the thresholds efface and forelimb clonus, myoclonic twitch,
21 running and bouncing clonus, and tonic hindlimb extension. In a study of adult male
22 Wistar rats, Pb administration by bolus injection (200 mg/kg Pb acetate or 50 mg/kg
23 Pb nitrate, single injection, 2 days, blood Pb levels >20 (ig/dL) also induced epileptic
24 form activity or seizures (Krishnamoorthy et al.. 1993).
November 2012 5-198 Draft - Do Not Cite or Quote
-------
5.3.10 Neurodegenerative Diseases
5.3.10.1 Alzheimer's Disease
1 Higher bone Pb level in NAS men (Wang et al.. 2007a: Weisskopf etal.. 2004; Wright et
2 al.. 2003) but not blood Pb level in adults in Sweden (Nordberg et al.. 2000) has been
3 associated lower scores on the MMSE, which is widely used as a screening tool for
4 dementia. Direct evidence regarding the effects of Pb exposure on Alzheimer's disease is
5 limited to studies reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b). which did not find
6 higher occupational exposure to Pb (Graves etal.. 1991) or Pb level in the brains
7 (Haraguchi et al.. 2001) in Alzheimer's disease cases than healthy controls. Overall, the
8 latter studies have sufficient limitations (e.g., case-control design that may be subject to
9 reverse causation, lack of blood or bone Pb measures, limited consideration for potential
10 confounding) such that evidence is inconclusive regarding the effect of Pb exposure on
11 Alzheimer's disease.
12 Despite inconclusive epidemiologic evidence, toxicological evidence indicates that Pb
13 exposure in early life promotes Alzheimer's disease-like pathologies in the brains of aged
14 adult animals. Alzheimer's disease is characterized by amyloid-beta peptide (Ab)
15 plaques, hyper-phosphorylation of the tau protein, neuronal death and synaptic loss. In
16 the last decade, the developmental origins of adult health and disease paradigm and the
17 similar Barker hypothesis have indicated that early life exposures can produce aberrant
18 effects in adults. Bolin et al. (2006) demonstrated the connection between developmental
19 exposure to Pb in the rat and inflammation-associated DNA damage with
20 neurodegenerative loss in the adult brain. Wu and colleagues (2008a) had similar findings
21 in a study examining infantile Pb exposure of monkeys. These results suggest the need to
22 directly examine the long-term effects of developmental exposure to toxicants rather than
23 relying on adult exposure alone to predict potential health risks in adults (Dietert and
24 Piepenbrink. 2006).
25 The fetal basis of amyloidogenesis has been examined extensively by the Zawia
26 laboratory in both rodents and nonhuman primates. Mechanistically, Ab plaques originate
27 from the cleavage of the amyloid precursor protein (APP). In male rodents exposed to Pb
28 as infants (200 ppm Pb acetate PND1-PND20 in dam drinking water, resulting in pup
29 PND20 blood Pb level of 46 (ig/dL and cortex 0.41 (ig/g wet weight of tissue) or as
30 adults, infancy Pb exposure induced APP gene expression in the aged animal brains. A
31 bimodal response was observed, with a significant increase in APP expression above that
32 in control animals first manifesting in infancy and again in old age (82 weeks) (Basha et
33 al.. 2005). A concomitant bimodal response was observed in specificity protein 1 (Spl), a
November 2012 5-199 Draft - Do Not Cite or Quote
-------
1 transcription factor involved in gene expression in the early development of an organism
2 and known to be related to APP expression. Ab was also significantly elevated in the
3 aged animals developmentally exposed to Pb. Adult-only (18-20 weeks) exposure to Pb
4 did not alter APP or Sp 1 expression or Ab production.
5 Consistent with findings in rodents, Wu et al. (2008a) found that Pb exposure in infancy
6 (PND1-PND400, 1.5 mg/kg/day in infant formula) resulted in significantly higher gene
7 expression of APP and Spl and significantly higher protein expression of APP and Ab in
8 aged female monkey cortex tissue (23 year-old Macaca fasclculans) from a cohort of
9 animals established in the 1980s by Rice (1992a. 1990). At PND400, the monkeys had
10 blood Pb levels of 19-26 (ig/dL. In old age when amyloid plaques had manifested, blood
11 Pb levels and brain cortex Pb levels had returned to control levels. Together, the rodent
12 and nonhuman primate evidence concurs, and indicates that developmental Pb exposure
13 and not adult-only exposure induces elevations in neuronal Alzheimer's Disease-related
14 plaque proteins in aged animals.
15 Mechanistic understanding of Ab production and elimination after Pb exposure was
16 examined in human SH-SY5Y neuroblastoma cells exposed to Pb concentrations of 0, 5,
17 10, 20, and 50 (iM for 48 hours. Pb was found to affect two separate pathways to increase
18 Ab. Pb exposure induced both the overexpression of APP and repression of neprilysin, a
19 rate-limiting enzyme involved in Ab metabolism or removal (Huang et al.. 201 la).
20 Further mechanistic understanding of how Ab peptide formation is affected by Pb
21 exposure was provided by Behl et al. (2009). The choroid plexus is capable of removing
22 beta-amyloid peptides from the brain extracellular matrix. Pb was shown to impair this
23 function, possibly via the metalloendopeptidase, insulin-degrading enzyme (IDE), which
24 metabolizes Ab (Behl et al., 2009). In another study, lactational Pb exposure of Long -
25 Evans hooded rat pups induced perturbations in DNA binding of SP 1 via its Zn finger
26 protein motif (Basha et al.. 2003). This effect of Pb was ameliorated by exogenous Zn
27 supplementation.
28 An additional study with gestational plus lactational Pb exposure (1,000-10,000 ppm,
29 dam drinking water, resultant offspring blood Pb levels: 40-100 (ig/dL) showed that the
30 rodent hippocampus as early as PND21 contained neurofibrillary changes, commonly
31 used a marker for Alzheimer's disease. These changes manifested with hyper-
32 phosphorylated Tau, which comprises neurofibrillary tangles, and increased tau and beta
33 amyloid hippocampal protein levels (Li et al., 2010b).
34 In summary, recent studies showed that Pb exposure of rats and monkeys during infancy
35 or during gestation/lactation induced significant increases in neuronal plaque associated
36 proteins such as Ab-peptide, activation of Ab-supporting transcription factors, and
37 hyperphosphorylation of tau, all of which are pathologies found in humans with
November 2012 5-200 Draft - Do Not Cite or Quote
-------
1 Alzheimer's disease. These pathologies were not found with adult-only Pb exposure of
2 animals, further demonstrating that early life Pb is a critical window for Pb-induced
3 Alzheimer's-like pathologies in animals. The few epidemiologic studies have not linked
4 higher Pb exposure with Alzheimer's disease. These case-control studies lacked
5 assessment of blood or bone Pb levels or consideration for potential confounding. The
6 animal evidence indicates that epidemiologic studies assessing concurrent brain Pb levels
7 or occupational Pb exposure may not have examined the etiologically relevant exposure
8 period. However, the observations that were made in experimental animals with high Pb
9 exposure and blood Pb levels (>40 (ig/dL) may have uncertain relevance to humans.
10 Further, animals were not behaviorally assessed for dementia.
5.3.10.2 Amyotrophic Lateral Sclerosis
11 The 2006 Pb AQCD (U.S. EPA. 2006b) reported mixed epidemiologic findings for an
12 association between Pb and ALS based on case-control studies, several of which relied on
13 indirect methods of assessing Pb exposure. Case-control studies that measured blood Pb
14 levels produced contrasting results. A study of 16 ALS cases (mean blood Pb level:
15 12.7 (ig/dL) and 39 controls (mean blood Pb level: 10.8 (ig/dL) found a small difference
16 in the mean concurrent blood Pb level (Vinceti etal.. 1997). A larger study of 109 cases
17 and 256 controls that examined concurrent blood and bone Pb levels in a New England-
18 area population found higher odds of ALS among subjects with concurrent blood Pb
19 levels > 3 (ig/dL (e.g., OR: 14.3 [95% CI: 3.0, 69.3] for n = 55 blood Pb levels 3-4 (ig/dL
20 compared with blood Pb levels 10 (ig/g
22 patella Pb and > 8 (ig/g tibia Pb), but lacked precision. For example, compared with
23 subjects with tibia Pb level <8 (ig/g, the OR for tibia Pb levels 8-14 (ig/g was 1.6 (95%
24 CI: 0.5, 5.6). Results were adjusted for age, education, and hours/day inactive. Potential
25 confounding by smoking was not considered. Also in this population, an estimate of
26 cumulative Pb exposure based on occupational history was found to be associated with
27 ALS (Kamel et al.. 2002). The stronger findings for blood Pb level were surprising given
28 that bone Pb level is a better biomarker of cumulative Pb exposure. One explanation for
29 these findings is reverse causation. Blood was collected from people who already had
30 ALS, and reduced physical activity among those with ALS could lead to more bone
31 turnover and greater release of Pb from bones into circulation in ALS cases than controls.
32 Since the 2006 Pb AQCD, a few additional studies of ALS have been conducted with the
33 same New England-area case-control group. Kamel et al. (2005) reported that the
34 association between blood Pb level and ALS was not modified by the ALAD genotype,
35 and Kamel et al. (2008) found that higher tibia and patella Pb levels were associated with
November 2012 5-201 Draft - Do Not Cite or Quote
-------
1 longer survival time among 100 of the original 110 ALS cases with adjustment for age,
2 sex, and smoking. Results were not altered by the additional adjustment for education,
3 BMI, or concurrent physical activity. Higher blood Pb levels were associated weakly with
4 longer survival time. These paradoxical findings that point to a protective effect of Pb are
5 not easily explainable but find coherence with results for Pb-induced increased survival
6 time in an ALS mouse model (see below). On one hand, the cases with longer survival
7 time may have higher bone Pb levels because they reflect a longer period of cumulative
8 exposure. On the other hand, with longer survival time, there could be greater progression
9 of disease and less mobility. Decreased mobility would tend to increase bone resorption,
10 lower bone Pb levels, and increase blood Pb levels over time. The latter hypothesis is a
11 less likely explanation for findings in this New England cohort because higher bone Pb
12 levels were more strongly associated with longer survival time than was blood Pb level.
13 Another case-control study examined concurrent blood Pb levels and ALS among 184
14 cases (33 were either progressive muscular atrophy or primary lateral sclerosis, mean
15 blood Pb level: 2.41 (ig/dL) and 194 controls (mean blood Pb level: 1.76 (ig/dL) (Fang et
16 al.. 2010). The cases were recruited from the National Registry of U.S. Veterans with
17 ALS, and controls were recruited from among U.S. Veterans without ALS and frequency
18 matched by age, gender, race, and past use of the Veterans Administration system for
19 health care. A doubling of concurrent blood Pb level was associated with ALS with an
20 OR of 2.6 (95% CI: 1.9, 3.7) with adjustment for age and a collagen protein as an
21 indicator of bone formation. Associations did not differ substantially by indicators of
22 bone turnover but were slightly higher among ALAD 1-1 carriers. The association with
23 blood Pb level was similar in analyses that excluded the progressive muscular atrophy
24 and primary lateral sclerosis cases. The similar results by degree of bone turnover suggest
25 that reverse causation is not likely explaining the association between blood Pb level and
26 ALS. However, as in other ALS case-control studies, the directionality of effects is
27 difficult to establish. This study did not have measures of bone Pb to assess the
28 association with biomarkers of longer-term Pb exposure.
29 Although epidemiologic studies have provided inconsistent evidence for associations of
30 Pb biomarker levels with ALS in adults, toxicological studies have found that Pb
31 exposure affects neurophysiologic changes associated with ALS. For example, chronic
32 postnatal Pb exposure from weaning onward (200 ppm Pb acetate in drinking water,
33 resultant blood Pb level: 27 (ig/dL) reduced astrocyte reactivity and induced increased
34 survival time in the superoxide dismutase transgenic (SOD1 Tg) mouse, which has SOD
35 mutations found in humans with familial ALS (Barbeito et al.. 2010). In this model, Pb
36 exposure did not significantly increase the onset of the ALS disease. These findings
37 provide coherence with the association observed between bone Pb level and longer
38 survival time in patients diagnosed with ALS (Kamel et al.. 2008).
November 2012 5-202 Draft - Do Not Cite or Quote
-------
1 Research outside of the Pb field has suggested different mechanisms for ALS initiation
2 versus ALS progression, i.e., motor neuron function versus astrocyte and microglia
3 function (Yamanaka et al., 2008; Boillee et al., 2006). Astrocyte vascular endothelial
4 growth factor (VEGF) was examined for its involvement in the effects of Pb on
5 increasing survival time in the ALS mouse model. Lower VEGF expression has been
6 linked with risk of ALS in humans and ALS-like symptoms in animals. Baseline VEGF
7 levels were elevated in astrocytes from the ventral spinal cord of untreated SOD1 Tg
8 mice versus untreated nontransgenic mice. VEGF was not induced in the astrocytes of
9 Pb-treated nontransgenic mice. In comparison, Pb-exposed SOD1 Tg mice, which had
10 longer survival time, also had significant elevations in astrocyte VEGF (Barbeito et al.,
11 2010). These findings for Pb-induced effects on astrocytes in a mouse model for ALS
12 may provide a mechanistic explanation for Pb effects on survival time in ALS.
13 Others reported that VEGF administration to the SOD1 Tg mice significantly reduced
14 glial reactivity, a marker or neuroinflammation (Zheng et al.. 2007). Using a cell-based
15 co-culture system of neurons and astrocytes isolated from Pb-exposed SOD1 Tg mice,
16 Barbeito et al. (2010) found that an up-regulation of VEGF production by astrocytes was
17 protective against motor neuron death in the SOD1 Tg mouse cells. Thus, in vivo and in
18 vitro results indicate that chronic Pb exposure resulted in increased survival time in an
19 ALS mouse model and was correlated with higher spinal cord VEGF levels, which made
20 astrocytes less cytotoxic to surrounding motor neurons (Barbeito et al.. 2010).
21 In summary, there is inconsistent evidence of association between indicators of Pb
22 exposure (history of occupational exposure, Pb biomarker levels) and ALS prevalence
23 and survival time in humans. Because of the potential for reverse causality and bias due
24 to survival time in the case-control studies, and the lack of objective assessment of
25 occupational exposure, firm conclusions are not warranted. While several studies have
26 considered potential confounding by age, education, and physical activity, few have
27 considered smoking. Toxicological evidence also points to Pb exposure increasing
28 survival in a mouse model of ALS and has suggested explanations including Pb-induced
29 increases in VEGF expression and subsequent reduction in glial activity and protection of
30 motor neurons against inflammation.
November 2012 5-203 Draft - Do Not Cite or Quote
-------
5.3.10.3 Parkinson's Disease
1 Previous Pb AQCDs reviewed a few studies, some ecological (Rybicki etal.. 1993;
2 Aquilonius and Hartvig. 1986) and some case-control relying on questionnaire data or
3 occupational history (Gulson et al.. 1999; Gorelletal.. 1997; Tanner etal.. 1989) that
4 indicated associations between exposure to heavy metals, particularly Pb, and risk of
5 Parkinson's disease. The limited number of previous studies, weak study designs, and
6 lack of examination of Pb biomarkers did not permit firm conclusions. Recent studies
7 maintain several of these limitations but have indicated associations with bone Pb levels.
8 A recent large case-control study (330 cases, 308 controls) examined a population in the
9 Boston, MA area with virtually no occupational exposures to Pb (Weisskopf et al., 2010).
10 Subjects in the highest quartile of tibia Pb level (>16.0 (ig/g) had higher odds of
11 Parkinson's disease compared to those in the lowest quartile (< 5 (ig/g) (OR: 1.91 [95%
12 CI: 1.01, 3.60]) with adjustment for age, race, pack-years smoking, education, and
13 recruitment site. Cases and controls were recruited from several different sources
14 including movement disorder clinics and the NAS, which could have introduced biased
15 participation by Pb exposure or reduced representativeness to the target population. In the
16 NAS, cases were ascertained from self-report, which may introduce measurement error.
17 However, when analyses were restricted to cases recruited from movement disorder
18 clinics and their spouse, in-law, or friend as controls, the results were even stronger (OR:
19 3.21 [95% CI: 1.17, 8.83]). Although the use of spouse, in-law, and friend controls can
20 introduce bias, this is expected to be toward the null as these groups are likely to share
21 many exposures. Manganese (Mn) exposure has been associated with Parkinsonian
22 symptoms and could potentially confound associations between Pb and Parkinson's
23 disease. Weisskopf et al. (2010) did not adjust results for Mn exposure. However, unlike
24 occupational exposure to Pb, general environment exposure to Pb is less likely to be
25 correlated with environmental Mn exposure. Thus, it is less likely that the observed
26 associations with Pb were confounded by co-occurring Mn exposure.
27 Coon et al. (2006) conducted a smaller case-control study of 121 Parkinson's disease
28 patients and 414 controls frequency-matched by age, sex, and race, all receiving health
29 care services from the Henry Ford Health System in Michigan. Subjects in the highest
30 quartile of both tibia (OR: 1.62 [95% CI: 0.83, 3.17] for levels > 15 (ig/g) and calcaneus
31 (OR: 1.50 [95% CI: 0.75, 3.00] for levels > 25.29 (ig/g) bone Pb levels had higher odds
32 of Parkinson's disease compared to those in the lowest quartiles (0-5.91 (ig/g for tibia and
33 0-11.70 (ig/g for calcaneus). Subjects in the highest quartile of whole-body lifetime Pb
34 level (> 80.81 (ig/g, estimated using PBPK modeling) had the highest OR: 2.27 (95% CI:
35 1.13, 4.55) versus the lowest quartile, 0-40.04 (ig/g. These results were adjusted for age,
36 race, sex, pack-years smoking, regular coffee consumption, and regular alcohol use, but
November 2012 5-204 Draft - Do Not Cite or Quote
-------
1 Mn exposure was not considered. It was not clear what the extent of occupational
2 exposure to Pb was among the participants; however, a previous Henry Ford Health
3 System study had linked occupational Pb exposure to Parkinson's disease (Gorell et al.,
4 1997). Thus, it is uncertain whether the observed associations were confounded by
5 co-occurring Mn exposure.
6 In summary, a small number of recent case-control studies expand on previous evidence
7 by finding associations of tibia and calcaneus bone Pb levels, biomarkers of cumulative
8 Pb exposure, with Parkinson's disease in adults. The associations observed with
9 biomarkers of cumulative Pb exposure increase confidence that associations are not
10 explained by reverse causality. However, firm conclusions are not warranted. While
11 associations were adjusted for potential confounding by age, sex, race, and education, Mn
12 co-exposure was not considered.
5.3.10.4 Essential Tremor
13 The few available case-control studies of essential tremor have found associations with
14 concurrent blood Pb levels. The 2006 Pb AQCD (U.S. EPA. 2006b) described case-
15 control studies that found associations between concurrent blood Pb levels and essential
16 tremor in New York City metropolitan area populations (Louis et al., 2005; Louis et al.,
17 2003). In the larger study, mean (SD) blood Pb levels were 3.3 (2.4) (ig/dL in the 100
18 essential tremor cases and 2.6 (1.6) (ig/dL in the 143 controls (Louis et al., 2003). In the
19 other study, mean (SD) blood Pb levels were 3.5 (2.2) (ig/dL in the 61 essential tremor
20 cases and 2.6 (1.5) (ig/dL in the 101 controls (Louis et al., 2005). In Louis et al. (2005).
21 the magnitude of association was larger among the 35 ALAD-2 carriers than among 129
22 adults with the ALAD-1 genotype. History of occupational Pb exposure was similarly
23 rare in cases and controls (2%).
24 Recently, Dogu et al. (2007) reported on 105 essential tremor cases selected from a
25 movement disorder clinic in Turkey and 105 controls (69 spouses and 36 other relatives)
26 living in the same district. With adjustment for age, sex, education, smoking status,
27 cigarette pack-years, and alcohol use, a 1 ug/dL higher blood Pb level (measured at the
28 time of study recruitment) was associated with essential tremor with an OR of 4.19 (95%
29 CI: 2.59, 6.78). This OR was much larger than that obtained in the New York area study
30 (OR: 1.19 [95% CI: 1.03, 1.37]) (Louis et al.. 2003). The magnitude of association in
31 Dogu et al. (2007) is even more striking because so many of the controls were spouses
32 who are expected to share many environmental exposures as cases. Most of the essential
33 tremor cases were retired at the time of the study; however past occupational history was
34 not examined.
November 2012 5-205 Draft - Do Not Cite or Quote
-------
1 In summary, a small body of studies indicates associations between blood Pb level
2 measured at the time of the study and prevalence of essential tremor in adults. However,
3 because of the case-control design, reverse causation cannot be excluded as a potential
4 explanation for the observed associations since loss of physical activity and subsequent
5 bone resorption may lead to an increase in blood Pb level. Further, the level, timing,
6 frequency, and duration of Pb exposure associated with essential tremor are uncertain.
7 History of occupational Pb exposure was not consistently examined, and potential
8 confounding by Mn exposure was not examined.
5.3.10.5 lexicological Studies of Cell Death Pathways
9 A common element of the neurodegenerative diseases described above is neuronal cell
10 death. Studies reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b) documented that Pb
11 exposure induced cell death or apoptosis in various models including rat brain (Tavakoli-
12 Nezhadetal., 2001), retinal rod cells (He et al., 2003; He et al.. 2000), cerebellar neurons
13 (Oberto et al.. 1996). and PC12 cells (Sharifi and Mousavi. 2008). Recent studies
14 produced similar findings, in most cases, in animals with higher blood Pb levels than
15 those relevant to humans. Long-term (40 days) exposure to 500 ppm Pb in drinking water
16 was found to increase pro-apoptotic Bax protein levels and the number of apoptotic cells
17 in the hippocampus in young (exposure starting at 2-4 weeks of age) and adult (exposure
18 starting at 12-14 weeks of age) male rats with blood Pb levels 98 (ig/dL (Sharifi et al..
19 2010). Apoptosis was verified by light and electron microscopy. Another study followed
20 the developmental profile of changes in various apoptotic factors in specific brain regions
21 of animals exposed to 2,000 ppm Pb acetate during lactation (to PND20) via drinking
22 water of dams (Chao et al.. 2007). At the end of lactation, male offspring blood Pb level
23 was 80 (ig/dL. The data showed that hippocampal mRNA for various apoptotic factors
24 including caspase-3, Bcl-x, and Brain-derived neurotrophic factor (BDNF) was
25 significantly upregulated on PND12, PND15 and PND20. The cortex of these male pups
26 showed upregulation of Bcl-x and BDNF on PND15 and PND20. The cerebellum did not
27 have elevated apoptotic mRNA levels in this model. Thus, in this study, Pb-induced
28 apoptosis varied by age and brain region in male offspring.
29 Pb exposure also has been shown to induce apoptosis of spinal cord cells during spinal
30 cord development in chicks treated with 150 or 450 (ig Pb acetate in ovo at
31 embryonic day 3 or 5 and visualized six days later (Muller et al.. 2012). TUNEL positive
32 cells, indicating DNA fragmentation induced by apoptosis, were at significantly higher
33 levels in Pb-exposed animals and were visualized in all layers of the developing spinal
34 cords. Also, levels of glial fibrillary acidic protein (GFAP), a factor important in neuronal
35 migration and cellular differentiation during nervous system development, was
November 2012 5-206 Draft - Do Not Cite or Quote
-------
1 significantly attenuated in spinal cords of Pb-exposed chicks. Liu et al. (201 Ob) examined
2 apoptotic effects in 30 day-old male rats that were treated with Pb acetate once daily for
3 6 weeks via intragastric infusion. Four groups: control, low (2 mg/kg BW), medium (20
4 mg/kg BW), and high (200 mg/kg BW) had blood Pb levels of 1.0-7.5 ug/dL;
5 4.5-11 ug/dL; 9-42 ug/dL; and 48-73 ug/dL, respectively. Pb induced hippocampal
6 neuronal apoptosis (TUNEL positive staining, statistically significant at all Pb doses)
7 with hippocampal XIAP (significant at high dose only) and Smac (statistically
8 nonsignificant trend) downregulation at the termination of the 6 week treatment. In
9 another study, Pb exposure (500 ppm Pb acetate in drinking water for 8 weeks) of adult
10 male rats induced regional-specific changes in brain apoptotic proteins poly(ADP-ribose)
11 polymerase, Bel-2, and caspase-3 with a greater effect observed in the hippocampus and
12 cerebellum and a lesser effect observed in the brainstem and the frontal cortex (Kiran
13 Kumar etal.. 2009).
14 In summary, a small body of epidemiologic studies found Pb-associated increases in
15 essential tremor and Parkinson's Disease in adults. However, limitations such as the
16 potential for reverse causation to explain cross-sectional associations observed with blood
17 Pb level, and the potential for confounding by Mn exposure preclude firm conclusions.
18 However, toxicological evidence supports an effect of Pb on neurodegeneration by
19 demonstrating that Pb exposure during various lifestages, early postnatal or adulthood,
20 induces neuronal apoptosis in animals. Several of these observations were made with
21 routes of Pb exposure (i.e., i.p.) that may not be relevant to those in humans.
5.3.11 Modes of Action for Pb Nervous System Effects
5.3.11.1 Effects on Brain Physiology and Activity
22 The 2006 Pb AQCD (U.S. EPA. 2006b) reviewed a small body of available
23 epidemiologic studies demonstrating associations of Pb biomarkers with
24 electrophysiologic and physical changes in the brains of young adults (Yuan et al.. 2006;
25 Cecil et al.. 2005) and children (Meng etal.. 2005; Trope etal.. 2001) as assessed by
26 magnetic resonance imaging (MRI) or spectroscopy (MRS). The implications of previous
27 findings were limited by the small sample sizes (n = 12-45) and limited consideration for
28 potential confounding. Recent studies examining MRI data were limited largely to the
29 Cincinnati cohort as adults (ages 19-24 years). In addition to supporting associations of
30 childhood blood Pb levels with physiological changes in the brain of adults, these recent
31 analyses expanded on previous studies by including larger sample sizes, aiming to
32 characterize important lifestages of Pb exposures, and evaluating potential links between
November 2012 5-207 Draft - Do Not Cite or Quote
-------
1 changes in brain activity and functional neurodevelopmental effects. While there are
2 overall few studies in few populations, by showing physical and physiologic changes in
3 areas of the brain associated with neurodevelopmental function, the evidence provides
4 biological plausibility for the associations observed between Pb biomarker levels and
5 cognitive function and behavior.
6 In prospective analyses of the Cincinnati cohort as adults (ages 20-23 years, n = 35, 42),
7 Cecil et al. (2005) and Yuan et al. (2006) conducted functional MRI during a verb
8 generation language task and found that higher age 3-78 month average blood Pb level
9 (mean 14.2 (ig/dL) was associated with decreased activation in the left frontal gyrus and
10 left middle temporal gyrus, regions implicated in semantic language function. Yuan et al.
11 (2006) considered birth weight, marijuana consumption, sex, SES, gestational age, and
12 IQ as potential confounding factors. Whereas previous analyses of the Cincinnati cohort
13 focused on activity in specific regions of the brain, Cecil et al. (2011) examined brain
14 metabolites. Higher age 3-78 month average blood Pb levels (mean: 13.3 (ig/dL) were
15 associated with lower levels of N-acetylaspartate (NAA) and creatine (Cr) in the basal
16 ganglia and lower levels of choline in white matter in 159 adults, ages 19-23 years. These
17 results were adjusted for age and FSIQ; however, several other unspecified factors were
18 considered. Lower levels of NAA, Cr, and choline are linked to decreased neuronal
19 density and alteration in myelin. A recent prospective analysis of 31 men in the NAS
20 cohort similarly reported an association between biomarkers of cumulative, long-term Pb
21 exposure and changes in brain metabolites in older adults. Weisskopf et al. (2007b) found
22 higher tibia and patella Pb levels to be associated with a higher myoinositol/Cr ratio in
23 the hippocampus measured more than 10 years after bone Pb and adjusted for age.
24 Myoinositol/Cr ratio may be indicative of glial activation and is a signal reportedly found
25 in the early stages of HIV-related dementia and Alzheimer's disease.
26 Other studies in the Cincinnati cohort as young adults found that childhood average blood
27 Pb levels were associated with altered brain architecture. Among 91 adults ages 20-26
28 years, Brubaker et al. (2009) found associations of age 3-78 month average blood Pb
29 levels (mean: 13.3 (ig/dL) with diffusion parameters that were indicative of less
30 organization of fibers throughout white matter. Results were adjusted for maternal IQ,
31 prenatal alcohol and tobacco exposure, and adult marijuana use. In regions of the corona
32 radiata, higher blood Pb levels were associated with less myelination axonal integrity. In
33 regions of the corpus callosum, higher blood Pb levels were associated with greater
34 myelination and axonal integrity. The differential impact among neural elements may be
35 related to the stage of myelination development present at various time periods.
36 Another study of 157 Cincinnati cohort adults ages 19-24 years provided evidence of
37 region-specific reductions in gray matter volume in association with age 3-78 month
November 2012 5-208 Draft - Do Not Cite or Quote
-------
1 average blood Pb levels (mean: 13.3 (ig/dL) with adjustment for sex (Cecil et al., 2008).
2 The most affected regions included frontal gray matter, specifically the anterior cingulate
3 cortex, and the ventrolateral prefrontal cortex (i.e., areas related to executive functions,
4 mood regulation, and decision-making). Further, fine motor factor scores were positively
5 correlated with gray matter volume in the cerebellar hemispheres; adding blood Pb level
6 as a variable to the model attenuated this correlation. These findings suggested that
7 changes observed with MRI may mediate the association between blood Pb levels and
8 decrements in motor function. The functional relevance of these structural changes in the
9 brain also is supported by observations from other studies that link changes in brain
10 architecture and activity to changes in cognitive function (e.g., visuoconstruction, visual
11 memory, eye-hand coordination) (Schwartz et al.. 2007) and behavioral problems
12 (impulsivity, aggression, violence) (Yang et al.. 2005; Raine et al., 2000). In a subsequent
13 comparison of blood Pb levels measured at various lifestages in 157 Cincinnati cohort
14 adults ages 19-24 years, Brubaker et al. (2010) found that blood Pb levels at older ages
15 (annual means from 3-6 years, means: 9.6-16.3 (ig/dL) were associated with greater
16 losses in gray matter volume than were age 3-78 month average or maximum blood Pb
17 levels (mean: 23.1 (ig/dL). Both Cecil et al. (2008) and Brubaker et al. (2010) found that
18 Pb-associated reductions in gray matter were more pronounced in males than females in
19 the Cincinnati cohort.
20 Studies of Pb-exposed workers (n = 15-532) also found associations of concurrent blood
21 (means: 17-63.5 (ig/dL) and tibia (mean 14.5 (ig/dL) Pb levels with changes in brain
22 structure and physiology, supporting the effects of chronic Pb exposure. Pb-associated
23 changes included white matter lesions, smaller brain volumes, less total gray matter, and
24 lower levels of brain metabolites such as NAA and Cr (Hsieh et al.. 2009b: Jiang et al..
25 2008; Bleecker et al.. 2007b; Stewart et al., 2006) with adjustment for similar factors as
26 associations for cognitive function. Other occupational exposures were not examined. In
27 a few of these occupational groups, Pb-associated brain changes were linked to poorer
28 performance in cognitive function tests (Caffo et al.. 2008; Bleecker et al.. 2007b).
29 Higher concurrent blood Pb level also was associated with lower NAA/Cr ratio in small
30 cross-sectional studies that included children (n = 6, 16, ages 4-21 years), although
31 neither study considered potential confounding (Meng et al.. 2005; Trope et al.. 2001).
32 All subjects had normal MRIs with no evidence of structural abnormalities. Thus, the
33 biological relevance of the observed physiological changes is unclear. Additionally, the
34 representativeness of findings is uncertain because results were based on comparisons of
35 subjects with relatively high blood levels (23-65 (ig/dL) to those with blood Pb levels
36 <10 (ig/dL.
November 2012 5-209 Draft - Do Not Cite or Quote
-------
1 In summary, results in a few populations indicate associations of childhood blood or adult
2 tibia Pb levels with changes in brain structure and physiology in adults assessed by MRI
3 or MRS. Associations were found in children, but implications are limited because of
4 small samples sizes, lack of consideration of potential confounding, and high blood Pb
5 levels of the children examined. Evidence from the prospective Cincinnati cohort studies
6 improves characterization of the temporal sequence between Pb exposure and changes in
7 brain structure and physiology. Several studies linked these changes to functional
8 changes in cognitive performance or motor skills. Because of the small samples sizes of
9 several studies and limited consideration for potential confounding, firm conclusion
10 regarding the effects of Pb exposed on changes in brain structure and physiology is not
11 warranted. However, the evidence provides biological plausibility for the associations
12 observed between Pb biomarker levels and cognitive function and behavioral problems.
5.3.11.2 Oxidative Stress
13 Because the brain has the highest energy demand and metabolism of any organ, energy
14 homeostasis is of utmost importance. Energy imbalance can increase the susceptibility of
15 the highly energetic brain tissue to stressors and cell death. Pb has been shown to induce
16 energy imbalance by inhibiting various enzymes involved in energy production or
17 glucose metabolism including glyceraldehydes-3 phosphate dehydrogenase, hexokinase,
18 pyruvate kinase, and succinate dehydrogenase (Verma et al.. 2005; Yun and Hover. 2000;
19 Regunathan and Sundaresan. 1984; Sterling et al., 1982). Mitochondria produce ATP or
20 energy through oxidative phosphorylation. Aberrant mitochondrial function can decrease
21 the energy pool and contribute to ROS formation via electron transport chain disruption.
22 ATP depletion can also affect synaptic and extracellular neurotransmission. The
23 mitochondrial Na+/K+ATPase is important in maintaining the inner mitochondrial
24 membrane potential A*Pm (delta psifsub m]) and the functioning of the mitochondria.
25 Gestational Pb exposure was found to impair mitochondrial function and energy
26 production in neuronal cells from mice and produce concomitant increases in
27 mitochondrial and cellular ROS production. The effect of Pb exposure on these
28 mitochondrial parameters were examined in the brains of mice after prenatal Pb exposure
29 (1,000 ppm Pb acetate in dam drinking water, resulting in offspring blood Pb levels of
30 4 (ig/dL and cerebella Pb levels of 7.2 (ig/g dry weight) (Baranowska-Bosiacka et al.,
31 201 Ib). Cerebellar granular cells were harvested from control and Pb-exposed animals at
32 PND8. These neuronal cells were cultured for 5 days in vitro, at which point various
33 mitochondrial parameters were measured. With Pb exposure, ROS were significantly
34 increased in both the cortical granule cells and in the mitochondria. Intracellular ATP
35 concentration and adenylate energy charge values were significantly decreased in cells of
November 2012 5-210 Draft - Do Not Cite or Quote
-------
1 Pb-exposed mice versus controls. Neuronal Na+/K+ATPase activity was significantly
2 lower in cortical granule cells from Pb-exposed mice versus cells from controls.
3 Mitochondrial mass was unaffected with Pb treatment, but mitochondrial membrane
4 potential was significantly decreased with Pb exposure. Energy imbalances also were
5 found in Wistar rats (PND15) of each sex injected daily for 2 weeks with Pb acetate
6 (15mg/kg BW, i.p., resulting in a mean blood Pb level of 30 (ig/dL; control blood Pb
7 level 3 (ig/dL) (Baranowska-Bosiacka et al., 201 la). ATP and ADP were significantly
8 decreased in various brain regions with Pb exposure, with the cerebellum and
9 hippocampus more strongly affected than the forebrain cortex. Also, expression of the
10 pro-inflammatory P2XR receptor was enhanced in the glial fraction, indicating the
11 astrocyte pool may be involved in the pathological changes found in Pb-exposed
12 immature rat brains. Mitochondrial energy imbalances also were found in Pb-exposed
13 crayfish that were placed under hypoxic conditions which induced a decrease in
14 metabolism (Morris et al., 2005).
15 In rats, Pb exposure has been shown to induce oxidative stress, in some cases, with
16 concomitant functional CNS changes. Exposure of adult male rats to 4,000 ppm
17 Pb acetate in drinking water for 6 weeks increased brain levels of lipid peroxides (LPO)
18 and lowered levels of antioxidants including nitric oxide (NO), total antioxidant capacity
19 (TAC), glutathione (GSH), glutathione-S-transferase (GST), and superoxide dismutase
20 (SOD). Whole blood Pb levels were positively correlated with brain tissue LPO levels
21 and negatively correlated with NO levels. Evidence also indicated a role for oxidative
22 stress in mediating the effects of Pb on cognition as evidenced by changes in synaptic
23 plasticity (Hamed et al., 2010). These effects of Pb on oxidative stress parameters were
24 attenuated with co-exposure to green tea extract (1.5%), which reduced brain (1.9 to
25 1.2 ppm) and blood Pb levels of rats (0.773 to 0.654 ppm). In a study of adult male
26 Wistar albino rats, Pb acetate treatment by i.p. (20 mg/kg, 5 days) elevated lipid
27 peroxidation, neuronal damage, and brain tissue DNA fragmentation and decreased
28 antioxidant GSH levels and antioxidant enzyme activity, (Abdel Moneim et al.. 201 la).
29 These effects were attenuated with co-administration of the polyunsaturated fatty acid
30 flaxseed oil (oral gavage l,000mg/kg body weight for 5 days, 1 hour prior to Pb dosing).
31 Flaxseed oil co-treatment also significantly attenuated the blood Pb level of Pb-exposed
32 animals (~31 (ig/dL the day after the last Pb injection to -12 (ig/dL) and control animals,
33 indicating that flaxseed oil may alter Pb toxicokinetics in animals. Another study
34 provided indirect evidence of Pb-induced oxidative stress with observations that
35 Pb-induced (2,000 ppm Pb acetate in drinking water PND1-PND67) impairments in long-
36 term potentiation (LTP), paired-pulse reactions, and input/output functions in the DG of
37 male and female Wistar rats were significantly attenuated with treatment with the
38 antioxidant quercetin (30 mg/kg BW, PND60-PND67) at PND67 (Hu et al.. 2008a).
November 2012 5-211 Draft - Do Not Cite or Quote
-------
1 Quercetin-treated animals had significantly less hippocampal Pb than did the animals
2 exposed only to Pb.
3 Oxidative stress may be involved in neurodegenerative pathologies including Alzheimer's
4 disease. Hydrogen peroxide-induced oxidative stress has been shown to induce
5 intracellular accumulation of Ab in human neuroblastoma cells (Misonou et al.. 2000).
6 Oxidative stress-induced DNA damage can be measured as the ratio of the adduct
7 8-hydroxy-2'-deoxyguanosine to 2-deoxyguanosine (8-oxo-dG/2-dG). 2-dG is a product
8 of oxidative cleavage and is oxidized to form 8-oxo-dG. Pb-induced changes in the
9 8-oxo-dG to 2-dG ratio were examined recently as a mechanism underlying
10 neurodegeneration. Similar to Ab levels, changes in the 8-oxo-dG to 2-dG ratio showed a
11 biphasic relationship in the brains of rats exposed to 2,000 ppm Pb acetate via drinking
12 water of dams from PND1-PND20 (blood Pb level 46 ng/dL) (Bolin et al.. 2006). The
13 8-oxo-dG to 2-dG ratio decreased early in exposure (PND5) but increased at age
14 20 months. No increase was found in animals exposed to Pb from age 18 to 20 months
15 (blood Pb level: 60 (ig/dL). Activity of the base-excision DNA repair enzyme oxoguanine
16 glycosylase was unaffected by Pb exposure. Similar findings were reported in a monkey
17 study (Wu et al., 2008a). The ratio of 8-oxo-dG to 2-dG in the brains of aged monkeys
18 (23 years) was significantly elevated above that in controls only with Pb exposure in
19 infancy (PND1-PND400, infant formula, blood Pb levels: 19-26 (ig/dL) but not as aged
20 adults (Wu et al.. 2008a). Thus, evidence in rats and monkeys suggests a possible role for
21 oxidative stress in Pb-induced neurodegenerative effects and indicates that early life but
22 not adult-only Pb exposure induces oxidative DNA damage and amyloidogenesis.
5.3.11.3 Nitrosative Signaling and Nitrosative Stress
23 The NO system is increasingly being recognized as a signaling system in addition to its
24 more classical role as a marker of cellular stress. Pb exposures during the gestational-
25 early postnatal period (GD6-PND21) (Chetty etal.. 2001) and during the postnatal period
26 only (Fan et al.. 2009a) were found to reduce hippocampal levels of NO or neuronal NO
27 synthase. In the hippocampus, NO mediates LTP, which is considered to be a major
28 cellular mechanism underlying learning and memory. Thus, observations of Pb-induced
29 changes in hippocampal NO may provide a mechanistic explanation for the effects of Pb
30 on cognitive function decrements. Fan et al. (2009a) found reduced hippocampal NOS
31 and NO in weanling male rats after either 4 or 8 weeks of Pb exposure resulting in blood
32 Pb levels of 47 and 66 (ig/dL, respectively. In the same study, dietary supplementation
33 with taurine or glycine concomitant with 8 weeks of Pb exposure induced significant
34 increases in hippocampal NOS, whereas Pb plus dietary supplementation with vitamin C,
35 methionine, tyrosine, or vitamin Bl decreased hippocampal NOS. In this study,
November 2012 5-212 Draft - Do Not Cite or Quote
-------
1 co-exposure of specific nutrients also prevented Pb-induced impairments in learning as
2 evidenced by lack of increased escape latency in the Morris water maze. Dietary
3 supplementation with tyrosine, methionine, or ascorbic acid after 4 weeks of Pb exposure
4 in weanling males (blood Pb levels upon cessation of exposure and after 4-week lag:
5 47.6 and 8.1 (ig/dL, respectively) reversed Pb-induced decrements in NO/NOS. Zn
6 supplementation given after Pb exposure had no effect on the NO system.
5.3.11.4 Synaptic Changes
7 Previous toxicological studies pointed to an effect of developmental Pb exposure on
8 synapse development, which mechanistically may contribute to multiple Pb-related
9 aberrant effects, including changes in LTP and facilitation. Facilitation of a neuronal
10 terminal is defined as the increased capability to transmit an impulse down a nerve due to
11 prior excitation of the nerve. Earlier work showed that developmental Pb exposure
12 resulted in altered density of dendritic hippocampal spines (Kiraly and Jones. 1982; Petit
13 and Leboutillier. 1979). aberrant synapse elimination (Lohmann and Bonhoeffer. 2008).
14 and abnormal long-term and short-term plasticity (MacDonald et al.. 2006). In a recent
15 study, Li et al. (2009c) focused on inflammatory endpoints and synaptic changes after
16 gestational plus lactational Pb exposure (1,000-10,000 ppm Pb acetate via drinking water
17 of dams, producing offspring blood Pb levels 40-100 (ig/dL, respectively at PND21).
18 Hippocampal TNF-a was significantly elevated with Pb exposure, and proteins that
19 comprise the SNARE complex were all changed with Pb exposure. The SNARE complex
20 of synaptic proteins includes SNAP-25, VAMP-2 and Syntaxin la and is essential in
21 exocytotic neurotransmitter release at the synapse. Thus, Li et al. (2009c) found
22 significant differences in hippocampal synaptic protein composition and increased pro-
23 inflammatory cytokine levels in the brains of Pb-exposed offspring.
24 Recent research using the Drosophila larval neuromuscular junction model showed that
25 compared with unexposed controls, Pb-exposed larvae had significant increases in
26 intracellular calcium and significant delays in calcium decays back to baseline levels at
27 the pre-synaptic neuronal bouton (as stimulated with multiple action potentials, also
28 called AP trains). Pb-exposed larvae had reduced activity of the plasma membrane
29 Ca2+ATPase, which is responsible for extravasations of calcium from the synaptic
30 terminal (He et al.. 2009). Intracellular calcium in Pb-exposed larvae was no different
31 from that in controls under resting conditions or in neurons with stimulation by a single
32 action potential. Pb media concentrations in these experiments were 100 or 250 (iM with
33 the body burden of Pb from the lower dose calculated to be 13-48 (iM per larvae. After
34 stimulation of the axon, facilitation of the excitatory post-synaptic potential, which is
35 dependent on residual terminal calcium, was significantly elevated in Pb-exposed larvae
November 2012 5-213 Draft - Do Not Cite or Quote
-------
1 versus control (He et al., 2009). The data from this synapse study demonstrate that
2 developmental Pb exposure affected the plasma membrane Ca2+ATPase, induced changes
3 in the intracellular calcium levels during impulse activation, and produced changes in
4 facilitation of the neuronal networks of Drosophila. Thus, the neuromuscular junction is a
5 potential site of Pb interaction.
6 Neurotransmission is an energy-dependent process as indicated by the presence of
7 calcium-dependent ATP releases at the synaptic cleft. At the synapse, ATP is
8 metabolized by ectonucleotidases. Acute exposure (96 hours) of male and female
9 zebrafish to Pb acetate (2 (ig/dL) in their water induced significant decreases in ATP
10 hydrolysis in brain tissue (Senger et al.. 2006). This dose is deemed to be
11 environmentally relevant. With chronic exposure (30 days), Pb acetate promoted the
12 inhibition of ATP, ADP and AMP hydrolysis. These findings were consistent with
13 findings in rodents (Baranowska-Bosiacka et al.. 201 la). The authors hypothesized that at
14 30 days, this Pb-induced change in nucleotide hydrolysis was likely due to post-
15 translational modification because expression of enzymes responsible for the hydrolysis,
16 NTPDasel and 5'-nucleotidase, were unchanged (Senger et al.. 2006). Thus, Pb has been
17 shown to affect nucleotidase activity in the CNS of zebrafish, possibly contributing to
18 aberrant neurotransmission.
19 Another enzyme important in synaptic transmission at cholinergic junctions in the CNS
20 and at neuromuscular junctions is acetylcholinesterase (AChE). After 24 hours of
21 exposure to Pb acetate (2 (ig/dL water), AChE activity was significantly inhibited in
22 zebrafish brain tissue (Richetti et al., 2010). AChE activity returned to baseline by 96
23 hours and maintained baseline activity after 30 days of exposure. Thus, Pb was shown to
24 affect synaptic homeostasis of AChE in the brains of zebrafish only transiently.
25 Pb has been shown to act as an antagonist of the NMDA receptor (NMDAR). The
26 NMDAR is essential for proper presynaptic neuronal activity and function. Primary
27 cultures of mouse hippocampal cells exposed to 10 or 100 (iM Pb during the period of
28 synaptogenesis had loss of two proteins necessary for presynaptic vesicular release,
29 synaptophysin (Syn) and synaptobrevin (Syb) but no change in a similar protein
30 synaptotagmin (Syt) (Neal etal. 2010a). This deficit was found in both GABAergic and
31 glutamatergic neurons. Pb also induced an increase in the number of presynaptic contact
32 sites. But, these sites may have been nonfunctional as they lacked the protein receptor
33 complexes necessary for proper vesicular exocytosis. Another factor involved in
34 maturation and signaling of presynaptic neurons is brain-derived neutrotrophic factor
35 (BDNF), which is synthesized and released by postsynaptic neurons regulated by the
36 NMDAR. In hippocampal cells, both pro-BDNF and BDNF release were significantly
37 attenuated with Pb exposure (Neal etal., 2010a). Further, exogenous BDNF
November 2012 5-214 Draft - Do Not Cite or Quote
-------
1 administration rescued the aforementioned Pb-related effects on presynaptic proteins.
2 Thus, this cell culture model showed that Pb-related presynaptic aberrations are
3 controlled by NMDAR-dependent BDNF effects on synaptic transmission.
4 Glutamate is another neurotransmitter that is released from presynaptic neurons and via
5 interactions with the NMDAR causes postsynaptic neuron depolarization. A recent study
6 of Wistar albino rats exposed to Pb postnatally from birth to age 12 weeks (drinking
7 water 3 x 104 (ig/dL Pb acetate, resulting in blood Pb levels of 17 (ig/dL at age 6 weeks)
8 showed decreased learning ability, decreased hippocampal glutamate at 6, 8, 10 and
9 12 weeks of age, as well as significant decrements in the hippocampal glutamate
10 synthesis-related enzymes aspartate aminotransferase and alanine aminotransferase (Niu
11 et al.. 2009).
5.3.11.5 Blood Brain Barrier
12 Two barrier systems exist in the body to separate the brain or the CNS from the blood.
13 These two barriers are the blood brain barrier (BBB) and the blood cerebrospinal fluid
14 barrier (BCB). The BBB, formed by tight junctions at endothelial capillaries forming the
15 zonulae occludens (occludins, claudins, and cytoplasmic proteins), separates the brain
16 from the blood and its oncotic and osmotic forces, allowing for selective transport of
17 materials across this barrier.
18 Pb exposure during various developmental windows has been shown to increase the
19 permeability of the BBB of animals (Dvatlov et al., 1998; Struzvnskaet al.. 1997b;
20 Moorhouse et al.. 1988; Sundstrom et al.. 1985). Possibly due the underdevelopment of
21 the BBB early in life, prenatal and perinatal Pb exposure has been found to result in
22 higher brain Pb accumulation than have similar exposures later in life (Moorhouse et al..
23 1988). The choroid plexus and cerebral endothelial cells that form the BBB and BCB
24 tight junctions have been shown to accumulate Pb more than other cell types and regions
25 of the CNS. Studies reviewed in earlier Pb AQCDs showed that the chemical form of Pb
26 and its capability to interact with proteins and other blood components affects its
27 capability to penetrate the BBB (U.S. EPA. 2006b). Pb also has been shown to
28 compromise the BCB and decrease the cerebrospinal fluid level of transthyretin, which
29 binds thyroid hormone in the cerebrospinal fluid. Low thyroid hormone levels in
30 pregnant women have been linked with IQ deficits in their children (Lazarus. 2005).
31 Recent research with male weanling rats exposed to Pb acetate via drinking water showed
32 leaky cerebral vasculature, an indication of a compromised BBB, as detected
33 histologically with lanthanum nitrate staining of the brain parenchyma (Wang et al..
34 2007b). Cerebral vasculature leakiness was ameliorated or resembled that of controls
November 2012 5-215 Draft - Do Not Cite or Quote
-------
1 after iron supplementation. The cerebral vasculature leakiness may by explained by
2 observations of significant Pb-induced decreases in the BBB tight junction protein
3 occludin in the hippocampus, brain cortex, and cerebellum in these weanling animals.
4 Occulin levels were rescued to control levels with iron supplementation. This loss of
5 integrity at the junctional protein level was affirmed with additional experiments using
6 the rat brain vascular endothelial cell line RBE4, in which 10 (iM Pb acetate exposure for
7 2, 4, 8, 16 and 24 hours resulted in decreases in junctional proteins occludin and claudin
8 5 as well as scaffold proteins ZO1 and ZO2 (Balbuena et al.. 2011). Because gene
9 expression for these junctional and scaffold proteins did not show decrements, it was
10 determined that these protein decrements were due to post-translational modifications.
11 Pb exposure also was found to contribute to leakiness of the BBB by decreasing the
12 resistance across the junction (Balbuena et al.. 2010). An in vitro co-culture system
13 employing endothelial cells (RBE4 or bovine brain microvascular endothelial cells) and
14 astrocytes (primary Sprague-Dawley neonatal pup astrocytes, GD21) served as the barrier
15 between Pb-containing media and neurons. Pb acetate exposure (1 and 10 (iM) for 14
16 hours significantly impaired transendothelial electrical resistance (TEER), a marker of
17 BBB integrity, in a concentration-dependent manner.
18 Long-term Pb exposure of adult mice was found to increase regional edema and BBB
19 permeability (Lopez-Larrubia and Cauli. 2011). Adult male rats exposed to Pb acetate in
20 drinking water for 4 or 12 weeks (50 or 500 ppm, resulting in blood Pb levels of 12 and
21 55 (ig/dL, respectively) were assessed by diffusion weighted imaging for changes in
22 apparent diffusion coefficent (ADC), a measure of tissue water diffusivity that changes
23 under pathological conditions like cerebral edema. With 12 weeks of exposure, 50 ppm
24 Pb increased the ADC values in the cerebellum and mesencephalic reticular formation,
25 and 500 ppm Pb exposure significantly increased ADC in the corpus callosum and
26 caudate putamen. With 4 weeks of exposure, 500 ppm Pb significantly increased the
27 water ADC in the hippocampus, mesencephalic reticular formation, and cerebellum but
28 not in other brain areas. The brain areas with elevated ADC also showed increased BBB
29 permeability as measured with evans blue dye.
November 2012 5-216 Draft - Do Not Cite or Quote
-------
5.3.11.6 Cell Adhesion Molecules
1 Classic cell adhesion molecules including neural cell adhesion molecule (NCAM) and the
2 cadherins are junctional or cell surface proteins that are critical for cell recognition and
3 adhesion. While direct effects of Pb on cell adhesion molecules have not been described,
4 the calcium-dependency of these molecules suggests that interaction from competing
5 cations like Pb can potentially contribute to nervous system barrier function disruption,
6 neurite outgrowth, synaptic plasticity, learning and memory (Prozialeck et al., 2002).
5.3.11.7 Effects on Glial Cells
7 Astroglia and oligodendroglia are supporting cells in the nervous system that maintain the
8 extracellular space in the brain and provide structural support to neurons, deliver
9 nutrients to neurons, and promote myelination. Glial cells provide immune surveillance
10 in the brain and contribute to inflammation-mediated pathologies. In Wistar rats, Pb
11 treatment (15 mg/kg of Pb acetate, i.p.) during early postnatal maturation was observed to
12 produce chronic glial activation with inflammation and neurodegeneration (Struzynska et
13 al., 2007). Among the cytokines detected in the brains of these Pb-treated rats were
14 IL-1P, TNF-a and IL-6. Glial cells have been shown to serve as Pb sinks in the
15 developing and mature brain by sequestering Pb (Tiffany-Castiglioni etal.. 1989). This
16 glial sequestration of Pb was accompanied by a decrease in brain glutamine
17 concentrations at doses of 0.25 ±1.0 (iM Pb acetate and a reduction in glutamine
18 synthetase activity in the astroglia; astroglia take up released glutamate and convert it to
19 glutamine. Pb has been shown to induce hypomyelination and demyelination (Coria et
20 al.. 1984) mediated through the oligodendrocytes with younger animals found to be more
21 susceptible to the effects of Pb (Tiffany-Castiglioni etal.. 1989). Pb accumulation in
22 young glial cells may contribute to a lifelong exposure of neurons to Pb as Pb is released
23 from the sink over time. Thus, Pb accumulation in glial cells can contribute to continual
24 damage of surrounding neurons (Holtzman et al.. 1987).
Glial transmitters
25 Evidence indicates that glial transmission is affected with Pb exposure and that the
26 NMDAR may be involved in this aberrant glial transmission. To determine the
27 contribution of the gliotransmitter serine to Pb-mediated changes in LTP, Sun et al.
28 (2007) exposed rats to Pb acetate from gestation through lactation to PND28 via maternal
29 drinking water and collected hippocampal sections. CA1 section LTPs were examined
30 using in vitro patch clamp monitoring. Chronic Pb exposure impaired the magnitude of
November 2012 5-217 Draft - Do Not Cite or Quote
-------
1 hippocampal LTPs, but the magnitude of long-term depression was restored with
2 supplementation with D-serine (Sun et al.. 2007). which is known to be regulated by the
3 NMDAR (Bear and Malenka. 1994). The use of 7-chlorokynurenic acid, an antagonist of
4 the glycine binding site of the NMDAR, which also is the binding site of D-serine,
5 effectively abolished the rescue of LTP by D-serine. NMDAR-independent LTP
6 hippocampal neurotransmission was inhibited in slices of Pb-exposed mossy-CA3
7 synapses and was not rescued by exogenous D-serine supplementation.
5.3.11.8 Neurotransmitters
8 Pb has been shown to compete with calcium for common binding sites and second
9 messenger activation. When Pb activates a calcium-dependent system in the nervous
10 system, it can contribute to aberrant neurotransmitter regulation and release because this
11 system intimately relies on calcium signaling for its homeostasis. Pb also has been shown
12 to interfere with other physiological divalent cations. Pb-related alterations in
13 neurotransmission are discussed in further detail below.
Monoamine Neurotransmitters and Stress
14 The monoamine neurotransmitters include dopamine (DA), serotonin (5HT), and
15 norepinephrine (NE). Combined exposures of maternal stress and Pb exposure can
16 synergistically enhance neurobehavioral impairments in offspring of exposed animals and
17 can sometimes potentiate an effect that would otherwise be sub-threshold. Virgolini et al.
18 (2008a) found enhanced DA and NE release in male rats and enhanced NE release in
19 female rats after developmental Pb exposure (50 or 150 ppm via drinking water,
20 2 months prior to mating through lactation, resulting in blood Pb levels of 11 (ig/dL and
21 35 (ig/dL, respectively) and combined maternal and offspring stress. In most cases, stress
22 potentiated the effects of Pb exposure on offspring NE and DA concentrations. Regional
23 5HT levels were unaffected in offspring with Pb exposure alone. Pb (50 and 150 ppm)
24 combined with stress (maternal and/or offspring stress) significantly potentiated 5HT
25 levels in the frontal cortex in females and in the nucleus accumbens (NAC) and striatum
26 in male offspring. The concentration of 5-Hydroxyindoleacetic acid (5HIAA), the main
27 metabolite of 5HT, was significantly increased in the striatum of male offspring with
28 150 ppm Pb exposure alone. With 50 ppm Pb, stress potentiated striatal and frontal cortex
29 5HIAA in males. Potentiation of 5HIAA levels in females was significant in the NAC
30 with 50 ppm Pb exposure; stress alone also significantly increased 5HIAA levels in the
31 NAC of females with no Pb exposure. Pb-induced changes in brain neurochemistry with
November 2012 5-218 Draft - Do Not Cite or Quote
-------
1 or without concomitant stress exposure are complex with differences varying by brain
2 region, neurotransmitter type, and sex of the animal.
Monoamine Neurotransmitters and Auditory Function
3 Earlier work showed that perinatal Pb exposure of rats induced increased tyrosine
4 hydroxylase, increased DA, and increased cerebral cortex catecholamine
5 neurotransmission (Devi et al.. 2005; Leret et al.. 2002; Bielarczyk et al.. 1996). Earlier
6 publications detailing important time windows, durations, and doses of Pb exposure
7 indicated varying effects on monoamine neurotransmitters. In recent work, these
8 neurotransmitters, among others, have been implicated in Pb effects on auditory function
9 in various integration centers of the brainstem including the lateral superior olive (LSO)
10 and the superior olivary complex (SOC). Among various functions, the SOC is vital for
11 sound detection in noisy settings. A recent study in mice found significant decreases in
12 immunostaining of LSO and SOC brainstem sections for monoamine vesicular
13 transporter VMAT2, 5HT, and dopamine beta-hydroxylase (DbH, a marker for NE) after
14 gestational-lactational Pb exposure (10 or 100 (iM Pb acetate from the formation of
15 breeding pairs to PND21). This exposure period corresponds to the period of auditory
16 development in the mouse. Statistically significant decreases in VMAT2 and DbH were
17 found in mice with blood Pb levels of 8.0 and 42.2 (ig/dL; however, decrements in 5HT
18 were statistically significant only in mice with 8.0 (ig/dL blood Pb level. Immunostaining
19 for tyrosine hydroxylase and transporters including VGLUT1, VGAT, VAChAT
20 indicated that they were unaffected by developmental Pb exposure. These data provide
21 evidence that specific regions of the brainstem involved in auditory integration are
22 affected by developmental Pb exposure via effects on monoamine neurotransmitters
23 (Fortune and Lurie. 2009). The Pb-induced effects on the monoamine system of the
24 auditory portion of the brainstem provide possible mechanistic explanation for the
25 epidemiologic and toxicological evidence for Pb-associated decrements in auditory
26 processing (Section 5.3.7).
Dopamine
27 The 2006 Pb AQCD (U.S. EPA. 2006b) detailed evidence for Pb-related decreased
28 dopaminergic cell activity in the substantia nigra and ventral segmental areas. Earlier
29 studies with postnatal or adult Pb exposure reported changes in DA metabolism, as
30 indicated by changes in DA and DOPAC, a DA metabolite. Expanding upon these
31 findings, a recent study measured DA and DOPAC in various brain regions of year-old
32 male C57BL/6 mice to examine if gestational plus lactational Pb exposure affected DA
33 metabolism (Leasure et al.. 2008). Exposure of males to 27 and 109 ppm Pb acetate
November 2012 5-219 Draft - Do Not Cite or Quote
-------
1 induced significant elevations in DOPAC concentration and the DOPAC to DA ratio in
2 the forebrain. In the forebrain, DA was significantly decreased with the lower dose and
3 significantly elevated with the higher dose compared to controls. In the striatum, DOPAC
4 was significantly elevated with both doses, but DA concentration was only significantly
5 elevated with the higher dose. The striatum ratio of DOPAC to DA was not significantly
6 different from that in controls. These recent data expand upon the monoamine literature
7 base which indicates that Pb exposure of rats during the gestational/lactational,
8 lactational, or postweaning period producing blood Pb levels 9-34 (ig/dL induces
9 increased sensitivity of the dopamine receptors (D2 and D3) (Gedeon etal. 2001; Cory-
10 Slechtaetal.. 1992). produces higher DA levels (Devi etal.. 2005; Leret et al.. 2002). and
11 enhances catecholamine neurotransmission in the cerebral cortex, cerebellum, and
12 hippocampus (Devi et al.. 2005).
13 The interaction of DA and the NO system in the striatum was studied after prenatal Pb
14 exposure (Nowak et al.. 2008). Blood Pb levels were not reported in this study, but
15 similarly treated Wistar rat pups had blood Pb levels at parturition in range of
16 50-100 (ig/dL (Grant etal. 1980). 7-nitroinidazole (7-NI), a selective inhibitor of nNOS,
17 enhanced amphetamine-evoked DA release in the rat striatum (Nowak et al.. 2008).
18 Prenatal Pb exposure attenuated the facilitatory effect of 7-NI on DA release in the
19 striatum. This interaction was ROS-independent; using spin trap measurements,
20 investigators found no significant concentration changes in hydroxyl radical with Pb
21 exposure (Nowak et al.. 2008). Thus, the neuronal NO system appears to be involved in
22 specific aspects of Pb-related dopaminergic changes.
23 In various animal models, the loss of retinal DA, dopamine turnover (DOPAC:DA ratio),
24 or Zn was associated with abnormal rod-mediated scotopic ERGs. These effects may
25 explain observations of Pb-associated subnormal or supernormal retinal ERGs observed
26 in animals and children (Rothenberg et al., 2002b; Lilienthal etal.. 1994; Lilienthal et al..
27 1988; Alexander and Fishman. 1984) (Sections 5.3.7.1 and 5.3.7.3). although the
28 biological relevance of the variable effects of Pb exposure on subnormal versus
29 supernormal ERGs is not clear.
NMDA Receptors
30 The glutamate receptor, NMDAR, has been shown to contribute to synaptic plasticity,
31 and Pb exposure at different developmental stages has been shown to contribute to
32 aberrations in LTP or long term depression (LTD) in the hippocampus via reduced
33 NMDA current, among other mechanisms (Liu et al.. 2004). The 2006 Pb AQCD (U.S.
34 EPA. 2006b) indicated that Pb attenuated the stimulation of glutamate release, which in
35 turn, affected LTP. Further, the effects of Pb exposure on decreasing the magnitude of
November 2012 5-220 Draft - Do Not Cite or Quote
-------
1 LTP and increasing the threshold of the LTP in the hippocampus were found to be
2 biphasic or nonlinear. NMDAR subtypes have been shown to be significantly decreased
3 with developmental Pb exposure (Guilarte and McGlothan, 1998). Recent evidence
4 indicated Pb-related decreases in the gene expression and protein level of NMDAR
5 subunits NR1, NR2A, and NR2B in weanling male rats exposed to 4 x 104 (ig/dL
6 Pb acetate in drinking water for 8 weeks. Several of these responses were attenuated with
7 methionine-choline co-exposure (Fan et al., 2010). Other recent mechanistic studies
8 found that pretreatment of primary fetal brain neuronal rat cultures with glutamic acid, a
9 NMDAR agonist, reversed Pb-induced reductions in NMDAR subunits (Xu and Raj anna.
10 2006) whereas pretreatment with the NMDA antagonist MK-801 exacerbated Pb-induced
11 NMDAR deficits (Xu and Rajanna. 2006). Further strengthening the link among Pb
12 exposure, NMDAR function, and learning, Guilarte et al. (2003) demonstrated that rats
13 exposed to 1,500 ppm Pb during gestation and lactation then reared in isolation, had
14 reduced expression of hippocampal NMDA receptor subunit 1, reduced induction of
15 BDNF mRNA, and learning impairment. These effects were attenuated in Pb-exposed
16 rats reared in an enriched environment with toys.
Other Glutamate Receptors
17 The metabotropic glutamate receptor (mGluR) is another well-recognized target of Pb
18 toxicity. In vitro (GD18 fetal rat cultures, 0.01, 100, 1 (iM Pb chloride in culture media)
19 and in vivo studies (gestational and lactational Pb acetate exposure; 500, 2,000,
20 5,000 ppm in dam drinking water, with respective weanling blood Pb levels of 18, 57,
21 186 (ig/dL) showed that Pb exposure induced mGluR5 mRNA and protein decrements in
22 a concentration-dependent manner (Xu et al.. 2009c). Recent evidence indicates a role for
23 mGluR5 in synaptic plasticity, LTP, and LTD; thus, the Pb-related attenuation of mGlu5
24 expression may represent a mechanism by which Pb impairs learning and memory.
5.3.11.9 Neurogenesis
25 Studies continue to show that Pb exposure decreases neurogenesis (i.e., proliferation of
26 neuronal cells) in the hippocampus, which is important in LTP, spatial learning, neuronal
27 outgrowth, and possibly mood disorders such as schizophrenia. Coherence for these
28 findings is provided by evidence for Pb-induced decreases in NMDAR, which mediates
29 the integration of new neurons into existing neuronal pathways in the adult hippocampal
30 DG. Earlier work by Schneider et al. (2005) showed that postnatal Pb exposure (PND25
31 to PND50 or PND55, 1,500 ppm Pb acetate in chow, resulting in blood Pb level of
November 2012 5-221 Draft - Do Not Cite or Quote
-------
1 20 (ig/dL) of male Lewis rats induced significant decrements in BrdU incorporation (an
2 indicator of DNA replication) at PND5 0-PND5 5.
3 Recent publications affirm this previous finding with different sex of animals, dosing and
4 exposure time windows. Postnatal Pb exposure of Wistar rat pups from PND1-PND30
5 (2,000 ppm Pb acetate, resulting in blood Pb levels of 34 and 6.5 (ig/dL at PND21 and
6 PND80, respectively) induced a statistically significant decrement in the number of new
7 cells (BrdU positive cells) in the DG at PND80 (Fox etal.. 2010) (Figure 5-14). In
8 another study, lifetime Pb exposure beginning in gestation (1,500 ppm Pb acetate in chow
9 from 10 days before mating to PND50 or PND78, resulting in blood Pb levels 26 (ig/dL)
10 of female Long-Evans rats induced significant decrements in hippocampal granule cell
11 neurogenesis in adult rats (Verina et al., 2007). Also, Pb-exposed animals had significant
12 decreases in brain volume in the stratum oriens (SO) region of the hippocampus,
13 specifically in the mossy fiber terminals of the SO. Pb-exposed animals also showed a
14 significant decrease in the length-density of immature or newly-formed neuron in the
15 outer portion of the DG. These findings show that Pb exposure at doses relevant to
16 humans induced significant decreases in adult hippocampus granule cell neurogenesis
17 and morphology, potentially providing mechanistic explanations for Pb-induced neuronal
18 aberrations and downstream effects such as learning and memory. Exposure of zebrafish
19 embryos to Pb (50-700 (iM Pb acetate in embryo medium from 0 to 6 days post hatch)
20 caused significant apoptosis of brain cells (increased TUNEL positive brain cells) and
21 decreased brain levels of some (gfap and huC) but not all (crestin and neurogeninl) genes
22 involved in neurogenesis (Dou and Zhang. 2011).
November 2012 5-222 Draft - Do Not Cite or Quote
-------
'
6000
5000
4000
3000
2000
1000
0
Con
Volume (mm3) i
» e •» -
3 cn O t
Con
Source: Reprinted with permission of Elsevier Science, Fox et al. (2010)
Note: Light micrograph pictures of Brd-U positive cells (proliferating cells undergoing DMA replication), black dots, in control (A) and
Pb-exposed rats (B). Counts of Brd-U positive cells (C) and Volume of hippocampus dentate gyrus (D) in control (white bars) and
Pb-exposed animals (black bars). *p <0.05 vs. control. Rats were exposed to 2,000 ppm Pb acetate from postnatal day 1 -30 (blood
Pb levels 34 ug/dL) and were examined at postnatal day 80.
Figure 5-14 Neurogenesis (production of new cells) in the rat hippocampal
dentate gyrus after early postnatal Pb exposure.
5.3.11.10 Neurite Outgrowth
3
4
5
6
7
8
9
10
11
12
13
14
As described in the 2006 Pb AQCD (U.S. EPA. 2006b). Pb was shown to decrease
neurite outgrowth in vitro and mediate such effects via protein kinase mediated pathways
(MAPK/ERK); earlier work had documented decreased primary DA neuron outgrowth
with 0.001 (iM Pb exposure (Lidsky and Schneider. 2004). A recent study showed that
gestational exposure of female Wistar rats to 500-4,000 (iM Pb chloride (resulting in
offspring blood Pb levels up to 12 (ig/dL) significantly decreased offspring hippocampal
neurite outgrowth and reduced the expression of hippocampal polysialylated neural cell
adhesion molecule (PSA-NCAM), NCAM, and sialytransferase (Hu et al.. 2008b). PSA-
NCAM is transiently expressed in newly formed neurons during the period of neurite
outgrowth from embryogenesis until the early postnatal period and is down-regulated in
the brains of adults except in areas known to exhibit synaptic plasticity (Seki and Arai.
1993). NCAM is important for memory formation, plasticity and synapse formation, and
its suppression by early-life Pb exposure may represent a mechanism mediating
Pb-associated impairments in cognitive function.
November 2012
5-223
Draft - Do Not Cite or Quote
-------
5.3.11.11 Epigenetics
1 Many investigators are beginning to show that environmental chemical exposures and air
2 pollution exposure are associated with epigenetic changes in humans (Baccarelli and
3 Bollati. 2009: Pavanello et al. 2009: Tarantini et al.. 2009: Bollati et al.. 2007).
4 Epigenetic changes involve changes in DNA expression without changes in the DNA
5 sequence, and these changes may be heritable. Epigenetic changes include histone
6 modification, DNA methylation, miRNA changes, or pathways that affect these
7 processes. Differential epigenetic modification has the potential to contribute to disease
8 by silencing or activating genes in an aberrant manner. For example, a recent study
9 identified differential methylation of a specific locus in monozygotic twins discordant for
10 schizophrenia (Dempster et al., 2011): Pb was not examined in this study.
11 DNA methyltransferases catalyze the transfer of a methyl group to DNA and are
12 important in epigenetics (i.e., silencing of genes like tumor suppressors) and imprinting.
13 DNA methyltransferase activity was significantly decreased in cortical neurons from
14 aged monkeys at ages 20-23 years after infancy exposure (PND1-PND400, blood Pb
15 level 19-26 (ig/dL) and fetal mouse brain cells exposed to Pb in culture (0.1 (iM Pb) (Wu
16 et al.. 2008b). Changes in DNA methyltransferases (Dnmtl, DnmtSa) were noted in
17 control monkey brains as they aged and these changes were exacerbated by early
18 postnatal Pb exposure (Bihaqi et al.. 2011). Another enzyme involved in DNA
19 methylation, methyl CpG binding protein 2 MECP2, showed a similar trend as the
20 Dnmts. Profiles of the histone modifying gene H34mc2 increased with age in control
21 animals. This age-related increase was significantly attenuated in Pb-exposed animals.
22 The cerebral cortex tissue used in this experiment was obtained from female primates
23 who had received 1.5 mg/kg Pb acetate via diet per day from birth until 400 days of age
24 (resulting in blood Pb levels 19-26 (ig/dL at age 400 days) (Rice. 1990).
25 Methyltransferases catalyze biological methylation reactions using cofactor S-adenosyl
26 methionine (SAM) as the methyl donor. In rats, SAM exposure after gestational-
27 lactational Pb exposure (1,500 ppm Pb acetate via drinking water of dams followed by
28 20-22 days of daily 20 mg/kg BW SAM exposure of offspring) improved hippocampal
29 LTP and Morris water maze performance at PND44-PND54 (Cao et al.. 2008). Thus, the
30 improved cognition and synaptic plasticity observed with co-exposure to Pb and the
31 methyl donor SAM suggest that methylation reactions may be involved in Pb-associated
32 effects on cognition.
November 2012 5-224 Draft - Do Not Cite or Quote
-------
5.3.11.12 Cholesterol and Lipid Homeostasis
1 Various pathological conditions are associated with elevated plasma free fatty acids or
2 elevated cholesterol. Adult male rats exposed to Pb acetate (200, 300, or 400 ppm) in
3 their drinking water for 12 weeks had increased cholesterogenesis and phospholipidosis
4 in brain tissue (Ademuyiwa et al., 2009). Pb-induced changes in brain cholesterol showed
5 an inverse U concentration-response relationship, with the largest increase in brain
6 cholesterol observed with 200 ppm Pb followed by 300 ppm Pb. Animals exposed to
7 400 ppm Pb did not have significant changes in brain cholesterol. In a separate study, Pb
8 treatment (single dose 100 (imol/kg, i.v.) was shown to depress the activity of
9 cholesterol-7-a-hydroxylase, an enzyme involved in biosynthesis of bile acid, which
10 mediates elimination of cholesterol from the body (Kojima et al., 2005). In Ademuyiwa
11 et al. (2009). Pb exposure significantly increased brain triglycerides by 83% at 300 ppm
12 and by 108% at 400 ppm. At 200 ppm, Pb exposure induced a statistically nonsignificant
13 decrease in brain triglycerides. Pb exposure across all three dose groups induced
14 significantly increased brain phospholipids. Interestingly, plasma free fatty acids were
15 significantly elevated in a concentration-dependent manner; plasma triglycerides and
16 cholesterol were unaffected by Pb exposure. The molar ratio of brain cholesterol to
17 phospholipids, an indicator of membrane fluidity, was significantly increased at 200 and
18 300 ppm Pb exposure, indicating increased membrane fluidity. Brain Pb in all dose
19 groups was below the limit of detection (0.1 ppm). Blood Pb levels at 0, 200, 300, and
20 400 ppm were 7, 41, 61, and 39 (ig/dL, respectively, higher than those relevant to
21 humans. In summary, a recent study found that adult 12-week Pb exposure significantly
22 increased brain cholesterol, triglycerides, and phospholipids as well as significantly
23 increased plasma free fatty acids in rats. These effects were sometimes more prominent at
24 the lower 200 ppm Pb dose. The impacts of these Pb-related changes in phospholipidosis
25 and cholesterogenesis in the brain on downstream nervous system effects are not well
26 characterized.
5.3.12 Lifestage of Pb Exposure and Neurodevelopmental Deficits
27 Environmental exposures during critical lifestages can affect key physiological systems
28 that orchestrate plasticity (Feinberg. 2007). Exposures during the prenatal and/or early
29 postnatal period may be especially detrimental for neurodevelopmental effects because of
30 active neuronal growth and/or synaptogenesis/pruning structure that occur during these
31 periods (Rice and Barone. 2000; Landrigan et al.. 1999). However, brain development
32 has been shown to continue throughout adolescence. MRI studies in children and adults
33 ages 3-30 years have shown that total cerebral volume peaks at age 10.5 and 14.5 years in
34 females and males, respectively (Giedd et al., 2009; Lenroot and Giedd. 2006). The
November 2012 5-225 Draft - Do Not Cite or Quote
-------
1 volume of the cerebellum was found to peak two years later. Lateral ventricular volume
2 and white and gray matter volume also were found to increase throughout adolescence.
3 Gray matter volume peaked 1 to 3 years earlier in females than males. These observations
4 that brain development is active throughout childhood and in adolescence indicate the
5 potential for neurodevelopment to be altered later in childhood.
6 Epidemiologic studies consistently show that blood Pb levels measured during various
7 lifestages and time periods, including the prenatal period, early childhood, later
8 childhood, and averaged over multiple years, are associated with cognitive function
9 decrements and increases in behavioral problems. These observations of Pb-associated
10 elevated risk of neurodevelopmental deficits in children are well supported by findings in
11 animals that prenatal and/or postweaning Pb exposure alters brain development via
12 changes in synaptic architecture (Section 5.3.11.4) and neuronal outgrowth
13 (Section 5.3.11.10) and leads to impairments in memory and learning (Section 5.3.2.3)
14 and increases in impulsivity (Section 5.3.3.1). In monkeys, Pb exposures during multiple
15 lifestages and time periods, including lifetime, lactational, or postlactation to adulthood,
16 resulted in impaired cognitive function, although not on all tests (Rice. 1992b. 1990; Rice
17 and Karpinski. 1988). On one test of executive function in the same monkeys at ages 5-6
18 years, impairments were found with lifetime Pb exposure starting from birth or starting
19 after weaning but not infancy-only exposure (Rice and Gilbert. 1990b). The latter
20 observations indicate that gestational or early infancy Pb exposures are not necessary to
21 induce cognitive function decrements in juvenile animals.
22 Unlike other organ systems, the unidirectional nature of CNS development limits the
23 capability of the developing brain to compensate for cell loss, and environmentally-
24 induced cell death can result in a permanent reduction in cell numbers (Bayer, 1989).
25 Hence, when normal development is altered, the early effects have the potential to persist
26 into adult life even in the absence of concurrent exposure, magnifying the potential public
27 health impact. Some epidemiologic evidence indicates associations of earlier childhood
28 blood or tooth Pb levels with cognitive function decrements, increases in inattention, and
29 increases in misconduct in adolescents or adults (Mazumdar et al.. 2011; Fergusson et al.
30 2008; Wright et al.. 2008; Ris et al.. 2004; Bellinger et al.. 1994a: Stiles and Bellinger.
31 1993). These epidemiologic studies did not examine adult blood Pb levels, thus the
32 relative influence of adult Pb exposure cannot be ascertained. In the Boston cohort,
33 stronger associations observed for age 2 year blood Pb level than concurrent blood Pb
34 level with FSIQ decrements at ages 57 months and 10 years indicated an effect of earlier
35 rather than later childhood Pb exposures (Bellinger et al.. 1992; 1991). The persistence of
36 effects of early exposures is supported by findings of impaired learning in adults
37 monkeys that had juvenile Pb exposure (Rice. 1992b. 1990). A few available recent
38 toxicological studies also found that infancy Pb exposure but not adult-only Pb exposure
November 2012 5-226 Draft - Do Not Cite or Quote
-------
1 led to neurodegenerative amyloid plaque formation in the brains of aged rodents and
2 monkeys (Section 5.3.10.1).
3 With repeated assessments of children prenatally to later childhood and early adulthood,
4 the prospective cohort studies have provided data to compare the neurodevelopmental
5 effects associated with blood Pb levels measured at different lifestages and time periods.
6 In the collective body of evidence, cognitive function decrements in children have been
7 associated with prenatal, early childhood, childhood average, and concurrent blood Pb
8 levels, without clear indication of a single critical lifestage or duration of Pb exposure
9 related to risk of neurodevelopmental effects in children. In prospective studies, the
10 identification of critical developmental periods with regard to risk of neurodevelopmental
11 decrements from Pb exposure has been complicated by the high degree of correlation of
12 the blood Pb levels of children over time and the confounding of age and peak blood Pb
13 levels (Lanphear et al., 2005; Dietrich et al.. 1993a; Needleman et al.. 1990).
14 As described in detail in the 2006 Pb AQCD (U.S. EPA. 2006b). several studies with
15 varying lengths of follow-up demonstrated associations of prenatal blood Pb levels
16 (maternal and umbilical cord) with decrements in cognitive function throughout
17 childhood and into early adulthood (Section 5.3.2). These findings are consistent with the
18 observations of active CNS development occurring during prenatal development as
19 described above. Substantial fetal Pb exposure may occur from mobilization of maternal
20 skeletal Pb stores, which may be related to past maternal Pb exposures (Gulson et al..
21 2003; Hu and Hernandez-Avila. 2002). Pb can cross the placenta to affect the developing
22 fetal nervous system (Rabinowitz. 1988). Among 94-211 mother-child pairs in Albany,
23 NY, maternal-cord blood Pb level correlations of 0.53-0.81 were reported, depending on
24 the stage of pregnancy, indicating the influence of maternal blood Pb levels on newborn
25 blood Pb levels (Schell et al.. 2003). Depending on the magnitude of child exposure, the
26 contribution of maternal blood Pb levels on child blood Pb levels may wane early, and by
27 age 9 months, child blood Pb levels may be influenced mainly by child Pb exposures
28 (Section 4.4.1). Thus, associations of neurodevelopmental outcomes assessed after
29 infancy with postnatal blood Pb levels may reflect effects of postnatal Pb exposures.
30 In most studies of very young children, ages <2 years, decrements in MDI score were
31 associated with higher prenatal (maternal or cord) and concurrent blood Pb levels (Table
32 5-14). Among studies that examined blood Pb levels at multiple time periods, several
33 found larger blood Pb-associated decrements in MDI for prenatal blood Pb than
34 concurrent blood Pb (Hu et al.. 2006; Gomaaet al.. 2002; Bellinger etal.. 1987; Dietrich
35 etal.. 1986). In the Yugoslavia cohort, per log increase in blood Pb level, the MDI
36 decrement at age 2 years was larger for concurrent blood Pb than for prenatal cord blood
37 Pb (Wasserman et al.. 1992). Concurrent blood Pb levels were higher than prenatal cord
November 2012 5-227 Draft - Do Not Cite or Quote
-------
blood Pb levels. The collective evidence indicates that both prenatal and postnatal child
Pb exposures may contribute to neurodevelopmental effects in children from infancy to
age 2 years, with some indication that prenatal Pb exposure has a stronger effect.
Table 5-14 Associations of cognitive function with blood Pb levels measured at
various lifestages and time periods in prospective studies.
Study3
Study Population and
Methodological Details
(Within studies, effect estimates are
presented in order of lifestage or
time period of blood Pb
measurement, with largest effect
estimate in bold)
Blood Pb
Levels
(ug/dL)
Outcome
Effect Estimate
(95% Cir
Cognitive Assessment at Age 2 Years and Younger
Bellinger et 249 children followed from birth
al. (1987) (1979-1981) to age 3 yr, Boston
area, MA
Moderate participation rate, high
follow-up retention. Participants had
higher cord blood Pb levels, higher
SES, maternal IQ, HOME score.
Regression model adjusted for the
maternal age, race, IQ, education,
years cigarette smoking,3rd
trimester alcoholic drinks/ week,
study period mean SES, HOME,
child sex, birth weight, gestational
age, birth order.
Prenatal
(cord)
Mean (SD):
6.6 (3.2)
Overall
Bayley MDI
among
Ages 6, 12,
18, and 24
mo
vs. prenatal blood Pb level <3 ug/dL:
Prenatal 6-7 |jg/dL:
-3.8 (-6.3,-1.3)
Prenatal > 10 |jg/dL:
-4.8 (-7.3, -2.3)
Concurrent reported to not to be
associated with overall MDI, no
quantitative data reported.
Dietrich et 280 children followed prenatally to
al. (1986) age 6 mo, Cincinnati, OH
No information on participation rate.
Log linear regression model adjusted
for birth weight, gestation, sex. Also
considered potential confounding by
SES, HOME score, prenatal
smoking and alcohol use, maternal
Fe binding.
Prenatal
(maternal)
Mean (SD):
8.0 (3.8)
Concurrent
Mean (SD):
4.5 (2.9)
Bayley MDI
Age 6 mo
Prenatal: -0.6 (-1.1, -0.09)
Concurrent: -0.23 (-0.58, 0.12)
Huetal. 146 children born 1997-1999
(2006) followed prenatally to age 2 yr,
Mexico City, Mexico
Moderate follow-up participation.
Eligible similar to non-eligible. Log
linear regression model adjusted for
sex, maternal age, current weight,
height-for-age Z score, maternal IQ,
concurrent blood Pb (in models
examining blood Pb at other
lifestages). Considered potential
confounding by other unspecified
factors.
Prenatal
(maternal 1st
trimester):
Mean(range):
7.1 (1.5-43.6)
Prenatal avg:
NR
Earlier
childhood at
12 mo:
Mean (SD):
5.2 (3.4)
Concurrent
Mean (SD):
4.8 (3.7)
Bayley MDI
Age 24 mo
Per log increase in blood Pb:
Prenatal 1st trimester: -4.1 (-8.1, -0.17)
Prenatal avg: -3.5 (-7.7, 0.63)
Age 12 month: -2.4 (-6.2, 1.49)
Concurrent: -1.0 (-3.9, 1.9)
November 2012
5-228
Draft - Do Not Cite or Quote
-------
Study3
Study Population and
Methodological Details
(Within studies, effect estimates are
presented in order of lifestage or
time period of blood Pb
measurement, with largest effect
estimate in bold)
Blood Pb
Levels
(ug/dL)
Outcome
Effect Estimate
(95% Cl)b
Gomaa et 197 children followed prenatally to
al. (2002) age 2 yr, Mexico City, Mexico
Moderate participation but high
retention. No selective attrition. Log
linear regression model adjusted for
maternal IQ, maternal age, sex,
parental education, marital status,
breastfeeding duration, child
hospitalization status. Did not
consider potential confounding by
parental caregiving quality.
Prenatal
(cord)
Mean (SD):
6.7 (3.4)
Concurrent
Mean (SD):
8.4 (4.6)
Bayley MDI
Age 24 mo
Per log increase in blood Pb:
Prenatal: -2.1 (-3.9, -0.39)
Concurrent: -0.09 (-0.58, 0.42)
Wasserman 392 children followed prenatally to
et al. (1992) age 24 mo, Kosovo, Yugoslavia (K.
Mitrovica, Pristina)
High follow-up participation, no
selective attrition. K. Mitrovica near
smelter. Log linear regression model
adjusted for sex, birth order, birth
weight, ethnicity, HOME, maternal
education, age, and IQ.
Prenatal
(cord)
Mean (SD):
14.4(10.4)
Concurrent
Means:
K. Mitrovica:
35.4,
Pristina: 8.5
Bayley MDI
At age 24
Per log increase in blood Pb:
Concurrent: -4.1 (-6.2, -2.0)
Prenatal: -3.2 (-7.2, 0.86)
359 children, followed prenatally to
age Syr, Cleveland, OH
Prospective. Recruitment at birth
hospital. High follow-up participation,
more white, higher IQ, nonalcoholic
mothers not followed. 50% born to
alcoholic mothers. Linear regression
adjusted for age, race, sex, birth
order, parental education, maternal
IQ, Authoritarian Family Ideology,
HOME.
Means (SD):
Prenatal
cord:
6.0(2.1)
6 mo:
10.1 (3.3)
Concurrent:
16.7(6.5)
Bayley MDI Variance estimates:
Age2yr Prenatal: 0.0003, t =-0.21d
Age 6 mo: 0.00, p = 0.95d
Concurrent: 0.00, p = 0.95d
Cognitive Function Assessments at School Age
Canfield et 172 children born 1994-1995
al. (2003a) followed from age 6 mo to 5 yr,
Rochester, NY
Recruitment from study of dust
control. 73% nonwhite. High follow-
up participation, no selective
attrition. Linear regression model
adjusted for maternal race, IQ,
education, and prenatal smoking
status, household income, HOME
score, child sex, Fe status, birth
weight.
Means (SD):
Infancy avg
(6-24 mo):
7.0 (3.8)
Peak:
11.1 (7.1)
Concurrent:
5.8(4.1)
Lifetime (to
age 5 yr)
avg:
7.4 (4.3)
FSIQ
Stanford-
Binet
Age 5 yr
Infancy avg: -0.53 (-0.95, -0.13)
Peak: -0.26, (-0.47, -0.05)
Concurrent: -0.61 (-0.99, -0.24)
Lifetime avg: -0.57 (-0.93, -0.20)
Wasserman 332 children followed prenatally to
et al. (1994) age 3-4 yr, Kosovo, Yugoslavia (K.
Mitrovica, Pristina)
High follow-up participation. More
participants were male, Albanian,
and had lower maternal IQ and
HOME. Log linear regression model
adjusted for HOME score, maternal
age, intelligence, and education,
language, birth weight, child sex.
Prenatal
(cord)
Mean (SD):
14.4(10.4)
Concurrent
means:
K. Mitrovica:
39.9
Pristina:
9.6
Overall mean
NR
FSIQ
McCarthy
General
Cognitive
Index
Age 3-4 yr
Per log increase in blood Pb:
Prenatal: -7.1 (-11.8, -3.1)
Age 2 yr: -10.4 (-15.2, -5.7)
Concurrent:-9.4 (-14.2, -4.6)
November 2012
5-229
Draft - Do Not Cite or Quote
-------
Study3
Study Population and
Methodological Details
(Within studies, effect estimates are
presented in order of lifestage or
time period of blood Pb
measurement, with largest effect
estimate in bold)
Blood Pb
Levels
(ug/dL)
Outcome
Effect Estimate
(95% Cl)b
Bellinger et 148 children followed from birth
al. (1992) (1979-1981) to age 10yr, Boston,
MA area
Moderate follow-up participation.
Participants had higher SES and
HOME scores. Linear regression
model adjusted for HOME score
(age 10 and 5 yr), maternal race, IQ,
and marital status, SES, child sex,
birth order, and stress, # residence
changes. Also considered potential
confounding by family stress,
maternal age, psychiatric factors,
and child serum ferritin levels.
Age 6 mo
Mean (SD):
6.7 (7.0)
Earlier
childhood
Age 2 yr
Mean (SD):
6.5 (4.9)
Concurrent
Mean (SD):
2.9 (2.4)
FSIQ
Wechsler
Intelligence
Scale for
Children-
Revised
Age 10 yr
Age 6 mo:-0.13 (-0.42, 0.16)
Age 2 yr: -0.58 (-0.99, -0.18)
Concurrent -0.46 (-1.5, 0.56)
Dietrich et 253 children followed from birth
al. (1993b) (1979-1985) to age 6.5 yr,
Cincinnati, OH
High follow-up participation.
Participants had slightly higher age 1
yr blood Pb levels. Linear regression
model adjusted for HOME score,
maternal IQ and prenatal cigarette
smoking, child birth weight, birth
length, sex. Also considered
potential confounding by perinatal
complications, prenatal maternal
substance abuse, nutritional status.
Prenatal
(maternal)
Mean (SD):
8.3 (3.7)
Age 5 yr
Mean (SD):
11.8(6.3)
Concurrent
NR
Lifetime (to
age 6.5 yr)
avg:
NR
FSIQ
Wechsler
Intelligence
Scale for
Children-
Revised
Age 6.5 yr
Prenatal: 0.15 (-0.26, 0.56)
Concurrent: -0.33 (-0.60, -0.06)
Lifetime avg: -0.13 (-0.35, 0.09)
Baghurst 494 children followed from birth
et al. (1992) (1979-1982) to age 7 yr, Port Pirie,
Australia
Moderate follow-up participation.
Participants had higher SES and
breastfeeding, less maternal
smoking. Log linear regression
model adjusted for sex, birth weight,
birth order, feeding method,
breastfeeding duration, parental
education and smoking, maternal
age and IQ, SES, HOME, parents
living together.
Prenatal
(maternal)
Mean 2nd
quartile:
7.4
Earlier
childhood
Age 2 yr
Mean 2nd
quartile:
16.6
Lifetime (to
age 7 yr) avg
Mean 2nd
quartile:
15.7
FSIQ
Wechsler
Intelligence
Scale for
Children-
Revised
Age 7-8 yr
Prenatal: 0.26 (-0.67, 1.5)
Age 2 yr: -2.0 (-3.8, -0.21)
Lifetime avg:-1.6 (-3.7, 0.52)
Schnaas et 150 children followed from prenatally
al. (2006) (1987-1992) to age 6-10 yr, Mexico
City, Mexico
Low follow-up participation.
Participants had higher SES, FSIQ,
higher blood Pb level before age 5
yr, lower at older ages. Log linear
mixed effects regression model
adjusted for SES, maternal IQ,
HOME score, child sex, birth weight,
indicator of first FSIQ measurement,
random slope for subject. Most
covariates assessed in pregnancy or
within child age 6 mo.
Geometric
Mean (range)
Prenatal
(maternal
28-36 week
gestation):
7.8 (2.5-
24.6)
Age 5 yr: 9.3
(3.8-18.0)
Age 6-10 yr
avg: 6.2
(2.2-18.6)
FSIQ
Wechsler
Intelligence
Scale for
Children-
Revised
Ages 6-10
yr
Per log increase in blood Pb:
Prenatal: -4.0 (-6.4, -1.7)
Age 5 yr: -0.32 (-4.3, 3.4)
Age 6-10 yr avg: -2.5 (-4.1, -0.81)
November 2012
5-230
Draft - Do Not Cite or Quote
-------
Study3
Study Population and
Methodological Details
(Within studies, effect estimates are
presented in order of lifestage or
time period of blood Pb
measurement, with largest effect
estimate in bold)
Blood Pb
Levels
(ug/dL)
Outcome
Effect Estimate
(95% Cl)b
Ris et al. 195 children followed prenatally
(2004) (1979-1985) to age 15-17 yr,
Cincinnati, OH
Prospective. High follow-up
participation, no selective attrition.
Mostly African-American. Linear
regression model adjusted for SES,
maternal IQ, average HOME,
adolescent marijuana use, and
obstetrical complications. Also
considered potential confounding by
birth outcomes, maternal age,
prenatal smoking, alcohol,
marijuana, and narcotics use, #
previous abortions, stillbirths,
gravidity, parity, caregiver education,
public assistance, child age, sex,
health, and Fe status
Prenatal,
Earlier
childhood
Age 6.5 yr,
Earlier
childhood
avg (Age
3-78 mo):
NR
Learning/IQ
composite
Wechsler
Intelligence
Scale for
Children
indices
normal
scores
Age 15-17
yr
Prenatal: -0.08 (-0.18, 0.03)
Age 6.5 yr: -0.08 (-0.17, 0.003)
Age 3-78 mo avg: -0.03 (-0.18, 0.03)
Lanphear 1,333 children pooled from Boston,
et ai. (2005) Cincinnati, Cleveland, Mexico City,
Port Pirie, Rochester, and
Yugoslavia cohorts.
Uniform analysis of cohorts from
diverse locations and SES. Blood Pb
levels and FSIQ measured at
different ages. Several sensitivity
analyses to examine heterogeneity
of results by cohort, model
specification, and confounding. Log
linear regression model adjusted for
HOME score, birth weight, maternal
IQ, maternal education. Also
considered potential confounding by
child sex, birth order, maternal age,
marital status, prenatal smoking
status, prenatal alcohol use.
Median
(5th-95th)
Early
childhood
(mean ages
6-24 mo):
12.7
(4.0-34.5)
Peak:
18.0
(6.2-47.0)
Lifetime avg
(to ages
4.8-1 Oyr):
12.4
(4.1-34.8)
Concurrent:
9.7 (3.5-33.2)
FSIQ
Various
tests
Ages 4.8-10
yr
Mean ages 6-24 mo: -0.14 (-0.23, -0.06)
Peak:-0.20 (-0.29,-0.11)
Concurrent: -0.23 (-0.29, -0.11)
Lifetime avg: -0.15 (-0.22, -0.09)
Pocock et Meta-analysis of 5 prospective
al. (1994) studies (over 1,100 children) from
Port Pirie and Sydney, Australia,
Cincinnati, Cleveland, Boston
Meta-analysis of combining
covariate-adjusted effect estimates
from individual studies.
Earlier
childhood
(2 yr) range
in means:
6.8-21.2
Around birth
and
Postnatal:
NR
FSIQ
Various
tests
Ages 5-10
yr
Around birth: 0.26 (-1.5, 2.0)
Age 2 yr: -2.7 (-4.1, -1.2)
Postnatal mean: -1.3 (-2.9, 0.37)
MDI = Mental Development Index, FSIQ = full-scale IQ, NR = Not reported
"Results are presented first for MDI in children up to age 3 years, then for FSIQ in school-aged children. Within studies, effect
estimates are presented in order of increase lifestage or time period of blood Pb measurement, with the largest effect estimate in
bold.
bEffect estimates are standardized to a 1 ug/dL increase in blood Pb level in analyses of blood Pb as a linear continuous variable.
°Effect estimates represent comparisons between children in different categories of blood Pb level, with children in the lower
blood Pb category serving as the reference group.
Sufficient data were not provided in order to calculate 95% Cl.
November 2012
5-231
Draft - Do Not Cite or Quote
-------
1 Prenatal and early postnatal (age 6 month) blood Pb levels also were associated with
2 cognitive function in children examined at school-age (ages 4-17 years) (Table 5-14).
3 However, most of these studies also found cognitive function decrements in association
4 with postnatal blood Pb levels, and results did not identify an individual postnatal time
5 period of blood Pb measurement associated with cognitive function decrements.
6 Increases in concurrent blood Pb levels were associated with larger decrements in FSIQ
7 in the Cincinnati and Yugoslavia cohorts at ages 3-6.5 years than were prenatal blood Pb
8 levels (Wasserman et al.. 1994; Dietrich et al.. 1993b). In the Cincinnati cohort as
9 adolescents ages 15-17 years, increases in both prenatal and higher earlier childhood (age
10 6.5 years) were associated with decrements in a learning/memory composite score (Ris et
11 al.. 2004). In the Boston and Port Pirie cohorts, increases in age 2 year blood Pb levels
12 were associated with larger FSIQ decrements at ages 7 and 10 years, respectively, than
13 were concurrent or lifetime average blood Pb levels (Baghurst et al.. 1992; Bellinger et
14 al., 1992). However, in the Port Pirie cohort, an association also was found lifetime (to
15 age 7 years) average blood Pb level. Among children ages 6-10 years in Mexico City, per
16 unit increase, prenatal blood Pb levels were associated with larger FSIQ decrements than
17 individual blood Pb levels between ages 1-5 or the age 6-10 year average (Schnaas et al..
18 2006). In contrast, results from the Rochester cohort indicated that increases in lifetime
19 average and concurrent blood Pb level were associated with larger FSIQ decrements at
20 ages 5 years than were increases in peak blood Pb level (Canfield et al., 2003a).
21 Collectively, the epidemiologic findings indicate that blood Pb levels measured at various
22 postnatal time periods, earlier childhood, childhood average, later childhood, and
23 concurrent blood Pb levels are associated with decrements in cognitive function when
24 assessed in school-aged children.
25 Consistent with individual studies, analyses combining studies pointed to associations of
26 FSIQ in school-aged children with blood Pb levels measured at various lifestages and
27 time periods. The analysis pooling data from seven prospective studies found that
28 increases in infancy average (age 6-24 months), peak, concurrent, and lifetime average
29 peak blood Pb levels were associated with decreases in FSIQ in children ages
30 4.8-10 years (Table 5-14). Investigators reported that the model with concurrent blood Pb
31 level explained the largest proportion of variance in FSIQ (R2) (Lanphear et al., 2005). In
32 a meta-analysis of results from five cohort studies (Pocock et al.. 1994). a larger decrease
33 in FSIQ was estimated for an increase in peak (around age 2 years) blood Pb level than
34 for blood Pb level measured around birth or after age 2 years. Deciduous tooth Pb levels
35 have been associated with decrements in cognitive function and increases in attention-
36 related behavioral problems in children and young adults (Table 5-8. Table 5-9. and
37 Table 5-11). These results indicate cumulative Pb exposure over several years may
38 contribute to neurodevelopmental effects in children. In school-aged children, concurrent
39 blood Pb levels reflect past Pb exposures that are mobilized from bone remodeling to
November 2012 5-232 Draft - Do Not Cite or Quote
-------
1 blood and recent exposures. Thus, associations with concurrent blood Pb levels also may
2 reflect an effect of cumulative past and more recent Pb exposures.
3 Studies conducted in the Cincinnati cohort examined diverse neurodevelopmental effects
4 and found that prenatal and neonatal blood Pb levels were associated with impaired
5 auditory processing in children ages 5 years and increases in parental ratings of
6 delinquent behavior in adolescents ages 15-17 years (Dietrich etal. 2001; Dietrich et al.,
7 1992) but not cognitive function or motor function decrements in children at age 6 years
8 (Bhattacharya et al., 2006; Bhattacharya et al.. 1995; Dietrich et al., 1993b; Dietrich et
9 al.. 1993a). These findings suggest that the critical lifestage of Pb exposure may vary
10 among nervous system effects.
11 Some studies have aimed to improve the characterization of important lifestages and time
12 periods of Pb exposure by examining children in whom blood Pb levels are not strongly
13 correlated over time (i.e., children whose blood Pb level ranking changed over time)
14 (Hornung et al.. 2009; Schnaas et al.. 2006; Chen etal.. 2005; Tongetal.. 1998; Bellinger
15 et al.. 1990). Collectively, most results indicated FSIQ decrements in association with
16 concurrent blood Pb levels but did not conclusively demonstrate stronger findings for
17 early or concurrent blood Pb levels (Table 5-15).
November 2012 5-233 Draft - Do Not Cite or Quote
-------
Table 5-15 Comparisons of blood Pb-FSIQ associations in groups of children
with different temporal trends in blood Pb levels.
Study
Hornung
etal.
(2009)
Bellinger
etal.
(1990)
long et
al.
(1998)
Study Population and
Methodological Details
397 children followed from
birth (1979-1 984) to age 6 yr,
Rochester, NY and
Cincinnati, OH.
High follow-up participation.
No selective attrition. Linear
regression model adjusted
for city, HOME score, birth
weight, maternal IQ,
maternal education.
170 children followed
prenatally to age 57 mo,
Boston area, MA
High follow-up participation.
No report on characteristics
of subjects followed. Log
linear regression adjusted for
HOME score, SES, maternal
IQ, maternal age, sex,
ethnicity
375 children followed from
birth (1979-1 982) to age
11-1 Syr, Port Pirie, Australia
Moderate follow-up
Darticioation. Particioants
Blood Pb Levels
(ug/dL)
Geometric mean
(5th-95th):
Earlier childhood
age 2 yr:
8.9 (3.0-23.8)
Concurrent; 6.0
(1.9-17.9)
High prenatal (cord)
>10
Low concurrent: <3
Medium concurrent:
3-10
High concurrent >10
Means
Earlier childhood
age 2 yr: 21 .2
Age 11-1 Syr: 7.9
Outcome
FSIQ
Wechsler Intelligence
Scale for Children-
Revised (WISC-R)
Age 6 yr
Change in McCarthy
General Cognitive Index
(GCI)z-score
Between ages 24 and 57
mo
Change in cognitive
function z-scores
between ages 2 and 1 1 yr
n— ., .i_, . it n r\\
Effect Estimate(95% Cl)
Difference in FSIQ at age 6 yr
with an age
6:2 yr blood Pb level ratio = 0.5 as
the reference
Ratio age 6:2 yr
blood Pb level = 2.0
-7.0 (-10, -4.0)
High prenatal/Low concurrent
0.42 (-0.1 5, 0.99)
High prenatal/Medium concurrent
0.15 (-0.14, 0.44)
High prenatal/High concurrent
-0.1 5 (-0.56, 0.26)
<10.2 ug/dL decline:
0.03 (-0.1 5, 0.21)
10.2-1 6.2 ug/dL decline:
0.04 (-0.1 5, 0.23)
had lower early blood
Pb-cognitive function
association. Log linear
regression model adjusted
for sex, birth weight, birth
rank, feeding style,
breastfeeding duration,
maternal IQ, maternal age,
SES, HOME score, parental
smoking, parents living
together. ANOVA to assess
association of change in IQ
with change in blood Pb
across time intervals
Bayley MDI,
age 2 yr
McCarthy GCI,
age4yr
WISC-R,
ages 7,
and 11-13yr
>16.2 ug/dL decline:
-0.01 (-0.20,0.18)
Chen et 780 children participating in
al. the TLC trial from age
(2005) 12-33 mo to age 7 yr,
Baltimore, MD; Cincinnati,
OH; Newark, NJ;
Mostly African-American.
50% given chelation at ages
12-33 mo, blood Pb levels
20-44 ug/dL. No information
on participation rate.
Regression-based path
analysis adjusted for city,
race, sex, language, parental
education and employment,
single parent, age at blood
Pb measurement, caregiver
IQ. Considered potential
confounding by chelation but
not parental caregiving
quality.
Mean (SD):
Age 2 yr: 26.2 (5.1)
Age Syr: 12.0(5.2)
Age 7 yr: 8.0 (4.0)
Low age 2 yr: <24.9
Low age 7 yr: <6.2
WISC-lllatage7yr
Difference in FSIQ vs. Low age 2,
Low age 7 as the reference
Low age 2, High age 7:
-0.27 (-0.48, -0.05)
High age 2, Low age 7:
0(-0.21, 0.20)
High age 2, High age 7:
-0.28 (-0.47,-0.10)
November 2012
5-234
Draft - Do Not Cite or Quote
-------
1 Schnaas et al. (2006) followed children in Mexico City prenatally to age 10 years and
2 found maternal blood Pb levels at 28-36 weeks of pregnancy to be weakly correlated with
3 repeated measures of blood Pb between ages 1 and 10 years (Pearson r < 0.23). In models
4 analyzing different lifestages of blood Pb level individually and in a mixed effects model
5 that included prenatal and multiple postnatal blood Pb measures, maternal 28-36 week
6 blood Pb level was associated with a larger decrement in FSIQ (Table 5-15). In the model
7 with multiple lifestages of blood Pb level, analysis of variance inflation factors indicated
8 a lack of collinearity among the serial blood Pb measures.
9 Pooling the Cincinnati and Rochester cohorts (n = 397), Hornung et al. (2009) created a
10 new indicator of Pb exposure: the ratio of blood Pb level at 6 years of age to that at
11 2 years of age. As illustrated in Figure 5-15. the three groups of children representing the
12 three temporal trends in blood Pb levels: no change ages 2-6 years (ratio = 1), higher
13 blood Pb at age 6 years than 2 years (ratio = 1.25), higher blood Pb levels at age 2 years
14 than 6 years (ratio = 0.5), have similar areas under the curve, indicating that cumulative
15 blood Pb levels were similar in the three groups. Thus, differences in FSIQ are more
16 likely to be attributable to differences in temporal trends. The lowest FSIQ at age 6 years
17 was found in children with an age 6:age 2 year blood Pb ratio 1.25, i.e., children who had
18 an increase in blood Pb level from 2 to 6 years of age (Figure 5-15 and Table 5-15).
November 2012 5-235 Draft - Do Not Cite or Quote
-------
18
16
14
12
6-year:2-year
ratio = 0.5
IQ = 89.0
6-year:2-year
ratio = 1.25
= 83.7
3 4
Age (years)
Note: In combined Cincinnati and Rochester cohorts, FSIQ was compared among three different patterns of blood Pb level changes
over time: peak at 2 years and age 6:2 year ratio = 0.5 (blue diamonds), peak at 5 years and age 6:2 year ratio = 1.25 (black
triangles), and constant blood Pb level and age 6:2 year ratio = 1 (white squares). All three patterns have a similar cumulative blood
Pb level (10 ug/dL) as indicated the areas under the curve. Children whose blood Pb levels peaked at age 5 years had the lowest
FSIQ at age 6 years.
Source: Hornung et al. (2009)
Figure 5-15 Estimated FSIQ for three patterns of temporal trends in blood Pb
level from ages 2 to 6 years in the Rochester and Cincinnati
cohorts.
i
2
o
3
4
5
6
7
8
9
10
n
12
13
U
15
In the Boston cohort with comparable blood Pb levels to those in the Rochester cohort,
Bellinger et al. (1990) found that at age 57 months, FSIQ, as assessed by McCarthy GCI,
was similar between children in the high (> 10 (ig/dL) and low (<3 (ig/dL) prenatal cord
blood Pb groups. Additionally, children with high prenatal and high concurrent blood Pb
level (> 10 (ig/dL) had a decrease (-0.15 standard deviation [95% CI: -0.46, 0.26]) in
FSIQ from age 24 to 57 months. In contrast, children with high prenatal blood Pb but low
concurrent blood Pb level (<3 (ig/dL) had an increase (0.42 standard deviation [95% CI:
-0.15, 0.99]) in FSIQ from age 24 to 57 months (Table 5-15). These findings indicated
that by age 5 years, children with higher prenatal blood Pb levels appeared to recover the
Pb-associated decrements in cognitive function unless concurrent blood Pb levels
remained high. The investigators also demonstrated that optimal sociodemographic
characteristics (e.g., higher HOME score, SES, maternal IQ and age, female) also
protected against decrements in cognitive function associated with higher postnatal blood
Pb levels. Collectively, these results suggest that cognitive development is not fixed early
in childhood and can be affected negatively or positively by postnatal influences.
November 2012
5-236
Draft - Do Not Cite or Quote
-------
1 Chen et al. (2005) also found a stronger influence of higher concurrent blood Pb levels on
2 FSIQ at age 7 years among children participating in a multi-city chelation trial. Children
3 with higher concurrent blood Pb levels (> median 7.2 (ig/dL) had lower IQ at age 7 years,
4 regardless of whether blood Pb level at age 2 years was low or high (less than or greater
5 than the median of 24.9 (ig/dL, respectively). Blood Pb levels at ages 2 and 7 years were
6 weakly correlated (r = 0.27). Because these children had been treated with chelators due
7 to high blood Pb levels (20-44 (ig/dL) at ages 12 to 33 months, the findings may have
8 limited generalizability to the general population of children currently living in the U.S.
9 In contrast with the aforementioned studies, Tong et al. (1998) found that higher early-
10 life blood Pb level was associated with a larger decrease in FSIQ than was concurrent
11 blood Pb level in the Port Pire, Australia cohort at age 11-13 years (Table 5-15). This
12 conclusion was based on the analysis of groups of children with different degrees of
13 decline in blood Pb levels between ages 2 and 11-13 years. Although the mean blood Pb
14 level in the study population declined overall from 21.2 (ig/dL at age 2 years to 7.9 (ig/dL
15 at age 11-13 years, the magnitude of decline varied among children. The change in FSIQ
16 between ages 2 and 11-13 years did not significantly differ between children with the
17 largest decline (>16 (ig/dL) in blood Pb level and children with the smaller decline
18 (<10 (ig/dL) (Table 5-15). These findings indicated an influence of higher blood Pb
19 levels early in life despite declines in blood Pb with age and a persistence of Pb effects.
20 The results do not preclude an independent association with concurrent blood Pb level.
21 A common limitation of studies that examined different temporal trends in blood Pb
22 levels is the higher blood Pb levels of the study populations compared to those currently
23 measured in most U.S. children. Additionally, in several study populations, children
24 experienced large changes in blood Pb levels overtime, for example, 50% decline or 25%
25 increase in four years in Hornung et al. (2009). It is unclear whether these findings would
26 apply to children currently in the U.S. within the same age range who would be expected
27 to have smaller changes in blood Pb levels over time.
28 To conclude, the collective body of epidemiologic evidence does not strongly identify an
29 individual critical lifestage or duration of Pb exposure with regard to neurodevelopmental
30 effects in children. Cognitive function decrements and behavioral problems have been
31 associated with prenatal, early childhood, lifetime average, and concurrent blood Pb
32 levels as well as with childhood tooth Pb levels. The identification of critical lifestages of
33 Pb exposure is complicated further by the fact that blood Pb levels in older children,
34 although affected by recent exposure, are also influenced by Pb stored in bone due to
35 rapid growth-related bone turnover in children relative to adults. Thus, associations of
36 neurodevelopmental effects with concurrent blood Pb level in children may reflect the
37 effects of past and/or recent Pb exposures (Section 4.3.5.1). Evidence indicates that
November 2012 5-237 Draft - Do Not Cite or Quote
-------
1 prenatal blood Pb levels are associated with mental development in very young children
2
-------
1 3-5 (ig/dL (Bellinger. 2008; Canfield. 2008; Hornung. 2008; Tellez-Roio. 2008). Except
2 for the pooled analysis, the lower strata of blood Pb levels comprised >30% of the study
3 population, indicating that the blood Pb-FSIQ relationships calculated for the lower strata
4 likely are not outliers or unrepresentative of the overall study population. Studies in the
5 Boston and Rochester cohorts each examined different ages of blood Pb and FSIQ but
6 lower blood Pb levels compared to other studies (means ~6 (ig/dL), and found nonlinear
7 blood Pb-FSIQ concentration-response relationships with respect to children whose peak
8 blood Pb levels did not exceed 10 (ig/dL (Bellinger and Needleman. 2003; Canfield et al..
9 2003a; Bellinger etal.. 1992). Pooled analyses of seven prospective studies, involving a
10 wider range of blood Pb levels, 5th-95th percentiles 2.4-33.1 (ig/dL, found that nonlinear
11 models (e.g., log-linear, piecewise linear) fit the relationship between blood Pb level and
12 FSIQ better than a linear model did (Lanphear et al., 2005; Rothenberg and Rothenberg.
13 2005).
November 2012 5-239 Draft - Do Not Cite or Quote
-------
Study Blood Pb Timing Outcome Blood Pb
(2003)
Canfieldetal.(2003)
Lanphearetal. (2005)
Age2yr
Concurrent
Concurrent
Cross-sectional
Tellez-Rojoetal. (2006) Concurrent
Lanphearetal. (2000)
Kordasetal. (2006)
Concurrent
Concurrent
""-""« stratum (ug/dL)
FSIQ All subjects
Peak<10a*
FSIQ All subjects
Peak< 10=
FSIQ Peak>10=
Peak<10a
Peak>7.5=
Peak<7.5=
BayleyMDI >10b
<10b
5-1 Ob
<5b
Reading score All subjects
<10
<7.5
<5
<2.5
Math score All subjects
<10
1230 -4-
532 -4-
Change in cognitive function test score perl ug/dL
increase in blood Pb level (95% Cl)
Note: Results are presented first for prospective studies then for cross-sectional studies. FSIQ = full-scale IQ, MDI = mental
development index. Effect estimates (concentration-response) are presented for a 1 ug/dL increase in blood Pb level. Black symbols
represent effect estimates among all subjects or subjects in the higher blood Pb stratum. Blue symbols represent effect estimates in
lower blood Pb strata. aStrata were defined by the peak blood Pb level measured in child at any point during follow up. b95% Cl
estimated from reported p-value.
Figure 5-16 Comparison of associations between blood Pb level and cognitive
function among various blood Pb strata.
November 2012
5-240
Draft - Do Not Cite or Quote
-------
Table 5-16 Additional characteristics and quantitative results for studies
presented in Figure 5-16.
Study
Bellinger et
al. (1992)
Bellinger
and
Needleman
(2003)
Bellinger
(2008)
Canfield et
al. (2003a)
Canfield
(2008)
Jusko et al.
(2008)
Lanphear et
al. (2005)
Hornung
(2008)
Study Population and
Methodological Details
148 children followed from
birth (1979-1 981) to age 10
yr, Boston area, MA.
Prospective. Recruitment at
birth hospital. Participation by
59% of original cohort but
88% of eligible. Participants
had higher SES. Linear
regression model adjusted for
HOME score (age 10 and 5),
child stress events, race,
maternal IQ, SES, sex, birth
order, marital status,
# residence changes before
age 5 yr.
172 children born 1994-1995
followed from age 6 mo to
age 5 yr, Rochester, NY.
Prospective. Recruitment
from study of dust control.
73% nonwhite. Moderate
follow-up participation but no
selective attrition. Mixed
effects models adjusted for
child sex, Fe status, and birth
weight, maternal race,
education, IQ, and prenatal
smoking, household income,
HOME score.
174 children born 1994-1995
followed from age 6 mo to
age 6 yr, Rochester, NY.
Prospective. Same cohort as
above. High follow-up
participation. Linear
regression model adjusted for
sex, birth weight, transferrin
saturation, maternal race, IQ,
education, and prenatal
smoking status, HOME score
(6 yr), family income.
1 ,333 children pooled from
Boston, Cincinnati,
Cleveland, Mexico City,
Port Pirie, Rochester, and
Yugoslavia cohorts.
Prospective. Large, uniform
analysis pooling diverse
cohorts. Included 84% of
eligible children. Linear
regression model adjusted for
HOME score, birth weight,
maternal IQ and education.
Considered child sex, marital
status, birth order, prenatal
alcohol consumption and
smoking, maternal age.
Blood Pb
Timing and
Levels
(ug/dL)
Earlier
childhood
(age 2 yr)
Mean (SD)
All subjects:
6.5 (4.9)
Peak <10:
3.8
(range: 1-9.3)
Concurrent
Mean (SD)
All subjects:
5.8 (4.1)
Peak<10:
3.3
(range:
0.5-8.4)
IV/I i n i m 1 1 m
IVIII III 1 IUI 1 1
below limit of
e ec ion
Peak
(6 mo-6 yr)
Mean (SD):
11.4(7.3)
Concurrent
Mean (95th)
Peak> 10:
13.9(35.4)
Peak<10:
4.3 (8.0)
Peak > 7.5:
12.9(34)
Pesk ^7 5'
3.2(6.0)'
Outcome
FSIQ
WISC-R
Age 1 0 yr
FSIQ
Stanford-
Binet
Age 5 yr
FSIQ
WPPSI-R
Age 6 yr
FSIQ
Various
tests
Ages
4.8-1 Oyr
Blood Pb stratum
(ug/dL)
All 148 subjects
48 subjects peak <10
All 172 subjects
101 subjects peak <10
Peak 20-30, n not given
Peak 10-20, n not given
96 subjects peak <1 0
1 ,089 subjects peak > 10
244 subjects peak <10
1 ,203 subjects peak > 7.5
1 03 subjects peak <7.5
Effect Estimate
/OCO/ ^l\^
(95% Cl)
-0.58 (-1 .0, -0.2)b
-1.56 (-2.9, -0.2)b
-0.61 (-0.99, -0.24)
-1.79 (-3.00, -0.60)
-0.15°
-0.32°
-1.2°
-0.1 3 (-2.3, -0.03)
-0.80 (-1.74, 0.14)
-0.1 6 (-0.24, -0.08)
-2.94 (-5. 16, -0.71)
November 2012
5-241
Draft - Do Not Cite or Quote
-------
Study
Tellez-Rojo
et al. (2006)
Tellez-Rojo
(2008)
Lanphear et
al. (2000)
Kordas et al.
(2006)
Study Population and
Methodological Details
384 children followed from
birth (1994-1995, 1997-1999)
to age 2 yr, Mexico City,
Mexico.
Cross-sectional.
Recruitment from prenatal
clinic or birth hospital.
Participants had higher
maternal education, lower
blood Pb level. Linear
regression model adjusted for
sex, birth weight, maternal IQ.
Considered maternal age and
other unspecified factors.
4,853 children ages 6-16 yr,
NHANES 1988-1994.
Cross-sectional. Large study
of multiple exposures and
outcomes. Linear regression
model adjusted for sex,
race/ethnicity, poverty index
ratio, reference adult
education level, serum ferritin
level, serum cotinine level.
Did not consider parental IQ
or caregiving quality.
532 children in 1st grade,
Torreon, Mexico
Cross-sectional.
Recruitment at prenatal clinic.
Residence near metal
foundry. High participation.
Linear regression model
adjusted for sex, age,
hemoglobin, family
possessions, forgetting
homework, house ownership,
crowding, maternal
education, birth order, family
structure, urine As, tester,
school. Did not consider
parental IQ or caregiving
quality.
Blood Pb
Timing and
Levels
(ug/dL)
Concurrent
Mean (SD)
> 10:
MR
Nrx
<10'
4.28 (2.25)
5-10:
6.9 (1 .4)
<5:
2.9(1.1)
Concurrent
Overall
II /!__._ /Q CV
Mean (oh).
1.9(0.1)
Subgroups:
NR
Concurrent
Overall
Mean (SD):
11.4(6.1)
Subgroups:
NR
Outcome
Bayley
MDI
Age 2 yr
Reading
Score
WRAT
Ages 6-1 6
vr
/ '
Math
score
Achieve-
ment test
1 st grade
Blood Pb stratum
(ug/dL)
90 subjects > 10
294 subjects <10
1 01 subjects 5-1 0
1 93 subjects <5
All 4,853 subjects
4681 subjects <10
4,526 subjects <7.5
4,043 subjects <5
2,467 subjects <2.5
All 532 subjects
293 subjects <10
Effect Estimate
/OCO/ ^l\^
(95% Cl)
0.07 (-10,9.2)"
-1.04 (-1.8, -0.30)b
-0.94 (-2.1, 0.2)b
-1.71 (-3.0, -0.42)b
-0.70 (-1 .03, -0.37)
-0.89 (-1 .52, -0.26)
-1 .06 (-1 .82, -0.30)
-1.06 (-2.00, -0.12)
-1 .28 (-3.20, -0.64)
-0.1 7 (-0.28, -0.06)
-0.42 (-0.92, 0.08)
FSIQ = Full-scale IQ, WISC-R = Wechsler Intelligence Scale for Children-Revised, WPPSI-R = Wechsler Preschool and Primary
Scale of Intelligence-Revised, MDI = Mental Developmental Index, NR = Not reported, WRAT = Wide Range Achievement Test.
"Effect estimates are derived from linear models and are presented for a 1 ug/dL increase in blood Pb level.
b95% CIs calculated from reported p-value.
°Results not included in Figure 5-16 because nonparametric analysis did not produce 95% CIs for various strata of blood Pb levels.
1
2
3
4
5
6
7
A few cross-sectional studies demonstrated larger Pb-associated decreases in cognitive
function with concurrent blood Pb levels <5 (ig/dL. Tellez-Rojo et al. (2006) estimated a
larger decrement in age 2 year Bayley MDI per unit increase in blood Pb level for
children with concurrent blood Pb levels <5 (ig/dL compared with children with blood Pb
levels 5-10 (ig/dL, and >10 (ig/dL (Figure 5-16 and Table 5-16). However, it is not clear
what the implications of age 2 year MDI results may be on cognitive function at later
ages. Among children ages 5-16 years participating in NHANES 1989-1994, Lanphear et
al. (2000) found larger decrements in reading and math skills and memory per unit
increase in blood Pb level in children with concurrent blood Pb levels <2.5 (ig/dL than
November 2012
5-242
Draft - Do Not Cite or Quote
-------
1 children with levels <5 (ig/dL, <7.5 (ig/dL, <10 (ig/dL, and all subjects. However, higher
2 Pb exposures earlier in childhood may have contributed to associations.
3 Several (Min et al.. 2009: Jusko et al.. 2008: Schnaas et al.. 2006) but not all
4 (Palaniappan et al., 2011) recent studies found a nonlinear blood Pb-cognitive function
5 relationship in nonparametric regression analyses using splines or lowess with smoothing
6 parameters that did not produce quantitative results for each blood Pb group. Similar to
7 the pooled analyses of the seven prospective cohorts, these relationship were evaluated
8 for a wide range of blood Pb levels. In the Rochester and Mexico City cohorts, the blood
9 Pb-FSIQ relationship was more negative for children with lower blood Pb levels,
10 specifically for peak blood Pb levels <10 (ig/dL (range: 2.1-45.7 (ig/dL) in the Rochester
11 cohort at age 6 years and (Jusko et al.. 2008) and for prenatal maternal week 28-36 blood
12 Pb levels <6(ig/dL (5th-95th percentile 2.5-24.6 (ig/dL) in the Mexico City cohort at ages
13 6-10 years (Schnaas et al.. 2006). In a formal test of nonlinearity, Schnaas et al. (2006)
14 found the nonlinear blood Pb term to fit the data better than a linear term. Among 267
15 children ages 4 years (blood Pb range: 1.3-23.8 (ig/dL) who had high prenatal alcohol
16 and drug exposure, Min et al. (2009) reported a p-value of 0.19 for a restricted cubic
17 spline term for blood Pb level and described the covariate-adjusted concurrent blood Pb
18 level-FSIQ curve to be more negative at blood Pb levels <7 (ig/dL. Among 814 children
19 in India ages 3-7 years, Palaniappan et al. (2011) mostly found linear associations
20 between concurrent blood Pb level (range: 2.6-40.5 (ig/dL) and indices of cognitive
21 function. The exception was visual-motor skills, for which a greater blood Pb-associated
22 decline was found with blood Pb levels >30 (ig/dL. The linearity versus nonlinearity of
23 the blood Pb-FSIQ concentration-response relationship within a lower, more narrow
24 range of blood Pb levels has not been examined in detail.
25 Few studies of adults have examined whether the relationship between blood or bone Pb
26 level and cognitive function is described better with a linear or nonlinear function. In
27 analyses of adults in NHANES, only log-linear models were used to fit the data (Krieg et
28 al..201Q: Krieg and Butler. 2009: Krieg et al.. 2009). Nonlinearity in the BMS and NAS
29 cohorts was examined with the use of quadratic terms, penalized splines, or visual
30 inspection of bivariate plots (Bandeen-Roche et al.. 2009: Weisskopf et al.. 2007b: Shih
31 et al.. 2006). There was some evidence for nonlinearity in prospective analyses of the
32 NAS cohort (Figure 5-8 and Figure 5-9), but not all results indicated greater declines in
33 cognitive function per unit increase in bone Pb level in the lower bone Pb groups. Wang
34 et al. (2007a) found that among NAS men with an HFE variant, there was a larger decline
35 in MMSE score per unit increase in tibia Pb level at higher tibia Pb levels, 20-25 ug/g
36 (Figure 5-9). In the BMS cohort, linear relationships were indicated for various tests of
37 cognitive function by a statistically nonsignificant quadratic term (Shih et al.. 2006) or
38 spline (Bandeen-Roche et al.. 2009) for tibia Pb level.
November 2012 5-243 Draft - Do Not Cite or Quote
-------
1 Although not specific to Pb exposure, attenuation of concentration-response relationships
2 at higher exposure or dose levels has been reported in the occupational literature, and
3 explanations have included greater exposure measurement error and saturation of
4 biological mechanisms at higher levels and larger proportions of at-risk populations at
5 lower exposure levels (Stavner et al.. 2003). Hypotheses for nonlinearity in the
6 relationship between Pb and cognitive function have included a lower incremental effect
7 of Pb due to covarying risk factors such as low SES, poor caregiving environment, higher
8 exposure to other environmental factors (Schwartz. 1994). different mechanisms
9 operating at different exposure levels, and confounding by omitted or misspecified
10 variables. The contribution of these factors to the supralinear relationship between blood
11 Pb levels and cognitive function in children has not been examined in many
12 epidemiologic studies to date. Several studies found that risk factors such as SES,
13 parental education, and parental caregiving quality explain a greater proportion of
14 variance in cognitive function than does blood Pb level (Wasserman et al.. 1997; Greene
15 et al.. 1992). Recently, among 57,678 fourth grade children across North Carolina,
16 Miranda et al. (2009) found that lower parental education and enrollment in a
17 free/reduced fee lunch program accounted for larger decrements in EOG scores than did
18 blood Pb level across the various quantiles of EOG score distribution (Figure 5-7).
19 Few studies have examined effect modification of the blood Pb level-cognitive function
20 relationship by covarying risk factors such as sociodemographic factors, and the limited
21 evidence is inconclusive. None of these studies examined effect modification within
22 specific strata of blood Pb levels. In the Boston cohort at age 57 months, a greater
23 Pb-associated FSIQ decrement was reported in the group that was female and had higher
24 HOME score, SES, and maternal IQ (Bellinger et al.. 1990). However, the Boston cohort
25 overall had higher SES and parental education, and the group that included higher SES
26 may not be comparable to other cohorts. In the Port Pirie, Australia cohort, larger blood
27 Pb-associated FSIQ decrements were found in groups with lower SES (Tong et al., 2000)
28 but not lower HOME score (McMichael et al.. 1992). Overall, evidence does not clearly
29 indicate whether the blood Pb-IQ relationship is modified by factors such as SES or other
30 sociodemographic characteristics or whether these differences can explain the observed
31 nonlinear concentration-response relationship.
32 Results from the pooled analysis by Rothenberg and Rothenberg (2005) do not indicate
33 that residual confounding by covariates explains the nonlinear blood Pb-FSIQ
34 relationship. Modeling maternal IQ, HOME score, and maternal education as spline
35 functions (df = 2) did not significantly improve model fit either with a linear blood Pb
36 term or log blood Pb term, which indicated that the improved model fit with log-
37 specification of blood Pb level was not influenced by the modeling of covariates as linear
38 or nonlinear functions.
November 2012 5-244 Draft - Do Not Cite or Quote
-------
1 Bowers and Beck (2006) postulated that a supralinear concentration-response function
2 necessarily will be found in a model with a log-normally distributed independent variable
3 and a normally distributed outcome variable. However, as discussed in the
4 2006 Pb AQCD, this modeling strategy was not employed in the epidemiologic analyses
5 showing a supralinear concentration-response function. FSIQ scores generally were not
6 forced into a normal distribution. Normalized FSIQ scores were not the basis for
7 individual findings from four of the seven studies included in the pooled analysis by
8 Lanphear et al. (2005) or the results pooling the seven cohorts (Hornung et al. 2006).
9 Further, a log-linear model (a linear relationship between IQ and the log of blood Pb)
10 provided a better fit of the pooled data.
11 Results from prospective analyses in the Boston and Rochester cohorts for associations
12 between blood Pb level as a continuous variable and FSIQ in groups of children in the
13 lower segment of the population blood Pb distribution have not identified a threshold in
14 the range of blood Pb levels examined. In the Boston cohort, higher age 2 year blood Pb
15 levels were associated with lower FSIQ at age 10 years in children with blood Pb levels
16 1-9.3 (ig/dL whose peak blood Pb levels never exceeded 10 (ig/dL (Bellinger. 2008;
17 Bellinger and Needleman. 2003). Schwartz (1994) explicitly assessed evidence for a
18 threshold in the Boston cohort data by regressing FSIQ and blood Pb level on age, race,
19 maternal IQ, SES, and HOME score and fitting a nonparametric smoothed curve to the
20 residuals of each regression model (variation in FSIQ or blood Pb level not explained by
21 covariates). A 7-point decrease in FSIQ was found over the range of blood Pb residuals
22 below 0 (corresponding to the mean blood Pb level of 6.5 (ig/dL), indicating an
23 association between blood Pb level and FSIQ down to a blood Pb level of 1 (ig/dL. In the
24 Rochester cohort, higher peak blood Pb levels were associated with lower FSIQ at ages 3
25 and 5 years in children with peak blood Pb levels < 10 (ig/dL (Canfield et al., 2003a). A
26 threshold also was not identified for the association between concurrent blood Pb level
27 and MDI score at age 2 years among children in Mexico City with blood Pb levels in the
28 range of 0.8-9.8 (ig/dL (Tellez-Roio. 2008: Tellez-Roio et al.. 2006).
29 In conjunction with downward trends in population blood Pb distributions (Figure 4-16).
30 more sensitive quantification methods have improved the detection limits for blood Pb
31 measurements (e.g., in NHANES, from 0.6 (ig/dL in 1999-2002 to 0.025 (ig/dL in
32 2003-2004). Consequently, the examination of groups of children (ages 8-11 years) with
33 lower blood Pb levels, overall range <1 to 16 (ig/dL, has indicated Pb-associated
34 cognitive function decrements or increases in attention-related behavioral problems, at
35 lower blood Pb levels (Cho et al.. 2010: Kim et al.. 2009b: Miranda etal.. 2009: 2007a).
36 In the studies examining concurrent blood Pb levels, the potential contribution of higher
37 past Pb exposures obscures assessment of a threshold. However, Miranda et al. (2009)
38 examined blood Pb levels measured between ages 6 and 36 months during 1995-1999
November 2012 5-245 Draft - Do Not Cite or Quote
-------
1 and found lower 9th grade EOG scores in 57,678 children in North Carolina with early
2 childhood blood Pb levels of 2 (ig/dL compared with children with blood Pb levels of
3 1 (ig/dL. Other studies did not identify a threshold for Pb-associated cognitive function
4 decrements in children using nonparametric regression analyses, but results have weaker
5 implications because the blood Pb levels in the examined populations of children were
6 higher than those in the current U.S. population (e.g., minimum 2.1, 5th percentiles 2.5
7 and 4.0 (ig/dL) (Jusko et al.. 2008; Schnaas et al.. 2006; Lanphear et al.. 2005).
8 Analyses of blood Pb level as a categorical variable did not clearly address the
9 identification of a threshold for the blood Pb-cognitive function relationship; however,
10 such analyses are not as sensitive as those of blood Pb level as a continuous variable. In
11 the analysis of large numbers (>600) of children participating in NHANES with blood Pb
12 levels <1 (ig/dL, Braun et al. (2008; 2006) found higher odds of parental reports of
13 conduct disorder and ADHD among children ages 4-15 years with concurrent blood Pb
14 levels ~1.0 (ig/dL compared with children with blood Pb levels <0.8 (ig/dL. However,
15 higher past Pb exposures may have contributed to associations found with concurrent
16 blood Pb levels because of the older age of some subjects and the birth of study
17 adolescents in the 1970s during the use of leaded gasoline. Other analyses of blood Pb
18 level categories indicated that cognitive function decrements were limited to children
19 ages 7-8 years with age 30-month blood Pb levels >10 (ig/dL (Chandramouli et al., 2009)
20 or children ages 6-10 years with concurrent blood Pb levels 5-10 (ig/dL (Surkan et al..
21 2007).
22 Some toxicological studies found nonlinear relationships between Pb exposure and
23 effects related to impaired learning and memory in animals. These results are distinct
24 from epidemiologic results as toxicological studies often show that lower and higher Pb
25 exposures have effects in opposite directions (U- or inverse U-shaped curves). Results
26 summarized across multiple studies in multiple species demonstrated that lower Pb
27 exposures increased FI response rates relative to controls, and higher Pb exposures
28 decreased FI response rates (Cory-Slechta. 1994). Increased FI response rates indicate
29 impaired learning by reflecting the impaired ability of animals to respond according to a
30 fixed schedule of reinforcement (Section 5.3.3.1). Consistent with previous findings,
31 Rossi-George et al. (2011) found that 50 ppm gestational plus lactational Pb exposure
32 when combined with stress increased FI responses of 2-month old rats whereas 150 ppm
33 Pb exposure with stress did not affect FI responses. Nonlinear effects of Pb on learning
34 are less consistently observed with longer duration exposures (e.g., 8-11 months) (Rossi-
35 George et al.. 2011; Cory-Slechta. 1990). These nonlinear effects of Pb on impaired
36 learning were supported by evidence in animals indicating that lower and higher Pb
37 exposures differentially activate underlying mechanisms. Gilbert et al. (1999) found
38 reduced LTP in adult rats exposed to 1,000 and 5,000 ppm but not 10,000 ppm Pb acetate
November 2012 5-246 Draft - Do Not Cite or Quote
-------
1 in drinking water (from GDI6). LTP is one indication of synaptic plasticity
2 (Section 5.3.11.4). which is considered to contribute to learning and memory. Learning
3 and memory have also been affected by glutaminergic neurotransmission via its NMDA
4 receptor (Section 5.3.11.8). and reduced glutamate release in the hippocampus was found
5 in adult rats exposed to Pb acetate from GDI5-GDI6 with blood Pb levels 27-40 (ig/dL
6 but not with blood Pb levels of 62-117 (ig/dL (Laslev and Gilbert. 2002).
7 Dopaminergic neurotransmission is involved in many CNS processes including
8 cognition, behavior, and motor function. The shape of the Pb-DA concentration-response
9 relationship varied among toxicological studies. Some studies found that lower Pb
10 exposures (~50 ppm) did not affect or increased DA activity relative to controls and
11 higher Pb exposure (109-250 ppm) (Leasure et al.. 2008; Virgolini et al.. 2005; Lewis and
12 Pitts. 2004). Others found higher Pb exposures (109 or 150 ppm) to increase or impair
13 DA activity (Leasure et al.. 2008; Virgolini et al.. 2005). These differential responses of
14 DA may be related to the diverse CNS effects of DA in different regions of the brain. For
15 example, the increased forebrain dopamine turnover with 27 ppm gestational/lactational
16 Pb acetate exposure was accompanied by less spontaneous activity in male mice
17 compared with male mice exposed to 109 ppm Pb (Leasure et al.. 2008).
18 In vitro results indicated differential effects on calcineurin enzyme activity, with inhibited
19 activity resulting from higher Pb exposure (>2 x 10"4 (iM) and stimulated activity from
20 lower Pb exposure (Kern and Audesirk. 2000). While calcineurin activity has been found
21 to modulate learning, LTP, and behavior in animals, studies have found lower calcineurin
22 activity to be associated with both impaired and improved effects related to learning
23 (Zeng et al.. 2001). Thus, it is uncertain whether altered calcineurin activity contributes to
24 the nonlinear relationships observed between Pb exposure and impaired learning in
25 animals. At lower concentrations, Pb may displace calcium at its binding sites on
26 calmodulin and by acting as a calmodulin agonist at the catalytic A subunit of calcineurin
27 and stimulate calcineurin activity. At higher Pb exposure, Pb may bind directly to a
28 separate calcium-binding B subunit, override the calmodulin-dependent effect and turn
29 off the activity of calcineurin. Lasley and Gilbert (2002) found that 2,000 ppm but not
30 5,000 or 10,000 ppm Pb acetate exposure of rats (in drinking water starting at
31 GD15-GD16) inhibited glutamate release by acting as a calcium mimetic.
32 Some toxicological studies have found nonlinear relationships for non-cognitive
33 outcomes in animals. U-shaped Pb concentration-response relationships were found for
34 spontaneous motor activity level and latency to fall from rotarod (Leasure et al.. 2008).
35 Inverted U-shaped relationships were found for hippocampal neurogenesis (Fox et al..
36 2008; Gilbert et al.. 2005). Evidence also points to differences in hormone production by
37 Pb exposure concentration. In male mice with long-term Pb exposure (PND21-9 months
November 2012 5-247 Draft - Do Not Cite or Quote
-------
1 of age), basal corticosterone levels were significantly lower with 50 ppm Pb than with
2 150 ppm Pb or controls (Cory-Slechta et al.. 2010). Visual system effects in animals also
3 have shown to be affected differentially by lower versus higher Pb exposure (GD1-
4 PND10, pup blood Pb levels 12, 24, and 46 (ig/dL). Inverted U-shaped concentration-
5 response curves were observed for rod photoreceptor numbers or neurogenesis
6 (Giddabasappa et al.. 2011) and retinal thickness (Fox et al.. 2010). These dichotomous
7 histological findings may give insight to the complex Pb-associated changes in ERG
8 wave amplitudes that vary by exposure window and dose (Section 5.3.7.3).
9 To conclude, several studies found a supralinear blood Pb-cognitive function
10 concentration-response relationship in children but not adults based on comparisons of
11 effect estimates in lower and higher strata of blood Pb level and nonparametric
12 regression. Explanations for this supralinear relationship have not been well characterized
13 by epidemiologic studies. Evidence from the prospective studies in the Boston and
14 Rochester cohorts has not identified a threshold for Pb-associated cognitive function
15 decrements in the range of blood Pb levels examined. Increases in childhood blood Pb
16 levels in the range of <1.0-9.8 ug/dL (means: 2.9 and 3.8 ug/dL) were associated with
17 cognitive function decrements at ages 3 to 10 years in children whose peak blood Pb
18 levels did not exceed 10 ug/dL (Bellinger. 2008: Canfield. 2008: Bellinger and
19 Needleman. 2003: Canfield et al.. 2003a). Further, a recent study found an association
20 between higher ages 6-36 month blood Pb levels (1995-1999) and lower 4th grade EOG
21 scores in 57,678 children in North Carolina with blood Pb levels 1-16 ug/dL (Miranda et
22 al.. 2009). Concurrent blood Pb levels in the range of 0.8-9.8 ug/dL were associated with
23 MDI decrements in children age 2 years in Mexico City (Tellez-Rojo, 2008: Tellez-Rojo
24 et al.. 2006). The lack of a reference population with blood Pb levels reflecting pre-
25 industrial Pb exposures limits the ability to identify a threshold. Analysis of ancient bones
26 in pre-industrialized societies suggests that "background" blood Pb levels in preindustrial
27 humans was approximately 0.016 (ig/dL (Flegal and Smith, 1992). approximately 65-fold
28 lower than that of the current U.S. population and lower than the levels at which
29 neurodevelopmental effects have been examined. Thus, the current evidence does not
30 preclude the possibility of a threshold for neurodevelopmental effects in children existing
31 with lower blood levels than those currently examined. While distinct from supralinear
32 relationships observed in epidemiologic studies, toxicological studies showed that lower
33 Pb exposures (e.g., 50 ppm in drinking water) induced learning and memory impairments
34 in animals compared to control exposures or higher Pb exposures (e.g., 150 ppm).
35 Additional toxicological evidence suggests that differentially activated mechanisms at
36 lower and higher Pb exposures and reduced LTP and hippocampal glutamate release with
37 lower Pb exposures may provide explanation for impaired learning observed with lower
38 Pb exposures in animals.
November 2012 5-248 Draft - Do Not Cite or Quote
-------
5.3.14 Confounding in Epidemiologic Studies of Nervous System Effects
1 In addition to Pb exposure, many factors influence cognitive function and behavior in
2 children, including parental IQ and education, SES of the family, quality of the
3 caregiving environment, and other environmental exposures (Wasserman and Factor-
4 Litvak. 2001). These other risk factors often are correlated with blood, tooth, and bone Pb
5 levels, thus, a major challenge to observational studies examining associations of Pb
6 biomarker levels with cognitive function and behaviors in children has been the
7 assessment and control for potential confounding factors. By definition, a confounder is
8 associated with both the independent variable and the outcome and consequently has the
9 potential to bias the association between the independent variable of interest and the
10 outcome. Most epidemiologic studies of Pb biomarkers in children have examined
11 potential confounding by parental IQ and SES-related variables such as parental
12 education, household income, and the Hollingshead Four-Factor Index of Social Position,
13 which incorporates education and income of both parents. Fewer but still several studies
14 have examined confounding by quality of the caregiving environment (i.e., HOME
15 score), birth weight, and smoking exposure. A relatively smaller number of studies have
16 considered nutritional status, other environmental exposures, parental substance abuse, or
17 parental psychopathology. Studies have varied with respect to the number of potential
18 confounding factors examined, with some studies considering multiple SES-related
19 variables and other studies focusing on a smaller set. The extent of confounding by a
20 particular factor likely varies across studies, depending on the population examined.
21 Thus, the impact of adjustment for specific covariates on the Pb effect estimate also
22 likely varies across studies.
23 Various methods have been used to control for potential confounding, including
24 examining a population relatively homogeneous in SES, examining populations in which
25 factors are not correlated, conducting multivariate regression, characterizing the change
26 in the blood Pb level effect estimate with adjustment for a covariate, and examining
27 associations in different strata of a covariate. The evidence derived from each of these
28 control strategies is discussed below. No single method is without limitation and
29 adjustment for SES is difficult as it is highly correlated with Pb exposure and there is no
30 single measure that represents SES. Residual confounding also is likely by factors not
31 considered. The combination of evidence from prospective studies that considered
32 several well-characterized potential confounding factors plus evidence that Pb exposure
33 induces impairments in cognitive function in animals, in particular, visual-spatial
34 memory and executive function, which are also found to be affected in children, increase
35 confidence that the associations observed between blood Pb and tooth Pb levels and
36 cognitive function in children represent a relationship with Pb exposure.
November 2012 5-249 Draft - Do Not Cite or Quote
-------
1 In the Boston prospective study, potential confounding by SES was controlled for by
2 study design and statistical adjustment for SES. The study subjects were from middle- to
3 upper-middle-class families, a majority with married, college-educated parents. Hence,
4 the potential for confounding by SES in this study was considerably less compared to
5 other studies examining similar outcomes. In this cohort, higher prenatal and concurrent
6 blood Pb levels were associated with FSIQ decrements at age 57 months, and higher age
7 2 year blood Pb levels were associated decrements in FSIQ and executive function at age
8 10 years (Stiles and Bellinger. 1993; Bellinger et al.. 1992; Bellinger et al. 1990). In
9 contrast, blood Pb levels were weakly associated with cognitive function decrements in
10 the Sydney, Australia cohort of middle-SES children (i.e., 20% mothers with greater than
11 high school education) (Cooney et al.. 1991). However, a relationship between Pb
12 exposure and cognitive function decrements is supported by a similar magnitude of blood
13 Pb-associated FSIQ decrement found in the Boston and low-SES Rochester cohort
14 (majority of mothers with less than college education and annual income <$15,000), with
15 adjustment for similar covariates (Figure 5-2 and Table 5-3) (Canfield et al.. 2003a). For
16 some outcomes, larger effects were estimated in the Boston cohort than in other cohorts.
17 Blood Pb levels also were associated with cognitive function decrements in populations
18 in which blood Pb levels were not correlated with SES-related factors (Factor-Litvak et
19 al.. 1999; Bellinger et al.. 1987). In the Yugoslavia cohort, blood Pb levels at age 4 years
20 were higher in groups with higher maternal education, maternal IQ, and HOME score in
21 one city near Pb sources and were lower in the distant city. Among all children, higher
22 blood Pb level was associated with lower FSIQ and learning and memory scores and with
23 higher ratings of internalizing behaviors (Factor-Litvak et al.. 1999). In the Boston
24 cohort, parental education, social class, and HOME score were similar among low
25 (<3 (ig/dL), medium (6-7 (ig/dL), and high (> 10 (ig/dL) cord blood Pb level groups.
26 Further, adjusting for these and other demographic variables, Bellinger et al. (1987)
27 found that children in the high cord blood Pb group had a 4.8-point lower Bayley MDI
28 score at age 2 years than did children in the low cord blood Pb group.
29 The primary method used by epidemiologic studies to control for potential confounding,
30 in particular recent studies of children with blood Pb levels more similar to current U.S.
31 levels, has been multivariate regression. Some studies modeled a set of covariates based
32 on a priori evidence, whereas others selected specific covariates based on their
33 association with the outcome in a model with all potential covariates and/or a greater than
34 10% change in the blood Pb level effect estimate. Studies also varied in the number of
35 potential confounding factors included in models. Some included multiple SES-related
36 variables, whereas others analyzed one or two factors. Regardless of the method used to
37 select model covariates or the number of covariates included, studies consistently found
38 associations of higher blood Pb level with cognitive function decrements and behavioral
November 2012 5-250 Draft - Do Not Cite or Quote
-------
1 problems. The evidence suggests that confounding by particular factors may vary across
2 populations and increases confidence that the associations observed with
3 neurodevelopmental effects in children represent a relationship with Pb exposure
4 The consistency of association across populations with different SES and co-exposures
5 and across studies examining different covariates was reinforced in pooled and meta-
6 analyses (Marcus etal. 2010; Lanphear et al., 2005; Schwartz. 1994). Pooling data from
7 seven international prospective cohorts, Lanphear et al. (2005) found similar FSIQ
8 decrements per log increase in blood Pb level (-2.6 to +8.6% difference) by excluding
9 one study at a time. These results indicated a relatively robust pooled estimate despite
10 between-study differences in population characteristics, including SES. In a meta-
11 analysis, Schwartz (1994) found a relatively narrow range of blood Pb-FSIQ effect
12 estimates among studies despite large between-study differences in the correlation
13 between blood Pb level and SES. A wider range of effect estimates would be expected if
14 omitted SES factors confounded the association. A recent meta-analysis of the
15 association between blood Pb level and conduct problems in earlier and recent studies of
16 children (Marcus etal. 2010) found that adjustment for SES and HOME score did little
17 to attenuate the association.
18 Among the several studies that provided both unadjusted and adjusted effect estimates,
19 most indicated that blood Pb level was a statistically significant predictor of cognitive
20 function (e.g., FSIQ, executive function, learning, memory) in children ages 5-10 years
21 before and after adjusting for potential confounders. Although most effect estimates
22 changed by 20-50% in multivariate models, they remained within the 95% CI of the
23 unadjusted estimate (Min et al.. 2009; Kordas et al.. 2006; Schnaas et al.. 2006; Canfield
24 et al.. 2003a; Dietrich et al.. 1993a; Bellinger etal., 1992). Such observations were made
25 in previous analyses of the Boston and Rochester cohort with mean blood Pb levels 6.5
26 and 5.8 (ig/dL, respectively, with adjustment for SES, maternal IQ and education, and
27 HOME score (Canfield et al.. 2003a; Bellinger et al.. 1992). These analyses also adjusted
28 for or considered potential confounding by nutritional factors. Recent studies of children
29 of a similar age range and mean blood Pb levels also found statistically significant
30 associations (as indicated by correlation and/or regression coefficients) between blood Pb
31 level and cognitive function before and after adjustment for similar covariates; however,
32 these populations had high prevalence of prenatal alcohol or drug use which may limit
33 the representativeness of their results (Min et al.. 2009; Chiodo et al.. 2007).
34 Blood Pb level also was a statistically significant predictor of cognitive function after
35 adjustment for covariates such as maternal education and IQ, SES, and HOME score in
36 with children with higher mean blood Pb levels, 8-14 (ig/dL (Kordas et al.. 2006; Schnaas
37 et al.. 2006; Tong and Lu. 2001; Dietrich et al.. 1993b). Exceptions include multiple
November 2012 5-251 Draft - Do Not Cite or Quote
-------
1 analyses of the Cleveland cohort, in which blood Pb level was estimated to have a weak
2 and imprecise or null effect after adjustment for potential confounding factors (Greene et
3 al.. 1992; Ernhart et al.. 1989; Ernhart et al.. 1988). Analyses of the Cleveland cohort
4 considered similar potential confounding factors as other studies, with the exception of
5 Greene et al. (1992) who also adjusted for pica and home conditions. However, these
6 latter variables each accounted for only a small partial correlation with FSIQ. HOME
7 score was the major factor accounting for the attenuation of the effect of Pb in the
8 Cleveland cohort. An analysis of the Yugoslavia cohort, which adjusted for most of the
9 same covariates as several of the Cleveland analyses reported larger magnitude blood
10 Pb-cognitive function associations in covariate-adjusted models (Factor-Litvak et al..
11 1999). The collective findings in children indicate potential confounding by the SES-
12 related and demographic factors examined in the literature base but also demonstrate that
13 blood Pb level is an independent predictor of cognitive function decrements in children
14 with adjustment for these factors.
15 A challenge to separating the effects of Pb exposure from those related to SES and
16 parental caregiving quality is their frequently high correlation with blood Pb levels. In
17 such cases, it is difficult to know how much variation in the outcome to attribute to each
18 of the risk factors (Needleman and Bellinger. 2001). For example, due to the high
19 correlation between blood Pb level and SES, a model that includes SES may
20 underestimate the Pb effect because some of the variance in outcome due to Pb is
21 mistakenly attributed to the variance due to SES. This misattribution may be exacerbated
22 when multiple correlated variables are included in the same model (i.e., overcontrol). The
23 relationships observed for Pb biomarker levels with SES and parental caregiving quality
24 may indicate that they are proxies or determinants of Pb exposure rather than a
25 confounder of the association of interest. Lower SES in urban children is closely linked
26 to residence in older, poorer condition housing that, in turn, may increase exposure of
27 children to environmental Pb and risk of cognitive deficits (Clark et al.. 1985). In such
28 cases where Pb exposure is a mediator of the SES effect, statistical adjustment for SES
29 will result in overcontrol of the Pb effect (Bellinger. 2004a). This type of overcontrol
30 could explain results from the New Zealand cohort, which were adjusted for residence in
31 older wooden housing, which is associated with higher exposure to Pb paint and
32 accumulated dust and soil and higher child tooth Pb levels (Fergusson et al.. 1988a. b).
33 However, even in models with older wooden housing, Pb remained a statistically
34 significant predictor of poorer reading skills and teacher ratings of school performance.
35 SES has been shown to be an effect modifier of the Pb-child cognitive function
36 relationship. Larger blood Pb-associated decreases in cognitive function were found with
37 lower SES in some studies (Ris et al.. 2004; Tong et al.. 2000; Bellinger et al.. 1990) and
38 higher SES in a meta-analysis (Schwartz. 1994). In cases of effect modification, potential
39 confounding by SES is less likely.
November 2012 5-252 Draft - Do Not Cite or Quote
-------
1 In summary, the collective epidemiologic evidence consistently demonstrates
2 associations of higher blood and tooth Pb levels with cognitive function decrements and
3 behavioral problems in children. These associations have been observed in diverse
4 populations in the U.S., Mexico, Europe, Asia, and Australia. Associations have been
5 observed across studies that use different methods to control for confounding and adjust
6 for different potential confounding factors but commonly, maternal IQ and education,
7 SES, and HOME score. Several studies have found associations with additional
8 adjustment for smoking exposure, child birth outcomes, and nutritional factors. No single
9 method to control for potential confounding is without limitation, and there is potential
10 for residual confounding by unmeasured factors. However, the consistency of findings
11 among different populations and study methods with consideration of several well-
12 characterized potential confounding factors as described above increases confidence that
13 the associations observed between blood Pb level and neurodevelopmental effects in
14 children represent a relationship with Pb exposure. Biological plausibility is provided by
15 the coherence with extensive evidence in animals with Pb exposures that produce blood
16 Pb levels relevant to humans and that is not subject to confounding by factors such as
17 social class and correlated environmental factors. Further, Pb exposure has been shown to
18 induce impairments in visual-spatial memory, rule learning and reversal, and response
19 inhibition, which also have been associated with blood or tooth Pb levels in children.
20 Additional support for the epidemiologic evidence is provided by extensive toxicological
21 evidence describing modes of action for Pb-induced cognition and behavioral problems,
22 including changes in neurogenesis, synaptic pruning, and neurotransmitter function in the
23 hippocampus, prefrontal cortex, and nucleus accumbens of the brain (Section 5.3.11).
5.3.15 Public Health Significance of Associations between Pb Biomarkers and
Neurodevelopmental Effects
24 As described in Section 5.3.2.1. most studies found that a 1 (ig/dL increase in blood Pb
25 level was associated with decrements in FSIQ in school-aged children in the range of <1
26 to 2 points, depending on the model and blood Pb level range examined (Figure 5-2 and
27 Table 5-3). Similarly, a 1 (ig/dL increase in blood Pb level typically was associated with
28 lower scores on tests of executive function (Table 5-8) and academic performance (Table
29 5-9). and higher ratings of behavioral problems (Figure 5-10. Table 5-11. Table 5-12) on
30 the order of less than 1 standard deviation. Such findings prompt consideration of the
31 public significance of blood Pb level-associated effects on cognitive function and
32 behavioral problems in children, specifically, whether the magnitudes of change have
33 consequences on the health and life-success of individuals. According to the WHO,
34 "Health is a state of complete physical, mental and social well-being and not merely the
35 absence of disease or infirmity" (WHO. 1948). By this definition, even decrements in
November 2012 5-253 Draft - Do Not Cite or Quote
-------
1 health status that are not severe enough to meet diagnostic criteria might be undesirable if
2 they reflect a decrement in the well being of an individual. Deficits in health indices or
3 life-success may not be observable except at the population level. The American
4 Thoracic Society discussed the need to consider the prevalence of exposures in the
5 population and exposure to other risk factors in evaluating whether shifts in the
6 population-level risk are adverse (ATS. 2000). Neurodevelopmental deficits measured in
7 childhood may set affected children on trajectories more prone toward lower educational
8 attainment and financial well-being. Thus, early deficits in children may have lifetime
9 consequences. There also may be groups in the population at increased risk of
10 neurodevelopmental deficits from Pb exposure. For example, some evidence points to
11 larger blood Pb-associated decrements in cognitive function in children with lower SES
12 (Ris et al.. 2004; Tong et al.. 2000; Bellinger et al.. 1990). whereas a meta-analysis found
13 a larger effect estimate for studies with higher SES (Schwartz. 1994).
14 It has been argued that blood Pb-associated decrements in IQ points less than 3 or 4
15 points are meaningless given that such changes are within the standard error of a single
16 test (i.e., the statistic that defines the range within which the true value of an individual is
17 likely to lie) (Kaufman. 2001). However, this argument incorrectly assumes that
18 conclusions drawn from individual-level data apply to populations. Evidence does not
19 indicate that the standard error is nonrandom, i.e., biased in one direction. Hence, there is
20 no reason to expect that children with higher blood Pb levels systematically test lower
21 than their true IQ value and that children with lower blood Pb levels test higher than their
22 true IQ value. Thus, in a population of children, on a given assessment, some children
23 will test lower than their true value and others will test higher than their true value. In
24 such cases, between-group differences will be measureable on a population basis. Error in
25 the measurement of IQ in an individual will contribute nondifferential error on a
26 population-level and bias the association to the null.
27 The issue of individual-level versus population-level risk also pertains to the relevance of
28 the magnitude of decrease in cognitive function or increase in behavioral problems per
29 unit increase in blood Pb level. Although fractional changes in IQ, memory, or attention-
30 related behavioral problems may not be consequential for an individual, they may be
31 consequential on a population level, especially in the two tails of the distribution
32 (Bellinger. 2007. 2004b). For example, interventions that shift the population mean, in a
33 beneficial direction, by an amount that is without clinical consequence for an individual
34 have been shown to produce substantial decreases in the percentage of individuals with
35 values that are clinically significant (Bellinger. 2007. 2004b). In statistical exercises not
36 specific to Pb or analysis of data collected from individuals, Weiss (1990. 1988)
37 predicted that a downward shift of five points in mean IQ, if the amount of dispersion in
38 the distribution remained the same, should be accompanied by a doubling of the numbers
November 2012 5-254 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
of individuals with scores two or more standard deviations below the mean. With a
reduction in population mean IQ from 100 to 95, the percentage of individuals predicted
to score above 130 (two standard deviations above the mean) decreases from 2.3% to
0.99%. Weiss (1988) stated that the implication of such a loss transcends the current
circumscribed definitions of risk. In addition to implications on the loss of intellectual
ability, the loss of a few IQ points potentially could result in the loss of academic
opportunities in children. For example, schools or programs for the gifted have used IQ
cut-offs (e.g., score of 130) to screen or accept applicants. In another hypothetical
analysis presented in the 2006 Pb AQCD (U.S. EPA. 2006b). based on a blood Pb-IQ
effect estimate of-0.9 points/(ig/dL (the median for blood Pb levels <10 (ig/dL), the
fraction of the population with an IQ <80 was estimated to more than double from 9%
with a blood Pb level of 0 (ig/dL to 23% with a blood Pb level of 10 (ig/dL (Figure 5-17).
The proportion with an IQ <70, which often requires community support to live (WHO.
1992). is predicted to increase from a little over 2% with a blood Pb level of 0 (ig/dL to
about 8% with a blood Pb level of 10 (ig/dL [(Figure 5-17) and (U.S. EPA.2006b)1.
These theoretical exercises estimate that for an individual in the low range of the IQ
distribution, a Pb-associated decline of a few points might be sufficient to drop that
individual into the range associated with increase risk of educational, vocational, and
social failure.
0.40
0.35
0.30
f 0.25
~ 0.20
c 0.15
B 0.10
O
™ 0.05
0.00
234567
Blood lead (pg/dL)
10
3 4 S 6 7
Blood Lead (pg/dL)
10
Note: The results presented in the figure are based on a theoretical analysis of changes in population IQ using a concentration-
response estimate of-0.9 IQ points/ug/dL, which was the median estimate from studies reviewed in the 2006 Pb AQCD for blood Pb
levels <10 ug/dL .
Source: 2006 Pb AQCD (U.S. EPA, 2006b).
Figure 5-17 Hypothetical effect of increasing blood Pb level on the proportion
of the population with IQ <70 and <80 points.
November 2012
5-255
Draft - Do Not Cite or Quote
-------
1 The hypothetical predictions presented in Weiss (1990. 1988) and the 2006 Pb AQCD
2 (U.S. EPA. 2006b) have been supported by findings from analyses using data from
3 studies of blood Pb levels and FSIQ decrements in children. Among children in 1st and
4 2nd grades from towns around Boston, Needleman et al. (1982) found that a 4-point
5 downward shift in the study population mean IQ estimated for tooth Pb levels >24 ppm
6 was associated with a 3-fold increase in the percentage of children with an IQ of <80 and
7 a decrease in the percentage achieving an IQ >125 from 5% to 0%.
8 The aforementioned hypothetical analyses and those using data collected from
9 individuals that estimate Pb-associated changes in the population IQ distribution assume
10 that the magnitude of change is equal across segments of the IQ distribution. Few studies
11 of Pb and cognitive function have examined whether the effect of Pb varies across the
12 distribution of cognitive function. However, in a recent study of fourth graders across the
13 entire state of North Carolina, Miranda et al. (2009) found that higher blood Pb level
14 measured once in each child between age 9 months and 3 years was associated with
15 larger decreases in fourth grade EOG scores in the lower segment of the EOG
16 distribution. An increase in blood Pb level from 1 to 10 (ig/dL was estimated to decrease
17 EOG score by 0.8 points in the 95th percentile of EOG scores but by 2.3 points in the 5th
18 percentile of EOG score (Figure 5-7). These findings by Miranda et al. (2009) based
19 analysis of a large database representative of fourth graders in North Carolina indicate
20 that a shift in the population mean from increased Pb exposure may increase the
21 proportions of children at the lower end of the cognitive function over that estimated by
22 theoretical analyses.
23 In summary, the public health significance of evidence demonstrating associations
24 between increases in blood Pb levels and decrements in IQ of children in the range of a
25 few points is supported by hypothetical predictions that a shift in the population mean
26 increases the proportion of individuals in the lower range of cognitive function and
27 decreases proportion of individual in the upper range of cognitive function. These
28 changes in the population distribution also were found in children in 1st and 2nd grade in
29 Massachusetts in whom higher tooth Pb level was associated with decrements in IQ
30 (Needleman et al.. 1982). Further support for the public health significance is provided by
31 findings that the blood Pb-associated decrement in cognitive function may be larger in
32 children in the lower range of cognitive function (Miranda et al., 2009) and in specific
33 groups within the populations such as those with lower SES (Ris et al.. 2004; Tong et al..
34 2000; Bellinger et al.. 1990). On a population-level, small Pb-associated decreases in
35 cognitive function could increase the number of individuals at increased risk of
36 educational, vocational, and social failure and decrease the number of individuals with
37 opportunities for academic and later-life success.
November 2012 5-256 Draft - Do Not Cite or Quote
-------
5.3.16 Summary and Causal Determination
1 The collective body of epidemiologic and toxicological evidence integrated across that
2 reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b) and recent studies indicates
3 relationships between Pb exposure and a range of nervous system effects. In children,
4 effects on cognitive function include FSIQ, learning, memory, executive function, and
5 academic performance. Outcomes evaluated related to attention-related behavioral
6 problems include inattention, impulsivity, hyperactivity, and ADHD. Effects on conduct
7 problems in children comprise aggression, delinquency, and criminal offenses. Effects on
8 internalizing behaviors include withdrawn behavior, depression-like symptoms,
9 fearfulness, and anxiety. Other nervous system effects evaluated in children are sensory
10 function and motor function. Relationships for Pb exposure with cognitive function and
11 sensory function also were evaluated in adults. Other nervous system effects in adults
12 examined in relation to Pb exposure include psychopathological effects such as
13 depression-like symptoms, anxiety, and panic disorder. Additionally, effects on
14 neurodegenerative diseases include Alzheimer's disease, ALS, Parkinson's disease, and
15 essential tremor. The subsequent sections describe the evaluation of evidence for each of
16 these outcome groups with respect to causal relationships with Pb exposure using the
17 framework described in Table II of the Preamble. The application of the key supporting
18 evidence to the causal framework is summarized in Table 5-17.
5.3.16.1 Evidence for Cognitive Function in Children
19 A causal relationship between Pb exposure and cognitive function decrements in children
20 is supported by evidence from prospective studies in diverse populations consistently
21 demonstrating associations of higher blood and tooth Pb levels with lower FSIQ and
22 performance on tests of executive function and academic performance in children ages
23 4-17 years (Section 5.3.2). coherence with evidence in animals for impairments in
24 learning, memory, and executive function with relevant Pb exposures, and evidence
25 describing modes of action (Table 5-17).
26 Clear support for Pb-associated cognitive function decrements in children, as described in
27 the 2006 Pb AQCD (U.S. EPA, 2006b). was provided by prospective epidemiologic
28 studies indicating associations of higher earlier childhood, concurrent, and childhood
29 average blood and tooth Pb levels with lower FSIQ in children ages 4-17 years (Table
30 5-17 and Section 5.3.2.1). Across studies, FSIQ was measured with similar instruments
31 (i.e., WISC-R, WISC-III, WPPSI, Stanford-Binet) scored on a similar scale with similar
32 measurement error. Associations were found in most of the prospective studies,
33 conducted in the U.S., Mexico, Europe, and Australia in representative populations, most
November 2012 5-257 Draft - Do Not Cite or Quote
-------
1 of which had moderate to high follow-up participation without indication of selective
2 participation among children with higher blood Pb levels and lower cognitive function
3 (Table 5-3). The prospective studies found associations of blood or tooth Pb levels
4 measured earlier in life (i.e., prenatal cord or maternal, age 2 year, age 6 year) and
5 averaged over multiple years with FSIQ decrements later in childhood (i.e., ages 4-17
6 years), better establishing the temporal sequence between Pb exposure and decrements in
7 cognitive function compared with cross-sectional analyses. Another strength of the
8 evidence from prospective studies was the consideration for several potential
9 confounding factors. As indicated in Table 5-3. results from most cohort studies were
10 adjusted for maternal IQ and education, child sex and birth weight, SES, and HOME
11 score. Although not considered as frequently, some studies also found associations with
12 adjustment for parental smoking and nutritional factors. The consistency and
13 reproducibility of the blood Pb-FSIQ association in children were substantiated in a
14 pooled analysis of seven prospective studies by Lanphear et al. (2005) as well as multiple
15 meta-analyses that combined results across various prospective and cross-sectional
16 studies (Pocock et al., 1994; Schwartz. 1994; Needleman and Gatsonis. 1990). with
17 Schwartz (1994) demonstrating the robustness of evidence to potential publication bias.
18 Among individual studies, a wide range of blood Pb-FSIQ effect estimates was obtained,
19 which is not unexpected given the wide range of blood Pb levels examined and modeling
20 methods used (i.e., linear, log-linear). The pooled analysis of seven prospective cohorts
21 demonstrated precision of effect estimates by applying a uniform method across
22 populations (Lanphear et al.. 2005). A narrow range of estimates was obtained by
23 excluding one study at a time, -2.4 to -2.9 points per log increase in concurrent blood Pb
24 level. Results from several individual studies indicated a supralinear concentration-
25 response relationship which estimated a greater decrement in cognitive function per unit
26 increase in blood Pb level among children in lower strata of blood Pb levels than children
27 in higher strata of blood Pb levels (Figure 5-16 and Table 5-16). Among the largest effect
28 estimates were found in the Boston and Rochester cohorts (Canfield et al.. 2003a;
29 Bellinger et al.. 1992). which had relatively smaller sample sizes but considered several
30 potential confounding factors as listed above and in Table 5-3. examined lower blood Pb
31 levels than did other prospective studies, and examined cohorts of different SES. Thus,
32 their results may be more generalizable. In the Boston cohort, a l(ig/dL increase in age 2
33 year blood Pb level was associated with a -1.6 (95% CI: -2.9, -0.2) point change in FSIQ
34 at age 10 years in 48 children with blood Pb levels 1-9.3 (ig/dL whose peak blood Pb
35 levels never exceeded 10 (ig/dL (Bellinger. 2008; Bellinger and Needleman. 2003). In the
36 Rochester cohort, a 1 (ig/dL increase in concurrent blood Pb level was associated a -1.8
37 (95% CI: -3.0, -0.60) change in FSIQ at age 5 years in 101 children with concurrent
38 blood Pb levels 0.5-8.4 (ig/dL, and peak blood Pb levels < 10 (ig/dL (Canfield. 2008;
39 Canfield et al.. 2003a). In the pooled analysis, a 1 (ig/dL increase in concurrent blood Pb
November 2012 5-258 Draft - Do Not Cite or Quote
-------
1 level was associated a -2.9 (95% CI: -5.2, -0.71) change in FSIQ at ages 4.8-10 years in
2 103 children with concurrent blood Pb levels 0.9-8.4 (ig/dL and peak blood Pb levels
3 <7.5 (ig/dL (Hornung. 2008; Lanphear et al.. 2005). These observations do not provide
4 evidence for a threshold in the ranges of blood Pb level examined. Null or weak
5 associations were limited to a few cohorts, namely, the Cleveland and Sydney cohorts
6 (Greene et al.. 1992: Coonevetal.. 1991: 1989a. b; Ernhart et al. 1988). The Cleveland
7 and Sydney studies were not outliers with respect to population mean blood Pb levels or
8 the specific confounding factors considered (Table 5-3). and the Cleveland cohort had
9 high prevalence of maternal prenatal substance abuse which may limit the
10 representativeness of results.
11 Other previous studies estimated smaller magnitude of effects for the blood Pb-FSIQ
12 association with either linear or log-linear models but examined higher blood Pb levels
13 (means: 8-16 (ig/dL) without analysis of the concentration response at the lower range of
14 the study population blood Pb distribution (Table 5-3). Recent cross-sectional studies
15 supported associations between higher concurrent blood Pb levels and decrements in
16 FSIQ. Among studies that examined populations with mean blood Pb levels <5 (ig/dL,
17 some lacked representative populations due to high prevalence of prenatal alcohol and/or
18 drug exposure (Chiodo et al.. 2007) or had limited consideration for potential
19 confounding (Zailina et al.. 2008). Kim et al. (2009b) estimated similar effects as the
20 Boston and Rochester studies although the log-linear model makes comparisons difficult.
21 Among children ages 8-11 years in Korea with mean blood Pb level 1.73 (ig/dL, a
22 1 (ig/dL increase in concurrent blood Pb level was associated with a 3.2-point decrease
23 (95% CI: -6.1, -0.23) in FSIQ among children ages 8-11 years in Korea with blood Mn
24 levels >1.4 (ig/dL in the 10th-90th percentile interval of blood Pb level (0.9-2.8 (ig/dL).
25 In this study, the potential influence of higher past Pb exposures cannot be excluded.
26 Among children ages 6-10 years in New England with a mean concurrent blood Pb level
27 2.2 (ig/dL, lower FSIQ was found in the group with blood Pb levels 5-10 (ig/dL (Surkan
28 et al.. 2007). While results from these studies were adjusted for SES and parental IQ or
29 education, parental caregiving quality was not examined.
30 A causal relationship between Pb exposure and cognitive function decrements in children
31 also is supported by previous prospective studies (several of which contributed to the
32 FSIQ evidence) that found associations of blood or tooth Pb level with decrements in
33 executive function and academic performance in children ages 4-18 years
34 (Sections 5.3.2.4. 5.3.2.5. and Table 5-17). The bodies of evidence for executive function,
35 and academic performance are smaller than that for FSIQ but consistently indicate
36 associations with blood or tooth Pb level. Associations with performance on tests of
37 learning and memory were less consistently found across populations (Section 5.3.2.3).
38 In most studies, previous and recent, multiple testing was common; however, the
November 2012 5-259 Draft - Do Not Cite or Quote
-------
1 consistent pattern of association observed across the ages of blood Pb level and/or
2 cognitive test examined increases confidence that the evidence is not unduly biased by an
3 increased probability of associations found by chance alone. While recent studies of
4 executive function and academic performance adjusted for SES and parental education
5 and/or IQ, few examined potential confounding by parental caregiving quality.
6 Adding to the evidence for Pb-associated cognitive function decrements in children were
7 recent prospective studies that indicated associations between higher earlier childhood
8 blood Pb level and poorer academic performance in school-aged children (Chandramouli
9 et al.. 2009; Miranda et al.. 2009). Among 57,678 children in North Carolina, lower
10 fourth grade EOG scores were found in children with age 3-36 month blood Pb levels
11 2 (ig/dL compared with children with blood Pb levels 1 (ig/dL, with adjustment for
12 parental education and enrollment in a free lunch program as an indicator of SES
13 (Miranda et al., 2009). In addition to finding associations with lower early childhood
14 blood Pb levels, this study indicated a greater incremental association of blood Pb level
15 with decrement in EOG score among children in the lower end of the EOG distribution.
16 Chandramouli et al. (2009) found decrements in school achievement tests in 488 children
17 ages 7-8 years in U.K. in children with age 30 month blood Pb levels >5 (ig/dL. These
18 results were adjusted for several potential confounding factors, including SES, parental
19 education, SES, home facilities score, and family adversity. Recent cross-sectional
20 studies conducted in the U.S. (Krieg etal.. 2010; Surkan et al.. 2007) found associations
21 of concurrent blood Pb level with decrements in executive function and academic
22 performance in children, including the large analysis of >700 children ages 12-16 years
23 participating in NHANES (Krieg etal.. 2010). Cho et al. (2010) did not find an
24 association between concurrent blood Pb level and executive function among children in
25 Korea ages 8-11 years with a mean blood Pb level 1.9 (ig/dL.
26 Several studies found associations of higher prenatal, earlier infancy, and concurrent
27 blood Pb levels with lower Bayley MDI scores in children ages 2 and 3 years (Table 5-4).
28 Similar to studies of FSIQ, Tellez-Rojo et al. (2006) estimated a larger decrement in age
29 2 year MDI per unit increase in concurrent blood Pb level for children in Mexico City
30 with blood Pb levels <5 (ig/dL compared with children with blood Pb levels 5-10 (ig/dL,
31 and >10 (ig/dL (Figure 5-16 and Table 5-16). MDI is a well-standardized measure of
32 current infant mental development. However, the test of MDI is not an intelligence test,
33 and MDI scores, particularly before ages 2-3 years, are not necessarily correlated with
34 later measurements of FSIQ in children with normal development.
35 A causal relationship between Pb exposure and cognitive function decrements in children
36 is further supported by consistent observations in animals of decrements in learning,
37 memory, and executive function with relevant dietary Pb exposures. In particular,
November 2012 5-260 Draft - Do Not Cite or Quote
-------
1 coherence was found between evidence in children and animals of Pb-associated
2 decrements in visual-spatial memory, working memory (Section 5.3.2.3) and rule
3 learning and reversal (Section 5.3.2.4). Previous studies in monkeys demonstrated
4 impairments in learning, memory, and executive function with dietary Pb exposures
5 during infancy only, lifetime after infancy, and lifetime from birth that produced blood
6 Pb levels of 19 and 26 ug/dL (Rice. 1992b. 1990: Rice and Karpinski. 1988).
7 Several recent toxicological studies added to the evidence for impaired learning and
8 memory in animals with lower blood Pb levels, 8-17 (ig/dL after gestational-lactational,
9 lactational, or lifetime (with and without gestational) Pb exposure (Cory-Slechta et al..
10 2010: Niu et al.. 2009: Virgolini et al.. 2008a: Stangle et al.. 2007). Together, the
11 prospective epidemiologic and toxicological studies provide evidence for the temporal
12 sequence between Pb exposure and decrements in cognitive function. Additional
13 biological plausibility for Pb-associated cognitive function decrements was provided by
14 toxicological evidence for the effects of Pb on modes of action for cognitive function
15 (Section 5.3.11). Pb has been shown to increase the permeability of the blood-brain
16 barrier and deposit in the target CNS. Pb has been shown to impair neurogenesis,
17 synaptic architecture, and neurite outgrowth. The high activity of these processes during
18 fetal and infant development provides biological plausibility for the effects of childhood
19 Pb exposure on decrements in cognitive function. Cognitive function is mediated by the
20 cortical and subcortical structures of the brain that integrate function in the hippocampus,
21 prefrontal cortex, and nucleus accumbens using dopamine and glutamate as primary
22 neurotransmitters. Experimental studies have shown that Pb induces changes in dopamine
23 and glutamate release in these regions and decreases the magnitude of LTP, which is a
24 major cellular mechanism underlying synaptic plasticity and learning and memory.
25 With regard to critical lifestages of Pb exposure, toxicological evidence clearly
26 demonstrates impaired learning and memory in animals exposed to Pb gestationally with
27 or without lactational exposure that produced blood Pb levels 8-17 (ig/dL. This evidence
28 is well supported by knowledge that processes such as neurogenesis, synaptogenesis, and
29 synaptic pruning are very active during this developmental period. However, evidence in
30 monkeys also indicates impaired cognitive function at ages 5-8 years with Pb exposure
31 starting after weaning (Rice. 1992b: Rice and Gilbert. 1990a: Rice. 1990: Rice and
32 Gilbert, 1990b). Epidemiologic studies also found cognitive function decrements
33 associated with blood Pb levels measured during various lifestages and time periods.
34 Distinguishing among the effects of Pb exposures at different time periods is difficult in
35 epidemiologic studies due to the high correlations commonly found among blood Pb
36 levels within children over time. Among studies of young children ages 6 months to
37 3 years, several found larger magnitudes of associations of MDI with prenatal maternal or
38 cord blood Pb than with postnatal child blood Pb (Hu et al.. 2006: Bellinger etal., 1987:
November 2012 5-261 Draft - Do Not Cite or Quote
-------
1 Dietrich et al., 1987a; Vimpani et al. 1985). However, in older children, ages 4-17 years,
2 in whom cognitive function is more stable and reliably measured, larger decrements in
3 cognitive function were found in association with postnatal blood Pb levels,
4 i.e., concurrent, earlier childhood, and cumulative average blood Pb levels as well with
5 tooth Pb levels. There was no clear indication of an individual critical lifestage, timing, or
6 duration of Pb exposure associated with cognitive function decrements in children.
7 Because of the contribution of bone Pb levels to concurrent blood Pb levels in children,
8 associations with concurrent blood Pb levels may reflect an effect of past and/or recent
9 Pb exposures.
10 The consideration for potential confounding varied among studies. Most studies adjusted
11 for SES-related variables such as the Hollingshead Index, household income, and/or
12 parental education. Several, in particular the prospective studies, additionally adjusted for
13 parental cognitive function and caregiving quality commonly evaluated as HOME score.
14 A few studies considered nutritional factors. Few recent studies considered potential
15 confounding by parental caregiving quality. The adjustment for SES is difficult as it is
16 highly correlated with Pb exposure and there is no single measure that represents SES.
17 Residual confounding also is possible by factors not considered. The combination of
18 evidence from prospective studies that considered several well-characterized potential
19 confounding factors plus evidence that Pb exposure induces impairments in cognitive
20 function in animals, in particular, for similar constructs as those associated with blood Pb
21 levels in children, increase confidence that the associations observed between blood Pb
22 levels and cognitive function in children represent a relationship with Pb exposure.
23 In conclusion, multiple prospective studies conducted in diverse populations consistently
24 demonstrate associations of higher blood and tooth Pb levels with lower FSIQ, executive
25 function, and academic performance and achievement. Most studies examined
26 representative populations and had moderate to high follow-up participation without
27 indication of selective participation among children with higher blood Pb levels and
28 lower cognitive function. Associations between blood Pb level and cognitive function
29 decrements were found with adjustment for several potential confounding factors, most
30 commonly, SES, parental IQ, parental education, and parental caregiving quality. In
31 children ages 4-11 years, associations were found with prenatal, early childhood,
32 childhood average, and concurrent blood Pb levels in populations with mean blood Pb
33 levels in the range 2-8 (ig/dL. Neither epidemiologic nor toxicological evidence has
34 identified an individual critical lifestage or duration of Pb exposure within childhood that
35 is associated with cognitive function decrements. Several epidemiologic studies found a
36 supralinear concentration-response relationship. Examination of children with blood Pb
37 levels in the range <1 to 10 (ig/dL, with consideration of early or peak childhood blood
38 Pb levels, has not identified a threshold for cognitive function decrements in the range of
November 2012 5-262 Draft - Do Not Cite or Quote
-------
1 blood Pb levels examined. Evidence in children was clearly supported by observations of
2 Pb-induced impairments in learning, memory, and executive function in juvenile animals.
3 Several studies in animals indicated learning impairments with prenatal, lactational, and
4 lifetime (with or without prenatal) Pb exposures that resulted in blood Pb levels of 8-
5 26 (ig/dL. The mode of action for Pb-associated cognitive function decrements is
6 supported by observations of Pb-induced impairments in neurogenesis, synaptogenesis
7 and synaptic pruning, LTP, and neurotransmitter function in the hippocampus, prefrontal
8 cortex, and nucleus accumbens. The associations consistently found for FSIQ and other
9 measures of cognitive function in prospective studies of children with adjustment for
10 SES, parental education, and caregiving quality and the biological plausibility provided
11 by evidence in animals for impairments in learning, memory, and executive function with
12 relevant Pb exposures and evidence describing modes of action is sufficient to conclude
13 that there is a causal relationship between Pb exposure and decrements in cognitive
14 function in children.
5.3.16.2 Evidence for Attention-related Behavioral Problems in Children
15 A causal relationship between Pb exposure and attention-related behavioral problems in
16 children is supported by evidence from prospective studies in diverse populations for
17 associations of blood or tooth Pb levels with inattention, impulsivity, and hyperactivity,
18 coherence with evidence in animals with relevant Pb exposures, and evidence describing
19 modes of action (Table 5-17). Although attention-related behavioral problems have been
20 examined less extensively than cognitive function, several epidemiologic and
21 toxicological studies have found associations with Pb. Prospective studies provided key
22 evidence for associations of childhood blood and tooth Pb levels with increases in
23 inattention, impulsivity, and hyperactivity in children ages 6-17 years and young adults
24 ages 19-24 years. These associations were found in populations in the U.S., U.K.,
25 Australia, and New Zealand (Table 5-17). Most studies had population-based recruitment
26 from prenatal clinics, hospitals at birth, or schools and had moderate to high participation.
27 A few prospective studies had increased loss-to-follow-up in certain groups, for example,
28 lower SES, earlier FSIQ, or HOME score. This potential selection bias may have reduced
29 the generalizability of findings to the original study population, but there was not a strong
30 indication that participation was biased to those with higher blood Pb levels and greater
31 behavioral problems. The most compelling evidence was that for inattention, impulsivity,
32 and hyperactivity assessed with neuropsychological testing or rated by parents or teachers
33 with widely-used structured questionnaires. A few studies found associations between
34 blood Pb level and diagnosis of ADHD but in studies that did not consider potential
35 confounding by parental caregiving quality.
November 2012 5-263 Draft - Do Not Cite or Quote
-------
1 Key evidence provided by prospective studies supported associations of blood and tooth
2 Pb levels with inattention and impulsivity as assessed using neuropsychological tests or
3 parent or teacher ratings (Section 5.3.3.1). Thus, the collective evidence does not appear
4 to be unduly influenced by biased reporting of such behaviors by parents of children with
5 higher blood Pb levels. Most studies that examined inattention with the continuous
6 performance test found associations with blood Pb level (Figure 5-10 and Table 5-11).
7 including previous prospective studies that indicated associations of higher prenatal or
8 earlier childhood blood Pb levels or tooth (from ages 5-8 years) Pb levels with increases
9 in commission and omission errors or reaction time in adolescents ages 15-17 years in
10 Cincinnati (Ris et al.. 2004) and young adults 19-24 years in Chelsea/Sommerville, MA
11 (Bellinger etal.. 1994a). Results from prospective studies also indicated associations with
12 parental and teacher ratings of inattention and impulsivity in children ages 8-13 years in
13 Australia, New Zealand, and Boston, MA (Burns etal.. 1999; Fergusson et al.. 1993;
14 Leviton et al., 1993). The mean blood Pb levels (prenatal cord, early childhood, lifetime
15 average) of populations examined in the prospective studies were in the range of
16 7-14 (ig/dL. The prospective findings better established the temporal sequence between
17 Pb exposure and inattention than did cross-sectional studies. Although the specific factors
18 varied by study, prospective studies of inattention and impulsivity considered several
19 potential confounding factors, including SES, parental IQ, maternal education, HOME
20 score, self drug use, prenatal drug and alcohol exposure, and birth outcomes. Evidence
21 did not strongly indicate associations between concurrent blood Pb levels and ratings of
22 inattention in younger children ages 4-5 years in Rochester and Yugoslavia (Canfield et
23 al.. 2003b: Wasserman et al.. 2001). These groups of younger children had lower blood
24 Pb levels, mean 6.5 (ig/dL; however, inattention may be less reliably rated in younger
25 children.
26 Consistent with previous prospective studies, recent cross-sectional studies found
27 associations of higher concurrent blood Pb level with increases in inattention as measured
28 by CPT, impulsivity using the stop task, and higher ratings of inattention and impulsivity
29 in children ages 8 to 12 years with mean concurrent blood Pb levels of 1 to 4 (ig/dL (Cho
30 etal.. 2010; Nicolescu etal.. 2010; Nigg et al.. 2008). However, the contribution of
31 higher Pb exposures earlier in childhood cannot be excluded. Further, while these recent
32 studies considered potential confounding by parental education, they had less consistent
33 consideration for other SES-related factors or parental caregiving quality than did
34 prospective studies. Some considered parental history of psychopathology; however, its
35 relationship with parental caregiving quality is not well characterized (Cho etal.. 2010;
36 Nicolescu et al.. 2010). Chiodo et al. (2007; 2004) found associations between concurrent
37 blood Pb level and increases in inattention as measured by the CPT and by independent
38 examiner ratings. Mean blood Pb levels were ~5 (ig/dL, and results were adjusted for
39 SES, parental education, and HOME score. However, the study population lacked
November 2012 5-264 Draft - Do Not Cite or Quote
-------
1 representativeness because of the high prevalence of prenatal alcohol and drug use.
2 Recent cross-sectional studies that included younger children (ages 3-5 years) also found
3 associations between concurrent blood Pb level and higher inattention as rated by
4 teachers or study examiners (Plusquellec et al. 2010; Roy et al.. 2009a): however, ratings
5 in young children may be less reliably measured.
6 A causal relationship between Pb exposure and attention-related behavioral problems also
7 is supported by a recent prospective study that found higher teacher ratings of
8 hyperactivity among children ages 7-8 years in the U.K. with age 30 month blood Pb
9 levels >10 (ig/dL (Chandramouli et al.. 2009). Previous findings were limited to cross-
10 sectional and case-control studies (Section 5.3.3.2). A strength of the recent prospective
11 study was the adjustment for several potential confounding factors, including maternal
12 education and smoking, SES, home facilities score, and family adversity index. Among
13 the recent cross-sectional studies, associations were found with consideration for
14 potential confounding by SES and maternal education; however, parental caregiving
15 quality was examined infrequently. Recent cross-sectional studies also found associations
16 of concurrent blood Pb level with higher parental ratings of a composite index of ADHD-
17 related behaviors, including the large U.S. representative analysis of 2,588 children
18 participating in NHANES which used DSM-IV criteria (Froehlich et al.. 2009). With the
19 exception of findings from the Rochester cohort (Canfield et al., 2003b). studies
20 generally found associations between blood Pb level and attention-related behavioral
21 problems with adjustment for child IQ or other measure of cognitive function (Cho et al..
22 2010: Chandramouli et al.. 2009: Nigg et al.. 2008: Silvaetal.. 1988). These findings add
23 support for higher Pb exposures having effects on attention-related behavioral problems,
24 independent of effects on cognitive function.
25 In the few available studies, concurrent blood Pb levels were associated with ADHD
26 prevalence in children (Section 5.3.3.4). Because of the cross-sectional or case-control
27 design of studies and lack of consideration for potential confounding by parental
28 caregiving quality or attention-related problems, the ADHD evidence is not a major
29 consideration in drawing conclusions about the relationship between Pb exposure and
30 attention-related behavioral problems.
31 Further support for a causal relationship between Pb exposure and attention-related
32 behavioral problems is provided by observations of impulsivity in animals in tests of
33 response inhibition (e.g., discrimination reversal learning, FR/waiting for reward) with
34 relevant dietary Pb exposures. In particular, coherence is found with observations in
35 children of associations between blood Pb levels and performance on the stop signal task,
36 which also measures response inhibition. Impulsivity in rodents and monkeys was found
37 with gestational and lactational dietary Pb exposures that resulted in blood Pb levels of 10
November 2012 5-265 Draft - Do Not Cite or Quote
-------
1 to 31 (ig/dL (Table 5-17 and Section 5.3.3.1). In animals, the effects of Pb exposure on
2 sustained attention were inconsistent as assessed with a signal-detection test with
3 distracting stimuli. In monkeys ages 9-10 years, lifetime dietary Pb exposure from birth
4 producing blood Pb levels 15 and 25 (ig/dL induced distractibility as assessed by poorer
5 performance on discrimination reversal tasks with distracting stimuli (Gilbert and Rice.
6 1987). Relevant dietary Pb exposures (i.e., producing blood Pb level of 10 (ig/dL) also
7 were found to increased activity in male (not female) mice with but only with
8 amphetamine co-treatment (Leasure et al.. 2008): thus, the findings may not be directly
9 comparable to observations of Pb-associated increases in hyperactivity in children.
10 Additional support for a causal relationship between Pb exposure and attention-related
11 behavioral problems is provided by evidence describing modes of action. Attention-
12 related behavioral problems have been linked with changes in the prefrontal cerebral
13 cortex, cerebellum, and hippocampus, and Pb exposure has been found to affect
14 development and neuronal processes in these regions. For example, Pb has been found to
15 affect dopaminergic neurons of the frontal cortex and striatum of the brain by altering
16 dopamine release and receptor density. Other lines of evidence supporting the mode of
17 action for the effects of Pb exposure on attention-related behavioral problems include
18 Pb-induced changes in neurogenesis, synapse formation, and synaptic plasticity.
19 In conclusion, although examined less extensively than cognitive function, several
20 prospective studies demonstrate associations of earlier childhood (e.g., age 6 years) and
21 lifetime average blood Pb levels or tooth (from ages 5-8 years) Pb levels with inattention,
22 impulsivity, and hyperactivity in children 7-17 years and young adults ages 19-24 years
23 as assessed using objective neuropsychological tests and rated by parents and teachers.
24 Most of these prospective studies examined representative populations without indication
25 of participation conditional on blood Pb levels and behavioral problems. The results from
26 prospective studies were adjusted for potential confounding by SES and parental
27 caregiving quality, with a few studies also considering substance abuse and nutritional
28 status. Blood Pb-associated increases in attention-related behavioral problems were found
29 in populations with earlier childhood (age 6 years) or lifetime average (to age 11-13
30 years) mean blood Pb levels of 7 and 14 (ig/dL and groups with earlier childhood (age
31 30 months) blood Pb levels >10 (ig/dL. Several cross-sectional studies found associations
32 between concurrent blood Pb level (means 1-4 (ig/dL) and attention-related behavioral
33 problems in children ages 8-12 years but had less consistent adjustment for SES and
34 parental caregiving quality. Biological plausibility for observations in children is
35 provided by several findings in animals for increases in impulsivity or impaired response
36 inhibition with relevant prenatal, lactational, and lifetime Pb exposures that resulted in
37 blood Pb levels of 10 to 31 (ig/dL. The mode of action for Pb-associated attention-related
38 behavioral problems is supported by observations of Pb-induced impairments in
November 2012 5-266 Draft - Do Not Cite or Quote
-------
1 neurogenesis, synaptic pruning, and dopamine transmission in the prefrontal cerebral
2 cortex, cerebellum, and hippocampus. The consistency of epidemiologic evidence,
3 particularly for inattention, impulsivity, and hyperactivity in prospective studies, and the
4 biological plausibility provided by evidence for Pb-induced impulsivity in animals and
5 for underlying modes of action is sufficient to conclude that there is a causal relationship
6 between Pb exposure and attention-related behavioral problems in children.
5.3.16.3 Evidence for Conduct Problems in Children and Young Adults
7 Epidemiologic evidence indicates that a causal relationship is likely to exist between Pb
8 exposure and misconduct in children and young adults. Key evidence is provided by
9 recent prospective studies finding associations of higher earlier childhood blood and
10 tooth (from ages 6-8 years) Pb levels with criminal offenses in young adults in Cincinnati
11 and Christchurch, New Zealand, ages 19-24 years, as assessed through government
12 records (Fergusson et al., 2008; Wright et al., 2008). In the Cincinnati cohort, a 1 (ig/dL
13 increase in age 6 year blood Pb level (mean 6.8 (ig/dL) was associated with an increased
14 risk of criminal arrests at age 19-24 years with an RR of 1.05 (95% CI: 1.01, 1.09).
15 Additional support was provided by most previous and recent prospective studies that
16 found associations of blood or tooth Pb levels with higher parent and teacher ratings of
17 delinquent behavior, aggression, antisocial activities, and destructive behavior in children
18 ages 8-17 years from diverse locations and SES (i.e., U.K., Cincinnati, Port Pirie,
19 Australia) (Table 5-12 and Table 5-17). Associations were found with lifetime average
20 blood Pb levels in boys ages 11-13 years in Port Pirie, Australia with a mean blood Pb
21 level 14 (ig/dL (Burns etal.. 1999) and with age 30 month blood Pb levels >10 (ig/dL in
22 children ages 7-8 years in the U.K. (Chandramouli et al.. 2009). The moderate to high
23 follow-up participation and associations found with parent and teacher ratings of conduct
24 problems do not provide strong evidence for biased participation or reporting of conduct
25 problems for children with higher blood Pb levels. Studies of criminal offenses and
26 ratings of conduct problems found associations with adjustment for several potential
27 confounding factors such as SES, smoking, drug, and alcohol exposure, and parental
28 caregiving quality.
29 Supporting evidence was provided by the large cross-sectional analysis of 2,867
30 adolescents ages 8-15 years participating in NHANES, which found that compared with
31 children with concurrent blood Pb levels <0.8 (ig/dL, children with concurrent blood Pb
32 levels 0.8-1.0 (ig/dL had higher odds of conduct disorder as assessed by parents with
33 adjustment for age, sex, race, poverty to income ratio, and smoking exposure (Braun et
34 al., 2008). Parental caregiving quality was not examined. These associations observed in
35 adolescents with relatively low concurrent blood Pb levels could have been influenced by
November 2012 5-267 Draft - Do Not Cite or Quote
-------
1 higher past Pb exposures. Further supporting the consistency of association between
2 blood Pb levels and conduct problems in children, a recent meta-analysis found that
3 evidence was robust to heterogeneity in study design, definition and assessment method
4 of conduct problems, potential confounding variables examined, and range of blood Pb
5 levels (Marcus etal.. 2010). Evidence of Pb-induced aggression in animals was
6 inconsistent, with increases in aggression found in some studies of adult animals with
7 gestational plus lifetime Pb exposure but not juvenile animals.
8 Associations of conduct problems in children and young adults with earlier childhood,
9 earlier childhood average, and lifetime average blood Pb levels, tooth Pb levels, and bone
10 Pb levels point to the effects of early childhood or cumulative Pb exposures. Most
11 prospective studies did not analyze Pb biomarker levels at multiple lifestages or time
12 periods and thus did not provide information on potential associations with more recent
13 blood Pb measurements or differences in association among Pb biomarkers at various
14 time periods. With respect to blood Pb levels, an association with criminal offenses was
15 found in young adults ages 19-24 years with a mean age 6 year blood Pb level of
16 6.8 (ig/dL, and associations with ratings of conduct problems were found in children ages
17 7-8 years with age 30 month blood Pb levels >10 (ig/dL and boys ages 11-13 years with a
18 mean lifetime average blood Pb level of ~14 (ig/dL (Table 5-17).
19 In conclusion, the few prospective studies consistently indicate that earlier childhood (age
20 30 months) or lifetime average (to age 11-13 years) blood Pb levels or tooth (from ages
21 6-8 years) Pb levels are associated with criminal offenses in young adults ages 19-24
22 years and with higher parent and teacher ratings of conduct problems in children ages
23 7-17 years. These associations were found without indication of strong selection bias and
24 with adjustment for SES, parental education and IQ, parental caregiving quality, family
25 functioning, smoking, and substance abuse. Supporting evidence is provided by cross-
26 sectional evidence of children participating in NHANES and a meta-analysis of
27 prospective and cross-sectional studies. Evidence for Pb-induced aggression in animals is
28 mixed. The consistent epidemiologic evidence from prospective and cross-sectional
29 studies for criminal offenses and ratings of misconduct but lack of clear evidence for
30 aggression in animals is sufficient to conclude that a causal relationship is likely to exist
31 between Pb exposure and conduct problems in children and young adults.
5.3.16.4 Evidence for Internalizing Behaviors in Children
32 Epidemiologic and toxicological evidence indicates that a causal relationship is likely to
33 exist between Pb exposures in children and internalizing behaviors, including withdrawn
34 behavior, fearfulness, and symptoms of depression and anxiety. Internalizing behaviors
November 2012 5-268 Draft - Do Not Cite or Quote
-------
1 have been examined to a lesser extent than cognitive function and attention-related
2 behaviors. However, supporting evidence is provided by a few previous prospective
3 studies that found associations of higher lifetime average blood or tooth Pb levels with
4 higher parent and teacher ratings of withdrawn behavior or symptoms of depression in
5 school-aged children, 8-13 years (Burns etal.. 1999; Bellinger et al.. 1994b) (Table
6 5-17). These prospective studies followed children from birth and had moderate follow-
7 up participation to later childhood. Participation was not conditional on early childhood
8 blood Pb levels, and associations were found with both parent and teacher ratings of
9 internalizing behaviors, reducing the likelihood of undue influence by biased
10 participation and ratings of internalizing behaviors by parents of children with higher Pb
11 exposures. Internalizing behaviors were assessed with widely-used structured
12 questionnaires such as the Child Behavior Checklist but not assessed as clinically-
13 diagnosed conditions such as depression.
14 The analysis of the Port Pirie, Australia cohort had the most extensive consideration of
15 potential confounding. Among only the 163 girls, ages 11-13 years, Burns et al. (1999)
16 found that 1 (ig/dL increase in lifetime average blood Pb level was associated with an
17 increased odds of an anxious/depressed parental rating above the median of 1.04 (95%
18 CI: 1.0, 1.09) with the adjustment for several SES-related variables and factors related to
19 parental caregiving including HOME score, family functioning score, and current
20 maternal psychopathology. In a Boston-area cohort, Pb level in deciduous teeth collected
21 around age 6 years was associated with a higher teacher rating of a composite of anxious
22 and social withdrawn behaviors in children ages 8 years with adjustment for receiving
23 public assistance at birth and maternal education but not parental caregiving quality
24 (Bellinger etal.. 1994b). In the Yugoslavia cohort, higher lifetime average blood Pb
25 levels were associated with higher maternal ratings of anxious-depressed and withdrawn
26 behaviors in 191 children ages 4-5 years with a mean blood Pb level ~8 (ig/dL, with
27 stronger associations found with delinquent behaviors (Wasserman et al.. 2001).
28 Behavior ratings may be less reliable in these younger children. With respect to critical
29 lifestages and durations of Pb exposure, evidence from prospective studies for
30 associations with tooth Pb levels and lifetime average blood Pb levels indicates an effect
31 of cumulative childhood exposure on increasing internalizing behaviors in children.
32 Cross-sectional studies, including several recent studies, indicated associations between
33 concurrent blood Pb levels and internalizing behaviors in children ages 3-16 years, but
34 most did not consider potential confounding by parental caregiving quality
35 (Section 5.3.5.1). Previously, Chiodo et al. (2004) found that among children age 7 years
36 in Detroit, MI (mean blood Pb level: 5 (ig/dL), HOME score, SES, maternal education,
37 and prenatal alcohol and drug exposure did not influence associations between blood Pb
38 level and internalizing behaviors; however, the population lacks representativeness
November 2012 5-269 Draft - Do Not Cite or Quote
-------
1 because of the high prevalence of prenatal alcohol or drug exposure (Table 5-17). A
2 recent study reported a lack of association between concurrent blood Pb level and
3 internalizing behaviors in Inuit children age 5 years in Quebec, Canada with a mean
4 blood Pb level of ~5 (ig/dL with consideration of potential confounding by HOME score,
5 caregiver education and IQ, blood Hg levels, and prenatal smoking and alcohol exposure
6 (Plusauellecetal.. 2010).
7 Supporting evidence for a relationship between Pb exposure and internalizing behaviors
8 is provided by the coherence of epidemiologic findings in children with evidence in
9 rodents that dietary prenatal plus lactational or lactational only Pb exposure resulted in
10 depression-like and loss of motivation behavior in rodents, in some cases with blood Pb
11 levels relevant to humans (13-17 (ig/dL) (Beaudin et al.. 2007; Dyatlov and Lawrence,
12 2002). Other studies found Pb-induced increases in emotionality, depression, and tactile
13 defensiveness in animals with blood Pb levels >30 (ig/dL after gestational and/or
14 lactational Pb exposure (Section 5.3.5.2). Biological plausibility for Pb-associated
15 increases in internalizing behaviors also is provided by evidence that describes mode of
16 action, including Pb-induced changes in the HPA axis (Section 5.3.2.3) and dopaminergic
17 and GABAergic systems (Sections 5.2.2.2. 5.3.11.4. and 5.3.11.8). which are found to
18 affect mood and emotional state.
19 In conclusion, prospective studies in a few populations demonstrate associations of
20 higher lifetime average blood (mean: ~14 (ig/dL) or childhood tooth (from ages 6-8
21 years) Pb levels with higher parent and teacher ratings of internalizing behaviors such as
22 depression, anxiety, and withdrawn behavior in children ages 8-13 years. The lack of
23 selective participation by blood Pb level and associations found with parental and teacher
24 ratings do not provide strong indication of biased reporting of behaviors for children with
25 higher blood Pb levels. While results were adjusted for maternal education and SES-
26 related variables, consideration for potential confounding by parental caregiving quality
27 was inconsistent. The biological plausibility for the effects of Pb on internalizing
28 behaviors is provided by consistent findings in animals with dietary prenatal plus
29 lactational or lactational only Pb exposure, with some evidence at blood Pb levels
30 relevant to humans. Additional toxicological evidence supports modes of action,
31 including Pb-induced changes in the HPA axis and dopaminergic and GABAergic
32 systems. The evidence from prospective studies in a few populations and the supporting
33 toxicological evidence with some uncertainty related to potential confounding by parental
34 caregiving quality in studies of children is sufficient to conclude that a causal relationship
35 is likely to exist between Pb exposure and internalizing behaviors in children.
November 2012 5-270 Draft - Do Not Cite or Quote
-------
5.3.16.5 Evidence for Sensory Function in Children
1 A causal relationship is likely to exist between Pb exposure and sensory function
2 decrements in children based on evidence from a previous prospective study indicating
3 associations of increased hearing thresholds with early childhood and lifetime average
4 blood Pb levels in 215 children age 5 years (Dietrich et al.. 1992) and previous large
5 (n = >3,000) cross-sectional NHANES and HHANES studies for concurrent blood Pb
6 levels in children ages 4-19 years (Schwartz and Otto. 1991. 1987). The high follow-up
7 participation from birth in the Cincinnati cohort and the examination of multiple
8 exposures and outcomes in NHANES and HHANES reduce the likelihood of biased
9 participation by children with high blood Pb levels.
10 The epidemiologic evidence in children is strengthened by the consideration for several
11 potential confounding factors. In the Cincinnati cohort at age 5 years, higher prenatal
12 maternal, neonatal (10 day, mean 4.8 (ig/dL), and lifetime average (mean: 17.4 (ig/dL)
13 blood Pb levels were associated with higher hearing thresholds with adjustment for SES,
14 HOME score, several birth outcomes, and maternal alcohol consumption and
15 consideration for factors such as maternal smoking and child health (Dietrich et al..
16 1992). In NHANES and HHANES, higher concurrent blood Pb levels (median: 8 (ig/dL)
17 were associated with increased hearing thresholds with adjustment for age, sex, race,
18 family income, parental education, and nutritional factors (Schwartz and Otto. 1991.
19 1987).
20 Additional support for a relationship between Pb exposure and sensory function
21 decrements in children is provided by evidence supporting modes of action. A previous
22 prospective study in children in Mexico City (n = 100, 113) found associations of
23 prenatal maternal (1-8 (ig/dL) and age 1 and 4 year blood Pb levels (age 2 year mean:
24 10.8 (ig/dL) with lower auditory evoked potentials (Rothenberg et al.. 2000). Increased
25 thresholds and increased latencies in brainstem auditory evoked potentials were also
26 found in nonhuman primates ages 8-13 years with long-term (multiple years) postnatal Pb
27 exposure beginning at birth (Rice. 1997; Lilienthal and Winneke. 1996). Pb exposure
28 from gestation through age 5 months was found to have weaker effects (Laughlin et al..
29 2009). In animals, auditory effects were examined with higher Pb exposures than those
30 relevant to the current U.S. general population (i.e., resultant blood Pb levels
31 33-170 (ig/dL); thus, it is difficult to assess coherence with observations in children.
32 Toxicological studies demonstrated a range of effects on the visual system including
33 impaired visual function, and potential mechanisms such as alterations in morphology
34 and cell architecture, signaling, enzyme inhibition, neurotransmitter levels,
35 neuroanatomical development, cell proliferation, and retinal cell apoptosis. An
36 epidemiologic study in children (Rothenberg et al.. 2002b) and toxicological studies in
November 2012 5-271 Draft - Do Not Cite or Quote
-------
1 rats found Pb-associated supernormal retinal ERGs (Fox et al.. 2008). Animal studies
2 also showed subnormal ERGs depending on lifestage of Pb exposure and blood Pb level.
3 Because the implications of supernormal ERGs on vision are not clear, the retinal ERG
4 findings are not a major consideration in drawing conclusions about Pb exposure and
5 sensory function decrements in children.
6 In conclusion, evidence from a prospective study and cross-sectional studies in a few
7 populations indicates associations of higher blood Pb level with increases in hearing
8 thresholds and decreases in auditory evoked potentials with adjustment for potential
9 confounding by SES in most studies and by child health and nutritional factors in some
10 studies. The high participation rates, particularly in the prospective study with follow-up
11 from birth, reduce the likelihood of biased participation by children with higher blood Pb
12 levels. Across studies, associations were found with blood Pb levels measured at various
13 time periods, including prenatal maternal, neonatal (10 day, mean 4.8 (ig/dL), lifetime
14 average (to age 5 years, mean 17.4 (ig/dL), and concurrent (ages 4-19 years) blood Pb
15 levels (median 8 (ig/dL). Findings in monkeys ages 8-13 years indicate increases hearing
16 thresholds and latencies for auditory evoked potentials with lifetime postnatal dietary Pb
17 exposure, albeit with higher blood Pb levels (i.e., 33-107 (ig/dL) than those relevant to
18 humans. The evidence in children, particularly that from a prospective study, but
19 uncertainties related to effects on auditory function in animals with relevant Pb
20 exposures, is sufficient to conclude that a causal relationship is likely to exist between Pb
21 exposure and decrements in sensory function in children.
5.3.16.6 Evidence for Motor Function in Children
22 A causal relationship is likely to exist between Pb exposure and motor function
23 decrements in children based on evidence from previous prospective epidemiologic
24 studies and supporting toxicological evidence. In the Cincinnati cohort, higher neonatal,
25 concurrent, and lifetime average blood Pb levels were associated with poorer fine and
26 gross motor function in 245 children ages 6 years (Dietrich et al., 1993a). and higher age
27 0-5 year average and 78 month blood Pb levels were associated with poorer fine motor
28 function in 195 children ages 15-17 years (Ris et al., 2004). In the Yugoslavian cohort,
29 higher lifetime average blood Pb level was associated with decrements in fine but not
30 gross motor function at age 4.5 years in 283 children (Wasserman et al., 2000). These
31 studies had high follow-up participation from birth or infancy reducing the likelihood of
32 biased participation by children with higher blood Pb levels. Motor function was assessed
33 using varied but widely-used, structured tests. The evidence from the Cincinnati and
34 Yugoslavia cohorts is substantiated by the consideration of several potential confounding
35 factors such as SES, parental caregiving quality, child health, and in adolescents,
November 2012 5-272 Draft - Do Not Cite or Quote
-------
1 marijuana use (Ris et al., 2004). In the prospective studies, mean concurrent and
2 childhood average blood Pb levels mostly ranged from 11 to 28 (ig/dL, higher than those
3 in most of the current U.S. population. Recent cross-sectional studies examining lower
4 concurrent blood Pb levels, means 2-5 (ig/dL, produced contrasting associations (Surkan
5 et al., 2007; Despres et al., 2005). An association between concurrent blood Pb level and
6 poorer motor function was found with adjustment for several potential confounding
7 factors including SES, parental caregiving quality, and blood levels of Hg and
8 polychlorinated biphenyls in 110 Inuit children living in subsistence fishing communities
9 (Despres et al., 2005). Higher blood Pb level was associated with improved motor
10 function in a more representative population of 534 children ages 6-10 years in New
11 England (Surkan et al. 2007).
12 Epidemiologic evidence is supported by observations of poorer performance on the
13 rotarod balance test in male (not female) mice with relevant blood Pb levels,
14 i.e., 10 (ig/dL after dietary Pb exposure from gestation to PND10 (Leasure et al.. 2008).
15 Other toxicological studies produced mixed results for effects on endurance, balance, and
16 coordination (Section 5.3.8) but are less relevant to humans because of the higher
17 concentrations of Pb exposure examined, i.e., those producing blood Pb levels
18 >30 (ig/dL.
19 In conclusion, evidence from prospective and cross-sectional studies in a few populations
20 indicates associations of decrements in fine and gross motor function with higher blood
21 Pb levels measured earlier in childhood (ages 0-5 year average, age 78 months) in
22 children ages 15-17 years or lifetime average blood Pb levels in children ages 4.5 years
23 with adjustment for several potential confounding factors, including SES, parental
24 caregiving quality, and child health. The prospective studies had high follow-up
25 participation from birth or early infancy, reducing the likelihood of biased participation
26 by children with higher blood Pb levels. The biological plausibility for associations
27 observed in children is provided by a study that found poorer balance in male mice with
28 relevant gestational to early postnatal (PND10) Pb exposures. The evidence in children,
29 particularly from a few prospective studies, and the coherence with limited available
30 findings in mice is sufficient to conclude that a causal relationship is likely to exist
31 between Pb exposure and decrements in motor function in children.
5.3.16.7 Evidence for Cognitive Function in Adults
32 Epidemiologic and toxicological evidence indicates that a causal relationship is likely to
33 exist between Pb exposure and cognitive function decrements in adults based primarily
34 on recent prospective and cross-sectional studies that indicate associations with bone Pb
November 2012 5-273 Draft - Do Not Cite or Quote
-------
1 level (Table 5-17). Key evidence for bone Pb levels comprised prospective analyses of
2 the BMS and NAS, with support provided by the cross-sectional Nurses' Health Study.
3 The multiple risk factors and health outcomes examined in these studies reduces the
4 likelihood of biased participation and/or follow-up by adults with higher Pb exposure and
5 lower cognitive function. While the NAS and Nurses' Health Study examined primarily
6 white men and white women, respectively, the BMS examined a more ethnically diverse
7 population of men and women, increasing the generalizability of findings. There was
8 variability in associations across the various domains of cognitive function tested within
9 studies; however, bone Pb levels were associated with decrements in most
10 neuropsychological tests performed. In many studies, bone Pb levels were associated
11 with poorer executive function, visuospatial skills, learning, and memory.
12 Recent evidence from prospective analyses of the NAS and BMS cohorts expanded upon
13 previous cross-sectional evidence by improving characterization of the temporal
14 sequence between Pb exposure and cognitive function declines in adults (n = 405-943,
15 mean ages 60 and 69 years) by demonstrating that higher tibia (means 19, 20 (ig/g) or
16 patella (mean 25 (ig/g) bone Pb levels measured at baseline were associated with
17 subsequent declines in cognitive function over 2- to 4-year periods (Bandeen-Roche et
18 al.. 2009; Weisskopf et al.. 2007b). The specific potential confounding factors considered
19 differed between studies; both studies adjusted for age and education. Additional
20 adjustment was made for income in the BMS and current alcohol use and current
21 smoking in the NAS.
22 Evidence from most cross-sectional analyses supported associations between higher bone
23 Pb level and decrements in cognitive function in adults. A strength of cross-sectional
24 studies overall was the adjustment for the same potential confounding factors described
25 above and also dietary factors, physical activity, medication use, and comorbid conditions
26 (Rajan et al.. 2008; Weuve et al.. 2006). Cross-sectional studies generally demonstrated
27 larger decrements in cognitive function in adults in association with tibia or patella Pb
28 levels than with concurrent blood Pb levels. Results from the NAS and Nurses' Health
29 Study did not clearly indicate a difference in association with cognitive performance
30 between tibia and patella Pb levels (Weuve et al.. 2009; Weisskopf et al.. 2007b). In
31 NHANES analyses, higher concurrent blood Pb levels were associated with lower
32 cognitive function in particular age and genetic variant subgroups but not consistently
33 across the various cognitive tests conducted (Krieg etal. 2010; Krieg and Butler. 2009;
34 Krieg et al.. 2009). NHANES did not have bone Pb measures for comparison.
35 Because bone Pb is a major contributor to blood Pb levels, blood Pb level also can reflect
36 longer term exposures, including higher past exposures, especially in adults without
37 occupational exposures. Thus, in the NHANES results, it is difficult to characterize the
November 2012 5-274 Draft - Do Not Cite or Quote
-------
1 relative contributions of recent and past Pb exposures to the associations observed
2 between concurrent blood Pb level and cognitive function. The discrepant findings for
3 blood and bone Pb levels indicate that cumulative Pb exposure that likely included higher
4 past exposures may be a better predictor of cognitive function in adults than is concurrent
5 blood Pb level.
6 Additional support for the effects of cumulative or past Pb exposure is provided by
7 analyses of a few child cohorts as adults, which indicate that childhood tooth (from ages
8 5-8 years) and blood (e.g., age 10 years) Pb levels are associated with decrements in
9 cognitive function in adults ages 19-30 years (Mazumdar et al.. 2011; Bellinger et al..
10 1994a). An uncertainty in the evidence for bone Pb levels is potential residual
11 confounding by age. Increasing age is highly correlated with increasing bone Pb level
12 (Section 4.3.5.2). and distinguishing Pb-related declines in cognitive function from age-
13 related declines with model adjustment is difficult. One explanation for the more variable
14 findings in adults than in children may be that cognitive reserve may compensate for the
15 effects of Pb exposure on learning new information. Compensatory mechanisms may be
16 overwhelmed with age and with higher long-term or cumulative Pb exposure represented
17 by higher bone Pb levels.
18 Higher blood and bone Pb levels were associated with cognitive function decrements in
19 adults with current or former occupational Pb exposures. Some studies examined current
20 workers with blood Pb level means 26 or 31 ug/dL (Table 5-17). Among adults with
21 current occupational Pb exposures, both concurrent and cumulative exposures may affect
22 cognitive function. Several of these studies considered potential confounding by a similar
23 set of factors as did studies of adults without occupational Pb exposures but did not
24 examine other occupational exposures. In the prospective study of adults with former
25 occupational Pb exposure, peak tibia Pb levels were associated more strongly with
26 cognitive performance than were concurrent blood Pb levels (Khalil et al., 2009a). Thus,
27 in the absence of higher current Pb exposures, cumulative Pb exposures may have a
28 greater effect on cognitive function in adults.
29 Additional support for a relationship between Pb exposure and cognitive function
30 decrements in adults is provided by the coherence with evidence in adult animals that
31 lifetime Pb exposure of animals starting from gestation, birth, or after weaning induces
32 learning impairments (Table 5-17 and Section 5.3.2.3). Biological plausibility also is
33 provided by evidence describing the effects of Pb on modes of action underlying
34 cognitive function. Cognitive function is mediated by actions of the neurotransmitters
35 dopamine and glutamate in the hippocampus, prefrontal cortex, and nucleus accumbens.
36 Experimental studies have shown that Pb induces changes in neurotransmitter release in
37 these regions. Studies also have shown Pb-induced decreases in the magnitude of LTP.
November 2012 5-275 Draft - Do Not Cite or Quote
-------
1 In conclusion, in adults without occupational exposure, recent prospective studies in the
2 NAS and BMS cohorts indicate associations of higher baseline tibia (means 19, 20 (ig/g)
3 or patella (mean 25 (ig/g) Pb levels with declines in cognitive function in adults (>age 50
4 years) over 2- to 4-year periods. While the specific covariates differed between studies,
5 these tibia Pb-associated cognitive function decrements were found with adjustment for
6 potential confounding factors such as age, education, SES, current alcohol use, and
7 current smoking. Supporting evidence is provided by cross-sectional analyses of the
8 NAS, BMS, and the Nurses' Health Study, which found stronger associations with bone
9 Pb level than concurrent blood Pb level. Cross-sectional studies also considered more
10 potential confounding factors, including dietary factors, physical activity, medication use,
11 and comorbid conditions. The multiple exposures and health outcomes examined in many
12 studies reduces the likelihood of biased participation by adults with higher Pb exposure
13 and lower cognitive function. The collective evidence indicates associations in cohorts of
14 white men and women and a cohort of more ethnically diverse men and women. The
15 specific timing, frequency, duration, and magnitude of Pb exposures contributing to the
16 associations observed with bone Pb levels are uncertain. Also uncertain is the potential
17 for residual confounding by age. The effects of recent Pb exposures on cognitive function
18 decrements were indicated in Pb-exposed workers by associations found with blood Pb
19 levels, although these studies did not consider potential confounding by other workplace
20 exposures. The biological plausibility for the effects of Pb exposure on cognitive function
21 decrements in adults is provided by findings that lifetime Pb exposures from gestation,
22 birth, or after weaning induce learning impairments in adult animals and by evidence for
23 the effects of Pb altering neurotransmitter function in hippocampus, prefrontal cortex,
24 and nucleus accumbens. The associations between bone Pb level and cognitive function
25 decrements consistently found in the few prospective and cross-sectional studies of adults
26 without occupational Pb exposure, the coherence with animal findings, and toxicological
27 evidence supporting modes of action but uncertainties related to potential residual
28 confounding by age in epidemiologic studies are sufficient to conclude that a causal
29 relationship is likely to exist between long-term cumulative Pb exposure and cognitive
30 function decrements in adults.
5.3.16.8 Evidence for Psychopathological Effects in Adults
31 Evidence indicates that a causal relationship is likely to exist between Pb exposure and
32 psychopathological effects in adults, based on the cross-sectional associations found
33 between concurrent blood Pb level (Bouchard et al. 2009) or bone Pb level (Rajan et al..
34 2008) and self-reported depression, anxiety, and panic disorder in adults participating in
35 NHANES and NAS, respectively and supporting toxicological evidence. Higher prenatal
November 2012 5-276 Draft - Do Not Cite or Quote
-------
1 blood 5-ALA level was associated with schizophrenia in adults in California (Opler et al..
2 2008; 2004). but because of the lack of assessment of blood or bone Pb levels, results
3 were not a major consideration in conclusions. In both NHANES and NAS, the high
4 participation rates and examination of multiple exposures and outcomes reduces the
5 likelihood that findings are influenced by biased participation or reporting of symptoms
6 by subjects with higher Pb exposures. Depression and anxiety were assessed with widely-
7 used structured questionnaires such as the Profile of Mood States, but there is uncertainty
8 regarding the effects of Pb exposure on clinically-diagnosed conditions.
9 Epidemiologic evidence for associations between blood or bone Pb level and
10 psychopathological effects is strengthened by the consideration of several potential
11 confounding factors. Among adults ages 20-39 years participating in NHANES, 369
12 adults with concurrent blood Pb level > 2.11 (ig/dL had the highest OR for self-reported
13 major depressive disorder (OR: 2.32 [95% CI: 1.13, 4.75]) and panic disorder (OR: 4.94
14 [95% CI: 1.32, 18.48]) compared with the 449 adults with blood Pb levels <0.7 (ig/dL
15 with adjustment for age, sex, race, education, and poverty to income ratio (Bouchard et
16 al.. 2009). Among 526 NAS men ages 48-70 years, a 27 (ig/g increase in tibia Pb level
17 was associated with a combined index of self-reported anxiety, depression, and phobic
18 anxiety with an OR of 2.08 (95% CI: 1.06, 4.07) with adjustment for age, grams/day
19 alcohol ingested, education, and employment status (Rhodes et al.. 2003). Because of the
20 cross-sectional design of studies, the temporal sequence between Pb exposure and
21 psychopathological symptoms in adults is uncertain. This uncertainty is somewhat
22 reduced with results for tibia Pb, since it is an indicator of cumulative Pb exposure. For
23 results with blood and bone Pb level, there is uncertainty regarding the critical level,
24 timing, frequency, and duration of Pb exposure associated with psychopathological
25 effects.
26 The epidemiologic evidence for Pb-associated psychopathological effects is supported by
27 the coherence with findings in rodents that dietary prenatal/lactational or lactational Pb
28 exposure resulted in depression-like and loss of motivation behavior in rodents, with
29 some evidence at blood Pb levels relevant to humans (13-17 (ig/dL) (Beaudin et al..
30 2007; Dvatlov and Lawrence. 2002). Other studies found Pb-induced increases in
31 depression-like behavior in animals with higher blood Pb levels (Section 5.3.5.2). Further
32 support for Pb-associated increases in psychopathological effects in adults is provided by
33 evidence that describes modes of action, including Pb-induced changes in the HPA axis
34 (Section 5.3.2.3) and dopaminergic and GABAergic systems (Sections 5.2.2.2 and
35 5.3.11.8). which are found to affect mood and emotional state.
36 In conclusion, cross-sectional studies in a few populations demonstrate associations of
37 higher concurrent blood or tibia Pb levels with self-reports of depression and anxiety in
November 2012 5-277 Draft - Do Not Cite or Quote
-------
1 adults. The examination of multiple exposures and outcomes in the available studies does
2 not provide strong indication of biased reporting of psychopathological effects by adults
3 with higher Pb exposures. In adults, Pb-associated increases in depression and anxiety
4 were found with adjustment for age, SES, and in the NAS, daily alcohol intake. The
5 biological plausibility for epidemiologic evidence is provided by observations of
6 depression-like behavior in animals with dietary prenatal/lactational or lactational Pb
7 exposure, with some evidence at blood Pb levels relevant to humans and by toxicological
8 evidence supporting modes of action, including Pb-induced changes in the HPA axis and
9 dopaminergic and GABAergic systems. The associations of blood and bone Pb level with
10 self-reported psychopathological effects found in the few studies of adults without
11 occupational Pb exposure, the biological plausibility provided by the coherence of
12 findings in animals and underlying modes of action, but uncertainties related to residual
13 confounding of bone Pb results by age in epidemiologic studies are sufficient to conclude
14 that a causal relationship is likely to exist between Pb exposure and psychopathological
15 effects in adults.
5.3.16.9 Evidence for Sensory Function Decrements in Adults
16 The small body of epidemiologic and toxicological evidence is suggestive of a causal
17 relationship between Pb exposure and sensory function decrements in adults. Key
18 evidence in humans is provided by the recent analysis of NAS males in which a 15 (ig/g
19 higher tibia Pb level at mean age 64.9 years was associated with a 0.05 dB/year (95% CI:
20 0.017, 0.083) increase in hearing threshold for a pure tone average frequency (Park et al..
21 2010). Results were adjusted for baseline age, race, education, occupational noise, BMI,
22 pack-years smoking, noise notch, diabetes, and hypertension. Although the
23 generalizability of results in this primarily white population of men is limited, high
24 follow-up participation and the examination of multiple exposures and outcomes in this
25 cohort reduces the likelihood that findings are biased by selective participation of men
26 with higher Pb exposures. Bone Pb levels were measured up to 20 years after the initial
27 hearing measurement; however, tibia Pb level is considered an indicator of cumulative Pb
28 exposure since the half-life of Pb in bone is on the order of decades (Section 4.3). Bone
29 Pb levels increase with age, and although age was included as a model covariate, residual
30 confounding by age is possible. Supporting evidence was provided by a recent case-
31 control study of adults attending a hospital for occupational health exams. Despite
32 limitations of a nonrandom population and uncertain comparability of controls, the
33 examination of multiple metals reduces the likelihood of biased participation by higher
34 Pb exposure. Higher concurrent blood Pb level was associated with hearing loss with
November 2012 5-278 Draft - Do Not Cite or Quote
-------
1 adjustment for several factors (Table 5-17) including blood levels of Mn, As, and Se
2 (Chuang et al.. 2007).
3 These epidemiologic findings are supported by the coherence of findings for Pb-induced
4 increases in hearing thresholds in animals, albeit at higher blood Pb levels than those
5 relevant for humans. Monkeys that were exposed to Pb in drinking water from gestation
6 through testing at age 13 years and had blood Pb levels 33-107 (ig/dL were found to have
7 elevated thresholds and increased latencies in brainstem auditory evoked potentials (Rice.
8 1997; Lilienthal and Winneke. 1996). A recent study found lack of persistence of effects
9 in monkeys tested at age 13 years that had shorter duration exposure, gestation through
10 age 5.5 months (Laughlin et al.. 2009).
11 In conclusion, the evidence provided by the analysis of NAS men for associations of
12 higher tibia Pb level with a greater rate of elevations in hearing threshold over 20 years
13 and the biological plausibility provided by the evidence for Pb-induced decreases in
14 auditory evoked potentials in animals but at higher blood Pb levels than those relevant to
15 humans, is suggestive of a causal relationship between Pb exposure and sensory function
16 decrements in adults.
5.3.16.10 Evidence for Neurodegenerative Diseases in Adults
17 Epidemiologic and toxicological studies have found associations between indicators of
18 Pb exposure and neurodegenerative diseases such as Parkinson's disease and essential
19 tremor, but evidence is inconclusive for Alzheimer's disease and ALS. Despite the
20 evidence for some neurodegenerative diseases, because of limitations as described below,
21 the evidence is inadequate to determine that there is a causal relationship between Pb
22 exposure and neurodegenerative diseases.
23 The few case-control studies of essential tremor found higher concurrent blood Pb levels
24 in cases than controls (Section 5.3.10.4). A common limitation of these studies was the
25 potential for reverse causation. Reduced physical activity among cases could result in
26 greater bone turnover and greater release of Pb from bones into blood in cases than
27 controls. Some case-control studies found adults with Parkinson's disease to have higher
28 bone Pb levels, which are not likely to increase with decreases in physical activity
29 ("Weisskopf et al.. 2010; Coon et al.. 2006). While some of these studies of Parkinson's
30 disease and essential tremor considered potential confounding by factors such as age, sex,
31 race, education, and alcohol consumption, they did not consider Mn co-exposure.
32 Epidemiologic findings for Parkinson's disease are supported by limited available mode
33 of action evidence for Pb-induced decreased dopaminergic cell activity in the substantia
34 nigra, which contributes to the primary symptoms of Parkinson's disease.
November 2012 5-279 Draft - Do Not Cite or Quote
-------
1 The few case-control studies of Alzheimer's disease did not find higher prevalence of
2 occupational Pb exposure or higher brain Pb levels in cases but did not measure Pb in
3 blood or bone (Section 5.3.10.1). Toxicological studies indicated that infancy exposure
4 during lactation but not adult-only Pb exposures of monkeys and rats induced pathologies
5 that underlie Alzheimer's disease, including the formation of amyloid plaques and
6 neurofibrillary tangles in the brains of aged animals (Section 5.3.10.1). While these
7 results suggest the need to consider early-life Pb exposure in epidemiologic studies, some
8 indicate that effects may be attributable to the high Pb exposure concentrations tested,
9 i.e., producing blood Pb levels >40 (ig/dL in rats (Li et al., 2010; BashaetaL 2005).
10 Studies of ALS have not consistently found higher blood Pb levels among ALS cases and
11 controls (Section 5.3.10.2). and a recent study found that higher tibia and patella Pb
12 levels were associated with longer survival time among ALS cases (Kamel et al., 2008).
13 In conclusion, while evidence is inconclusive for ALS and Alzheimer's disease, a few
14 case-control studies each found higher blood Pb levels in adults with essential tremor and
15 higher bone Pb levels in adults with Parkinson's disease. Because of the inconclusive
16 evidence for some diseases and limitations such as reverse causation for essential tremor
17 and the lack of consideration for potential confounding by Mn exposure for both essential
18 tremor and Parkinson's disease, the evidence is inadequate to determine that there is a
19 causal relationship between Pb exposure and neurodegenerative diseases.
November 2012 5-280 Draft - Do Not Cite or Quote
-------
Table 5-17 Summary of Evidence Supporting Nervous System Causal
Determinations.
Attribute in
Causal
Framework3
Key Supporting Evidence13 References'3
Pb Biomarker Levels
Associated with
Effects0
Cognitive Function Decrements in Children - Causal
Consistent Evidence from prospective studies for
associations from decrements in FSIQ in association
multiple, high with prenatal, earlier childhood, peak,
quality concurrent, lifetime average blood Pb
epidemiologic levels and tooth Pb levels in children
studies with ages 4-17 yr in multiple U.S. locations,
relevant blood Pb Mexico, Europe, Australia
levels
Most studies adjust for confounding by
SES, maternal IQ and education,
HOME score. Several adjust for birth
weight, smoking. A few, nutritional
factors.
Pooled analysis of seven cohorts
indicates precise effect estimates,
-2.36 to -2.94 FSIQ points per log
increase in blood Pb level, excluding
one study at a time
Meta-analyses demonstrate the
consistency of association
Evidence from prospective studies for
lower scores on tests of executive
function and academic performance in
association with earlier childhood or
lifetime average blood Pb levels or
tooth Pb levels in children ages 5-18
yrin multiple U.S. locations, U.K, New
Zealand. Associations less consistent
for learning and memory.
Supporting evidence from cross-
sectional studies of children ages 3-16
yr, but most did not consider potential
confounding by parental caregiving
quality. Includes large NHANES III
analysis.
Studies had population-based
recruitment, most with moderate to
high follow-up participation not
conditional on blood or tooth Pb level.
Outcomes assessed using widely-
used, structured questionnaires.
Several studies indicate supralinear
C-R relationship, with larger
decrements in cognitive function per
unit increase in blood Pb at lower
blood Pb levels in children ages
5-1 Oyr
Canfield et al. (2003a),
Bellinger etal. (1992).
Jusko et al. (2008).
Dietrich et al. (1993b),
Schnaaset al. (2006).
Wasserman et al. (1997),
long et al. (1996)
Section 5.3.2.1
Lanphear et al. (2005)
Pococket al. (1994).
Schwartz (1994)
Bellinger etal. (1991).
Canfield et al. (2004).
Risetal. (2004),
Stiles and Bellinger (1993).
Miranda et al. (2009: 2007a).
Fergusson et al. (1997. 1993),
Leviton et al. (1993).
Chandramouli et al. (2009)
Sections 5.3.2.3. 5.3.2.4. 5.3.2.5
Surkan et al. (2007).
Kim et al. (2009b).
Roy etal. (2011).
Lanphear etal. (2000),
Froehlich et al. (2007)
Table 5-3. Table 5-5:
Table 5-8. Table 5-9
Canfield et al. (2003a).
Bellinger etal. (1992).
Jusko et al. (2008).
Kordasetal. (2006).
Lanphear et al. (2005)
Blood Pb (various
lifestages and time
periods): means 3-
14ug/dL
Blood Pb (various
lifestages and time
periods):
Means 4.8-8 ug/dL,
Groups with early
childhood blood Pb
>2 and >5 ug/dL
Tooth Pb (ages 6-8 yr):
means 3.3, 6.2 ug/g
Concurrent blood Pb :
Means 1.7-11.4 ug/dL,
Groups with blood Pb
>10 ug/dL
Groups with peak blood
Pb <10 ug/dL: concurrent
mean 3.3 ug/dL, age 2
year mean 3.8 ug/dL
November 2012
5-281
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Epidemiologic
evidence supported
by consistent
toxicological results
with relevant
exposures
Impaired learning in juvenile and adult
animals as indicated by performance
in Y maze, Operant Schedules of
Reinforcement with relevant dietary
Pb exposure.
Impaired learning, memory, executive
function in juvenile and adult animals
as indicated by poorer performance on
spatial delayed alternation and
discrimination reversal learning tasks
with dietary Pb exposures.
Stangleetal. (2007).
Niuetal. (2009).
Cory-Slechta et al. (2010).
Virgolini et al. (2008a).
Altmannetal. (1993).
Section 5.3.2.3
Gilbert and Rice (1987).
Rice and Gilbert (1990b).
Rice (1992b).
Rice and Gilbert (1990a),
Rice and Karpinski (1988).
Sections 5.3.2.3 and 5.3.2.4
Blood Pb (after prenatal/
lactation, lactation only,
postlactation,
prenatal/lifetime Pb
exposure): 8-31 ug/dL
Blood Pb (after lifetime
Pb exposure after
lactation): 15-26 ug/dL
Evidence clearly
describes mode of
action
Impaired neuron
development
Synaptic changes
LTP
Neurotransmitter
changes
Decreased neurogenesis in
hippocampus DG, which is involved in
LTP and learning, with lactational,
postlactational (25 days), lifetime from
gestation dietary Pb exposures.
Decreased NMDAR, which is involved
in integration of new neurons into
existing neuronal pathways with
postlactational (8 weeks) and lifetime
from gestation dietary Pb exposures.
Decreased neurite outgrowth in
animals with gestational Pb exposure
Decreased synaptic development with
gestational-lactational dietary Pb
exposures.
Changes in synaptic protein
composition with gestational-
lactational Pb exposure.
Decreased ATP, AchE, which mediate
neurotransmission with gestational Pb
exposure
Decreased magnitude, increased
threshold of LTP with gestational-
lactational Pb exposure.
Decreased dopamine in substantia
nigra with gestational-lactational
dietary Pb exposure.
Increased sensitivity of dopamine
receptor with gestational-lactational,
lactational, or postlactational Pb
exposure.
Increased catecholamine transmission
in cerebral cortex, cerebellum,
hippocampus with gestational-
lactational Pb exposure.
Decreased glutamate and expression
of glutamate receptor, NMDAR in vitro
and in rats with gestational-lactational
Pb exposure.
Section 5.3.11.9 and 5.3.11.10
Section 5.3.11.4
Section 5.3.11.8
Section 5.3.11.8
November 2012
5-282
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Attention-related Behavioral Problems in Children (e.g., inattention, impulsivity, hyperactivity, ADHD)
- Causal
Consistent Evidence from prospective studies for
associations from inattention, impulsivity, and
multiple, high hyperactivity in association with
quality prenatal (maternal or cord), earlier
epidemiologic childhood, and lifetime avg blood Pb
studies with and tooth Pb levels in children ages 7-
relevant blood Pb 17 yr and young adults 19-24 yr in
levels U.S., U.K., Australia, New Zealand.
Most studies adjusted for SES,
maternal education, and parental
caregiving quality. Some also
considered parental IQ, smoking, birth
outcomes. A few considered
substance abuse, nutritional factors.
Studies had population-based
recruitment with moderate to high
follow-up participation not conditional
on blood or tooth Pb level.
Associations found with
neuropsychological tests (CPT) and
teacher and parent ratings using
widely-used, structured
questionnaires.
Associations with inattention ratings
inconsistent in prospective studies
examining children with lower blood
Pb levels, but children were younger,
<5yr
Supporting evidence from cross-
sectional studies for associations of
concurrent blood Pb level with
inattention, impulsivity, and
hyperactivity, and total ADHD rating in
children ages 8-15 yr.
Cross-sectional studies had less
extensive consideration for potential
confounding, particularly parental
caregiving quality.
Risetal. (2004),
Fergusson et al. (1993).
Bellinger et al. (1994a).
Chandramouli et al. (2009).
Leviton et al. (1993)
Burns et al. (1999) with the most
extensive consideration for potential
confounding
Sections 5.3.3.1. 5.3.3.2. 5.3.3.3
Blood Pb:
means 6.8 ug/dL
(prenatal cord),
14ug/dL
(lifetime avg to age 11 -
13yr), Group with age
30 mo >10 ug/dL
Tooth Pb (age 5-8 yr)
means: 3.3, 6.2, 14 ug/g
Wasserman et al. (2001)
Canfield et al. (2003b)
Section 5.3.3.1
Choetal. (2010).
Nicolescu et al. (2010).
Froehlichetal. (2009).
Silva et al. (1988)
Sections 5.3.3.1. 5.3.3.2. 5.3.3.3
Concurrent blood Pb
means 6.5 ug/dL
Concurrent Blood Pb
means 1.9-11.1 ug/dL
Epidemiologic
evidence supported
by toxicological
results with relevant
exposures
Impulsivity indicated by premature
responses, increased perseveration,
decreased pause time between events
on tests of response inhibition in
rodents and monkeys with relevant
dietary postnatal Pb exposures.
Increased distractibility found in adult
monkeys with relevant lifetime dietary
Pb exposure as assessed by poorer
performance on discrimination
reversal learning tests with distracting
stimuli.
Relevant postnatal dietary Pb
exposure not found consistently to
affect sustained attention in rats as
assessed using signal detection test
with distracting stimuli.
Stangleetal. (2007).
Brockel and Cory-Slechta (1998),
Rice (1985).
Brockel et al. (1999b)
Section 5.3.3.1
Gilbert and Rice (1987)
Section 5.3.3.1
Brockel and Cory-Slechta (1999a)
Stangle et al. (2007)
Section 5.3.3.1
Blood Pb: 15, 25 ug/dLin
monkeys after infancy
only exposure, 11,
29 ug/dL in rats after 40-
day postweaning
exposure, 10, 26 ug/dL
after 3, 7 mo postweaning
exposure
Blood Pb after lifetime
(after birth) exposure: 15,
25 ug/dL
Blood Pb: 16,28 ug/dL
after 34-day postweaning
exposure, 13, 31 ug/dL
after lactational exposure.
November 2012
5-283
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Evidence clearly
describes mode of
action
Same as above for cognitive function
in children
Internalizing Behaviors in Children (e.g., withdrawn behavior, symptoms of depression, anxiety, tearfulness)
- Likely Causal
Associations found
in high-quality
epidemiologic
studies with
relevant exposures
Evidence from prospective studies for
higher ratings of internalizing
behaviors in children ages 8-13 yr in
Boston and Port Pirie cohorts in
association with tooth or lifetime
average blood Pb levels.
Results adjusted for SES, birth
outcomes, parental education. Port
Pirie also adjusted for HOME, current
maternal psychopathological
symptoms.
Burns etal. (1999).
Bellinger etal. Q994b)
Section 5.3.5.1
Blood Pb lifetime (to age
11-13 yr) average mean:
~14ug/dL
Tooth Pb (age 6 yr) mean:
3.4 ug/g
Associations also found in children
age 4-5 yr in Yugoslavia in association
with lifetime average blood Pb level;
stronger association found for
delinquent behavior.
Results adjusted for similar covariates
as above plus maternal history of
smoking, residence type.
Studies had population-based
recruitment with moderate follow-up
participation. Participation not
conditional on tooth/blood Pb levels
and behavior.
Associations found with teacher and
parent ratings on widely used,
structured questionnaires.
Wasserman et al. (2001)
Blood Pb lifetime (to age
4-5 yr) average mean:
9.6 ug/dL
Cross-sectional studies found
associations with concurrent blood Pb
level but had limited consideration for
potential confounding and/or
nonrepresentative populations (e.g.,
prenatal drug exposure).
Section 5.3.5.1
Epidemiologic
evidence supported
by toxicological
evidence with
relevant exposures
Postnatal dietary Pb exposure
increased emotionality, loss of
motivation in response to reward
omission in juvenile female rats
Postnatal dietary Pb exposure
increased sickness behavior due to
bacteria infection in juvenile mice.
No specific mode of action examined
with Pb exposure
Stangleetal. (2007).
Beaudinetal. (2007)
Dyatlov and Lawrence (2002)
Section 5.3.5.2
Blood Pb at PND45 after
PND1-PND30 exposure:
13, 31 ug/dL
Blood Pbat PND22 after
PND1-PND22 exposure:
17ug/dL
November 2012
5-284
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Conduct Problems in Children and Young Adults (e.g., criminal offenses, delinquent behavior, aggression, antisocial
behavior)
- Likely Causal
Consistent results
from high-quality
epidemiologic
studies with
relevant blood or
tooth Pb levels
Evidence from prospective studies for
criminal offenses in young adults ages
19-24 yr in Cincinnati and New
Zealand in association with earlier
childhood blood or tooth Pb levels.
Evidence from prospective studies for
higher parent and teacher ratings of
aggression, antisocial behavior,
delinquent behavior in children ages 8-
17 yr in U.S., U.K., Australia in
association with earlier childhood or
lifetime average blood Pb and tooth
Pb levels.
Associations also found in children
age 4-5 yr in Yugoslavia in association
with lifetime average blood Pb level;
stronger association found for
delinquent behavior.
Studies had moderate to high follow-
up participation, not conditional on
blood Pb level.
Most studies considered potential
confounding by SES, parental
education and IQ, other SES factors,
parental caregiving/functioning,
smoking, substance abuse.
Supporting evidence for parental
report of conduct disorder in
association with concurrent blood Pb
in cross-sectional study of children
ages 8-15 yr participating in
NHNANES. Examination of multiple
exposures and outcomes reduces
likelihood of selection bias.
Consistency supported by meta-
analysis indicating similar effect
estimates by study design, potential
confounding factors considered
Teacher and parental ratings derived
from widely-used, structured
questionnaires.
Wright et al. (2008).
Fergusson et al. (2008),
Section 5.3.4.1
Dietrich et al. (2001).
Burns etal. (1999).
Chandramouli et al. (2009).
Bellinger etal. (1994b),
Section 5.3.4.1
Wasserman et al. (2001)
Braun et al. (2008)
Section 5.3.4.1
Marcus et al. (2010)
Section 5.3.4.1
Age 6 yr blood Pb mean
6.8 ug/dL
Tooth Pb (age 6-8 yr)
mean: 6.2 ug/g
Blood Pb: lifetime (to age
11-13 yr) avg mean:
14 ug/dL, age 30 month
group with blood Pb
>10ug/dL
Tooth Pb (age 6 yr) mean
3.4 ug/g
Mean lifetime (to age 4-5
yr) average: 9.6 ug/dL
Groups with concurrent
blood Pb >0.8 ug/dL.
Blood Pb range of study
means (concurrent or
lifetime avg): 1.0-26 ug/dL
Inconsistent
evidence in animals
for aggression at
relevant exposures
Aggression observed in adult
hamsters with gestational-lifetime
dietary Pb exposure. Other evidence
in adult animals with similar duration
exposure inconsistent.
Aggression generally not found in
juvenile animals with lactational Pb
exposure.
No specific mode of action examined
with Pb exposure
Delville (1999)
Section 5.3.4.2
Blood Pb after
gestational-lifetime
exposure: 10-15 ug/dL
November 2012
5-285
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Sensory Function Decrements in Children - Likely Causal
Consistent findings Prospective study indicated
from high-quality associations of prenatal (maternal),
epidemiologic neonatal, yearly age 1 to 5 yr, lifetime
studies with avg blood Pb levels with poorer
relevant blood Pb auditory processing in children at age
levels Syr in Cincinnati.
Information was not provided on
participation rates.
Results were adjusted for SES,
HOME, birth weight, gestational age,
obstetrical complication, maternal
smoking. Several other factors
considered.
Supporting evidence from cross-
sectional studies for increased hearing
thresholds in children ages 4-19 yr
participating in NHANES and HHANES
in association with higher concurrent
blood Pb levels.
Examination of multiple exposures and
outcomes in NHANES and HHANES
reduces likelihood of selection bias.
Studies considered potential
confounding by age, sex, race,
income, parental education, nutritional
factors.
Dietrich et al. (1992)
Section 5.3.7.1
Schwartz and Otto (1991. 1987)
Section 5.3.7.1
Blood Pb means:
neonatal (10 day)
4.8 ug/dL, yearly age 1 to
5 year 10.6-17.2 ug/dL,
lifetime (to age 5 yr) avg
NR
Concurrent blood Pb
median in HHANES:
8 ug/dL, NHANES: NR
Limited
toxicological results
at relevant
exposures
Increased hearing thresholds in
monkeys age 13 years with lifetime
dietary Pb exposure.
Supernormal or subnormal retinal
ERGs in rats depending on timing and
dose of Pb exposure. Uncertain
biological relevance.
Rice (1997).
Section 5.3.7.3
Foxetal. (1997).
Section 5.3.7.3
Blood Pb after lifetime
(from birth) exposure: 33-
170ug/dL
Blood Pb: 12, 24 ug/dL
with gestational-
lactational exposure,
19 ug/dL with lactational
exposure
Evidence describes
mode of action
Decreased auditory evoked potentials
with lifetime Pb exposure of monkeys
ages 13 years.
Rod cell proliferation, retinal cell
apoptosis, dopamine changes
Section 5.3.7.3
November 2012
5-286
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Motor Function Decrements in Children - Likely Causal
Consistent findings Evidence from prospective studies for
from high-quality fine and gross motor function
epidemiologic decrements in children ages 4.5-17 yr
studies with in Cincinnati, Yugoslavia in association
relevant blood Pb with neonatal, earlier childhood,
levels concurrent, lifetime avg blood Pb
levels.
High follow-up participation, no
selective attrition in Cincinnati cohort,
higher loss-to-follow-up in Yugoslavia
cohort with lower maternal IQ, HOME.
Both studies adjusted for maternal IQ,
parental education, SES, HOME score
Supporting evidence from cross-
sectional studies in children ages 3-7
yr in India, Canada. In Inuit Canadian
children, potential confounding factors
varied by outcome but included
HOME, maternal education, weight,
prenatal alcohol exposure, Hg,
polychlorinated biphenyls.
No decrease in motor function found in
children ages 6-10 yr in New England
with lower concurrent blood Pb levels
with adjustment for age, race, sex,
caregiver education, SES.
Studies used various, widely-used
tests to assess outcomes.
Risetal. (2004).
Dietrich et al. Q993a),
Bhattacharya et al. (2006),
Wasserman et al. (2000)
Section 5.3.8
Cincinnati: blood Pb
means: neonatal
4.8 ug/dL, age 6 yr
11.6 ug/dL, lifetime (to
age 15-17 yr) avg
12.3ug/dL
Yugoslavia: NR
Despres et al. (2005).
Palaniappan et al. (2011)
Section 5.3.8
Surkan et al. (2007)
Concurrent blood Pb
means: 4.1 ug/dL
Canada, 11.5 ug/dL India
Concurrent blood Pb
mean: 2.2 ug/dL
Limited
toxicological
evidence at
relevant exposures
Poorer balance (fell off rotarod more
quickly) in adult mice with gestational-
lactation dietary Pb exposure
Leasure et al. (2008)
Section 5.3.8
Peak blood Pb after
gestational -lactational
exposure ~10 ug/dL
postnatal exposures
postnatal Pb exposure
>60 ug/dL
November 2012
5-287
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Cognitive Function Decrements in Adults - Likely Causal
Consistent results Prospective analyses in MAS cohort of
from high-quality white men and BMS cohort of men
epidemiologic and women of diverse ethnicities
studies with found cognitive function decrements
relevant bone Pb over 2 to 4 years in association with
levels bone Pb levels.
Baseline participation rates differed
but high follow-up participation, not
conditional on bone Pb levels.
Different potential confounding factors
considered. Adjustment for age and
education in both cohorts, SES in
BMS, smoking and alcohol use in
MAS.
Supporting cross-sectional evidence
from MAS, BMS, and also women in
Nurses' Health Study with adjustment
for additional potential confounding
factors, including dietary factors,
medications, physical activity,
comorbid conditions.
Associations with blood Pb level found
in men and women participating in
NHANES in certain genetic variant
groups with adjustment for age, sex,
education, income, race/ethnicity,
alcohol use, computer/video game
familiarity.
Several studies found associations
with blood Pb levels and bone Pb
levels in former and current
Pb-exposed workers. Most studies
adjusted for age and education. Some
also adjusted for depression and/or
alcohol use, but none considered
other occupational exposures.
Outcomes assessed using various but
widely used, structured instruments.
Uncertainty regarding potential
residual confounding of bone Pb
results by age.
Weisskopfetal. (2007b),
Bandeen-Roche et al. (2009)
Table 5-10 and Section 5.3.2.7
Nurses Healthy Study:
Weuve et al. (2009)
Section 5.3.2.7
Table 5-10
Krieg et al. (2009).
Krieg et al. (2010).
Krieg and Butler (2009)
Section 5.3.2.7
Khaliletal. (2009a).
Dorsey et al. (2006).
Bleeckeret al. (2007a)
Stewart et al. (2002)
Section 5.3.2.7
Baseline tibia Pb means:
18.8, 20 ug/g, patella
mean 25 ug/g
Concurrent tibia Pb
Mean: 10.5 ug/g
Concurrent blood Pb
means: 3-4 ug/dL
Concurrent blood Pb: 12
(former workers)-
31 ug/dL.
Peak tibia Pb: mean
26.2 ug/g, median
57 ug/g.
Epidemiologic
evidence supported
by consistent
toxicological results
with relevant
exposures
Impaired learning, memory, and
executive function in adult monkeys
with lifetime dietary Pb exposures after
weaning.
Impaired learning in animals with
lifetime dietary Pb exposures starting
in gestation.
Rice (1992b),
Rice and Gilbert (1990a).
Rice (1990)
Section 5.3.2.3
See above for cognitive function in
children.
Blood Pb after post-
weaning exposure to age
7-1 Oyr means 19,
26 ug/dL
Evidence clearly
describes mode of
action
Same as above for cognitive function
in children
November 2012
5-288
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Psychopathological Effects in Adults (e.g., self ratings of depression, anxiety, panic disorder)
- Likely Causal
Consistent findings
from high-quality
epidemiologic
studies with
relevant blood and
bone Pb levels
A few cross-sectional studies indicate
associations of higher concurrent
blood or tibia Pb level with increased
reporting of depression, anxiety, panic
disorder in adults without occupational
Pb exposures.
Studies examine multiple exposures
and outcomes.
Association with blood Pb level among
1,987 adults participating in NHANES;
adjustment for age, sex, race,
education, poverty to income ratio.
Associations with blood and bone Pb
level among MAS men (mostly white)
with high follow-up participation and
adjustment for education, age,
employment, pack-years smoking,
alcohol use.
Higher ratings of disorders also in
Pb-exposed workers with higher blood
Pb levels.
Studies used widely used, structured
instruments to assess outcomes but
not diagnosed conditions.
Bouchard et al. (2009),
Rhodes etal. (2003).
Rajanetal. (2008)
Section 5.3.6.1
Bouchard et al. (2009)
Section 5.3.6.1
Rhodes etal. (2003),
Rajanetal. (2007)
Section 5.3.6.1
Section 5.3.6.1
Group with concurrent
blood Pb>2.11 ug/dL
Concurrent blood Pb
mean: 6.3 ug/dL,
concurrent tibia Pb mean:
21.9 ug/g
Concurrent or peak blood
Pb: means 31 -79 ug/dL
Supporting
toxicological
evidence at
relevant Pb
exposures
Same as above for internalizing
behaviors in children
Sensory Function Decrements in Adults - Suggestive
Limited but high-
quality
epidemiologic
evidence with
relevant bone or
blood Pb levels
Prospective study indicates
associations between tibia Pb level
and faster rate of increase in hearing
threshold over 23 yr, among MAS male
adults.
Population comprises only males,
primarily white, but study examines
multiple exposures and outcomes and
has high follow-up participation.
Results adjusted for age, race,
education, BMI, pack-years smoking,
diabetes, hypertension, occupational
noise.
Supporting evidence from case-control
study finding higher blood Pb levels in
workers from various occupations with
hearing loss with adjustment for age,
smoking, alcohol consumption, years
of noise exposure, blood Mn, As, Se
Park etal. (2010)
Section 5.3.7.2
Tibia Pb mean: 22.5 ug/g,
measured near end of
follow-up
Chuang et al. (2007
Section 5.3.7.2
Concurrent blood Pb
mean in cases:
10.7ug/dL
Supporting
toxicological
evidence with
relevant exposures
Same as above for sensory function
decrements in children, including
mode of action
November 2012
5-289
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Biomarker Levels
Associated with
Effects0
Neurodegenerative Diseases (e.g..Alzheimer's disease, ALS, Parkinson's disease, Essential tremor)
- Inadequate
The available The few available case-control studies
evidence is not found higher blood or bone Pb levels
sufficiently and history of occupational exposure
informative in cases with Parkinson's disease and
essential tremor. For ALS, association
also found with increased survival
time.
Studies subject to selection and recall
bias and reverse causation (blood Pb)
that could produce artifactual
associations.
Some studies consider potential
confounding by age, smoking,
education, BMI, and activity levels,
occupational studies did not consider
Mn co-exposures.
Parkinson's disease:
Gorell et al. (1997).
Gulson et al. (1999).
Tanner et al. (1989).
Weisskopfetal. (2010).
Coon et al. (2006)
Section 5.3.10.3
Essential tremor:
Louis etal. (2005: 2003),
Dogu et al. (2007)
Section 5.3.10.4
ALS:
Kamel et al. (2002).
Kamel et al. (2005).
Vinceti et al. (1997).
Fang et al. (2010)
Section 5.3.10.2
Parkinson's disease:
groups with tibia Pb levels
>15 ug/g
Essential tremor:
concurrent blood Pb
means 3-4 ug/dL
ALS:
groups with blood Pb
>3 ug/dL,
groups with tibia Pb
>8 ug/g
Case-control studies did not show
associations between occupational
history of Pb exposure or brain Pb
levels and Alzheimer's disease.
Graves et al. (1991).
Hariguchi et al. (2001)
Section 5.3.10.1
Some evidence
describes mode of
action
Amyloid plaques found in brains of
adult monkeys and rodents with
infancy-only lactational Pb exposures.
In monkeys (ages 20-23 yr), no effect
with adult-only Pb exposure
In rodents, adult blood Pb levels at
testing had returned to baseline.
Increases in neuronal cell apoptosis
found in vitro with Pb exposure
Section 5.3.10.1
Basha et al. (2005)
Section 5.3.10.5
Blood Pb with lactational
exposure: 19-26 ug/dL
Blood Pb with lactational
exposure: 46 ug/dL
"Described in detail in Table II of the Preamble.
bDescribes the key evidence and references contributing most heavily to causal determination. References to earlier sections
indicate where full body of evidence is described.
""Describes the blood Pb levels in children with which the evidence is substantiated and blood Pb levels in animals most relevant to
humans.
November 2012
5-290
Draft - Do Not Cite or Quote
-------
5.4 Cardiovascular Effects
5.4.1 Introduction
1 The 2006 Pb AQCD (U.S. EPA. 20061)) concluded that both epidemiologic and animal
2 toxicological studies support the relationship between increased Pb exposure and
3 increased cardiovascular effects, in particular, increased blood pressure (BP) and
4 increased incidence of arterial hypertension. Although fewer in number, epidemiologic
5 studies demonstrated associations of blood and bone Pb levels with other cardiovascular
6 diseases (CVDs) in adults, such as ischemic heart disease, cerebrovascular disease,
7 peripheral vascular disease, and CVD-related mortality. As the cardiovascular and renal
8 systems are intimately linked, cardiovascular effects can arise secondarily to Pb-induced
9 renal injury (Section 5.5). Toxicological studies also provided compelling evidence
10 supporting the biological plausibility for Pb-associated cardiovascular effects by
11 characterizing a number of the underlying mechanisms by which Pb exposure can lead to
12 human cardiovascular health effects. Such studies demonstrated that the Pb content in
13 heart tissue of animals reflects the increases in blood Pb levels (Lai et al., 1991).
14 indicating that the cardiovascular morbidity associated with blood Pb levels may
15 represent the effects of the bioavailable Pb in the target tissue. The strongest evidence
16 supported the role of oxidative stress in the pathogenesis of Pb-induced hypertension.
17 Additionally, several toxicological studies characterized other pathways or cellular,
18 molecular, and tissue events promoting the Pb-induced increase in BP. These
19 mechanisms included inflammation, adrenergic and sympathetic activation, renin-
20 angiotensin-aldosterone system (RAAS) activation, vasomodulator imbalance, and
21 vascular cell dysfunction.
22 With regard to the concentration-response relationship, a meta-analysis of human studies
23 found that each doubling of blood Pb level (between 1 and >40 ug/dL measured
24 concurrently in most studies) was associated with a 1 mmHg increase in systolic BP and
25 a 0.6 mmHg increase in diastolic BP (Nawrot et al.. 2002). On a population-wide basis,
26 the estimated effect size could translate into a clinically significant increase in the
27 segment of the population with the highest BP. In a moderately-sized population, a
28 relatively small effect size thus has important health consequences for the risk of
29 sequelae of increased BP, such as stroke, myocardial infarction, and sudden death. It was
30 also noted that most of the reviewed studies examining bone Pb levels, biomarkers of
31 cumulative Pb exposure, also showed increased BP (Cheng etal.. 2001; Huetal.. 1996a)
32 or increased hypertension with increasing bone Pb level (Lee etal.. 200 la). Across
33 studies, over a range of bone Pb concentrations (<1.0 to 96 ug/g), every 10 ug/g increase
34 in bone Pb was associated with increased odds ratios of hypertension between 1.28 and
November 2012 5-291 Draft - Do Not Cite or Quote
-------
1 1.86. Studies observed an average increase in systolic BP of-0.75 mmHg for every
2 10 ug/g increase in bone Pb concentration over a range of <1 to 52 ug/g.
3 With regard to etiologically-relevant timing of Pb exposure, toxicological evidence
4 demonstrated increases in BP after long-term (>4 weeks) Pb exposure. In epidemiologic
5 studies, cardiovascular outcomes were most often examined in cross-sectional studies
6 with one or a limited number of Pb biomarker measurements, so uncertainty exists as to
7 the specific Pb exposure level, timing, frequency, and duration that contributed to the
8 observed associations. While associations of adult bone Pb (particularly tibia Pb) with
9 health outcomes in adults are indicative of effects related to past or cumulative exposures,
10 interpretation of similar associations involving adult blood Pb levels, especially those
11 measured concurrently with outcomes, is complicated by the higher past exposures
12 generally observed in U.S. adults populations. Detailed interpretation of Pb in blood and
13 bone are provided in Sections 4.3 and 4.7.3. Briefly, higher past Pb exposures in adults
14 increased their bone Pb stores which contribute to current blood Pb levels through the
15 normal process of bone remodeling, as well as periods of increased bone remodeling and
16 loss (e.g., osteoporosis and pregnancy). Due to the long latency period for the
17 development of increased BP and CVD, associations of cardiovascular effects with low
18 concurrent blood Pb levels (e.g., population means 1.6-4 ug/dL) in adults may be
19 influenced by higher past Pb exposures (Section 4.4.1).
20 Past air Pb concentration and blood Pb data provide context for the cardiovascular
21 studies. Section 3.2 notes that the peak U.S. use of Pb anti-knock additives in automobile
22 gasoline occurred between 1968 and 1972 and was finally banned from use in 1996.
23 Section 3.5 shows that air Pb measured at trends monitors across the U.S. decreased from
24 1.3 (ig/m3 in 1980 to 0.14 (ig/m3 in 2010. Many of the monitors reporting to the trends
25 network were more recently influenced only by Pb sources; the mean 2010 3-month
26 rolling average for non-source monitors was an order of magnitude lower than the 2010
27 trends site average. Collective review of blood Pb studies from the late 1960s and 1970s,
28 including NHANES II (1976-1980) suggest that blood Pb levels ranged from roughly 10
29 to 30 (ig/dL (Pirkle et al.. 1994; Billick et al.. 1979; Tepper and Levin. 1975; Fine et al..
30 1972).
November 2012 5-292 Draft - Do Not Cite or Quote
-------
1 This section reviews the published studies pertaining to the cardiovascular effects of Pb
2 exposure in humans, experimental animals, isolated vascular tissues, and cultured
3 vascular cells. With the large and strong existing body of evidence serving as the
4 foundation, emphasis was placed on studies published since the 2006 Pb AQCD.
5 Epidemiologic and toxicological studies continued to augment the evidence for increases
6 in BP and hypertension development associated with long-term Pb exposure and
7 expanded the evidence for the biological pathways of these effects. Epidemiologic studies
8 strengthened the evidence for associations between Pb biomarkers and cardiovascular
9 effects after adjusting for potential confounding factors such as age, SES, diet, alcohol
10 use, BMI, comorbidities, and smoking. Emphasis was placed on studies that had
11 extensive consideration for confounding and prospective study designs. The
12 epidemiologic evidence was substantiated with results from several available prospective
13 studies demonstrating the directionality of effects by indicating associations between Pb
14 biomarkers and the subsequent incidence of cardiovascular health effects.
5.4.2 Blood Pressure and Hypertension
5.4.2.1 Epidemiology
15 The most commonly used indicator of cardiovascular morbidity was increased BP and its
16 derived index, hypertension. Hypertension in these studies was defined as diastolic and/or
17 systolic BP above certain cut-points or use of anti-hypertensive medicines. The BP cut-
18 points were established by reference to informed medical opinion, but BP cut-points
19 defining hypertension have been lowered over time, as medical knowledge has improved.
20 Consequently, different studies using "hypertension" as a cardiovascular outcome may
21 have assigned different cut-points, depending on the year and location of the study and
22 the individual investigator. All of the recent studies in the current review used the same
23 criteria for hypertension (e.g., systolic BP at or above 140, diastolic BP at or above 90, or
24 use of anti-hypertensive medications). Studies in the medical literature show that elevated
25 BP is associated with increased risk of CVD including coronary disease, stroke,
26 peripheral artery disease, and cardiac failure. Coronary disease (i.e., myocardial
27 infarction, angina pectoris, and sudden death) is the most lethal sequelae of hypertension
28 (Ingelsson et al., 2008; Chobanian et al., 2003; Pastor-Barriuso et al., 2003; Prospective
29 Studies Collaboration. 2002; Kannel. 2000a. b; Neatonetal. 1995).
30 Earlier, U.S. EPA (1990a) reviewed the then available studies examining Pb exposure
31 and BP and hypertension outcomes which included evaluation of several studies
32 conducting analysis of the data in NHANES II (1976-80). They noted that across a range
November 2012 5-293 Draft - Do Not Cite or Quote
-------
1 of 7 to 34 (ig/dL, no evident threshold was found below which blood Pb was not
2 significantly related to blood pressure. U.S. EPA Q990a) concluded that a small but
3 positive association exists between blood Pb levels and increases in blood pressure.
4 Quantitatively, the relationship appears to hold across a wide range of blood Pb values
5 and, furthermore, an estimated mean increase of about 1.5-3.0 mmHg in systolic blood
6 pressure appears to occur for every doubling of blood Pb concentration in adult males and
7 something less than 1.0-2.0 mmHg for adult females. U.S. EPA Q990a) further
8 concluded that the plausibility of these relationships observed in epidemiologic studies of
9 human populations being of a causal nature is supported by controlled experimental
10 animal studies demonstrating increased blood pressure clearly attributable to Pb.
11 Subsequently, the 2006 Pb AQCD (U.S. EPA. 2006b) reviewed the literature examining
12 Pb exposure and effects of BP and hypertension, published after the 1990 document as
13 discussed in Section 5.4.1.
14 Several recent general population and occupational cohort and cross-sectional studies
15 strengthened the evidence for associations of blood and bone Pb levels with measures of
16 BP (Figure 5-18 and Table 5-18) and with the prevalence and incidence of hypertension
17 (Figure 5-19 and Table 5-19). Further, recent studies expanded evidence, finding
18 differences in association among racial/ethnic groups, perceived stress, diet, and genetic
19 variants, and thus, identified populations potentially at increased risk of Pb-associated
20 cardiovascular effects.
21 In a cross-sectional analysis, Martin et al. (2006) examined the associations of concurrent
22 blood and tibia Pb levels with BP and hypertension in a large, community-based study of
23 older adults (n = 964, age ranging from 50 to 70 years) in Baltimore, MD. Although
24 cross-sectional in design, a key strength of this study was the extensive consideration of
25 potential confounding variables. Four models evaluated associations for BP and
26 hypertension. The base model included age, sex, BMI, sodium intake, potassium intake,
27 total cholesterol, time of day, testing technician, and hypertensive medication use. Other
28 models added SES, race/ethnicity, or both as covariates. Blood Pb but not tibia Pb level
29 was a strong predictor of BP in all models; a 1 (ig/dL increase in concurrent blood Pb
30 level was associated with an approximately 1 mmHg increase in systolic BP and an
31 approximately 0.5 mmHg increase in diastolic BP. Tibia Pb but not blood Pb was
32 associated with hypertension in logistic regression models. The authors applied
33 propensity analysis to their models to better account for the effect of other risk factors for
34 hypertension such as race/ethnicity, age, and SES that were strongly associated with tibia
35 Pb level. The propensity score analysis and model adjustment did not substantially
36 change the numerical findings and conclusions (e.g., tibia Pb and hypertension were
37 positively associated independently of race/ethnicity and SES), indicating that neither
38 SES nor race/ethnicity confounded the association between tibia Pb level and
November 2012 5-294 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
hypertension. No evidence for effect modification by race/ethnicity was found either.
Martin et al. (2006) concluded that Pb in blood has a short term effect on BP and that Pb
contributes to hypertension risk as a function of cumulative, chronic exposure (as
represented as bone Pb in this population). While different aspects of Pb exposure may
contribute differentially to increases in BP and hypertension, it is important to note that
concurrent blood Pb levels in adults also reflect cumulative Pb exposure. Thus, its
association with BP may not reflect a short term effect but may also reflect an effect of
cumulative Pb exposure.
Reference
Martin etal. (2006)
Glenn etal. (2006)
Weaver etal. (2008)
Scinicariello etal. (2010)
Martin etal. (2006)
Glenn etal. (2006)
Peters etal. (2007)
Elmarsafawy etal. (2006)
Weaver etal. (2008)
Peters etal. (2007)
Population
Adults, Baltimore, MD
Korean Pb Workers
Korean Pb Workers
NHANES III - Blacks
NHANES III - Mexicans
Adults, Baltimore, MD
Korean Pb Workers
NASmen, High Stress
NASmen, Low Stress
NASmen, High calcium
NASmen, Low calcium
Korean Pb Workers
NASmen, High Stress
Pb Distribution a
2.9(2.0,4.4)
27.2(19.3,28.3)
1.4(0.6,3.6)
2.0(1.0,3.9)
15.7(10.5,23.5)
18.1(12.2,26.9)
18.1(12.2,26.9)
21.6(12.0)
21.6(12.0)
74.3(67.3,82.0)
26.9(18.4,39.3)
PbBiomarker SBP
Blood Pb — . —
Blood Pb (concurrent) ™«
Blood Pb (longitudinal) — .... —
Blood Pb — .,—
In Blood Pb — —
In Blood Pb — —
Tibia Pb — ' —
Tibia Pb (historical) — —
Tibia Pb ' —' • —
Tibia Pb — '- —
Tibia Pb — '.—
Tibia Pb . — ' —
Patella Pb
Patella Pb • — - —
Martin etal. (2006)
Scinicariello etal. (2010)
Martin etal. (2006)
Perlstein etal. (2007)
Zhang etal. (2010)
Adults, Baltimore, MD
NHANES III-Whites
NHANES III - Blacks
NHANES III - Mexicans
Adults, Baltimore, MD
NAS men
NAS men
N AS men
N AS men
N AS men
N AS men
N AS men
N AS men
NASmen, HFE Wild-type
NASmen, HFE H63D
NASmen, HFE C282Y
NAS men, Any HFE variant
NASmen, HFE Wild-type
NASmen, HFE H63D
NASmen, HFE C282Y
NAS men, Any HFE variant
2.9(2.0,4.4)
1.6(0.8,3.3)
1.4(0.6,3.6)
2.0(1.0,3.9)
15.7(10.5,23.5)
12.4(4.4)
7.4 (0.6)
5.4 (0.5)
3.9(0.3)
40.9(14)
29.4(2.2)
18.9(1.4)
14.1(1.4)
18(12,27)
19(14,26)
20(14,27)
19(14,27)
26(17,34)
27(19,37)
25(17,37)
26(18,37)
Blood Pb
In Blood Pb
In Blood Pb
In Blood Pb
Tibia Pb
Blood Pb
Blood Pb
Blood Pb
Blood Pb
Tibia Pb
Tibia Pb
Tibia Pb
Tibia Pb
Tibia Pb
Tibia Pb
Tibia Pb
Tibia Pb
Patella Pb
Patella Pb
Patella Pb
Patella Pb
DBF
PP
5-6-4-20 2 4 6 £
Change in BP (mmHg 95% Cl) per
lu.g/dL increase in blood Pb or 10 u.g/gbone Pb
aPb distributions present the median (IQR), which were estimated from the mean and SD assuming a normal distribution.
bEffect estimates were standardized to 1 ug/dL blood Pb or 10 |jg/g bone Pb.
Note: In general, results are categorized by specific BP parameter, then by Pb biomarker. For categories with multiple studies, the
order of the studies follows the order of discussion in the text. Results display associations (95% Cl) of a 1 ug/dL increase in blood
Pb level (closed circles) or 10 |jg/g increase in bone Pb (open circles) with systolic BP (SBP; blue), diastolic BP (DBP; red), and
pulse pressure (PP; purple) in adults.
Figure 5-18 Associations of blood and bone Pb levels with systolic BP,
diastolic BP, and pulse pressure in adults.
November 2012
5-295
Draft - Do Not Cite or Quote
-------
Table 5-18 Additional characteristics and quantitative data for associations of
blood and bone Pb with BP measures for studies presented in
Figure 5-18.
Study
Martin et al.
(2006)
Glenn et al.
(2006)
Weaver et
al. (2008)
Study Population
/ Methodology
Cross-sectional
964 men and
women, 50-70 yr,
40% African
American, 55%
White, 5% other, in
Baltimore, MD
Longitudinal
575 Pb exposed
workers, age 18-65
yr, in South Korea
(10/1997-6/2001)
Cross-sectional
652 current and
former Pb workers
in South Korea
(12/1999-6/2001)
Same cohort as
Glenn et al. (2006)
Parameter Pb Data
BP Concurrent Mean
Blood Pb:
Mean (SD): 3.5
(2.3) ug/dL
African American: 3.4
(2.3)
White: 3.5 (2.4)
Tibia Pb:
Mean(SD): 18.8
(12.4) ug/g
African American:
21.5(12.6)
White: 16.7(11.9)
BP Blood Pb mean (SD):
Visit 1 : 20.3 (9.6),
Women
Visit 2: 20.8(10.8),
Women
Visits: 19.8(10.7),
Women
Visit 1: 35.0(13.5),
Men
Visit 2: 36.5 (14.2),
Men
Visit 3: 35.4 (15.9),
Men
Tibia Pb, mean (SD):
Visit 1: 28.2(19.7),
Women
Visit 2: 22.8 (20.9),
Women
Visit 1 : 41 .7 (47.6),
Men
Visit 2: 37.1 (48.1),
Men
Patella Pb, mean
(SD):
Visit 3 49.5 (38.5)
Women
Visit 3 87.7 (11 7.0)
Men
BP Concurrent Blood Pb:
Mean (SD):
30.9(16.7) ug/dL
Concurrent Patella
Pb:
Mean (SD):
75.1 (101.1) ug/g
Statistical Analysis
Extensive analysis of
potential confounding
factors. Multiple linear
regression base model
adjusted forage, sex, BMI,
a nti hypertensive medication
use, dietary sodium intake,
dietary potassium intake,
time of day, testing
technician, serum total
cholesterol. SES, race/
ethnicity also included in
models that are presented in
Figure 5-18. and tabulated
here.)
Multivariable models using
GEE were used in
longitudinal analyses.
Models were adjusted for
visit number, baseline age,
baseline age squared,
baseline lifetime alcohol
consumption, baseline body
mass index, sex, baseline
BP lowering medication use,
alcohol consumption, BMI,
sex, BP lowering medication
use.
Linear regression model
adjusted forage, sex, BMI,
diabetes, antihypertensive
and analgesic medication
use, Pb job duration, work
status, tobacco and alcohol
use
Effect Estimate
P (95% Cl)
Blood Pb:
SBP: 1.05(0.53,1.58)
DBP: 0.53 (0.25,
0.81)
mmHg per ug/dL
blood Pb
Tibia Pb:
SBP: 0.07 (-0.05,
0.14)
DBP: 0.05 (-0.02,
0.08)
mmHg per ug/g bone
Pb
Model 1 (short-term)
Blood Pb
(longitudinal):
0.09(0.01, 0.16)
Blood Pb
(concurrent):
0.08 (-0.01, 0.1 6)
Model 4 (short and
longer-term)
Blood Pb
(longitudinal):
0.09(0.01, 0.16)
Blood Pb
(concurrent):
0.10(0.01,0.19)
mmHg per 10 ug/dL
blood Pb
SBP
Patella Pb:
0.0059 (-0.008, 0.02)a
Blood Pb:
0.1007(0.02, 0.1 8)a
mmHa oer 1 ua/dL
blood Pb or 1 ug/g
patella Pb
Interaction between
blood Pb/patella Pb
with ALAD and
vitamin D receptor
polymorphisms not
significant.
November 2012
5-296
Draft - Do Not Cite or Quote
-------
Study
Perlstein et
al. (2007)
Peters et al.
(2007)
Elmarsafawy
et al. (2006)
Study Population
/ Methodology
Cross-sectional
593 predominantly
white men from
MAS in
Greater Boston,
MA area
(1991-1997)
Longitudinal and
Cross-sectional
513 elderly men
(mean 67 yr) from
MAS in Greater
Boston, MA area
Cross-sectional
471 elderly men
(mostly white,
mean age 67 yr)
from MAS in
Greater Boston,
MA area
Parameter Pb Data
PP Blood Pb:
Overall mean (SD):
6.12(4.03)ug/dL
Mean (SD) quintiles:
Q1: 2.3(0.8) ug/dL
Q2: 3.9 (0.3) ug/dL
Q3: 5.4 (0.5) ug/dL
Q4: 7.4 (0.6) ug/dL
Q5: 12.4(4.4) ug/dL
Tibia Pb:
Median: 19 ug/g
Mean (SD) quintiles:
Q1: 7.4 (3.2) ug/g
Q2:14.1 (1.4) ug/g
Q3: 18.9(1.4) ug/g
Q4: 24.9 (2.2) ug/g
Q5: 40.9 (14) ug/g
BP Tibia Pb:
mean (SD):
21 .5 (13.4) ug/g
Patella Pb:
Mean (SD):
31 .5 (19.3) ug/g
BP Blood Pb:
Mean (SD):
6.6 (4.3) ug/dL
Tibia Pb:
Mean (SD):
21 .6 (12.0) ug/g
Patella Pb:
Mean (SD):
31 7 (183) uq/q
\/r^yy
Statistical Analysis
BP association assessed
using Spearman correlation
coefficients.
PP association (adjusted
mean difference) assessed
using multiple linear
regression model adjusted
forage, height, race, heart
rate, waist circumference,
diabetes, family history of
hypertension, education level
achieved, smoking, alcohol
intake, fasting plasma
glucose, and ratio of total
cholesterol to HDL
cholesterol
Logistic and linear
regression models adjusted
for age, age squared,
sodium, potassium, and Ca2+
intake, family history of
hypertension, BMI,
educational level, pack-years
of smoking, alcohol
consumption, and physical
activity
Linear regression models
adjusted forage, BMI, family
history of hypertension,
history of smoking, dietary
sodium intake, and
cumulative alcohol ingestion
Lack of consideration for
potential confounding by
SES-related variables.
Effect Estimate
o in co/ f*t\
p (95% Cl)
PP
4.2(1.9, 6.5) mmHg
higher in men with
tibia Pb >19 ug/g
(median) compared
with men with tibia Pb
800 mg/day):
0.40.(0.1 1,0.70)
2+
Low Ca group
(<800 mg/day):
0.19(0.01,0.37)
mmHg per ug/g tibia
Pb
November 2012
5-297
Draft - Do Not Cite or Quote
-------
Study Population
Study / Methodology
Zhang et al. Cross-sectional
(20K)a) 61 9 older adult
males (mostly
white, mean
age 67 yr) enrolled
in the MAS in
Greater Boston,
MA area
Scinicariello Cross-sectional
etal. (2Q10) 6,016 NHANES III
(1988-1994)
participants a 17 yr
Parameter Pb Data
PP Wild type HFE
Tibia Pb:
Median (IQR):
8(1 2-27) ug/g
Patella Pb:
Median (IQR):
26(1 7-37) ug/g
C282Y HFE
Tibia Pb:
Median (IQR):
20 (14-27) ug/g
Patella Pb:
Median (IQR):
25(1 7-37) ug/g
H63D HFE
Tibia Pb:
Median (IQR):
19(1 4-26) ug/g
Patella Pb:
Median (IQR):
27(19-37)ug/g
BP Concurrent Blood Pb:
Overall Mean (SE):
2.99 (0.09) ug/dL
Non-Hispanic Whites:
2.87 (0.09)
Non-Hispanic Blacks
3.59 (0.20)
Mexican American
3.33(0.11)
Statistical Analysis
Linear mixed effects
regression models with
repeated measurements
adjusted for age; education;
alcohol intake; smoking;
daily intakes of Ca2+, sodium,
and potassium; total calories;
family history of
hypertension; diabetes;
height; heart rate; high-
density lipoprotein (HDL);
total cholesterol:HDL ratio;
and waist circumference
Multivariable linear
regression of log-
transformed blood Pb level
adjusted for age, sex,
education, smoking status,
alcohol intake, BMI, serum
creatinine levels, serum
Ca2+, glycosylated
hemoglobin, and hematocrit
Effect Estimate
o in co/ f*t\
p (95% Cl)
PP
mmHg
per 13 ug/g Tibia Pb:
Wild Type HFE:
0.38(0,1.96)
H63D HFE:
3.30(0.16, 6.46)
pOQOY HFF-
w^o^ i nrQ.
0.89 (0, 5.24)
Any HFE variant:
2.90(0.31, 5.51)
mmHci
per 19 ug/g Patella
Pb:
Wild Type HFE:
0.26(0, 1.78)
H63D HFE:
2.95 (0, 5.92)
C282Y HFE:
0.55(0, 1.66)
Any HFE variant:
2.83 (0.32,5.37)
SBP
Non-Hispanic whites:
1 .05 (0.32, 1 .78)
Non-Hispanic blacks:
2.55(1.59, 3.51)
Mexican Americans:
0.84 (-0.06, 1 .74)
DBP
Non-Hispanic whites:
-0.14 (-1.1, 0.82)
Non-Hispanic blacks:
1.99(1.13,2.85)
Mexican Americans:
0.74 (-0.005, 1.48)
mmHg per unit
increase in
In [Blood Pb]
Significant
interactions with
blood PbandALAD
observed in relation
to SBP for non-
Hispanic whites and
non-Hispanic blacks
November 2012
5-298
Draft - Do Not Cite or Quote
-------
Study Population
Study / Methodology Parameter Pb Data
Navas-Acien Longitudinal and BP
et al. (2008)" Cross-sectional
Meta-analysis of
studies using bone
Pb as an exposure
metric and BP as
the outcome
(8 studies)
Yazbeck et Cross-sectional BP Midpregnancy Blood
al. (2009)' 971 pregnant Pb:
women, age 18-45 PIH group mean
yr, (SD):
in France 2.2 (1 .4)
No PIH group mean
(SD):
1.9(1.2)
Statistical Analysis
Inverse variance weighted
random-effects meta-
analyses
Multivariable logistic
regression models adjusted
for maternal age; Cd, Mn,
and Se blood levels;
hematocrit; parity; BMI;
pregnancy weight gain;
gestational diabetes;
educational level; SES;
geographic residence; and
smoking status and alcohol
consumption before and
during pregnancy
Effect Estimate
P (95% Cl)
BP Pooled Estimates
mmHg
per 10 ug/g Tibia Pb
Prospective SBP
0.33 (-0.44, 1.11)
Cross-sectional SBP
0.26 (0.02, 0.50)
Cross-sectional DBP
0.02 (-0.15, 0.19)
Hypertension
per 10 ug/g patella
Pb
Cross-sectional
hypertension
OR: 1.04 (1.01, 1.07)
Pooled Estimate
hypertension
OR: 1.04(0.96, 1.12)
Log-transformed
blood Pb at mid-
pregnancy
SBP:
r = 0.08; p = 0.03
DBP:
r = 0.07; p = 0.03
Significant
correlations also
observed after
24 weeks of gestation
and after 36 weeks of
gestation.
a95% CIs estimated from given p-value.
bReference not included in Figure 5-18. because it is a meta-analysis.
""Reference not included in Figure 5-18. because only correlations were reported
November 2012
5-299
Draft - Do Not Cite or Quote
-------
Martin etal. (2006) Baltimore, MD
Elmarsafawy etal. (2006) NAS
Yazbecketal. (2009)
Muntner etal. (2005)
Scinicarielloet al. (2010) NHANES III
1998-1994
Park etal. (2009)
Study Location Population Blood Pbc(ug/dL) Comparison
perlu.g/dL
perlu.g/dL
perlu.g/dL
Reference
Q2vQl
Q3vQl
Q4vQl
Reference
Q2vQl
Q3vQl
Q4VQ1
Reference
Q2vQl
Q3vQl
Q4vQl
Reference
Q2vQl
Q3vQl
Q4vQl
Reference
Q2VQ1
Q3vQl
Q4vQl
ALAD2vlb
Reference
Q2VQ1
Q3vQl
Q4vQl
ALAD2vlb
Reference
Q2vQl
Q3vQl
Q4vQl
ALAD2vlb
perlu.g/dL
White Men
BlackMen
White Women
BlackWomen
Men<50
Men>50
Women <50
Women >50
LowCalcium
High Calcium
Pregnant Women
Non-Hispanic Whites
Non-Hispanic Blacks
Mexican Americans
Non-Hispanic Whites
Non-Hispanic Blacks
Mexican Americans
Overall
3.5(2.3)
6.6(4.3)
6.6(4.3)
1.2-1.7
1.71-2.30
>2.30
<1.06
1.06-1.63
1.63-2.47
>2.47
<1.06
1.06-1.63
1.63-2.47
>2.47
<1.06
1.06-1.63
1.63-2.47
>2.47
0.7-1.4
1.5-2.3
2.4-3.7
3.8-52.9
2.4-3.7
0.7-1.4
1.5-2.3
2.4-3.7
3.8-52.9
2.4-3.7
0.7-1.4
1.5-2.3
2.4-3.7
3.8-52.9
2.4-3.7
3.52(0.10)
Elmarsafawy etal. (2006)
Martin etal. (2006) Baltimore, MD
Petersetal. (2007) Boston, MA
Petersetal.(2007) Boston, MA
Elmarsafawy etal. (2006) NAS
LowCalcium
High Calcium
High Stress
High Stress
LowCalcium
High Calcium
Tibia Pbd(ug/g)
21.6(12.0)
21.6(12.0)
18.8(12.4)
21.5(13.4)
Patella Pb d (ug/g)
31.5(19.3)
31.7(18.3)
31.7(18.3)
perlO|ag/g
perlO|ag/g
perlO|ag/g
perlO|ag/g
perlO|ag/g
perlO|ag/g
perlO|ag/g
-O*-
23456
Odds Ratio (95% Cl)
Note: Studies are categorized by Pb biomarker. Within each category, studies generally are presented in order of discussion in the
text.
a: The outcomes plotted are hypertension prevalence with the exception of Yazbeck et al. (2009) which measured pregnancy
induced hypertension and Peters et al. (2007) which measured hypertension incidence.
b: ALAD2 vs. 1 indicates comparison between ALAD 2 carriers (e.g., ALAD1-2 and ALAD2-2) and ALAD 1 homozygotes
(e.g., ALAD1-1).
c: Effect estimates were standardized to a 1 ug/dL increase in blood Pb (closed circles).
d: Effect estimates were standardized to a 10 ug/g increase in bone Pb (open circles).
Figure 5-19 Odds ratios (95% Cl) for associations of blood (closed circles)
and bone (open circles) Pb with hypertension prevalence and
incidence9.
November 2012
5-300
Draft - Do Not Cite or Quote
-------
Table 5-19 Additional characteristics and quantitative data for results presented
in Figure 5-19 for associations of blood and bone Pb with
hypertension measures.
Study
(same order
as in text)
Martin et al.
(2006)
Weaver et
al. (2008)a
Peters et al.
(2007)
Elmarsafawy
et al. (2006)
Study
Population and
Methodology
Cross-sectional
964 men and
women, 50-70 yr,
40% African
American, 55%
White,
5% other,
in Baltimore, MD
Cross-sectional
652 current and
former Pb
workers in
South Korea
(12/1999-6/2001)
Longitudinal
51 3 elderly men
(mean 67 yr)
from MAS in
Greater Boston,
MA area
Cross-sectional
471 elderly men
(mean 67 yr)
from MAS in
Greater Boston,
MA area
Parameter
Hypertension
(current use of
antihypertensive
medication,
mean SBP
a 140 mmHg or
DBP > 90
mmHg)
Hypertension
(mean SBP
> 140 mmHg,
DBP > 90
mmHg; and/or
use of
antihypertensive
medications; or
physician
diagnosis)
Hypertension
(mean SBP
>140 mmHg,
DBP >90
mmHg; or
physician
diagnosis)
Hypertension
(mean SBP
> 160 mmHg,
DBP > 95
mmHg; and/or
physician
diagnosis with
current use of
antihypertensive
medications)
Pb Data
Blood Pb:
Mean (SD):
3.5 (2.3) ug/dL
Tibia Pb:
Mean (SD):
18.8(1 2.4) ug/g
Blood Pb:
Mean (SD):
31.9(14.8)ug/dL
Patella Pb:
Mean (SD):
37.5 (41 .8) ug/g
Tibia Pb:
mean (SD):
21 .5 (13.4) ug/g
Patella Pb:
Mean (SD):
31 .5 (19.3) ug/g
Blood Pb:
Mean (SD):
6.6 (4.3) ug/dL
Tibia Pb:
Mean (SD):
21 .6 (12.0) ug/g
Patella Pb:
Mean (SD):
31 .7 (18.3) ug/g
Statistical Analysis
Logistic regression
models adjusted for age,
sex, BMI,
antihypertensive
medication use, dietary
sodium intake, dietary
potassium intake, time
of day, testing
technician, and serum
homocysteine
Lack of consideration for
potential confounding by
SES-related variables.
Logistic regression
models adjusted for age,
sex, BMI, diabetes,
antihypertensive and
analgesic medication
use, Pb job duration,
work status, tobacco
and alcohol use
Cox proportional
hazards models
adjusted for age, age
squared, sodium,
potassium, and Ca +
intake, family history of
hypertension, BMI,
educational level,
smoking, alcohol
consumption, baseline
SBP and DBP, and
physical activity
Logistic regression
models adjusted for age,
BMI, family history of
hypertension, history of
smoking, dietary sodium
intake, and cumulative
alcohol ingestion
Effect Estimate
(95% Cl)
Blood Pb level:
OR=1.02(0.87,
1.19)
Tibia Pb:
OR=1.24(1.05,
1.47)
Quantitative results
not reported. None
of the examined Pb
exposure metrics
(blood, patella, and
In patella) were
significantly
associated with
hypertension
Risk of
Hypertension
Incidence
High Stress
RR=2.66(1.43,
4.95) per SD
increase in tibia Pb
RR=2.64(1.42,
4.92) per SD
increase in patella
Pb
Low Ca2+ group
(<800 mg/day):
Blood Pb OR:
1.07(1.00,1.15)
Tibia Pb OR:
1.02(1.00, 1.04)
Patella Pb OR:
1.01 (1.00,1.03)
High Ca2+ group
(>800 mg/day):
Blood PbOR:
1.03(0.97,1.11)
Tibia Pb OR:
1.01 (0.97,1.04)
Patella Pb OR:
1.01 (0.99, 1.03)
Per ug/dL blood Pb
or ug/g tibia or
patella Pb
November 2012
5-301
Draft - Do Not Cite or Quote
-------
Study
(same order
as in text)
Study
Population and
Methodology
Parameter
Pb Data
Statistical Analysis
Effect Estimate
(95% Cl)
Yazbeck et
al. (2QQ9)a
Cross-sectional
971 pregnant
women, age
18-45yr, in
France
PIH
(SBP> 140
mmHg or DBP
> 90 mmHg
after the
22nd week of
gestation)
Blood Pb:
PIH group mean (SD):
2.2 (1 .4) ug/dL
No PIH group mean
(SD):
1 .9 (1 .2) ug/dL
Q1: <1.20 ug/dL
Q2: 1.20-1.70 ug/dL
Q3: 1.71-2.30 ua/dL
Multivariable logistic
regression models
adjusted for maternal
age, Cd, Mn, and Se
blood levels, parity,
hematocrit, BMI,
gestational diabetes,
educational levels, SES,
geographic residence,
and smoking status
during pregnancy
PIH
Blood Pb
OR=
3.29(1.11,9.74)
per 1 unit increase
in log maternal
blood Pb level
Q1 : Reference
arouo
Q4: >2.30 ug/dL
Q2: OR 1.84 (0.77,
4.41)
Q3: OR=2.07 (0.83,
5.13)
Q4: OR=2.56 (1.05,
6.22)
Muntner et
al. (2005)
Cross-sectional
9,961 NHANES
(1999-2002)
participants
Hypertension
(current use of
antihypertensive
medication,
SBP> 140
mmHg, or DBP
> 90 mmHg)
Concurrent Blood Pb:
Overall Mean (Cl):
1.64 (1.59-1.68) ug/dL
quartile 1:
<1.06 ug/dL,
quartile 2:
1.06-1.63 ug/dL,
quartile 3:
1.63-2.47 ug/dL, and
quartile 4:
> 2.47 ug/dL
Multivariable logistic
regression models
adjusted forage, sex,
diabetes mellitus, BMI,
cigarette smoking,
alcohol consumption,
high school education,
and health insurance
status
Monotonic increase in
OR across blood Pb
level groups.
Non-Hispanic white:
Q1: Reference
group
Q2: OR=1.12(0.83,
1.50)
Q3:OR=1.03(0.78,
1.37)
Q4: OR=1.10(0.87,
1.41)
Non-Hispanic black
Q1: Reference
group
Q2:OR=1.03(0.63,
1.67)
Q3: OR=1.12(0.77,
1.64)
Q4: OR=1.44 (0.89,
2.32)
Mexican American
Q1: Reference
group
Q2: OR=1.42 (0.75,
2.71)
Q2: OR=1.48 (0.89,
2.48)
Q3:OR=1.54(0.99,
2.39)
p for trend=0.04
November 2012
5-302
Draft - Do Not Cite or Quote
-------
Study
(same order
as in text)
Study
Population and
Methodology
Parameter
Pb Data
Statistical Analysis
Effect Estimate
(95% Cl)
Scinicariello
et al. (2010)
Cross-sectional
6,016 NHANES
111(1988-1994)
participants
> 17yr
Hypertension
(current use of
antihypertensive
medication,
SBP> 140
mmHg, or DBP
> 90 mmHg)
Concurrent Blood Pb:
Mean (SE):
2.99 (0.09) ug/dL
Q1:0.7-1.4ug/dL,
Q2: 1.5-2.3 ug/dL,
Q3: 2.4-3.7 ug/dL,
Q4: 3.8-52.9 ug/dL
Non-Hispanic Whites:
2.87 (0.09)
Non-Hispanic Blacks:
3.59 (0.20)
Mexican American:
3.33(0.11)
Multivariable logistic
regression model
adjusted for
race/ethnicity, age, sex,
education, smoking
status, alcohol intake,
BMI, serum creatinine
levels, serum Ca2+,
glycosylated
hemoglobin, and
hematocrit
Non-Hispanic
whites:
Q1: Reference
group
Q2: POR=1.21
(0.66, 2.24)
Q3: POR=1.57
(0.88, 2.80)
Q4: POR=1.52
(0.80, 2.88)
ALAD1-2/2-2:
POR= 0.76 (0.17,
3.50)
ALAD-1: Reference
group
Non-Hispanic
blacks:
Q1: Reference
Q2: POR=1.83
(1.08, 3.09)
Q3: POR=2.38
(1.40, 4.06)
Q4: POR=2.92
(1.58,5.41)
ALAD1-2/2-2:
POR= 3.40 (0.05,
219.03)
ALAD-1: Reference
group
Mexican Americans:
Q1: Reference
Q2: POR=0.74
(0.24, 2.23)
Q3: POR=1.43
(0.61, 3.38)
Q4: POR=1.27
(0.59, 2.75)
ALAD1-2/2-2:
POR= 0.49 (0.08,
3.20)
ALAD-1: Reference
group
POR for
hypertension with
ALAD2 carriers
across quartiles of
blood Pb level also
reported. ALAD2
carriers associated
with hypertension in
non-Hispanic
whites.
November 2012
5-303
Draft - Do Not Cite or Quote
-------
Study
(same order
as in text)
Parket al.
(2QQ9c)
Study
Population and
Methodology Parameter
Cross-sectional Hypertension
12,500 NHANES
111(1988-1994)
participants
Pb Data
NHANES III Concurrent
Blood Pb Mean and SE
3.52(0.10)
White men
<50yr4.02(0.16)
> 50 yr 4.92 (0.1 8)
Rlspk' mpn
DlaUt\ 1 1 Id 1
<50yr4.55(0.15)
> 50 yr 7.57 (0.22)
White women
<50 yr 2.09 (0.07)
> 50 yr 3.53 (0.1 2)
Black women
<50 yr 2.52 (0.09)
> 50 yr 4.49 (0.1 6)
Statistical Analysis
Logistic regression
models adjusted for age,
education, smoking
status, cigarette
smoking, BMI,
hematocrit, alcohol
consumption, physical
activity, antihypertensive
medication use, and
diagnosis of type-2
HJ3 hpfpQ
UICI UClCO
Effect Estimate
(95% Cl)
OR per SD
(0.75 ug/dL) in log
blood Pb:
Overall: 1.12(1.03,
1.23).
White men:
1.06(0.92,1.22)
Black men:
1.17(0.98, 1.38)
White women:
1.16(1.04,1.29)
Black women:
1.19(1.04, 1.38)
Men <50 yr:
0.98 (0.80, 1 .22)
Men >50 yr:
1.20(1.02,1.41)
Women <50 yr:
1 .23 (1 .04, 1 .46)
Women >50 yr:
1 .09 (0.94, 1 .26)
aNot included in Figure 5-19 because OR data were not reported.
1
2
o
5
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
In an occupational cohort in South Korea, Glenn et al. (2006) simultaneously modeled
multiple Pb dose measures of individuals collected repeatedly over four years of follow
up. Thus, through the assessment of cross-sectional and longitudinal relationships with
BP, this study provided key insight on potentially important time periods of Pb exposure
and also informed the directionality of association. The initial blood Pb level was used as
a baseline covariate and the difference in blood Pb level between visits was computed for
each subsequent visit. The bone Pb measures (tibia Pb at visits 1 and 2, patella Pb at visit
3) were used to indicate historical exposure and cumulative dose. Four models were
specified: Model 1 was conceptualized to reflect short-term changes in BP associated
with recent dose; Model 2 to reflect longer-term changes associated with cumulative dose
controlling for the association of baseline BP with recent dose; Model 3 to reflect longer-
term changes associated with cumulative dose controlling for cross-sectional influence of
cumulative dose on baseline BP; and Model 4 to reflect both short-term change with
recent dose and longer-term change with cumulative dose. Concurrent blood Pb and
increases in blood Pb between visits were associated with increases in systolic BP in
Model 1 (short-term dose) and Model 4 (short- and longer-term dose). No association
was observed between BP and tibia Pb at baseline while higher tibia Pb was associated
with a decrease in systolic BP in each of the models.
Glenn et al. (2006) was strengthened by the analysis of associations between changes in
blood Pb and changes in BP overtime within individual subjects. These results indicate
that circulating Pb (e.g., blood Pb) may act continuously on systolic BP and reduction in
blood Pb may contribute to reductions in BP, while cumulative Pb exposure (represented
November 2012
Draft - Do Not Cite or Quote
-------
1 by bone Pb in this study) may contribute to hypertension incidence by different
2 mechanisms over longer time periods and in older subjects. This analysis in relatively
3 young subjects (mean [SD] age at baseline 41.4 [9.5] years) with a low prevalence of
4 hypertension suggests that at least one of the biological pathways that influences how
5 systolic BP responds to Pb operates over a relatively rapid timeframe. This may reflect an
6 immediate response to Pb at a biochemical site of action as a consequence of the
7 biologically available Pb circulating in blood. A persistent effect of cumulative doses
8 over a lifetime may occur via other mechanisms. Bone Pb level may exert influence on
9 blood Pb levels and consequently on BP in an aging population with prolonged Pb
10 exposure. Thus, the findings contribute important information regarding the various short
11 and long-term exposure relationships with increases in BP and hypertension. It is
12 important to acknowledge the uncertainty regarding the applicability of these findings
13 regarding short-term and long-term effects in Pb workers with relatively high current Pb
14 exposures contributing to blood Pb levels (mean blood Pb levels over time: 20-37 (ig/dL)
15 to adults in the U.S. general population whose concurrent blood Pb levels are influenced
16 more by Pb mobilized from bone stores. Further, for bone Pb analysis, the potential for
17 bone Pb BP and hypertension findings in older populations to be impacted by residual
18 confounding by age may be a factor to consider since exposure studies of older cohorts
19 (NAS/ mean age >60 years; (Wilker et al.. 2011; Kimetal. 1997)) indicate that bone Pb
20 is correlated with age.
21 In a separate cross-sectional analysis of the same occupationally exposed group in year
22 three of follow-up, Weaver et al. (2008) examined associations of concurrent patella Pb
23 and blood Pb level with systolic BP, diastolic BP, and hypertension and effect
24 modification by ALAD and vitamin D receptor (VDR) polymorphisms. None of the Pb
25 biomarkers were associated with diastolic BP. Patella Pb alone was not significantly
26 associated with systolic BP. However, blood Pb, either alone or with patella Pb, was
27 significantly associated with higher systolic BP. The patella Pb-age and blood Pb-age
28 interactions were not statistically significant. There were no significant associations of
29 blood Pb or patella Pb with hypertension status or effect modification by age or sex.
30 Further, interactions between polymorphisms of the VDR and of ALAD with blood Pb
31 and patella Pb on systolic BP were not statistically significant. Mean blood Pb level was
32 high (30.9 (ig/dL) compared to non-occupational groups.
33 Weaver et al. (2010) provided the results of further analysis of this Korean worker cohort,
34 with a focus on determining the functional form of the concentration-response
35 relationships. In a log linear model, the coefficient indicated that every doubling of blood
36 Pb level was associated with a systolic BP increase of 1.76 mmHg. The J test, a statistical
37 test for determining which, if either, of two functional forms of the same variable
38 provides a superior fit to data in non-nested models (Davidson and MacKinnon. 1981).
November 2012 5-305 Draft - Do Not Cite or Quote
-------
1 returned a p-value of 0.013 in favor of the natural log blood Pb level over the linear blood
2 Pb level specification. This analysis indicates that the systolic BP increase in this cohort
3 is better described as a logarithmic function of blood Pb level within the range of the
4 study than by a linear function.
5 Several analyses in the NAS cohort of predominantly white older men in the greater
6 Boston area found associations of blood and bone Pb level with BP and hypertension, and
7 they indicated effect modification by calcium intake, perceived stress, and HFE gene
8 variants. In a cross-sectional analysis, Perlstein et al. (2007) found a statistically
9 significant association between blood Pb and diastolic BP in adjusted models. The
10 subjects in this study had at least one bone Pb measurement during the years 1991-1997
11 and were not on antihypertensive medication at the time of the measurement. While tibia
12 Pb was not significantly associated with BP, it was associated with pulse pressure (PP).
13 Men with tibia Pb above the median (19 (ig/g) had a higher mean PP (4.2 mmHg [95%
14 CI: 1.9, 6.5]) compared to men with tibia Pb below the median. The trend toward
15 increasing PP with increasing quintile of tibia Pb was statistically significant although
16 none of the confidence intervals for PP referenced to the lowest quintile of tibia Pb
17 (<7.4 (ig/g) excluded the null value.
18 Peters et al. (2007) examined cross-sectionally the modification of the associations of
19 tibia and patella Pb with BP and hypertension by self-reported stress (assessed by
20 questionnaire) in NAS men. High stress also has been linked with higher BP, potentially
21 via activation of sympathetic pathways, ROS, and the HPA axis. Among all subjects,
22 higher bone Pb level was associated (statistically nonsignificant) with greater odds of
23 hypertension status and higher systolic BP. As indicated in Figure 5-20. the association
24 between systolic BP and tibia Pb differed between those with high and low self-reported
25 stress ((3 for tibia Pb x stress interaction = 3.77 [95% CI: 0.46, 7.09]) per SD increase in
26 tibia Pb. Stress also was found to modify the patella Pb-BP association ((3 for patella Pb x
27 stress interaction = 2.60 [95% CI: -0.95, 6.15] per SD increase in patella Pb). Neither
28 bone, self-reported stress, nor their interaction was associated significantly with diastolic
29 BP. Peters et al. (2007) also used Cox proportional hazards models to assess the
30 interaction of stress and bone Pb level in the development of hypertension among those
31 free of hypertension at baseline. The results of this analysis showed that increasing tibia
32 and patella Pb were associated with greater risk of developing hypertension among those
33 with high stress compared to those with lower perceived stress (RR of developing
34 hypertension among those with high stress: 2.66 [95% CI: 1.43, 4.95] per SD increase in
35 tibia Pb and 2.64 [95% CI: 1.42, 4.92] per SD increase in patella Pb). These results
36 provide evidence supporting adults with higher stress as a population at increased risk of
37 Pb-associated cardiovascular effects. Earlier, Cheng et al. (2001) examined the NAS
38 cohort in 474 subjects without hypertension (mean [SD] blood Pb level: 5.87 [4.01]) at
November 2012 5-306 Draft - Do Not Cite or Quote
-------
1
2
3
baseline measurement and analyzed linear models with patella Pb and reported that only
patella Pb level was associated with a significant increase in the rate ratio for
hypertension using a Cox's proportional hazards model.
140-
135-
"5> 130-
E
£ 120-
115-
110
-1
-
<% * ?'
£ilftfr**9~~r~~'
^'^ ' ^^ • • O High perceived stress
9 • Low perceived stress
Trend (high stress)
• Trend (low stress)
0 0 10 30 50 70 90
4
5
6
7
8
9
10
11
12
13
14
15
16
17
Tibia lead (ug/g)
Source: Peters et al. (2007)
Figure 5-20 The relationship between tibia Pb and estimated systolic BP
(SBP) for those with high self-reported stress versus those with
low self-reported stress.
Elmarsafawy et al. (2006) examined the modification of the relationship between Pb and
hypertension by dietary calcium, with 467 subjects from the NAS. Responses on a semi-
quantitative dietary frequency questionnaire with one-year recall were used to estimate
calcium intake. Effect modification by calcium intake (dichotomized at 800 mg/day) was
examined using interaction terms in logistic regression models and by conducting
analyses stratified on the calcium variable. Increasing bone and blood Pb increased the
odds of hypertension, particularly among subjects with low dietary calcium.
Zhang et al. (2010a) examined the effect of polymorphisms of the hemochromatosis gene
(HFE) on the relationship of bone Pb with PP in NAS men. HFE polymorphisms promote
Fe absorption and have been shown to modify the impact of adult cardiac function.
Subjects had up to three PP measurements during the 10 year study period. The overall
results demonstrated a strong relationship between bone Pb and PP in this study, similar
to an earlier cross-sectional PP study of many of the same subjects (Perlstein et al.. 2007).
Zhang et al. (2010a) extended these findings by demonstrating larger increases in PP per
November 2012
5-307
Draft - Do Not Cite or Quote
-------
1 unit increase in tibia and patella Pb level among those with the H63D variant compared
2 to those with the wild-type or the C282Y variant.
3 A small number of cross-sectional studies examined and found that blood Pb level was
4 associated with hypertension in pregnancy. Yazbeck et al. (2009) examined a
5 community-based group of pregnant women in France and unlike most other studies,
6 adjusted for potential confounding by blood concentrations of Cd, Mn, and Se. Pregnancy
7 induced hypertension (PIH) was defined as systolic BP >140 mmHg and/or diastolic BP
8 >90 mmHg during at least two clinic visits after week 22 of gestation. Patients with pre-
9 existing chronic hypertension were excluded. The mean (SD) blood Pb levels measured
10 during pregnancy were 2.2 (1.4 (ig/dL) in PIH cases and 1.9 (1.2) (ig/dL in normotensive
11 women. An association between blood Pb and PIH was observed (OR 3.29 [95% CI:
12 1.11, 9.74] per unit increase in log-transformed blood Pb level). Cd and Se concentrations
13 were comparable between PIH and no PIH groups. Adjustment for the metals slightly
14 attenuated but did not eliminate the association between blood Pb levels and the risk of
15 PIH. Investigators observed no significant interactions among blood Pb level, any of the
16 other elements, and maternal characteristics in predicting the risk of PIH. Interaction
17 between blood Se and Pb concentrations was not significant, and the putative protection
18 effects of Se through antioxidative properties were not found in this study.
19 Wells et al. (20 lib) measured the relationship of cord blood Pb with BP in 285 women at
20 admission to the Johns Hopkins Hospital in Baltimore, MD, during labor and delivery.
21 Women with cord blood Pb levels in the highest quartile for the study group
22 (>0.96 (ig/dL) had significantly higher systolic and diastolic BP (upon admission and for
23 maximum BP) compared to women in the first quartile (<0.46 (ig/dL). The level of
24 uncertainty at these levels of exposure is difficult to estimate. The authors used
25 Benchmark Dose Software V2.1, developed by the EPA, to estimate the blood Pb level
26 (benchmark dose or BMD) and the associated lower confidence limit (BMDL) that was
27 associated with one standard deviation (SD) increase in BP. In this study group, one SD
28 is approximately equivalent to a 10% increase above the mean for the first quartile blood
29 Pb reference group. The BMD approach was used only as a means of quantifying the
30 relationship of blood Pb with BP in this population. This analysis indicated that the 95%
31 lower bound confidence limit on the maternal blood Pb level (estimated from cord blood
32 Pb levels) that was associated with a 1 SD increase in all blood pressure outcomes was
33 about 1.4 ug/dL. These reported results are similar to those reported in the
34 2006 Pb AQCD as well as those found 25 years ago but with blood Pb levels an order of
35 magnitude lower in the more recent study. However, uncertainty exists as to the specific
36 Pb exposure level, timing, frequency, and duration that contributed to the observed
37 associations.
November 2012 5-308 Draft - Do Not Cite or Quote
-------
1 Recent analyses using NHANES data continued to indicate associations of Pb biomarkers
2 with BP and hypertension. Muntner et al. (2005) previously used the NHANES
3 1999-2002 data to indicate that concurrent blood Pb levels were associated with
4 hypertension, peripheral artery disease (PAD), and chronic kidney disease. The PAD
5 results are discussed later in Section 5.4.3.5. and chronic kidney disease results are
6 discussed in Section 5.5.2. Blood Pb increased regularly with age (geometric means [95%
7 CIs]: 1.28 (ig/dL [1.23, 1.33] in the 18-39 age group to 2.32 (ig/dL [2.20, 2.44] in the 75
8 and older age group). Associations were observed between concurrent blood Pb level and
9 hypertension across race/ethnicity groups with significant trends observed for
10 non-Hispanic blacks and Mexican Americans.
11 In the NHANES III 1988-1994 population, Scinicariello et al. (2010) found a gene-
12 environment interaction between blood Pb level and ALAD genotype (the genotypes
13 have different affinities for Pb) in relation to systolic BP and diastolic BP in a cross-
14 sectional analysis. These interactions varied across race/ethnicity strata. The strongest
15 associations were observed among non-Hispanic blacks (Figure 5-18. Table 5-18). A
16 statistically significant interaction was observed between concurrent blood Pb level and
17 ALADl-2/2-2b among non-Hispanic whites and non-Hispanic blacks. Scinicariello et al.
18 (2010) also found an interaction between ALAD genotype and blood Pb level in the
19 association with hypertension. Statistically significant associations between concurrent
20 blood Pb level and hypertension were observed among non-Hispanic blacks and
21 nonsignificant increases were observed among non-Hispanic whites and Mexican
22 Americans (with the exception of Mexican Americans in the second quartile of blood Pb
23 level) (Figure 5-19. Table 5-19). In addition, non-Hispanic white ALAD2 carriers in the
24 highest blood Pb level quartile 3.8-52.9 (ig/dL) had a significantly higher association
25 with hypertension compared with ALAD 1 homozygous individuals in the highest quartile
26 of blood Pb. In the same NHANES population, Park et al. (2009c) predicted bone Pb
27 levels using a model developed with NAS data. Concurrent blood Pb was associated with
28 hypertension overall in the NHANES population, with larger associations observed
29 among black men and women as well as older adults (Figure 5-19. Table 5-19).
30 Associations also were observed with estimated bone Pb.
5.4.2.2 Toxicology
31 Studies on the effect of Pb (as blood Pb level) on systolic BP in unanesthetized adult rats
32 consistently reported an increase in BP with increasing blood Pb level as shown in Figure
33 5-21 (results summarized in Table 5-20). An array of studies has provided evidence that
34 long-term Pb exposure (>4 weeks), resulting in blood Pb levels relevant to humans,
35 i.e., below 10 (ig/dL can result in the onset of hypertension (after a latency period) in
November 2012 5-309 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
experimental animals that persists long after the cessation of Pb exposure (U.S. EPA.
2006b). Tsao et al. (2000) presented evidence for increased systolic and diastolic BP in
rats with blood Pb levels similar to the current U.S. population (mean [SD]: 2.15
[0.92] Lig/dL blood Pb; 140 [7] mmHg systolic BP, 98 [7] mmHg diastolic BP) compared
to untreated controls (mean [SD]: 0.05 [0.05] Lig/dL blood Pb; 127 [7] mmHg systolic
BP, 88 [7] mmHg diastolic BP). As this was the lowest Pb level tested, no evidence of a
threshold was evident. Further, a test for linear trend revealed a statistically significant,
positive trend for increasing BP with increasing blood Pb levels up to 56 Lig/dL
(e.g., mean [SD]: 5.47 [2.1] Lig/dL blood Pb; 143 [6] mmHg systolic BP, 97 [8] mmHg
diastolic BP), with the effect leveling off at higher blood Pb levels.
120
15 20 25
Blood Pb Level (ng/dL)
-Bravoetal. 2007
-Ri Hi eta I. 2009
-Changetal. 1997 Changetal. 2005
Rizzietal. 2009 Tsao etal. 2000
-Heydarietal. 2006 Nakhouletal. 1992
-Zhangetal. 2009 ^—Fiorimet al. 2011
Note: Crosses represent standard error for blood Pb and BP measurements. If no crossbar is present, error results were not
reported. Arrows represent higher doses tested.
Figure 5-21 Changes in BP after Pb exposure (represented as blood Pb level)
in unanesthetized adult rats across studies.
November 2012
5-310
Draft - Do Not Cite or Quote
-------
Table 5-20 Characteristics of studies of blood Pb with BP measures in animals
presented in Figure 5-21.
Reference3
Fiorim et al.
(2011)
Nakhoul et al.
(1992)
Chang et al.
(2005)
Tsao et al.
(2000)
Rizzi et al.
(2009)
Chang et al.
(1997)
Heydari et al.
(2006)
Bravo et al.
(2007)
Zhang et al.
(2009a)
Lifestage; Exposure Exposure Level;
Sex Duration Route
Adult; M 7 days 4 ug/1 00 g followed
by 0.05 ug/1 00 g
daily; intramuscular
Adult; M 8 weeks 100 ppm;
drinking water
Adult; M 8 weeks 20,000 ppm then
removal and
measurements
1-7 mo after; drinking
water
Adult 8 weeks 100 - 20,000 ppm;
drinking water
Adult; M 8 weeks 30 -90 ppm;
drinking water
Adult; M 8 weeks 500 ppm; drinking
water
Adult; M 12 weeks 100 ppm; drinking
water
Adult; M 14 weeks 100 ppm; drinking
water
Adult; M 40 weeks 100 ppm; drinking
water
Mean
[SEM]b
Blood Pb
Level
(ug/dL)
9.98 [1 .7]
5.3 [3]
Range:
4.5 to 83
Range of
means: 2.15
[0.29] to
85.76 [1 .29]b
7.6 [1.3],
19.3 [3.4]
29.1 [0.6]b
26.8 [2.2]
23.7[1.9]b
28.4[1.1]b
ASBP
(mmHg;
lowest blood
Pb level
compared
n with control)"
12 16
7 28
5 13.8
10 13
11 13.3
10 58
6 25.8
12 30
8-10 15.3
Comments
Spontaneously
hypertensive
rat model
aStudies are presented in order of increasing duration of exposure.
""Standard deviation converted to SEM.
°Difference in systolic BP (SBP) between group means not within one exposure group.
1
2
o
3
4
5
6
7
8
9
10
11
12
13
14
Experimental animal studies continued to provide evidence that long-term Pb exposure
results in sustained arterial hypertension after a latency period. Systolic BP increased in
rats after exposure to 90-10,000 ppm Pb (as Pb acetate in drinking water) for various time
periods that resulted in blood Pb levels between 19.3-240 (ig/dL (Mohammad et al..
2010; Zhang et al.. 2009a: Badavi et al.. 2008; Grizzo and Cordellini. 2008; Rezaetal..
2008: Bravo et al.. 2007: Vargas-Robles et al.. 2007: Hevdari et al.. 2006: Bagchi and
Preuss. 2005). Past studies have shown statistically significant elevations in BP in rats
with lower blood Pb levels. For example, long-term Pb exposure to spontaneously
hypertensive rats (resulting in mean [SEM] blood Pb level: 5.3 [3] (ig/dL) led to
increased BP (Nakhoul et al.. 1992). Consistent with measurements of systolic BP by tail-
cuff plethysmography, Pb exposure (100 ppm for 14 weeks; mean blood Pb level:
24 (ig/dL) also caused an increase in intra-aortic mean arterial pressure (Bravo et al..
2007). In a study that tested low levels of Pb exposure (30 ppm; mean blood Pb level:
7.6 (ig/dL), a statistically significant increase in systolic BP was not observed despite
November 2012
5-311
Draft - Do Not Cite or Quote
-------
1 elevated blood Pb level after 8 weeks of treatment. Nonetheless, there was a trend of
2 higher BP with higher blood Pb levels (Rizzi et al. 2009V
3 Studies found that Pb-induced increases in BP persisted long after cessation of Pb
4 exposure. Bagchi and Preuss (2005) found that elevated systolic BP was maintained for
5 210 days after cessation of Pb exposure (10,000 ppm Pb acetate in water, 40 days,
6 monitored for one year). However, chelation therapy using Na2CaEDTA returned systolic
7 BP to levels comparable to those in rats not treated with Pb (Bagchi and Preuss. 2005).
8 Chang et al. (2005) reported a partial reversibility of effect after cessation of Pb exposure,
9 where Pb-induced elevated BP decreased but did not return to control levels 7 months
10 post Pb exposure. After Pb exposure was removed, blood, heart, aorta, and kidney Pb
11 levels decreased quickly within the first three months (Chang et al.. 2005). Pb-induced
12 elevated systolic BP persisted for one month following Pb exposure cessation, followed
13 by obvious decreases in BP until 4 months after Pb exposure cessation. Between 4 and
14 7 months after Pb exposure cessation, the still-elevated BP did not decrease further, thus
15 never returning to control BP levels. Decreases in BP were strongly correlated with
16 decreases in blood Pb level after exposure cessation.
17 The aforementioned studies all assessed the relationship between long-term exposure
18 (>4 weeks) of rats to Pb and measures of BP. However, recent research also investigated
19 BP elevation occurring after short-term treatment with Pb (<4 weeks). Studies found
20 increased systolic BP after 7 days of Pb treatment (daily injections resulting in mean
21 [SEM] blood Pb levels of 9.98 [1.7] (ig/dL) (Fiorim et al.. 2011) and after 2 weeks of Pb
22 exposure (100 ppm via drinking water) (Sharifi et al.. 2004). A study utilizing intra-
23 arterial pressure measurements found that a single high-dose Pb injection in rats
24 (resulting in mean [SEM] blood Pb levels of 37 [1.7] (ig/dL) increased systolic arterial
25 pressure after only 60 minutes (Simdes et al.. 2011). The injection of Pb into the rat may
26 not allow for extrapolation of these results to humans since this is not a comparable Pb
27 exposure method. These studies suggest that there is the potential for increase in BP
28 following short-term Pb treatment. It is possible that the increases in BP following short-
29 and long-term Pb exposures are occurring through separate mechanisms; however,
30 studies using both short- and longer-term Pb exposure have correlated increased BP with
31 an activation of the renin-angiotensin system (i.e., increase in angiotensin converting
32 enzyme (ACE) activity) (Section 5.4.2.3). Several of these aforementioned studies used
33 the injection route of Pb administration, and the relevance of these bolus doses over short
34 periods of time to human routes of short-term exposure is uncertain. However, it is
35 important to acknowledge that the results were similar to those from the study that
36 examined short-term exposure to Pb via drinking water,
November 2012 5-312 Draft - Do Not Cite or Quote
-------
5.4.2.3 Hypertension Modes of Action
1 The 2006 Pb AQCD (U.S. EPA. 2006b) examined a number of mechanisms leading to
2 Pb-induced hypertension, including oxidative stress, hormonal and blood pressure
3 regulatory system dysfunction, vasomodulation, and cellular alterations. As described
4 below, recent studies in experimental animals and cells further supported roles for these
5 potential mechanisms in mediating hypertension from Pb exposure.
Oxidative Stress Response - Reactive Oxygen Species and Nitric Oxide
6 Several studies discussed in the 2006 Pb AQCD demonstrated a role for oxidative stress
7 in the pathogenesis of Pb-induced hypertension, mediated by the inactivation of nitric
8 oxide (NO) and downregulation of soluble guanylate cyclase (sGC) (Dursun et al.. 2005;
9 Attri etal.. 2003; Gonicketal.. 1997; Vaziri etal.. 1997; Khalil-Manesh et al.. 1994;
10 Khalil-Manesh et al.. 1993b). Pb-induced reduction of biologically active NO was found
11 not to be due to a reduction in NO-production capacity (Vaziri and Ding. 2001; Vaziri et
12 al.. 1999a); instead it was found to result from inactivation and sequestration of NO by
13 ROS (Malvezzi et al.. 2001; Vaziri etal.. 1999b). Oxidative stress from Pb exposure in
14 animals may be due to upregulation of NAD(P)H oxidase (Ni et al.. 2004; Vaziri et al..
15 2003). induction of Fenton and Haber-Weiss reactions (Ding etal.. 2001; Ding et al..
16 2000). and failure of the antioxidant enzymes, CAT and GPx, to compensate for the
17 increased ROS (Farmand et al.. 2005; Vaziri et al.. 2003). Many biological actions of
18 NO, such as vasorelaxation, are mediated by cGMP, which is produced by sGC from the
19 substrate GTP. Oxidative stress also has been found to play a role in Pb-induced
20 downregulation of sGC (Farmand et al.. 2005; Courtois et al.. 2003; Marques et al..
21 2001). Thus, the reduction of the vasodilator NO from inactivation and sequestration by
22 Pb-induced ROS leads to increased vasoconstriction and BP.
23 Pb-induced oxidative stress also has been found to induce renal tubulointerstitial
24 inflammation which plays a crucial role in models of hypertension (Rodriguez-Iturbe et
25 al.. 2005; Rodriguez-Iturbe et al.. 2004). Tubulointerstitial inflammation from treatment
26 with Pb has been coupled with activation of the redox sensitive NF-KB (Ramesh et al..
27 2001). Pb-induced hypertension, inflammation, and NF-KB activation can be ameliorated
28 by antioxidant therapy (Rodriguez-Iturbe et al.. 2004). There is mixed evidence to
29 suggest that Pb-induced hypertension may also be promoted by activation of PKC leading
30 to enhanced vascular contractility (Valencia et al.. 2001; Watts etal.. 1995).
31 Recent studies continued to provide evidence for the role of ROS and NO metabolism in
32 Pb-induced hypertension and vascular disease. Increased systolic BP after Pb exposure
33 was accompanied by increased superoxide (O2~) and O2~ positive cells (Bravo et al.. 2007;
November 2012 5-313 Draft - Do Not Cite or Quote
-------
1 Vargas-Robles et al.. 2007). elevated urinary malondialdehyde (MDA, a measure of lipid
2 peroxidation) (Bravo et al.. 2007). and increased 3-nitrotyrosine (Vargas-Robles et al..
3 2007). Inhibition of NAD(P)H oxidase, an enzyme that generates O2 and hydrogen
4 peroxide, was able to block Pb-induced (1 ppm) aortic contraction
5 to 5-hydroxytryptamine (5-HT) (Zhang et al.. 2005). Increases in systolic BP, intra-aortic
6 mean arterial pressure, and MDA after Pb exposure (100 ppm; mean blood Pb level:
7 23.7 (ig/dL) were also prevented by treatment with the immunosuppressant,
8 mycophenolate mofetil (MMF) (mean blood Pb level in MMF-treated animals: 27 (ig/dL)
9 (Bravo et al.. 2007). MMF has been shown to inhibit endothelial NAD(P)H oxidase,
10 which could explain how it decreases Pb-induced increases in oxidative stress and BP.
11 MMF was not found to alter blood Pb levels of animals. Red grape seed extract and
12 ascorbic acid supplementation were also able to protect rats from Pb-induced (100 ppm)
13 increased BP and heart rate, perhaps through the antioxidant properties of the extract
14 (Badavi et al.. 2008) and vitamin C (Mohammad et al.. 2010). Red grape seed extract did
15 not alter the accumulation of Pb in blood, indicating that its protective effect was not
16 mediated through altered Pb toxicokinetics; however, internal doses of Pb were not
17 measured in the vitamin C study to clarify the mechanism of action of vitamin C. Another
18 study found that the antioxidant, anti-inflammatory chemical, curcumin, as well as
19 physical exercise training reversed Pb-induced increases in serum creatinine kinase-MB
20 (CK-MB), low density lipoprotein (LDL), heart high-sensitivity C-reactive protein
21 (hs-CRP), and MDA. Pb-induced decreases in serum total antioxidant capacity, high
22 density lipoprotein (HDL), and heart glutathione peroxidase (GPx) were also reversed by
23 curcumin and exercise. However, internal doses of Pb were not measured to clarify the
24 mechanism of action in this study (Roshan et al.. 2011).
25 Exposure to Pb can also affect the activity and levels of antioxidant enzymes. Male ($)
26 and female (9) rats exposed to Pb for 18 weeks (100-1,000 ppm) had altered responses in
27 antioxidant enzymes in heart tissue (Sobekova et al.. 2009; Alghazal et al.. 2008a). Pb
28 exposure in female rats increased the activity of cardiac SOD, GST, GR, and GPx
29 (>100 ppm) and increased cardiac thiobarbituric acid reactive substances (TEARS, a
30 measure of lipid peroxidation) (1,000 ppm). Pb exposure in male rats did not affect the
31 activity of SOD or production of TEARS, however decreased the activity of GST and GR
32 (>100 ppm). Male and female rats also accumulated different amounts of Pb in the
33 cardiac tissue after similar Pb exposure (<$ 100 ppm: 205% of control, 1,000 ppm: 379%;
34 9 100 ppm: 246%, 1,000 ppm: 775%), which could explain the sex differences observed
35 in antioxidant enzyme responses.
36 Oxidative stress can trigger a cascade of events that promote cellular stress, renal
37 inflammation, and hypertension. As was shown previously (Rodriguez-Iturbe et al..
38 2005). Pb exposure can increase renal NF-KB, which was associated with
November 2012 5-314 Draft - Do Not Cite or Quote
-------
1 tubulointerstitial damage and infiltration of lymphocytes and macrophages (Bravo et al.,
2 2007). These events could also be ablated by MMF treatment, likely due to its anti-
3 inflammatory and antioxidant properties. Pb also was found to induce inflammation in
4 human endothelial cells as a model for vessel intima hyperplasia (Zeller et al.. 2010). The
5 pro-inflammatory cytokine, interleukin (IL)-8 protein and mRNA were increased,
6 concentration- and time-dependently, after in vitro Pb exposure (5-50 (JVI). Enhanced
7 IL-8 production was mediated through activation of the transcription factor Nrf2 (but not
8 NF-KB, hypoxia inducible factor-1, or aryl hydrocarbon receptor), as shown through
9 increased nuclear translocation and Nrf2 cellular knockdown experiments. Additionally,
10 measures of endothelial stress, NQO1 and HO-1 protein, were induced by Pb exposure
11 (Zeller et al.. 2010). Pb treatment (20 ppm, i.p., 3 days/week, 8 weeks) increased the
12 inflammatory markers hs-CRP and CK-MB in rat hearts (Roshan etal.. 2011).
13 Oxidative stress affects vascular reactivity and tone through inactivation and
14 sequestration of NO, causing a reduction in biologically active NO. Recent studies
15 affirmed past conclusions on the interplay of ROS and NO metabolism in the
16 cardiovascular effects of Pb. Elevated systolic BP and altered vasorelaxation after Pb
17 exposure was accompanied by a decrease in total nitrates and nitrites (NOX) (Mohammad
18 etal.. 2010: Zhang et al.. 2007a: Hevdari et al.. 2006). Serum NOX levels in Pb-treated
19 rats remained depressed for 8 weeks and then reversed after 12 weeks, despite continued
20 elevation in systolic BP (Hevdari et al.. 2006). This return of serum NOX levels to levels
21 similar in controls could be a result of compensatory increases in endothelial NOS
22 (eNOS) attempting to replenish an over-sequestered NO supply. With this in mind,
23 studies showed increased eNOS protein expression after long-term Pb exposure in kidney
24 (Zhang et al.. 2007a) and isolated cultured aorta (Vargas-Robles et al.. 2007). No change
25 in inducible NOS was observed in isolated cultured aorta after 1 ppm Pb exposure (Zhang
26 et al.. 2007a). In contrast to long-term exposure, Pb treatment over a short time period
27 (daily injections resulting in mean [SEM] blood Pb levels of 9.98 [1.7] (ig/dL) was found
28 to increase iNOS and phosphorylated eNOS protein (Fiorimet al.. 2011) which may
29 cause an increase in NO production and a short-term increase in NO bioavailability. This
30 increase in NO bioavailability early after Pb exposure could be the immediate
31 compensatory mechanism against the elevation in BP.
32 NO, also known as endothelium-derived relaxing factor, is a potent endogenous
33 vasodilator. Toxicological studies continued to investigate the effects of Pb on
34 NO-dependent vascular reactivity by using NO stimulating vasodilators, such as
35 acetylcholine (ACh) and sodium nitroprusside (SNP), and NO inhibiting
36 vasoconstrictors, such as L-NAME. Studies provided mixed evidence; however, results
37 suggested that Pb disrupts the vasorelaxant response to NO in the aorta due to damage to
38 the endothelium. Pb exposure (1 ppm and 100 (iM, 1 hour) decreased ACh-induced
November 2012 5-315 Draft - Do Not Cite or Quote
-------
1 vasorelaxation, which triggers the release of NO from the endothelial cell, in isolated rat
2 tail artery, suggesting damage to the endothelium (Silveira et al.. 2010; Zhang et al..
3 2007a). In aortic rings of perinatally exposed rats (1,000 ppm through pregnancy and
4 lactation, mean blood Pb level: 58.7 (ig/dL), blocking NOS with L-NAME abolished the
5 relaxant response evoked by ACh (Grizzo and Cordellini. 2008). However, there was no
6 change observed in the relaxation response to ACh by Pb alone (Fiorim et al.. 2011; Rizzi
7 et al., 2009; Grizzo and Cordellini, 2008). Conversely, Skoczynska and Stojek (2005)
8 found that Pb exposure (50 ppm; blood Pb level 11.2 (ig/dL) enhanced NO-mediated
9 vasodilation by ACh in rat mesenteric arteries, and NOS inhibition enhanced the ACh
10 relaxant response. A number of studies found that Pb exposure did not affect smooth
11 muscle integrity since SNP-induced vasorelaxation, which is endothelium independent,
12 was unchanged (Fiorim et al., 2011; Silveira et al.. 2010; Rizzi et al.. 2009; Grizzo and
13 Cordellini. 2008).
14 NO also was found to play a role in the interaction between Pb and the vasoconstrictor
15 response. Blocking NOS with L-NAME or inhibiting iNOS specifically, which decreases
16 NO production, increased the contraction of aortic rings in response to the
17 vasoconstrictor phenylephrine (PHE), and Pb exposure potentiated this response (Fiorim
18 et al.. 2011). Also, L-NAME increased the Pb pressor response to PHE after perinatal Pb
19 exposure (1,000 ppm through pregnancy and lactation, blood Pb level 58.7 (ig/dL)
20 (Grizzo and Cordellini. 2008). Conversely, in rat renal interlobar arteries, Pb exposure
21 blunted the increase in renal angiotensin II (Angll)-mediated contraction from NOS
22 inhibition by L-NAME (Vargas-Robles et al.. 2007). Treatment with the SOD mimetic
23 tempol, which would increase NO bioavailability, decreased, but did not eliminate, the Pb
24 pressor response (Silveira et al.. 2010).
25 In summary, recent studies continued to provide evidence for the role of ROS in
26 Pb-induced hypertension and vascular disease by indicating Pb-induced increases in ROS
27 and modulation of cardiovascular responses by antioxidant substances. Additionally,
28 recent studies continued to show that Pb-induced hypertension and vascular responses are
29 mediated primarily via inactivation of NO not via inhibition of NO production.
Vascular Reactivity
30 Alteration of the adrenergic system from Pb exposure, which can increase peripheral
31 vascular resistance, and thereby arterial pressure, may be one mediator of Pb-induced
32 hypertension. Pb exposure in animals can increase stimulation of the sympathetic nervous
33 system (SNS), as shown by increased plasma levels of norepinephrine (NE) and other
34 catecholamines (Carmignani et al.. 2000; Chang et al.. 1997) and decreased (3 adrenergic
35 receptor density and (3 agonist-stimulated cAMP production in the aorta and heart (Tsao
November 2012 5-316 Draft - Do Not Cite or Quote
-------
1 et al.. 2000; Chang etal., 1997). These stimulatory effects on the SNS paralleled the
2 effects of Pb on BP, cardiac contractility, and carotid blood flow. Pb-induced elevations
3 in arterial pressure and heart rate were abrogated by ganglionic blockade (Simdes et al.,
4 2011; Lai et al.. 2002). Arterial pressure and heart rate gradually decreased 7 months
5 after Pb exposure cessation as did the Pb-induced SNS alterations (Chang et al., 2005).
6 Increases in BP can be caused by activation of the SNS, which can lead to vascular
7 narrowing, in turn, resulting in increased total peripheral resistance. In this neural
8 mechanism, activation of the SNS leads to vasoconstriction, whereas inhibition leads to
9 vasodilation. It has been suggested that Pb leads to increased vascular reactivity to
10 catecholamines (i.e., epinephrine, NE, and dopamine), hormones of the SNS. Indeed, the
11 isolated mesenteric vessel bed from Pb-treated rats (50 ppm with blood Pb level:
12 11.2 (ig/dL, but not 100 ppm with blood Pb level: 17.3 (ig/dL) exhibited increased
13 reactivity to NE (Skoczynska and Stojek. 2005). However, in another study, 100 ppm Pb
14 did not affect the NE-induced contractile response after 10 months of exposure (Zhang et
15 al., 2009a). suggesting a small range of Pb doses affects pressor response to NE.
16 Catecholamines act primarily through the adrenergic and dopaminergic receptors.
17 Antagonists of a 1-adrenergic, a2-adrenergic, (3-adrenergic, and dopamine Dl receptors
18 were found to abolish Pb-induced aortic contraction (Fazli-Tabaei et al.. 2006; Heydari et
19 al., 2006). However, the a 1-adrenergic receptor agonist, PHE, induced aortic contractions
20 and these were enhanced by treatment with Pb (100 ppm; blood Pb level: 26.8 (ig/dL),
21 indicating a specific role for the a 1-adrenergic receptor (Silveira et al., 2010; Grizzo and
22 Cordellini. 2008; Heydari et al.. 2006). Removal of the endothelium blunted the PHE-
23 induced contraction. Conversely, short-term Pb treatment (7 days, i.p.) decreased the
24 contractile response induced by PHE in rat aortas resulting in a decreased vascular
25 reactivity (Fiorim etal.. 2011). This decrease may be playing a compensatory role in
26 attempting to correct the Pb-induced BP elevation. Additionally, Pb blunted the
27 isoproterenol-induced relaxation, supporting a role for the (3-adrenoceptors (Vassallo et
28 al.. 2008; Hevdari et al.. 2006).
29 Recently, there was mixed evidence for Pb disrupting vascular reactivity to other pressor
30 agents. Pb (1 ppm) treatment of isolated rat thoracic aorta increased 5-HT induced
31 contraction, which was endothelium dependent, but not due to 5-HT2B receptor
32 expression (Zhang et al., 2005). Follow-up of this study in whole animals found, on the
33 contrary, that Pb (100 ppm; blood Pb level: 28.4 (ig/dL) decreased the maximum
34 contractile response to 5-HT, but did not affect 5-HT plasma levels or 5-HT2B receptor
35 expression (Zhang et al.. 2009a). In addition, Pb exposure (100 ppm, 12 weeks) increased
36 the renal vascular response to Angll in isolated perfused kidneys from Pb-exposed rats
37 (Vargas-Robles et al.. 2007).
November 2012 5-317 Draft - Do Not Cite or Quote
-------
1 Studies continued to investigate the effects of Pb on NO-dependent vascular reactivity by
2 using NO stimulating vasodilators, such as ACh and SNP, and NO inhibiting
3 vasoconstrictors, such as L-NAME. These studies were discussed in the preceding
4 subsection (Oxidative Stress Response).
Renin-Angiotensin-Aldosterone and Kininergic Systems
5 The adrenergic system also affects the renin-angiotensin-aldosterone system (RAAS),
6 which is responsible for fluid homeostasis and BP regulation, and has been shown to be
7 affected by Pb exposure. A meta-analysis found that Pb exposure (resulting in blood Pb
8 levels: 30-40 (ig/dL) increased plasma renin activity and renal tissue renin in young but
9 not old rats fVander. 1988). Exposure of experimental animals to Pb also induced
10 increases in plasma, aorta, heart, and kidney angiotensin converting enzyme (ACE)
11 activity; plasma kininase II, kininase I, and kallikrein activities; and renal Angll positive
12 cells (Rodriguez-Iturbe et al.. 2005; Sharifi et al.. 2004; Carmignani etal.. 1999). ACE
13 activity declined over time while arterial pressure stayed elevated, suggesting that the
14 RAAS may be involved in the induction, but not the maintenance of Pb-induced
15 hypertension in rats.
16 Recent studies continued to implicate the RAAS in the development of Pb-induced
17 hypertension, especially during early exposure in young animals. Angll, a main player in
18 the RAAS, induces arteriolar vasoconstriction leading to increased BP. Pb exposure
19 increased the vascular reactivity to Angll (Vargas-Robles et al.. 2007). Acute
20 (60 minutes) or short-term (7 days) treatment of rats to Pb increased the plasma ACE
21 activity (Fiorim et al.. 2011; Simdes et al.. 2011). and Fiorim et al. (2011) additionally
22 found this increase to be correlated with the Pb-induced increase in systolic BP.
23 However, at these short time points there were no changes in the Angll receptors 1 or 2
24 protein levels or expression. Treatment with the Angll receptor (ATiR) blocker,
25 Losartan, or the ACE inhibitor, Enalapril, blocked the Pb-induced systolic BP increase
26 (Simdes et al.. 2011) and decreased the PHE-induced vasoconstrictor response in
27 Pb-treated aortas (Fiorim et al.. 2011). Similarly, treatment with Losartan resulted in a
28 greater decrease in systolic BP in highly Pb-exposed rats (10,000 ppm Pb, 40 days; blood
29 Pb level >240 (ig/dL after exposure, 12-13 (ig/dL after chelation after 1 year) compared
30 to control rats that continued into later periods of follow-up (day 283) (Bagchi and
31 Preuss. 2005). Increased systolic BP after early exposure to Pb corresponded with
32 increased water intake, urine output, potassium excretion, and decreased urinary sodium
33 and urine osmolality. These functional changes in renal behavior are consistent with the
34 actions of a stimulated RAAS. Lower level Pb (100 ppm, 14 weeks; range of blood Pb
35 levels: 23.7-27 (ig/dL) exposure increased renal cortical Angll content and the number of
36 tubulointerstitial Angll-positive cells (Bravo et al.. 2007). This heightened intrarenal
November 2012 5-318 Draft - Do Not Cite or Quote
-------
1 angiotensin corresponded with sodium retention and increased systolic BP and was
2 ablated by the anti-inflammatory antioxidant, MMF. Sodium reabsorption is important
3 for the maintenance of BP, and Na+ transporters play a key role in this process. In other
4 studies, Pb exposure increased activity and levels of the a-1 subunit protein of
5 Na+/K+ATPase, which plays a major role in Na+ reabsorption and is regulated by the
6 RAAS (Fiorim et al.. 2011; Simdes et al.. 2011). These studies point to the activation of
7 the RAAS in the course of Pb-induced hypertension, particularly in the early stages of
8 elevated BP.
Vasomodulators
9 The balance between production of vasodilators and vasoconstrictors is important in the
10 regulation of BP and cardiovascular function. The 2006 Pb AQCD reported that Pb did
11 not affect all vasomodulators in the same way. Urinary excretion of the vasoconstrictor,
12 thromboxane (TXB2), and the vasodilatory prostaglandin, 6-keto-PGFla, was unchanged
13 in rats with Pb-induced hypertension (Gonick et al.. 1998). However, in vitro Pb
14 exposure promoted the release of the prostaglandin precursor, arachidonic acid, in
15 vascular smooth muscle cells (VSMCs) via activation of phospholipase A2 (Dorman and
16 Freeman. 2002). Plasma concentration and urinary excretion of the vasoconstrictive
17 peptide, endothelin (ET) 3 was increased after low (100 ppm), but not high-level
18 (5,000 ppm) Pb exposure in rats (Gonick et al.. 1997; Khalil-Manesh et al.. 1994; Khalil-
19 Manesh et al.. 1993b). Antagonism of the ET receptor A blunted the downregulation of
20 sGC and cGMP production by Pb in isolated rat artery segments, suggesting that some of
21 the hypertensive effects of Pb exposure may be mediated through ET (Courtois et al..
22 2003). Additionally, Pb-exposed animals exhibited fluid retention and a
23 concentration-dependent decline in the vasodilator, atrial natriuretic factor (ANF)
24 (Giridhar and Isom. 1990). Results from these studies suggest that Pb may interfere with
25 the balance between vasodilators and vasoconstrictors that contribute to the complex
26 hormonal regulation of vascular contraction and BP.
27 The imbalance in vasomodulators is one explanation for the concentration-dependent
28 vasoconstriction observed in some animals after Pb exposure (Valencia et al.. 2001;
29 Watts et al.. 1995; Piccinini et al.. 1977). However, vasoconstriction after Pb exposure
30 was not reported in all studies (Shelkovnikov and Gonick, 2001) and is likely varied
31 depending on the type of vessel used, the Pb concentration employed, and the animal
32 species being studied. Studies have reported Pb-induced attenuation of ACh- and NO-
33 mediated vasodilation (Marques et al.. 2001; Oishi et al.. 1996) in some, but not all
34 vascular tissues and in some, but not all studies (Purdy et al.. 1997). These effects have
35 been variably attributed to Pb-mediated activation of PKC and direct action on the
November 2012 5-319 Draft - Do Not Cite or Quote
-------
1 VSMCs through the Ca2+ mimetic properties of Pb among other possibilities (Valencia et
2 al..2Q01: Watts etal. 1995: Piccinini et al.. 1977).
3 A recent study investigated the role of the endothelial-derived vasoconstrictor, ET-1, in
4 Pb-induced hypertension. ET-1 from the endothelium acts on the ETA-type receptors
5 located on the vascular smooth muscle layer and may be involved in vascular reactivity
6 by NO and COX derivatives. Pb exposure (1 ppm, 24 hours) to rat aortic segments
7 decreased expression of sGC-(31 subunit, an enzyme involved in NO-induced
8 vasodilation, and increased expression of COX-2 in an endothelium-dependent manner
9 (Molero et al.. 2006). Even though Pb treatment did not alter ET-1 or ETA-type receptor
10 protein expression in this system, blocking the ETA-type receptors partially reversed
11 Pb-induced changes in sGC and COX-2 in vascular tissue. These results suggest that the
12 endothelium and ET-1 may contribute to Pb-induced hypertension through activation of
13 ETA-type receptors that alter expression of COX-2 and sGC-(31 subunit, which affects
14 NO signaling.
15 COX-2 blockade has been shown to prevent Pb-induced downregulation of sGC
16 expression (Courtois et al., 2003). Inhibition of COX-2 also decreased the Pb-induced
17 pressor response to ACh (Grizzo and Cordellini. 2008) and PHE (Silveira et al.. 2010) in
18 experimental animals. These results suggest that Pb-induced vascular reactivity may
19 depend on the participation of a COX-derived vasoconstrictor, such as prostaglandins,
20 prostacyclins, orthromboxanes.
21 In summary, recent studies continued to show that Pb exposure affects vasomodulatory
22 pathways that are important for the maintenance of vascular tone; however, results
23 indicated that not all vascular cell types are similarly affected by Pb exposure. Further,
24 effects appeared to vary according to the concentration of Pb exposure. Pb exposure has
25 been shown to interrupt baseline or endogenous NO-mediated vasodilation of vessels via
26 alterations in PKC, sGC, VSMC, endothelial cells, NADPH oxidase, and Ca2+ levels.
27 Recent studies indicated that Pb exposure may affect vascular reactivity by increasing
28 COX-2 and COX-2-dependent vasoconstrictors. Also, the vasoconstrictor endothelin may
29 contribute to Pb-induced vasomodulation via similar pathways as NO including effects
30 on sGC and COX-2.
5.4.2.4 Summary of Blood Pressure and Hypertension
31 The 2006 Pb AQCD (U.S. EPA. 2006b) reported a clear association between higher
32 blood Pb levels and higher BP. The effect was modest, but robust, as determined by a
33 meta-analysis (TSfawrot et al.. 2002) of over 30 cross-sectional and prospective studies
34 comprising over 58,000 adults (Figure 5-22). In the meta-analysis, each doubling of
November 2012 5-320 Draft - Do Not Cite or Quote
-------
1 concurrent blood Pb was associated with a 1 mmHg increase in systolic BP and a 0.6
2 mmHg increase in diastolic BP. Recent epidemiologic studies supported this association
3 at lower concurrent blood Pb levels (in populations with mean blood Pb levels <2 (ig/dL)
4 and added to the evidence base regarding populations potentially at increased risk
5 (i.e., high stress, genetic variants) and regarding associations of bone Pb levels with BP
6 and hypertension in populations with mean bone Pb levels less than 20 (ig/g. As these
7 studies were mostly cross-sectional in design and were conducted in adults whose
8 concurrent blood Pb levels are influenced both by current Pb exposures and past Pb
9 exposures mobilized from bone, uncertainty exists over the Pb exposure conditions that
10 contributed to the associations observed between concurrent blood Pb level with
11 increased BP and hypertension (Sections 4.3 and 4.7.3). However, U.S. EPA (1990a)
12 reviewed studies available prior to 1990, a period when Pb exposures from air were
13 probably at the highest level, that examined Pb exposure and BP outcomes which
14 included evaluation of several studies of the population represented in NHANES II
15 (1976-80). They noted that across a range of 7 to 34 (ig/dL, no evident threshold was
16 found below which the blood Pb level was not significantly related to BP. U.S. EPA
17 (1990a) concluded that a small but positive association exists between blood Pb levels
18 and increases in BP.
19 A recent prospective study in Pb workers found independent associations of both baseline
20 blood Pb level and subsequent changes in blood Pb over follow-up with changes in BP
21 over follow-up and bone Pb level with hypertension (Glenn et al.. 2006). Although these
22 Pb workers had higher current Pb exposure compared with nonoccupationally-exposed
23 adults, the results indicated that different mechanisms may mediate shorter-term
24 Pb-associated increases in BP and longer-term Pb-associated development of
25 hypertension.
26 Key evidence was further provided by a recent cross-sectional study in an ethnically
27 diverse community-based cohort of women and men aged 50-70 years of age that found
28 associations of both blood and tibia Pb levels with BP with extensive consideration of
29 potential confounding factors (Martin et al.. 2006). Additionally, a recent epidemiologic
30 study provided evidence for associations in an adult cohort between blood Pb level and
31 BP and hypertension with relatively low blood Pb levels; a positive relationship was
32 found in the NHANES adult data (1999-2002) with a geometric mean blood Pb level of
33 1.64 (ig/dL (Muntner et al.. 2005). However, as noted above, in adults, uncertainty exists
34 regarding the magnitude, timing, frequency, and duration of Pb exposure that contribute
35 to the associations observed with concurrent blood Pb levels.
November 2012 5-321 Draft - Do Not Cite or Quote
-------
POCOCK81.
KRCMHOUT 85
ORSSAUD 85
WEISS 86
DE KORT 87
PARKINSON 87 K
RABINOWITZ87
ELWOOD (Cl 88 i-
ELWOOO [HP) 88 H
ELWOOO [HP) 88 *
GARTSIOE IW) 88
GART5IOE [W18B t-
NERI 88 i
GRANDJEAN89 1—
GRANDJEAN89 i —
APOSTOL1 90
MORRIS 90
MORRIS 90
SHARP [BI90
M01LER92
HEN5E 93
HENSE93 i-
MENDITTD9*.
PROCTOR 96 i—
STAESSEN \p) 96 \ »
STAESSEN (P) 96 i
SQf Ac |W) 97 i •
CHU99
CHU 99 I
ROTHENBERG INI] 99 i-
ROTHENBERG [I) 99
SCHWARTZ 00
DEN HOND (W) 01 i-
DEN HOND (B) 01
DEN HOND (W) 01 •
DEN HOND IB) 01
ALL
* o-
* o-
h*H $
H O*
H er
H 9
H^H 9
-• < 9
• 9
*~ 9
• 4 C
I c^
-1-A f
+—* cr
g
J B
• O*
9
MH 9
-•H (f
%
i^^ 0
• o-9
f I 1 1 1 1 1 1 1
-15 -10 -50 5 10 15 20 25 30
SBP
7379
15?
89
105
116
3851
1136
665
B56
350
28Z7
288
W
58
2?r
US
106
in
132
398
133
(39
1703
809
1319
798
339
345
'86
2563
2763
K7]
mi
(.39
lisa
51.3
1,685
1761
5138
Z197
58518
Source: Reprinted with permission of MacMillan Press, Nawrot et al. (2002)
Study Key: C - Caerphilly Study; HP - Welsh Heart Program; W - Whites; B - Blacks; Nl - Non-immigrants; I - Immigrants;
FW- Foundry Workers; CS - Civil Servants; P - PheeCad (Public Health and Environmental Exposure to Cadmium) Study.
Note: Individual study results are presented in each row. The rightmost columns indicate the sex of subjects and study sample size.
Circles represent individual groups and squares represent the combined association sizes. Open circles denote a nonsignificant
association size that was assumed to be zero.
Figure 5-22 Meta-analysis of change in systolic BP (SBP), in mmHg with 95%
Cl, associated with a doubling in the blood Pb concentration.
i
2
3
4
5
6
7
In concordance with epidemiologic evidence, collectively, the animal toxicological
studies providing blood Pb level and BP measurements reported higher BP with higher
blood Pb levels in adult rodents (Figure 5-21). While the contribution of low concurrent
blood Pb levels to the findings is difficult to ascertain in adult humans, animal
toxicological studies provide support for low blood Pb level effects with increases in BP
observed in groups of animals with long-term dietary Pb exposure resulting in blood Pb
levels as low as 2 (ig/dL (Rizzi et al.. 2009: Tsao et al.. 2000: Nakhoul et al.. 1992).
However, the majority of animal toxicological studies showing Pb-induced hypertension
November 2012
5-322
Draft - Do Not Cite or Quote
-------
1 were conducted at higher Pb exposure levels that result in blood Pb levels >10 (ig/dL. In
2 addition, recent animal evidence suggests the potential for increased BP following short-
3 term (4 weeks) Pb treatment that included injected bolus doses that may have uncertain
4 relevance to human routes of Pb exposure (Fiorim et al.. 2011; Simdes et al.. 2011;
5 Sharifi et al., 2004). A recent study also demonstrated partial reversibility (not to levels in
6 controls) of Pb-induced elevations in BP following Pb exposure cessation or chelation
7 (Chang et al.. 2005).
8 Epidemiologic studies continued to investigate the relationship between bone Pb and BP.
9 A recently published meta-analysis (Figure 5-23) (Navas-Acien et al.. 2008) included
10 several studies (three prospective, five cross-sectional) that individually showed that bone
11 Pb level was associated with systolic BP but not diastolic BP. In the cross-sectional
12 studies, a pooled estimate indicated an increase in systolic BP of 0.26 mmHg (95% CI:
13 0.02, 0.50) per 10 (ig/g tibia Pb. In the longitudinal studies, a 0.33 mmHg (95% CI: -0.44,
14 1.11) increase was estimated per 10 (ig/g bone Pb. Most studies also reported associations
15 of bone Pb with hypertension. Pooled odds ratios for hypertension of 1.04 (95% CI: 1.01,
16 1.07) per 10 (ig/g increase in tibia Pb and 1.04 (95% CI: 0.96, 1.12) per 10 (ig/g increase
17 in patella Pb were reported.
November 2012 5-323 Draft - Do Not Cite or Quote
-------
First author, year
Tibia lead
Prospective
Glenn 2006"
Cheng 2001 '3
Glenn 200312
Cross-sectional
Lee 2001'"
Hu 96'VCheng 01
Martin'8 2008
SchwartzK2000
KorrickJ51999
Patella lead
Cheng'3 2001
Hu 96's/Cheng 01
Korrick'5 1999
Mean Increase in SBP (95%CI) Increase in DBP (95%CI) Hypertension RR or OR (95%CI)
Lead
fuo/a)
3S-4 -0.02 (-0.03 to 0.004) 1
21-9 --
14.7 0.78 (0.24 to 1.31)
Overall: 0.33 (-0.44 to 1. 11)
-1
Increase i
37.2 0.20 (-0.05 to 0.45}
5 32.1 1.01 (0.01 to 2. 02)
18.8 Q.20(-Q.SQto 1.10)
14.4 0.74 (-0.73 to 2. 21)
13.3 -
Overall: 0.26 {0.02 to 0.50)
31 .4 --
5 32.1 0.29 (-0.36 to 0.95)
17.3 -
1
• 0.07 (-0.30 to 0.45) ^|
II 1 1 1 1 1 1 1 1
17. -1 fl 1 5 OSS 1 1 5 1 S
i SBP (mmHg / year) Increase in DBP {mrnHg / year) Hypertension RR
-R- -0.02 (-0.20 to 0.1 7) ^
• 0 "0 ' 0 nn to 0 ^0)
f Cj 05 f.f) 7K [Q -5 ^gl
0.02 (-0.1 5 to 0.19)
— •
__
^ 1.05(1.00101.11)
1.15 (0.97 to 1.35}
"• 1.13 (0.98 !o 1.29}
— • 0.90 (0.70 to 1 .17} ^ — •
1.03(1.00101.05)
1.04 (1.01 10 1.07)
1.14 (1.01 to 1.28}
1.09(0.98101.22}
1 .00 (0.98 to 1 .03} |
1.04(0.96101.12)
-•—
•
•
•
•
1
-1012 -1012 0.85 1 1.2 1.5
increase in SBP (mrnHg) Increase in DBP (mrnHg) Hypertension OR
In the Normative Aging Study, Hu et al. (1996a) reported the cross-sectional association between bone Pb levels and the
prevalence of hypertension and Cheng et al. (2001) reported the cross-sectional association between bone Pb levels and systolic
BP in study participants free of hypertension at baseline.
Note: The studies are ordered by increasing mean bone Pb levels. The area of each square is proportional to the inverse of the
variance of the estimated change or log relative risk. Horizontal lines represent 95% confidence intervals. Diamonds represent
summary estimates from inverse-variance weighted random effects models. Because of the small number of studies, summary
estimates are presented primarily for descriptive purposes. RR indicates risk ratio.
Source: Reprinted with permission of Elsevier Publishers, Navas-Acien et al. (2008)
Figure 5-23 Meta-analysis of an increase in systolic BP (SBP) and diastolic BP
(DBP) and relative risk of hypertension per 10 ug/g increase in
bone Pb levels.
i
2
o
6
4
5
6
9
10
A few recent epidemiologic studies also emphasized the potential interaction between
measures of long-term Pb exposure, i.e., bone Pb levels, and factors such as chronic
stress and HFE genetic variants to moderate or modify the relationship of BP and
hypertension with Pb. For example, among NAS men, tibia Pb level was associated with
a larger risk of developing hypertension in an originally nonhypertensive group among
men with higher self-reported stress (Peters et al.. 2007).
In addition to stress, recent epidemiologic studies investigated effect modification by
race/ethnicity and genetic variants. In the NHANES 1988-1994 population of adults, the
association of concurrent blood Pb with systolic BP was higher among Mexican
Americans. In the same NHANES population, the association between blood Pb level and
November 2012
Draft - Do Not Cite or Quote
-------
1 hypertension was higher among non-Hispanic Blacks with the ALAD2 allele (see Figure
2 5-18 and Figure 5-19 for results) (Scinicariello et al.. 2010). Additionally, the association
3 between blood Pb and PP was larger among NAS men with the HFE H63D variant
4 (Figure 5-18) (Zhang etal.. 2010a). PP represents a good predictor of cardiovascular
5 morbidity and mortality and an indicator of arterial stiffness. The aforementioned genes
6 are related to iron metabolism and have been linked with differences in Pb distribution in
7 blood and bone. Park et al. (2009b) provided further evidence of variants in iron
8 metabolism genes impacting the association of bone Pb levels with QT interval changes
9 (see Table 5-21 for results).
10 Animal toxicological evidence continued to build on the evidence characterizing the
11 mechanisms leading to these Pb-induced cardiovascular alterations. Biological
12 plausibility for the consistent associations observed between blood and bone Pb and
13 cardiovascular effects is provided by enhanced understanding of Pb-induced oxidative
14 stress including NO inactivation, endothelial dysfunction leading to altered vascular
15 reactivity, activation of the RAAS, and vasomodulator imbalance.
5.4.3 Vascular Effects and Cardiotoxicity
16 Not only has Pb been shown to increase BP and alter vascular reactivity, but Pb can alter
17 cardiac function, initiate atherosclerosis, and increase cardiovascular mortality. Past
18 toxicological studies have reported that Pb can increase atheromatous plaque formation in
19 pigeons, increase arterial pressure, decrease heart rate and blood flow, and alter cardiac
20 energy metabolism and conduction (Prentice and Kopp. 1985; Revis etal.. 1981). A
21 limited number of available epidemiologic studies discussed in the 2006 Pb AQCD (U.S.
22 EPA. 2006b) provided evidence of associations of blood Pb level with ischemic heart
23 disease (IHD) and peripheral artery disease (PAD).
5.4.3.1 Effects on Vascular Cell Types
24 The endothelial layer is an important constituent of the blood vessel wall, which regulates
25 macromolecular permeability, VSMC tone, tissue perfusion, and blood fluidity. Damage
26 to the endothelium is an initiating step in development of atherosclerosis, thrombosis, and
27 tissue injury. Given that epidemiologic and toxicological evidence suggests that long-
28 term Pb exposure is associated with a number of these conditions, numerous
29 toxicological studies have investigated and found an effect of Pb on endothelial
30 dysfunction. A recent occupational study found that endothelial function assessed by
November 2012 5-325 Draft - Do Not Cite or Quote
-------
1 flow-mediated dilatation was impaired in highly Pb-exposed workers (mean blood Pb
2 levels: 24.1 in workers versus 7.8 (ig/dL in unexposed controls) (Por^baet al.. 2010).
3 The endothelial layer makes up only a small part of the vascular anatomy; the majority of
4 the vessel wall is composed of VSMCs, which work in concert with the endothelial cells
5 (EC) in contraction and relaxation of the vessel, local BP regulation, and atherosclerotic
6 plaque development. Since Pb has been shown repeatedly to result in hypertension and
7 vascular disease in experimental animals, studies continued to investigate and find an
8 effect of Pb on VSMCs.
9 In in vitro assays, Pb (50 (iM, 2 weeks) stimulated VSMC invasiveness in isolated human
10 arteries leading to the invasion of medial VSMC into the vessel intima and development
11 of intimal hyperplasia, a key step in atherosclerotic progression (Zeller et al.. 2010). In
12 addition, treatment with Pb (50 (iM, 12 hours) promoted VSMC elastin expression and
13 increased arterial extracellular matrix in isolated human arteries. VSMC invasiveness was
14 also increased in culture by treatment with supernatant of Pb-treated human EC (50 (JVI),
15 suggesting that Pb-exposed ECs secrete an activating compound. This compound was
16 confirmed to be IL-8. Pb exposure (5-50 (iM) was able to, in a concentration-dependent
17 manner, increase IL-8 synthesis and secretion in human umbilical vein EC cultures
18 through activation of the transcription factor Nrf2. Neutralization of IL-8 could block
19 VSMC invasion and arterial intima thickening (Zeller et al.. 2010). This study provides
20 evidence that Pb exposure stimulates ECs to secrete IL-8 in an Nrf2-dependent manner
21 which stimulates VSMC invasion from the vessel media to intima leading to a vascular
22 thickening and possibly atherogenesis.
23 A number of CVDs, including atherosclerosis, are characterized by increased
24 inflammatory processes. Numerous studies have shown that Pb exposure is associated
25 with an inflammatory environment in vascular tissues of humans and animals as indicated
26 by higher levels of inflammatory mediators like prostaglandin E2 (PGE2). Human aortic
27 VSMCs treated with Pb (1 (JVI, 1-12 hours) exhibited increased secretion of PGE2 time-
28 dependently through enhanced gene transcription (Chang et al.. 2011). This was preceded
29 by a Pb-induced increase in the gene expression of cytosolic phospholipase A2 (cPLA2)
30 and COX-2, two rate limiting enzymes in the regulation of prostaglandins. The induction
31 of these enzymes was mediated by activation of ERK1/2, MEK1, and MEK2. Further
32 investigation of the entrance of Pb into the cell revealed that inhibition of the store-
33 operated calcium channels (SOC) could only partially suppress cPLA2 and COX
34 activation by Pb; however inhibition of epidermal growth factor receptor (EGFR)
35 attenuated Pb-induced PGE2 secretion and activation of cPLA2 and COX. A follow-up to
36 this study found that Pb treatment (1(JVI) of a human epithelial cell line increased COX-2
37 gene expression, promoter activity, and protein (Chou et al.. 2011). Inhibition of NF-KB
November 2012 5-326 Draft - Do Not Cite or Quote
-------
1 decreased the Pb-induced COX-2 activation; whereas EGFR inhibition blocked COX-2
2 upregulation and NF-KB nuclear translocation. Overall these results suggest that Pb can
3 induce pro-inflammatory events in VSMC in the form of increased PGE2 secretion and
4 expression of cPLA2 and COX-2 through activation of EGFR via ERK1/2 and NF-KB
5 pathways.
6 Damage to the endothelium is a hallmark event in the development of atherosclerosis.
7 Past studies have shown that Pb exposure results in de-endothelialization, impaired
8 proliferation, and inhibition of endothelium repair processes after injury (Fujiwara et al.,
9 1997; Uedaetal.. 1997; Kajietal.. 1995; Kishimoto et al. 1995). However, Pb exposure
10 was not found to lead to nonspecific cytotoxicity at low exposure levels (2-25 (iM) as
11 shown by the lack of release of lactate dehydrogenase (LDH) from Pb-treated bovine
12 aortic EC (Shinkai etal. 2010). Instead, Pb induced specific apoptosis (caspase3/7
13 activation) through endoplasmic reticulum (ER) stress that was protected against by the
14 ER chaperones glucose-regulated protein 78 (GRP78) and glucose-regulated protein 94
15 (GRP94). GRP78 and GRP94 play key roles in the adaptive unfolded protein response
16 that serves as a marker of and acts to alleviate ER stress. Exposure of ECs to Pb induced
17 GRP78 and GRP94 gene (2-25 \M) and protein (GRP78 [5-25 \M\ and GRP94
18 [10-25 \\M\) expression through activation of the IREl-JNK-AP-1 pathways (Shinkai et
19 al., 2010). This finding suggests that the functional damage in ECs caused by Pb
20 exposure may be partly attributed to induction of ER stress.
5.4.3.2 Cholesterol
21 As blood cholesterol rises so does the risk of coronary heart disease. Previous
22 occupational studies (Ademuviwa et al., 2005a; Beneretal.. 200Ib; Kristal-Boneh et al.,
23 1999) examining higher than current adult blood Pb levels (>40 (ig/dL) reported higher
24 total cholesterol levels related to Pb exposure, but mixed results for HDL, LDL, and
25 triglycerides. More recently, Poreba et al. (2010). in an occupational study, reported no
26 significant differences in parameters of lipid metabolism between Pb exposed workers
27 (mean blood Pb level: 25 (ig/dL) and unexposed individuals. Conversely, Kamal et al.
28 (2011) reported that occupational Pb exposure (mean blood Pb level: >40 (ig/dL) was
29 associated with higher levels of triglycerides, total cholesterol, and LDL, and decreased
30 HDL-C. Other Pb studies adjusted models for total cholesterol to control for this coronary
31 heart disease risk factor. Higher mean total cholesterol with higher blood Pb levels has
32 been reported in aNHANES study (Menke et al., 2006). In developing models to predict
33 bone Pb levels, Park et al. (2009c) noted in a NAS study that total and HDL cholesterol
34 were selected as 2 of 18 predictors for the bone Pb level model. Their findings suggested
35 that higher Pb exposure in nonoccupationally-exposed men may be associated with
November 2012 5-327 Draft - Do Not Cite or Quote
-------
1 higher total and HDL cholesterols. In support of epidemiologic evidence, a recent
2 toxicological study reported increased LDL and decreased HDL in rats treated with Pb
3 (20 ppm, i.p., 3 days/week, 8 weeks) (Roshan et al., 2011). The major risk factor that
4 lipids represent for heart disease make relating lipid levels to Pb exposures an interesting
5 but challenging hypothesis to test.
5.4.3.3 Atherosclerosis
6 A small number of toxicological and cross-sectional epidemiologic studies provide
7 evidence for increased atherosclerosis and intimal medial thickening (IMT) due to Pb
8 exposure. The association of stroke subtypes and severity of cerebral atherosclerosis was
9 examined in relation to a single concurrent blood Pb level and total 72-hour urinary Pb
10 level (body Pb store-EDTA mobilization test) in a cross-sectional study of 153 patients
11 (mean age 63.7 years) receiving digital subtraction angiography in Chang Gung
12 Memorial Hospital in Taiwan from 2002 to 2005 (Lee et al., 2009). In an analysis
13 adjusted for age, sex, hypertension, diabetes, triglyceride, uric acid, smoking, and alcohol
14 consumption, a 1 (ig increase in urine Pb was associated with > 50% stenosis in the
15 intracranial carotid system with an OR of 1.02 (95% CI: 1.00, 1.03). Urine Pb was not
16 associated with greater stenosis in the extracranial or vertebrobasilar systems. Blood Pb
17 level was not associated with greater stenosis in any region. As the development of
18 atherosclerosis is a lifelong process, body Pb stores, analyzed by total 72-hour urine Pb
19 amount, may more strongly be associated with atherosclerosis than are single blood Pb
20 measurements.
21 A recent study correlated greater carotid artery IMT with higher concurrent serum Pb
22 levels (mean [SD] 0.41 [0.38] ng/dL) in hemodialysis patients (Ari et al.. 2011). A few
23 available recent occupational studies also presented evidence for increased measures of
24 atherosclerosis in highly Pb-exposed adult populations with mean blood Pb levels around
25 25 (ig/dL. Por^ba et al. (2011 a) reported increased local arterial stiffness and more
26 frequent left ventricular diastolic dysfunction in Pb-exposed workers with hypertension
27 compared to nonexposed controls with hypertension. Occupational exposure to Pb (mean
28 blood Pb levels: 24 (ig/dL in workers, 8.3 (ig/dL in nonexposed group) was also
29 associated with greater IMT and atherosclerotic plaque presentation, analyzed by Doppler
30 ultrasound (Porebaet al., 2011).
31 Zeller et al. (2010) examined human radial and internal mammary arteries exposed to Pb
32 in culture and reported a concentration-dependent increase in arterial intimal thickness
33 (statistically nonsignificant at 5 (iM Pb, significant at 50 (iM Pb, 2 week treatment) and
34 intimal extracellular matrix accumulation (50 (JVI). Also, Pb promoted EC proliferation
November 2012 5-328 Draft - Do Not Cite or Quote
-------
1 (5 and 50 (JVI, 72 hours) and VSMC elastin expression (50 (JVI, 12 hours), as discussed
2 above (Section 5.4.3.1) (Zeller et al.. 2010). Another study showed that Pb exposure
3 (100 ppm in drinking water for 10 months; mean blood Pb level 28.4 (ig/dL) of rats also
4 increased the aortic media thickness, media-lumen ratio, and medial collagen content
5 (Zhang et al., 2009a). These morphological changes to the vessel due to Pb exposure
6 indicate initiation of arteriosclerosis and could be the cause of decreased contractile
7 response of the vessel due to altered visco-dynamic vessel properties. Alternatively, these
8 vascular changes could be an effect of Pb-induced hypertension.
5.4.3.4 Heart Rate Variability
9 HRV and BP are regulated, in part, by the sympathetic and parasympathetic nervous
10 systems. Changes in either may increase the risk of cardiovascular events. HRV is
11 defined as the oscillation in the interval between consecutive heart beats and between
12 consecutive instantaneous heart rates. Decreases in HRV have been associated with
13 cardiovascular mortality/morbidity in older adults and those with significant heart disease
14 [(1996). Task Force of the European Society of Cardiology and the North American
15 Society of Pacing and Electrophysiology]. In addition, decreased HRV may precede
16 some clinically important arrhythmias, such as atrial fibrillation, as well as sudden
17 cardiac death, in high risk populations (Chen and Tan. 2007; Sandercock and Brodie.
18 2006).
19 Pb has been shown not only to affect vascular contractility in animals, but also is
20 associated with cardiac contractility. The 2006 Pb AQCD (U.S. EPA. 2006b) described
21 one study that investigated Pb-induced alterations in HRV (Cheng etal. 1998). Cheng et
22 al. (1998) found increasing duration of corrected QT interval (QTc) with increasing bone
23 Pb levels in men <65 years, but not in men > 65 years. Bum et al. (2011) and Park et al.
24 (2009b) followed up this previous NAS cohort (Cheng etal., 1998) (details found in
25 Table 5-21). Bum et al. (2011) prospectively examined the association between blood and
26 bone Pb levels and the development of electrocardiographic (ECG) conduction
27 abnormalities among 600 men who were free of ECG abnormalities at the baseline
28 assessment. A second ECG was obtained for 496 men 8.1 (SD: 3.1) years later on
29 average. Baseline Pb concentrations in blood (mean [SD]: 5.8 [3.6] (ig/dL), patella bone
30 (mean [SD]: 30.3 [17.7] (ig/g), and tibia bone (mean [SD]: 21.6 [12.0] (ig/g) were similar
31 to those found in other samples from the general U.S. adult population and much lower
32 than those reported in occupationally exposed groups. Higher tibia Pb was associated
33 with increases in QTc interval and QRSc duration. Compared with those in the lowest
34 tertile of baseline tibia Pb (<16 (ig/g), participants in the highest tertile (>23 (ig/g) had a
35 7.94 msec (95% CI: 1.42, 14.45) greater increase in QTc interval and a 5.94 msec (95%
November 2012 5-329 Draft - Do Not Cite or Quote
-------
1 CI: 1.66, 10.22) greater increase in QRSc duration over 8 years after adjusting for
2 covariates: age, education, smoking, BMI, albumin-adjusted serum calcium, and diabetes
3 status at baseline, and years between ECG tests and QT-prolongation drugs at the time of
4 ECG measurement. There were no statistically significant associations with patella or
5 blood Pb levels. These associations with tibia bone Pb levels were observed in men with
6 relatively low blood and bone Pb concentrations who were free of cardiac conduction
7 abnormalities at baseline and were examined prospectively. Thus, they indicate that long-
8 term cumulative Pb exposure may increase the risk of developing cardiac abnormalities.
9 Uncertainty exists as to the specific Pb exposure level, timing, frequency, and duration
10 contributing these associations observed for tibia Pb levels. A recent occupational study
11 reported lower HRV and abnormal parameters of heart rate turbulence in Pb-exposed
12 workers (mean blood Pb levels: -25 (ig/dL) compared to control subjects (Por^ba et al.,
13 201 Ib).
14 Park et al. (2009b) cross-sectionally examined whether polymorphisms in genes known
15 to alter iron metabolism (HFE, transferrin [TF] C2, heme oxygenase-1 [HMOX-1])
16 modify the association between Pb biomarker levels and the QT interval. Investigators
17 examined associations in data stratified on polymorphisms in the three genes. They also
18 analyzed interaction models with cross-product terms for genotype and the Pb biomarker.
19 The distributions of all genotypes but the HFE variant, H63D, were in Hardy-Weinberg
20 equilibrium. Subjects homozygous for the other HFE variant, C282Y, had higher bone Pb
21 levels and those homozygous for H63D and heterozygous with both C282Y and H63D
22 had lower bone Pb levels. The antioxidant HMOX-1 L variant (longer repeats of GT,
23 associated with lower enzyme inducibility) alone, compared to the wild type, showed a
24 statistically significant interaction with tibia Pb (11.35 msec longer QTc interval for each
25 13 (ig/g increase in bone Pb in L-allele variants). No other gene variant alone showed
26 different Pb-associated QTc intervals from those in wild types, either for tibia and patella
27 Pb or for (linear) concurrent blood Pb. Lengthening of QTc with higher tibia and blood
28 Pb was more pronounced with an increase in the total number of gene variants, driven by
29 a joint effect between HFE variant and HMOX-1 L allele. There was a trend observed
30 with blood and tibia Pb-associated QTc interval increasing with increasing number of
31 gene variants from 0 to 3. This study provided further evidence of gene variants
32 modifying associations of Pb biomarkers with cardiovascular effects.
33 The interaction of key markers of the metabolic syndrome with bone Pb levels in
34 affecting HRV was cross-sectionally investigated in a group of 413 older adults with
35 patella Pb measurements in the NAS (Park et al.. 2006). Metabolic syndrome was defined
36 to include three or more of the following: waist circumference >102 cm,
37 hypertriglyceridemia (>150 mg/dL), low HDL cholesterol (<40 mg/dL in men), high BP
38 >130/85 mmHg, and high fasting glucose (>110 mg/dL). Men using antihypertensive
November 2012 5-330 Draft - Do Not Cite or Quote
-------
1 medication or diabetes medications were counted as high BP or high fasting glucose,
2 respectively. The strongest relationships between patella Pb levels and lower HRV were
3 observed among those with three or more metabolic abnormalities. A trend was observed
4 for larger patella Pb-associated decreases in HRV with increasing number of metabolic
5 abnormalities. These results suggest multiplicative effects of cumulative Pb exposure and
6 metabolic abnormalities on key predictors of CVD. Park et al. (2006) also reported the
7 penalized spline fits to bone Pb in models assessing only main effects of bone Pb. The
8 optimal degree of smoothing determined by the generalized cross-validation criterion for
9 all HRV measures was 1, which indicated that the associations were nearly linear. The
10 spline fits and associated statistics showed that the bone Pb main effects on HRV
11 measures were linear. However, the relationship with LF/HF was linear with log(LFTHF).
12 Increased incidence of arrhythmia and atrioventricular conduction block was found in
13 rats after 12 weeks of Pb exposure (100 ppm; mean blood Pb level 26.8 (ig/dL) (Reza et
14 al.. 2008). Also, Pb exposure for 8 weeks increased heart rate and systolic BP. These
15 increases corresponded with increased cardiac contractile force and prolonged ST
16 interval, without alteration in QRS duration or coronary flow. In contrast, another study
17 using rat right ventricular strips found that Pb (100 (iM) exposure, in a concentration-
18 dependent manner, reduced myocardial contraction by reducing sarcolemmal Ca2+ influx
19 and myosin ATPase activity (Vassallo et al.. 2008). This study also found that Pb
20 exposure changed the response to inotropic agents and blunted the force produced during
21 contraction. Conversely, past studies have found that Pb exposure increases intracellular
22 Ca2+ content (Laletal.. 1991: Favalli et al.. 1977: Piccinini et al.. 1977). which could
23 result in increased cardiac output and hypertension.
5.4.3.5 Peripheral Artery Disease
24 Peripheral artery disease (PAD) is an indicator of atherosclerosis and measured by the
25 ankle brachial index, which is the ratio of BP between the posterior tibia artery and the
26 brachial artery. PAD is typically defined as an ankle brachial index of less than 0.9.
27 Muntner et al. (2005). whose results describing the association of blood Pb and
v / 7 O
28 hypertension in the NHANES 1999-2002 data set for adults were discussed previously,
29 also examined the association of blood Pb with PAD (details found in Table 5-21). The
30 authors observed an increasing trend in the odds of PAD with increasing concurrent
31 blood Pb level. The OR for PAD comparing the fourth quartile of blood Pb (>2.47 (ig/dL)
32 to the first quartile of blood Pb (<1.06 (ig/dL) was 1.92 (95% CI: 1.02, 3.61). Key
33 potential confounding factors were adjusted for in the analysis. These results are
34 consistent with those from a previous NHANES analysis by Navas-Acien et al. (2004)
35 reviewed in the 2006 Pb AQCD.
November 2012 5-331 Draft - Do Not Cite or Quote
-------
1 Navas-Acien et al. (2004) reported a trend of increasing OR for PAD with increasing
2 quartile of concurrent blood Pb or Cd in adults who were 40 years of age in the
3 1999-2000 NHANES population. These authors tested both Pb and Cd in separate
4 models, tested the metals simultaneously, and tested the interaction between the metals.
5 The correlation coefficient between natural log Pb and natural log Cd was 0.32
6 (p <0.001). Although the interaction was not statistically significant, when blood Pb and
7 blood Cd were in the same model, the ORs were diminished slightly. Both showed
8 statistically significant trends of increasing OR with increasing quartile of the metal.
9 These results indicate that blood Cd levels did not confound the association between
10 blood Pb level and PAD. In a subsequent analysis, Navas-Acien et al. (2005) used the
11 same 1999-2000 NHANES dataset, but constructed PAD models using a suite of urine
12 metal concentrations. Power was reduced in this study because only 659-736 subjects
13 (compared to 2,125) had spot urine metal tests in the data set. Urinary Cd, but not urinary
14 Pb, was consistently associated with PAD in all models. Associations also were observed
15 with urinary antimony and tungsten. Spot urine Pb measurements are less reliable
16 compared to blood Pb measurements. In Navas-Acien et al. (2005). the urinary Pb level
17 association with PAD was sensitive to adjustment for urinary creatinine, indicating that
18 spot urine Pb measurements are affected by differences in urine dilution. This finding
19 illustrates the limited reliability of spot urine Pb measurements compared to blood Pb
20 measurements.
5.4.3.6 Ischemic Heart Disease
21 A few cross-sectional studies discussed in the 2006 Pb AQCD (U.S. EPA. 2006b)
22 indicated associations between Pb biomarker levels and increased risk of cardiovascular
23 outcomes associated with IHD, including left ventricular hypertrophy (Schwartz. 1991)
24 and myocardial infarction (Gustavsson et al.. 2001). Recently, Jain et al. (2007) provided
25 prospective evidence for the incidence of IHD (physician confirmed MI, angina pectoris)
26 among older adult males enrolled in the NAS that were followed during the period of
27 September, 1991 to December, 2001 (details found in Table 5-21). All subjects had blood
28 Pb and bone Pb measurements with no IHD at enrollment. Fatal and nonfatal cases were
29 combined for analysis. Baseline blood, tibia, and patella Pb levels were log-transformed.
30 Blood Pb level and patella Pb level were associated with increased risk of IHD over the
31 10-year follow-up period. When blood Pb and patella Pb were included simultaneously in
32 the model, each of their HRs was only moderately attenuated (HR: 1.24 [95% CI: 0.80,
33 1.93] per SD increase in blood Pb and HR: 2.62 [95% CI: 0.99, 6.93] per SD increase in
34 patella Pb). When blood Pb and tibia Pb were included simultaneously in the model, their
35 risk estimates were only moderately attenuated (HR: 1.38 [95% CI: 0.89, 2.13] per SD
November 2012 5-332 Draft - Do Not Cite or Quote
-------
1 increase in blood Pb and HR: 1.55 [95% CI: 0.44, 5.53] per SD increase in tibia Pb).
2 These findings indicate that both blood and bone Pb levels are independently associated
3 with IHD incidence.
4 IHD, characterized by reduced blood supply to the heart, may result from increased
5 thrombosis. In support of the epidemiologic evidence, a recent animal study suggested
6 that Pb exposure promotes a procoagulant state that could contribute to thrombus
7 formation (Shin et al.. 2007). In a rat model of venous thrombosis, Pb treatment (i.v.
8 25 mg/kg) resulted in increased thrombus formation, although i.v. Pb treatment may have
9 uncertain relevance to human routes of Pb exposure. Additionally, Pb treatment to human
10 erythrocytes (red blood cells, RBCs) increased coagulation at a dose of 5 (iM and
11 thrombin generation in a concentration-dependent manner at doses from 2-5(iM. This
12 enhanced procoagulant activity in Pb-treated RBCs was the result of increased outer cell
13 membrane phosphatidylserine (PS) surfacing (human RBCs: 2-5 (iM Pb; rat RBCs: 5 (iM
14 Pb). Similar to these in vitro results, PS externalization on erythrocytes was increased in
15 Pb-treated rats (i.v. 50-100 mg/kg, not 25 mg/kg). Increased PS externalization was likely
16 the result of increased intracellular calcium (5 (iM Pb), enhanced scramblase activity
17 (5-10 (iM Pb), inhibited flippase activity (5-10 (iM Pb), and ATP depletion (1-5 (iM Pb)
18 after Pb exposure (Shin et al.. 2007).
November 2012 5-333 Draft - Do Not Cite or Quote
-------
Table 5-21 Characteristics and quantitative data for associations of blood and
bone Pb with other CVD measures HRV, PAD, and IHD in recent
epidemiologic studies.
Study
(Ordered as Study
they appear in Population/
the text)
Heart rate
Eum et al.
(2011)
Parket al.
(2009b)
Methodology
variability
Longitudinal
600 men free of
electrographic
abnormalities at
the time of
baseline ECG
from MAS in
Greater Boston,
MA area (496
with follow-up
ECG 8 years
later)
Cross-sectional
61 3 men from
MAS in Greater
Boston MA area
(8/1 991'-
12/1995)
Parameter Pb Data
ECG Baseline Blood Pb:
conduction" Mean (SD): 5.8
(QTc, QRSc, (3.6) ug/dL
JTc,
QT prolongation,
JT prolongation, Baseline Patella Pb:
IVCD° AVCD Mean (SD): 30.3
Arrhythmia) ' (17.7) ug/g
Baseline Tibia Pb:
Mean (SD): 21.6
(12.0) ug/g
Q1: <16 ug/g (n = 191)
Q2: 16.0-23 ug/g
(n = 208)
Q3: >23 ug/g (n = 195)
QTcb interval Blood Pb:
Median (IQR):
5 (4-7) ug/dL
Patella Pb:
Median (IQR):
26 (18-37) ug/g
Tibia Pb:
Median (IQR):
19 (14-27) ug/g
Statistical Analysis
Repeated measures
linear regression
adjusted forage,
education, smoking,
BMI, albumin-adjusted
serum Ca +, and
diabetes status at
baseline, and years
between ECG tests
and QT-prolongation
drugs at the time of
ECG measurement.
Linear regression
models adjusted for
age, BMI, smokina
status, serum Ca ,
and diabetes. No SES
indicator was
considered.
Effect Estimate (95%
r*na
Cl)
Tibia Pb:
Adjusted 8-year
change
(95% Cl):
QTc:
Q2 vs. Q1 (reference):
7.49(1.22, 13.75)
msec,
Q3vs. Q1:
7.94(1.42,14.45)
msec
p for trend = 0.03
QRSc:
Q2vs. Q1:
0.52 (-3.60, 4.65)
msec
Q3vs. Q1:
5.94(1.66,10.22)
msec
p for trend = 0.005
No associations with
patella or blood Pb
Per IQR (3 ug/dL)
increase in blood Pb:
1.3 (-0.76, 3.36) msec
after 8-year follow up
Per IQR (19 ug/g)
increase in patella Pb:
2.64(0.13,5.15) msec
Per IQR (13 ug/g)
increase in tibia Pb:
2.85 (0.29, 5.40) msec
November 2012
Draft - Do Not Cite or Quote
-------
Study
(Ordered as
they appear in
the text)
Study
Population/
Methodology
Parameter
Pb Data
Statistical Analysis
Effect Estimate (95%
Cl)a
Parket al.
(2006)
Cross-sectional
413 men from
MAS in Greater
Boston, MA area
(11/14/2000-
12/22/2004)
HRV
(SDNN, HF,
r~irnormi Lr,
LFnorm, LF/HF)
Patella Pb (measured
within 6 mo of HRV:
Median (IQR):
23.0 (15-34) ug/g
Estimated3: Median
(IQR):
16.3(10.4-25.8) ug/g
Tibia Pb:
Median (IQR):
19.0(11-28) ug/g
Log linear regression
models adjusted for
age, cigarette
smoking, alcohol
consumption, room
temperature, season
(model 2) BMI, fasting
blood glucose, HDL
cholesterol,
triglyceride, use of p-
blockers, Ca2+ channel
blockers, and/or ACE
inhibitors. No SES
indicator was
considered.
Tibia Pb: Model 2
Change (95%CI)
HF:
-0.9 (-3.8, 2.1)
normalized units (nu)
LF:
0.9 (-2.0, 3.9) nu
Log LF/HF:
3.3 (-10.7, 19.5) (%)
Per 17 ug/g tibia Pb
Patella Pb:
Model 2 Change
(95%CI)
HF:-0.6 (-3.1, 1.9)
nu
LF: 0.6 (-1.9, 3.1)nu
Log LF/HF: 3.0 (-8.7,
16.2) (%)
Per 15.4 ug/g patella
Pb
Effect estimates were
more pronounced
among those with
greater* metabolic
abnormalities.
Peripheral artery disease
Muntner et al. Cross-sectional PAD
(2005) g 961 NHANES
(1999-2002)
participants
Navas-Acien Cross-sectional PAD
et al. (2QQ5) 7go partjcjpants,
age > 40 yr,
from NHANES
(1 999-2000)
Range Concurrent Blood
Pb:
Q1:<1.06ug/dL,
Q2: 1 .06-1 .63 ug/dL
Q3: 1 .63-2.47 ug/dL
Q4: >2.47 ug/dL
Concurrent urinary Pb:
Mean (1 Oth-90th
percentile)):
0.79 ug/L (0.2-2.3)
Logistic regression
models adjusted for
age, race/ethnicity,
sex, diabetes mellitus,
BMI, cigarette
smoking, alcohol
consumption, high
school education,
health insurance
status
Logistic regression
adjusted for the
following:
Model 1: age, sex,
race, and education
Model 2: covariates
above plus smoking
status
Model 3:covariates
above plus urinary
creatinine
OR (95% Cl):
Q1 : 1 .00 (Reference),
Q2: 1.00(0.45,2.22),
Q3: 1.21 (0.66,2.23),
Q4: 1.92(1.02,3.61)
Model 1:
OR: 1.17(0.81, 1.69)
Model 2:
OR: 1.1 7 (0.78, 1.76)
Model 3:
OR: 0.89 (0.45, 1.78)
Per IQR increase in
i irinsrx/ Ph
ui M icti y ~u
Array of metals in
urine also evaluated.
November 2012
5-335
Draft - Do Not Cite or Quote
-------
Study
(Ordered as
they appear in
the text)
Study
Population/
Methodology
Parameter
Pb Data
Statistical Analysis
Effect Estimate (95%
Cl)a
Ischemic Heart Disease
Jain et al. Longitudinal IHD
t^uU' ' 837 men from (Ml or anoina
NAS in Greater pectoris)
Boston, MA area
(1991-2001)
Baseline Blood Pb Mean
(SD):
Non-cases
6.2 (4.3) ug/dL;
Cases
7.0 (3.8) ug/dL
Baseline Patella Pb
Mean (SD):
Non-cases
30.6(19.7) ug/dL;
Cases
36.8 (20.8) ug/dL
Cox proportional
hazards models
adjusted forage, BMI,
education, race,
smoking status, pack-
years smoked, alcohol
intake, history of
diabetes mellitusand
hypertension, family
history of
hypertension, DBP,
SBP, serum
triglycerides, serum
HDL, and total serum
cholesterol
Blood Pb level
> 5 ug/dL
OR over 10-year
follow-up: 1.73(1.05,
2.87)
Ln [blood Pb] OR:
1.45(1.01,2.06)
Ln [patella Pb level]
OR'
2.64(1.09,6.37)
Ln [tibia Pb level ]OR:
1.84(0.57,5.90)
Baseline Tibia Pb Mean
(SD):
Non-Cases
21.4 (13.6) ug/g;
Cases
24.2(15.9) ug/g
Cases:
Blood Pb range:
1.0 to 20.0 ug/dL
Patella Pb range:
5.0 to 101 ug/g
Tibia Pb range:
-5 to 75 ug/g
Per 1 SD increase in
Pb biomarker
"Estimated patella Pb accounts for declining trend in patella Pb levels between analysis of bone Pb and HRV.
bHeart-rate-corrected QT interval calculated by Bazett's formula
°IVCD, intraventricular conduction defect; AVCD, atrioventricular conduction defect
1
2
o
J
4
5
6
7
10
11
12
5.4.3.7 Summary of Vascular Effects and Cardiotoxicity
There are a limited number of studies in a limited number of populations that investigate
the associations between Pb biomarkers and cardiovascular effects other than BP or
hypertension (Table 5-21). As presented in Table 5-21. these studies demonstrated
associations between various biomarkers of Pb exposure and clinical cardiovascular
outcomes such as atherosclerosis, IHD, PAD, and HRV occurrence in adult populations
after adjusting for potential confounding by variables such as age, sex, education, BMI,
smoking, alcohol consumption, and diabetes. In a limited body of studies, mixed
evidence of association between occupational exposure to Pb and altered cholesterol was
reported.
Few studies have evaluated markers of subclinical atherosclerosis such as PAD and IMT
following Pb exposure in humans or animals. Concurrent blood Pb levels (population
means >2.5 (ig/dL) were associated with greater odds of PAD in adults in NHANES
November 2012
5-336
Draft - Do Not Cite or Quote
-------
1 analyses (Muntner et al.. 2005; Navas-Acien et al., 2004). Since these effects are
2 observed in adults that may have had higher past exposure to Pb, there is uncertainty as to
3 the specific Pb exposure level, timing, frequency, and duration that contributed to the
4 observed associations. A recent study involving both human and toxicological studies
5 observed Pb-mediated arterial IMT, an early event in Pb-induced atherogenesis (Zeller et
6 al.. 2010). A second study in rats report increased aortic media thickness following Pb
7 exposure (Zhang et al., 2009a). Toxicological studies of Pb-induced endothelial
8 dysfunction, VMSC invasiveness, and inflammation in isolated vascular tissues and cells
9 provide mechanistic evidence to support the biological plausibility of these vascular
10 effects and cardiotoxicity. Studies in isolated tissues and cells found that Pb stimulated
11 the synthesis and secretion of IL-8 in ECs, which was responsible for stimulating VSMC
12 invasion into the vessel intimal layer. Pb treatment also increased extracellular matrix and
13 elastin, primary sites for lipid deposition in the vessel wall.
14 Several studies report associations between biomarkers of Pb exposure and diseases
15 associated with coronary heart disease (CHD), such as HRV, IHD, and MI. A prospective
16 NAS study reported that higher baseline tibia Pb was associated with increases in QTc
17 interval and QRSc duration over an 8-year follow-up period (Eumet al., 2011). In
18 addition, in the NAS cohort of older adult men, blood Pb (> 5 (ig/dL) and patella Pb
19 levels were associated with increased incidence of IHD (Jain et al., 2007). A recent study
20 provided evidence for the interaction between biomarkers of Pb exposure and the HFE
21 C282Y and HMOX-1 L variant on the prolonged QT interval in
22 nonoccupationally-exposed older men (Park et al.. 2009b). Also, in the NAS population,
23 bone Pb levels were associated with larger decreases in HRV parameters among subjects
24 identified as having metabolic abnormalities (Park et al.. 2006). These metabolic
25 abnormalities, abdominal obesity, hypertriglyceridemia, low HDL cholesterol, high
26 BP/medication use, or high fasting glucose, have been shown to be associated with
27 increased risk of cardiovascular events.
28 Overall, the relatively few available studies provide support for associations between Pb
29 biomarkers and other cardiovascular conditions including subclinical atherosclerosis and
30 CHD. A number of these are quality studies from two cohorts, NAS and NHANES with
31 adequate sample size that account for potential confounding, with some being conducted
32 prospectively.
November 2012 5-337 Draft - Do Not Cite or Quote
-------
5.4.4 Cardiovascular Function and Blood Pressure in Children
5.4.4.1 Introduction
1 The study of cardiovascular function effects in relation to blood Pb levels in children
2 potentially offers unique information on several topics. First, by examining endpoints
3 predictive of future cardiovascular pathology, these studies may offer information on the
4 potential cardiovascular effects of Pb exposure in an understudied population. Second,
5 examination of cardiovascular changes that are antecedent to increased BP and changes
6 in other CVD-related endpoints at later lifestages may inform uncertainties in regards to
7 the time course of cardiovascular changes associated with Pb exposure. Finally, these
8 studies address gaps in knowledge regarding Pb exposure effects in populations of
9 children with mean blood Pb levels in the range of <10 (ig/dL.
10 An important aspect to the literature about the association between cardiovascular effects
11 and blood Pb levels in children is that the blood Pb levels of children may better reflect
12 relatively recent Pb exposure and its effect on CVD than blood Pb levels do in adults
13 because of the much longer exposure history of adults during which Pb exposures were
14 commonly much higher than they are today. However, in older children there is still
15 uncertainty regarding the frequency, duration, timing, and magnitude of exposure
16 contributing to the blood Pb levels measured. The much lower prevalence of
17 cardiovascular effects in children, however, poses a challenge to investigations of
18 potential relationships with Pb exposures. For example, the prevalence of hypertension in
19 children (9 to 10 years old) ranges from to 2 to 5 percent (Daniels. 2011; Steinthorsdottir
20 et al.. 2011). while more than half of people aged 60 to 69 years have hypertension
21 (Chobanian et al., 2003). Accordingly, much larger study populations are required to
22 provide similar statistical power for such studies in children as compared to adult studies.
23 Further, in drawing interpretations from such studies with regard to potential effects of Pb
24 exposures at later ages, it is additionally important to recognize that compensatory
25 mechanisms in children may be more active than in adults, and the cardiovascular tissue
26 of the young may be less susceptible to damage than that of adults.
November 2012 5-338 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
The limited numbers of cardiovascular studies published on children have examined
endpoints such as total peripheral resistance (TPR), BP, and autonomic nervous system
activation. These recent and earlier studies are presented in Table 5-22. Multiple single
pollutant studies in New York State evaluated two child cohorts born in the 1990s after
Pb was removed from gasoline in the U.S. with mean blood Pb levels of 4.62 and
1.01 (ig/dL (Gump etal.. 2011: Gump et al.. 2009: Gump et al. 2007: Gump et al.. 2005).
Zhang et al. (2011 a) examined children in Mexico City born from 1994 to 2003, when Pb
was being taken out of gasoline in Mexico as indicated by Martinez et al. (2007). The
geometric means for cord and concurrent blood Pb levels of the children in the Mexico
City cohort were 4.67 and 2.56 (ig/dL.
Table 5-22 Studies of child cardiovascular endpoints and Pb biomarkers.
Study
(Ordered as
they appear in
the text)
Gump et al.
(2005)
Gump et al.
(2007)
Study
Population/
Methodology Parameters
Prospective SBP, TPR
122 children (total
age 9.5 yr in peripheral
Oswego, NY vascular
(born at a resistance)
single hospital
in New York
from 1991-94)
Prospective SBP, TPR
122 children
age 9.5 yr in
Oswego, NY
Blood Pb Data3
Cord blood Pb:
GM (GSD):
2.56ug/dL(1.16)
Childhood (mean
age of measurement:
2.6 yr) blood Pb: GM
/'OQnv
^ooUJ.
4.06ug/dL(1.14)
Childhood (mean
age of measurement:
2.6 yr) blood Pb:
GM (GSD):
4.06ug/dL(1.14)
Statistical Analysis
Multivariate linear
regression models
examined the
relationship of blood Pb
with change in z-score
for outcome (post- and
pre-stress). Potential
confounders
considered: HOME
score, SES, birth
weight, child BMI, child
sex.
Linear regression
models adjusting for
the same covariates as
in Gump et al. (2005).
Separate models
testing whether Pb is a
mediator of SES
associations, (Sobel
test) and whether Pb
moderates SES
associations (Pb-SES
interaction).
Effect Estimates/Results
Per 1 ug/dL increase in
childhood blood Pb level, 0.088
(95% Cl: 0.023, 0.153) dyne-
s/cm5 change in TPR
Per 1 ug/dL increase in cord
blood Pb level, 12.16 (95% Cl:
2.44, 21 .88) mmHg higher SBP
Blood Pb was a mediator of the
SES-TPR relationship
SES alone: -0.62 dyne-s/cm5
(p <0.05)
SES with Blood Pb: -0.40
dyne-s/cm5 (p >0.10), change
in R2 attributable to SES:
-55.3%
Blood Pb was a potential
moderator of the SES-TPR
relationship. Blood Pb x SES
interaction: p = 0.07 .
Blood Pb was a moderator of
SES-SBP relationship
Pb x SES interaction:
p = 0.007
At blood Pb levels >4 ug/dL,
SES not significantly
associated with SBP
November 2012
5-339
Draft - Do Not Cite or Quote
-------
Study
(Ordered as Study
they appear in Population/
the text) Methodology Parameters
Blood Pb Data3
Statistical Analysis Effect Estimates/Results
Gump et al.
(2009)
Prospective Salivary
122 children cortisol
age 9.5 yr in
Oswego, NY
Cord blood Pb:
GM (GSD):
2.56ug/dL(1.16)
Childhood (mean
age of measurement:
2.6 yr) blood Pb: GM
(GSD): 4.06 ug/dL
(1.14)
Linear regression to
examine whether blood
Pb level mediates or
moderates the
relationship between
SES and salivary
cortisol as in Gump et
al. (2007)
Blood Pb was a mediator of the
SES-cortisol association. SES
was no longer significantly
associated with cortisol after
adjusting for blood Pb level. R2
for SES decreased by 40, 33,
and 50% for cortisol measured
at 21, 40, and 60 min.
Blood Pb was not a significant
n-irtrlarotrt r r\f Q P Q_*^rt rtiort 1
association. Blood Pb x SES
interaction term was not
statistically significant
Gump et al.
(2011)
Cross-
sectional
140 children
ages 9-11 yr
Oswego, NY
SBP, TPR,
HRV (heart
rate
variability) in
response to
acute stress
(mirror
tracing task)
Concurrent blood Pb:
GM: 1.01 ug/dL
Quartiles:
Q1: 0.14-0.68 ug/dL
Q2: 0.69-0.93 ug/dL
Q3: 0.94-1.20 ug/dL
Q4: 1.21-3.76 ug/dL
Outcomes were
analyzed as continuous
variables for the pre-
stress values or the
change post- and pre-
stress. Regression
models were adjusted
for sex, SES, BMI, and
age.
Blood Pb levels associated
with autonomic and
cardiovascular dysregulation in
response to stress -greater
vascular resistance, reduced
stroke volume, and cardiac
output
Change in SBP (mmHg) across
quartiles: Q1: 5.30, Q2: 7.33,
Q3: 7.07, Q4: 7.23, p for
trend = 0.31
Change in TPR (%) across
quartiles: Q1: 2.91, Q2: 8.18,
Q3: 9.55, Q4: 9.51, p for
trend = 0.03
Change in Stroke Volume (%)
across quartiles: Q1: 2.23, Q2:
0.91, Q3:-3.47, Q4:-0.89, p
for trend = 0.04
Zhang et al.
(2011 a)
Prospective SBP
457 mother
child pairs in a
birth cohort,
born 1 994 to
2003 in
Mexico City.
Children were
evaluated
2008-201 0 at
ages 7-15 yr
Cord blood Pb:
GM (GSD):
4.67ug/dL(1.18)
(N=323)
Concurrent blood Pb:
GM (GSD):
2.56ug/dL(1.16)
(N=367)
Maternal post-
partum bone Pb:
Multiple regression
models and
generalized estimating
equations (log linear for
cord blood, linear for
concurrent blood and
maternal bone). The
base model considered
maternal education,
birth weight, BMI, sex,
and child concurrent
age as covariates.
Prenatal Pb exposure may be
associated with higher BP in
female offspring.
Among girls, an IQR (13 ug/g)
increase in maternal tibia Pb
was associated with a 2.11
(95% Cl: 0.69, 3.52) mmHg
increase in SBP
IQR (16 ug/g) increase in
maternal patella Pb was
associated with a 0.87 (95%
Cl: -0.75, 2.49) mmHg
increase in SBP
Median (IQR):
Tibia Pb: 9.3 (3.3,
16.1) ug/g
Patella Pb: 11.6(4.5,
19.9) ug/g
IQR (4 ug/dL) increase in cord
blood Pb was associated with
a 0.75 (95% Cl:-1.13, 2.63)
mmHg increase in SBP
Factor-
Litvak et al.
(1999: 1996)
Cross-
sectional
260 children
ages 5.5 years
old in
K. Mitrovica
and Pristina,
Yugoslavia
SBP
Concurrent blood Pb
range:
4.1 to 76.4 ug/dL
Linear regression
analysis. Potential
confounders
considered: sex,
maternal education,
birth weight, HOME
score, and BMI.
Per 1 ug/dL increase in
concurrent blood Pb level, 0.05
(95% Cl:-0.02, 0.13) mmHg
higher SBP
Blood Pb level at birth and
cumulative blood Pb level were
not as strongly associated with
SBP at age 5.5 yr.
November 2012
5-340
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
Study
(Ordered as Study
they appear in Population/
the text) Methodology Parameters
Gerretal. Cross- BP
(2002) sectional
508 young
adults age
19-29 years,
born
1965-1975,
male and
female; half of
the subjects
had grown up
around an
active Pb
smelter in
Silver Valley,
Idaho
aBlood Pb data are estimates of geometric mean
Blood Pb Data3
While the concurrent
mean blood Pb level
was 3.15 ug/dL for
the highest bone Pb
category (>10 ug/g),
early childhood
mean blood Pb
levels in this group
were substantially
elevated for all bone
Pb level categories
and were highest
among participants
in the highest bone
Pb level category.
The mean blood Pb
level was 65 ug/dL
among participants
with bone Pb level
>10 ug/g. Bone Pb
was measured at the
time of entry into this
study.
Statistical Analysis
Multiple linear
regression models
always included age,
sex, height, BMI,
current smoking status,
frequency of alcohol
consumption, current
use of birth-control
medication,
hemoglobin level,
serum albumin, and
income, regardless of
significance levels.
Both blood Pb (as a
linear term) and bone
Pb (a four category
ordinal variable from
<1 ug/g to >10 ug/g)
were tested together.
Effect Estimates/Results
Group in highest quartile of
tibia Pb level (>10 ug/g) had
4.26 (95% Cl: 1.36, 7.16)
mmHg higher SBP and 2.80
(95% Cl: 0.35, 5.25) mmHg
higher DBP compared to the
lowest tibia Pb group (<1 ug/g).
(GM) and geometric standard deviation (GSD) using the arithmetic mean and SD.
5.4.4.2 Cardiovascular Functioning in Children
The relationship between cardiovascular functioning (TPR, BP, stroke volume, and
cardiac output,) and blood Pb levels was examined prospectively by Gump et al. (2007;
2005) in a cohort born at a single New York hospital. Higher early childhood Pb levels
(average age 2.6 years) were associated with greater TPR response to acute stress induced
by mirror tracing on a computer at age 9.5 years as shown in Figure 5-24. Testing blood
Pb with linear, quadratic, and cubic terms did not produce significantly different Pb-TPR
associations, and the authors suggested that these effects were concentration-dependent
and notably, were not emergent at a specific exposure threshold. TPR increased with
increasing quartile of blood Pb level. A mediational analysis indicated that Pb was a
significant mediator of the SES-TPR reactivity association; some evidence also suggested
moderation, whereby the inclusion of blood Pb into the model reduced the effect estimate
for SES. Observations that Pb exposure increases TPR in toxicological studies and
mechanistic evidence indicating that Pb-induced changes in SNS activity may mediate
such effects (Section 5.4.2.3) provides some biological plausibility for a role of Pb in
affecting the TPR response to acute stress in this child population. Additionally, higher
blood Pb level measured at age 2.6 years was associated with a smaller stroke volume
and cardiac output responses to acute stress at age 9.5 years (Gump et al.. 2007). In a
further analysis in this cohort, Gump et al. (2009) examined the possibility that Pb may
mediate an association between SES and cortical responses to acute stress. Elevated
November 2012
5-341
Draft - Do Not Cite or Quote
-------
cortisol has been associated with hypertension (Whitworth et al., 2000). Gump et al.
(2009) found that lower family income was associated with greater cortisol levels
following an acute stress task and that blood Pb was a mediator of this association.
30
25
e
8 H3 20
c »
SS
ff$
'1 | 15
1 If 10
•S.3
II 5
Ji »
-5
-10
(
r-
-
» »
* » »*
* * 4 * * ^
•.^H^T^-
•S'..-'-. •
:*•
*
i i i i i i i
) 2 4 6 8 10 12 14
Blood lead level ({ig/dL)
Source: Reprinted with permission of Elsevier (Gump et al.. 2005)
Figure 5-24 Children's adjusted total peripheral resistance (dyn-s/cm5)
responses to acute stress tasks, as a function of childhood Pb
levels.
4
5
6
7
8
9
10
11
12
13
14
15
16
In a different cohort of 140 children 9 to 11 years of age recruited from local pediatrician
offices and from mailings to homes with children in this age group, Gump et al. (2011)
used a similar acute stress-producing paradigm as in previous studies to examine the
cross-sectional associations of concurrent blood Pb with cardiovascular responses. TPR
significantly increased in a concentration-dependent relationship with blood Pb, with
most of the increase occurring between the first quartile blood Pb (0.14-0.68 (ig/dL) and
the second quartile blood Pb (0.69-0.93 (ig/dL). This result is consistent with those of
Gump et al. (2005). Also, these newer findings provided evidence of associations with
concurrent blood Pb levels and with lower blood Pb levels (Gump et al.. 2011) than were
previously examined by Gump et al. (2005) and in a large group of children without
higher Pb exposures earlier in childhood.
Studies in adults and animals indicate Pb-associated decreases in HRV (Section 5.4.3.4).
In Gump et al. (2011). cardiac autonomic regulation decreased in a
November 2012
5-342
Draft - Do Not Cite or Quote
-------
1 concentration-dependent manner with increasing concurrent blood Pb quartile, with the
2 largest change relative to the first quartile (0.14-0.68 (ig/dL) measured in the highest
3 blood Pb quartile (1.21-3.76 (ig/dL). Also, high frequency HRV, decreased more with
4 acute stress in the highest Pb quartile group (1.21-3.76 (ig/dL). In the earlier cohort, early
5 childhood (mean age at collection: 2.6 years) blood Pb level was associated with reduced
6 stroke volume and cardiac output (Gump et al.. 2007; Gump et al.. 2005). In this recent
7 study, Gump et al. (2011) found the same but for concurrent blood Pb level and at lower
8 blood Pb levels.
5.4.4.3 Blood Pressure in Children
9 Zhang et al. (2011 a) conducted a longitudinal study that examined changes in BP in 323
10 girls and boys aged 7 to 15 years old in a Mexico City cohort and associations with
11 maternal bone Pb measured one month post-partum (a measure of cumulative exposure
12 that could expose fetuses to Pb through accelerated mobilization of bone Pb during
13 pregnancy) and with cord blood Pb at delivery. This was the first study to examine the
14 association of maternal bone Pb, as a marker of prenatal exposure, with offspring BP.
15 The model including both girls and boys (without adjustment for concurrent blood Pb)
16 showed no statistically significant association overall for any Pb biomarker with child
17 BP. A significant interaction was found between maternal tibia Pb and sex, and in models
18 stratified by sex, maternal tibia Pb was associated with adjusted systolic and diastolic BP
19 in females, but not males. Maternal post-partum median tibia Pb was 9.3 (ig/g (IQR: 3.3,
20 16.1 (ig/g) with no significant differences between mothers of male and female offspring.
21 Suboptimal growth in utero is associated with accelerated weight gain in offspring during
22 childhood and greater risk of later hypertension (Barker and Bagby. 2005; te Velde et al..
23 2004; Barker et al.. 1989). The relationship between birth weight and Pb biomarkers is
24 discussed in Section 5.8.3. These may represent biologically plausible mechanisms by
25 which prenatal Pb exposure may result in increased BP later in childhood as was
26 demonstrated in female offspring.
27 Gump et al. (2011; 2005) examined the relationship of blood Pb level with BP in their
28 two cohorts of contemporary children around age 10 years in New York State. Gump et
29 al. (2005) reported an association of cord blood levels with systolic BP (12.16 mmHg
30 [95% CI: 2.44, 21.88] increase per 1 (ig/dL increase in cord blood Pb level). Gump et al.
31 (2011) found that with acute stress, children in higher quartiles of concurrent blood Pb
32 level (>0.69 (ig/dL) had larger increases in systolic BP. For example, children with blood
33 Pb levels between 1.21 and 3.76 (ig/dL had a 7.23 mmHg change, and children with
34 blood Pb levels between 0.14 and 0.68 (ig/dL had a 5.30 mmHg change. A linear trend
35 was not observed across quartiles. An interaction between long-term perceived stress and
November 2012 5-343 Draft - Do Not Cite or Quote
-------
1 bone Pb levels in association with BP and hypertension also was reported in a study of
2 adults (Peters et al.. 2007) (described in Section 5.4.2.1). An earlier study (Factor-Litvak
3 et al., 1999; Factor-Litvak et al.. 1996) of children with higher blood Pb levels ranging
4 from 4.1 to 76.4 (ig/dL found that a 1 (ig/dL increase in concurrent blood Pb was
5 associated with a 0.05 (95% CI: -0.02, 0.13) mmHg increase in systolic BP. An
6 additional study (Gerr et al.. 2002) reported that systolic BP for young adults (ages 19-29
7 years) with bone Pb levels greater than 10 (ig/g (mean concurrent blood Pb = 65 (ig/dL)
8 was 4.26 mmHg higher compared with young adults with bone Pb levels <1 (ig/dL
9 compared to young adults with bone Pb levels <1 (ig/dL.
10 The pathogenesis of CVD has been hypothesized to begin in childhood (Kapuku et al..
11 2006). Early markers observable in youth in association with Pb biomarkers include
12 increased BP during stress, reduced HRV, increased IMT, and vascular endothelium
13 dysfunction. Kapuku et al. (2006) state that endothelial dysfunction is the center of the
14 CVD paradigm. The factors measured in childhood or as a cumulative burden since
15 childhood are predictors of outcomes in young adults who are still too young to
16 experience coronary events (Li et al.. 2003). and early-life exposures may induce changes
17 in arteries that contribute to the development of atherosclerosis (Raitakari et al.. 2003).
18 Berenson et al. (2002) observed that the effects of multiple risk factors on coronary
19 atherosclerosis support evaluation of cardiovascular risk in young people. Thus, evidence
20 relating levels of biomarkers of Pb exposure in children to cardiovascular function in the
21 groups of studies presented in the preceding text when combined with the evidence for
22 the potential pathogenesis of CVD starting in childhood that yield effects in adulthood,
23 provides coherence with evidence in adults supporting the effects of long-term,
24 cumulative Pb exposures in the development of cardiovascular effects.
25 Few animal studies have examined the effect of Pb exposure during pregnancy and
26 lactation on BP in offspring as adults and those that have used high levels of exposure.
27 Recently, pups of Pb-exposed dams (1,000 ppm through pregnancy and lactation)
28 exhibited increased blood Pb level (mean blood Pb level: 58.7 (ig/dL) and increased
29 arterial systolic BP after weaning (Grizzo and Cordellini. 2008) suggesting a role for
30 childhood Pb exposure leading to adult disease.
5.4.4.4 Summary of Child Cardiovascular Studies
31 The 2006 Pb AQCD (U.S. EPA. 2006b) described three studies on the effects of Pb on
32 cardiovascular function in children; however, no conclusions were made as to the
33 strength of the evidence. Studies have reported antecedent cardiovascular changes such as
34 TPR responses to acute stress tasks as a function of childhood blood Pb levels. Also, a
November 2012 5-344 Draft - Do Not Cite or Quote
-------
1 study reported associations with acute stress-induced autonomic and cardiovascular
2 dysregulation responses. Biomarkers of prenatal Pb exposure (maternal post-partum
3 patella and tibia Pb levels) were related to later higher BP. Other lines of evidence have
4 linked increased intrauterine growth restriction to later accelerated weight gain in
5 childhood, and this may indicate greater risk of hypertension later in life. The results are
6 not uniform with respect to the important lifestages of Pb exposure and can differ by sex
7 and other factors. Uncertainties in these studies may be related to sample size, single
8 measures of BP, variation in the age of onset of puberty, and cross-sectional design.
9 However, some of these uncertainties may result in the attenuation of observed
10 associations rather than the generation of spurious associations. Overall, recent study
11 findings indicate that in children with mean blood Pb levels in the range of <10 (ig/dL,
12 increasing blood Pb level may be associated with small increases in BP and changes in
13 the cardiovascular system that may be related to later development of CVD.
14 Factors may limit the ability of studies to detect statistically significant Pb-associated
15 changes with BP. The relatively young age of the subjects may have limited the ability of
16 these studies to detect significant BP effects (as opposed to early function effects) if
17 longer duration Pb exposure is necessary to produce the cardiovascular changes
18 considering the lower prevalence and strength of compensatory mechanisms in children.
19 There is uncertainty in the shape of the concentration-response relationship to
20 cardiovascular endpoints at lower blood Pb levels since most studies modeled a linear
21 relationship. A nonlinear concentration-response relationship has been found for Pb with
22 other outcomes in children, most notably, decrements in cognitive function (See
23 Section 5.3.2).
24 Cardiovascular endpoints other than baseline BP may be more sensitive outcomes for
25 measuring Pb-associated cardiovascular effects in very young children. The series of
26 studies by Gump et al. (2011; 2009; 2007; 2005) evaluating much smaller samples than
27 did the adult studies, was able to demonstrate statistically significant relationships of
28 blood Pb levels with cardiovascular responses such as TPR, related to acute stress. These
29 results suggest that the stress paradigm may be useful to detect associations of blood Pb
30 levels with effects on the cardiovascular system of children. Selection of the appropriate
31 cardiovascular outcome in children is an important factor to consider in the design of
32 future studies. Rather than using indicators of cardiovascular effects, such as BP,
33 evaluation of cardiovascular changes that are antecedent to increased BP and changes in
34 other CVD-related endpoints that present at later lifestages be informative to
35 understanding the time course of cardiovascular changes that may be associated with
36 early Pb exposure.
November 2012 5-345 Draft - Do Not Cite or Quote
-------
1 Overall this small body of evidence, based on different cohorts, locations, and study
2 designs, begins to form a literature base that suggests a relationship between biomarkers
3 of Pb exposure and cardiovascular effects in children. One longitudinal study ties in
4 maternal bone Pb level, and cord and concurrent blood Pb level for the children.
5 Limitations exist in the studies. While blood pressure increases are more prevalent in
6 older adults than in children, BP increases have been related to higher blood Pb level in
7 earlier studies of children and young adults (Gerr et al., 2002; Factor-Litvak et al., 1999;
8 Factor-Litvak et al.. 1996). The recent Gump studies of children provide information in
9 populations with mean blood Pb levels in the range of <10 (ig/dL for BP and potential
10 antecedents for CVD such as increases in TPR and changes in cardiac autonomic
11 regulation.
5.4.5 Mortality
12 The 2006 Pb AQCD (U.S. EPA. 2006b) stated that available evidence suggested an effect
13 of Pb on cardiovascular mortality in the general U.S. population but cautioned that these
14 findings should be replicated before these estimates for Pb-induced cardiovascular
15 mortality could be used for quantitative risk assessment purposes (U.S. EPA, 2006b).
16 Previous results involved NHANES II and III analyses that examined prospectively the
17 association of adult blood Pb measured at the time of the study with all cause and cause-
18 specific mortality ascertained 8-16 years later (Schober et al.. 2006; Lustberg and
19 Silbergeld. 2002). As blood Pb levels in adults reflect contributions from both recent Pb
20 exposure and mobilization of historic Pb from bone, it is unclear to what extent recent,
21 past, or cumulative Pb exposures contributed to the observed associations. Given the
22 decline in ambient air Pb concentrations and population blood Pb levels, it is likely that
23 study subjects had a much higher past Pb exposure compared to exposure during the
24 study period. Using NHANES II (1976-1980) data, Lustberg and Silbergeld (2002) found
25 significant increases in all-cause mortality, circulatory mortality, and cancer mortality,
26 comparing adults with blood Pb levels of 20-29 (ig/dL to those with blood Pb levels less
27 than 10 (ig/dL (measured 12-16 years before ascertainment of vital status). Using
28 NHANES III data, Schober et al. (2006) found significant increased all-cause,
29 cardiovascular, and cancer mortality comparing adults with blood Pb levels
30 from 5-9 (ig/dL and above 10 (ig/dL compared to those with blood Pb levels less than
31 5 (ig/dL (measured a median of 8.8 years before ascertainment of vital status).
32 Several recent studies substantially strengthen the evidence base for Pb-associated
33 mortality. A further analysis of the NHANES III database by a different research group
34 using different methods addressed uncertainties from earlier analyses by considering a
35 greater number of potential confounding factors and by characterizing concentration-
November 2012 5-346 Draft - Do Not Cite or Quote
-------
1 response relationships. Additionally, two longitudinal prospective studies in different
2 U.S. cohorts conducted by different researchers with different methods demonstrate
3 consistency within the evidence base for blood Pb and add new evidence for mortality
4 associated with bone Pb levels.
5 Menke et al. (2006) examined all-cause and cause-specific mortality using NHANES III
6 data. Subjects at least 18 years of age were followed up to 12 years after their blood Pb
7 was measured, and 1,661 deaths were identified. Those with baseline blood Pb levels
8 from 3.63 to 10 (ig/dL had significantly higher risks of all-cause (HR: 1.25 [95% CI:
9 1.04, 1.51]), cardiovascular (HR: 1.55 [95% CI: 1.08, 2.24]), MI (HR: 1.89 [95% CI:
10 1.04, 3.43]), and stroke (HR: 2.51 [95% CI: 1.20, 2.26]) mortality compared to those with
11 baseline blood Pb levels less than 1.93 (ig/dL and increased risk of cancer mortality (HR:
12 1.10 [95% CI: 0.82, 1.47]). Effect estimates adjusted for demographic characteristics
13 were robust to the additional adjustment for factors such as smoking, alcohol
14 consumption, diabetes, BMI, hypertension, and level of kidney function. The consistency
15 of HRs across models with a varying number of control variables indicated little residual
16 confounding. Hazard ratios were not higher comparing adults with blood Pb levels from
17 1.94 to 3.62 (ig/dL to those with blood Pb levels <1.93 (ig/dL. However, tests for linear
18 trend were statistically significant for all mortality outcomes except for cancer mortality.
19 Menke et al. (2006) evaluated several of the model covariates (e.g., diabetes,
20 hypertension, and glomerular filtration rate [GFR]) in a subgroup analysis. The
21 comparisons for these are shown in Figure 5-25. The authors reported that there were no
22 interactions between blood Pb and other adjusted variables.
23 The results from Menke et al. (2006) generally were consistent with those from the
24 previous NHANES III analysis of the association of blood Pb with mortality by Schober
25 et al. (2006) that included participants greater than 40 years of age (N = 9686) and
26 adjusted for covariates including age, sex, ethnicity, and smoking rather than the full suite
27 of covariates evaluated by Menke et al. (2006). Schober et al. (2006). which was
28 discussed in the 2006 Pb AQCD (U.S. EPA. 2006b). reported increased HRs comparing
29 adults with blood Pb levels > 10 (ig/dL to those with blood Pb levels <5 (ig/dL for all-
30 cause (HR: 1.59 [95% CI: 1.28, 1.98]), CVD (HR: 1.55 [95% CI: 1.16, 2.07]), and cancer
31 (HR: 1.69 [95% CI: 1.14, 2.52]) mortality. In general, HRs were higher but
32 nonsignificant, comparing adults with blood Pb levels from 5-9 (ig/dL to those with
33 blood Pb levels <5 (ig/dL. The median follow-up time between measurement of blood Pb
34 and death ascertainment was 8.55 years.
November 2012 5-347 Draft - Do Not Cite or Quote
-------
Subgroup
Hazard ratio of all-cause mortality (95% C!)
Hazard ratio of cardiovascular mortality (95% Ci)
Age (years)
< 60
>=60
Race-ethnicity
Non-Hispanic white
Non-Hispanic black
Mexican- American
Sex and menopausai status
Male
Female
Pre-menopausai
Post-menopausai
Residence
Rural
Urban
Smoking
Never
Former
Current
Body mass index (kg/m2)
<25
>=25
Hypertension
No
Yes
Diabetes
No
Yes
Estimated glomerular filtration
rate (ml/mi n/1.73m2)
< 60
>=60
Overall
1 ,75 (1.25
1.31 (1.08
1.32(1.09
1 .23 (0,99
1.17(0.86
1.41 (1.11
1.24(1.00
1 .02 (0.54
1.24(1.00
1.28(1.05
1.42(1.18
1.21 (0.93
1.61 (1.33
1 .34 (0.96
1.51 (1.16
1.28(1.03
1.31 (1.08
1.32(1.09
1.37(1.19
1.12(0.73
1 .44 (1.01
1.32(1.12
1.34(1.16
2.44)
1,58)
1.60)
1.52)
1.60) —
1.78)
1.54)
1 .95)
1.54)
1.54)
1.72)
1 .58)
1.94)
1 .87)
1.96)
1.58)
1,58)
1.60)
1.58)
1.71)
2.06)
1,56)
1,54)
—4—
— ^ —
— •-! —
I
!•
— *r~
— »j—
— •; —
— !• —
— »-^ —
i~*
— 1_ • —
— * —
— * —
— ^ —
p
— * — ;
-V
+-
0,5 1 23
1,49(1.12
1,490.12
1.63(1.25
1.49(1.15
1.59(1.31
1.49(1.18
1.53(1.21
1.99)
1,99)
2.11)
1,94)
1.92)
1.89)
1.94)
m
•
_™_fl
•
•^
—
*
•—
—
*-
0,5
Note: Hazard ratios were calculated for a 3.4 ug/dL increase in blood Pb level with log-blood Pb as a continuous variable. This
increase corresponds to the difference between the 80th and 20th percentiles of the blood Pb distribution (4.92 ug/dL versus
1.46 ug/dL, respectively).
Source: Reprinted with permission of Lippincott Williams & Wilkins, Menke et al. (2006)
Figure 5-25 Multivariate adjusted relative hazards of all-cause and
cardiovascular mortality per 3.4 ug/dL increase in blood Pb.
1 Both Menke et al. (2006) and Schober et al. (2006) presented mortality curves that plot
2 the HRs against blood Pb level. Figure 5-26 shows the mortality hazard ratio curves (not
3 absolute cases of mortality) for both stroke and MI reported by Menke et al. (2006).
4 Nonlinear associations were modeled. The curves were fitted using predetermined
5 restricted quadratic splines with knots at the 10th percentile (1.00 (ig/dL), the 50th
6 percentile (2.67 (ig/dL), and the 80th percentile (5.98 (ig/dL) blood Pb levels. The
7 authors did not explain the shape of the blood Pb-mortality curves in detail; however, the
8 knots corresponded with the inflection points in the curve. In the tails of the blood Pb
9 distribution, hazard ratios decreased with increasing blood Pb level. However, hazard
10 ratios remained above 1 over most of the blood Pb distribution (blood Pb level greater
11 than 2 (ig/dL and between 2 and 3 (ig/dL for stroke and myocardial infarction,
12 respectively), and in the most heavily populated portion of the blood Pb distribution,
13 hazard ratios increased with increasing blood Pb level. Using a referent group of persons
14 with blood Pb level less than 1.94 (ig/dL, the hazard ratio for persons with blood Pb level
November 2012
5-348
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
greater than 3.63 (ig/dL was significant at the 5% level 1.51 (1.07-2.14), but not
significant for persons with blood Pb level in the range of 1.94 to 3.62 (ig/dL. Hazard
ratios peaked for all outcomes at a blood Pb level of approximately 6 (ig/dL. Lower
concentration-response functions at higher blood Pb levels also have been found for
blood Pb-cognitive function relationships in children (Section 5.3.2).
2.0 1
— 1.5
O
•**
TO
B
I
1.0-
All cause
Myocardial infarction
Stroke
Cancer
0.7
12 5 10
Blood Lead, pg/dL
Source: Reprinted with permission of Lippincott Williams & Wilkins, Menke et al. (2006)
Note: A histogram of blood Pb levels is superimposed in the background and displayed on the right axis.
Figure 5-26 Multivariate-adjusted relative hazard (left axis) of mortality
associated with blood Pb levels between 1 ug/dL and 10 ug/dL.
6 Schober et al. (2006) examined proportional hazard assumptions, tested for a linear trend
7 across blood Pb tertiles, and evaluated log-transformed continuous blood Pb level as
8 a 5-knot cubic spline (position of knots not reported). A statistically significant increasing
9 linear trend for mortality was observed across blood Pb tertiles. The results of the spline
10 fit of the continuous blood Pb level term to relative hazard of all cardiovascular diseases
11 reported by Schober et al. (2006) are shown in Figure 5-27. Schober et al. (2006) shows
12 the upper 95% confidence band (dashed lines) of the relative risk for all cause mortality
13 spline is greater than 1 for all blood Pb levels greater than 1.5 (ig/dL using the referent
14 group of persons with blood Pb levels less than 1.5. The hazard ratio was fixed at 1.0 for
15 the referent blood Pb level of 1.5 (ig/dL. Also, the lower 95% confidence band is greater
November 2012
5-349
Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
than 1 when the blood Pb levels is greater than about 4.5 (ig/dL. Using a referent group of
persons with blood Pb levels less than 5 (ig/dL, they found statistically significant
relative risks of CVD for persons with blood Pb levels in the range of 5 to 9, and those
with blood Pb levels greater than 10. In contrast to the curve presented by Menke et al.
(2006). Schober et al. (2006) found the relative hazard axis and the blood Pb axis largely
to be linear (solid line). Both Menke et al. (2006) and Schober et al. (2006) agree that
persons with blood Pb levels greater than 4.5 (ig/dL are at increased risk for mortality;
however, these studies report different shapes for the concentration-response curves.
Despite differences in the age groups included, follow-up time, categorization of blood
Pb levels, and differences in hazard ratio across the blood Pb range, results reported by
Menke et al. (2006) and Schober et al. (2006) are find associations between higher blood
Pb and increased CVD mortality (see Figure 5-30).
2.0
1.8
1.6
•21.4
J 1.2
g 1.0
J 0.8
K 0.6
0.4
0.2
0.0
1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0
Blood lead (ug/dL)
Source: Schober etal. (2006)
Note: The solid line shows the fitted five-knot spline relationship; the dashed lines are the point-wise upper and lower 95% CIs.
Figure 5-27 Relative risk of all cause mortality for different blood Pb levels
compared with referent level of 1.5 ug/dL (12.5th percentile).
13 In addition to the NHANES analyses described above, studies of older adult, primarily
14 white, males (Weisskopf etal.. 2009) and older adult females (Khalil. 2010; Khalil et al..
15 2009b) were conducted recently. Weisskopf et al. (2009) used data from the NAS to
16 determine the associations of blood, tibia, and patella Pb with mortality. The authors
17 identified 241 deaths over an average observation period of 8.9 years (7,673 person-
18 years). The strongest associations were observed between mortality and baseline patella
November 2012
Draft - Do Not Cite or Quote
-------
1 Pb concentration. Baseline tibia Pb levels were more weakly associated with CVD
2 mortality. Tibia bone Pb level is thought to reflect a longer cumulative exposure period
3 than is patella bone Pb level because the residence time of Pb in trabecular bone is shorter
4 than that in cortical bone. IHD contributed most to the relationship between patella Pb
5 and all CVD death with an individual HR of 2.69 (95% CI: 1.42, 5.08). Although there
6 was high correlation between tibia and patella Pb (Pearson r = 0.77), compared with
7 cortical bone Pb, trabecular bone Pb may have more influence on circulating blood Pb
8 level and thus, local organ concentration of Pb because of its shorter residence time in
9 bone. In contrast to the NHANES analyses, the NAS study found that baseline blood Pb
10 was not significantly related to cardiovascular mortality. This discrepancy may be related
11 to differences in sample size and resulting power, modeling strategies (e.g., linear versus
12 log-linear blood Pb level terms), or age range of the study populations. The duration of
13 follow-up was similar across studies. In the Weisskopf et al. (2009) study of NAS data,
14 the youngest subjects at baseline were approximately 50-55 years old, compared to the
15 youngest in the Menke et al. (2006) and Schober et al. (2006) NHANES studies, who
16 were 18 and 40 years, respectively. Further, the blood Pb tertile analysis of Weisskopf
17 et al. (2009) could have been affected if the majority of a hypothesized nonlinear
18 association was contained largely in the lowest (reference) blood Pb tertile.
19 Weisskopf et al. (2009) also conducted a concentration-response analysis. A linear trend
20 was observed for increasing HR across tertiles of both tibia and patella Pb levels. The
21 linear relationship using tertile patella Pb was confirmed in other models in which
22 continuous patella Pb and nonlinear penalized spline terms (higher order terms) were not
23 statistically significant. The number of knots and their placement within the Pb variable,
24 which can influence these results, were determined by an iterative best fit procedure.
25 Concentration-response relationships shown in Figure 5-28 were approximately linear for
26 patella Pb on the log HR scale for all CVD, but appeared nonlinear for IHD (p <0.10).
27 The peak HR is shown around 60 (ig/g, beyond which the HR tends to decrease. It is
28 important to note the wide confidence limits, which increase uncertainty at the lower and
29 upper bounds of patella Pb levels.
November 2012 5-351 Draft - Do Not Cite or Quote
-------
-------
1 The association of adult blood Pb with mortality has also been examined among women
2 enrolled in the Study of Osteoporotic Fractures (SOF) (Khalil et al. 2009b). This
3 prospective cohort (N = 533) enrolled female volunteers (age 65-87 years) from two U.S.
4 locations, Baltimore, MD and Monongahela Valley, PA and followed women for an
5 average of 12 years after blood Pb measurement. All-cause mortality is significantly
6 higher comparing women with blood Pb levels >8 (ig/dL to those with blood Pb levels
7 <8 (ig/dL (HR: 1.59 [95% CI: 1.02, 2.49]). Hazard ratios for combined cardiovascular
8 disease mortality (HR: 1.78 [95% CI: 0.92, 3.45]), coronary heart disease mortality (HR:
9 3.08 [95% CI: 1.23, 7.70]), but not stroke mortality (HR: 1.13 [95% CI: 0.34, 3.81]) were
10 higher among the women enrolled in this study with blood Pb levels >8 (ig/dL. In
11 addition, analyses of blood Pb tertiles and quintiles indicated that blood Pb-mortality HRs
12 were consistently elevated in groups with blood Pb levels >7 (ig/dL (Khalil 2010). The
13 findings for elevated mortality HRs with the highest blood Pb levels are reinforced by the
14 results displayed in Figure 5-29. The HR curve for all-cause mortality is relatively flat
15 over most of population blood Pb distribution (represented by the blue dots) and
16 increases only in the upper tail of the blood Pb distribution where there are relatively few
17 subj ects (i. e., fewer dots).
18 Other studies also reported Pb-associated increased in mortality but have limited
19 implications due to their weaker analytic methods. Two studies reported standardized
20 mortality ratios (SMR) to compare observed deaths in a Pb-exposed population versus
21 expected deaths, calculated from a reference group (Neuberger et al.. 2009; Cocco et al..
22 2007). Mortality studies that compare populations by calculating SMRs based on an
23 "exposed group" versus the population within which the exposed group resides have
24 several drawbacks, including the ecologic nature of the analysis and the absence of Pb
25 exposure data or biological markers of Pb exposure. Neuberger et al. (2009)carried out a
26 retrospective mortality study of a Superfund site that was highly contaminated with heavy
27 metals, principally Zn, Pb, and Cd. Not knowing the metal concentrations in the
28 population obscures interpretation of the significantly elevated county-state SMRs and
29 the insignificant or significantly lowered SMRs in the county comparison.
30 A retrospective study of causes of death among Pb smelter workers in Sardinia, Italy
31 followed 933 male production and maintenance workers (Cocco et al.. 2007). SMRs for
32 cardiovascular disease-related deaths were calculated based on age-specific and calendar-
33 year specific mortality of the entire region. Significantly reduced mortality was reported
34 in the worker groups. The authors attributed the results to the healthy worker effect based
35 on health criteria applied at hiring and the small size of the cohort. The usual caveats
36 regarding population comparison mortality studies apply.
November 2012 5-353 Draft - Do Not Cite or Quote
-------
00 -
o -
5 10
LEAD VALUE
15
relative hazard
Median spline
Source: Khalil et al. (2010)
Figure 5-29 Multivariate adjusted relative hazard (left axis) of mortality as a
function of blood Pb levels between 1 ug/dL and 15 ug/dL.
i
2
o
6
4
5
6
7
8
9
10
11
5.4.5.1 Summary of Mortality
The mortality results in this review supported and expanded upon findings from the
2006 Pb AQCD (U.S. EPA. 2006b). which included a few NHANES mortality studies
(Schober et al., 2006; Lustberg and Silbergeld. 2002). The recent NHANES mortality
study discussed above (Menke et al.. 2006) addressed many of the limitations of the
earlier studies, including control for a wider range of potential confounders, testing for
interactions with Pb, consideration of concentration-response relationships, extensive
model evaluations, and examination of mortality from specific CVDs. Further, an
association with increased mortality was observed at lower mean population blood Pb
levels. The mean blood Pb level of the NHANES III population was 2.58 (ig/dL. In the
recent analysis, the Pb risk of increased cardiovascular mortality increased with
increasing blood Pb level over the most heavily populated portion of the blood Pb
November 2012
Draft - Do Not Cite or Quote
-------
1 distribution, with maximum blood Pb levels between 6 and 7 (ig/dL. It is important to
2 note that the relative contributions of recent, past, and cumulative Pb exposure to
3 associations observed with the baseline blood Pb levels is uncertain. In addition, the first
4 evidence that bone Pb, a metric of cumulative Pb exposure, is associated with increased
5 mortality was reported recently among NAS men (Weisskopf et al.. 2009).
6 Quantitative differences in Pb-associated hazard for death between studies may be
7 influenced by age range of the study groups, follow up time to death, variation in model
8 adjustment, central tendency and range of the Pb biomarker levels, assumptions of
9 linearity in relationship with Pb biomarkers, and choice of Pb biomarker. Quantitative
10 differences in Pb-associated mortality across NHANES II and NHANES III studies or
11 between different NHANES III analyses may be explained by the use of continuous or
12 ordered blood Pb terms and different data selection strategies. Further, studies using
13 ordered categories of blood Pb level may obtain different results, as the range of blood Pb
14 level represented in the reference category will affect the calculated coefficients of the
15 remaining percentiles or groups.
16 Specifically, Menke et al. (2006) is the strongest study presently published for estimating
17 the effects of Pb on cardiovascular disease-related mortality. The study uses the
18 nationally representative NHANES III (1988-1994) sample of men and women. The
19 results corroborate of earlier published NHANES studies but address some of the key
20 weaknesses noted in those studies. For example, Menke et al. (2006) examined potential
21 confounding by a large number of factors, including hypertension and kidney function.
22 Weisskopf et al. (2009) is the first published mortality study using bone Pb as an
23 exposure index. The study is a prospective study with nearly 100% successful follow-up
24 of deaths. This rigorous study found increased cardiovascular disease mortality in
25 association with patella bone Pb with weaker associations for tibia Pb level. The Khalil
26 et al. (2010; 2009b) study of SOF subjects provides supporting results for a cohort
27 consisting of white females aged 65-87 years. Further, a number of prior studies found
28 associations between accumulated Pb reflected in bone Pb measurements and higher
29 CVD morbidity (Sections 5.4.2.1 and 5.4.3). This evidence base is augmented with new
30 findings indicating that biomarkers of longer-term cumulative Pb exposure increases
31 CVD mortality. The NAS and SOF examine only men and women, respectively.
32 However, the consistency of findings between the two studies indicates that the results of
33 either study may be applicable widely. Despite the differences in design and methods
34 across studies, associations between higher levels of Pb biomarkers and higher risk of
35 mortality were generally observed (Figure 5-30 and Table 5-23). One exception is that
36 stroke mortality was not significantly elevated in the SOF study although it was positive.
37 Mortality from specific CVD causes, MI and IHD mortality, which are related to higher
38 BP and hypertension, were elevated with higher Pb biomarker levels.
November 2012 5-355 Draft - Do Not Cite or Quote
-------
Reference
Menkeetal. (2006)
n = 13,946
mn age=44
mn blood Pb=2.58
Schoberetal. (2006)
n = 9757
age > 40 y
Lustberg & Silbergeld
[2002]; n = 4,190
mn age=54 y
mn blood Pb=14,0
Weisskopf etal. (2009)
n = 868 men
mn age-67.3 y
mn blood Pb 5.7
Khalil etal. (2009)
n - 533 women
rnn age=70 y
mn blood Pb=5.3
Outcome
All Cause
CVD
Ml
Stroke
All Cause
CVD
All Cause
CVD
All Cause
CVD
IHD
All Cause
CVD
IHD
All Cause
CVD
IHD
All Cause
CVD
CHD
Stroke
Study Pb Biomarker Comparison
Groups
Blood Pb (ug/dL)
NHANESIII 23,63 vs. S1.93
1.94-3.62 vs. S1.93
>3.63 vs. S1.93
1.94-3.62 vs. <1.93
>3.63vs. S1.93
1.94-3.62 vs. Sl.93
£3.63 vs. £1.93
1.94-3.62 vs. <1.93
>10vs. <5
5-9 vs. < 5
HO vs. <5
5-9 vs. < 5
NHANESII 20-29 vs. < 10
10-19 vs. < 10
20-29 vs. < 10
10-19 vs. < 10
NAS
SOF
>6vs. < 4
4-6 vs. < 4
>6 vs. < 4
4-6 vs. < 4
>6 vs. < 4
4-6 vs. < 4
Tibia Pb (M«/g)
Tertile 3 vs. 1 (NR)
Tertile 3 vs. 1 (NR)
Tertile 3 vs. 1 (NR)
Patella Pb (u.g/g)
> 35 vs. < 22
22-35 vs. < 22
> 35 vs. < 22
22-35 vs. < 22
> 35 vs. < 22
22-35 vs. < 22
Z 8 vs. < 8
> 8 vs. < 8
1986-1988 a 8 vs. < 8
2 8 vs. < 8
0.0 1.0 2.0 3.0 4.0 5.0 6.0 7.0 8.0 9.0 10.0
Hazard Ratio (95% Cl)
Note: Studies are presented in order of strength of study design and follow the order of discussion in the preceding text. Hazard
ratios represent the hazard in the higher blood or bone Pb group relative to that in the lowest blood or bone Pb group (reference
group).
Blood Pb (closed markers), or Bone Pb (open markers) associations with All-cause mortality (black diamonds) or
Cardiovascular mortality (blue circles).
Figure 5-30 Hazard ratios for associations of blood Pb or bone Pb with
all-cause mortality and cardiovascular mortality.
November 2012
Draft - Do Not Cite or Quote
-------
Table 5-23 Additional characteristics and quantitative data for associations of
blood and bone Pb with CVD mortality for studies presented in
Figure 5-30.
Study
Menke et al.
(2006)
Schober et
al. (2006)
Lustberg
and
Silbergeld
(2002)
Study
Population /
Methodology
Longitudinal
13,946 adult
participants of
NHANES III ,
> 17yr
(1988-1994)
Longitudinal
9,686 adult
participants of
NHANES III,
^ 40 yr
Longitudinal
4,190 adult
participants of
NHANES III,
yr (1976-1 980)
aged 30 to 74,
Studied
through
December 31,
1992
Parameter
All cause and cause-
specific mortality
Studied through
December 31, 2000
CVD:ICD-9 390-434;
ICD-10 IOO-I99), Ml
(ICD-9410-414and
429.2; ICD-10 I20-I25),
stroke (ICD-9 430-434
and 436-438; ICD-10
I60-I69).
All cause and cause-
specific mortality
All cause and cause-
specific mortality
Pb Data
Baseline
Blood Pb
(measured an
average of 12
yr before
mortality):
Mean:
2.58 ug/dL
Tertiles:
<1.93ug/dL,
1 .94-3.62 ug/d
L,
> 3.63 ug/dL
Ordered
categorical
blood Pb level,
measured a
median of 8.55
yr prior to
death
<5 ug/dL
5-9 ug/dL
> 10 ug/dL
Categorical
blood Pb level
Mean: 14.0
(5.1)
Msdisrr
13 ug/dL
Isttertile:
<10 ug/dL
(Reference)
2nd tertile:
10-19ug/dL
3rd tertile:
20-29 ug/dL
Statistical Analysis
Survey-design adjusted Cox
proportional hazard
regression analysis (up to 12
yr follow-up) adjusted for
Model 1 : age, race/ethnicity,
sex, Model 2: urban
residence, cigarette smoking,
alcohol consumption,
education, physical activity,
household income,
menopausal status, BMI,
CRP, total cholesterol,
diabetes mellitus, Model 3:
hypertension, GFR category
Survey-design adjusted Cox
proportional hazard adjusted
for sex, age, race/ethnicity,
smoking, education level Did
not evaluate BMI nor
cormorbidities
Proportional Hazard model,
RRs adjusted for age, sex,
location, education, race,
income, smoking, BMI,
exercise
Hazard Ratio or
SMR (95% Cl)
All-cause (3rd vs.
Isttertile):
1.25(1.04, 1.51)
CVD (3rd vs. 1st):
1 .55 (1 .08, 2.24)
Ml (3rd vs. 1st):
1 .89 (1 .04, 3.43)
Stroke (3rd vs. 1st):
2.51 (1.20,5.26)
Cancer (3rd vs.
1st):
1.10(0.82, 1.47)
All-cause (2nd vs.
1st):
1 .24 (1 .05, 1 .48)
All-cause (3rd vs.
1st):
1.59(1.28, 1.98)
CVD (2nd vs. 1st):
1.20(0.93, 1.55)
CVD (3rd vs. 1st):
1.55(1.16,2.07)
Cancer (2nd vs.
1st):
1.44(1.12, 1.86)
Cancer (3rd vs.
1st):
1.69(1.14,2.52)
All-cause (2nd vs.
1st):
1.40(1.16-1.69)
All-cause (3rd vs.
1st)'
i 01;.
2.02 (1 .62-2.52)
Circulatory (2nd vs.
1st):
1 .27 (0.97-1 .57)
Circulatory (3rd vs.
1st):
1 .74 (1 .25-2.40)
Cancer (2nd vs.
1st):
1 .95 (1 .28-2.98)
Cancer (3rd vs.
1st):
2.89 (1 .79-4.64)
November 2012
Draft - Do Not Cite or Quote
-------
Study
Weisskopf et
al. (2009)
Khalil et al.
(2QQ9b)
Study
Population /
Methodology Parameter
Longitudinal All cause and cause-
868 men, >55 specific mortality
yr, 95% white,
from MAS in
Greater
Boston
area, MA
Longitudinal All cause and cause-
533 women, specific mortality
65-87 yr, from
Study of
Osteoporotic
Fractures
cohort in
Baltimore, MD
and
Monongahela
Valley, PA
Pb Data
Pb biomarkers
collected an
average of 8.9
years before
death
Blood Pb:
Mean (SD):
5.6 (3.4) ug/dL
Patella Pb:
Mean (SD):
31.2
(19.4) ug/g
Tertiles:
<22 ug/g,
22-35 ug/g,
>35 ug/g
Tibia Pb:
Mean (SD):
21.8
(13.6) ug/g
Blood Pb
measured an
average 12
(SD; 3) yr
before death:
Mean (SD;
range): 5.3
(2.3;
1-21)ug/dL
Statistical Analysis
Cox proportional hazard
regression analysis adjusted
forage, smoking, education.
Additional models adjusted
for alcohol intake, physical
activity, BMI, total cholesterol,
serum HDL, diabetes
mellitus, race, and
hypertension
Cox proportional hazards
regression analysis adjusted
forage, clinic, BMI,
education, smoking, alcohol
intake, estrogen use,
hypertension, total hip bone
mineral density, walking for
exercise, and diabetes
Hazard Ratio or
SMR (95% Cl)
All-cause (3rd vs.
1st patella Pb
tertile):
1 .76 (0.95, 3.26)
All CVD (3rd vs. 1st
tertile):
2.45 (1 .07, 5.60)
IHD(3rdvs. 1st):
8.37 (1 .29, 54.4)
Cancer (3rd vs.
1st):
0.59(0.21, 1.67)
After excluding 154
subjects with CVD
and stroke at
baseline:
All-cause (3rd vs.
1st):
2.52(1.17-5.41)
All CVD (3rd vs.
1st):
5.63(1.73, 18.3)
All-cause (3rd vs.
1st blood Pb tertile):
0.93(0.59, 1.45)
All CVD (3rd vs.
1st):
0.99(0.55, 1.78)
IHD(3rdvs. 1st):
1.30(0.54,3.17)
> 8 ug/dL vs.
<8 ug/dL
All cause:
1 .59 (1 .02, 2.49)
CVD: 1 .78 (0.92,
3.45)
Coronary Heart
Disease:
3.08 (1 .23, 7.70)
Stroke: 1.13 (0.34,
3.81)
Cancer: 1.64(0.73,
3.71)
November 2012
Draft - Do Not Cite or Quote
-------
Study
Study
Population /
Methodology Parameter
Pb Data
Statistical Analysis
Hazard Ratio or
SMR (95% Cl)
"Neuberger
et al. (2009)
Ecological
Residents at
or near Tar
Creek
Superfund
site, Ottawa
County, OK
(exposed pop.
5,852,
unexposed
pop. 16,210)
Cause-specific mortality
No biomarker
measurements
Standardized mortality ratio
(SMR) based on 2000 U.S.
Census data
Heart disease:
Both sexes:
114.1 (113.1, 115.2)
Men:
118(116.4, 119.6)
Women:
111 (109.5, 112.5)
Stroke:
Both sexes:
121.6(119.2,123.9)
Men:
146.7(107.4, 195.7)
Women:
106.5(80.2,138.6)
aCocco et al.
(2007)
Ecological
933 male Pb
smelter
workers from
Sardinia, Italy
(1973-2003)
All cause and cause-
specific mortality
No biomarker SMR
measurements
All cause: 56 (46,
68)
CVD: 37 (25, 55)
"These references not included in Figure 5-30 because they reported standardized mortality ratios.
5.4.6
Air Pb-PM Studies
i
2
3
4
5
6
7
8
9
10
11
12
13
14
15
5.4.6.1 Cardiovascular Morbidity
A relatively small number of studies used Pb measured in PMi0 and PM2 5 ambient air
samples to represent Pb exposures. However, given that size distribution data for Pb-PM
are fairly limited, it is difficult to assess the representativeness of these concentrations to
population exposure (Section 3.5.3). Moreover, data illustrating the relationships of
Pb-PMio and Pb-PM2 5 with blood Pb levels are lacking. A few available studies exposed
rats, dogs, or humans to concentrated ambient air particles (CAPS) in which Pb and
several other components were measured. Consistent with epidemiologic studies of blood
and bone Pb and with studies of animals exposed to Pb, these studies show exposure to
Pb-containing CAPS resulted in various changes related to increased vasoconstriction
(Urch et al.. 2004: Wellenius etal.. 2003: Batalha et al.. 2002). While Pb-containing
CAPS indicate cardiovascular effects with short-term exposure (2-6 hours over
multiple days), they cannot be attributed specifically to the Pb component of the mixture.
It is important to note that Urch et al. (2004) estimated the Pb effect on brachial artery
diameter based on the ambient concentration of Pb, not direct exposure to Pb isolated
from CAPs.
November 2012
Draft - Do Not Cite or Quote
-------
1 A U.S. time-series study of almost 3 million pregnant women found that increases in
2 ambient Pb-TSP concentrations were associated with increased odds of pregnancy
3 induced hypertension (PIH) assessed at delivery (Chen et al., 2006c). In contrast,
4 epidemiologic studies provide weak evidence for an association between short-term
5 changes (daily average) in ambient air concentrations of Pb- PM25 and cardiovascular
6 morbidity in adults adjusting for weather and time trends. Some of these time-series
7 studies analyzed Pb individually, whereas others applied source apportionment
8 techniques to analyze Pb as part of a group of correlated components. In a time-series
9 study of 106 U.S. counties, Bell et al. (2009) found that an increase in lag 0 Pb- PM2 5
10 was associated with an increased risk of cardiovascular hospital admissions among adults
11 ages 65 years and older. Quantitative results were not presented; however, the 95% CI:
12 was wide and included the null value. In this study, statistically significant associations
13 were observed for other PM metal components such as nickel, vanadium, and Zn. In the
14 absence of detailed data on correlations among components or results adjusted for
15 copollutants, it is difficult to exclude confounding by ambient air exposures to these other
16 components or copollutants. To address correlations among PM chemical components,
17 some studies applied source apportionment techniques to group components into
18 common source categories. In these source-factor studies, it is not possible to attribute the
19 observed association (Sarnat et al.. 2008) or lack of association (Andersen et al.. 2007)
20 specifically to Pb.
5.4.6.2 Mortality
21 Time-series epidemiologic studies of ambient air Pb- PM25 reported positive associations
22 with mortality. Although limited in number, these studies indicated associations in
23 multiple cities across the U.S. In the Harvard Six Cities Study, Laden et al. (2000) found
24 a 1.16% (95% CI: 0.20, 2.9%) increased risk in all-cause mortality per 461.4 ng/m3
25 (5th-95th percentile) increase in Pb-PM2 5. In six California counties, Ostro et al. (2007)
26 found that a 5 ng/m3 (interquartile range) increase in Pb-PM2 5 was associated with a
27 1.89% (95% CI: -0.57, 4.40%) increased risk of cardiovascular mortality and a 1.74%
28 (95% CI: 0.24, 3.26%) increased risk of all-cause mortality during the cool season. The
29 limitations of air-Pb studies were described in Section 5.4.6.1 above and also are relevant
30 to the interpretation of these findings for mortality.
November 2012 5-360 Draft - Do Not Cite or Quote
-------
5.4.7 Summary and Causal Determination
1 Large bodies of epidemiologic and toxicological evidence indicate effects of Pb exposure
2 on a range of related cardiovascular effects. For evaluation of causal relationships with
3 Pb exposure, evidence was grouped in categories using the U.S. Surgeon General's
4 Report on Smoking as a guideline (CDC. 2004). The categories include hypertension,
5 subclinical atherosclerosis, coronary heart disease, and cerebrovascular disease. The
6 causal determination for hypertension and increased BP is not only informed by evidence
7 for hypertension and blood pressure, but also cardiovascular mortality. Coronary heart
8 disease is informed by evidence for HRV, MI, IHD, mortality from MI, IHD, and CHD,
9 and in animals, increased thrombosis, coagulation, and arrhythmia. The biological
10 plausibility and mode of action for these cardiovascular effects is provided by evidence
11 for oxidative stress, inflammation, vascular cell activation or dysfunction. The sections
12 that follow describe the evaluation of evidence for these four groups of outcomes,
13 hypertension, subclinical atherosclerosis, coronary heart disease, and cerebrovascular
14 disease, with respect to causal relationships with Pb exposure using the framework
15 described in Table II of the Preamble. The key supporting evidence to the causal
16 framework is summarized in Table 5-24.
5.4.7.1 Evidence for Hypertension and Increased Blood Pressure
17 The 2006 Pb AQCD concluded that there was a relationship between higher blood Pb and
18 bone Pb and cardiovascular effects in adults, in particular increased BP and increased
19 incidence of hypertension (U.S. EPA, 2006b). and recent evidence strengthens this
20 conclusion. This conclusion is informed by the coherence of effects observed between
21 epidemiologic and toxicological findings and among related endpoints. Prospective
22 evidence and animal toxicology studies demonstrate the temporal relationship of the
23 exposure to effect, while meta-analyses provide indications of consistency and strength,
24 and cross-sectional evidence support the consistently observed results. Consideration of
25 numerous potential confounding factors in both the prospective and cross-sectional
26 studies limit uncertainty from bias and other lines of evidence characterizing modes of
27 action provide biological plausibility to the associations.
28 Longitudinal prospective studies clearly support the relationship between biomarkers of
29 Pb exposure and hypertension incidence and BP changes establishing the directionality of
30 effects. High-quality studies are replicated by different investigators using different
31 designs and in large cohorts in different locations (Peters et al.. 2007; Glenn et al.. 2006;
32 Cheng etal.. 2001). Bone Pb coupled with high perceived stress was associated with an
33 increased risk of developing hypertension in an originally nonhypertensive group of
November 2012 5-361 Draft - Do Not Cite or Quote
-------
1 adults (Peters et al.. 2007). Cheng et al. (2001) examined subjects from the NAS cohort
2 without hypertension at baseline measurement and reported a significant increase for
3 hypertension with patella Pb analyzed by linear models. A recent prospective study in Pb
4 workers found independent associations of both baseline blood Pb level and subsequent
5 changes in blood Pb over follow-up with changes in BP over follow-up and bone Pb level
6 with hypertension (Glenn et al.. 2006). The results indicated that different mechanisms
7 may mediate short-term Pb-associated increases in BP and long-term Pb-associated
8 development of hypertension. Consideration for key potential confounding factors was
9 appropriate including baseline age, alcohol consumption, BMI, and use of BP lowering
10 medications. Other factors such as smoking and education were evaluated but did not
11 predict systolic BP. When subjects with hypertension were excluded from the model, the
12 predicted change was not altered. Thus, chance, bias, and confounding can be ruled out
13 with reasonable confidence based on the consistent, positive, statistically significant
14 results indicated in these studies. Figure 5-18 and Figure 5-19 and the meta-analysis
15 indicate that results for effects of Pb exposure on BP and hypertension are positive and
16 precise. This provides more confidence in this relationship and reduces the level of
17 uncertainty.
18 The prospective evidence is supported by meta-analyses that underscore the consistency
19 and reproducibility of Pb-associated increases in BP and hypertension across diverse
20 populations and different study designs (Navas-Acien et al.. 2008; Nawrot et al.. 2002).
21 Nawrot et al. (2002) found that each doubling of concurrent blood Pb level (between 1
22 and >40 ug/dL) was associated with a 1 mmHg increase in systolic BP and a 0.6 mmHg
23 increase in diastolic BP. Navas-Acien et al. (2008) found that all included studies showed
24 a relationship between higher bone Pb levels and higher BP. Also, all but one that
25 characterized hypertension showed higher relative risks or odds ratios associated with
26 higher bone Pb levels.
27 Further support for a causal relationship between blood and bone Pb levels and increased
28 BP and hypertension is provided by many cross-sectional analyses conducted by
29 numerous researchers using different study designs and analyses in large, diverse cohorts
30 in different locations. A recent study in an ethnically diverse community-based cohort of
31 women and men aged 50-70 years found hypertension risk to be associated with blood
32 and tibia Pb levels (Martin et al.. 2006). Recent epidemiologic studies in adults found
33 associations with hypertension in populations with relatively low mean blood Pb levels.
34 For example, a positive relationship was found in the nationally representative NFfANES
35 III (1988-1994), in which the population geometric mean blood Pb level was 1.64 (ig/dL
36 (Muntner et al., 2005). Despite the extensive evidence for associations at relatively low
37 concurrent blood Pb levels, these cardiovascular outcomes were most often examined in
38 adults that have been exposed to higher levels of Pb earlier in life, and uncertainty
November 2012 5-362 Draft - Do Not Cite or Quote
-------
1 remains concerning the Pb exposure level, timing, frequency, and duration contributing to
2 the observed associations. However, evidence presented in the 1990 Pb Supplement to
3 the Addendum Q990a), indicated that in populations aged 20-74 years during 1976-1980
4 in NHANES II (Schwartz. 1991) across the range of 7-34 (ig/dL no evident threshold was
5 found below which the blood Pb level was not significantly related to BP. Further, as
6 described in Section 5.4.1. general population blood Pb levels across those aged 20 to 74
7 years as indicated by NHANES II and other studies probably peaked in the time frame of
8 1978-1988 achieving levels that were likely to persist over the long-term ranging from 10
9 to 30 (ig/dL.
10 Further, recent cross-sectional epidemiologic studies also emphasized the interaction
11 between Pb biomarker levels and factors, such as genetic variants, race/ethnicity, and
12 metabolic syndrome, in modifying the association with BP or hypertension. Evidence
13 was presented for a larger blood Pb-associated increase in BP in carriers of the ALAD2
14 allele, which is associated with greater binding affinity for Pb in the bloodstream (see
15 Figure 5-18 for results) (Scinicariello et al., 2010). Additionally, bone Pb level was
16 associated with larger increases in PP, which represents a good predictor of
17 cardiovascular morbidity and mortality and an indicator of arterial stiffness, among NAS
18 adults with the HFE H63D and/or C282Y variant (Zhang etal.. 2010a) (Figure 5-18 for
19 results). Park et al. (2009b) provided further evidence of HFE and transferrin gene
20 variants, related to iron metabolism, impacting the associations of bone Pb levels with
21 cardiovascular effects, evaluated by QT interval changes in the NAS cohort.
22 Combined evidence from prospective and cross-sectional studies helps limit the level of
23 uncertainty for bias from confounding with reasonable confidence. While the adjustment
24 for specific factors varied by study, the collective body of evidence adjusted for multiple
25 potential known key confounding factors, including age, diet, sex, BMI, blood pressure
26 lowering medication use, SES, race/ethnicity, alcohol consumption, cholesterol, smoking,
27 pre-existing disease (i.e., diabetes), measures of renal function, and copollutant exposures
28 (i.e., Cd).
29 Cardiovascular effects of Pb exposure in children are discussed in Section 5.4.4. Overall
30 this body of evidence, based on different cohorts, locations, and study designs provides a
31 preliminary literature base examining the potential for a relationship between biomarkers
32 for Pb exposure and cardiovascular effects in children. Recent studies provide
33 information for BP and antecedents for cardiovascular disease such as increases in TPR
34 and changes in cardiac autonomic regulation.
35 A causal relationship is further supported by coherence between epidemiologic and
36 toxicological evidence for the effects of long-term exposure on BP. Collectively, all
37 animal toxicological studies providing blood Pb level and BP measurements reported
November 2012 5-363 Draft - Do Not Cite or Quote
-------
1 increases in BP with increasing blood Pb level in the range relevant to humans (Figure
2 5-21). Whereas the majority of studies examined long-term Pb exposures that resulted in
3 mean blood Pb levels >10 (ig/dL, one animal toxicological study found a continuous
4 monotonic increase in BP in animals with a mean blood Pb level from 0.05 to 29 (ig/dL
5 with no evidence of a threshold (Tsao et al. 2000). Thus, most evidence demonstrated
6 such effects in adult animals with blood Pb levels >10 (ig/dL. Also, recent studies
7 demonstrated only partial reversibility of Pb-induced increased BP following Pb exposure
8 cessation or chelation and the possibility for short-term Pb exposure-induced increases in
9 BP. The short-term effects were found with routes of Pb exposure that may have
10 uncertain relevance to humans.
11 Coherence for BP and hypertension evidence was also provided by epidemiologic
12 evidence indicating associations with related CV conditions. Studies in the medical
13 literature show that increasing BP, even within the nonhypertensive range, is associated
14 with increased rates of death and cardiovascular disease, including CHD, stroke, and
15 cardiac failure (Ingelsson et al., 2008; Chobanian et al., 2003; Pastor-Barriuso et al.,
16 2003; Prospective Studies Collaboration. 2002; Kannel. 2000a. b; Neatonetal.. 1995).
17 Evidence for Pb-induced hypertension and increased BP is supported by, consistently
18 observed associations between Pb biomarkers and both cardiovascular and all-cause
19 mortality in prospective studies with follow-up periods ranging between 8 and 12 years.
20 A recent analysis of the NHANES III sample reported associations of adult blood Pb
21 level with cardiovascular mortality (Menke et al., 2006). These findings were supported
22 by a community-based cohort of women age 65-87 years, in which higher effect
23 estimates were observed for mortality from cardiovascular disease (Khalil et al., 2009b).
24 Weisskopf et al. (2009) published the first mortality study using bone Pb as an exposure
25 index. This prospective study found that patella bone Pb levels were associated with
26 increased mortality from cardiovascular disease.
27 Animal toxicology studies further indicate coherence and strengthen the evidence for
28 causality by providing strong biological plausibility for Pb-associated increases in BP and
29 hypertension. Hypertension results from dysfunction in the regulation of blood flow and
30 vascular resistance. Many systems, including the central and sympathetic nervous
31 systems, the contractile processes in the vasculature, and various hormonal regulators,
32 contribute to the maintenance of BP and disruption of these systems will alter BP
33 homeostasis. Studies demonstrate that oxidative stress produced following Pb exposure
34 inactivates the vasodilator NO which may lead to increased vasoconstriction and
35 increased BP, leading to hypertension. In addition, oxidative stress can damage the
36 endothelium, further disrupting endothelium-dependent vascular relaxation and
37 increasing the contractile response. Studies also suggest Pb exposure disrupts normal
38 contractile processes by altering the sympathetic nervous system, the renin-angiotensin-
November 2012 5-364 Draft - Do Not Cite or Quote
-------
1 aldosterone system, and the balance between production of vasodilators and
2 vasoconstrictors (Section 5.4.2.3).
3 Associations between biomarkers of Pb exposure and increased BP and hypertension
4 have been observed in a number of populations, including the large nationally
5 representative NHANES cohort (Menke et al.. 2006; Muntner et al.. 2005). In addition,
6 associations are found in other cohorts that include both men and women (Martin et al..
7 2006). Further, the meta-analyses assess cohorts both within the U.S. and international,
8 further supporting the generalizability of the relationship between Pb exposure and
9 increased BP and hypertension.
10 Changes in BP that have been associated with biomarkers of Pb exposure indicate a
11 modest change for an individual; however, these modest changes can have a substantial
12 public health implication at the population level. The reported effects represent a central
13 tendency of Pb-induced cardiovascular effects among individuals; some individuals may
14 differ in risk and manifest effects that are greater in magnitude. For example, a small
15 increase in BP may shift the population distribution and result in considerable increases
16 in the percentages of individuals with BP values that are clinically significant, i.e., an
17 indication of hypertension and medication use.
18 Overall, evidence in epidemiologic and toxicological studies demonstrates consistent
19 effects of long-term Pb exposure on increased BP and hypertension in adults; however,
20 uncertainty remains concerning the Pb exposure level, timing, frequency, and duration
21 contributing to the effects. The epidemiologic studies are of high-quality, have been
22 replicated by different researchers in different cohorts, and have adjusted for numerous
23 potential confounding factors. Thus, collectively, they help limit the level of uncertainty
24 for bias from confounding with reasonable confidence. In addition, a biologically
25 plausible potential mode of action is described. Thus, the overall evidence is sufficient to
26 conclude that there is a causal relationship between Pb exposure and hypertension.
5.4.7.2 Evidence for Subclinical Atherosclerosis
27 Measures of subclinical atherosclerosis provide the opportunity to assess the pathogenesis
28 of vascular disease at an earlier stage. Studies that discuss markers of subclinical
29 atherosclerosis, such as PAD (i.e., ankle-brachial index) and generalized atherosclerosis
30 (i.e., IMT), are included in this category. A limited number of studies have evaluated
31 markers of subclinical atherosclerosis following Pb exposure in adult humans or animals.
32 One study described in the 2006 Pb AQCD (U.S. EPA. 2006b) indicated that Pb was
33 associated with PAD in the NHANES population and coexposure with Cd did not
34 confound the association (Navas-Acien et al., 2004). Recent epidemiologic findings are
November 2012 5-365 Draft - Do Not Cite or Quote
-------
1 limited to cross-sectional analyses, so uncertainty exists as to the specific Pb exposure
2 level, timing, frequency, and duration that contributed to the observed effects. One study
3 reported an increasing trend in the odds of PAD and concurrent blood Pb level in adults
4 within the NHANES population (Muntner et al.. 2005). which is consistent with the
5 results from the previous Navas-Acien et al. (2004) analysis. An occupational study of
6 Pb-exposed adults with a mean blood Pb level around 25 (ig/dL presented evidence for
7 increased measures of atherosclerosis analyzed by Doppler ultrasound (i.e., greater IMT
8 and atherosclerotic plaque presentation) in the Pb-exposed population (Poreba et al..
9 2011). Similarly, toxicological studies have provided limited evidence to suggest long-
10 term Pb exposure may initiate atherosclerotic vessel disease. Pb exposure to human radial
11 and internal mammary arteries resulted in a concentration-dependent increase in arterial
12 intimal thickness (Zeller et al.. 2010). Also, exposure to Pb in rats increased aortic medial
13 thickness (Zhang et al.. 2009a).
14 Toxicological studies also present evidence to clearly describe a plausible biological
15 mechanism. Atherosclerosis is considered an inflammatory disease with a clear role for
16 oxidative stress in the pathogenesis of the disease. There is consistent evidence that Pb
17 exposure promotes oxidative stress and increased inflammation in animal and cell culture
18 models (Section 5.4.2.3). In addition, there is evidence that Pb will stimulate vascular cell
19 activation and lead to endothelial cell dysfunction. Both events are key to the
20 development and progression of atherosclerosis. Also, epidemiologic and animal
21 toxicology studies have related higher blood Pb levels with higher cholesterol; high
22 cholesterol is one of the principal risk factors for atherosclerosis (Section 5.4.3.3).
23 In summary, the evidence includes one high-quality epidemiologic study with control for
24 numerous potential confounders (Muntner et al.. 2005) and biological plausibility for the
25 effects observed in humans. Thus, the evidence for subclinical atherosclerosis is
26 suggestive of a causal relationship.
5.4.7.3 Evidence for Coronary Heart Disease
27 Coronary heart disease (CHD) results from interruption of the blood supply to a part of
28 the heart resulting from atherosclerosis of the coronary arteries, with acute injury and
29 scarring leading to permanent damage to the heart muscles. A disrupted HRV has been
30 associated with a higher mortality after MI and is used as a predictor of the physiological
31 processes underlying CHD (Buccelletti et al.. 2009). Studies that discuss incidence of MI,
32 IHD, HRV, and mortality from CHD, MI, or IHD are included in this category.
33 There were a small number of studies discussed in the 2006 Pb AQCD (U.S. EPA.
34 2006b) that indicated associations between Pb biomarker levels and increased risk of
November 2012 5-366 Draft - Do Not Cite or Quote
-------
1 cardiovascular outcomes associated with CHD. However, recent longitudinal studies in
2 cohorts in different locations with follow-up periods ranging between 8 and 12 years
3 report that biomarkers of Pb exposure are associated with risk of mortality from
4 cardiovascular disease, specifically MI, IHD, or CHD. A recent analysis of the NHANES
5 III sample reported associations of adult blood Pb level with cardiovascular mortality,
6 with stronger associations observed with MI mortality (Menke et al.. 2006). These
7 findings were supported by a community-based cohort of women age 65-87 years, in
8 which higher effect estimates were observed for mortality from CHD (Khalil et al..
9 2009b). Weisskopf et al. (2009) published the first mortality study using bone Pb as an
10 exposure index. This prospective study found that patella bone Pb levels were associated
11 with increased mortality from IHD. Despite the differences in design and methods across
12 studies, with few exceptions associations between higher levels of Pb biomarkers and
13 higher risk of mortality were consistently observed (Figure 5-30 and Table 5-23).
14 The body of evidence demonstrating associations with mortality from CHD is
15 substantiated by several findings indicating associations between biomarkers of Pb and
16 incidence of CHD-related outcomes. A prospective analysis examined the incidence of
17 IHD (physician confirmed MI, angina pectoris) in the NAS cohort and reported findings
18 indicating that both blood and bone Pb levels contribute independently to IHD incidence
19 (Jain et al.. 2007). Earlier studies reported associations of increased Pb biomarkers with
20 increased risk of left ventricular hypertrophy (Schwartz. 1991). Coherence for the
21 associations in humans is provided by a recent animal study that suggested that Pb
22 exposure promotes a procoagulant state that could contribute to thrombus formation
23 which could reduce the blood supply to the heart (Shin et al.. 2007).
24 Further support for a relationship between Pb exposure and CHD is provided by evidence
25 from the NAS cohort for effects on disrupted HRV (Bum et al.. 2011; Park et al.. 2009b:
26 Park et al.. 2006). which has been associated with a higher mortality from MI and is used
27 as a predictor of the physiological processes underlying CHD. A prospective analysis
28 reported that higher tibia Pb, but not blood or patella Pb, was associated with increases in
29 QTc interval and QRSc duration (Eumetal.. 2011). Park et al. reported associations of
30 bone Pb with HRV measures and effect modification by increasing number of iron
31 metabolism gene variants from 0 to 3. Park et al. (2006) reported associations of bone Pb
32 with HRV measures and effect modification by increasing number of iron metabolism
33 gene variants from 0 to 3. Park et al. (2006) reported the strongest relationships between
34 patella Pb levels and lower HRV among those with three or more metabolic
35 abnormalities. Also, bone Pb level was associated with larger decreases in HRV among
36 adults with metabolic syndrome, which like reduced HRV is associated with increased
37 risk of cardiovascular events (Park et al.. 2006).
November 2012 5-367 Draft - Do Not Cite or Quote
-------
1 As CHD is the result of vascular blockage, the suggestive evidence for subclinical
2 atherosclerosis supports the observations of increased CHD morbidity and mortality. In
3 addition, the strong and consistent evidence for Pb-induced hypertension serves as further
4 biological plausibility for CHD. Hypertension may contribute to CHD development in a
5 number of ways. Hypertension may lead to thickening of the vascular wall or
6 exacerbation of atherosclerotic plaque development and thus contribute to plaque
7 instability. In addition, hypertension may increase the myocardial oxygen demand
8 priming for potential myocardial ischemia (Olafiranye et al.. 2011). Both subclinical
9 atherosclerosis and hypertension are supported by consistent evidence describing the
10 mode of action including Pb-induced oxidative stress, inflammation, cellular activation
11 and dysfunction, altered vascular reactivity, RAAS dysfunction, and vasomodulator
12 imbalance.
13 Building on this strong body of evidence, recent epidemiologic and toxicological studies
14 substantiated the evidence that long-term Pb exposure is associated with CHD in adults;
15 however, uncertainty remains concerning the Pb exposure level, timing, frequency, and
16 duration contributing to the effects. Overall, high-quality studies examining CHD
17 morbidity and mortality and contributing cardiovascular effects have been replicated by
18 different researchers in different cohorts and report consistent associations that increase
19 the confidence that a relationship exists between Pb exposure and CHD. In addition, both
20 animal and human studies describe a biologically plausible potential mode of action.
21 Thus the overall evidence is sufficient to conclude that there is a causal relationship
22 between Pb exposure and coronary heart disease.
5.4.7.4 Evidence for Cerebrovascular Disease
23 Cerebrovascular disease describes a group of conditions involving the cerebral blood
24 vessels that result in transient or permanent disruption of blood flow to the brain. These
25 conditions include stroke, transient ischemic attack, and subarachnoid hemorrhage. Both
26 hypertension and atherosclerosis are risk factors for Cerebrovascular disease and the
27 mechanisms for these outcomes also apply to Cerebrovascular disease. Despite strong
28 evidence for hypertension and CHD and long-term Pb exposure, very few studies have
29 examined the effects of Pb exposure on Cerebrovascular disease. Lee et al. (2009)
30 examined 153 patients in Taiwan cross-sectionally while adjusting for key confounders
31 and reported increased stenosis greater than 50% in the intracarotid system related to
32 urine Pb but not blood Pb level. Two epidemiologic studies prospectively evaluated
33 mortality from stroke. Menke et al. (2006) reported a positive relationship with wide
34 confidence intervals compared to other outcomes considered for blood Pb levels with
35 stroke mortality in the NHANES study. Khalil et al. (2009b) provides a non significant
November 2012 5-368 Draft - Do Not Cite or Quote
-------
1
2
3
4
result with imprecise confidence intervals. These few studies provide insufficient
evidence to inform the causal relationship between cerebrovascular disease and long-term
Pb exposure. Thus, the evidence at this time is inadequate to determine that a causal
relationship exists between Pb exposure and cerebrovascular disease.
Table 5-24 Summary of evidence supporting cardiovascular causal
determinations.
Attribute in Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or Blood
Levels Associated with
Effects0
Hypertension - Causal
Consistent associations
with relevant blood Pb
levels from multiple,
high quality
epidemiologic studies
Longitudinal prospective evidence for
associations with incidence of
hypertension and increase in blood
pressure in adults.
Large body of supportive cross-sectional
studies applying differing designs across
multiple cohorts of adults in different
locations.
Meta-analyses provide further support
Associations found while adjusting for
numerous potential confounding factors
Studies provide C-R information
Peters et al. (2007).
Glenn et al. (2006).
Cheng et al. (2001)
Martin et al. (2006).
Scinicariello et al. (2010).
Park et al. (2009c)
Navas-Acien et al. (2008).
Nawrot et al. (2002)
Section 5.4.2.1
Adult, prospective :
Blood Pb level >20 ug/dL;
Bone Pb level >20 ug/g
Adult, concurrent:
Blood Pb level >2 ug/dL;
Bone Pb level >19 ug/g
Consistent toxicological
results provide
coherence with
epidemiologic evidence
Consistent cross-sectional evidence for
increases in BP in adults are supported by
studies in adult rodents with relevant
dietary long-term Pb exposure
Rodents:
Rizzi et al.(2009).
Bravo et al.(2007),
Chang et al.(2005),
Tsao et al. (2000)
Rat, adult:
Blood Pb level >10 ug/dL
Section 5.4.2.2
Consistent associations
with relevant Pb levels
in blood and/or bone
and cardiovascular
mortality from multiple,
high quality
epidemiologic studies
Longitudinal, prospective studies find
consistent associations of blood and/or
bone Pb levels in adults with risk of
cardiovascular mortality applying differing
designs across multiple cohorts in different
locations while controlling for potential
confounding.
Khalil et al. (2009b).
Weisskopfetal. (2009).
Menke et al. (2006)
Schoberet al. (2006)
Lustberg and Silbergeld
(2002)
Section 5.4.5
Adult, prospective:
Blood Pb level >4 ug/dL
November 2012
5-369
Draft - Do Not Cite or Quote
-------
Attribute in Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or Blood
Levels Associated with
Effects0
Evidence clearly
describes mode of
action
Oxidative Stress
Alteration of vascular
reactivity
Renin-angiotensin-
aldosterone system
dysfunction
Vasomodulator
imbalance
Consistent evidence of increased
oxidative stress leading to inactivation of
'NO and downregulation of sGC in
animals with relevant dietary Pb
exposures and cultured vascular cells.
Toxicological evidence for activation of the
sympathetic nervous system, increased
reactivity to catecholamines, and
activation of the adrenergic and
dopaminergic receptors in rats, isolated
vessels, and cultured cells.
Mixed evidence for reactivity to other
pressor agents (e.g., 5-HT) in rats.
Toxicological evidence that activation of
the RAAS may be involved in
development of Pb-induced hypertension
Evidence for increased RAAS activity in
rats and decreased BP following RAAS
inhibition and Pb exposure.
Limited available toxicological evidence
reporting vasomodulator imbalance in
Pb-exposed rats and cells.
Section 5.4.2.3
Subclinical Atherosclerosis - Suggestive
Limited evidence in
humans of an
association with
subclinical
atherosclerosis and
peripheral artery
disease
One NHANES analysis reported
associations with PAD at relevant adult
blood Pb levels with control for potential
confounding.
Limited evidence for increased IMT or
arterial stiffness in adult human
populations.
Occupational studies report increased IMT
and atherosclerotic plaque presentation in
highly exposed adult populations.
Muntner et al. (2005)
Ari et al. (2011
Poreba et al. (2011: 2011 a)
Sections 5.4.3.3 and
5.4.3.5
Adult, concurrent:
Blood Pb level >2.5 ug/dL
Adult, concurrent:
Serum Pb level >0.4 ug/dL
Adult workers:
Blood Pb level >24 ug/dL
Limited evidence in
animals of initiation or
progression of
atherosclerosis after Pb
exposure
Limited studies reporting increased IMT,
vascular morphological changes, and
endothelial and SMC alterations in rats
and human tissue.
Zelleretal. (2010).
Zhang et al. (2009a)
Section 5.4.3.3
Rat: 28.4 ug/dL
Human Tissue: 50 uM
Evidence clearly
describes mode of
action
Oxidative Stress
Inflammation
Vascular Cell
Activation and
Endothelial
Dysfunction
Consistent evidence of increased
oxidative stress in animals with relevant
dietary Pb exposures and cultured
vascular cells.
Toxicological evidence of increased
inflammation as indicated by increased
production of TNF-a, IL-6, IL-8, and PGE2
by macrophages and vascular cells.
Toxicological evidence of VSMC
stimulation and endothelial dysfunction
and damage in culture.
Limited available evidence of impaired
flow-mediated dilatation in Pb exposed
workers.
Section 5.4.2.3
Section 5.4.3.1
November 2012
5-370
Draft - Do Not Cite or Quote
-------
Attribute in Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or Blood
Levels Associated with
Effects0
Coronary Heart Disease - Causal
Consistent associations
with relevant bone
and/or blood Pb levels
and mortality from Ml,
IHD, CHD, and
cardiovascular disease
from multiple, high
quality epidemiologic
studies
Longitudinal, prospective studies find
consistent associations of bone and/or
blood Pb levels in adults with risk of
cause-specific cardiovascular mortality
applying differing designs across multiple
cohorts in different locations with control
for potential confounding.
Khalil et al. (2009b),
Weisskopfetal. (2009).
Menke et al. (2006)
Schoberetal. (2006)
Lustberg and Silbergeld
(2002)
Section 5.4.5
Adult, prospective:
Blood Pb level >4 ug/dL
Limited evidence in
humans of an
association with
ischemic heart disease,
myocardial infarction,
or HRV
One prospective study demonstrates an
association of adult blood and bone Pb
levels with incidence of IHD in the MAS
cohort
Associations of Pb levels in adults and left
ventricular hypertrophy and Ml
Prospective evidence of association of
HRV with tibia bone Pb level in adults.
Evidence for interaction of markers of
metabolic syndrome and genetic
polymorphisms with Pb-induced HRV.
Jainetal. (2007)
Schwartz (1991)
Eum et al. (2011)
Park et al. (2009b: 2006)
Sections 5.4.3.4 and
5.4.3.6
Adult, prospective:
Blood Pb level >5 ug/dL
Adult, prospective:
Bone Pb level >23 ug/g
Limited evidence in
animals of increased
thrombosis, enhanced
coagulation, and
arrhythmia
One study reporting increased thrombosis
and enhanced coagulation in rats and
cells.
One study reporting increased incidence
of arrhythmia and atrioventricular
conduction block in rats.
Rat
Blood Pb level: 26.8 ug/dL
Reza et al. (2008)
Sections 5.4.3.4 and
5.4.3.6
Evidence clearly
describes mode of
action
Oxidative Stress
Inflammation
Atherosclerosis
Hypertension
Consistent evidence of increased
oxidative stress in animals with relevant
dietary Pb exposures and cultured
vascular cells.
Toxicological evidence of increased
inflammation as indicated by increased
production of TNF-a, IL-6, IL-8, and PGE2
by macrophages and vascular cells.
Suggestive evidence of subclinical
atherosclerosis in humans and animals
with relevant Pb exposure resulting in
narrowing of the blood vessels to the
heart.
Consistent evidence of increased BP and
hypertension following Pb exposure in
humans and animals at relevant Pb levels
across numerous studies with control for
confounding.
Association of increased blood pressure
with manifestation of CHD has been well
documented.
Section 5.4.2.3
Section 5.4.3.1
Sections 5.4.3.3 and 5.4.3.5
Section 5.4.2
Cerebrovascular Disease - Inadequate
Evidence for
cerebrovascular
disease in humans and
animals is of insufficient
quality and quantity
One study reported an association of
intracranial carotid stenosis with urinary
Pb level.
Lee et al. (2009)
Section 5.4.3.3
Adult, concurrent:
Blood Pb level >5 ug/dL
November 2012
5-371
Draft - Do Not Cite or Quote
-------
Attribute in Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or Blood
Levels Associated with
Effects0
Limited evidence for
increased mortality
from stroke
Limited evidence for increased risk of
mortality from stroke across two cohorts in
different locations.
Menke et al. (2006),
Khalil et al. (2QQ9b)
Section 5.4.5
Adult, prospective:
Blood Pb level >4 ug/dL
Evidence for possible
mode of action
Hypertension
Atherosclerosis
Consistent evidence of increased BP and
hypertension following Pb exposure in
humans and animals at relevant Pb levels
across numerous studies with control for
confounding.
Association of increased blood pressure
with manifestation of CHD has been well
documented.
Suggestive evidence of subclinical
atherosclerosis in humans and animals
with relevant Pb exposure resulting in
narrowing of the blood vessels to the
heart.
Section 5.4.2
Sections 5.4.3.3 and 5.4.3.5
"Described in detail in Table II of the Preamble.
bDescribes the key evidence and references contributing most heavily to causal determination. Also noted are the sections where
full body of evidence is described.
°Describes the blood Pb levels in humans with which the evidence is substantiated and blood Pb levels in animals most relevant to
humans.
dBecause blood Pb level in nonoccupationally-exposed adults reflects both recent and past Pb exposures, the magnitude, timing,
frequency, and duration of Pb exposure contributing to the observed associations is uncertain.
5.5
Renal Effects
5.5.1
Introduction
i
2
o
J
4
5
6
7
8
9
10
11
This section summarizes key findings with regard to effects of Pb on the kidney in animal
toxicology and epidemiologic studies. Findings summarized across epidemiologic and
toxicological studies indicate that long-term Pb exposure is associated with pathological
changes in the renal system such as proximal tubule (PT) cytomegaly, renal cell
apoptosis, mitochondrial dysfunction, aminoaciduria, increased electrolyte excretion,
ATPase dysfunction, oxidant redox imbalance, altered glomerular filtration rate (GFR),
chronic kidney disease (CKD) development, and altered NO homeostasis with ensuing
elevated BP. As several of these outcomes are most often observed in adults with likely
higher past Pb exposures, uncertainty exists as to the Pb exposure level, timing,
frequency, and duration contributing to the associations observed with blood or bone Pb
levels.
November 2012
5-372
Draft - Do Not Cite or Quote
-------
1 The cardiovascular and renal systems are intimately linked. Homeostatic control at the
2 kidney level functions to regulate water and electrolyte balance via filtration,
3 re-absorption and excretion and is under tight hormonal control. Pb exposure has been
4 shown to damage the kidneys and its vasculature with ensuing effects on systemic
5 hypertension and effects on the cardiovascular (Section 5.4) and renal systems. Chronic
6 increases in vascular pressure can contribute to glomerular and renal vasculature injury,
7 which can lead to progressive renal dysfunction and kidney failure. In this manner,
8 Pb-induced hypertension has been regarded as one potential contributor of Pb-induced
9 renal disease. However, the relationship between BP and renal function is more
10 complicated. Not only does hypertension contribute to renal dysfunction but damage to
11 the kidneys can also cause increased BP. Long-term control of arterial pressure is
12 affected by body fluid homeostasis which is regulated by the kidneys. In examination of
13 the physiological definition of BP (i.e., mean BP equates to cardiac output multiplied by
14 total peripheral resistance [TPR]) the role of the kidneys in BP regulation is highlighted.
15 Cardiac output is driven by left ventricular and circulating blood volume. TPR is driven
16 by vasomodulation and electrolyte balance. Thus, it is possible to dissect the causes of
17 hypertension from features of primary kidney disease. Increased extracellular fluid
18 volume results in increased blood volume which enhances venous return of blood to the
19 heart and increases cardiac output. Increased cardiac output not only directly increases
20 BP, but also increases TPR due to a compensatory autoregulation or vessel constriction.
21 In addition, damage to the renal vasculature will alter the intra-renal vascular resistance
22 thereby altering kidney function and affecting the balance between renal function and BP.
23 The interactions between these systems can lead to further exacerbation of vascular and
24 kidney dysfunction following Pb exposure. As kidney dysfunction can increase BP and
25 increased BP can lead to further damage to the kidneys, Pb-induced damage to both
26 systems may result in a cycle of further increased severity of disease.
27 In general, associations between bone Pb (particularly in the tibia) and health outcomes in
28 adults indicate chronic effects of cumulative Pb exposure. In adults without current
29 occupational Pb exposure, blood Pb level represents both recent and cumulative Pb
30 exposure. In particular, blood Pb level may represent cumulative exposure in
31 physiological circumstances of increased bone remodeling or loss (e.g., osteoporosis and
32 pregnancy) when Pb from bone of adults contributes substantially to blood Pb
33 concentrations. Blood Pb level in children is also influenced by Pb stored in bone due to
34 rapid growth-related bone turnover in children relative to adults. Thus, blood Pb in
35 children is also reflective of cumulative dose. Additional details on the interpretation of
36 Pb in blood and bone are provided in Section 4.3.5. The toxicokinetics of Pb in blood and
37 bone are important considerations in making inferences about etiologically-relevant Pb
38 exposures that contributed to associations observed between blood and bone Pb levels
39 and health outcomes.
November 2012 5-373 Draft - Do Not Cite or Quote
-------
5.5.1.1 Kidney Outcome Measures
1 The primary function of the kidneys is to filter waste from the body while maintaining
2 appropriate levels of water and essential chemicals, such as electrolytes, in the body.
3 Therefore, the gold standard for kidney function assessment involves measurement of the
4 GFR through administration of an exogenous radionuclide or radiocontrast marker
5 (e-g-, 1251-iothalamate, iohexol) followed by timed sequential blood samples or, more
6 recently, kidney imaging, to assess clearance through the kidneys. This procedure is
7 invasive and time-consuming. Therefore, serum levels of endogenous compounds are
8 routinely used to estimate GFR in large epidemiologic studies and clinical settings.
9 Creatinine is the most commonly measured endogenous compound; blood urea nitrogen
10 (BUN) has also been examined. Increased serum concentration or decreased kidney
11 clearance of these markers both indicate decreased kidney function. The main limitation
12 of endogenous compounds identified to date is that non-kidney factors impact their serum
13 levels. Specifically, since creatinine is metabolized from creatine in muscle, muscle mass
14 and diet affect serum levels resulting in variation in different population subgroups
15 (e.g., women and children compared to men), that are unrelated to kidney function.
16 Measured creatinine clearance, involving measurement and comparison of creatinine in
17 both serum and urine, can address this problem. However, measured creatinine clearance
18 utilizes timed urine collections, traditionally over a 24-hour period, and the challenge of
19 complete urine collection over an extended time period makes compliance difficult.
20 Therefore equations to estimate kidney filtration that utilize serum creatinine but also
21 incorporate age, sex, race, and, in some, weight (in an attempt to adjust for differences in
22 muscle mass), have been developed. Although these are imperfect surrogates for muscle
23 mass, such equations are currently the preferred outcome assessment method.
24 Traditionally, the Cockcroft-Gault equation (Cockcroft and Gault. 1976). which estimates
25 creatinine clearance, a GFR surrogate, has been used. In the last decade, the abbreviated
26 Modification of Diet in Kidney Disease (MDRD) Study equation (Levey et al.. 2000;
27 Levey etal.. 1999). which estimates GFR, has become the standard in the kidney
28 epidemiologic and clinical communities. With widespread use of the MDRD equation, it
29 became clear that the equation underestimates GFR at levels in the normal range.
30 Therefore, the CKD-Epidemiology Collaboration (CKD-EPI) equation was recently
31 developed to be more accurate in this range (Levey et al.. 2009). This is a decided
32 advantage in nephrotoxicant research since most participants in occupational and many
33 even in general population studies have GFRs in a range that is underestimated by the
34 MDRD equation.
35 Both the MDRD and CKD-EPI equations use serum creatinine. Due to the inability to
36 adjust serum creatinine levels for muscle mass, alternative serum biomarkers have been
November 2012 5-374 Draft - Do Not Cite or Quote
-------
1 evaluated such as cystatin C, a cysteine protease inhibitor that is filtered, reabsorbed, and
2 catabolized in the kidney (Fried. 2009). It is produced and secreted by all nucleated cells
3 thus avoiding the muscle mass confounding that exists with serum creatinine (Fried.
4 2009). However, recent research indicates that serum cystatin C varies by age, sex, and
5 race (Kottgen et al.. 2008). Thus, a cystatin C-based eGFR equation was recently
6 developed that includes age, sex, and race (Stevens et al.. 2008).
7 Most of the kidney outcome measures discussed above were developed for use in the
8 clinical setting. Unfortunately, they are insensitive for detection of early kidney damage,
9 as evidenced by the fact that serum creatinine remains normal after kidney donation.
10 Therefore, in the last two decades, the utility of early biological effect (EBE) markers as
11 indicators of preclinical kidney damage has been of interest. These can be categorized as
12 markers of function (i.e., low molecular weight proteins that should be reabsorbed in the
13 PT such as p2-microglobulin and retinol-binding protein [RBP]); biochemical alteration
14 (i.e., urinary eicosanoids such as prostaglandin E2, prostaglandin F2 alpha, 6-keto-
15 prostaglandin FI alpha, and thromboxane B2); and cytotoxicity (e.g., N-acetyl-(3-D-
16 glucosaminidase [NAG]) (Cardenas et al.. 1993). Elevated levels may indicate an
17 increased risk for subsequent kidney dysfunction. However, most of these markers are
18 research tools only, and their prognostic value remains uncertain since prospective
19 studies of most of these markers in nephrotoxicant-exposed populations are quite limited
20 to date. Recently, microalbuminuria has been identified as a PT marker, not just
21 glomerular as previously thought (Comper and Russo. 2009). Kidney EBE markers are a
22 major recent focus for research in patients with acute kidney injury (AKI) and markers
23 such as neutrophil gelatinase-associated lipocalin (NGAL) and kidney injury molecule-1
24 (Kim-1), developed in AKI research, may prove useful for chronic nephrotoxicant work
25 as well (Ferguson et al.. 2008; Devarajan. 2007).
5.5.2 Nephrotoxicity and Renal Pathology
5.5.2.1 Epidemiology in Adults
26 A number of advances in research on the impact of Pb on the kidney in the 20 years
27 following the 1986 Pb AQCD (U.S. EPA. 1986a) were noted in the 2006 Pb AQCD (U.S.
28 EPA. 2006b). These included research in general and CKD patient populations at much
29 lower blood Pb levels (5-10 (ig/dL) at the time of evaluation than were previously
30 studied. These advances contributed to the understanding of the effects of Pb exposure on
31 kidney dysfunction overall in the population. Pb, at much lower doses than those causing
32 chronic Pb nephropathy, may act as a cofactor with other more established kidney risks to
November 2012 5-375 Draft - Do Not Cite or Quote
-------
1 increase the risk of CKD and disease progression in susceptible patients. Marie and Hall
2 (2011) note that data from basic and clinical studies suggest that obesity, hypertension,
3 hyperglycemia, hyperlipedemia, and other elements of the metabolic syndrome are highly
4 interrelated and contribute to the development and progression of diabetic nephropathy
5 and thus represent populations potentially at increased risk for kidney dysfunction.
6 In the 2006 Pb AQCD (U.S. EPA. 2006b). several key issues could not be completely
7 resolved based on the Pb-kidney literature published to date. These included
8 characterizing the lowest Pb dose at which altered kidney function effects occur, the
9 impact of higher past exposures on associations with concurrent Pb biomarker levels, the
10 impacts of Pb on the kidney in children, the use of paradoxical Pb-kidney associations on
11 risk assessment in the occupational setting, and the impact of co-exposure to other
12 environmental nephrotoxicants, such as Cd. In the intervening five years, relevant data
13 addressing several of these challenges have been published.
General Population Studies
14 The 2006 Pb AQCD (U.S. EPA. 2006b) reported studies that examined associations
15 between indicators of Pb exposure and kidney function in general populations. This was a
16 new approach to Pb-kidney research in the two decade time period covered by the
17 2006 Pb AQCD. As illustrated in Figure 5-31 and Table 5-25. studies consistently
18 demonstrate associations between higher blood Pb level and lower renal function in
19 adults. These general population studies provided critical evidence that the effects of Pb
20 on the kidney occur at much lower doses than previously appreciated based on
21 occupational exposure data. However, because blood Pb level in nonoccupationally-
22 exposed adults reflects both recent and past Pb exposures, the magnitude, timing,
23 frequency, and duration of Pb exposure contributing to the observed associations was
24 uncertain. The evidence of Pb-associated renal effects in general population studies was
25 substantiated by results that were adjusted for multiple potential confounding factors
26 including age, race, sex, education, household income, smoking, alcohol use, and various
27 health indicators such as diabetes, SBP, BMI, and history of cardiovascular disease. A
28 few studies also adjusted for Cd exposure.
29 The landmark Cadmibel Study was the first large environmental study of this type that
30 adjusted for multiple kidney risk factors, including urinary Cd (Staessen et al.. 1992). It
31 included 965 men and 1,016 women recruited from Cd exposed and control areas in
32 Belgium. Mean concurrent blood Pb was 11.4 ug/dL (range 2.3-72.5) and 7.5 ug/dL
33 (range 1.7-60.3) in men and women, respectively. After adjustment for covariates (Table
34 5-25). log transformed blood Pb was negatively associated with measured creatinine
35 clearance. A 10-fold increase in blood Pb was associated with a decrease in creatinine
November 2012 5-376 Draft - Do Not Cite or Quote
-------
1 clearance of 10 and 13 mL/min in men and women, respectively. Blood Pb was also
2 negatively associated with estimated creatinine clearance.
3 Multiple analyses assessing the kidney impact of Pb exposure have been conducted in the
4 NAS population (Tsaih et al.. 2004; Wu et al.. 2003a: Kimetal.. 1996; Pavton et al..
5 1994). Participants in this study were originally recruited in the 1960s in the Greater
6 Boston area. The inclusion criteria, male sex, age 21 to 80 years, and absence of chronic
7 medical conditions, limit the generalizability of the results to the rest of the U.S.
8 population. Longitudinal data contained in NAS publications remain essential to address
9 the dearth of prospective data on the kidney effects of Pb. The first of these included 459
10 men whose blood Pb levels from periodic examinations, conducted every 3 to 5 years
11 during 1979-1994, were estimated based on measurements in stored packed red blood
12 cell samples adjusted for hematocrit level (Kim etal.. 1996). Participants were randomly
13 selected to be representative of the entire NAS population in terms of age and follow-up.
14 Kidney function was assessed with serum creatinine. Data from four evaluations were
15 available for the majority of participants. At baseline, mean (SD) age, blood Pb level, and
16 serum creatinine, at baseline, were 56.9 (8.3) years, 9.9 (6.1) ug/dL, and 1.2 (0.2) mg/dL,
17 respectively. In the longitudinal analysis with random-effects modeling of repeated
18 measures, In-transformed blood Pb was associated with an increase in serum creatinine
19 from the previous to current follow-up period in the 428 participants whose highest blood
20 Pb level was < 25 ug/dL (|3= 0.027 mg/dL [95% CI: 0.0, 0.054] per unit increase in In
21 blood Pb); effect estimates in the entire group and subsets with different peak blood Pb
22 levels (< 10 or 40 ug/dL) also were positive (and larger for blood Pb levels < 10 ug/dL).
23 This study made two other key contributions. In order to address the question of whether
24 nephrotoxicity observed at current blood Pb levels is due to higher blood Pb levels from
25 past exposure, these authors performed a sensitivity analysis in participants whose peak
26 blood Pb levels, dating back to 1979, were < 10 ug/dL. A statistically significant positive
27 association between blood Pb and concurrent serum creatinine remained in a cross-
28 sectional analysis. These authors evaluated reverse causality, which attributes increased
29 blood Pb levels to lack of kidney excretion rather than as a causative factor for CKD, by
30 showing in adjusted plots that the association between blood Pb and serum creatinine
31 occurred over the entire serum creatinine range (0.7-2.1 mg/dL), including the normal
32 range where reverse causality would not be expected.
33 Cortical and trabecular bone Pb measurements were obtained in addition to whole blood
34 Pb in evaluations performed in the NAS between 1991 and 1995. Associations between
35 baseline blood, tibia, and patella Pb and change in serum creatinine over an average of 6
36 years in 448 men were reported in a subsequent NAS publication (Tsaih et al.. 2004). At
37 baseline, eligible participants were similar to nonparticipants with regard to age, BMI,
November 2012 5-377 Draft - Do Not Cite or Quote
-------
1 alcohol consumption, smoking status, diabetic status, hypertensive status, baseline SCr,
2 and blood and bone Pb levels, indicating lack of selective follow-up by blood/bone Pb
3 level or kidney function. At baseline 6 and 26% of subjects had diabetes and
4 hypertension, respectively. Mean blood Pb levels and serum creatinine decreased
5 significantly over the follow-up period in the group. In cross-sectional analyses, both
6 patella and tibia Pb, but not blood Pb level, were positively but nonsignificantly
7 associated with serum creatinine. Baseline blood Pb level was not significantly associated
8 with change in creatinine in all participants. However, diabetes was observed to be an
9 effect modifier of the relations of blood and tibia Pb with change in serum creatinine. Per
10 unit increase in In blood Pb, the increase in serum creatinine between follow-up periods
11 was substantially stronger in diabetics ((3 = 0.076 mg/dL [95% CI: 0.031, 0.121])
12 compared to non-diabetics ((3 = 0.006 mg/dL [95% CI: -0.004, 0.016]). A similar
13 relationship was observed for tibia Pb. An interaction was also observed between tibia Pb
14 and hypertension, although it is possible that many of the 26 diabetics were also included
15 in the hypertensive group and were influential there as well. A sensitivity analysis was
16 conducted to evaluate the potential for reverse causality by examining participants whose
17 serum creatinine was <1.5 mg/dL; the authors reported that longitudinal associations did
18 not materially change.
19 In modeling the association between blood Pb level and change in serum creatinine,
20 Tsaih et al. (2004) adjusted for baseline serum creatinine. Glymour et al. (Glymour et al..
21 2005) discusses how such adjustment may introduce bias. If there is no interaction
22 between Pb exposure and unmeasured causes of kidney disease, the model is linear, and
23 the slope does not change direction prior to and during the study period, the bias should
24 be to underestimate the effect. However, Glymour (2012). noted that the direction of the
25 bias is difficult to predict when the model is nonlinear or the data are restricted to a
26 specific stratum of outcome.
November 2012 5-378 Draft - Do Not Cite or Quote
-------
Study
Population
Blood Pb Median Blood Pb 10th-90th Outcome
(IQR) (ug/dL)1 percentile d
POSITIVE EFFECT ESTIMATES INDICATE POORER f UNCTION
LONGITUDINAL RESULTS
Kim et al, (1996) NAS men, blood Pb < 40 ng/dL 8.4(5.7,12.4) 4.0-17,5
NAS men, blood Pb < 25 ng/dL
NAS men, blood Pb < 10 [ig/dL
Change in Scr between visits
(mg/dL) x 10
3 9 (2 8 5 6) 21-76
Tsiah et al. (2004) NAS men with diabetes
NAS men without diabetes
NAS men with hypertension
NAS men without hypertension
CROSS-SECTIONAL RESULTS
Kim etal. (1996) NAS men, blood Pb < 40 Mg/dL 8.4(5.7,12.4) 4.0-17.5 5cr(mg/dL)
NAS men, blood Pb < 25 ug/dL
NAS men, blood Pb s 1
chs"Be '" Scr Per Vear (mg/dl)
Tsaih et al. (2004)
NAS men with diabetes
NAS men without diabetes
NAS men with hypertension
NAS men without hypertension
Baseline Blood Pb
5.5(3.7,8.1) 2.6-11.5 Scr (mg/dL)
Repeated measures Blood Pb
IIM:J men wan aidoeiei
NAS men without diabetes
NAS men with hypertension
NAS men without hypertension
DeBurbure et al. (2006) Ctech, French, Polish Children
NEGATIVE EFFECT ESTIMATES INDICATE POORER FUNCT
LONGITUDINAL RESULTS
Yu et al. (2004) CKD Patients
CROSS-SECTIONAL RESULTS
Akesson et al. (2005) Swedish Women
Staessen etal. (1992) Belgian Women
Paytonet ol(1996) NAS Men
Akesson et al. (2005) Swedish Women
Fadrowski et al. (2010) NHANES III Adolescents
3.3 (i.O, 3.OJ 4.J-/.D
3.9 (2.6, 5,7) 1.8-8.1
ION
3.2 (2.5, 4.1) 2.0-5.1
2.2 (1.7, 3.0) 1.3-3.8
7.5 (5.2, 10.9) 3.7-15 1
7.3 (5.4, 9.9) 4.1-12.9
2.2 (1.7, 3.0) 1.3-3.8
1.4 (0.7, 2.9) 0.4-5.4
Log Scr (rng/L)b •« — »
Change In GFR over A yr/100 >
(rnL/mln)
Creatinine C lea ra nee/ 1 DO
(mL/min)
Creatinine Clearance/ 100
(mL/min)
Log Creatinine Clearance
(mL/min)
GFR/100(mL/rnin) -•-
GFR/100(mL/min/1.73m2) — •—
^
-•
0.1
0.0
0.1
0.2
Change in Kidney Function Per 1 ng/dL increase in
blood Pb level within the 10th-90th percentile interval
"Blood Pb data are presented as median and (IQR) in u.g/dL for blood Pb. For uniform presentation, median and IQR were
estimated from the given distributional statistics by assuming normal distributions.
bThe cross product of logged blood Pb and ranked urine Hg was included in the regression to model the interaction between these
two variates. The significant hyperfiltrative effect to these children could be due to a biphasic time course sometimes seen in early
exposure.
Note: Results are presented first for kidney function tests where an increase is considered impaired function (black circles) then for
tests where a decrease is considered impaired function (blue circles, outlined in box). Within a category, results are presented first
for longitudinal analyses followed by cross-sectional analyses. To compare results for linear and nonlinear modeling, effect
estimates were standardized to a 1 ug/dL increase in blood Pb level within the 10th-90th percentile interval. Magnitudes of the effect
should not be compared among different kidney metrics.
Figure 5-31 Concentration-response relationships for associations between
blood Pb level or bone Pb level and kidney function outcomes.
November 2012
5-379
Draft - Do Not Cite or Quote
-------
1 The impact of Pb on the kidney has been examined in multiple NHANES datasets
2 obtained over the last few decades (Figure 5-32 and Table 5-25). NHANES data analyses
3 benefit from a number of strengths including large sample size, ability to adjust for
4 numerous potential confounding factors, and the fact that the study population is
5 representative of the U.S. non-institutionalized, civilian population. The results, covering
6 different time frames, have been consistent in providing support for Pb as a CKD risk
7 factor, including NHANES III, conducted from 1988-1994, in which adults with
8 hypertension and diabetes were observed to be potentially at-risk populations (Muntner et
9 al.. 2003) and NHANES 1999-2002 (Muntner et al.. 2005). However, because the various
10 NHANES analyses were cross-sectional in design, examining associations between
11 concurrent measures of kidney function and blood Pb levels, a common limitation is the
12 uncertainty regarding the temporal sequence between Pb exposure and renal function and
13 the magnitude, timing, frequency, and duration of Pb exposure that contributed to the
14 observed associations.
15 A recent publication examined NHANES data collected from 1999 through 2006 (Navas-
16 Acien et al.. 2009). The geometric mean concurrent blood Pb level was 1.58 ug/dL in
17 14,778 adults aged > 20 years. After adjustment for survey year, sociodemographic
18 factors, CKD risk factors, and blood Cd, the odds ratios for albuminuria (> 30 mg/g
19 creatinine), reduced eGFR (<60 mL/min/1.73 m2), and both albuminuria and reduced
20 eGFR were 1.19 (95% CI: 0.96, 1.47), 1.56 (95% CI: 1.17, 2.08), and 2.39 (95% CI:
21 1.31, 4.37), respectively, comparing the highest (>2.4 ug/dL) to the lowest (< 1.1 ug/dL)
22 blood Pb quartiles. Thus, in the subset of the population with the most severe kidney
23 disease (both reduced eGFR and albuminuria), the magnitude of association with
24 concurrent blood Pb was greater. When blood Cd was included as a covariate, blood Pb
25 remained significantly associated with renal function. In fact, the most important
26 contribution of this recent NHANES analysis was the evaluation of joint Pb and Cd
27 exposure (discussed in Section 5.5.4.1).
28 An important contribution of all NHANES publications is that they provide evidence that
29 blood Pb remains associated with reduced kidney function (<60 mL/min/1.73 m2 as
30 estimated with the MDRD equation cross-sectionally) despite steadily declining blood Pb
31 levels in the U.S. population during the time periods covered. Other studies of adults
32 participating in NHANES have also reported worse kidney function related to blood Pb
33 levels (Lai et al.. 2008a: Hernandez-Serrato et al.. 2006; Goswami et al.. 2005).
November 2012 5-380 Draft - Do Not Cite or Quote
-------
Study
Quartiles of Blood Pb Distribution Used
Muntner et al. (2003)
Elevated Serum Creatinine
Hypertensive
Normotensive
Navas-Acien et al. (2009)
Albuminuria >30 mg/g creatinine
eGFR <60 mL/min/1.73m2
AL
J 1
Al
1
1
ug/dL Blood Pb
-40 -10 10 50
% Change per ug/dL Blood Pb
Note: These results depicted are from studies that reported ORs of kidney function measures by grouping the population into
quartiles of blood Pb and then comparing each group to the quartile with the lowest blood Pb (reference group). The blood Pb
distribution of the examined group is shaded black and the reference group is shaded gray. To express these odds ratios in terms of
blood Pb concentration, a log normal distribution was fit to the statistics presented and then the medians of each group were
determined. The adjusted OR was the exponentiated quantity (log(OR) divided by the difference in the medians of the groups
compared). The resulting odds ratio is presented in terms of percent change=100*(OR-1).
Figure 5-32 Percent change in kidney outcomes across quartiles of blood Pb
level in NHANES.
November 2012
5-381
Draft - Do Not Cite or Quote
-------
Table 5-25 Additional characteristics and quantitative data for associations of blood and bone Pb with kidney
outcomes for results presented in Figure 5-31 and Figure 5-32.
Reference
Population
Study
Location;
Time Period
N
Pb Biomarker
Data
Outcome
Statistical Analysis
Effect Estimate
(95% Cl)
Results for Figure 5-31: Positive Effect Estimates Indicate Poorer Function
Longitudinal Results
Per 1 jjg'dL increase in blood Pb
within the 10th-90th percentile
interval
Adult males
Boston, MA;
Multiple
examinations
1979-1994
459 Median baseline
blood = 8.6 ug/dL
10th-90th percentile:
4.0-17.5
Change in serum
creatinine
between visits x
10(mg/dL)
Random-effects modeling
adjusted for baseline age,
time since initial visit, BMI,
smoking status, alcohol
ingestion, education level,
hypertension, baseline serum
creatinine, and time between
visits
Peak blood Pb < 40 ug/dL: 0.012
(-0.0001, 0.025)
Peak blood Pb < 25 ug/dL: 0.015
(0.0002, 0.03)
Peak blood Pb < 10 ug/dL: 0.021
(-0.005, 0.048)
Tsaih et al.
(2004)
Adult males
Boston, MA;
8/1991-1995
with mean
6 year
follow-up
448 Mean (SD) Baseline
Blood Pb = 6.5
(4.2) ug/dL
10th-90th percentile:
2.1-7.6
Tibia Pb = 21.5
(13.5)ug/g
Patella Pb = 32.4
(20.5) ug/g
Change in serum
creatinine per
year
x 10(mg/dL)
Log linear regression adjusted
forage, age squared, BMI,
hypertension, diabetes,
smoking status, alcohol
consumption, analgesic use,
baseline serum creatinine,
serum creatinine squared
With diabetes: 0.18 (0.07, 0.29)
Without diabetes: 0.014 (-0.009,
0.037)
With hypertension: 0.019 (-0.027,
0.065)
Without hypertension: 0.021
(-0.007, 0.049)
Per unit increase in In-transformed
tibia Pb
With diabetes: 0.082 (0.03, 0.14)
Without diabetes: 0.005 (-0.01,
0.02)
With hypertension: 0.023 (0.003,
0.04)
Without hypertension: 0.0004
(-0.01,0.01)
November 2012
5-382
Draft - Do Not Cite or Quote
-------
Reference Population
Study
Location;
Time Period N
Pb Biomarker
Data
Outcome
Statistical Analysis
Effect Estimate
(95% Cl)
Cross-Sectional Results
Kim et al. (1 996) Adult males
Tsaih et al. Adult males
(2004)
De Burbure et al. Children, mean
(2006) age = 1 0 years,
age
range = 8.5-12.3
years
Boston, MA; 459
Multiple
examinations
1979-1994
Boston, MA; 448
8/1991-1995
with mean 6
yr follow-up
France, 804
Czech
Republic, and
Poland; dates
not provided
Median baseline
blood = 8.6 ug/dL
10th-90th percentile:
4.0-17.5
Mean (SD) baseline
Blood
Pb = 6.5 (4.2) ug/dL
10th-90th percentile:
2.6-11 .5
Repeated measures
10th-90th percentile:
2.1-7.6
Tibia Pb = 21 5
(13.5)ug/g
Patella Pb = 32.4
(20.5) ug/g
Concurrent Blood Pb
Median (IQR) = 3.9
(2.6, 5.7) ug/dL
10th-90th percentile:
1.8-8.1
Serum creatinine
(mg/dL)
Serum creatinine
(mg/dL)
Log-transformed
serum creatinine,
cystatin C, and
p2-microglobulin
Random-effects modeling
adjusted for baseline age,
time since initial visit, BMI,
smoking status, alcohol
ingestion, education level, and
hypertension.
Log linear regression adjusted
forage, age squared, BMI,
hypertension, diabetes,
smoking status, alcohol
consumption, analgesic use
Log linear regression adjusted
for Cd, urinary creatinine,
urinary Hg
Peak blood Pb < 40 ug/dL: 0.0017
(0.0005, 0.003)
Peak blood Pb < 25 ug/dL: 0.0021
(0.0007, 0.0035)
Peak blood Pb < 10 ug/dL: 0.0033
(0.0012,0.0053)
Baseline blood Pb
With diabetes: -0.009 (-0.038,
0.020)
Without diabetes: -0.004
(-0.010, 0.003)
With hypertension: 0 (-0.013,
0.013)
Without hypertension: -0.005
(-0.011,0.002)
Follow-up blood Pb
With diabetes: 0.053 (-0.032,
0.138)
Without diabetes: 0.034 (0.007,
0.061)
With hypertension: 0.083 (0.038,
0.128)
Without hypertension: 0.014
(-0.016, 0.044)
Log serum creatinine (mg/L):
-0.062 (-0.1 06, -0.01 7)a
Log Cystatin C: -1 .3 (-2.4,
-0.21 )a
Log p2-microglobulin: -2.2
(-4.0, -0.54)3
November 2012
5-383
Draft - Do Not Cite or Quote
-------
Reference
Results for
Longitudinal
Population
Study
Location;
Time Period N
Fiqure 5-31: Neqative Effect Estimates
Results:
Yu et al. (2004) Adult CKD
patients
Taipei, 121
Taiwan;
48 month
longitudinal
study period
Pb Biomarker
Data
Indicate Poorer
Mean (SD) Baseline
blood = 4.2
(2.2) ug/dL
10th-90th percentile
2.0-5.1
Outcome
Function
Change in MDRD
eGFRover4
yr/100
(mL/min/1.73m2
body surface area)
Statistical Analysis
Cox proportional hazard
model examined whether a
predictor was associated
with renal function including
age, sex, BMI,
hyperlipidemia,
hypertension, smoking, use
of ACE inhibitor, baseline
serum creatinine, daily
protein excretion, daily
protein intake, underlying
kidney disease
Effect Estimate
(95% Cl)
Per 1 ug/dL increase
in blood Pb within the
10th-90th percentile interval
-0.040 (-0.072, -0.008)3
Cross-Sectional Results:
Akesson et al.
(2005)
Staessen et al.
(1992)
WHILA,
adult women
Adults
Sweden; 820
6/1999-1/2000
Belgium; 1,981
1985-1989
Median (5-95%)
concurrent
blood = 2.2 (1.1,
4.6) ug/dL
10th-90th percentile:
1.3-3.8
Creatinine
clearance/100
(mL/min)
Cystatin C-based
eGFR (Larsson et
al.. 2004V100
(mL/min)
Concurrent Blood Pb Creatinine
Mean (SD) clearance/100
Males:11.4ug/dL (mL/min)
Linear regression adjusted
for age, BMI, diabetes,
hypertension, regular use of
status
Log linear regression
adjusted for age, age
squared, sex, BMI, BP,
-0.01 8 (-0.03, -0.006)
-0.02 (-0.03, 0.007)
Females: -0.067
(-0.108, -0.027)3
Males- -O rWI C-D DQ7 -D ClA7\B
Payton et al.
(1994)
Adult males
Boston, MA;
1988-1991
744
Females: 7.5 ug/dL
10th-90th percentile:
3.7-15.1
Mean (SD)
concurrent
blood = 8.1
(3.9) ug/dL
10th-90th percentile:
4.1-12.9
Log-transformed
creatinine
clearance (mL/min)
ferritin level, smoking status,
alcohol ingestion,
rural/urban residence,
analgesic and diuretic use,
blood and urinary Cd,
diabetes, occupational
exposure to heavy metals,
and gamma glutamyl
transpeptidase
Log linear regression
adjusted forage, BMI,
analgesic and diuretic use,
alcohol consumption,
smoking status, SBP, DBP
-0.040 (-0.079, -0.0015)
November 2012
Draft - Do Not Cite or Quote
-------
Study
Location;
Reference Population Time Period
Fadrowski et al. NHANES, U.S.;
(2010) adolescents 19881994
Results for Figure 5-32: Analysis of Blood
Muntner et al. NHANES III, U.S.;
(2003) adults 1988-1994
Navas-Acien et NHANES III, U.S.;
al. (2009) adults 1999-2006
Pb Biomarker
N Data
769 Median concurrent
blood = 1.5 ug/dL
10th-90th percentile:
0.4-5.4
Q1: <1 .0
Q2: 1.0 to 1.5
Q3: 1.6 to 2.9
Q4: >2.9
Pb Quartiles
4813 Mean(SD)
concurrent blood Pb
With Hypertension:
4.2(0.14)ug/dL
Q1: 0.7 to 2.4
Q2: 2.5 to 3.8
Q3: 3.9 to 5.9
Q4: 6.0 to 56.0
Without
Hypertension: 3.3
(0.10) ug/dL
Q1: 0.7 to 1.6
Q2: 1.7 to 2.8
Q3: 2.9 to 4.6
Q4: 4.7 to 52.9
14,778 Geometric
concurrent blood
mean = 1.58 ug/dL
Q1: £ 1 .1
Q2: 1.2 to 1.6
Q3: 1.7 to 2.4
Q4: >2.4
Outcome
Cystatin C-based
eGFR/100
(mL/min/1.73 m2;
calculated using
the Filler and
Lepage equation)
Elevated Serum
Creatinine
(99th percentile of
each race-sex
specific distribution
for healthy young
adults)
CKD
eGFR <60
mL/minute/1.73 m2
Albuminuria and
eGFR <60
mL/minute/1 .73 m
Statistical Analysis
Log linear regression
adjusted for age, sex,
race/ethnicity, urban/rural
residence, smoking, obesity,
household income,
education level of family
reference person, BP, lipid
levels, glucose levels
Logistic regression adjusted
for age, race, sex, diabetes,
SBP, smoking, history of
CVD, BMI, alcohol
consumption, household
income, education level,
marital status, health
insurance
Logistic regression adjusted
for survey year, age, sex,
race/ethnicity, BMI,
education, smoking,
cotinine, alcohol intake,
- hypertension, diabetes,
menopausal status
Effect Estimate
(95% Cl)
-0.022 (-0.038, -0.0054)
Q1 : Referent
Q2: -1.4 (-7.4, 4.5)
Q3: -2.6 (-7.3, 2.2)
Q4: -6.6 (-12.6, -0.07)
% change in kidney outcome
Q1 : Referent
With hypertension
Q2:47%(3, 110)
Q3: 80% (34, 1 42)
Q4: 1 41 % (46, 297)
Without hypertension
Q2:11%(-44, 121)
Q3: 19% (-38, 125)
Q4:9%(-47, 122)
With hypertension
Q2:44%(0, 109)
Q3:85%(32, 159)
Q4: 1 60% (52, 345)
Without hypertension
Q2:-10% (-63, 116)
Q3:0%(-55, 122)
Q4:9%(-59, 189)
Q1 : Referent
Q2: 10% (-20, 51)
Q3: 36% (-1 , 85)
Q4: 56% (17, 108)
Q2:53%(-15, 177)
Q3:57%(-17, 198)
Q4: 139% (31, 337)
a95% Cl: estimated from given p-value.
November 2012
5-385
Draft - Do Not Cite or Quote
-------
Patient Population Studies
1 CKD as defined by the National Kidney Foundation (NKF) - Kidney Disease Outcomes
2 Quality Initiative workgroup (NKF. 2002) is the presence of markers of kidney damage
3 or GFR <60 mL/min/1.73 m2 for > 3 months. The MDRD equation is the most common
4 one used in the eGFR determination for this definition. Notably, decreased GFR is not
5 required for the first criterion and markers of kidney damage are not required for the
6 second criterion.
7 Several key studies in CKD patients provide prospective data that indicate that higher
8 baseline blood Pb level is associated with greater CKD progression over time (kidney
9 function decline) in patient populations (Table 5-26). Yu et al. (2004). discussed in the
10 2006 Pb AQCD, followed 121 patients over a four year period. Eligibility required well-
11 controlled CKD with serum creatinine between 1.5 and 3.9 mg/dL. Importantly, EDTA-
12 chelatable Pb <600 ug/72 h, a level below that traditionally thought to indicate risk for
13 Pb-related nephrotoxicity, was required at baseline. Patients with potentially unstable
14 kidney disease were excluded (i.e., due to systemic diseases such as diabetes). Mean
15 blood Pb and EDTA-chelatable Pb levels were 4.2 ug/dL and 99.1 ug/72 hours,
16 respectively. Cox proportional hazard modeling indicated lack of significant association
17 between serum creatinine changes and various potential confounding factors (Table
18 5-25). examined one at a time. Only chelatable Pb (body Pb burden indicator) was
19 significantly associated with overall risk for the primary endpoint (doubling of serum
20 creatinine over the 4-year study period or need for hemodialysis). When the group was
21 dichotomized by EDTA chelatable Pb level, Kaplan-Meier analysis demonstrated that
22 significantly more patients (15/63) in the high-normal group (EDTA chelatable Pb level >
23 80 but <600 ug/72 hours) reached the primary end point than did those in the lower
24 EDTA chelatable Pb levels (<80 ug Pb/72 hours) group (2/58). Associations between
25 baseline chelatable or blood Pb level and change in serial measurements of eGFR
26 (estimated by the MDRD equation (Levey etal. 1999)) were modeled separately using
27 generalized estimating equations. Based on these models, a 10 ug higher chelatable Pb
28 level or 1 ug/dL higher blood Pb level reduced the GFR by 1.3 and 4.0 mL/min/1.73 m2,
29 respectively, during the 4-year study period. The use of estimated GFR provides a better
30 estimate of progressive changes of renal function than creatinine clearance used in the
31 other related studies. Recent studies expanded the CKD patient populations in which this
32 effect was observed to include those with diabetic nephropathy (Lin et al.. 2006b) and
33 with the lowest blood Pb levels studied to date (Lin et al.. 2006a). Results of these
34 observational studies have been summarized in Table 5-26 (Weaver and Jaar. 2010).
November 2012 5-386 Draft - Do Not Cite or Quote
-------
Table
Study
Linet
al.
(2003)
Yuet
al.
(2004)
Linet
al.
(2006b)
Lin et
al.
(2006a)
5-26 Prospective patient population studies: kidney function decline.
Baseline
mean
(SD)
blood
Pb
n (ua/dL)
202 5.3 (2.9)
121 4.2(2.2)
87 6.5 (3.4)
108 2.9(1.4)a
"Notably, mean blood Pb level
in Baltimore, MD (Martin et al..
Baseline mean
(SD) chelatable
Pb(ug/72 hours)
104.5(106.3)
99.1 (83.4)
108.5(53.8)
40.2(21.2)
(all <80)
Baseline
mean(SD)
eGFR
(mL/min
/1.73 m2)
41.6(14.4)
36.0 (9.8)
35.1 (9.0)
47.6 (9.8)
in this study was below that observed in a
2006).
Decline in
eGFR perl SD
higher Pb dose
Years of at baseline
follow-up per year
2 0.16
4 2.7 (chelatable)
2.2 (blood Pb)
1 3.87
2 1.1
Comments
Largest study to date
Longest follow-up;
1 jxg/dL higher blood
Pb, at baseline,
associated with
4.0mL/min/1.73m2
reduction in eGFR over
4 years
Type II diabetics with
nephropathy
Lowest Pb exposed
CKD patients
recent large general population study of 50- to 70-year olds
Source: Reprinted with permission of UpToDate.com, Weaver and Jaar (2010)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
A recent population-based case-control study examined occupational Pb exposure as a
risk factor for severe CKD (Evans et al.. 2010). The study included 926 cases with first
time elevations of serum creatinine >3.4 mg/dL for men and >2.8 mg/dL for women and
998 population-based controls. Occupational Pb exposure was assessed using an expert
rating method based on job histories; no biomarkers of Pb exposure were measured. In
multivariable logistic regression modeling, the OR for CKD (adjusted for age, sex,
smoking, alcohol consumption, diabetes, education, and BMI) was 0.97 (95% CI: 0.68,
1.38) in Pb-exposed compared to non-exposed participants. In addition, the CKD patients
were followed prospectively for a mean of 2.5 years for the 70 Pb-exposed patients and
2.4 years for the 731 patients without past occupational Pb exposure. Mean eGFRs (using
the MDRD equation) were 16.0 and 16.6 mL/min/1.73 m2 in exposed and non-exposed
patients, respectively, indicating severe disease in both groups. The results overall did not
provide strong evidence that Pb exposure was associated with renal effects. The expert
ratings used in this study may have lower validity and reliability as compared to other
exposure assessment methods (Teschke et al.. 2002) including blood and bone
measurements used in the majority of well-conducted studies. Strengths included
virtually complete case ascertainment and minimal loss to follow-up. Exposure
assessment was listed as both a strength and a limitation. Expert rating methods are
commonly used when biological monitoring is not an option and in case-control studies
where many occupational exposures are considered. In Pb-kidney research, this approach
is uncommon except in the case-control setting. However, given the challenges of
November 2012
5-387
Draft - Do Not Cite or Quote
-------
1 interpreting blood Pb in dialysis patients (discussed below), this approach may have
2 advantages in this study of such severe CKD. Other case-control studies examining
3 occupational risk factors for CKD found Pb exposure to be a risk factor (Nuvts et al..
4 1995: Steenland et al.. 1990). Nuyts et al. (1995) found adults with history of
5 occupational Pb exposure to have elevated odds of CKD (OR for ever- versus never-
6 exposed: 2.11 [95% CI: 1.23, 4.36]). The association was weaker in Steenland et al.
7 (1990) (OR for ever- versus never-exposed: 1.73 [95% CI: 0.82, 3.65]). Regular
8 moonshine consumption, also a potential source of Pb exposure, was a stronger risk
9 factor for CKD (OR: 2.42 [95% CI: 1.10, 5.36]).
10 The prospective observational aspect of Evans et al. (2010) is similar in design to the
11 work of Lin and colleagues but differs in several important respects. In Evans et al.
12 (2010). only occupational Pb exposure was considered whereas the work in Taiwan
13 excluded occupational exposure and used blood and chelatable Pb measures. In the past
14 in developed countries, environmental exposures were substantial. For example, mean
15 tibia Pb levels were 21.5 and 16.7 |o,g/g bone mineral, in environmentally-exposed 50- to
16 70-year-old African-Americans and whites, respectively, in Baltimore (Martin et al..
17 2006). In Korean Pb workers, mean baseline tibia Pb level was only twofold higher (35.0
18 Mg/g) (Weaver et al.. 2003a) which illustrates the substantial body burden in middle- and
19 older-aged Americans from lifetime Pb exposure. Declines in blood Pb levels in Sweden
20 have been reported and attributed to the leaded gasoline phase-out (Stromberg et al..
21 1995: Blinder etal.. 1986). although blood Pb levels were lower than those noted during
22 the U.S. phase-out. Finally, the severe degree of CKD among subjects in Evans et al.
23 (2010) creates a survivor bias at enrollment and limits the eGFR decline possible during
24 follow-up, thus limiting the ability to identify factors that influence that decline.
ESRD Patient Studies
25 End stage renal disease (ESRD) is a well-established public health concern, and is
26 characterized by the use of dialysis to perform the normal functions of the kidney.
27 Incidence and prevalence in the U.S. continue to increase resulting in rates that are the
28 third highest among nations reporting such data (U.S. Renal Data System. 2009). Studies
29 in patients with CKD requiring chronic hemodialysis have also been published in the past
30 five years. A study of 271 adult patients on regular thrice weekly dialysis reported much
31 higher blood Pb levels than had been appreciated by the treating clinicians (Davenport et
32 al.. 2009). Blood Pb levels ranged from 3 to 36.9 ug/dL; 25.5% had levels >20 ug/dL,
33 59% had values of 10-20 ug/dL, and 15.5% were <10 ug/dL. Few details on the statistical
34 analysis were provided which complicates interpretation of the findings. However, blood
35 Pb was positively correlated with hemodialysis vintage (months on dialysis; Spearman
November 2012 5-388 Draft - Do Not Cite or Quote
-------
1 r = 0.38, p <0.001); negatively correlated with urine output (r = -0.44, p <0.001) and
2 higher in patients using single carbon filter and reverse osmosis water purification
3 devices. Another recent publication reported higher Pb in dialysate than in the tap water
4 used in its preparation (Chen et al.. 2009a). A systematic review of a wide range of trace
5 elements in hemodialysis patients reported higher Pb levels in patients compared to
6 controls although the difference was not large (Tonelli et al.. 2009). These data suggest
7 that blood Pb monitoring in dialysis patients may be useful.
8 Interpretation of blood and bone Pb in patients on dialysis is challenging for several
9 reasons. First, renal osteodystrophy, the bone disease related to kidney disease, may
10 result in increased release of Pb from bone stores. Thus, interpretation of blood and even
11 bone Pb levels may require adjustment with one or more of a range of osteoporosis
12 variables. Secondly, as observed above (Davenport et al.. 2009). residual kidney function
13 may have a substantial impact on blood Pb levels in populations with such minimal
14 excretion. Third, as illustrated in the studies cited above (Chen et al.. 2009a: Davenport et
15 al.. 2009), water and concentrates used in dialysis may be variable sources of Pb. A
16 recent study reported decreased blood Pb in post-dialysis compared to pre-dialysis
17 samples (Kazi et al.. 2008). Thus, substantial fluctuations in blood Pb are possible while
18 on dialysis. Finally, anemia is common in CKD and Pb is stored in red blood cells. Thus,
19 measurement of blood Pb in anemia may require adjustment for hemoglobin; no
20 standardized approach to this currently exists.
21 Given these caveats, a small cross-sectional pilot study observed higher median blood Pb
22 levels in 55 African-American dialysis patients compared to 53 age- and sex-matched
23 controls (6 and 3 ug/dL respectively; p <0.001) (Muntner et al.. 2007). However, median
24 tibia Pb was higher in ESRD patients although the difference did not reach statistical
25 significance (17 and 13 ug/g bone mineral, respectively [p = 0.13]). Further, the authors
26 note the limitation related to the sample size based on too few cases needed to achieve
27 statistical significance from power calculations.
28 In order to determine the potential impact of renal osteodystrophy, median blood and
29 tibia Pb levels in the dialysis patients were compared by levels of serum parathyroid
30 hormone, calcium, phosphorus, and albumin and were not found to be significantly
31 different (Ghosh-Narang et al.. 2007). A study of 211 diabetic patients on hemodialysis
32 (Lin et al.. 2008) found parathyroid hormone and serum creatinine to be associated with
33 blood Pb level in crude but not adjusted associations. In contrast, a study of 315 patients
34 on chronic peritoneal dialysis observed parathyroid hormone to be positively correlated
35 and residual renal function to be negatively correlated with logarithmic-transformed
36 blood Pb levels after adjustment (Lin et al.. 2010). In the prospective portion of this
37 study, blood Pb levels at baseline were categorized by tertile (range of 0.1 to 29.9 ug/dL
November 2012 5-389 Draft - Do Not Cite or Quote
-------
1 with cut points of 5.62 and 8.66 ug/dL). Cox multivariate analysis, after adjustment for
2 parathyroid hormone level, residual renal function, and 20 other variables, showed
3 increased all-cause mortality in the middle (5.62-8.66 ug/dL) and highest (>8.66 ug/dL)
4 compared to the lowest (<5.62 ug/dL) tertiles after 18 months of follow-up (hazard ratio=
5 2.1 [95% CI: 2.0, 2.2] and 3.3 [95% CI: 1.3, 13.5], respectively). A recent publication of
6 an 18-month follow-up of 927 patients on maintenance hemodialysis also reported
7 increased hazard ratios for all-cause (4.7 [95% CI: 1.9, 11.5]), cardiovascular-cause (9.7
8 [95% CI: 2.1, 23.3]), and infection-cause (5.4 [95% CI: 1.4, 20.8]) 18-month mortality in
9 the highest (>12.64 ug/dL) compared to the lowest tertile (<8.51 ug/dL) of baseline blood
10 Pb level, after adjustment for sex, urban residence, hemodialysis vintage, hemoglobin,
11 serum albumin, and ferritin (Lin et al.. 2011). Given other recent publications in
12 hemodialysis patients by this group, it would be valuable to examine these risks after
13 adjustment for hemoglobin A1C (Lin-Tan et al.. 2007a). and blood Cd (Yen et al.. 2011;
14 Hsu et al.. 2009a).
Clinical Trials in Chronic Kidney Disease Patients
15 Randomized chelation trials in CKD patients, uncommon in nephrotoxicant research,
16 provide unique information on the impact of Pb on the kidney. These studies have been
17 performed by Lin and colleagues at the Chang Gung Memorial Hospital in Taipei,
18 Taiwan and involve similar study designs. Initially, patients were observed to compare
19 CKD progression prior to chelation. Then, CKD patients whose diagnostic EDTA
20 chelatable Pb levels were within certain ranges (generally 60-600 |og/72 hours and thus
21 below the level commonly considered for chelation) were randomized. The treated group
22 received weekly chelation with 1 g EDTA intravenously for up to 3 months. The control
23 group received placebo infusions. In the follow-up period, chelation was repeated for
24 defined indications such as increased serum creatinine or chelatable Pb levels above
25 specified cut-offs. Placebo infusions were repeated in the controls as well. The results of
26 the most recent of these trials are summarized in Table 5-27 below.
November 2012 5-390 Draft - Do Not Cite or Quote
-------
Table 5-27 Clinical randomized chelation trials in chronic kidney disease
patients.
Baseline
mean(SD)
blood Pb
Reference Group n (ug/dL)
Lin et al. Chelated 32 6.1 (2.5)
(2003)
Control 32 5.9 (3.0)
Linetal. Chelated 15 7.5(4.6)
(2006b)
Control 15 5.9(2.2)
Linetal. Chelated 16 2.6(1.0)a
(2006a)
Control 16 3.0(1.1)
Lin-Tan et Chelated 58 5.0 (2.2)
al. (2007b)
Control 58 5.1 (2.6)
Baseline mean(SD)
chelatable Pb (ng/72 hr)
150.9(62.4)
144.5(87.9)
148.0(88.6)
131.4(77.4)
43.1 (13.7)
47.1 (15.8)
164.1 (111.1)
151.5(92.6)
aNotably, mean blood Pb level in this study was below that observed in a
olds in Baltimore, MD (Martin et al.. 2006).
Baseline Months
mean(SD) of
eGFR treatment
(mL/min / follow-
/1.73m2) up
32.0(12.1) 27
31.5(9.0)
22.4(4.4) 15
26.3 (6.2)
41.2(11.2) 27
42.6 (9.7)
36.8(12.7) 51
36.0(11.2)
recent large general population
Change
in eGFR
peryr
(mL/min
/1.73m2)
+ 1.07
-2.7
-3.5
-10.6
+3.0
-2.0
-0.3
-2.9
Comments
Subjects
with Type II
diabetes
and
nephropathy
Lowest Pb
exposed
and treated
range
Body Pb
Burden (72
h urinary Pb
excretion)
> 20- <80
^9
Subjects
without
diabetes
study of 50- to 70-year
1
2
3
4
5
6
7
10
11
12
13
14
15
The unique body of work in patient populations by Lin and co-workers, both
observational and experimental, has numerous strengths including prospective study
design, randomization, Pb assessment that includes estimates of the bioavailable dose,
longitudinal statistical analysis, and control for multiple kidney risk factors. However, the
generalizability of the results to broader populations is unknown. In addition, the
association observed between Pb dose and decline in GFR has been variable; the annual
decline in eGFR per standard deviation higher Pb dose at baseline was much lower in the
2003 study than in subsequent publications (Table 5-27 above). Small sample sizes and
differences in renal diagnoses between groups may be factors in this variability.
The studies presented in Table 5-26 and Table 5-27 have a number of potential
limitations. These include small sample size and lack of blinding and placebo control
except for Lin et al. (2003). which attempted to address this potential limitation. Another
possible limitation may be the shorter follow-up time in some of the studies. The use of
creatinine clearance to assess changes in renal function may limit interpretation of results
as discussed in Section 5.5.1.1. Also, the effects observed following chelation therapy
November 2012
5-391
Draft - Do Not Cite or Quote
-------
1 may result from removal of other ions such as Zn, Cu, and Fe. In addition, changes in
2 kidney function after treatment with Pb chelating agents may be by mechanisms other
3 than reduction in Pb body burden. Chelating agents have been shown to act as
4 antioxidants. DMSA abolished reactive oxygen species formation (i.e., MDA and
5 nitrotyrosine in interlobular arteries) and was protective against nonPb-induced
6 nephrosclerosis in rats (Gonick et al.. 1996). EDTA administration enhanced endothelial
7 NO production and reduced kidney damage in a rat model of ischemia-induced acute
8 renal failure (Foglieni et al.. 2006). Improved renal function following administration of
9 chelating agents have been reported in rodent models of Pb-induced nephrotoxicity
10 (Sanchez-Fructuoso et al., 2002a; Sanchez-Fructuoso et al., 2002b; Khalil-Manesh et al.,
11 1992a). Chelation did not appear to improve Pb-induced structural damage (Khalil-
12 Manesh et al., 1992a); again suggesting that improved hemodynamics may be a result of
13 reduction in reactive oxidant species, which could be due to reduced Pb level and/or
14 directly to the chelating agent (Gonick et al.. 1996). Despite these uncertainties and
15 limitations, the most prudent explanation for the combination of the observational and
16 experimental chelation work of Lin and colleagues is that reduced Pb is the underlying
17 reason for improved kidney function. This study design requires replication in larger
18 populations at multiple clinical centers to confirm that the change in renal function may
19 be due to removal of Pb.
Occupational Studies
20 The vast majority of studies in the literature on the impact of Pb on the kidney have been
21 conducted in the occupational setting. In general, study size and extent of statistical
22 analysis are much more limited than those in general population studies. Publications in
23 few populations have reported adjusted results in occupationally exposed workers in the
24 five years since the 2006 Pb AQCD. In a two-year prospective cohort study, generalized
25 estimating equations were used to model change in kidney function between each
26 evaluation in relation to tibia Pb and concurrent change in blood Pb in 537 current and
27 former Pb workers (Weaver et al., 2009). Tibia Pb was evaluated at the beginning of each
28 follow-up period (yearly on average) and Pb biomarker levels were adjusted for baseline
29 levels and other covariates. In males, serum creatinine decreased and calculated
30 creatinine clearance increased over the course of the study; these changes were largest in
31 participants whose blood Pb declined concurrently or whose tibia Pb was lower at the
32 beginning of the follow-up interval. In females, decreasing serum creatinine was
33 associated with declining blood Pb (as in males); however, increasing blood Pb was
34 associated with a concurrent increase in serum creatinine. Women (25.9% of the study
35 population) were older and more likely to be former Pb workers than were men which
36 may have been important factors in the effect modification observed by sex.
November 2012 5-392 Draft - Do Not Cite or Quote
-------
1 Chia and colleagues observed a significant, positive association between concurrent
2 blood Pb and urine NAG in linear regression models after adjustment for age, sex, race,
3 exposure duration, ALAD G177C polymorphism and the interaction between ALAD
4 genotype and blood Pb (Chiaetal.. 2006). Similar positive associations were observed
5 between blood Pb and a wider range of EBE markers in models that adjusted for age, sex,
6 race, exposure duration, and the HpyCH4 ALAD polymorphism (Chia et al.. 2005).
7 Other studies published in the last 5 years also focused on ALAD polymorphisms but did
8 not find effect modification to be in a consistent direction (Gao etal. 2010a: Wang et al..
9 2009a: Weaver et al.. 2006; Weaver et al.. 2005b). In adults with the ALAD2 genotype,
10 Pb has been associated with better and poorer renal function in separate cohorts of Pb
11 workers.
12 Two studies of occupationally-exposed adults have performed benchmark dose
13 calculations for the effect of Pb on the kidney. Both used only EBE markers and found
14 NAG to be the most sensitive outcome; reported lower confidence limits on the
15 benchmark doses were 10.1 ug/dL (Sun et al., 2008b). and 25.3 ug/dL (Lin and Tai-yi.
16 2007).
17 A number of other publications in the five years since the 2006 Pb AQCD, have reported
18 significantly worse kidney outcomes in unadjusted analyses in occupationally-exposed
19 adults compared to unexposed controls (Onuegbu et al.. 2011; Patil et al.. 2007) and/or
20 significant correlations between higher levels of Pb biomarkers and worse kidney
21 function (Alasiaet al.. 2010; Khan et al.. 2008; Garcon et al.. 2007; Lin and Tai-yi. 2007;
22 Alinovi et al.. 2005). A study of 155 male workers reported significant, positive
23 correlations between blood and urine Pb and urine NAG and albumin after controlling for
24 age and job duration (Sun et al.. 2008b). One small study found no significant differences
25 (Orisakwe et al.. 2007). In a study of 108 Pb workers with mean blood Pb level of
26 36.2 (ig/dL, no significant correlations were observed between blood Pb concentration
27 and GFR, creatinine clearance, uric acid clearance or uric acid excretion fraction
28 (Karimooy et al.. 2010). However, interpretation of this study is limited by the fact that
29 "only 30 subjects had a correct 24 hours urine volume" and no methods are described for
30 kidney outcome measurement or analysis.
31 Overall, the occupational literature published in the last five years on the kidney impact
32 of Pb exposure has been more consistent in reporting statistically significant associations
33 than were data reviewed for the 2006 Pb AQCD. This may reflect increased reliance on
34 EBE markers as more sensitive outcome measures, publication bias, or multiple
35 comparisons due to a greater number of outcomes assessed.
November 2012 5-393 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
In a study of Korean Pb workers, Weaver et al., (2003a) reported inconsistent results with
higher Pb measures associated with worse renal function in some models and better renal
function in other models. In models of effect modification by age, a pattern emerged in
which higher Pb exposure and dose measures were associated with worse renal function
in older workers and better renal function in younger workers (Weaver et al., 2003a).
A small number of publications that include concentration-response information provides
evidence of Pb-related nephrotoxicity in the occupational setting across the blood Pb
ranges analyzed (Weaver et al., 2003a; Ehrlich et al., 1998). Data in 267 Korean Pb
workers in the oldest age tertile (mean age = 52 years) did not provide evidence of a
threshold for a Pb effect on serum creatinine levels (added variable plot shown in Figure
5-33) (Weaver et al., 2003a). It is important to note the uncertainty regarding whether the
concentration-response information provided in these studies applies to lower blood Pb
levels or to populations with lower current environmental Pb exposures.
O) ...
E, ^
CL>
•-£= o
CO ^
o
E
CO
O
CD
O
0
20
40
60
Adjusted blood Pb level (|jg/dL)
Note: Both the adjusted regression line (straight line) and the line estimated by the smoothing method of the S-PLUS statistical
software function lowess (line with curves) are displayed. Both have been adjusted for covariates. For ease of interpretation, axes
have been scaled, so that the plotted residuals are centered on the means, rather than zero.
Source: Reprinted with permission of the BMJ Publishing Group, Weaver et al. (2003a)
Figure 5-33 Added variable plot of association between serum creatinine and
blood Pb in 267 Korean Pb workers in the oldest age tertile.
November 2012
Draft - Do Not Cite or Quote
-------
1 A major challenge in interpretation of the occupational literature is the potential for
2 Pb-related hyperfiltration. Hyperfiltration involves an initial increase in glomerular
3 hypertension which results in increased GFR. If persistent, the risk for subsequent CKD
4 increases. This pattern has been observed in diabetes, hypertension, and obesity (Nenov
5 et al.. 2000). As discussed in the 2006 Pb AQCD (U.S. EPA. 2006b). findings consistent
6 with hyperfiltration have been observed in occupational populations (Weaver et al..
7 2003a; Hsiao et al., 2001; Roels et al.. 1994). a study of adults who were Pb poisoned as
8 children (Hu. 1991). and a study in European children (de Burbure et al.. 2006).
9 Longitudinal data in Pb-exposed rodents provide evidence of a hyperfiltration pattern of
10 increased, followed by decreased GFR, associated with Pb exposure and are critical in
11 interpretation of the human Pb-kidney literature (Khalil-Manesh et al.. 1992b). Pb could
12 induce glomerular hypertension resulting in hyperfiltration by several mechanisms
13 including increased ROS, changes in eicosanoid levels, and/or an impact on the renin-
14 angiotensin system (Vaziri. 2008b; Roels et al.. 1994). Whether hyperfiltration
15 contributes to pathology in humans is unclear; longitudinal studies are needed.
16 The 2006 Pb AQCD provided several explanations for this inconsistency (U.S. EPA.
17 2QQ6b) (Chapter 6, pp 99):
"Some are unique to the occupational literature, such as smaller sample sizes. In addition,
employed workers are typically healthier and younger than the general population—
resulting in the healthy worker bias. This is a particular problem as susceptible risk groups
are identified. Survivor bias in cross-sectional studies is also a concern, since workers
whose renal function has declined are generally removed from exposure, particularly if
they are followed in a medical surveillance program. Few studies have included former
workers. Also, statistical analyses have been more limited in occupational studies.
Analyses for some outcomes were limited to comparisons between exposed workers and
controls whose Pb levels were in the range associated with adverse renal outcomes in
environmental work. Use of multiple linear regression has generally involved more limited
adjustment for covariates than in most of the environmental studies. Many of these
limitations result in bias towards the null, which increases the risk that true associations
may not be detected."
18 Regardless, significant findings could be obscured if opposite direction associations are
19 present in different segments of the study population and interaction models are not
20 performed to address this. In the Korean Pb workers (Weaver et al.. 2003a; Weaver etal..
21 2003b). significant associations in opposite directions were observed only when relevant
22 effect modifiers such as age or genetic variants in ALAD, VDR, and NOS were included
23 in the model. This is a valid concern for risk assessment, since the factors involved in
24 these inverse associations in Pb-exposed workers are not well defined at present.
November 2012 5-395 Draft - Do Not Cite or Quote
-------
5.5.2.2 Epidemiology in Children
Pb Nephrotoxicity in Children
1 Both the 2006 and 1986 Pb AQCDs noted that the degree of kidney pathology observed
2 in adult survivors of untreated childhood Pb poisoning in the Queensland, Australia
3 epidemic (Inglis et al., 1978) has not been observed in other studies of childhood Pb
4 poisoning. Recent publications remain consistent with that conclusion; a recent study
5 observed an impact of childhood Pb poisoning on IQ but not kidney outcomes (Coria et
6 al., 2009). Chelation was raised as a potential explanation for this discrepancy in the
7 2006 Pb AQCD.
8 With declining Pb exposure levels, recent work has focused on studies in children with
9 much lower blood Pb levels. However, insensitivity of the clinical kidney outcome
10 (i.e., GFR) measures for early kidney damage is a particular problem in children who do
11 not have many of the other kidney risk factors that adults do, such as hypertension and
12 diabetes. As a result, such studies have utilized EBE markers. However, data to
13 determine the predictive value of such biomarkers for subsequent kidney function decline
14 in Pb exposed populations are extremely limited (Coratelli et al.. 1988) and may pose
15 particular challenges in children due to puberty-related biomarker changes (Sarasua et al.,
16 2003). The few studies included the 2006 Pb AQCD (U.S. EPA. 2006b) that analyzed
17 clinical kidney outcomes in children found associations with indicators of Pb exposure
18 that were inconsistent in direction. Pels et al. (1998) found no difference in mean serum
19 creatinine between 62 children living near Pb-producing factories and 50 control children
20 living in communities without Pb emission sources. In a study of 200 Belgian adolescents
21 aged 17 years, higher concurrent blood Pb level was associated with higher serum
22 cystatin-C (de Burbure et al.. 2006); however, among 300-600 European children (n
23 varied by outcome), higher concurrent blood Pb level was associated with lower serum
24 creatinine and cystatin C (Staessen et al.. 2001).
25 Recent studies of children with elevated Pb exposure did not consistently indicate that Pb
26 exposure was associated with reduced kidney function. A study in 123 children of
27 workers in Pakistani Pb smelters and battery recycling plants and 123 control children,
28 ages 1-6 years, reported elevated blood Pb levels, serum creatinine and urea in children of
29 Pb-exposed workers compared to controls (medians: 8.1 versus 6.7 (ig/dL; 56 versus
30 52 (iM; and 4,500 versus 4,300 (iM, respectively (p < 0.01 for all) in unadjusted analyses
31 (Khan et al.. 2010a). Blood Pb levels were correlated with serum creatinine (Spearman
32 r = 0.13; p = 0.05). However, a study of 77 participants, ages 10-25 years, who were
33 previously Pb poisoned through contaminated flour and chelated, reported no difference
34 in renal effects between children with blood Pb levels >48 (ig/dL and <43 (ig/dL although
November 2012 5-396 Draft - Do Not Cite or Quote
-------
1 lower IQ was observed in the subset who were exposed before the age of six years (Coria
2 et al.. 2009).
3 One of the key gaps identified in the 2006 Pb AQCD (U.S. EPA. 2006b) was limited data
4 in children and adolescents particularly with respect to GFR measures and in populations
5 without the elevated Pb exposure associated with Pb poisoning, living near a Pb source,
6 or having parents with occupational Pb exposures. A recently published NHANES
7 analysis in adolescents begins to fill this gap (Fadrowski et al.. 2010). Associations
8 between concurrent blood Pb and kidney function were investigated in 769 adolescents
9 aged 12-20 years in the U.S. NHANES III, conducted 1988-1994. Kidney function was
10 assessed with two eGFR equations. One utilized serum cystatin C and the other used the
11 more traditional marker, serum creatinine. Median concurrent blood Pb and cystatin C-
12 based eGFR levels were 1.5 ug/dL and 112.9 mL/min/1.73 m2, respectively. Cystatin C-
13 based eGFR was lower (-6.6 mL/min/1.73 m2 [95% CI: -0.7, -12.6]) in participants with
14 blood Pb levels in the highest quartile (> 3.0 ug/dL) compared with those in the lowest
15 (<1 ug/dL). A doubling of blood Pb level was associated with a -2.9 mL/min/1.73 m2
16 (95% CI: -0.7, -5.0) lower eGFR. In contrast, the association between blood Pb and
17 creatinine-based eGFR, although in the same direction, was not statistically significant.
18 As these children were born between 1968 and 1982, some likely had higher Pb
19 exposures in earlier childhood, although notably, not as high or as long in duration as did
20 older adults examined in aforementioned studies. Nonetheless, in this study of NHANES
21 adolescents, there also is uncertainty regarding the magnitude, timing, frequency, and
22 duration of Pb exposure that contributed to the observed associations. Additional research
23 in children is warranted, in particular studies with longitudinal follow-up, multiple
24 outcome assessment methods, and examination of children born after Pb was banned
25 from gasoline.
5.5.2.3 Associations between Pb Dose and New Kidney Outcome
Measures
26 As noted above, in an effort to more accurately estimate kidney outcomes, new equations
27 to estimate GFR based on serum creatinine have been developed, and the utility of other
28 biomarkers, such as cystatin C, as well as equations based on them, are being studied.
29 However, few publications have utilized these state-of-the-art techniques when
30 evaluating associations between Pb or Cd dose and renal function. In addition to the
31 study in NHANES adolescents discussed above (Fadrowski et al.. 2010). a cross-
32 sectional study of Swedish women reported that higher concurrent blood Pb (median:
33 2.2 (ig/dL) and Cd (median: 0.38 (ig/L) levels were associated with lower eGFR based on
34 serum cystatin C alone (without age, sex, and race) after adjustment for socio-
November 2012 5-397 Draft - Do Not Cite or Quote
-------
1 demographic and CKD risk factors (Akesson et al., 2005). Associations were comparable
2 to those using creatinine clearance as the kidney outcome for Pb; however associations of
3 Cd dose measures were stronger for the cystatin C based outcome. Staessen et al. (2001)
4 found a statistically significant association between concurrent blood Pb level and serum
5 cystatin C in a cross-sectional study of adolescents; creatinine-based measures were not
6 reported. However, in a cross-sectional study of 804 European children aged range 8.5 to
7 12.3 years, higher concurrent blood Pb levels were associated with lower serum cystatin
8 C and creatinine; these inverse associations were attributed to hyperfiltration (de Burbure
9 et al.. 2006). A recent publication compared associations of blood Pb and eGFR using the
10 traditional MDRD equation to those with four new equations: CKD-EPI, and cystatin C
11 single variable, multivariable, and combined creatinine/cystatin C, in 3,941 adults who
12 participated in the 1999-2002 NHANES cystatin C subsample (Spector et al.. 2011).
13 Similar to the NHANES adolescent analysis, associations with the cystatin C outcomes
14 were stronger. After multivariable adjustment, differences in mean eGFR for a doubling
15 blood Pb were -1.9 (95% CI: -3.2, -0.7), -1.7 (95% CI: -3.0, -0.5), and -1.4 (95% CI: -2.3,
16 -0.5) mL/min/1.73 m2, using the cystatin C single variable, multivariable and combined
17 creatinine/cystatin C equations, respectively, reflecting lower eGFR with increased blood
18 Pb. The corresponding differences were -0.9 (95% CI: -1.9, 0.02) and -0.9 (95% CI: -1.8,
19 0.01) using the creatinine-based CKD-EPI and MDRD equations, respectively.
5.5.2.4 Reverse Causality
20 The reverse causality hypothesis suggests that the associations between blood Pb and
21 kidney function may be due to reduced excretion of Pb rather than a causal association
22 between Pb exposure and this outcome. Cross-sectional studies of populations that
23 include participants with CKD frequently note the potential for their findings to be
24 explained by reverse causality as a limitation of the study (Muntner et al.. 2003). There
25 are several techniques that can be used to assess the potential for reverse causality to
26 underlie associations between higher Pb dose and worse kidney function. Prospective
27 studies in which associations between baseline measurements of Pb biomarkers and
28 subsequent changes in renal function are demonstrated provide the strongest evidence to
29 evaluate the possibility of reverse causality. In the NAS, baseline blood Pb levels were
30 associated with subsequent declines in renal function over follow-up periods ranging
31 from 3 to 6 years (Tsaih et al.. 2004; Kim et al.. 1996). Prospective data in CKD patients
32 also revealed an association between baseline Pb dose and decline in eGFR over follow-
33 up periods as long as four years (Yu et al.. 2004). Another approach involves sensitivity
34 analyses in which associations are explored in participants with normal glomerular
35 filtration. This approach has been used in the NAS with plots which revealed that the
November 2012 5-398 Draft - Do Not Cite or Quote
-------
1 association between blood Pb level and serum creatinine was present across the entire
2 range of serum creatinine levels, including those in the normal range where excretion is
3 not impaired (Tsaih et al., 2004; Kim et al., 1996). Analyses restricted to the population
4 with serum creatinine below 1.5 mg/dL were conducted in a later publication; the authors
5 reported that associations were consistent (Tsaih et al., 2004). The use of a serum
6 creatinine rather than an eGFR cut-off is a limitation since there can be substantial
7 decrements in renal function with 'normal' serum creatinine. The associations observed
8 in both NAS studies were not limited to the segment of the population with potentially
9 clinically significant renal dysfunction in whom reduced Pb excretion would be more
10 likely.
11 Research in which EDTA chelation decreases body Pb burden and improves kidney
12 function also provides evidence informing the possibility of reverse causality but is not
13 without limitation because EDTA may independently improve kidney function (Lin et al..
14 2006b; Lin et al.. 2006a; Lin et al.. 2003). Additional evidence evaluating reverse
15 causality was provided by findings among 153 adults with chronic kidney disease, in
16 which renal failure was not associated with increases in blood or bone Pb levels or
17 chelatable Pb levels (Van de Vvver et al.. 1988). Batuman et al. (1983) found that
18 chelatable Pb levels were similar in 27 adults with renal disease of unknown and known
19 non Pb-related causes where bone Pb levels (group means: 18 and 19 (ig/g) are in the
20 range of those measured in recent epidemiologic studies. A pilot study of 55 ESRD cases
21 at Tulane clinics (Muntner et al., 2007) reported the median blood Pb level was
22 significantly higher among the ESRD cases compared to their control counterparts. For
23 ESRD patients the distribution of blood Pb was shown by pre-defined levels: 18.5% less
24 than 5 ng/dl; 66.7% of ESRD cases had blood Pb levels between 5 and 9.9 (ig/dL; and
25 14.8% equal or greater 10 (ig/dL.
26 The reverse causality hypothesis can be broadened to include a physiologic process in
27 which Pb levels are influenced over the entire range of kidney filtration rates. If this
28 occurs, even normal kidney function would impact blood Pb levels such that higher GFR
29 would result in greater Pb excretion and lower blood Pb levels. Serum creatinine levels
30 are influenced in this way over the entire range of kidney function; as a result, these
31 levels are used to estimate kidney function. However, creatinine is produced and excreted
32 at a steady state in the body which is one reason it was selected as a biomarker to assess
33 kidney function. This expanded hypothesis implies that low blood and bone Pb levels
34 may reflect kidney function in addition to exposure. If so, this would increase
35 misclassification bias, with Pb biomarkers reflecting both exposure and kidney function.
36 Given the longstanding use of blood Pb as a dose marker in research for many non-
37 kidney outcomes this seems unlikely. Thus, published research has not directly addressed
38 this. One such approach involves comparing associations of blood and urine Pb in models
November 2012 5-399 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
of kidney function. If they are consistent, this hypothesis is not valid. However, urine Pb
is rarely used and may not be as reliable a biomarker as blood Pb (Gulson et al.. 1998c).
In summary, several lines of evidence support that reverse causality does not contribute
substantially to associations between higher blood Pb levels and worse kidney function.
These lines of evidence include prospective data observing that baseline Pb measures are
associated with subsequent declines in renal function, that associations in prospective
studies persist among adults with normal renal function, that renal failure does not
increase Pb biomarker levels and that reduction of Pb levels by chelation improves
kidney function. However, this bidirectional relationship is still possible and additional
research is needed to fully exclude the hypothesis. In particular prospective data are
required as is research to determine if normal kidney function influences blood Pb levels.
12
13
14
15
16
17
5.5.2.5 Toxicology
In animals, Pb has been found to induce changes in a wide range of indicators of renal
function. Most studies examined Pb exposure concentrations that resulted in higher blood
Pb levels (>20 (ig/dL) than those in the current U.S. general population. While
toxicological information on renal dysfunction with blood Pb levels <10 (ig/dL generally
is not available, dysfunction in kidney function measures, including urinary flow, ALP,
microalbumin, and NAG, was observed at blood Pb levels above 20 (ig/dL.
Table 5-28 Animal toxicological studies reporting the effects of Pb exposure
(as blood Pb level) on kidney function.
Reference
Berrahal et
al. (2011)
Masso-
Gonzalez
etal.
(2009)
Roncal et
al. (2007)
Species;
Lifestage;
Sex
Rat; Adult
Rat;
Weanling
pups
Rat; Adult;
Male
Pb Dose;
Exposure
Duration
50 ppm
Pb acetate in
drinking water;
lactation day 1
to PND40or
PND65
300 ppm
Pb acetate in
drinking water;
GD1 to PND21
150 ppm
Pb acetate in
drinking water;
16 weeks with
remnant kidney
surgery at week
4
Blood Pb
Level with
Response
(ug/dL)
PND40: 12.7
PND65: 7.5
23
26
Responses
Oxidative stress - Increased renal MDA (PND40 and PND65),
decreased renal SOD activity (PND40)
Morphology- Increased relative kidney weight (PND65)
Kidney function - Increased blood creatinine (PND40 and
PND65), increased BUN (PND40), decreased uric acid
(PND65), increased kidney proteins (PND65).
Oxidative stress - Elevated TEARS and catalase activity
Morphology- Elevated relative kidney weight at PND21
Inflammation - Increased the number of macrophages & renal
MCP-1 mRNA.
Morphology - Pb induced pre-glomerular vascular disease of
kidney (i.e., sclerosis, fibrosis, peritubular capillary loss)
Kidney function - Decreased creatinine clearance, increased
serum creatinine, increased BUN, and increased serum uric
acid.
November 2012
5-400
Draft - Do Not Cite or Quote
-------
Reference
Khalil-
Manesh et
al. (|993a)
Khalil-
Manesh et
al. (1992a)
Vyskocil et
al. (1995)
Ademuyiwa
etal.
(2009)
Navarro-
Moreno et
al. (2009)
Khalil-
Manesh et
al. (1992b)
Wang et al.
(201 Od)
Species;
Lifestage;
Sex
Rat; Adult;
Male
Rat; Adult;
Male
Rat, Adult,
Female
Rat; Adult
Rat; Adult;
Male
Rat; Adult;
Male
Rat; Adult;
Female
Pb Dose;
Exposure
Duration
100 ppm
Pb acetate in
drinking water;
12 months
5,000 ppm
Pb acetate in
drinking water;
1, 6, or
9 months
1,000 ppm
Pb acetate in
drinking water; 2
or 4 months
200, 300, and
400 ppm
Pb acetate in
drinking water;
12 weeks
500 ppm
Pb acetate in
drinking water;
28 weeks
5,000 ppm
Pb acetate in
drinking water;
12 months
300 ppm
Pb acetate in
drinking water;
8 weeks
Blood Pb
Level with
Response
(ug/dL)
Mean at
3 months
29.4
Mean at
12 months
22
Range 9-34
At 1 month
7.9
At 6 months
-30
At 9 months
52
37.6
200 ppm: 41
300 ppm: 61
400 ppm: 39
43
Max 125
Mean 45
20 (serum)
Responses
Morphology - Mild tubular atrophy and interstitial fibrosis seen
at 12 months, otherwise normal.
Kidney function -Increased GFR at 1 and 3 months, increased
NAG, and no change in GST.
At 1 month
Kidney function - no functional or pathological changes
At 6 months
Morphology - Prominent tubulointerstitial fibrosis and
segmental sclerosis. Increased kidney weight
Kidney function - Decreased GFR, increased serum creatinine
and SUN, no change urinary NAG or GST
At 9 months
Morphology - Severe tubulointerstitial disease
Kidney function - Decreased GFR, increased serum creatinine
and SUN
Kidney function - No change in kidney function or
nephrotoxicity
Kidney function - Renal phospholipidosis and depletion of
renal cholesterol.
Oxidative stress - Increased kidney lipid peroxidation
(i.e., TEARS)
Morphology - Electron micrography showed lumen reduction,
microvilli loss, brush border loss, and mitochondrial damage
Kidney function - Elevated urinary pH and protein, and glucose
and blood in the urine.
Morphology - Tubular atrophy and interstitial fibrosis after
6 months. Increased urinary brush border antigens.
Kidney function - Hyperfiltration at 3 months and decreased
GFR at 12 months. After 3 months, elevated urinary NAG and
GST.
Biomarker- Aberrant NAG, GGT, p2-microgobulin expression
Oxidative Stress- Increased lipid peroxidation (i.e., MDA
production), elevated kidney antioxidant enzymes (SOD, GPx,
CAT), and depleted GSH
Morphology - Electron micrography showed Pb damages
mitochondria, basement membrane, and brush border in
kidney tissue. Some focal tubal necrosis observed.
Kidney function - Elevated urinary total protein, urinary
albumin, and serum urea nitrogen.
1
2
o
J
4
5
Renal Function and Interstitial Fibrosis
Past studies have shown that chronic continuous or repeated Pb exposure can result in
interstitial nephritis and focal or tubular atrophy. A series of studies on Pb exposure in
rats (longitudinal 12-month exposure study to either 100 ppm or 5,000 ppm Pb in
drinking water) report an initially elevated GFR, consistent with hyperfiltration, and renal
hypertrophy (Khalil-Manesh et al.. 1993a: Khalil-Manesh et al.. 1992b: Khalil-Manesh et
November 2012
5-401
Draft - Do Not Cite or Quote
-------
1 al.. 1992a). After 6 months of exposure, GFR decreased, albuminuria was present, and
2 pathology ensued with focal tubular atrophy and interstitial fibrosis formation. This
3 pathology and functional decrement was persistent out to 12 months, and at 12 months
4 glomeruli developed focal and segmental sclerosis.
5 The toxicological evidence for differences in GFR according to duration of Pb exposure
6 (i.e., hyperfiltration with 3-month exposure versus decreased GFR with 6- or 12-month
7 exposure) provides biological plausibility for epidemiological studies that observed a
8 similar phenomenon by age in adults in association with Pb biomarker levels. However,
9 the toxicological studies report blood Pb levels at some exposure durations much higher
10 than relevant to blood Pb levels in current human populations. Still, these duration-
11 dependent dichotomous changes in GFR are consistent between the toxicological and
12 epidemiologic literature.
13 At exposure concentrations resulting in blood Pb levels within one order of magnitude of
14 the upper range of current human population blood Pb level, animal toxicological studies
15 present inconsistent results for the effects of Pb on kidney function. There are studies that
16 have corroborated the previously observed increase in serum creatinine following Pb
17 exposure in rats. Berrahal et al. (2011) reported on the effects of age-dependent exposure
18 to Pb on nephrotoxicity in male rats (Table 5-29). Pups were exposed to Pb lactationally
19 (as a result of dams consuming water containing 50 ppm Pb acetate) until weaning.
20 Thereafter, the offspring were exposed to the same solution from weaning (day 21) until
21 sacrifice. Male pups were sacrificed at age 40 days (puberty; blood Pb level 12.7 ug/dL)
22 and at age 65 days (post-puberty; blood Pb level 7.5 ug/dL). Serum creatinine was
23 elevated at both 40 days and 65 days (0.54 and 0.60 mg/dL compared to control values of
24 0.45 mg/dL [p <0.001]). Various parameters of Pb-induced renal dysfunction are listed in
25 Table 5-29 below. The elevated serum creatinine in the Pb-exposed animals compared to
26 controls suggests that animals exposed to low dose (i.e., 50 ppm) Pb from birth may
27 develop renal abnormalities. However, the lack of measurements of GFR or renal
28 pathology weakens the conclusions.
November 2012 5-402 Draft - Do Not Cite or Quote
-------
Table 5-29 Indicators of renal damage in male rats exposed to 50 ppm Pb for 40
and 65 days, starting at parturition.
Biomarker
(Mean ± SD)
Blood Pb level (|jg/dL)
Plasma Creatinine (mg/L)
Plasma Urea (mg/L)
Plasma Uric Acid (mg/L)
PND40 Control
1.8 ±0.33
4.5 ±0.21
0.37 ±0.01 9
7.51 ± 0.44
PND40 Pb
12.7± 1.7
5.35 ± 0.25a
0.47 ± 0.021 a
7.65 ± 0.32
PND65 Control
2.1 ±n0.35
4.55n± 0.27
0.29n± 0.009
9.39n± 0.82
PND65 Pb
7.5n± 0.78
6.04 ± 0.29a
0.29n± 0.009
5.91n±0.53a
ap <0.001
Source: Modified with permission of John Wiley & Sons, Berrahal et al. (2011)
1 Roncal et al. (2007) found that Pb accelerated renal function decrements,
2 tubulointerstitial injury, and arteriolopathy in non-Pb-related CKD. Sprague-Dawley rats
3 were administered Pb acetate at 150 ppm for 4 weeks, then subjected to remnant kidney
4 surgery (left kidney mass reduced by 2/3 and right kidney removed), and subsequently
5 exposed to Pb for an additional 12 weeks resulting in a blood Pb level of 26 (ig/dL.
6 Pb-treated rats had higher systolic BP, increased serum creatinine, lower creatinine
7 clearance, and higher proteinuria than did controls. Most striking was development of
8 worse arteriolar disease, peritubular capillary loss, tubulointerstitial damage, and
9 macrophage infiltration. Pb treatment was associated with significant worsening of pre-
10 glomerular vascular disease, as characterized by an increase in the media-to-lumen ratio.
11 There was also a higher percentage of segmental sclerosis within glomeruli and a
12 tendency for a higher number of sclerotic glomeruli. Additionally, a loss of peritubular
13 capillaries, as reflected by a reduction in thrombomodulin staining, was observed. This
14 was associated with worse tubular injury (osteopontin staining) due to more interstitial
15 fibrosis (type III collagen staining) and a greater macrophage infiltration in the
16 interstitium. The increase in macrophages was associated with higher renal MCP-1
17 mRNA. As a whole, these findings indicate that Pb exposure concomitant with existing
18 renal insufficiency due to surgical kidney resection accelerated vascular disease and
19 glomerular pathology. These findings are consistent with the previous work of Bagchi
20 and Preuss (2005) also showing that Pb-exposed animals with non-Pb-related CKD
21 (remnant surgery) had kidney dysfunction including impairment of the renin-angiotensin
22 system (Losartan challenge), elevated systolic BP, and alterations in renal excretion of
23 Pb, K+, and Na+. Thus, this model shows that Pb exposure may exacerbate pre-existing
24 underlying kidney disease.
25 Other investigators have shown that chronic Pb exposure has detrimental effects on renal
26 function at higher blood Pb levels. A number of studies report increased serum creatinine
27 following high level Pb exposure (e.g., blood Pb levels >55.6 (ig/dL) (Abdel Moneim et
28 al..2011b: Ozsov et al.. 2010: Javakumaretal.. 2009: Kharoubi et al.. 2008a). In
November 2012 5-403 Draft - Do Not Cite or Quote
-------
1 addition, studies reporting high blood Pb levels and high Pb exposure levels report
2 increased urine, serum, or blood urea nitrogen (Wang et al.. 2010d: El-Nekeetv et al..
3 2009; Javakumar etal..2009; Kharoubi et al., 2008a). Jayakumar et al. (2009) reported
4 alterations in other markers of kidney toxicity, lysosomal marker and brush border
5 enzymes (i.e., ALP, ACP, y-GT, NAG, (3-D-glucuronidase), following Pb exposure
6 (2,000 ppm for 6 weeks). Similarly, Wang et al. (2010d) reported time-related increases
7 in urinary alkaline phosphatase, urinary GGT, urinary NAG, urinary total protein, urinary
8 (3-2 microglobulin, and urinary microalbumin following Pb exposure (300 ppm in
9 drinking water, serum Pb level 20 (ig/dL). Pb-exposed male rats (500 ppm Pb acetate in
10 drinking water for 7 months, blood Pb level 43 (ig/dL) had elevated urinary pH, urinary
11 glucose, and proteinuria (Navarre-Moreno et al.. 2009).
12 Qiao et al. (2006) measured the effect of Pb on the expression of the renal nuclear factor-
13 kappa B (NF-KB), transforming growth factor (TGF-(3) and fibronectin in Sprague-
14 Dawley rat kidney. These growth (TGF-|3) and transcription (NF-KB) factors modulate
15 the progression of renal function decrements through promotion of extracellular matrix
16 (fibronectin) synthesis and promotion of fibrosis. Pb was administered at a dose of
17 5,000 ppm Pb acetate, continuously for either one, two, or three months. All factors
18 increased by the end of three months of treatment, but only NF-KB increased
19 progressively at each time period. These changes were hypothetically related to the
20 development of Pb-induced renal fibrosis in rats, but no histology was performed.
21 The renal effects of chronic Pb exposure as detailed above were partially rescued in rats
22 following lowering of blood Pb level with chelation therapy (i.e., DMSA) (Khalil-
23 Manesh et al.. 1992a) and after treatment with antioxidants (Abdel Moneim et al.. 201 Ib;
24 Ozsovetal. 2010; Wang etal.. 2010d; El-Nekeetv et al.. 2009; Javakumar et al.. 2009;
25 Kharoubi et al.. 2008a). DMSA treatment improved renal function; however, Pb-induced
26 pathology remained (Khalil-Manesh et al., 1992a). Improvements include increased GFR,
27 decreased albuminuria, and decreased inclusion body numbers but little change in
28 tubulointerstitial scarring. DMSA also acts as an antioxidant, so the protective effects
29 may not be entirely attributed to the lowering of blood Pb level. Similarly, several studies
30 found that treatment with antioxidant compounds could protect against Pb-induced
31 kidney dysfunction. Administration of flaxseed oil, L-carnitine, NAC, and several
32 medicinal plants including, Achyranthes aspera, Artemisia absinthium, and Aquilegia
33 vulgaris, to Pb-exposed rodents protected against injury to the kidney or restored kidney
34 function (Abdel Moneim et al., 201 Ib; Ozsoy etal., 2010; Wang etal., 2010d; E\-
35 Nekeetv et al.. 2009; Javakumar et al.. 2009; Kharoubi et al.. 2008a). These studies
36 suggest that a reduction in reactive oxygen species may attenuate the effects of Pb on
37 kidney function implicating oxidative stress as a predominant mechanism for Pb-induced
38 reduced kidney function.
November 2012 5-404 Draft - Do Not Cite or Quote
-------
Histological Changes
1 Earlier studies discussed in previous Pb AQCDs have identified Pb-related renal damage
2 by the presence of dense intranuclear inclusion bodies, which are capable of sequestering
3 Pb (Goveretal.. 1970b). Pb-induced formation of intranuclear inclusion bodies in the
4 proximal tubule (PT) is considered protective; Pb is sequestered such that it is not in its
5 bioavailable, free, lexicologically active form. Intranuclear inclusion bodies are found in
6 the kidney with short-term (i.e., <4 weeks) Pb exposure but present to a lesser degree
7 with chronic exposures (See Section 5.2.3 for further discussion). Chelators such as
8 CaNa2EDTA have removed these inclusion bodies from affected nuclei (Goyer et al..
9 1978).
10 Multiple ultrastructural changes indicate dysfunction in the PT and nephropathy after Pb
11 exposure, including changes to the PT epithelium, endoplasmic reticulum dilation,
12 nuclear membrane blebbing, and autophagosome enlargement (Fowler et al.. 1980; Goyer
13 et al.. 1970a). Indications similar to the PT transport-associated Fanconi syndrome appear
14 with Pb exposure, albeit often at high doses of Pb, i.e., Pb poisoning. These indications,
15 which include increased urinary electrolyte excretion (Zn), decreased Na+/K+ATPase
16 activity, mitochondrial aberrations, and aminoaciduria, also have been associated with
17 blood Pb levels in children.
18 Recent studies since the 2006 Pb AQCD are consistent with the earlier findings and build
19 upon the literature base by including the role of antioxidants. Jabeen et al. (2010) exposed
20 pregnant albino BALB/c mice to a daily oral dose of Pb acetate (10 mg/kg body weight,
21 daily throughout pregnancy) until GDI8, at which point the fetal kidneys were processed
22 for histological examination. Histology revealed Pb exposure induced decreased kidney
23 cortical thickness, decreased diameter of renal corpuscles, and increased renal tubular
24 atrophy (with desquamated epithelium and degenerated nuclei in the distal and proximal
25 tubules). Blood Pb levels were not reported in this study. Nonetheless, these data show
26 that in utero Pb exposure had significant histological effects on the fetal kidney, which
27 could contribute to altered renal function including clearance of waste products,
28 electrolyte balance, and vasoregulation.
29 Massanyi et al. (2007) reported on Pb-induced alterations in male Wistar rat kidneys after
30 single i.p. doses of Pb acetate (50, 25, and 12.5 mg/kg); kidneys were removed and
31 analyzed 48 hours after Pb administration. Qualitative microscopic analysis detected
32 dilated Bowman's capsules and dilated blood vessels in the interstitium with evident
33 hemorrhagic alterations. Quantitative histomorphometric analysis revealed increased
34 relative volume of interstitium and increased relative volume of tubules in the
35 experimental groups. The diameter of renal corpuscles and the diameter of glomeruli and
36 Bowman's capsule were significantly increased. Measurement of tubular diameter
November 2012 5-405 Draft - Do Not Cite or Quote
-------
1 showed dilatation of the tubule with a significant decrease of the height of tubular
2 epithelium compatible with degenerative renal alterations. These findings extend the
3 observations of Fowler et al. (1980) and Khalil-Manesh et al. (1992b; 1992a); in
4 particular, the enlarged glomeruli are consistent with the early hyperfiltration caused by
5 Pb.
6 Abdel Moneim et al. (20 lib) reported histological evidence of inflammation after Pb
7 treatment in rats (i.p. 20 ppm, 5 days). This evidence included increased inflammatory
8 cellular infiltrations, cytoplasmic vacuolation, and dilatation of some kidney tubules.
9 Inflammation was accompanied by an increase in apoptotic cells and increased oxidative
10 stress.
11 A recent study has also reported inclusion body formation in the nuclei, cytoplasm, and
12 mitochondria of PT cells of Pb-treated rats (50 mg Pb/kg bw i.p., every 48 hours for
13 14 days) (Navarre-Moreno et al.. 2009). These inclusion bodies were not observed in
14 chronically Pb-exposed rats (500 ppm Pb in drinking water, 7 months). However, chronic
15 Pb exposure resulted in morphological alterations including loss of PT apical membrane
16 brush border, collapse and closure of the PT lumen, and formation of abnormal
17 intercellularjunctions.
18 Vogetseder et al. (2008) examined the proliferative capacity of the renal PT (particularly
19 the S3 segment) following i.v. administration of Pb to juvenile and adult male Wistar
20 rats. Proliferation induction was examined by detection of Bromo-2'-deoxyuridine
21 (BrdU), Ki-67 (labels S, G2, and M phase cells), and cyclin Dl (an essential cell cycle
22 progression protein). The cycling marker Ki-67 revealed a much higher proliferation rate
23 in the S3 segment in control juvenile rats (4.8 ± 0.3%) compared with control adult rats
24 (0.4 ± 0.1%). Pb administration (3.8 mg/100 g bw) increased the proportion of Ki-67-
25 positive cells to 26.1 ± 0.3% in juvenile rats and 31.9±0.3%in adult rats. Thus, the
26 increased proliferation caused by Pb was age independent. The proliferation induction
27 caused by Pb administration may be a result of reduced cell cycle inhibition by p27klp"1.
28 Acute Pb treatment increased the incidence of cyclin D1 labeling in the BrdU-positive
29 cells suggesting Pb was able to accelerate re-entry of cells into the cell cycle and cause
30 proliferation in the PT. Pb-induced cellular proliferation has also been reported in the
31 retina with gestational and early postnatal rodent Pb exposure (Giddabasappa et al..
32 2011).
33 Ademuyiwa et al. (2009) examined Pb-induced phospholipidosis and cholesterogenesis in
34 rat tissues. Sprague-Dawley rats were exposed to 200, 300 and 400 ppm Pb acetate for
35 12 weeks. The Pb exposure resulted in induction of phospholipidosis in kidney tissue,
36 accompanied by depletion of renal cholesterol. The authors suggested that induction of
37 cholesterogenesis and phospholipidosis in kidney may be responsible for some of the
November 2012 5-406 Draft - Do Not Cite or Quote
-------
1 subtle and insidious cellular effects found with Pb-mediated nephrotoxicity. Drug-
2 induced PT phospholipidosis is seen clinically with use of the potentially nephrotoxic
3 aminoglycoside drugs, including gentamicin (Baronas et al., 2007).
4 Various antioxidants have been shown to attenuate Pb-induced histopathological changes
5 to the kidney. Ozsoy et al. (2010) found L-carnitine to be protective in a model of
6 experimental Pb toxicity in female rats. Markers of histopathological change in the
7 kidney, including tubule dilatation, degeneration, necrosis, and interstitial inflammation
8 were rescued by L-carnitine treatment in females. Male rats exposed to Pb (2,000 ppm for
9 6 weeks) also displayed tubular damage, whereas concomitant treatment with Pb and an
10 extract of Achyranthes aspera ameliorated the observed damage (Jayakumar et al.. 2009).
11 El-Sokkary et al. (2005) reported Pb-induced (100 ppm s.c. for 30 days) tubular
12 degeneration with necrotic cells that could be prevented with melatonin treatment.
13 Melatonin is known to be an efficacious free radical scavenger and indirect antioxidant.
14 El-Nekeety et al. (2009) found an extract of the folk medicine plant Aquilegia vulgaris to
15 be protective against Pb acetate-induced tubular dilatation, vacuolar and cloudy epithelial
16 cell lining, interstitial inflammatory cell infiltration, hemorrhage, cellular debris, and
17 glomerulus hypercellularity. Concomitant exposure to Pb and extract produced histology
18 indiscernible from that in controls. Post treatment with extract partially rescued the
19 Pb-induced histopathology. El-Neweshy and El-Sayed (2011) studied the influence of
20 vitamin C supplementation (20 mg/kg pretreatment every other day) on histopathological
21 alterations in Pb-exposed male rats (20 mg/kg by intragastric feeding once daily for
22 60 days). Control rats showed normal histology, while Pb-treated rats exhibited
23 karyomegaly with eosinophilic intranuclear inclusion bodies in the epithelial cells of the
24 proximal tubules. Glomerular damage and tubular necrosis with invading inflammatory
25 cells were also found. Rats treated with Pb acetate plus vitamin C exhibited relatively
26 mild or no karyomegaly with eosinophilic intranuclear inclusion bodies in the proximal
27 tubules. Normal glomeruli were noted in animals exposed to Pb and vitamin C. These
28 findings consistently show that some antioxidants are capable of preventing or rescuing
29 Pb-induced renal histopathological changes, suggesting a role for oxidative stress in the
30 development of Pb-induced nephropathy.
31 Table 5-30 presents the acute and chronic renal effects of Pb exposure observed in recent
32 and past animal toxicology studies.
November 2012 5-407 Draft - Do Not Cite or Quote
-------
Table 5-30 Effects of Pb on the kidney/renal system related to exposure
duration- evidence from animal toxicology studies.
Effects with less than 3 months of exposure
Effects with 6 or 12 months of exposure
Mitochondrial dysfunction
Renal cell apoptosis
Nuclear Inclusion Body Formation
Proximal Tubule Cytomegaly
Glomerular Hypertrophy
Increased GFR
Mitochondrial dysfunction
Renal cell apoptosis
Oxidant redox imbalance
Altered NO homeostasis
ATPase dysfunction
Aminoaciduria
Increased electrolyte excretion
Elevated blood pressure
Decreased GFR
5.5.3
Modes of Action for Pb-lnduced Nephrotoxicity
i
2
o
5
4
5
6
9
10
11
12
13
14
15
16
17
18
19
20
5.5.3.1 Oxidative Damage
A role for ROS in the pathogenesis of experimental Pb-induced hypertension and renal
disease has been well characterized (Vaziri. 2008a. b; Vaziri and Khan. 2007). The
production of oxidative stress following Pb exposure is detailed in respect to modes of
action of Pb (Section 5.2.4). Past studies have shown that Pb treatment (single or three
daily i.p. injections) can elevate kidney GST levels, affecting glutathione metabolism
(Daggett et al.. 1998: Moseretal.. 1995: Oberlev et al.. 1995).
Animal studies continue to provide evidence for increased oxidative stress playing a role
in the pathogenesis of Pb-induced renal toxicity. Increased ROS, serum NO, and renal
NO were observed after Pb injections in rats (i.p. 20 mg/kg, 5 days) (Abdel Moneim et
al., 20lib). Pb exposure to rat proximal tubular cells (0.5-1 (iM) also increased ROS
production, in a concentration-dependent manner (Wang et al.. 20 lib). Increased lipid
peroxidation (i.e., MDA) was demonstrated in serum and renal tissue after Pb exposure
(Abdel Moneim et al.. 201 Ib: Lodi etal.. 2011: Wang etal.. 20lib). Berrahal et al.
(2011) reported increased MDA in Pb-exposed (50 ppm Pb acetate pre- and post-natally)
rat kidney relative to controls at both 40 (puberty; blood Pb 12.7 ug/dL) and 65 (post-
puberty; blood Pb 7.5 ug/dL) days of age. In addition, total sulfhydryl groups were
significantly decreased at 65 days. These increases in oxidative stress were accompanied
by age-dependent Pb nephrotoxicity in male rats.
Alterations in endogenous antioxidants and antioxidant enzymes that may lead to
oxidative stress have also been reported after Pb exposure. Pb treatment decreased the
November 2012
5-408
Draft - Do Not Cite or Quote
-------
1 activity of the renal antioxidant enzymes, CAT, SOD, GST, GPx, and GR (Abdel
2 Moneim et al. 20 lib) and protein levels of CAT and GSH (Lodietal.. 2011).
3 Additionally, proteomic analysis of high-level Pb treated (1,500 ppm, 5 weeks; resulting
4 in blood Pb level of 53.4 (ig/dL) rat kidney identified decreased abundance of a rate-
5 limiting enzyme in the synthesis of GSH (glutamate cysteine ligase) (Chen etal.. 20 lib).
6 Conterato et al. (2007) examined the effect of Pb acetate on the cytosolic thioredoxin
7 reductase activity and oxidative stress parameters in rat kidneys. A single injection of
8 Pb acetate consisted of a single i.p. injection of 25 or 50 mg/kg Pb acetate, while repeated
9 injections consisted of one daily i.p. injection of Pb acetate (5 or 25 mg/kg) for 30 days.
10 Measured were thioredoxin reductase-1, a selenoprotein involved in many cellular redox
11 processes, SOD, 5-ALAD, GST, GPx, non protein thiol groups (NPSH), CAT, as well as
12 plasma creatinine, uric acid, and inorganic phosphate levels. The single injection at the
13 25 mg Pb dose level resulted in increased SOD and thioredoxin reductase-1 activity,
14 while the 50 mg dose level increased CAT activity and inhibited 5-ALAD activity in the
15 kidney. Repeated injections at the 5 mg dose level of Pb inhibited 5-ALAD and increased
16 GST, NPSH, CAT, and thioredoxin reductase-1. Repeated injections at the 25-mg dose
17 level reduced 5-ALAD but increased GST, NPSH, and plasma uric acid levels. No
18 changes were observed in TEARS, GPx, creatinine or inorganic phosphate levels after
19 either single or repeated injection dosing. As both dosing regimens increased thioredoxin
20 reductase-1 activity, the authors suggest that this enzyme may be a sensitive indicator of
21 renal changes with low dose Pb treatment.
22 Jurczuk et al. (2006) published a study of the involvement of some low molecular weight
23 thiols in the peroxidative mechanisms of action of Pb in the rat kidney. Wistar rats were
24 fed a diet containing 500 ppm Pb acetate for a period of 12 weeks and were compared to
25 a control group receiving distilled water for the same time period. GSH, metallothionein
26 (MT), total and nonprotein SH groups (TSH and NPSH) were measured, as were the
27 blood activity and urinary concentration of 5-ALA. The concentrations of GSH and
28 NPSH were decreased by Pb administration, while MT concentration was unchanged.
29 5-ALAD in blood was decreased, whereas urinary 5-ALA was increased by Pb
30 administration. Negative correlations were found between the kidney GSH concentrations
31 and previously reported concentrations of Pb and MDA in kidneys of these rats. It is
32 apparent from graphical presentation of the data that GSH was reduced by more than
33 50% following Pb administration, while TSH was reduced by approximately 15%. No
34 values for either blood or kidney Pb levels or kidney MDA were reported in this article.
35 In 2007, the same authors (Jurczuk et al.. 2007) reported on the renal concentrations of
36 the antioxidants, vitamins C and E, in the kidneys of the same Pb-treated and control rats.
37 Exposure to Pb significantly decreased vitamin E concentration by 13% and vitamin C
38 concentration by 26%. The kidney concentration of vitamin C negatively correlated with
November 2012 5-409 Draft - Do Not Cite or Quote
-------
1 MDA concentration. The authors concluded that vitamins E and C were involved in the
2 mechanism of peroxidative action of Pb in the kidney, and their protective effect may be
3 related to scavenging of free radicals.
4 Studies have used antioxidant compounds to investigate the role of oxidative stress in
5 Pb-induced nephrotoxicity. Abdel Moneim et al. (20lib) reported that flaxseed oil
6 treatment protected rats from Pb-induced (i.p. 20 mg/kg, 5 days) oxidative stress,
7 inflammation, and apoptosis. However, the flaxseed oil also decreased the accumulation
8 of Pb in renal tissue making it difficult to ascertain whether the protection was due to
9 decreased oxidative stress or to altered Pb uptake kinetics.
10 El-Neweshy and El-Sayed (2011) studied the influence of vitamin C supplementation on
11 Pb-induced histopathological alterations in male rats. Rats were given Pb acetate,
12 20 mg/kg by intragastric feeding once daily for 60 days. Control rats were given 15 mg of
13 sodium acetate per kg once daily, and an additional group was given Pb acetate plus
14 vitamin C (20 mg/kg every other day) 30 minutes before Pb feeding. Control rats showed
15 normal histology, while Pb-treated rats exhibited karyomegaly with eosinophilic
16 intranuclear inclusion bodies in the epithelial cells of the proximal tubules. Glomerular
17 damage and tubular necrosis with invading inflammatory cells were also seen in
18 Pb-treated animals. Among rats treated with Pb acetate plus vitamin C, five exhibited
19 relatively mild karyomegaly and eosinophilic intranuclear inclusion bodies of proximal
20 tubules and an additional five rats were normal. Normal glomeruli were noted in all.
21 Thus, vitamin C was shown to ameliorate the renal histopathological effects of Pb
22 intoxication, however no measures of Pb accumulation were provided to clarify the
23 mechanism of action of vitamin C.
24 Masso-Gonzalez and Antonio-Garcia (2009) studied the protective effect of natural
25 antioxidants (Zn, vitamin A, vitamin C, vitamin E, and vitamin B6) against Pb-induced
26 damage during pregnancy and lactation in rat pups. At weaning, pups were sacrificed and
27 kidneys were analyzed. Pb-exposed pups had decreased body weights. Blood Pb levels
28 were 1.43 ug/dL in the control group, 22.8 ug/dL in the Pb group, 21.2 ug/dL in the Pb
29 plus Zn plus vitamins group, and 0.98 ug/dL in the Zn plus vitamin group. The kidney
30 TEARS were significantly elevated in Pb exposed pups, while treatment with vitamins
31 and Zn returned TEARS to control levels. Kidney CAT activity was significantly
32 increased above control with Pb treatment; however supplementation with Zn and
33 vitamins reduced CAT activity toward normal. Pb exposure inhibited kidney Mn-
34 dependent SOD but not Cu-Zn-dependent SOD activity. Thus, supplementation with Zn
35 and vitamins during gestation and lactation was effective in attenuating the redox
36 imbalance induced by developmental, chronic low-level Pb exposure, likely through the
37 alteration of Pb accumulation.
November 2012 5-410 Draft - Do Not Cite or Quote
-------
1 Bravo et al. (2007) reported further that mycophenolate mofetil (an immunosuppressive
2 agent used in renal transplantation which inhibits T and B cell proliferation)
3 administration reduces renal inflammation, oxidative stress and hypertension in
4 Pb-exposed rats. Thus, an inflammatory immune and oxidative stress component can be
5 seen as contributing to Pb-induced renal effects and hypertension.
6 Although the majority of studies of the effects of Pb exposure have been conducted in
7 male rats, a couple of studies have compared the response of male rats with female rats
8 (Sobekova et al., 2009; Alghazal et al., 2008a). Sobekova et al. (2009) contrasted the
9 activity response to Pb on the antioxidant enzymes, GPx and GR, and on TEARS in both
10 male and female Wistar rats of equal age. Males weighing 412 ± 47 g and females
11 weighing 290 ± 19 g were fed diets containing either 100 ppm or 1,000 ppm Pb acetate
12 for 18 weeks. In the male rats, kidney Pb content increased by 492% on the 100 ppm Pb
13 diet and by 7,000% on the 1,000 ppm Pb diet. In the female rats, kidney Pb content
14 increased by 410% on the 100 ppm Pb diet and by 23,000% on the 1,000 ppm Pb diet.
15 There was virtually no change in GPx in the kidney of male rats given the 100 ppm Pb
16 diet but there was a significant reduction in GPx in the female rats on both the 100 ppm
17 diet and 1,000 ppm diet. In male rats, GR was increased from 182 units/gram of protein
18 in control kidneys to 220 units on the 100 ppm Pb diet and 350 units on the 1,000 ppm
19 diet. In female rats, kidney GR decreased from 242 units in control animals to 164 units
20 in animals on the 100 ppm Pb diet and 190 units in animals on the 1,000 ppm diet. In
21 male rats, kidney TEARS content increased from 7.5 units/gram protein to 10.0 units
22 (1,000 ppm Pb diet group). In female rats, there was a reduction in TEARS from 14.4
23 units per gram protein to 10.0 units in rats on the 100 ppm Pb diet and to 11 units in rats
24 on the 1,000 ppm Pb diet.
25 Alghazal et al. (2008a) compared the activity responses of the antioxidant enzyme, SOD
26 and the detoxifying enzyme, GST, of the same rats exposed to 100 ppm or 1,000 ppm
27 Pb acetate for 18 weeks. Similar to the previous study, kidney TEARS were increased
28 only in male rats given the higher dose of Pb. Kidney SOD activity, on the other hand,
29 was increased in both males and females at the higher dose of Pb, while GST activity was
30 increased in kidney of males at the higher dose of Pb and decreased at the lower dose, but
31 was decreased at both doses of Pb in females. Thus there were significant differences in
32 the responses of male and female rats to Pb exposure. Differences may be accounted for
33 in part due to the greater deposition of Pb in female rat kidneys. Another explanation,
34 offered by the authors, is that male rats are known to metabolize some foreign
35 compounds faster than do females, so the biological half-life of xenobiotics in the
36 females may be longer.
November 2012 5-411 Draft - Do Not Cite or Quote
-------
5.5.3.2 Hypertension and Alteration of Renal Vasculature and
Reactivity
1 As discussed in Section 5.5.1. changes in renal vasculature function or induction of
2 hypertension can contribute to further renal dysfunction. Pb exposure increases BP,
3 resulting in hypertension, through the promotion of oxidative stress and altered vascular
4 reactivity (Section 5.4). Antioxidants attenuated Pb-related oxidative/nitrosative stress in
5 the kidney and abrogated the Pb-induced increased BP (Vaziri etal.. 1999a). Chronic
6 increases in vascular pressure can contribute to glomerular and renal vasculature injury,
7 which can lead to progressive renal dysfunction and kidney failure. In this manner,
8 Pb-induced hypertension has been noted as one contributer to Pb-induced renal disease.
9 Also, Pb has been shown to act on known vasomodulating systems in the kidney. In the
10 kidney, two vascular tone mediators, NO and ET-1, are found to be affected by Pb
11 exposure. Administration of the vasoconstrictor endothelin-1 (ET-1) affected mean
12 arterial pressure (MAP) and decreased GFR (Novak and Banks. 1995). Acute high-dose
13 Pb exposure (24 nmol/min for 15 or 30 minutes) completely blocked this ET-1-mediated
14 GFR decrease but had no effect on MAP. Depletion of the endogenous antioxidant
15 glutathione using the drug buthionine sulfoximine, a GSH synthase inhibitor, increased
16 BP and increased kidney nitrotyrosine formation without Pb exposure, demonstrating the
17 importance of GSH in maintenance of BP (Vaziri et al., 2000). Multiple studies have
18 shown that Pb exposure depletes GSH stores. Catecholamines are vascular moderators
19 that are also affected by Pb exposure (Carmignani et al., 2000). The effect on BP with Pb
20 exposure is especially relevant to the kidney because it is both a target of Pb deposition
21 and a mitigator of BP. These earlier data detail the interaction of known modulators of
22 vascular tone with Pb.
23 The renin-angiotensin-aldosterone system plays an important role in kidney homeostasis
24 and alteration of this pathway may affect renal function. Simoes et al. (2011) reported
25 that acute Pb treatment (Pb acetate i.v. bolus dose of 320 (ig/kg bw, blood Pb of 37 (ig/dL
26 at 120 minutes after Pb administration) in adult male Wistar rats increased serum
27 angiotensin converting enzyme (ACE) activity. Systolic arterial pressure, but not diastolic
28 arterial pressure or heart rate, was also elevated 60 minutes after treatment. The
29 Pb-induced altered systolic BP attenuated in animals co-treated with Losartan (Ang II
30 receptor blocker) or Enalapril (ACE inhibitor), suggesting a regulatory role for the renin-
31 angiotensin system (Simdes et al.. 2011). These data agree with earlier reports of
32 Pb-related increases in ACE activity in young rats exposed to Pb for 2-8 weeks (Sharifi et
33 al.. 2004) and adult rats exposed to Pb for 10 months (Carmignani et al.. 1999).
34 Recently, Vargas-Robles et al. (2007) examined the effect of Pb exposure (100 ppm
35 Pb acetate for 12 weeks) on BP and angiotensin II vasoconstriction in isolated perfused
November 2012 5-412 Draft - Do Not Cite or Quote
-------
1 kidney and interlobar arteries. Vascular reactivity was evaluated in the presence and
2 absence of the nitric oxide synthase inhibitor L-NAME in both Pb-treated and control
3 animals. Pb exposure significantly increased BP (134 ± 3 versus 100 ± 6 mmHg), eNOS
4 protein expression, oxidative stress, and vascular reactivity to angiotensin II. L-NAME
5 potentiated the vascular response to angiotensin II in the control group, but had no effect
6 on the Pb-treated group. Conversely, passive microvessel distensibility, measured after
7 deactivation of myogenic tone by papaverine, was significantly lower in the arteries of
8 Pb-exposed rats. Nitrites released from the kidney under the influence of angiotensin II in
9 the Pb group were lower as compared to the control group whereas 3-nitrotyrosine was
10 higher in the Pb group. The authors concluded that Pb exposure increases vascular tone
11 through nitric oxide-dependent and -independent mechanisms, increasing renal vascular
12 sensitivity to vasoconstrictors.
5.5.3.3 Apoptosis and/or Ischemic Necrosis of Tubules and
Glomeruli
13 Apoptosis or programmed cell death in excess can cause cell atrophy while an
14 insufficiency can lead to uncontrolled cell proliferation, such as cancer. Pb exposure has
15 been shown to cause morphological changes to the kidney structure. Some of these
16 Pb-induced changes are a result of cellular apoptosis or necrosis. Past studies have shown
17 Pb-induced necrosis in proximal tubule cells (Fowler et al. 1980). Pb-induced apoptosis
18 is known to act through the mitochondria (Rana. 2008). Pb-induced calcium overload
19 may depolarize the mitochondria, resulting in cytochrome c release, caspase activation,
20 and apoptosis. The apoptosis is mediated by Bax translocation to the mitochondria and
21 can be blocked by overexpression of Bcl-xl. Also, Pb-induced ALA accumulation can
22 generate ROS, which may damage DNA leading to apoptosis.
23 Mitochondria are targets of Pb toxicity and often involved in apoptosis. Pb can induce
24 uncoupling of oxidative phosphorylation, decreased substrate utilization, and
25 modification of mitochondrial ion transport. ATP energetics are affected when ATP-Pb
26 chelates are formed and ATPase activity is decreased. ROS formation can contribute to
27 these mitochondrial changes and to other changes within the kidney. Antioxidant
28 supplementation after Pb exposure can remedy some changes. All of these outcomes, in
29 conjunction with Pb-related depletion of antioxidants (e.g., GSH) and elevation of lipid
30 peroxidation point to possible susceptibility of the kidney to apoptosis or necrosis.
31 Rodriguez-Iturbe et al. (2005) reported that chronic exposure to low doses of Pb
32 (100 ppm in drinking water for 14 weeks) results in renal infiltration of immune cells,
33 apoptosis, NF-KB activation and overexpression of tubulointerstitial Ang(II). Similarly,
November 2012 5-413 Draft - Do Not Cite or Quote
-------
1 higher level Pb treatment in rats (i.p. 20 mg/kg, 5 days) induced inflammatory cellular
2 infiltrations and an increase in apoptotic cells, accompanied by more pronounced BAX
3 staining in kidney tubule epithelial cells (Abdel Moneim et al., 201 Ib). Pb treatment
4 (0.5-1 (iM) of isolated rat proximal tubular cells increased cell death by apoptosis and
5 necrosis in a concentration- and time-dependent manner (Wang et al., 20 lib). This was
6 accompanied by increased morphological changes typical of apoptosis such as
7 fragmented chromatin, condensed chromatin, and shrunken nuclei. These cells also
8 exhibited decreased mitochondrial membrane potential, decreased intracellular pH,
9 inhibition of Na+/K+ATPase and Ca2+ATPase activity, and increased intracellular Ca2+
10 following Pb treatment.
11 Navarro-Moreno et al. (2009) examined the effect of 500 ppm Pb in drinking water over
12 7 months on the structure (including intercellular junctions), function, and biochemical
13 properties of PT cells of Wistar rats. Pb effects in epithelial cells consisted of an early
14 loss of the apical microvilli, followed by a decrement of the luminal space and the
15 respective apposition and proximity of apical membranes, resulting in the formation of
16 atypical intercellular contacts and adhesion structures. Inclusion bodies were found in
17 nuclei, cytoplasm, and mitochondria. Lipid peroxidation (TEARS measurement) was
18 increased in the Pb-treated animals as compared to controls. Calcium uptake was
19 diminished and neither proline nor serine incorporation that was present in controls was
20 noted in the PT of Pb-exposed animals. The authors speculated that Pb may compete with
21 calcium in the establishment and maintenance of intercellular junctions.
22 Tubular necrosis was also observed in rats treated with Pb acetate (100 ppm s.c.) for
23 30 days (El-Sokkary et al.. 2005). Histological sections of kidneys from Pb-treated rats
24 showed tubular degeneration with some necrotic cells. Similarly, El-Neweshy and
25 El-Sayed (2011) reported glomerular damage and tubular necrosis with invading
26 inflammatory cells after Pb treatment (20 mg/kg by intragastric feeding once daily for
27 60 days) to male rats. The incidence of necrosis was decreased in both of these studies by
28 pretreatment with either melatonin or vitamin C. Pretreatment with melatonin
29 (10 mg/kg), an efficacious free radical scavenger and indirect antioxidant, resulted in a
30 near normal tubular structure. The authors concluded that melatonin protected the liver
31 and kidneys from the damaging effects of exposure to Pb through inhibition of lipid
32 peroxidation and stimulation of endogenous antioxidative defense systems (El-Sokkary et
33 al.. 2005). Vitamin C supplementation (20 mg/kg pretreatment every other day) protected
34 the renal architecture and histology (El-Neweshy and El-Saved. 2011).
35 Wang et al. (2009c) examined the effect of Pb acetate (0.25, 0.5 and 1 (iM) on cell death
36 in cultured rat primary PT cells. A progressive loss in cell viability, due to both apoptosis
37 and necrosis, was observed in cells exposed to Pb. Apoptosis predominated and could be
November 2012 5-414 Draft - Do Not Cite or Quote
-------
1 ameliorated with concomitant N-acetylcysteine exposure, whereas necrosis was
2 unaffected. Elevation of ROS levels and intercellular calcium, depletion of mitochondrial
3 membrane potential, and intracellular glutathione levels was observed during Pb
4 exposure. Pb-induced apoptosis was demonstrated morphologically (Hoechst 33258
5 staining) with condensed/fragmented chromatin and apoptotic body formation. CAT and
6 SOD activities were significantly elevated, reflecting the response to accumulation of
7 ROS.
5.5.3.4 Renal Gangliosides
8 Gangliosides are constituents of the plasma membrane that are important for control of
9 renal GFR because they can act as receptors for various molecules and have been shown
10 to take part in cell-cell interactions, cell adhesion, cellular recognition, and signal
11 transduction. Aguilar et al. (2008) studied changes in renal gangliosides following Pb
12 exposure (600 ppm Pb acetate in drinking water for 4 months) in adult male Wistar rats.
13 Pb exposure caused an increase in blood Pb from 2.1 to 35.9 (ig/dL. There was no change
14 in serum creatinine or in hemoglobin, but there was an increase in urinary 5-ALA. The
15 following renal gangliosides were measured by immunohistochemistry and by thin layer
16 chromatography: GM1, GM2, GM4, and 9-O-acetylated modified form of the
17 GD3 ganglioside (9-O-Ac-GD3). The ganglioside pattern was mainly characterized by a
18 decrease in the GM1 ganglioside as well as by a mild increase in GM4 and GM2
19 gangliosides, while the strongest alteration was observed in the 9-O-Ac-GD3, which was
20 overexpressed. The latter was observed only in the glomerular zone. This was associated
21 with a decrease in apoptotic glomerular cells, as assessed by the TUNEL assay. The
22 authors hypothesized that the increase in GD3-O-acetylation could represent a strategy to
23 attenuate the normal renal apoptotic process and therefore contribute to cell survival
24 during Pb exposure.
5.5.3.5 Altered Uric Acid
25 Higher occupational Pb exposure or blood Pb levels have been linked to increased risk
26 for both gout and kidney disease (Shadick et al., 2000; Batuman. 1993). Pb is thought to
27 increase serum uric acid by decreasing its kidney excretion (Emmerson and Ravenscroft.
28 1975; Ball and Sorensen, 1969; Emmerson. 1965). Research during the past decade
29 indicates that uric acid is nephrotoxic at lower levels than previously recognized (Johnson
30 et al.. 2003). Therefore, the 2006 Pb AQCD (U.S. EPA. 2006b) reviewed literature
31 implicating increased uric acid as a mechanism for Pb-related nephrotoxicity (Weaver et
32 al., 2005a; Shadick et al., 2000). However, this does not appear to be the only
November 2012 5-415 Draft - Do Not Cite or Quote
-------
1 mechanism, since associations between blood Pb and serum creatinine have remained
2 significant even after adjustment for uric acid (Weaver et al.. 2005a).
3 Alterations in serum uric acid have been studied in animal models exposed to Pb. In male
4 rats exposed to Pb in drinking water from lactation to puberty (40 days) or post-puberty
5 (65 days), Berrahal et al. (2011) found that plasma urea levels increased after 40 days of
6 exposure (puberty blood Pb level of 12.7 ug/dL) but decreased after 65 days of Pb
7 exposure (post-puberty blood Pb level of 7.5 ug/dL) (Table 5-29). Serum uric acid was
8 increased in male rats following long-term Pb exposure (2,000 ppm for 6 weeks)
9 (Javakumar et al.. 2009). Conterato et al. (2007) followed various parameters of kidney
10 function after single or multiple Pb injections in rats. The single dosing regimen consisted
11 of a single i.p. injection of 25 or 50 mg/kg Pb acetate, while the multiple injections
12 involved once daily i.p. injection of either vehicle or Pb acetate (5 or 25 mg/kg) for
13 30 days. Single and multiple injections at both dose levels increased plasma uric acid
14 levels. Similarly, Abdel Moneim et al. (20lib) reported increased serum uric acid and
15 urea levels after 5 days of Pb acetate treatment (i.p. 20 mg/kg).
5.5.3.6 Role of Metallothionein
16 The metal-binding protein, metallothionein, may play a role in inclusion body formation
17 and thus block potential interaction of Pb with cellular targets. Yu et al. (2009) described
18 dichotomous effects of Pb acetate on the expression of MT in the liver and kidney of
19 mice. Male mice were i.p. injected with Pb acetate in doses of 100, 200, and 300 (imol/kg
20 and sacrificed 4, 8, and 24 hours after Pb treatment. Administration of Pb increased the
21 levels of MT-1 mRNA in the liver and kidneys but increased MT protein only in the
22 liver. Treatment of mouse PT cells in vitro with Pb also resulted in an increase in MT
23 mRNA but little increase in MT protein. Thus, Pb appears to exert a dual effect on MT
24 expression in the kidney: enhancement of MT gene transcription but suppression of MT
25 mRNA translation.
26 Zuo et al. (2009) examined the potential role of a-Synuclein (Sena) and MT in
27 Pb-induced inclusion body formation. Unlike the parental wild type (WT) cells, MT-I/II
28 double knockout (MT-null) cells did not form inclusion bodies after Pb treatment;
29 however, transfection of MT-1 into MT-null cells allowed inclusion body formation after
30 Pb treatment. As inclusion bodies formed during Pb treatment, soluble MT protein in WT
31 cells was lost. As Sena is a protein with a natural tendency to aggregate into oligomers,
32 Sena was measured in WT cells and MT-null cells after Pb treatment. In both cell lines
33 Pb-induced Sena expression rapidly increased and then decreased over 48 hours as
34 Pb-induced inclusion bodies were formed. Pb exposure caused increased colocalization
November 2012 5-416 Draft - Do Not Cite or Quote
-------
1 of MT and Sena proteins and MT was localized to the surface of inclusion bodies in WT
2 renal cortex samples following Pb treatment. Thus, Sena may be a component of
3 Pb-induced inclusion bodies and, with MT, may play a role in inclusion body formation.
5.5.4 Effects of Exposure to Pb Mixtures
4 The effect of Pb on other cations, specifically calcium, is well established in the kidney
5 literature. Calcium-mediated processes involving receptors, transport proteins, and
6 second messenger signaling among other endpoints have been shown to be significantly
7 affected by Pb exposure. The disposition of Pb in the soft tissues (kidney and spleen) can
8 change with exposure to Pb and other compounds. Pb plus Cd exposure changed Pb
9 disposition with increased blood Pb (versus Pb alone group) and decreased metal
10 concentration in the kidney and liver (versus Pb alone). An iron deficient diet
11 significantly increased Pb deposition in adult animals (Hashmi et al.. 1989). pregnant
12 dams, and maternally-exposed fetuses (Singh et al.. 1991). Dietary thiamine plus Zn
13 slightly reduced blood and kidney Pb in exposed animals (Flora etal.. 1989). Selenium
14 (Se), a cofactor for GPx, attenuated Pb-induced lipid peroxidation and abrogated the
15 Pb-induced attenuation of GR and SOD. Concomitant exposure to the cations aluminum
16 and Pb protected animals from ensuing nephropathy (Shakoor et al.. 2000). In summary,
17 Pb has been shown to affect processes mediated by endogenous divalent cations. In
18 addition, exposure to other metals or divalent cations can modulate Pb disposition and its
19 effects in the body.
5.5.4.1 Lead(Pb) and Cadmium(Cd)
20 Cd shares many similarities with Pb; it has been shown to be a ubiquitous PT
21 nephrotoxicant and accumulates in the body. Despite the similarities, few studies have
22 evaluated associations between Cd exposure and CKD or the impact of joint exposure of
23 Pb and Cd or other metals on CKD. As discussed in the 2006 Pb AQCD (U.S. EPA.
24 2006b). environmental exposure to Cd, at levels common in the U.S. and other developed
25 countries, has been shown to impact substantially associations between indicators of Pb
26 exposure and the kidney EBE marker, NAG, even in the presence of occupational level
27 Pb exposure. In an occupational study, mean NAG, although higher in the Pb-exposed
28 worker group compared to controls, was correlated with urine Cd but not blood or tibia
29 Pb (Roels et al.. 1994). In another occupational population where both metals were
30 significantly associated with NAG, a 0.5 ug/g creatinine increase in Cd had the same
31 effect on NAG as did a 66.9 ug/g bone mineral increase in tibia Pb (Weaver et al..
32 2003a).
November 2012 5-417 Draft - Do Not Cite or Quote
-------
1 The 2006 Pb AQCD noted that data examining the concentration-response relation
2 between environmental Cd and the kidney were too scarce to determine the impact of Cd
3 exposure on relations between Pb exposure and other kidney outcomes. A recent
4 publication in NHANES data collected from 1999 through 2006 addresses this need;
5 (results pertaining solely to Pb were discussed in Section 5.5.2.1) (Navas-Acien et al..
6 2009). Geometric mean concurrent blood Cd level was 0.41 ug/L in 14,778 adults aged
7 > 20 years. After adjustment for survey year, sociodemographic factors, CKD risk
8 factors, and blood Pb, the ORs for albuminuria (> 30 mg/g creatinine), reduced eGFR
9 (<60 mL/min/1.73 m2), and both albuminuria and reduced eGFR were 1.92 (95% CI:
10 1.53, 2.43), 1.32 (95% CI: 1.04, 1.68), and 2.91 (95% CI: 1.76, 4.81), respectively,
11 comparing the highest with the lowest blood Cd quartiles. Both Pb and Cd remained
12 significantly associated after adjustment for the other. Effect modification was not
13 observed; however, ORs were higher for adults in the highest quartiles of both metals
14 compared with the ORs for the highest quartiles of concurrent blood Cd or Pb alone
15 (Table 5-25). Compared with adults with blood Cd levels < 0.2 ug/L and blood Pb levels
16 < 1.1 ug/dL, adults with blood Cd levels >0.6 ug/L and blood Pb levels >2.4 ug/dL had
17 ORs (95% CIs) of 2.34 (95% CI: 1.72, 3.18) for albuminuria, 1.98 (95% CI: 1.27, 3.10)
18 for reduced eGFR, and 4.10 (95% CI: 1.58, 10.65) for albuminuria and reduced eGFR
19 together. These findings are consistent with other recent publications (Akesson et al.
20 2005; Hellstrom et al.. 2001). support consideration of both metals as independent CKD
21 risk factors in the general population, and provide novel evidence of increased risk in
22 those with higher environmental exposure to both metals.
23 However, a very recent study suggests that interpretation of Cd associations with GFR
24 measures may be much more complex. Conducted in Pb workers to address the fact that
25 few studies have examined the impact of environmental Cd exposure in workers who are
26 occupationally exposed to other nephrotoxicants such as Pb, the study assessed Cd dose
27 with urine Cd, which is widely considered the optimal dose metric of cumulative Cd
28 exposure. In 712 Pb workers, mean (SD) blood and tibia Pb, urine Cd, and eGFR using
29 the MDRD equation were 23.1 (14.1) ug/dL, 26.6 (28.9) ug/g, 1-15 (0.66) ug/g
30 creatinine, and 97.4 (19.2) mL/min/1.73m2, respectively (Weaver et al.. 2011). After
31 adjustment for age, sex, BMI, urine creatinine, smoking, alcohol use, education, annual
32 income, diastolic BP, current or former Pb worker job status, new or returning study
33 participant, and blood and tibia Pb, higher urine Cd was associated with higher calculated
34 creatinine clearance, eGFR (P = 8.7 mL/min/1.73 m2 [95% CI: 5.4, 12.1] per unit
35 increase in In-transformed urine Cd) and In-NAG, but lower serum creatinine. These
36 unexpected paradoxical associations have been reported in a few other publications (de
37 Burbure et al.. 2006; Hotz et al.. 1999) and have been observed in other populations.
38 Potential explanations for these paradoxical results included a normal physiologic
November 2012 5-418 Draft - Do Not Cite or Quote
-------
1 response in which urine Cd levels reflect renal filtration; the impact of adjustment for
2 urine dilution with creatinine in models of kidney outcomes; and Cd-related
3 hyperfiltration.
4 Wang et al. (2009c) studied the effects of Pb and/or Cd on oxidative damage to rat kidney
5 cortex mitochondria. In this study young female Sprague Dawley rats were fed for
6 8 weeks with either Pb acetate (300 ppm), Cd chloride (50 ppm), or Pb and Cd together
7 in the same dosage. Lipid peroxidation was assessed as MDA content. Renal cortex
8 pieces were also processed for ultrastructural analysis and for quantitative rtPCR to
9 identify the mitochondrial damage and to quantify the relative expression levels of
10 cytochrome oxidase subunits (COX-I/II/III). Cytochrome oxidase is the marker enzyme
11 of mitochondrial function, and COX-I, II, and III are the three largest mitochondrially-
12 encoded subunits which constitute the catalytic functional core of the COX holoenzyme.
13 Mitochondria were altered by either Pb or Cd administration, but more strikingly by Pb
14 plus Cd administration, as indicated by disruption and loss of mitochondrion cristae.
15 Kidney cortex MDA levels were increased significantly by either Pb or Cd, given
16 individually, but more so by Pb plus Cd. COX-I/II/III were all reduced by either Pb or Cd
17 administration, but more prominently by Pb plus Cd administration. This study adds to
18 knowledge of the synergistic effects of Pb and Cd on kidney mitochondria.
5.5.4.2 Lead(Pb), Cadmium(Cd), and Arsenic(As)
19 Wang and Fowler (2008) present a general review of the roles of biomarkers in
20 evaluating interactions among mixtures of Pb, Cd, and As. Past studies have found that
21 addition of Cd to treatment of rats with Pb or Pb and As significantly reduced the
22 histological signs of renal toxicity from each element alone, including swelling of the
23 proximal tubule cells and intranuclear inclusion body formation. On the other hand,
24 animals exposed to Cd in addition to Pb or Pb and As showed an additive increase in the
25 urinary excretion of porphyrins, indicating that, although measured tissue burdens of Pb
26 were reduced, the biologically available fraction of Pb is actually increased (Mahaffey et
27 al.. 1981: Mahaffev and Fowler. 1977).
November 2012 5-419 Draft - Do Not Cite or Quote
-------
1 Stress proteins were examined after exposure to mixtures of Pb and other metals.
2 Induction of MT was strongest in groups with Cd treatment. However, co-exposure to Pb
3 and As induced higher levels of MT protein than did either Pb or As exposure alone in
4 kidney tubule cells. Heat shock proteins (Hsps) are commonly altered with exposure to
5 metal mixtures. A study found in vitro (low dose) and in vivo that Pb induced Hsps in a
6 metal/metalloid-, concentration- and time-specific manner (Wang et al.. 2005a). Additive
7 or more than additive interactions occurred among Pb, Cd and As under combined
8 exposure conditions.
5.5.4.3 Lead (Pb) and Zinc (Zn)
9 Zinc has been investigated as a protective compound against the effects of Pb. Pb
10 treatment (35 mg/kg i.p. for 3 days) caused a significant fall in hemoglobin content,
11 significant increases in lipid peroxidation and decreased level of reduced glutathione in
12 liver, together with diminished total protein content in liver and kidney. Co-treatment of
13 Pb with Zn (10 mg/kg i.p.) or ascorbic acid (10, 20 and 30 mg/kg i.p.) showed a moderate
14 therapeutic effect when administered individually, but more pronounced protective
15 effects after combined therapy (Upadhyay et al., 2009).
16 Jamieson et al. (2008) studied the effect of dietary Zn content on renal Pb deposition.
17 Weanling Sprague Dawley rats were assigned to marginal zinc (MZ, 8 mg Zn/kg diet),
18 zinc adequate control (CT, 30 mg Zn/kg), zinc-adequate diet-restricted (30 mg Zn/kg), or
19 supplemental zinc (SZn, 300 mg Zn/kg) groups, with or without Pb acetate (200 ppm for
20 3 weeks). Pb exposure did not result in nephromegaly or histological alterations. The MZ
21 rats had higher renal Pb (35%) and lower renal Zn (16%) concentrations than did CT rats.
22 On the other hand, SZn was more protective than the CT diet was against renal Pb
23 accumulation (33% lower). Standard procedures for indirect immunoperoxidase staining
24 were used to determine MT localization in the kidney. Pb had no effect on MT staining
25 intensity, distribution, or relative protein amounts. Western blot analysis confirmed that
26 MT levels were responsive to dietary Zn but not to Pb exposure.
5.5.4.4 Lead(Pb) and Mercury(Hg)
27 Stacchiotti et al. (2009) studied stress proteins and oxidative damage in a renal-derived
28 cell line exposed to inorganic Hg and Pb. The time course of the expression of several
29 Hsps, glucose-regulating proteins and MTs in a rat proximal tubular cell line (NRK-52E)
30 exposed to subcytotoxic doses of inorganic mercury (HgCl2, 1-40 uM) and Pb (PbCl2,
31 2-500 uM) were analyzed. ROS and reactive nitrogen species (RNS) were detected by
November 2012 5-420 Draft - Do Not Cite or Quote
-------
1 flow cytometric analysis. Endogenous total GSH content and the enzymatic activity of
2 GST were determined in cell homogenates. Western blot analysis and
3 immunohistochemistry were used for quantification of hsps and MTs. Reverse
4 transcription PCR was used for quantification of metallothionein. The higher doses of Hg
5 (20 uM and 40 uM) were shown to markedly inhibit growth of the cell line while the
6 higher doses of Pb (60 uM to 500 uM) inhibited cell growth to a lesser degree. After 24
7 hours of exposure of 20 (iM Hg, the cells presented abnormal size and pyknotic nuclei,
8 swollen mitochondria and both apoptosis and overt necrosis. In the presence of 60 or
9 300 (iM Pb, the cells lost cell-cell and cell-matrix contacts, showed a round size, irregular
10 nuclear contour and often mitotic arrest, but no apoptosis or overt necrosis at 24 hours.
11 Mercury (Hg) induced a significant increase in both ROS and RNS, maximal RNS at
12 24 hours, and maximal ROS at 48 hours. Pb (60 or 300 (iM) did not cause an increase in
13 ROS or RNS beyond the levels measured in control cells. Total GSH significantly
14 increased in cells grown in the presence of Pb; the effect was concentration-dependent
15 and GSH reached its maximal value at a dose of 300 (iM Pb. The effect of Hg was
16 biphasic: 10 uM significantly enhanced GSH by 600%, while the amount of GSH
17 detected after 20 (iM Hg only increased by 50% compared to control levels. GST activity
18 was enhanced by both Pb and Hg. Hsp25 and Hsp72 were up-regulated by Hg but there
19 was no effect on Grp78 as compared to control. On the contrary, Pb treatment only
20 upregulated Grp78. Mercury (Hg) induced a time-dependent effect on MT mRNA
21 expression, which reached its maximal value 3 hours after beginning treatment and
22 reverted to control values at 24 hours. With Pb, on the other hand, mRNA transcription
23 was concentration- and time-dependent. The transcripts remained overexpressed
24 compared to controls up to 72 hours. The results of this study with regard to the Pb effect
25 on MT synthesis clearly differ from those of Jamieson et al. (2008). which found no
26 increase in MT following Pb exposure. This discrepancy remains to be clarified.
November 2012 5-421 Draft - Do Not Cite or Quote
-------
5.5.5 Summary and Causal Determination
1 A large body of epidemiologic evidence and limited toxicological evidence indicates Pb
2 exposure leads to reduced kidney function. The causal determination for reduced kidney
3 function is informed by evidence for reduced GFR, reduced creatinine clearance, and
4 increased serum creatinine. Biological plausibility and mode of action for these effects is
5 provided by evidence for hypertension, oxidative stress, inflammation, vascular reactivity
6 and injury, increased uric acid, morphological changes, and apoptosis or necrosis. The
7 section that follows describes the evaluation of evidence for reduced kidney function,
8 with respect to causal relationships with Pb exposure using the framework described in
9 Table II of the Preamble. The key supporting evidence to the causal framework is
10 summarized in Table 5-31.
5.5.5.1 Evidence for Reduced Kidney Function
11 The 2006 Pb AQCD (U.S. EPA. 2006b) concluded that "in the general population, both
12 circulating and cumulative Pb was found to be associated with a longitudinal decline in
13 renal function," evidenced by increased serum creatinine and decreased creatinine
14 clearance or eGFR over follow-up of 4 to 15 years in association with higher baseline
15 blood and bone Pb levels (U.S. EPA. 2006b). Data in general and patient populations of
16 adults provided consistent evidence of Pb-associated lower renal function in populations
17 with mean concurrent or baseline blood Pb levels of 2-10 ug/dL (Akesson et al., 2005;
18 Tsaih et al.. 2004; Yu et al.. 2004; Kimetal.. 1996); associations with lower eGFR were
19 observed in adults with hypertension with a mean concurrent blood Pb level of 4.2 ug/dL
20 (Muntner et al.. 2003). The conclusion from the 2006 Pb AQCD was substantiated by the
21 coherence of effects observed across epidemiologic and toxicological studies. However, a
22 number of the animal toxicological studies were conducted at Pb exposure concentrations
23 that resulted in blood Pb levels higher than what is relevant to the general U.S. adult
24 human population. Both human and animal studies observed Pb-associated
25 hyperfiltration. In animals during the first 3 months after Pb exposure, effects were
26 characterized by increased GFR and increased kidney weight due to glomerular
27 hypertrophy. However, exposure for 6 or 12 months resulted in decreased GFR,
28 interstitial fibrosis, and kidney dysfunction. Additionally, toxicological studies found that
29 early effects of Pb on tubular cells were generally reversible, but continued exposure
30 resulted in chronic irreversible damage. Toxicological studies provided mechanistic
31 evidence to support the biological plausibility of Pb-induced renal effects, including
32 oxidative stress leading to NO inactivation. Despite the strong body of evidence
33 presented in the 2006 Pb AQCD, uncertainty remained on the contribution of past Pb
November 2012 5-422 Draft - Do Not Cite or Quote
-------
1 exposures to associations observed in adults, the impact in children, the implication of
2 hyperfiltration, and reverse causality.
3 Consideration of results from recent epidemiologic studies does not alter the conclusions
4 of the previous AQCD. Prospective studies in adult men in the general population (Tsaih
5 et al.. 2004; Kimetal.. 1996): an adult patient population study (Yu et al. 2004): CKD
6 patient studies (Lin et al., 2006b: Lin et al., 2006a: Lin et al., 2003). and a Pb worker
7 cohort (Weaver et al.. 2009) provide evidence of a relationship between blood Pb level
8 and subsequent decreases in kidney function and better establish the temporal sequence
9 between Pb exposure and kidney function. The study designs and analysis are well
10 designed with careful measurement of exposure, outcomes, and covariates, and
11 adjustment for numerous potential confounding factors including age, race, sex,
12 education, household income, smoking, alcohol use, Cd exposure, and various health
13 indicators such as diabetes, systolic BP, BMI, and history of cardiovascular disease.
14 Large sample sizes provide strength to the general population studies, whereas some
15 caution may be appropriate for the CKD patient studies where the smallest study has 87
16 subjects. Confidence in the relationship between Pb exposure and renal effects is
17 provided by the combined results of a body of studies from different research groups
18 using different designs in different cohorts.
19 Limitations of these studies produce some uncertainty in the relationship between Pb
20 exposure and reduced kidney function. Since the prospective studies are reported in
21 patients, workers, and primarily white men with a mean age of 60, they may not be
22 generalizable to the entire U.S. population. Also, as these studies report effects most
23 often observed in adults with likely higher past Pb exposures, uncertainty exists as to the
24 Pb exposure level, timing, frequency, and duration contributing to the associations
25 observed with blood or bone Pb levels. In addition, it is possible that the CKD patient
26 studies (Lin et al., 2006b: Lin et al., 2006a: Lin et al., 2003). in which blood Pb level was
27 lowered compared to control groups by chelation, demonstrate an improvement in renal
28 function by reducing reactive oxygen species, blood Pb level or both. The uncertainty
29 related to this may reflect an involvement of both lowering of blood Pb levels and a
30 reduction of reactive oxygen species following chelation as both are possible to a varying
31 extent. The potential for a bidirectional relationship because of reverse causality is
32 possible in observational epidemiologic studies and must be weighed in this discussion
33 (see Section 5.5.2.4).
34 Cross-sectional studies add support to the associations observed in prospective
35 epidemiologic studies (Section 5.5.2.1). The majority of cross-sectional studies report
36 associations between higher measures of Pb exposure and worse renal function. These
37 studies include analyses from the NHANES cohort which provides a representative U.S.
November 2012 5-423 Draft - Do Not Cite or Quote
-------
1 population sample that may be generalizable to the total U.S. population. Re-examination
2 of a study from the 2006 Pb AQCD (U.S. EPA. 2006b) provided data to conclude that in
3 a population with likely higher past exposures to Pb, a 10-fold increase in concurrent
4 blood Pb was associated with an 18 mL/min decrease in estimated creatinine clearance or
5 a 25% decrease from the mean, and that an increase in blood Pb from the 5th to the 95th
6 percentile (3.5 ug/dL) had the same negative impact on eGFR as did an increase of 4.7
7 years in age or 7 kg/m2 in body mass index (Akesson et al.. 2005). However, a small
8 number of studies report findings that are less consistent with the body of evidence
9 (Section 5.5.2.1). Overall, a relationship between higher Pb exposure and various
10 indicators of lower kidney function is indicated by a set of high-quality studies that
11 control for important potential confounders such as age, sex, BMI, comorbid
12 cardiovascular conditions, smoking and alcohol use, and that are conducted with different
13 designs in different cohorts by different researchers.
14 At current blood Pb levels in the U.S. adult population, a downward shift in kidney
15 function of the entire population due to Pb may not result in CKD in identifiable
16 individuals; however, that segment of the population with the lowest kidney reserve may
17 be at increased risk for CKD when Pb is combined with other kidney risk factors. For
18 example, in adults with mean (concurrent or baseline measured 4-6 years before kidney
19 function tests) blood Pb levels that are comparable to that of the general U.S. population
20 (1.6 to 4.2 ug/dL), higher blood Pb level was found to be associated with clinically-
21 relevant effects (e.g., eGFR <60 mL/min/1.73 m2, doubling of serum creatinine)
22 (Fadrowski et al.. 2010; Yu et al.. 2004) and larger magnitudes of effect in potentially at-
23 risk populations with cormorbidities for CKD such as diabetes mellitus (Tsaih et al.,
24 2004) and hypertension (Tsaih et al.. 2004; Muntner et al.. 2003) or higher co-exposure to
25 other environmental nephrotoxicants such as Cd (Navas-Acien et al.. 2009).
26 Research in the occupational setting has traditionally been far less consistent than that in
27 environmentally exposed populations (Section 5.5.2.1). Limitations of the occupational
28 evidence, which are discussed earlier in this section have been used to explain this
29 inconsistency. The actual cause of paradoxical or inverse associations (higher Pb dose
30 with lower serum creatinine, and/or higher eGFR or calculated or measured creatinine
31 clearance) in several of these studies may not be known. If associations are in opposite
32 directions in different subgroups of the population and the relevant effect modifier is not
33 considered, null associations will be observed. For these reasons, nonsignificant
34 associations or paradoxical associations in the occupational setting cannot be used as a
35 rationale for discounting Pb-related nephrotoxicity at lower environmental levels.
36 CKD is an important risk factor for cardiac disease. As kidney dysfunction can increase
37 BP and increased BP can lead to further damage to the kidneys, Pb-induced damage to
November 2012 5-424 Draft - Do Not Cite or Quote
-------
1 either or both renal or cardiovascular systems may result in a cycle of further increased
2 severity of disease. Pb exposure has been causally linked to both increased BP and other
3 cardiovascular effects (Section 5.4). Interestingly, animal studies have shown Pb-induced
4 vascular injury in the kidney was associated with increased glomerular sclerosis,
5 tubulointerstitial injury, increased collagen staining, and an increase in macrophages
6 associated with higher levels of MCP-1 mRNA. It is possible that the cardiovascular and
7 renal effects of Pb observed are mechanistically linked and thus, Pb-induced hypertension
8 has been noted as one cause of Pb-induced renal disease.
9 Recently available animal toxicological studies strengthen the evidence regarding the
10 modes of action for Pb exposure leading to renal alterations, including the influence of
11 Pb-induced oxidative stress. The mode of action of Pb in the kidneys has been extended
12 to the field of immunology, evidenced by observations that Pb exposure resulted in
13 infiltration of lymphocytes and macrophages associated with increased expression of
14 NF-KB in proximal tubules and infiltrating cells (Roncal et al.. 2007). Additionally,
15 recent findings expand on the evidence of acute effects of Pb, including mitochondrial
16 dysfunction, renal cell apoptosis, and glomerular hypertrophy. These mechanisms are
17 useful in understanding the occurrence of acute hyperfiltration followed by chronic
18 kidney dysfunction. Lower concentration Pb exposures and lower blood Pb levels in
19 animals have not been widely examined. As indicated in Table 5-28. studies found
20 dysfunction in various kidney function measures, including urinary flow, ALP,
21 microalbumin, and NAG at blood Pb levels greater than 20 (ig/dL.
22 Changes in renal function that have been associated with biomarkers of Pb exposure may
23 indicate a modest change for an individual; however, these modest changes can have a
24 substantial public health implication at the population level. The reported effects
25 represent a central tendency of Pb-induced renal function effects among individuals;
26 some individuals may differ in risk and manifest effects that are greater in magnitude. For
27 example, a small worsening of renal function may shift the population distribution and
28 result in considerable increases in the percentages of individuals with worse renal
29 function that are clinically significant.
30 Overall, recent studies evaluated in the current review support and expand upon the body
31 of evidence presented in the 2006 Pb AQCD indicating that Pb exposure is associated
32 with reduced kidney function. In addition, animal studies provide biological plausibility
33 for the associations observed in epidemiologic studies between blood Pb levels and
34 reduced kidney function with evidence for Pb-induced hypertension, renal oxidative
35 stress, inflammation, apoptosis, and glomerular hypertrophy. However, a number of
36 limitations remain including the representativeness of worker studies and older all male
37 cohorts to the U.S. population, the potential for reverse causality to play a role in the
November 2012 5-425 Draft - Do Not Cite or Quote
-------
1 findings of cross-sectional studies, and inconsistent findings in occupational studies.
2 Collectively, the evidence integrated across epidemiologic and toxicological studies is
3 sufficient to conclude that a causal relationship is likely to exist between Pb exposures
4 and reduced kidney function.
November 2012 5-426 Draft - Do Not Cite or Quote
-------
Table 5-31 Summary of evidence supporting renal causal determinations.
Attribute in Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or Blood
Levels Associated
with Effects0
Reduced Kidney Function - Likely Causal
Consistent associations Multiple prospective epidemiologic
in multiple high quality
epidemiologic studies
with relevant blood Pb
levels
studies in nonoccupationally exposed
adults for associations with change in
serum creatinine and GFR in MAS,
Korean workers, and CKD patients
Studies provide concentration-response
information
Associations found with adjustment for
potential confounding factors including
age, pre-existing cardiovascular disease,
baseline kidney function, and SES.
Supporting cross-sectional evidence for
associations between concurrent blood
Pb level and serum creatinine, creatinine
clearance, and GFR
Studies had population based
recruitment (NHANES) with high follow-
up participation
Uncertainty related to reverse causality;
the bidirectional association is possible
Uncertainty due to baseline serum
creatinine adjustment
Section 5.5.2.1
Muntner (2003).
Navas-Acien (2009)
Section 5.5.2.1
Akesson (2005).
Staessen (1992)
Payton (1994)
Section 5.5.2.4
Blood Pb level:
Adults,
<10ug/dLd
Concurrent Blood Pb
level:
Adults, means
1.58-4.2 ug/dL
Concurrent Blood Pb
level:
Adults, medians
2.2-11.4 ug/dL
Limited toxicological
evidence to support
epidemiologic evidence
Mixed evidence in animals report
decreased creatinine clearance,
increased serum creatinine, and
decreased GFR at both relevant and high
level long-term exposures.
Berrahal et al. (2011).
Roncal et al. (2007).
Khalil-Manesh et al.
(1993a: 1992b: 1992a)
Blood Pb level:
Rodents: >7.5 ug/dL
<65 days from birth,
26 ug/dL
12 weeks as adults,
29-125 ug/dL
Evidence clearly
describes mode of action
Hypertension
Oxidative Stress
Increased Uric Acid
Inflammation
Morphology
Consistent evidence of increased BP and
hypertension following Pb exposure in
humans and animals at relevant Pb
levels across numerous studies with
control for confounding.
Association of increased blood pressure
with manifestation of CHD has been well
documented.
Consistent evidence for increased ROS,
enhanced lipid peroxidation, and
antioxidant enzyme disruption in Pb
exposed animals.
Evidence of increased plasma uric acid
and urea in animals.
Mixed results in humans
Lymphocyte and macrophage infiltration
Increased MCP-1 expression
Glomerular hypertrophy
Cellular apoptosis and necrosis leading
to PT damage
Sections 5.4 and 5.5.3.2
Section 5.5.3.1
Section 5.5.3.5
Sections 5.5.2.5 and 5.2.5
Sections 5.5.2.5 and 5.5.3.3
"Described in detail in Table II of the Preamble.
bDescribes the key evidence and references contributing most heavily to causal determination. Also noted are the sections where full body of
evidence is described.
°Describes the blood Pb levels in humans with which the evidence is substantiated and blood Pb levels in animals most relevant to humans.
dBecause blood Pb level in nonoccupationally-exposed adults reflects both recent and past Pb exposures, the magnitude, timing, frequency, and
duration of Pb exposure contributing to the observed associations is uncertain.
November 2012
5-427
Draft - Do Not Cite or Quote
-------
5.6 Immune System Effects
5.6.1 Introduction
1 With respect to studies conducted in laboratory animal and in vitro models, the immune
2 effects of Pb exposure have been extensively examined over several decades. Animal
3 studies of the effects of Pb exposure on host resistance date back to the 1960s while those
4 focusing on Pb-induced immune functional alterations, including developmental
5 immunotoxicity, were first conducted during the 1970s. Despite this long history of
6 research, Pb-associated immune effects in animals with blood Pb levels in the range of
7 current U.S. population levels (i.e., <10 (ig/dL), particularly early in life, have been
8 observed only within the last 10-15 years (Dietert and McCabe. 2007). Recent findings of
9 Pb-associated changes in immunological parameters in humans have increased
10 understanding of the immune effects of environmental exposure to Pb.
11 The pathways by which Pb exposure may alter immune cell function and consequently
12 increase the risk of immune-related diseases are presented in (Figure 5-34). Rather than
13 producing overt cytotoxicity, Pb exposure has been associated with functional alterations
14 in cellular and humoral immunity. In the 2006 Pb AQCD (U.S. EPA. 2006b). the
15 hallmarks reported for Pb-induced changes in immune functional pathways were:
16 (1) suppression of T cell-derived helper (Th)l-mediated immunity (i.e., suppressed Thl
17 cytokine production and delayed type hypersensitivity [DTH] response); (2) stimulation
18 of Th2 immunity (i.e., increased production of Th2 cytokines and immunoglobulin (Ig)E
19 antibody); and (3) altered macrophage function (U.S. EPA. 2006b). The latter was
20 characterized by increased production of reactive oxygen species (ROS), prostaglandin E2
21 (PGE2), and inflammatory cytokines such as tumor necrosis factor-alpha (TNF-a) and
22 interleukin (IL)-6 and decreased production of nitric oxide (NO). Changes in immune
23 cells can alter cell-to-cell interactions, multiple signaling pathways, and inflammation,
24 that in turn, can influence the risk of developing infectious, allergic, and autoimmune
25 diseases and exacerbate inflammatory responses in other organ systems. Studies
26 conducted in animal and in vitro models provided consistent evidence for Pb exposure
27 inducing effects on the range of immune effects presented in this continuum. In the much
28 smaller epidemiologic evidence base, most studies examined Pb-exposed male workers
29 and a limited range of immune-related endpoints.
November 2012 5-428 Draft - Do Not Cite or Quote
-------
Elevated IL-4 and IL-5
Suppressed INF-y
Macrophages and
Other Innate Immune Cells
I
Skewed
Th2-biased
responses
Elevated IL-10
Suppressed IL-12
Increased lipid
and DNA oxidation
in tissues
Elevated TNF-a
Overproduction of ROS
Depleted antioxidant
defenses
Suppressed
Thl- mediated
anti-tumor
hostdefense / B Cells
Increased tumor \.
cell formation Increased IgE
production
Increased tissue
inflammation
(e.g. lung, gut, skin)
Reduced:
Phagocytosis
Nitric oxide production
Peroxynitrite production
Lysosomal activity
Removal of normal
myelomonocytic
suppression
Damaged epithelia
andmucosal barriers
\
/productio
\
Tissue damage
and de novo
antigen
appearance
Inappropriate
Tcell proliferation
activation
Increased risk of
later-life cancer
Increased risk of atopy
and allergic disease
Increased risk of tissue
inflammatory diseases
Increased risk of
autoimmunity
Reduced host resistance
to bacterial infection
Figure 5-34 Immunological pathways by which Pb exposure may increase risk
of immune-related diseases.
i
2
o
J
4
5
6
7
8
9
10
11
12
13
14
15
Reflecting suppressed Thl activity, toxicological evidence presented in the
2006 Pb AQCD linked Pb exposure of animals to impaired host resistance to viruses and
bacteria (U.S. EPA. 2006b). Indicating a hyperinflammatory state and local tissue
damage, a few available toxicological studies found Pb exposure-induced generation of
auto-antibodies, suggesting an elevated risk of autoimmune reactions. Additionally, the
shift toward Th2 responses suggested that Pb could elevate the risk of atopic and
inflammatory responses. While the biological plausibility of such effects was supported
by toxicological evidence for Pb-induced increases in Th2 cytokines, IgE, and
inflammation, epidemiologic evidence was too sparse to draw conclusions about the
effects of Pb exposure on these broader indicators of immune dysfunction in humans.
However, in concordance with toxicological evidence, a shift to a Th2 phenotype was
indicated in the few available studies by associations observed between higher concurrent
blood Pb level and higher serum IgE levels in children. Because of lack of examination,
the immune effects of Pb exposure in adults without occupational exposures were not
well characterized.
November 2012
5-429
Draft - Do Not Cite or Quote
-------
1 Changes in the spectrum of immune endpoints were found in association with a wide
2 range of blood Pb levels. Juvenile and adult animals with Pb levels in the range of
3 7-100 (ig/dL were found to have suppressed DTH, elevated IgE, and changes in cytokine
4 levels. Most epidemiologic studies examined and found lower T cell abundance and
5 higher serum IgE levels in association with population mean (or group) concurrent blood
6 Pb levels >10 (ig/dL.
7 With respect to important lifestages of Pb exposure, animal studies provided strong
8 evidence for immune effects in juvenile animals induced by prenatal Pb exposures and in
9 adult animals by postnatal exposures. There was uncertainty regarding important
10 lifestages of Pb exposure in humans as epidemiologic studies of children primarily were
11 cross-sectional and examined concurrent blood Pb levels. Several other limitations of
12 epidemiologic studies were noted, including small sample sizes; little consideration for
13 potential confounding factors such as age, sex, smoking, SES indicators, and allergen
14 exposures; and comparisons of immune endpoints among groups with different blood Pb
15 levels that provided little information on the concentration-response relationship.
16 Collectively, the small numbers of toxicological and epidemiologic studies published
17 since the 2006 Pb AQCD, supported the previous findings of Pb-associated immune
18 effects. Epidemiologic studies supported previous findings in children and provided new
19 evidence for effects in nonoccupationally-exposed adults. Recent studies also expanded
20 on the array of immunological parameters affected by Pb exposure as presented in Figure
21 5-34. For example, a recent toxicological study indicated that Pb may modulate the
22 function of dendritic cells. Results from recent toxicological and epidemiologic studies
23 supported the link between Pb-associated effects on immune cells and immune- and
24 inflammatory-related diseases by providing evidence for changes in intermediary
25 signaling and inflammatory pathways (Figure 5-34). Several recent epidemiologic studies
26 examined signaling molecules such as pro-inflammatory cytokines to provide coherence
27 with toxicological findings. Recent toxicological studies further supported the broader
28 role of Pb-associated immune modulation in mediating Pb effects in nonlymphoid tissues
29 (e.g., nervous, reproductive, respiratory systems). Recent epidemiologic studies improved
30 on the design of previous studies through greater examination of children and adults with
31 blood Pb levels more comparable to current levels in the U.S. population and greater
32 consideration for confounding by age, sex, smoking, SES indicators, and allergen
33 exposures. This epidemiologic evidence particularly that from prospective studies,
34 combined with the extensive toxicological evidence formed the basis of conclusions
35 about the immune effects of Pb exposure (Section 5.6.8).
November 2012 5-430 Draft - Do Not Cite or Quote
-------
5.6.2 Cell-Mediated Immunity
5.6.2.1 T Cells
1 T cells have a central role in cell-mediated immunity including maturation of B cells,
2 activation of cytotoxic T cells and macrophages, and interactions with antigen presenting
3 cells (APCs). A majority of the evidence for the effects of Pb exposure on T cells was
4 provided by toxicological and epidemiologic studies reviewed in the 2006 Pb AQCD
5 (U.S. EPA. 2006b). Toxicological evidence consistently links Pb exposure with
6 alterations in T cells with observations of Pb-induced shifts in the partitioning of CD4+
7 (T helper) cell populations to favor Th2 cells in vitro (25-50 (iM Pb chloride, 3-5 days)
8 (Heoetal.. 1998; 1996). proliferation of Th2 cells over Thl cells in vitro (10, 100 (iM
9 Pb chloride, 7 days) (McCabe and Lawrence. 1991). and the production of Th2 cytokines
10 over Thl cytokines in vitro and in vivo (wide range of Pb concentrations,
11 Section 5.6.6.1). Epidemiologic findings are limited largely to associations observed
12 between higher concurrent blood Pb level and lower T cell abundance in children.
13 In vitro results indicated various mechanisms by which Pb exposure may induce a shift to
14 Th2 responses including activation of transcription factor NF-KB (regulates T cell
15 activation) in cultures of human CD4+ T cells (1 (iM Pb acetate, 30 minutes) (Pvatt et al..
16 1996) and a concentration-dependent (10, 50 (iM Pb chloride, 24 hours) increased
17 expression of MHC class II surface antigens (e.g., HLA-DR), which mediate the CD4+
18 response to exogenous antigens (Guo etal.. 1996b). The few available recent
19 toxicological studies described T cell-dependent and -independent pathways. Kasten-
20 Jolly et al. (2010) provided evidence in vivo and with relevant dietary Pb exposures.
21 While results were based on a microarray analysis of hundreds of genes, which is subject
22 to higher probability of chance findings, they were supported by the extant evidence base.
23 In this study, gestational-lactational Pb acetate exposure of BALB/c mice (100 (iM in
24 drinking water of dams GD8-PND21, resultant spleen homogenate level <3 (ig/dL)
25 altered splenic cell gene expression of cytokines well documented in the literature to be
26 affected by Pb, including the Th2 cytokine IL-4 and the Thl cytokine interferon-gamma
27 (IFN-y). These changes occurred with increases in adenylate cyclase 8 and
28 phosphatidylinositol 3-kinase in the absence of signaling molecules STAT4 or STAT6,
29 which comprise the preferential signaling pathway for T cells. Similarly, in cultures of
30 stimulated mouse T cells, Heo et al. (2007) showed that Pb chloride (25 (JVI, 12-24
31 hours) decreased the IFN-y to IL-4 ratio (indicating a shift to Th2) in the absence of
32 STAT6. Additionally, Pb blocked production of IFN-y not by affecting gene expression
33 but by suppressing translation of the protein. This blockage was rescued with the addition
34 of IL-12, which is a T cell stimulating factor. The STAT results indicated a T cell-
November 2012 5-431 Draft - Do Not Cite or Quote
-------
1 independent pathway to skewing toward Th2 responses whereas the IL-12 results pointed
2 to a T cell-dependent pathway.
3 While a few available recent epidemiologic studies found associations of blood Pb levels
4 with Thl and Th2 cytokines in humans (Section 5.6.6.1). the extant evidence for effects
5 on T cells in humans is derived largely from previous cross-sectional studies describing
6 differences in the abundance of several T cell subtypes that mediate acquired immunity
7 responses to antigens. Most studies of children found that higher blood Pb levels were
8 associated with lower T cell abundance in serum, primarily CD3+ cells. These
9 associations were observed in studies that adjusted for some potential confounding
10 factors (as described below) (Karmaus et al.. 2005; Sarasua et al.. 2000) and studies
11 without consideration for potential confounding (Zhao et al., 2004; Lutz et al., 1999). In
12 children, blood Pb level was less consistently associated with lower abundance of other T
13 cell subtypes such as CD4+ (helper T) or CD8+ (cytotoxic T). Some studies did not
14 provide evidence of blood Pb-associated decreases in T cell abundance but did not
15 consider potential confounding (Hegazy et al., 2011; Belles-Isles et al., 2002).
16 Associations between blood Pb level and T cell abundance were found in studies that
17 generally had population-based recruitment. Most studies did not provide sufficient
18 information to assess the potential for biased participation by Pb exposure and immune
19 conditions. Most studies had multiple comparisons; however, associations were not
20 isolated to T cell abundance. Most studies found lower T cell abundance in groups of
21 U.S. and non-U.S. children (ages 6 months-10 years, n = 73-331) with concurrent blood
22 Pb levels >10 ug/dL (Zhao et al.. 2004; Sarasua et al.. 2000; Lutz et al.. 1999).
23 Associations were inconsistent in comparisons of children with lower blood Pb levels.
24 Among 331 children in Germany living near (15 km) and distant from industrial
25 facilities, Karmaus et al. (2005) found that children (ages 7-10 years) with concurrent
26 blood Pb levels 2.2-2.8 (ig/dL (2nd quartile) had a 9 to 11% lower abundance of several
27 T cell subtypes (for some subtypes, p <0.05, t-test) compared with children with blood Pb
28 levels <2.2 (ig/dL (lowest quartile). This study recruited children from schools and
29 examined multiple exposures, reducing the likelihood of biased participation by Pb
30 exposure. Monotonic decreases were not found across blood Pb groups. Compared with
31 other studies of T cells, Karmaus et al. (2005) had greater consideration for potential
32 confounding, adjusting for sex, age, number of infections in the previous 12 months,
33 number of cigarettes/day smoked in the home in the previous 12 months, serum lipids,
34 and blood organochlorine levels. SES was not examined. Cord blood levels of
35 organochlorine and Hg but not Pb were associated with T cell abundance in 96 newborns
36 from a subsistence fishing community and an urban center in Quebec, Canada with a
37 population mean cord blood Pb level <2 (ig/dL (Belles-Isles et al.. 2002).
November 2012 5-432 Draft - Do Not Cite or Quote
-------
1 Another study of children from multiple unspecified U.S. locations with and without
2 mining and smelting operations that considered confounding also found an association
3 between higher concurrent blood Pb level and lower T cell abundance, albeit limited to
4 the youngest subjects (Sarasua et al. 2000). Among 241 children ages 6-35 months, a
5 1 (ig/dL higher blood Pb level was associated with a 0.18% (95% CI: -0.34, -0.02) lower
6 CD3+ cell count, a 0.10% (95% CI: -0.24, 0.04) lower CD4+ cell count, and a 0.04%
7 (95% CI: -0.15, 0.07) lower CD8+ cell count, with adjustment for location of residence,
8 age, and sex. In older age groups (36-71 months, 6-15 years), many effect estimates were
9 positive. Analysis of blood Pb level categories indicated that associations were influenced
10 by lower T cell abundance (3-6%) among children ages 6-35 months with blood Pb levels
11 >15 (ig/dL. Notably, 76% of subjects lived near a Pb smelting operation, were likely to
12 have higher blood Pb levels, and may have influenced the observed associations. Neither
13 Karmaus et al. (2005) nor Sarasua et al. (2000) found a monotonic decrease in T cell
14 abundance across blood Pb level groups. Neither of these studies adjusted for SES, which
15 has been associated with blood Pb levels and immune-related conditions such as asthma,
16 allergy, and respiratory infections.
17 In the few studies with nonoccupationally-exposed adults (U.S, Italy), higher concurrent
18 blood Pb levels were associated with higher T cell abundance (Boscolo et al.. 2000;
19 Sarasua et al.. 2000; Boscolo etal.. 1999). the functional relevance of which is unclear.
20 These studies included healthy subjects and those with allergies, a wide range of samples
21 sizes (17-433), ages (16-75 years), and mean blood Pb levels (4.3-11.4 (ig/dL).
22 Pb-exposed workers in the U.S. and Asia did not consistently have lower or higher
23 abundance of various T cell subtypes than unexposed controls (Mishra et al., 2010;
24 Pinkerton et al.. 1998; Yiicesoy et al.. 1997b; Undeger et al.. 1996; Fischbein et al..
25 1993). The inconsistency among studies was not related to differences in sample size
26 (20-145), age (means: 22-58 years), or blood Pb levels (14.6-132 (ig/dL) among Pb
27 workers. None of the studies of adults considered potential confounding factors,
28 including other workplace exposures in occupational studies.
29 In summary, toxicological studies provided clear evidence for the effects of Pb exposure
30 on T cells by demonstrating Pb-induced expansion of Th2 cells and increased Th2
31 cytokine production. Providing mechanistic evidence, a few recent toxicological studies
32 found that Pb-induced Th2 skewing may occur via T cell-dependent (Heo et al.. 2007)
33 and -independent pathways (Kasten-Jolly et al.. 2010; Heo et al.. 2007). The most
34 consistent epidemiologic findings were associations between higher concurrent blood Pb
35 level (>10 (ig/dL) and lower T cell abundance observed in children ages 6 months to 10
36 years (Karmaus etal.. 2005; Zhao et al.. 2004; Sarasua et al.. 2000; Lutzetal. 1999). An
37 association was found with lower blood Pb levels, i.e., <3 (ig/dL (Karmaus et al.. 2005).
38 albeit in children ages 7-10 years who may have had higher Pb exposures in earlier
November 2012 5-433 Draft - Do Not Cite or Quote
-------
1 childhood. The uncertainties regarding blood Pb levels and the timing and duration of Pb
2 exposure contributing to the associations with T cell abundance apply to the evidence as a
3 whole since concurrent blood Pb levels in children also reflect past Pb exposures. There
4 are several other uncertainties in this evidence, including the temporal sequence between
5 Pb exposure and T cell changes, potential selection bias, the concentration-response
6 relationship, and potential confounding by factors such as SES and other environmental
7 exposures. The implications of Pb-associated changes in T cell abundance are limited
8 further by the uncertain functional relevance of small magnitudes of change in T cell
9 abundance (1-9% lower CD3+ abundance in groups of children with higher blood Pb
10 levels) to downstream immune responses. Because toxicological studies examined effects
11 related specifically to Thl or Th2 responses, toxicological evidence is the major
12 consideration in drawing conclusions about the effects of Pb exposure on T cells.
5.6.2.2 Lymphocyte Activation
13 Lymphocytes (T, B, and natural killer [NK] cells) are activated by reversing the normal
14 suppression mediated by macrophage-like cells. Their activation is an indicator of
15 response to antigens. A majority of data on the effects of Pb exposure on lymphocyte
16 activation is provided by toxicological studies reviewed in the 2006 Pb AQCD that
17 showed both mitogen-induced expansion and suppression of alloreactive B and T
18 lymphocytes proliferation with Pb exposures in vivo and in vitro (U.S. EPA. 2006b).
19 Adding to the mixed nature of evidence, a recent study found that 4-week oral exposure
20 of 7 week-old Wistar rats to 200 ppm Pb acetate induced proliferation of lymphocytes
21 within the thymus and submaxillary lymph nodes, primarily by affecting B cells (Teijon
22 et al.. 2010). Overall T cell proliferation did not change or was suppressed. Specific T
23 cell subtypes, CD4+, CD8+ (decreased), CD4-CD8- (elevated) were affected only with
24 i.p. Pb dosing (p <0.05) and not oral exposure. Using the local lymph node assay, Carey
25 et al. (2006) found that Pb chloride increased antigen-induced (ovalbumin, OVA) T cell
26 proliferation in adult female BALB/c mice but administered Pb via injection (25-50 (ig).
27 The mechanistic basis for Pb effects on lymphocyte activation is not well characterized.
28 As discussed in Section 5.6.6.2. changes in NO production appear to be involved (Farrer
29 et al.. 2008). Gao et al. (2007) described a potential role for dendritic cells. Dendritic
30 cells that matured in the presence of 25 (iM Pb chloride enhanced alloreactive T cell
31 proliferation in vitro compared to control dendritic cells.
32 A few available cross-sectional epidemiologic studies in children and nonoccupationally-
33 exposed adults provided indirect evidence for Pb-associated lymphocyte activation.
34 Instead of directly measuring lymphocyte proliferation, these studies measured the
35 abundance of cells that expressed HLA-DR, a cell surface marker that indicates both
November 2012 5-434 Draft - Do Not Cite or Quote
-------
1 activated lymphocytes and monocytes. These studies had limited consideration for
2 potential confounding, which also limits the implications of findings. In the study of
3 children (ages 9 months-6 years, Missouri), the mean percentage of HLA-DR+ cells was
4 about 2-fold higher (p >0.05, Kruskal-Wallis) in the 19 children with concurrent blood Pb
5 levels 15-19 (ig/dL than in children with blood Pb levels 10-14 (ig/dL (n = 61) or
6 <10 (ig/dL (n = 178) with adjustment for age (Lutz et al.. 1999). However, activated cells
7 were not elevated in 16 children with blood Pb levels 20-44 (ig/dL. Small studies of
8 adults without occupational Pb exposure in Italy found that concurrent blood Pb level
9 was correlated positively with the percentage of HLA-DR expressing cells in men ages
10 19-52 years with and without allergies (Spearman r = 0.51, p <0.002, n=17 each, overall
11 median blood Pb level: 11 (ig/dL) (Boscolo etal.. 1999) but only in women ages 19-49
12 years without allergies (Spearman r = 0.44, p <0.05, n=25, median blood Pb level:
13 5.5 (ig/dL) (Boscolo et al.. 2000). Associations also were found with other metals.
14 Comparisons of Pb-exposed workers and unexposed controls indicated similar levels of
15 lymphocyte proliferation (< 1% difference) in Pb-exposed workers (n = 10-33, mean age:
16 32-40 years, blood Pb level range: 12-80 (ig/dL) (Queiroz et al.. 1994b: Cohen et al..
17 1989) or lower lymphocyte proliferation (8-25%) among Pb-exposed workers (n = 15-39,
18 mean age: 30-49 years, mean blood Pb level: 14.6-129 (ig/dL) (Mishraet al.. 2003;
19 Fischbein et al.. 1993; Alomran and Shleamoon. 1988; Kimber et al.. 1986). In the
20 combined epidemiologic and toxicological evidence, Pb was associated with both
21 activation and suppression of lymphocyte activation. None of the studies considered
22 potential confounding factors, including other workplace exposures, and inconsistency
23 among studies could not be explained by differences in sample size, age of subjects, or
24 blood Pb level either. None of the studies provided concentration-response information.
25 Toxicological studies have demonstrated the selective expansion of Th2 cells and
26 suppression of Thl cells (Section 5.6.2.1). Therefore, the differential activation of
27 specific subtypes may not be discernible in studies that measure overall lymphocyte
28 proliferation.
5.6.2.3 Delayed-type Hypersensitivity
29 Although not widely examined recently, several toxicological studies reviewed in the
30 2006 Pb AQCD (U.S. EPA. 2006b) and recent reviews (Mishra. 2009; Dietert and
31 McCabe. 2007) identified a suppressed DTH response as one of the most consistently
32 observed immune effects of Pb exposure in animal models. A recent study indicated that
33 this effect may be mediated by dendritic cells. The DTH assay commonly is used to
34 assess the T cell-mediated response to antigens, i.e., induration and erythema resulting
35 from T cell activation and recruitment of monocytes to the site of antigen deposition. The
November 2012 5-435 Draft - Do Not Cite or Quote
-------
1 DTH response is largely Thl-dependent in that Thl cytokines induce the production of T
2 cells specifically directed against the antigen (sensitization) and recruitment of antigen-
3 specific T cells and monocytes to the site of antigen deposition (elicitation phase).
4 Previous studies demonstrated suppressed DTH in animals after gestational (Chen et al..
5 2004: Bunnetal. 200la: 2001b. c; Lee et al.. 200Ib: Chenetal.. 1999: Miller et al.
6 1998: Faith etal.. 1979) and postnatal (McCabe et al.. 1999: Laschi-Loquerie et al.. 1984:
7 Muller et al.. 1977) Pb acetate exposures. Such observations were made in F344 and CD
8 rats, BALB/c and Swiss mice, and chickens. Most studies exposed animals to Pb in
9 drinking water and found suppressed DTH in animals with blood Pb levels relevant to
10 humans (means: 6.75, 25 (ig/dL) (Chen et al.. 2004: Bunnetal.. 200 la) and higher (51 to
11 >100 (ig/dL) (Bunnetal.. 200 Ib: Chenetal.. 1999: McCabe et al.. 1999). The
12 associations of DTH with lower blood Pb levels occurred with gestational Pb exposure.
13 In some studies that examined Pb exposures at multiple stages of gestation, exposures
14 later in gestation suppressed DTH in animals (Bunnetal.. 2001c: Lee etal.. 200 Ib).
15 These latter findings may reflect the status of thymus and T cell development. A recent
16 study contributed to the robust evidence by indicating a role for dendritic cells in the
17 Pb-induced suppression of the DTH response. Gao et al. (2007) exposed bone marrow-
18 derived dendritic cells in vitro to Pb chloride (25 (JVI, 10 days) then the antigen OVA and
19 injected the cells into naive adult mice. Mice treated with Pb-exposed dendritic cells had
20 a diminished OVA-specific DTH footpad response compared with mice treated with
21 dendritic cells not exposed to Pb.
22 Evidence indicates Pb-induced suppression of DTH in animals with blood Pb levels
23 relevant to humans (6.75-25 (ig/dL) produced by gestational Pb exposure via drinking
24 water of dams. The mode of action is strongly supported by observations that Pb
25 suppresses production of the Thl cytokine IFN-y (Section 5.6.6.1). IFN-y is the primary
26 cytokine that stimulates recruitment of macrophages, a key component of the DTH
27 response. In animal studies that also examined IFN-y, the suppressed DTH response was
28 accompanied by a decreased production of IFN-y (Lee etal.. 2001b: Chen et al.. 1999).
29 Observations of a concomitant decrease in IFN-y strengthen the link between Pb-induced
30 inhibition of Thl functional activities and suppression of the DTH response.
5.6.2.4 Macrophages and Monocytes
31 As reported in the 2006 Pb AQCD, based on a large body of toxicological evidence and
32 some supporting epidemiologic evidence, Pb-induced alteration of macrophage function
33 was considered to be a hallmark of Pb-associated immune effects (U.S. EPA. 2006b).
34 Macrophages, which are produced by the differentiation of blood monocytes in tissues,
November 2012 5-436 Draft - Do Not Cite or Quote
-------
1 mediate host defense through their role in phagocytosing pathogens and stimulating other
2 immune cells. Pb exposure was found to induce macrophages into a hyperinflammatory
3 phenotype as indicated by enhanced production of TNF-a, IL-6, and ROS and increased
4 metabolism of arachidonic acid into PGE2. Observations in macrophages of Pb-induced
5 enhanced production of ROS, suppressed production of NO, impaired growth and
6 differentiation, and potentially altered receptor expression [e.g., toll-like receptors])
7 provided coherence with the effects of Pb observed on tissue damage and diminished host
8 resistance in animals. Several of these findings are described in detail in Sections 5.6.6.2
9 and 5.6.6.3. Because macrophages are major resident populations in most tissues and
10 organs and also are highly mobile in response to microbial signals and tissue alterations,
11 their functional impairment in response to Pb exposure may serve as a link between
12 Pb-induced immune effects and impaired host defense, tissue integrity, and organ
13 homeostasis in numerous physiological systems.
14 Some rodent studies indicated reduced macrophage generation or phagocytosis with
15 gestational or postnatal dietary Pb acetate exposure that produced blood Pb levels (upon
16 cessation of exposure) relevant to humans, i.e., 8.2 (ig/dL in F344 rats (Bunn et al..
17 200Ic) and 18 (ig/dL in CBA/J mice (Kowolenko et al., 1991). Similarly, Knowles and
18 Donaldson (1997) found that Pb acetate trihydrate (PND1-PND21) induced a decrease in
19 macrophage phagocytosis but in turkeys with higher blood Pb levels, 42 (ig/dL. In one set
20 of experiments, CBA/J mice exposed to Pb acetate for 2 weeks with blood Pb levels of
21 18 (ig/dL had reduced macrophage generation (Kowolenko et al.. 1991) in response to
22 Listeria infection but no change in macrophage phagocytosis (Kowolenko et al.. 1988).
23 Other animal studies administered Pb through routes that may not be directly relevant to
24 those in humans. Effects such as decreased macrophage yield, viability, phagocytosis,
25 chemotaxis, and killing ability were reported in Swiss mice following bacterial infection
26 and Pb treatment by oral gavage (40 mg/kg Pb nitrate, oral gavage, 40 days) (Lodi et al..
27 2011) or injection 10 mg/kg, i.p., 15 days) (Bishayi and Sengupta. 2006). Lee et al.
28 (2002) found no change in monocyte abundance in 5-6 week-old chickens treated with
29 200 (ig Pb acetate via the air sac in ovo at embryonic day 5 or 12. Some (Bussolaro et al..
30 2008; Zhouetal.. 1985) but not all in vitro studies (De Guise et al.. 2000) also found
31 Pb-induced (0.2-1,000 (iM Pb chloride or Pb nitrate) reduced phagocytosis. In particular,
32 Bussolaro et al. (2008) found such effects with a relatively low concentration of Pb
33 exposure (0.2 (iM Pb nitrate, 72 hours).
34 The effects of Pb exposure on macrophages in humans have not been widely examined.
35 Pineda-Zavaleta et al. (2004) was unique in examining the hyperinflammatory state
36 specifically in macrophages, and consistent with the large body of toxicological studies,
37 found associations of higher concurrent blood Pb level with lower NO release and higher
38 superoxide anion release from macrophages isolated from child sera (Sections 5.6.6.2 and
November 2012 5-437 Draft - Do Not Cite or Quote
-------
1 5.6.6.3). Other studies in humans examined macrophage abundance in Pb-exposed
2 workers, and evidence overall did not clearly indicate an association with concurrent
3 blood Pb level. Pinkerton et al. (1998) considered potential confounding and found a
4
-------
1 neurodegeneration in brain tissue (Struzvnska et al.. 2007). Bone osteoblasts have been
2 shown to be affected by Pb exposure (Section 5.9.4). which, given the interactions
3 between osteoblasts and osteoclasts (Chang et al.. 2008a). could have implications for
4 development of arthritis [reviewed in Zoeger et al. (2006)1. In vitro, 1 (iM Pb acetate
5 elevated TGF-(3 production and cartilage formation in limb bud mesenchymal cells
6 (Zuscik et al.. 2007). Kaczynska et al. (2011) reported effects on alveolar macrophages
7 after Pb acetate treatment (i.p. 25 mg/kg, 3 days, resulting in blood Pb levels of
8 2.1 (ig/dL) in Wistar rats. Macrophages infiltrated airways, limiting air space available to
9 gas exchange and contained parts of phagocytized surfactant and alveolar lining. Resident
10 immune cells in reproductive organs have been shown to be affected by high
11 concentration Pb exposure. In male BALB/c mice, Pace et al. (2005) found that 0.1 ppm
12 Pb acetate exposure in drinking water PND1-PND42 (mean peak blood Pb level:
13 59.5 (ig/dL) resulted in sterility concomitantly with a decrease in the testicular
14 macrophage population and an increase in apoptotic testicular cells.
15 In summary, an extensive toxicological evidence base demonstrates that Pb exposure
16 decreases functionality of macrophages and promotes a hyperinflammatory phenotype.
17 Animals with dietary Pb exposure resulting in blood Pb levels (upon cessation of
18 exposure) relevant to humans, 8.2 and 18 (ig/dL, had reduced macrophage generation and
19 phagocytosis (Bunn et al., 200Ic; Kowolenko et al., 1991). Some in vitro studies
20 (Bussolaro et al.. 2008; Zhouetal.. 1985) provided supporting evidence. Several
21 observations link Pb exposure to impaired function and/or structure of specialized
22 macrophages in nonlymphoid tissue, including liver Kupffer cells and alveolar
23 macrophages. The results suggest that immune dysfunction may contribute to the effects
24 of Pb on dysfunction in nonlymphoid tissues and provides a link between immune
25 dysfunction and disease in other organ systems. However, the implications of findings to
26 effects in humans are uncertain because in several studies, animals were treated with Pb
27 by injection and/or had high blood Pb levels. Evidence for Pb-induced decreases in
28 macrophage functionality provides mode of action support for observations of
29 Pb-induced decreased host resistance in animals (Section 5.6.5.1). The sparse
30 epidemiologic evidence is not conclusive but was a lesser consideration than the
31 toxicological evidence in drawing conclusions about the effects of Pb exposure on
32 macrophages because most epidemiologic studies did not examine the functional state of
33 macrophages. The study that examined the functional state of macrophages,
34 i.e., mediators of host defense or inflammation, found an association with blood Pb level
35 in children (Pineda-Zavaleta et al., 2004) consistent with toxicological evidence.
36 Occupational studies examined abundance of monocytes or markers of activated
37 macrophages plus other antigen presenting cells, and evidence did not clearly indicate a
38 difference in Pb-exposed workers. Inconsistency among studies was not related to
39 differences in sample size, age, or blood Pb levels of Pb-exposed workers. None of the
November 2012 5-439 Draft - Do Not Cite or Quote
-------
1 studies considered the potential influence of other occupational exposures or provided
2 concentration-response information.
5.6.2.5 Neutrophils
3 In the 2006 Pb AQCD, Pb exposure was not judged to have a strong effect on
4 neutrophils, which comprise the majority of polymorphonuclear cells (PMNs) (U.S. EPA.
5 2006b). This conclusion was based on the limited available toxicological evidence as
6 compared with that for effects on other immune cells. However, the modulation of
7 neutrophil activity may have important consequences on the dysregulation of
8 inflammation and ability of organisms to respond to infectious agents. Studies of cultured
9 human PMNs (Governaet al.. 1987) and occupationally-exposed adults (Queiroz et al..
10 1994a; Queiroz et al., 1993; Valentino et al.. 1991; Bergeret et al.. 1990) found
11 Pb-associated reductions in PMN functionality, as indicated by reduced chemotactic
12 response, phagocytic activity, respiratory oxidative burst activity, or reduced ability to
13 kill ingested antigen. These observations were made in groups of Pb-exposed workers
14 (n = 10-60) with range of mean age 34-41 years and blood Pb levels 14.8-91.4 (ig/dL.
15 These studies were focused on neutrophil function, and while the evidence did not appear
16 to be influenced by multiple comparisons, it could have been influenced by publication
17 bias. In these cross-sectional studies, the temporal sequence between Pb exposure and
18 neutrophil function cannot be determined. Other limitations across all studies include the
19 lack of consideration for potential confounding factors, including other workplace
20 exposures, and high blood Pb levels of Pb-exposed workers.
21 Instead of examining neutrophil functional activities, the few available recent studies of
22 animals and occupationally-exposed adults examined neutrophil counts, an increase in
23 which has been interpreted by some investigators to be a compensatory response to
24 Pb-induced impairment in neutrophil chemotactic activity and a hyperinflammatory
25 response. A study in male Wistar rats found that 12 mg Pb spheres implanted in brains
26 (compared with control glass spheres) resulted in greater neutrophil filtration from day
27 7-28 with inflammatory-related damage that included apoptosis and indications of
28 neurodegeneration (Nakao et al., 2010). However, the route of Pb administration has
29 uncertain relevance to the typical routes of Pb exposure in humans.
30 Occupational studies produced contrasting results that were not related to the blood Pb
31 levels of workers. DiLorenzo et al. (2006) found a Pb-associated higher absolute
32 neutrophil count (ANC) in analyses that adjusted for potential confounding factors and
33 showed a concentration-dependent relationship. In an analysis combining 68 ceramic, Pb
34 recycling, or bullet manufacturing workers and 50 control food plant workers in Italy, a
November 2012 5-440 Draft - Do Not Cite or Quote
-------
1 1 (ig/dL higher concurrent blood Pb level was associated with a 21.8 cells/(iL (95% CI:
2 11.2, 32.4 cells/(iL) higher ANC. While these results were adjusted for age, current BMI,
3 and current smoking status, other workplace exposures were not examined. Pb-exposed
4 workers had a mean age 44 years and a geometric mean concurrent blood Pb level of
5 20.5 (ig/dL. Controls had a mean age 46.8 years and mean blood Pb level of 3.5 (ig/dL.
6 Neutrophilia (n >7,500 cells/mm3) was found in 8 workers described to have medium to
7 high Pb exposures (exact blood Pb levels not reported) but no controls had suggesting
8 that long-term, higher-level Pb exposures can lead to a biologically meaningful excess of
9 circulating neutrophils. Additionally, a blood Pb concentration-dependent relationship
10 was indicated by observations of a monotonic increase in ANC across increasing blood
11 Pb level groups: controls, workers with blood Pb levels < 30 (ig/dL, and workers with
12 blood Pb levels >30 (ig/dL. Results further indicated an interaction between concurrent
13 blood Pb level and current smoking. ANC increased across the three blood Pb groups
14 among current smokers but not nonsmokers. In contrast, in a study that did not consider
15 any potential confounding factors, Conterato et al. (In Press) found lower neutrophil
16 concentrations among 23 battery workers and 50 painters in Brazil with mean concurrent
17 blood Pb levels of 50.0 and 5.4 (ig/dL, respectively, than among 36 controls with a mean
18 blood Pb level of 1.5 (ig/dL. Pb-exposed workers did not consistently have higher levels
19 of eosinophils, basophils, monocytes, or total lymphocytes either.
20 Support for the decreased neutrophil function found in Pb-exposed workers is provided
21 by findings of Pb-associated increases in TNF-a (Section 5.6.6.1) and complement,
22 which are mediators of neutrophil proliferation, survival, maturation, and functional
23 activation. The complement system is a component of the innate immune system that is
24 involved in chemotaxis of macrophages and neutrophils and phagocytosis of antigens.
25 The few available occupational studies found lower complement in Pb-exposed workers
26 with mean blood Pb levels >60 (ig/dL (Undeger et al.. 1996; Ewers etal.. 1982). higher
27 than those relevant to the U.S. general population. Neither study considered potential
28 confounding factors, including other workplace exposures. The evidence has limited
29 implications also because the cross-sectional nature of studies cannot establish the
30 temporal sequence between Pb exposure and complement.
31 In summary, previous occupational studies provided evidence for the effects of Pb
32 exposure on neutrophils by finding that compared with controls, Pb-exposed workers had
33 lower neutrophil functionality (Queiroz et al.. 1994a: Queiroz etal.. 1993; Valentino et
34 al.. 1991; Bergeret et al.. 1990) and lower complement (Undeger et al.. 1996; Ewers et
35 al.. 1982). which is a mediator of phagocyte functionality. The limited number of recent
36 epidemiologic studies examined only neutrophil abundance and conclusively did not find
37 Pb-exposed workers to have higher or lower neutrophil abundance. While there is
38 evidence for Pb-associated reduced neutrophil functionality, firm conclusions are not
November 2012 5-441 Draft - Do Not Cite or Quote
-------
1 warranted because the results are based on cross-sectional examination of male workers
2 with relatively high blood Pb levels (range: 18.6-100 (ig/dL), and they lack consideration
3 for potential confounding factors including other occupational exposures, concentration-
4 response information, and analogous toxicological evidence.
5.6.2.6 Dendritic Cells
5 Whereas as research reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b) focused on
6 examining T cells (Section 5.6.2.1). recent ex vivo and in vitro results suggest that the
7 effects of Pb on suppressing Thl activity and promoting Th2 activity may be a
8 consequence of the direct action of Pb on the function of dendritic cells, a major APC.
9 Gao et al. (2007) found that 25 (iM Pb chloride exposure for 10 days stimulated dendritic
10 cell maturation in bone marrow cultures by changing the ratio of cell surface markers
11 (e-g-, CD86/CD80) that promote Th2 cell development. Additionally, upon activation
12 with LPS, Pb-matured dendritic cells produced less IL-6, TNF-a, and IL-12 (stimulates
13 growth and differentiation of T cells) than did control cells but the same amount of IL-10
14 (inhibits production of Thl cytokines). The effect of Pb in altering the cytokine
15 expression profile of dendritic cells, in particular, the lower IL-12/IL-10 ratio, may serve
16 as an important signal to shift naive T cell populations toward a Th2 phenotype.
17 Supporting a role for dendritic cells in skewing to a Th2 phenotype, ex vivo results from
18 the same study showed that Pb-naive adult BALB/c mice implanted with Pb-exposed
19 dendritic cells had suppressed DTH (Section 5.6.2.3) and IgG2a antibody (Section 5.6.3)
20 responses (Gao et al.. 2007).
5.6.2.7 Natural Killer Cells
21 Based mostly on studies reviewed in the 2006 Pb AQCD, evidence does not clearly
22 indicate that Pb exposure affects the innate immune NK cells, which mediate host
23 defense by killing infected cells. Some epidemiologic studies adjusted for factors such as
24 age, sex, and smoking but did not find differences in NK cell abundance or level of
25 functional activity by blood Pb level in children or adults with or without occupational
26 exposures (n = 145-675) (Karmaus et al.. 2005; Sarasuaet al.. 2000; Pinkerton et al..
27 1998). Studies in children did not examine potential confounding by SES, and the study
28 in Pb-exposed workers did not examine other workplace exposures. Other smaller
29 (n = 30-108) studies that did not consider potential confounding factors found a positive
30 correlation between blood Pb level and NK cell abundance in adults in Italy (Boscolo et
31 al.. 2000; 1999) or reported no significant association in children (quantitative results not
32 reported) (Belles-Isles et al.. 2002). Pb-exposed workers in the U.S., Europe, and Asia
November 2012 5-442 Draft - Do Not Cite or Quote
-------
1 (n = 25-141, mean ages: 26-49 years) with higher concurrent blood Pb levels (means:
2 6.5-128 (ig/dL) had similar means of NK cell abundance or functional activity as did
3 unexposed controls (n = 10-84, mean blood Pb levels: <2-16.7 (ig/dL, mean ages: 28-47
4 years) (Garcia-Leston et al.. 2011; Mishra et al.. 2003; Pinkerton et al.. 1998; Yiicesov et
5 al.. 1997b: Undeger et al.. 1996; Fischbein et al.. 1993; Kimberetal.. 1986).
6 The epidemiologic evidence is not sufficiently informative for drawing conclusions about
7 the effects of Pb exposure on NK cells because of its many limitations including cross-
8 sectional nature, limited consideration for potential confounding, and lack of
9 concentration-response information. However, toxicological evidence equally does not
10 clearly indicate an effect of Pb on NK cells. A decrease in NK cell activity was found in
11 6-8 week-old BALB/c mice but with higher Pb exposure than that relevant to humans
12 (1,300 ppm Pb acetate in drinking water, 10 days, blood Pb level -100 (ig/dL) (Queiroz
13 etal.. 2011). In an in vitro study, Fortier et al. (2008) found that Pb chloride
14 (7.5-20.7 (ig/dL) did not affect NK cytotoxicity compared with the control. However,
15 Pb chloride was not found to affect monocytes or lymphocytes either.
5.6.3 Humoral Immunity
16 The 2006 Pb AQCD (U.S. EPA. 2006b) described another hallmark effect of Pb on the
17 immune system to be an enhanced humoral immune response as characterized by
18 increased production of IgE antibodies (U.S. EPA. 2006b). Several toxicological and
19 epidemiologic studies (Table 5-32) demonstrated Pb-associated increases in IgE, which
20 mediates inflammation in allergic and allergic asthma responses by binding to mast cells
21 and releasing histamines, leukotrienes, and interleukins upon exposure to an allergen.
22 Neither toxicological nor epidemiologic evidence (Table 5-32) consistently indicated that
23 Pb exposure was associated with changes in other classes of Igs including IgG, IgM, and
24 IgA, which function in complement activation and host resistance or activation of
25 immune cells.
26 In toxicological evidence, there was a lack of coherence between results for IgE and
27 activation of B cells, which regulate IgE production through differentiation into antibody-
28 producing cells. Pb chloride exposure in vitro (10 (iM up to 5 days) was found to increase
29 markers of B cell activation, including cell surface markers and levels of plaque forming
30 cells (PFC), which are a measure of antibody-forming cells (McCabe and Lawrence.
31 1990; Lawrence. 198 la). However, several studies in animals (Swiss mice and rabbits)
32 found a wide range of Pb concentrations (0.5 to 250 ppm Pb acetate or tetraethyl Pb for
33 3-10 weeks, postnatal via drinking water) to decrease levels of PFC (Blakley et al.. 1980;
34 Koller and Kovacic. 1974; Koller. 1973). Among many mice strains tested, Mudzinski et
November 2012 5-443 Draft - Do Not Cite or Quote
-------
1 al. (1986) found an increase in PFC only in BALB/c mice with 8-week postnatal dietary
2 Pb acetate exposure that produced high blood Pb levels, 70 (ig/dL. Epidemiologic studies
3 of children (Table 5-32) and adults (Table 5-32) with group comparisons and (Boscolo et
4 al.. 2000; Boscolo et al.. 1999) with correlation analyses) did not find a consistent
5 association between blood Pb level and the abundance of B cells, which may not reflect
6 activation. Inconsistencies among studies did not appear to be related to differences in
7 age, group blood Pb levels, or the extent of consideration for potential confounding
8 (Table 5-32).
9 Most animal studies found Pb-induced increases in IgE, with key evidence provided by
10 studies that examined Pb acetate exposure through drinking water during the gestation
11 and/or lactation period and IgE (Snyder et al.. 2000; Miller et al.. 1998). In particular,
12 Snyder et al. (2000) found elevated IgE in juvenile BALB/c mice with relevant blood Pb
13 levels, means 5-20 (ig/dL measured 0-1 week after gestational and/or lactational Pb
14 exposure. In Miller et al. (1998). elevated IgE was found in adult mice exposed
15 gestationally to Pb via drinking water of dams who had blood Pb levels of 30-39 (ig/dL.
16 Chen et al. (2004) did not find gestational dietary Pb acetate exposure to result in an
17 increase in IgE in adult F344 rats who had blood Pb levels of 6.75 and 8 (ig/dL, measured
18 one week postweaning. In BALB/c and OVA-transgenic (produce OVA-specific T cells)
19 mice, Heo et al. (1997; 1996) found concomitant Pb-induced increases in IgE and IL-4,
20 consistent with the mode of action of IL-4 to induce class switching of B cells to IgE
21 producing cells. However, these effects were observed with Pb administered via s.c.
22 injection (50 (ig/100 (iL, 3 times per week for 3 weeks) and with higher blood Pb levels,
23 38 (ig/dL, than those relevant to humans. Some of these studies examined multiple
24 immune endpoints; however, chance findings due to multiple comparisons likely are not
25 responsible for the findings because the pattern of results consistently pointed to a shift
26 from Thl to Th2 responses (Miller etal.. 1998; Heoetal.. 1997; 1996).
November 2012 5-444 Draft - Do Not Cite or Quote
-------
Table 5-32 Comparison of serum immunoglobulin levels and B cell abundance
among various blood Pb groups.
Study
Study Population and
Methodological Details
Blood Pb
Level
(ug/dL)
|gEa
igGb
igMb
igAb
B cells0
Children
Karmaus et
al. (2005)
331 children, ages 7-1 0 yr, Hesse,
Germany
Cross-sectional. School-based
recruitment. No information of
participation rate. Most subjects live
<2.2
2.21 -2.83
2.84-3.41
>3.41
46 (1 .0)
30 (0.65)
59 (1 .28)
59 (1 .28)d
1210
1214
1241
1201
150
143
153
148
123
121
133
136
418e(1.0)
353 (0.84)
389 (0.93)
393 (0.94)
near industrial facilities. Multiple
exposures examined. Results adjusted
for age, sex, number cigarettes/day
smoked in home in previous 12 months,
number of infections in the previous
12 months, serum lipid level, and blood
organochlorine level. No consideration
for potential confounding by SES,
allergens. Monotonic C-R for IgE except
in highest blood Pb group.
Sarasua et
al. (2000)
Sarasua et
al. (2000)
Sarasua et
al. (2000)
Lutz et al.
(1999)
Hegazy et
al. (2011)
Zhao et al.
(2004)
382 children, ages 6-30 mo, Multiple
U.S. locations
Cross-sectional. No information on
participation rate. Large proportion with
residence near Pb sources.
Comparison group age- and
demographically-matched. Results
adjusted for age, sex, and study
location. No consideration for potential
confounding by SES, allergens.
Inconsistent C-R.
562 children, ages 36-71 mo, Multiple
U.S. locations
Same methodology as above.
675 children ages 5-16 yr, Multiple U.S.
locations
Same methodology as above.
279 children, ages 9 mo-6 yr,
Springfield, MO
Cross-sectional. Recruitment from
public assistance and Pb poisoning
prevention program. No information on
participation rate. Results adjusted for
age. Lack of rigorous statistical
methods. No consideration for potential
confounding by SES, allergens.
Monotonic C-R for IgE except in highest
blood Pb group.
31 8 children, ages 6 mo-7 yr, Egypt
Cross-sectional. Clinic-based
recruitment. No information on
participation rate. Lack of rigorous
statistical methods. Potential
confounding not considered. No
monotonic C-R.
73-75 children, ages 3-6 yr, Zhejiang
Province, China
0.6-4.9
5-9.9
10- 14.9
> 15
0.6-4.9
5-9.9
10- 14.9
> 15
0.6-4.9
5-9.9
10- 14.9
> 15
<10
10-14
15-19
20-69
<5
5-9
10-14
15-19
20-44
45-69
<10
> 10
609
666d
680d
630
817
813
856
835
1,031
1 ,094d
1,048
1,221
51.8(1.0)
74.0 (1 .43)
210.7(4.07)
63.7 (1 .23)d
13.0(1.0)
12.0(0.92)
20.8 (1 .60)
14.9(1.15)
20.4 (1 .57)
10.2(0.78)d
103 50.1
108 55.0
105 58.2
124d 61 .4d
120 88.6
116 90.9
125 96.3
121 94.1
128 140
131 143
136 140
106 108
19.1 (1.0)
20 (1 .05)
20.4 (1 .07)
22.2(1.16)
18.4(1.0)
17.6(0.96)
19.2(1.04)
18.6(1.01)
16.1 (1.0)
15.8(0.98)
15.3(0.95)
20.1 (1.25)
13.4(1.0)
12.6(0.94)
16.9(1.26)
11.1 (0.83)
16.6(1.0)
16.8(1.01)
November 2012
5-445
Draft - Do Not Cite or Quote
-------
Study
Sun et al.
(2003)
Study Population and
Methodological Details
Cross-sectional. School-based
recruitment. No information on
participation rate. Lack of rigorous
statistical methods. Potential
confounding not considered.
Blood Pb
Level
(ug/dL) |gEa lgGb
<10 32.50 40.53
>10 41.33f 34.76f
lgMb lgAb B cells0
41.53
31 .74f
Adults without Occupational Pb Exposures
Sarasua et
al. (2000)
Adults with
Pinkerton et
al. (1998)
Fischbein et
al. (1993)
Kimber et
al. (1986)
Heoetal.
(2004)
Anetorand
Adeniyi
(1998)
Ewers et al.
(1982)
433 children and adults, ages 16-75 yr,
Multiple U.S. locations
Same methodology as that in children.
Occupational Pb Exposures
84 hardware factory controls, mean age
30 yr
145 male Pb smelter workers, mean
age 33 yr, U.S., exact location NR
Cross-sectional. Results adjusted for
age, race, current smoking status, and
workshift. No consideration for potential
confounding by other workplace
exposures, SES
36 industrial worker controls, mean age
47 yr
36 firearms instructors, mean age 49 yr
15 firearms instructors, mean age 48 yr
New York metropolitan area
Cross-sectional. Lack of rigorous
statistical methods. Potential
confounding not considered.
21 unexposed male controls, ages
20-60 yr
39 male tetraethyl Pb plant workers,
ages 25-61 yr, U.K.
Cross-sectional. Lack of rigorous
statistical methods. Potential
confounding not considered.
606 Pb battery plant workers, Korea
Cross-sectional. Monotonic C-R found;
lack of rigorous statistical methods.
Potential confounding not considered.
50 male controls, ages 22-58 yr
80 male Pb-exposed workers, ages
21-66yr, Nigeria
Cross-sectional. Lack of rigorous
statistical methods. Potential
confounding not considered.
53 male various occupation controls,
ages 21-54yr
72 male Pb battery/smelter workers,
ages 16-58 yr, Germany
Cross-sectional. Lack of rigorous
statistical methods. Potential
confounding not considered.
0.6-4.9 1 ,099
5-9.9 1 ,085
10-14.9 1,231
>15 1,169
<2 1 ,090
Mean: 39 1,110
NR
Mean: 14.6
Mean: 31.4
Mean: 11. 8 1062
Mean: 38.4 1018
<10 112.5(1.0)
10-29 223.3(1.99)
> 30 535.8 (4.76)d
Mean: 30.4 1,997
Mean: 56.3 1,1 87d
Mean: 11. 7 1939
Mean: 59.0 171
175 252 13.9(1.0)
175 242 13.0(0.94)
262d 283 12.4(0.89)
139 193 14.8(1.06)
94.5 180 14.6(1.0)
106.2 202 13.2(0.90)
8.6(1.0)
10.5(1.22)
11.2(1.3)"
1294 2235
1040 2425
215 188
191 144d
1619 1409
127 128
November 2012
5-446
Draft - Do Not Cite or Quote
-------
Study Population and
Study Methodological Details
Blood Pb
Level
(ug/dL)
igEa
lgGb
lgMb lgAb B cells0
Undeger et 25 male university worker controls,
al. (1996) ages 22-56 yr
25 male Pb battery plant workers, ages
22-55 yr, Turkey
Cross-sectional. Lack of rigorous
statistical methods. Potential
confounding not considered.
Mean: 16.7
Mean: 74.8
1202.1 140.4 210.3 545.5e(1.0)
854.6d 93.3d 168.1 635.9e (1.2)
Alomran
and
Shleamoon
(1988)
18 management personnel controls
39 Pb battery workers, mean age 35 yr
Iraq
Cross-sectional. Controls age matched.
Lack of rigorous statistical methods.
Potential confounding not considered.
NR
NR
1713
1610
183
170
Note: Results are presented in order of quality of study design and methodology.
algE data are presented as lU/mL unless otherwise specified. (In parentheses are ratios of IgE in the higher blood Pb group to IgE in
the lowest blood Pb group.)
bOther Ig data are presented as mg/dL unless otherwise specified.
°B cell data are presented as the percentage of B cells among all lymphocytes unless otherwise specified. (In parentheses are the
ratio of B cells in the higher blood Pb group to B cells in the lowest blood Pb group.)
dp <0.05 for group differences.
eData represent the number of cells/uL serum.
'Data represent the mean rank for Mann-Whitney U test, p = 0.07 for IgE.
9Data are presented as lU/mL.
1
2
o
J
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
A small number of available recent toxicological studies examined IgG subtypes, and as
in previous studies, found inconsistent effects of Pb exposure. Kasten-Jolly et al. (2010)
examined 100 (iM Pb acetate in drinking water of BALB/c dams GD8-PND21 because it
produced blood Pb levels relevant to humans in another study, i.e., 10-30 (ig/dL (Snvder
et al.. 2000). Pb-exposed pups had increases in the expression of genes encoding Ig
antibodies and those involved in B lymphocyte function and activation. These genes
included those for the heavy chain of IgM, IL-4, IL-7 and IL-7 receptor, IL-21, RAG-2,
CD antigen 27, B-cell leukemia/lymphoma 6, RNA binding motif protein 24,
Histocompatibility class II antigen A (beta 1), Notch gene homolog 2, and histone
deacetylase 7A. These results were produced by amicroarray analysis of hundreds of
genes, which is subject to a higher probability of finding an effect by chance.
Other recent studies examined specific IgG subtypes, did not find Pb-induced changes in
a consistent direction, and thus did not clearly indicate a shift to Th2 responses. A
limitation of this evidence is the higher blood Pb levels of animals than those relevant to
humans. Fernandez-Cabezudo et al. (2007) reported evidence for a shift to Th2 responses
following Salmonella infection in C3H/HeN mice exposed postnatally to IxlO4 (iM
Pb acetate in drinking water for 16 weeks (resultant mean blood Pb level: 106 (ig/dL).
Relative to control mice, production of the Th2 cytokine IL-4 increased in spleen cells of
Pb-exposed mice after infection as did serum levels of Salmonella-specific IgGl.
Infection increased Thl-mediated IgG2a levels in control but not Pb-exposed mice.
November 2012
5-447
Draft - Do Not Cite or Quote
-------
1 In contrast, Gao et al. (2006) found a Pb-induced increased IgG2a/IgGl ratio, albeit via
2 i.p. injection (50 (ig Pb chloride, 3 times per week for 3 weeks), high blood Pb levels
3 (65 (ig/dL), and in a highly-specialized strain of adult knockout mice lacking the ability
4 to produce IFN-y. This result was surprising given evidence that IFN-y usually directs
5 secretion of IgG2a; however, the authors suggested that in these knockout mice, Pb may
6 initiate a Thl response via an IFN-y independent pathway to enhance IgG2a production.
7 Carey et al. (2006) found concentration-dependent increases in both IgG2a- and IgGl-
8 producing cells (after 7 days) in adult BALB/c mice treated with subsensitizing doses of a
9 T cell-independent (Trinitrophenyl-Ficoll [TNP-Ficoll]) or T cell-dependent (TNP-
10 ovalbumin [TNP-OVA]) hapten-protein conjugate and 25-50 (ig Pb chloride by bolus
11 injection. These results indicated stimulation of both Thl- and Th2-mediated
12 mechanisms. Pb treatment also increased the numbers of T and B cells and IgM-
13 producing cells in the lymph node against both TNP-Ficoll and TNP-OVA. The increase
14 in IgM-producing cells against TNP-Ficoll indicated a T-cell independent mechanism.
15 Despite finding increases in both IgGl- and IgG2a-producing cells, the authors concluded
16 that Pb skewed the response toward Th2 based on observations of Pb-induced increases
17 in T and B cells and suppression of DTH. Thus, the results indicated the potential for Pb
18 to promote allergic sensitization against T-dependent antigens.
19 Observations of Pb-induced increases in IgE in animals provide biological plausibility for
20 associations observed between higher Pb biomarkers levels and higher serum IgE levels
21 in various populations of children, although a monotonic concentration-dependent
22 increase was not consistently observed (Hegazy et al.. 2011; Hon et al.. 2010; Hon et al..
23 2009; Karmaus et al., 2005; Annesi-Maesano et al.. 2003; Sun et al., 2003; Lutz et al..
24 1999) (Table 5-32). The evidence was based on cross-sectional analyses which preclude
25 establishing the temporal sequence between Pb exposure and IgE. Associations between
26 blood Pb level and IgE were found in studies that generally had population-based
27 recruitment. Most studies did not provide sufficient information to assess the potential for
28 biased participation by Pb exposure and immune conditions. Most studies examined
29 multiple immune endpoints; however, associations were not isolated to IgE.
30 Karmaus et al. (2005) had greater adjustment for potential confounding factors and
31 examined associations with lower blood Pb levels. Compared with 82 children (ages 7-10
32 years) in Germany with concurrent blood Pb level <2.2 (ig/dL, children with blood Pb
33 levels 2.84-3.41 (ig/dL (n = 86) and >3.4 (ig/dL (n = 82) had 28% higher serum IgE
34 levels (p = 0.03, F-test). These differences were found with the adjustment for age, blood
35 organochlorine levels, serum lipid levels, number of infections in the previous 12 months,
36 and number of cigarettes/day smoked in the home in the previous 12 months. SES was
37 not examined. A monotonic increase in IgE was found across blood Pb quartiles, except
38 for the highest group (Table 5-32). Although IgE was elevated in children with relatively
November 2012 5-448 Draft - Do Not Cite or Quote
-------
1 low blood Pb levels (>2.84 (ig/dL), in these children ages 7-10 years, the contribution of
2 higher past Pb exposures cannot be excluded. Similar differences in IgE count on
3 basophils were not found among the blood Pb quartiles. Although serum IgE and
4 basophil-bound IgE have been correlated in adults (Malveaux et al.. 1978; Conrov et al..
5 1977). few data are available in children (Dehlink et al., 2010). A study in children found
6 that serum IgE levels were not correlated with basophil-bound IgE (Spearman r = -0.003)
7 but were correlated with other IgE receptor-expressing cells such as dendritic cells and
8 monocytes (Spearman r = 0.43 to 0.65, p <0.05) (Dehlink etal. 2010). The number of
9 IgE-bound basophils also has been highly variable across individuals, particularly
10 children (Hausmann et al., 2011; Dehlink et al., 2010). Thus, it is not unexpected that
11 higher blood Pb level was associated with higher serum IgE but not basophil-bound IgE
12 counts in Karmaus et al. (2005). In this study, blood Pb level was not associated with
13 serum levels of IgG, IgA, IgM or B cell abundance. Lutz et al. (1999) found higher serum
14 IgE in low SES children (9 months-6 years) on public assistance in Springfield, MO after
15 only adjusting for age, albeit with concurrent blood Pb levels >10 (ig/dL (n = 105/279).
16 Recent studies in children (non-U.S.) also reported associations between concurrent
17 blood Pb level and elevated serum IgE but did not adjust for potential confounding
18 factors (Hegazy et al.. 2011: Honetal.. 2010: Hon et al.. 2009). Ron et al. (2010: 2009)
19 demonstrated associations in 110 children (mean age 9.9 years) with atopic dermatitis in
20 Hong Kong with low blood Pb levels (range: 1.4-6.0 (ig/dL) and found that blood Pb
21 level also was correlated with severity of atopic dermatitis, a condition commonly
22 characterized by elevated IgE levels. Among 318 children, ages 6 months to 7 years, in
23 Egypt, Hegazy et al. (2011) did not find a monotonic increase across the blood Pb groups.
24 Sarasua et al. (2000) did not examine IgE but found associations of higher concurrent
25 blood Pb level with higher IgA, IgG, and IgM in 372 U.S. children ages 6-35 months but
26 not older (36-71 months, 6-15 years, 16-75 years, n = 433-673). In the youngest age
27 group, a 1 (ig/dL higher blood Pb level was associated with a 0.8 [95% CI: 0.2, 1.4], 4.8
28 [95% CI: 1.2, 8.4], and 1.0 [95% CI: 0.1, 1.9] mg/dL higher IgA, IgG, and IgM,
29 respectively, adjusted for age, sex, and location. In the youngest children, serum levels of
30 all three examined Igs were elevated among the 24 children with concurrent blood Pb
31 levels > 15 (ig/dL than among the 165 children with blood Pb levels <5 (ig/dL.
32 While most epidemiologic studies examined concurrent blood Pb levels, some studies
33 indicated that prenatal Pb exposure may impact Ig levels in newborns (Annesi-Maesano
34 et al.. 2003: Belles-Isles et al.. 2002). While these studies better indicated the temporal
35 sequence between Pb exposure and Ig levels, an important limitation was the lack of
36 extensive consideration for potential confounding factors. Belles-Isles et al. (2002)
37 examined 97 newborns in Quebec, Canada (from subsistence fishing communities and
November 2012 5-449 Draft - Do Not Cite or Quote
-------
1 two other towns, geometric mean blood Pb levels: 1.64 and 1.32 (ig/dL, respectively) and
2 found an association between higher cord blood Pb level and higher cord serum IgG
3 adjusted for prenatal maternal smoking status. However, IgG also was associated with
4 cord plasma organochlorines, which also are elevated with high fish diets. Annesi-
5 Maesano et al. (2003) found that Pb level in infant hair (mean: 1.38 ppm, n = 234) but not
6 cord blood (mean: 6.7 (ig/dL, n = 326) or placenta (mean: 9.6 (ig/dL, n = 332) was
7 associated with cord serum IgE in newborns in Paris. Potential confounding was not
8 considered. The authors inferred a stronger effect of Pb exposure integrated over the
9 entire gestational period compared to exposures closer to birth. However, an empirical
10 basis for interpreting Pb levels in hair has not been established (Section 4.3.4.2). Cotinine
11 was not associated significantly with Pb biomarker levels or IgE. The correlation
12 (Spearman r = 0.21, p <0.01) was larger among the 67% infants with mothers without
13 allergies than infants with maternal allergies (Spearman r = 0.12), pointing to the possible
14 masking of a blood Pb-IgE association by the stronger association of family history of
15 allergy.
16 Blood Pb level also was associated with IgE in adults without (Pizent et al.. 2008) and
17 with occupational Pb exposure (Heo et al.. 2004). Pizent et al. (2008) adjusted for
18 potential confounding by age, pack years smoking, and alcohol consumption and found
19 that higher concurrent blood Pb level was associated with higher IgE in 166 women in
20 Zagreb, Croatia of similar SES (i.e., white-collar office workers) (Pizent et al.. 2008).
21 Among women not on hormone replacement therapy or oral contraceptives, a 1 (ig/dL
22 higher blood Pb level was associated with a 0.60 (95% CI: 0.58, 1.18) higher log of IgE.
23 Concurrent blood Pb levels were low in these women who were aged 19-67 years (mean:
24 2.16 (ig/dL, range: 0.56-7.35 (ig/dL); however, the cross-sectional study design makes it
25 difficult to characterize the temporal sequence between exposure and outcome or the
26 timing, level, frequency, and duration of Pb exposure that contributed to the observed
27 association. Authors did not report an effect estimate in men because it did not attain
28 statistical significance. Without quantitative results, it is difficult to ascertain whether
29 there was suggestion of association in men but insufficient power to indicate statistical
30 significance due to the smaller number of men examined (50 men versus 166 females).
31 Another study of 34 men with and without allergy in Italy also did not report quantitative
32 results and only indicated a lack of statistically significant correlation between concurrent
33 blood Pb level (median: 11 (ig/dL) and IgE without considering potential confounding
34 (Boscolo et al.. 1999V
35 Limitations of the collective epidemiologic evidence for IgE include the cross-sectional
36 analyses with limited adjustment for potential confounding. Karmaus et al. (2005) found
37 a blood Pb-IgE association with adjustment for age, blood organochlorine levels, serum
38 lipid levels, number of infections in the previous 12 months, and number of cigarettes
November 2012 5-450 Draft - Do Not Cite or Quote
-------
1 a day smoked in the home in the previous 12 months. Lutz et al. (1999) comprised a low
2 SES population on public assistance; however, none of the studies adjusted for SES or
3 allergen exposure. Lower SES has been associated with poorer housing conditions,
4 higher exposures to Pb, allergens, and other factors associated with allergy and asthma.
5 Allergen exposure and lower SES are associated with higher IgE and related conditions
6 such as allergy and asthma (Bryant-Stephens. 2009; Dowd and Aiello. 2009; Aligne et
7 al., 2000). Most studies did not provide detailed demographic or residential information.
8 Thus, there is uncertainty as to the extent to which the blood Pb-IgE associations
9 observed in children may be confounded by unmeasured SES and/or allergen exposure.
10 The cross-sectional nature of evidence raises the possibility that associations are due to
11 reverse causality, i.e., children with higher IgE may have increased lung permeability,
12 and consequently, greater uptake of inhaled Pb into the blood. Studies have not directly
13 compared Pb uptake in groups with different IgE levels; however, animals sensitized with
14 allergens to produce higher IgE have not had greater uptake of radiolabeled particles than
15 controls (Turi et al., 2011; Erjefalt and Persson, 1991). Histamine was shown only
16 transiently to increase uptake of particles in a baboon (n = 1) (Yeates and Hameister.
17 1992) and in humans, in both those with and without asthma (Rees etal.. 1985; Braude et
18 al.. 1984; O'Byrne etal.. 1982). Histamine is released by IgE-bound mast cells and
19 basophils upon exposure to sensitized antigens and leads to inflammation. Compared
20 with healthy controls (n = 6-9), subjects with asthma (n = 9-13) did not consistently have
21 greater particle uptake into blood (Del Donno et al., 1997; Rees etal.. 1985; O'Bvrne et
22 al.. 1984; Elwoodetal.. 1983). Thus, evidence does not strongly link higher
23 inflammation with increased uptake of particles into the blood which reduces the
24 likelihood that blood Pb-IgE associations observed in children or adults are attributable to
25 reverse causality.
26 Most of the epidemiologic evidence about the effects of Pb on IgA, IgG, and IgM levels
27 is provided by previous studies of Pb-exposed workers (mostly males) from various
28 industries with mean ages 32-36 years and mean blood Pb levels 38-74.8 (ig/dL (Anetor
29 andAdeniyi. 1998; Pinkerton et al.. 1998; Undeger et al.. 1996; Queiroz et al.. 1994b:
30 Alomran and Shleamoon. 1988; Kimber et al., 1986; Ewers et al.. 1982). As are
31 toxicological findings for these other Ig classes, epidemiologic evidence is mixed, with
32 studies reporting higher, lower, and similar Ig levels in Pb-exposed workers (n = 25-145)
33 compared with unexposed controls (n = 18-84). Some studies reporting lower Ig levels in
34 Pb-exposed workers included workers with the highest mean blood Pb levels (>50 (ig/dL)
35 (Anetor and Adenivi. 1998: Undeger etal.. 1996: Ewers etal.. 1982). The lack of
36 analysis of potential confounding factors, including other workplace exposures, precludes
37 characterization of others factors that may contribute to inconsistent associations in
38 occupational studies.
November 2012 5-451 Draft - Do Not Cite or Quote
-------
1 In summary, evidence for the effects of Pb exposure on humoral immunity largely
2 comprises consistent toxicological and epidemiologic observations of Pb-associated
3 increases in IgE. The combined toxicological and epidemiologic results do not clearly
4 indicate whether Pb exposure affects IgG, IgM, or IgA. Collectively, epidemiologic
5 studies conducted in the U.S., Europe, and Asia, some with large study populations
6 (n = 279-331) indicate higher serum IgE in children with concurrent blood Pb levels
7 >10 (ig/dL. There was some evidence of associations in non-U.S. children with lower
8 blood Pb level; however, the contribution of higher Pb exposures earlier in childhood
9 cannot be excluded (HonetaL 2010; Karmaus et al.. 2005). Pb-associated increases in
10 IgE were found in children with atopic dermatitis (Hon et al.. 2010; 2009) and children
11 without immune conditions (Sun et al.. 2003). Other studies did not report the health
12 status of subjects. Thus, sufficient information was not provided to assess the potential
13 for selection bias. Among studies that provided concentration-response information,
14 some found serum IgE to increase across blood Pb level groups, except for the highest
15 blood Pb groups (Karmaus etal.. 2005: Lutzetal.. 1999). In Hegazy et al. (2011). IgE
16 did not increase monotonically across blood Pb groups. All evidence in humans is based
17 on cross-sectional analyses, making it difficult to establish the temporal sequence
18 between Pb exposure and increase in IgE. Findings of similar particle uptake in subjects
19 with and without acute inflammation of inflammatory conditions increase confidence that
20 blood Pb-IgE associations are not due to reverse causality. Most studies did not consider
21 potential confounding, and none adjusted for SES. A study in children in Germany and a
22 study in adults in Croatia adjusted for age and smoking, with additional adjustment for
23 blood organochlorine levels in children (Karmaus et al.. 2005) and alcohol consumption
24 in adults (Pizent et al., 2008). Blood Pb level was associated with IgE in a low SES
25 population of children in Michigan (Lutz etal.. 1999) and in a population of female
26 office workers of similar SES in Croatia (Pizent et al., 2008). However, uncertainty
27 remains regarding confounding in the associations observed between blood Pb level and
28 IgE in humans. Biological plausibility for the epidemiologic evidence is provided by the
29 Pb-induced increases in IgE observed in most of the animal studies, with some evidence
30 at blood Pb levels relevant to humans (Snvder etal.. 2000; Miller etal.. 1998).
31 Toxicological evidence indicates increases in IgE with gestational and/or postnatal
32 juvenile Pb exposure, whereas epidemiologic evidence points to associations with
33 concurrent blood Pb levels. Because concurrent blood Pb levels in children reflect both
34 recent and past Pb exposures, the combined evidence indicates that cumulative Pb
35 exposures during childhood may affect IgE levels. While evidence for B cell activation is
36 inconsistent, the mode of action for Pb-induced IgE production is well supported by
37 extensive toxicological evidence for Pb-induced increases in the Th2 cytokine, IL-4
38 (Section 5.6.6.1). The coherence between epidemiologic and toxicological findings for
November 2012 5-452 Draft - Do Not Cite or Quote
-------
1 IgE and evidence describing modes of action for increases in IgE supports a relationship
2 between Pb exposure and increases in IgE.
5.6.4 Inflammation
3 The 2006 Pb AQCD (U.S. EPA. 2006b) identified misregulated inflammation a major
4 immune-related effect of Pb based primarily on consistent toxicological evidence for
5 Pb-induced increases in pro-inflammatory cytokines (Section 5.6.6.1). PGE2, and ROS
6 (Section 5.6.6.3). Inflammation has been characterized as a major mode of action for Pb
7 effects in multiple organ systems such as the liver, kidney, and vasculature given that
8 immune cells make up permanent residents and infiltrating cell populations of these other
9 organ systems (Section 5.2.5). Inflammation also provides a link between the evidence
10 for the effects of Pb on modulating immune cell function and production of cytokines and
11 IgE and the evidence for the effects of Pb on immune-based conditions such as infections
12 and asthma and allergy. For example, IL-4-induces increases in IgE, which primes
13 basophils and mast cells to secrete histamine, leukotrienes, and cytokines, which in turn,
14 produce the inflammation associated with asthma and allergy, i.e., airway responsiveness,
15 mucus secretion, respiratory symptoms. Pb-induced inflammation also has been
16 associated with diminished host resistance by inducing local tissue damage. As described
17 in Section 5.6.6. the few available recent toxicological studies support the effects of Pb
18 exposure on inflammation with findings of Pb-induced increases in pro-inflammatory
19 cytokines, ROS and PGE2.
20 The few available epidemiologic studies have found Pb-associated changes in ROS
21 release from macrophages (Section 5.6.6.3) and cytokine levels (Section 5.6.6.1) in
22 children and adults. Adding to this evidence, recent cross-sectional studies found
23 associations between blood Pb level and indicators of inflammation that may be related to
24 multisystemic effects. As discussed in Section 5.6.3. evidence has not provided strong
25 evidence for increased particle uptake in subjects with acute inflammation or
26 inflammatory conditions, reducing the likelihood of reverse causality. However, because
27 of the cross-sectional design of studies, the temporal sequence between Pb exposure and
28 inflammation cannot be established. The most compelling epidemiologic evidence was
29 provided by studies in adults, which were larger had greater consideration for potential
30 confounding. The consistent pattern of association observed across endpoints reduces the
31 likelihood of chance findings due to multiple comparisons.
32 Strengths of the study of adults (age > 40 years, n = 4,663- 7,342) participating in
33 1999-2004 NHANES included the examination of several potential confounding factors,
34 multiple exposures and outcomes in predominately healthy adults and statistical analyses
November 2012 5-453 Draft - Do Not Cite or Quote
-------
1 to provide nationally-representative results. Higher concurrent blood Pb level was
2 associated with higher serum inflammation markers, C-reactive protein (CRP),
3 fibrinogen, and white blood cell (WBC) count, particularly among men (Songdej et al.,
4 2010). Results were adjusted for age; sex; race/ethnicity; education; current income;
5 physical activity; and several factors related to inflammation including, BMI, smoking
6 status, and history of diabetes, inflammatory disease, or cardiovascular disease. For
7 women, most ORs for associations between quintiles of blood Pb and tertiles of CRP,
8 fibrinogen, and WBC count were <1.0 whereas corresponding ORs in men mostly were
9 >1.0. For example, compared with men with concurrent blood Pb level <1.16 (ig/dL, men
10 with blood Pb levels of 1.16-< 1.63 ng/dL, 1.63-<2.17, 2.17-<3.09 (ig/dL, and
11 > 3.09 (ig/dL had elevated odds of higher CRP (OR [95% CI]: 2.22 [1.14, 4.32], 1.67
12 [0.85, 3.28], 2.12 [1.07, 4.21], and 2.85 [1.49, 5.45], respectively). For all inflammation
13 markers, although the highest OR was found in the highest quintile of blood Pb level
14 (> 3.09 (ig/dL), monotonic concentration-dependent increases were not observed.
15 Consistent with NHANES findings, higher concurrent blood Pb level was associated with
16 higher levels of WBCs and IL-6 with adjustment for age, BMI, and current smoking
17 status among 300 university students in Incheon, Korea (Kim et al.. 2007). Adults with
18 allergic conditions or using anti-inflammatory medication were excluded; however,
19 sufficient information was not provided to assess potential selection bias. Larger effects
20 were estimated for the 147 men in the upper two quartiles of blood Pb levels,
21 2.51-10.47 (ig/dL than for the full range of blood Pb levels (n = 150).
22 Low blood Pb levels also were associated with inflammation in a small genome-wide
23 association study that included 37 children with autism and 15 children without autism
24 (ages 2-5 years; blood Pb level range: 0.37 to 5.2 (ig/dL) in California who were unlikely
25 to have had higher past Pb exposures. In models that included age, sex, and autism
26 diagnosis, concurrent blood Pb level was associated with the expression of several genes
27 related to immune function and inflammation, including HLA-DRB and MHC Class II-
28 associated invariant chain CD74 (involved in antigen presentation) (Tian et al.. 2011).
29 Although blood Pb levels were similar between children with and without autism and
30 correlations with gene expression were observed in both groups, they were in opposite
31 directions (positive among children with autism and negative among children without
32 autism). With gene expression arrays, there is a higher probability of chance. Further,
33 there was limited consideration for potential confounding in this study, and the
34 representativeness of findings in children with autism may be limited. However, the
35 results are supported by observations that Pb chloride (10-100 (iM) increases MHC
36 molecule surface expression in mouse and human HLA antigen presenting cells (Guo et
37 al.. 1996a: McCabe and Lawrence. 1991).
November 2012 5-454 Draft - Do Not Cite or Quote
-------
1 In summary, Pb-associated increases in indicators of inflammation such as CRP, WBCs,
2 and IL-6 were found in populations mostly comprising healthy adults with concurrent
3 blood Pb levels 1.16-10 (ig/dL with adjustment for several potential confounding factors
4 (Songdei etal.. 2010; Kim et al. 2007). The analysis of adults participating in NHANES
5 was particularly noteworthy for its representative population and adjustment for age, sex,
6 race/ethnicity, education, current income, physical activity, and several inflammatory
7 conditions (Songdei et al., 2010). Because all evidence was based on cross-sectional
8 analyses, the relative contributions of recent and past Pb exposures to the observed
9 associations are uncertain. Other lines of evidence do not strongly support reverse
10 causality; however, the temporal sequence between Pb exposure and inflammation is
11 difficult to establish. Despite the limited extent and cross-sectional nature of
12 epidemiologic evidence, the biological plausibility is provided by findings of Pb-induced
13 increases in Th2 cell partitioning (Section 5.6.2.1) and IL-6 (Section 5.6.6.1) in
14 toxicological studies. Th2 cells produce IL-6 which is the primary stimulus for
15 expression of CRP and fibrinogen (Hage and Szalai. 2007; Fuller and Zhang. 2001).
5.6.5 Immune-based Diseases
5.6.5.1 Host Resistance
16 The capability of Pb to reduce host resistance of animals to bacteria has been recognized
17 for almost 40 years and was supported by several animal studies described in the
18 2006 Pb AQCD. Several studies demonstrated increased mortality following Pb exposure
19 through drinking water and infection with Listeria monocytogenes. Multiple
20 investigations in the same laboratory indicated increases in body burdens of viable
21 bacteria, mortality, and sickness behavior induced by Listeria exposure in juvenile or
22 adult BALB/c or CBA/J mice exposed postnatally to 500 to 2,000 (iM Pb acetate in
23 drinking water for 3 to 8 weeks (Dyatlov and Lawrence. 2002; Kim and Lawrence. 2000;
24 Kishikawaetal.. 1997; Lawrence. 198 Ib). Decreased bacterial resistance was observed
25 in mice with blood Pb levels (upon cessation of Pb exposure) relevant to humans,
26 i.e., 25 (ig/dL in BALB/c mice exposed PND1-PND22 (Dyatlov and Lawrence. 2002)
27 and 20 (ig/dL in adult C3H/HeN mice with 16-week exposure (Fernandez-Cabezudo et
28 al., 2007). Other studies found mortality from Salmonella or E. coli or reduced clearance
29 of Staphylococcus in mice or rats administered Pb acetate or nitrate via injection, a route
30 of Pb exposure less relevant to humans (Bishavi and Sengupta. 2006; Cook etal., 1975;
31 Hemphill etal.. 1971; Serve etal.. 1966). Although not examined as much, postnatal
32 dietary Pb (mostly Pb acetate) exposure for 4-10 weeks increased mortality of mice and
33 chickens from viral infection (Gupta et al., 2002; Youssef et al., 1996; Exon etal.. 1979;
November 2012 5-455 Draft - Do Not Cite or Quote
-------
1 Thind and Khan. 1978; Gainer. 1977). These effects were observed in animals with high
2 blood (71-313 (ig/dL) (Gupta et al. 2002: Thind and Khan. 1978) or tissue Pb levels
3 (0.12-0.71 ppm) (Exonetal.. 1979).
4 The mode of action for Pb-induced decreased host resistance is well characterized by
5 observations that Pb suppresses Thl-driven acquired immune responses and increases
6 inflammatory responses in target tissue, which may compromise host protective barriers.
7 Host resistance to bacteria such as Listeria requires effective Thl-driven responses
8 including the production of IL-12 and IFN-y (Lara-Tejero and Pamer. 2004) and these
9 have been found to be inhibited by Pb exposure (Section 5.6.6.1). The lack of IFN-y can
10 inhibit appropriate and timely macrophage activation. Strengthening the evidence for
11 Pb-induced decreased host resistance, Fernandez-Cabezudo et al. (2007) found both
12 increased mortality and decreased production of IL-12 and IFN-y (ex vivo in spleen cells)
13 in Salmonella-exposed CH3/HeN mice with blood Pb levels of 20 (ig/dL. Nitric oxide is
14 produced by activated macrophages and has been found to be suppressed by Pb exposure
15 (Section 5.6.6.2). Pb-induced decreases in bacterial clearance have been found in
16 conjunction with reduced NO and macrophage functionality (Bishayi and Sengupta.
17 2006). Further, Pb-induced inflammation has been demonstrated as increases in ROS and
18 PGE2 (Section 5.6.6.3). Additional mode of action evidence was provided recently with
19 observations that developmental Pb exposure of BALB/c mice (100 (iM Pb acetate in
20 drinking water of dams from GD8 to PND21) upregulated splenic gene expression of
21 caspase-12 (Kasten-Jolly et al.. 2010). Caspase-12 is a cysteine protease that functions in
22 apoptosis and activation of pro-inflammatory cytokines and has been linked with a role in
23 the inhibition of bacterial clearance both systemically and in the gut mucosa (Saleh et al..
24 2006).
25 In the few available epidemiologic studies, a range of Pb exposure indicators (i.e., cord or
26 concurrent blood Pb, Pb content in total deposition samples or lichen) was associated
27 with viral and bacterial infections in children. An increase in infections was associated
28 with cord blood Pb levels > 10 (ig/dL in children in Boston, MA (n = 283) (Rabinowitz et
29 al.. 1990) and a mean concurrent blood Pb level of 3.34 (ig/dL in children in Germany
30 (n = 311) (Karmaus et al.. 2005). Similarly, a study found higher frequency of self-
31 reported colds or influenza among 66 Pb battery or smelter plant workers with blood Pb
32 levels 21.3-85.2 (ig/dL than among 53 controls with blood Pb levels 6.6-20.8 (ig/dL
33 (Ewers et al.. 1982). Because of the many limitations, the lack of consideration for
34 potential confounding (Karmaus et al.. 2005; Rabinowitz et al.. 1990; Ewers etal.. 1982).
35 lack statistical rigor in comparisons of mean blood Pb levels by number of infections
36 (Karmaus et al.. 2005; Ewers et al.. 1982). and ecological design (Carreras et al.. 2009).
37 conclusions about the effects of Pb exposure on viral or bacterial infections cannot be
38 drawn based on epidemiologic evidence alone. And, the weak epidemiologic data do not
November 2012 5-456 Draft - Do Not Cite or Quote
-------
1 detract from the consistent findings in animals for Pb-induced decreased host resistance,
2 including those in animals with relevant blood Pb levels, and evidence for modes of
3 action including decreased macrophage function and Thl cytokine production.
4 With few studies available, the effect of Pb on resistance to eukaryotic parasites is not
5 clear. High concentration Pb acetate (> 30 (iM) diminished the ability of macrophages to
6 kill Leishmania enrietti protozoa in vitro (Mauel et al.. 1989). Survival of malaria-
7 infected mice was enhanced with 100 (iM Pb nitrate exposure via drinking water (Koka
8 et al.. 2007). which was attributed to Pb inducing eryptosis and removal of infected
9 erythrocytes and not to Pb-induced alterations in immune function. Nriagu et al. (2008)
10 found that higher blood Pb level was associated with lower malaria prevalence among
11 653 children (ages 2-9 years) from three Nigerian cities with a mean blood Pb level of
12 8.9 (ig/dL. Results were adjusted for age, sex, number of siblings, and other
13 comorbidities such as depressed mood, headaches, and irritability. Given the well-
14 characterized effect of Pb in promoting Th2 activity, it is plausible for Pb to enhance host
15 resistance to parasites that require robust Th2 responses such as helminths. However, this
16 relationship is not well characterized.
5.6.5.2 Asthma and Allergy
17 lexicological evidence and to a lesser extent, epidemiologic evidence, have supported
18 the effects of Pb exposure on stimulating Th2 activity, including increasing production of
19 Th2 cytokines such as IL-4 (Section 5.6.6.1). IgE antibody (Section 5.6.3). and
20 inflammation (Section 5.6.4). These endpoints comprise a well-recognized mode of
21 action for the development and exacerbation of asthma and allergy, which are atopic and
22 inflammatory conditions. Thus, this mechanistic evidence provides support for the small
23 body of epidemiologic evidence indicating associations of blood Pb levels with asthma or
24 allergy in children (Figure 5-35 and Table 5-33). Whereas such evidence reviewed in the
25 2006 Pb AQCD was too sparse to permit conclusions, findings from recent studies add
26 supporting evidence. Children examined in studies of asthma and allergy encompassed a
27 wide age range (i.e., <1-12 years) and across studies, blood Pb was measured at different
28 lifestages. Studies ascertained outcomes with parent report of doctor diagnosis but also
29 more objectively using a surveillance database or and clinical testing. This variability in
30 could contribute to between-study heterogeneity in results; however, the objective
31 assessment of outcomes in some studies is not likely to produce a spurious association.
32 Further, some of the evidence was provided by large studies that prospectively
33 ascertained outcome incidence after the measurement of blood Pb levels, did not indicate
34 selection bias, and considered potential confounding by SES and other environmental
35 exposures. These strengths reduce the likelihood of reverse causality and the influence of
November 2012 5-457 Draft - Do Not Cite or Quote
-------
other risk factors and increase confidence that the observed associations reflect a
relationship with Pb exposure.
Study Population Outcome
Jedrychowskietal. (2011) Children
Rabinowitzet al. (1990) Children Eczema
Blood Pb Level Mean
or Group (ug/dL)
Positive skin prick test prenatal: 1.16
concurrent: 2.02
>10vs. <10 —
i L i i /^nnr\ r*\--i i i~> • IncidentAsthma ,_ _
Josephetal. (2005) Children,Caucasian _ ....,.,_ >5vs. <5
Requiring Medical Care
Children, African American
Children, African American
Rabinowitzet al. (1990) Children
Pugh Smith and Nriagu
(2011)
Children
Asthma
Asthma
>5vs. <5
>10vs. <5
>10vs.
>10vs.
12345
Odds ratio (95% Cl)a
aFor analyses with blood Pb level as a continuous variable, odds ratios are standardized to a 1 ug/dL increase in blood Pb level.
Note: Results are presented first for allergy-related outcomes then for asthma. All results are from prospective analyses, except for
Pugh Smith and Nriagu (2011). Black diamond represents associations with concurrent blood Pb levels, green triangles represent
associations with prenatal (cord) blood Pb levels, and blue circles represent associations with blood Pb levels measured in
childhood up to 12 months prior to outcome assessment.
Figure 5-35 Associations of blood Pb levels with asthma and allergy in
children.
November 2012
5-458
Draft - Do Not Cite or Quote
-------
Table 5-33 Additional characteristics and quantitative results for studies
presented in Figure 5-35.
Study Population and
Study Methodological Details
Blood Pb Level Data (ug/dL)
Outcome
Odds Ratio or
Relative Risk
(95% Cl)
Jedrychowski 224 children followed prenatally to
et al. (2011) age 5 yr, Krakow, Poland
Prospective. Study of multiple
exposures and outcomes. Clinical
assessment of atopy. No information
on follow-up participation but no
selective attrition. Logistic regression
adjusted for sex, parity, maternal
age, education, and atopy, cord
blood cotinine, smoker in home
during follow-up. Also considered
potential confounding by
breastfeeding and allergen levels in
house dust.
Prenatal (cord): Geometric mean:
1.16(95% Cl: 1.12, 1.22)
Concurrent: Geometric means:
2.02 (95% Cl: 1.95,2.12)
Positive Skin
Prick Test
2.3(1.1,4.6)a
1.1 (0.7, 1.6)a
Rabinowitz et 159 children followed from birth to
al. (1990) unspecified age, Boston area, MA
Prospective. Low participation
among eligibles. No information on
differences with nonparticipants.
Logistic regression with parental
report of eczema. No consideration
of potential confounding factors.
Prenatal (cord) > 10 vs. <10
Eczema
1.0 (0.6, 1.6)b
Hon et al. 110 children with atopic dermatitis,
(2010: 2009)° mean (SD) age: 9.9 (4.6) yr, Hong
Kong, China,
Not clear whether subjects were free
of atopic dermatitis at time of blood
Pb measurement. Recruitment from
dermatology clinic. Clinical
assessment of atopic dermatitis. No
information on participation rate.
Examination of multiple metals. Lack
of rigorous statistical methods. No
consideration of potential
confounding factors.
Serum Pb mean (SD): 1.86 (0.83)
Atopic
dermatitis
severity
r = 0.329,
p O.001
Joseph et al. 4,634 children, ages 1 -3 yr followed
(2005) for 12 months, Southeastern Ml
Prospective. Large sample size.
Indirect assessment of asthma
diagnosis but ascertained from
managed care organization claims
database. Logistic regression
adjusted for sex, birth weight, and
average annual income available
only at census block level. Lack of
information on other environmental
exposures.
Caucasian a 5 vs. Caucasian <5
African American a 5 vs.
African American <5
African American > 10 vs.
African American <5
Measured up to 12 mo before
outcome
Incident
asthma
requiring
medical care
2.7(0.9,8.1)°
1.1 (0.8, 1.7)d
1.3(0.6, 2.6)d
Rabinowitz et 204 children followed from birth to
al. (1990) unspecified age, Boston area, MA
Same methodology as above.
Prenatal (cord blood)
> 10 vs. <10
Prevalent
asthma
1.3(0.8,2.0)"
November 2012
5-459
Draft - Do Not Cite or Quote
-------
Study Population and
Study Methodological Details
Blood Pb Level Data (ug/dL)
Outcome
Odds Ratio or
Relative Risk
(95% Cl)
Pugh Smith 356 children, ages 0-12 yr, Saginaw,
and Nriagu Ml
(2011) Cross-sectional. Recruitment from
blood Pb database. Moderate
participation rate. Parental report of
asthma diagnosis. Logistic
regression adjusted for age, sex,
family income, number of stories in
unit, cat in home, dog in home,
cockroach problem, number of
persons in home, smoker in home,
clutter, candles/incense, type of
cooking stove, main heating
source, months of residency, housing
tenure, type of air conditioning,
peeling paint, ceiling/wall damage,
age of housing, water
dampness/mold/mildew.
Highest blood Pb level at address
> 10 vs. <10
Levels ascertained from statewide
database, specific timing
unreported but varied among
subjects
Prevalent
asthma
diagnosed
within previous
12 months
7.5(1.3,42.9)"
aOdds ratio presented per 1 ug/dL increase in blood Pb level.
bOdds ratio in children with blood Pb level a 10 ug/dL with children with blood Pb level <10 ug/dL serving as the reference group.
°Results are not included in Figure 5-35 because only correlations are presented.
dRelative risk in each specified subgroup with children with blood Pb level <5 ug/dL serving as the reference group.
1
2
o
6
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
Key evidence for Pb-associated effects on allergy-related outcomes was provided by a
prospective study in 224 children in Poland at age 5 years that compared associations of
prenatal (cord and maternal) and concurrent blood Pb levels with incidence of allergic
sensitization, as ascertained by investigators (Jedrychowski et al., 2011). Subjects
assessed at five years did not differ from the full cohort, indicating lack of selective
attrition of subjects by blood Pb level or health status. The potential for selection bias
also is reduced because multiple exposures and outcomes were examined in this cohort.
A 1 (ig/dL increase in prenatal cord blood level was associated with greater risk of
positive skin prick test (SPT, rash/inflammatory reaction) to dust mite, dog, or cat
allergen with an RR of 2.3 (95% CI: 1.1, 4.6). Concurrent blood Pb level was more
weakly associated with risk of positive SPT (Figure 5-35 and Table 5-33). For prenatal
Pb biomarkers, similar effect estimates were obtained before and after adjustment for sex,
parity, maternal age, education, and atopy, and prenatal (cord blood cotinine) and
postnatal (smoker in the home) smoking exposure. Results were not altered by the
addition of house dust allergen levels. Cord and concurrent blood Pb levels were weakly
correlated (r = 0.29), providing support for an independent association for prenatal Pb
biomarkers. A relationship with Pb was substantiated by observations that indicators of
other exposures, including blood levels of Hg, poly cyclic aromatic hydrocarbon DNA
adducts, and residential levels of dust mite or pet allergen were associated with lower
risks of SPT than was blood Pb level. These associations were observed with relatively
low cord blood Pb levels (geometric mean: 1.16 (ig/dL [95% CI: 0.12, 1.22]). However,
cord blood Pb levels reflect the pregnancy blood Pb levels of mothers. Evidence indicates
November 2012
5-460
Draft - Do Not Cite or Quote
-------
1 increased mobilization of Pb from bone to blood in pregnant women (Sections 4.2.2.4
2 and 4.3.5.2). Thus, there is uncertainty regarding the Pb exposure scenarios that
3 contributed to associations between cord blood Pb level and allergic sensitization in
4 children examined in Jedrychowski et al. (2011).
5 Contrasting results were produced by prospective studies of eczema or atopic dermatitis,
6 which are reactions of the skin to sensitized allergens; however, neither study considered
7 potential confounding factors. Rabinowitz et al. (1990) found no elevated risk of parental
8 reported eczema in children (n = 159) in the Boston, MA area with cord blood Pb levels
9 > 10 (ig/dL. Hon et al. (2010; 2009) found a correlation between concurrent serum Pb
10 levels (mean: 1.86 (ig/dL) and clinical diagnosis of atopic dermatitis severity (e.g., skin
11 area affected, intensity of rash and inflammation, symptoms) in 110 children
12 approximately age 10 years in Hong Kong (Spearman r = 0.33, p O.005). The various
13 other metals were examined were negatively correlated with atopic dermatitis. Although
14 Hon et al. (2010; 2009) examined incidence of atopic dermatitis, subjects were selected
15 from patients referred to a dermatology clinic. The representativeness of the children to
16 the source population is uncertain, and allergies may already have developed by the time
17 serum Pb levels were measured.
18 Prospective and cross-sectional evidence indicated associations with blood Pb levels with
19 asthma in children. Among prospective studies, Joseph et al. (2005) accounted for
20 potential confounding factors. Asthma-free children, ages 1-3 years, (n = 4,634) all
21 members of the same managed care organization in southeastern Michigan were selected
22 based on availability of blood Pb level data in the database then tracked for the following
23 12 months for incidence of asthma. Incident asthma requiring a doctor visit or medication
24 was defined from the medical claims database as four or more asthma medication
25 dispensing events and one or more asthma emergency department visit, hospitalizations,
26 or outpatient visits with at least two asthma medication dispensing events in the previous
27 12 months. While this definition is not a direct diagnosis, it is used to define persistent
28 asthma by the Healthcare Effectiveness Data and Information Set, which most U.S.
29 health plans use to measure health care performance. The records-based analysis
30 precluded bias due to selective participation of subjects by blood Pb level and health
31 status. However, because a blood Pb measurement was required, it is uncertain whether
32 the study population is representative of the managed care organization population. In
33 analyses that adjusted for average annual income at the census block level, birth weight,
34 and sex, an elevated risk of incident asthma requiring a doctor visit or medication was
35 associated with blood Pb levels > 5 (ig/dL in Caucasian children (RR: 2.7 [95% CI: 0.9,
36 8.1] compared with Caucasian children with blood Pb levels <5 (ig/dL) (Figure 5-35 and
37 Table 5-33). In analyses restricted to African Americans, children with blood Pb levels
38 > 10 (ig/dL had an elevated risk of asthma requiring medical care (RR: 1.3 [95% CI: 0.6,
November 2012 5-461 Draft - Do Not Cite or Quote
-------
1 2.6] compared with children with blood Pb level <5 (ig/dL). There were small numbers of
2 children with asthma requiring medical care in the higher blood Pb level categories,
3 which could have accounted for the wide 95% CIs (5 Caucasian children with blood Pb
4 > 5 (ig/dL and 9 African American children with blood Pb level > 10 (ig/dL). In analyses
5 that used Caucasian children with blood Pb level <5 (ig/dL as the reference group, blood
6 Pb level was associated with increased risk of asthma requiring medical care among
7 African American children in all blood Pb level categories, which indicated a stronger
8 association with race. Nonetheless, results within race groups pointed to an association
9 with blood Pb level.
10 Similarly, the prospective study of 204 children in the Boston, MA area found a
11 Pb-associated increased risk of parental-reported asthma (age of assessment not
12 reported), specifically in children with cord blood Pb levels >10 (ig/dL relative to cord
13 blood Pb levels < 10 (ig/dL (Rabinowitz et al.. 1990) (Figure 5-35 and Table 5-33).
14 However, potential confounding factors were not examined.
15 Supporting the prospective evidence, a cross-sectional study conducted in Saginaw, MI
16 found a higher prevalence of parental report of doctor-diagnosed asthma in children (ages
17 < 12 years) with blood Pb levels > 10 (ig/dL (Pugh Smith and Nriagu. 2011V Similar to
18 Joseph et al. (2005). the study population was predominately African American (78% of
19 356). Children were randomly selected from a statewide database of initial blood Pb
20 measurements collected at unspecified ages, and a positive bias is possible if parents of
21 children with higher blood Pb levels and asthma were more likely to participate or recall
22 an asthma diagnosis. Data were collected on asthma diagnosis in the previous 12 months;
23 thus, for some subjects, blood Pb measurement likely preceded asthma diagnosis. A
24 strength of this study was the adjustment for a large number of potential confounding
25 factors such as age, sex, household pets, housing characteristics, and household smoking
26 family income. Compared with children with initial blood Pb levels <10 (ig/dL, children
27 with initial blood Pb levels > 10 (ig/dL had a higher odds of having a doctor diagnosis of
28 asthma within the past 12 months (OR: 7.5 [95% CI: 1.3, 42.9]). The results were
29 imprecise, and while this study had more children with blood Pb levels > 10 (ig/dL
30 (18.6%) than did Joseph et al. (2005). the analyses considered a much large number of
31 covariates.
32 As was discussed in Section 5.6.3. one may speculate that cross-sectional associations
33 could be attributed to reverse causality. Individuals with asthma and animal models of
34 asthma have been shown to have epithelial cell damage and exudation of cells and fluids
35 into airways, which are indicators of increased lung permeability. With increased lung
36 permeability, one may speculate the potential for greater uptake of Pb from airways into
37 blood. Most evidence does not demonstrate greater uptake of particles into the blood in
November 2012 5-462 Draft - Do Not Cite or Quote
-------
1 subjects with asthma than in healthy controls (Del Donno et al.. 1997; Rees et al.. 1985;
2 O'Byrne etal.. 1984; Elwoodetal.. 1983). Histamine was found to increase particle
3 uptake transiently. In Rees et al. (1985). histamine equally increased particle uptake in
4 subjects with and without asthma whose mean lung function decreased by 33% and 55%,
5 respectively. This evidence combined with that from prospective epidemiologic studies
6 and that characterizing modes of action (i.e., increases in IgE, Th2 cytokines, and
7 inflammation) increases confidence that the associations observed between blood Pb
8 level and asthma and allergy in children are not due to reverse causality.
9 Among the studies in children that found associations of blood Pb level with asthma and
10 allergy, several adjusted for potential confounding by indicators of SES and other
11 environmental exposures. Joseph et al. (2005) adjusted for census block annual income,
12 which may measure individual-level income with error. However, Pugh Smith and
13 Nriagu (2011). which examined a primarily low SES population of children in Michigan,
14 adjusted for family annual income. In this study, a Pb-associated higher asthma
15 prevalence was found with adjustment for various additional factors associated with SES
16 and allergen exposure, including multiple indices of housing condition and presence of
17 pets and cockroaches in the home. Jedrychowski et al. (2011) adjusted for maternal
18 education, and found similar magnitudes of association between cord blood Pb level and
19 positive SPT as those in the unadjusted analysis. Further, residential levels of dust mite or
20 pet allergen were associated with lower risks of SPT than was blood Pb level. Blood Pb
21 level also was associated with asthma or allergy after adjusting for concurrent exposure
22 to smoking in the home (Jedrychowski et al.. 2011; Pugh Smith and Nriagu. 2011). with
23 Jedrychowski et al. (2011) additionally adjusting for prenatal smoke exposure assessed as
24 cord blood cotinine levels. The studies varied in the specific confounding factors
25 considered, the method of measurement, and the method of control. Some studies
26 examined several potential confounding factors (Jedrychowski et al.. 2011; Pugh Smith
27 and Nriagu. 2011). which increases confidence that the observed associations with
28 asthma and allergy reflect a relationship with Pb. However, in the small evidence base,
29 uncertainty remains regarding residual confounding, particularly by SES. While there is
30 no single complete measure of SES, these studies adjusted for different indicators of SES
31 that may vary in the adequacy of control for confounding. Residual confounding also is
32 possible by factors not examined.
33 Cross-sectional evidence did not strongly indicate associations of biomarkers of Pb
34 exposure with asthma or allergy in nonoccupationally-exposed adults (Mendy et al..
35 2012; Pizent et al.. 2008). However, study limitations make the evidence less informative
36 for drawing conclusions about the effects of Pb on asthma compared with evidence in
37 children. Higher Pb level in a spot urine sample was not associated with an increase in
38 asthma prevalence (OR: 0.72 [95% CI: 0.46, 1.12] per 1 (ig Pb/g creatinine increase in
November 2012 5-463 Draft - Do Not Cite or Quote
-------
1 urine) in the large U.S. NHANES 2007-2008 analysis of 1,857 adults ages 20 years and
2 older (geometric mean urinary Pb level: 0.59 (ig/g creatinine (Mendv et al.. 2012).
3 Associations were not found with other respiratory conditions such as emphysema or
4 chronic bronchitis either. This study examined several potential confounding factors,
5 adjusting for sex, race/ethnicity, education, income to poverty ratio, number of alcoholic
6 drinks/day, and current smoking status. However, spot urinary Pb level has an uncertain
7 relationship with long-term Pb exposure (Section 4.3.3). In a study of adult (19-67 years)
8 office workers in Zagreb, Croatia, Pizent et al. (2008) found a lower concurrent blood
9 Pb-associated odds of positive SPT to common inhaled allergens among 50 men (median
10 blood Pb level: 3.2 (ig/dL) and lack of statistically significant association among 166
11 women (median blood Pb level: 2.2 (ig/dL), adjusting for age, smoking (current and
12 history), and number of alcoholic drinks/day. The findings in women appeared to be
13 discordant because there was an association between concurrent blood Pb level and
14 serum IgE, which commonly mediates the acute inflammatory response to allergens.
15 However, the interpretation of the findings is difficult because only statistically
16 significant effect estimates were reported; thus it is not known whether odds ratios were
17 in the same direction for SPT and IgE in women. Bener et al. (200la) found higher
18 prevalence of asthma and allergy-related conditions in 110 Pb industrial workers (mean
19 age: 35.5 years) than in 110 age-matched controls. However, the implications are limited
20 because blood Pb levels in both Pb-exposed workers and controls (geometric means: 77.5
21 and 19.8 (ig/dL, respectively) were higher than those in the current U.S. adult general
22 population, and potential confounding factors, including other occupational exposures,
23 were not evaluated.
24 In summary, evidence supports associations of higher Pb biomarker levels with asthma
25 and allergy (Jedrychowski et al.. 2011; Pugh Smith and Nriagu. 2011; Joseph et al.. 2005)
26 in children. Because of study limitations, the evidence in adults does not largely inform
27 the conclusion (Mendv etal., 2012; Pizent et al., 2008; Bener etal., 200la). In children,
28 evidence was limited to a few populations. Because of the heterogeneity in the relatively
29 small body of evidence, it was difficult to identify whether the strength of association
30 with asthma and allergy differed by age of children, lifestage of blood Pb measurement
31 (prenatal, sometime in childhood prior to outcome assessment, concurrent), or blood Pb
32 level. The prospective analysis in Jedrychowski et al. (2011) and Joseph et al. (2005)
33 increase confidence that the observed associations are not due to reverse causality. Lack
34 of reverse causality also is indicated by observations that particle uptake generally does
35 not differ between subjects with and without asthma and between animals sensitized with
36 allergens and unsensitized controls. In these studies, the lack of selective participation of
37 subjects and objective assessment of outcomes indicates lack biased reporting of asthma
38 and allergy in children with higher blood Pb levels. In some studies, the method of
39 recruitment of subjects from blood Pb surveillance databases may limit generalizability
November 2012 5-464 Draft - Do Not Cite or Quote
-------
1 of findings. The adjustment for maternal education and exposure to smoking or allergens
2 in Jedrychowski et al. (2011) and family income, smoking, housing conditions, pets, or
3 pests in Pugh Smith and Nriagu (2011) increase confidence that the observed associations
4 in these studies are not due to confounding by SES, smoking, or allergen exposure.
5 Further, biological plausibility is well supported by evidence describing modes of action
6 for asthma and allergy, including Pb-associated increases in IgE (Section 5.6.3).
7 Th2 cytokines (Section 5.6.6.1). and inflammation (Section 5.6.4). However, because
8 there are few studies, uncertainty remains regarding residual confounding, particularly by
9 SES, which was examined with different indicators across studies.
5.6.5.3 Other Respiratory Effects
10 Other respiratory effects associated with blood or air Pb levels were not well
11 characterized in the 2006 Pb AQCD (U.S. EPA. 2006b) but have been examined recently
12 in a small number of studies. As with asthma, associations between blood Pb level and
13 respiratory effects in adults is inconsistent. Studies were cross-sectional, included
14 similarly aged subjects, and considered similar confounding factors. Increased bronchial
15 responsiveness (BR) is a characteristic feature of asthma and other respiratory diseases
16 and can result from the activation of innate immune responses and increased airway
17 inflammation. In the larger study of 523 adults (ages 19-58 years) in Seoul, Korea, Min et
18 al. (2008a) found an association between concurrent blood Pb level and BR. A 1 (ig/dL
19 higher concurrent blood Pb level was associated with a higher BR index (log [% decline
20 in forced expiratory volume in 1 second (FEVi)/log of final methacholine concentration
21 in mg/dL]) of 0.018 (95% CI: 0.004, 0.03), with adjustment for age, sex, height, smoking
22 status, baseline FEVi, and presence of asthma (Min et al.. 2008a). The concurrent blood
23 Pb levels in these adults were low (mean [SD]: 2.90 [1.59] (ig/dL); however, it is
24 uncertain what timing, level, frequency, and duration of Pb exposures contributed to the
25 observed association. In contrast to Min et al. (2008a). Pizent et al. (2008) found that
26 higher concurrent blood Pb level was associated with lower BR in 47 men (2.4%
27 decrease [95% CI: -4.2, -0.52%] in percent change FEVi post-histamine challenge per
28 1 (ig/dL increase in blood Pb level adjusted for age and serum Se). Smoking intensity and
29 alcohol consumption were excluded as covariates by stepwise regression. Similarly,
30 among these men, higher blood Pb level was associated with lower odds of positive SPT.
31 Pb-associated respiratory effects were not clearly indicated in adults with occupational Pb
32 exposures either. However, the lack of direct analysis of blood Pb levels and
33 consideration for potential confounding limit the utility of this evidence in drawing
34 conclusion about the respiratory effects of Pb related to airway responses. In bus drivers
35 (mean age: 46 years) in Hong Kong (Jones et al.. 2008; Jones et al.. 2006). 129 drivers of
November 2012 5-465 Draft - Do Not Cite or Quote
-------
1 non-air conditioned buses had lower exposures to PM10, lower blood Pb levels (mean
2 3.7 (ig/dL versus 5.0 (ig/dL in air conditioned buses) but lower indices of lung function
3 than did 358 drivers of air conditioned buses (Jones et al., 2006). The authors attributed
4 the slightly higher blood Pb levels of air conditioned bus drivers to the poor efficiency in
5 the filters resulting in higher PM10 levels on those buses. Blood Pb levels and various
6 lung function parameters were similar between 33 roadside vendors and 31 adjacent
7 shopkeepers (mean ages: 45.1 and 42.8 years, respectively and mean blood Pb levels:
8 5.61 and 5.14 (ig/dL) (Jones et al.. 2008). Pb industrial workers (n = 100, mean age: 34.6
9 years) in the United Arab Emirates had higher prevalence of respiratory symptoms such
10 as cough, phlegm, shortness of breath, and wheeze than did 100 age- and sex-matched
11 unexposed controls (Bener etal. 200la). Blood Pb levels in both the Pb industrial
12 workers and the control group (geometric means: 77.5 and 19.8 (ig/dL, respectively) were
13 higher than those in most of the current U.S. adult general population.
14 An effect of Pb specifically on the lung was demonstrated in a recent study of Wistar rats
15 with low blood Pb levels (2.1 (ig/dL) but produced by Pb acetate given by injection
16 (25 mg/kg, 3 consecutive days) (Kaczynska et al.. 2011). The lungs of Pb-treated rats
17 exhibited pulmonary fibrosis, epithelial cell damage, an increase in mast cells, an
18 increased recruitment of monocytes and thrombocytes into capillaries, and increased
19 macrophage accumulation in the alveolar space. While these pulmonary changes have
20 been linked with functional pulmonary decrements and inflammation in other studies
21 (unrelated to Pb exposure), the implications are uncertain because the results were
22 obtained with a route of Pb exposure less relevant to those in humans.
Air-Pb Studies
23 The 2006 Pb AQCD (U.S. EPA. 2006b) did not review studies that represent Pb exposure
24 by Pb measured in PMi0 and PM2 5 air samples. However, recent studies have examined
25 the respiratory effects of PM-Pb by analyzing the Pb component individually or as part of
26 a group of correlated components using source apportionment or principal component
27 analysis. Daily ambient air Pb-PM concentrations were associated with daily respiratory
28 morbidity in children (Gent et al.. 2009; Hong et al.. 2007b). Gent et al. (2009) found that
29 increases in lag 0 and 0-2 average Pb-PM2 5 were associated with increases in respiratory
30 symptoms and asthma medication use in 149 children with asthma in Southern New
31 England (ages 4-12 years), adjusting for season, day of the week, and date. Hong et al.
32 (2007b) found that an increase in lag 1 Pb-PMi0 was associated with a decrease in lung
33 function in 43 mostly healthy children in Korea, adjusting for age, sex, height, weight,
34 household smoking, and weather. In support of these results in children, toxicological
35 studies found Pb-containing CAPs to induce pulmonary inflammation. Uzu et al. (2011)
36 found that Pb-rich PM from a Pb recycling plant increased the release of the cytokine
November 2012 5-466 Draft - Do Not Cite or Quote
-------
1 granulocyte-macrophage colony-stimulating factor from human epithelial cells.
2 Pulmonary inflammation was found in animals exposed to CAPs in which Pb was one of
3 numerous components (Wei et al., 2011; Duvall et al., 2008; Godleski et al., 2002;
4 Saldiva et al.. 2002).
5 As with blood Pb, daily ambient air Pb-PM2 5 concentrations were not consistently
6 associated with daily respiratory effects in older adults with adjustment for weather and
7 temporal trends. Among adults ages 65 years and older in 6 California counties, a 4
8 ng/m3 increase in lag 3 Pb-PM25 was associated with an relative risk of respiratory
9 mortality of 1.01 (95% CI: 0.99, 1.03) in the all-year analysis and in a summer-only
10 analysis (RR not reported) (Ostro et al.. 2007). However, among adults ages 65 years and
11 older in 106 U.S. counties, Bell et al. (2009) found that an increase in lag 0 Pb-PM2s was
12 associated with a decrease in respiratory hospital admissions.
13 Despite evidence that indicates a relationship between respiratory effects in children and
14 short-term (over a few days) changes in ambient air Pb-PM concentrations, uncertainties
15 limit the utility of these findings in evaluating Pb-associated respiratory effects. Few data
16 on size distribution of Pb-PM are available, so it is difficult to assess the
17 representativeness of these concentrations to population exposure (Section 3.5.3).
18 Moreover, few data are available on the relationship between blood Pb and air Pb for the
19 varying Pb-PM size distributions (see Section 4.5.1). In several air-Pb studies, other PM
20 components such as elemental carbon (EC), copper (Cu), and zinc (Zn) also were
21 associated with respiratory effects. In the absence of data on correlations among PM
22 components, measurements on co-occurring ambient pollutants, or results adjusted for
23 copollutants, it is difficult to exclude confounding by ambient air exposures to other PM
24 components or ambient pollutants. In several studies that analyzed PM component
25 mixtures, of which Pb-PM comprised one component, it is not possible to attribute the
26 observed associations or lack of associations specifically to Pb (Sarnat et al.. 2008;
27 Andersen et al.. 2007; Veranth et al.. 2006; Macieiczyk and Chen. 2005).
28 In summary, while air Pb-PM has been associated with respiratory effects in children,
29 main limitations of this recent evidence include the confounding by other PM
30 components and the uncertain representativeness of Pb-PM to population exposures. In
31 adults, neither blood Pb nor Pb-PM was consistently associated with respiratory effects.
32 Blood Pb studies of nonoccupationally-exposed adults were similar in cross-sectional
33 design, age of subjects, potential confounding factors examined, and respiratory
34 endpoints which exhibit short-term changes. Studies of Pb-exposed workers were
35 similarly limited by lack of rigorous statistical analysis with blood Pb levels and lack of
36 consideration for potential confounding factors, including other occupational exposures.
November 2012 5-467 Draft - Do Not Cite or Quote
-------
5.6.5.4 Autoimmunity
1 Autoimmunity is an immune response against self (e.g., generation of antibodies against
2 self antigens) and is linked with diseases such as lupus and rheumatoid arthritis. Evidence
3 for the effects of Pb on increasing the risk of autoimmunity is provided primarily by a
4 small number of toxicological studies reviewed in the 2006 Pb AQCD in which pre- and
5 post-natal Pb acetate exposure of animals, several by injection, was associated with the
6 generation of autoantibodies (Hudson et al., 2003; Bunn et al., 2000; El-Fawal et al.,
7 1999; Waterman et al.. 1994). El-Fawal et al. (1999) found elevated auto-antibodies in
8 F344 rats with blood Pb levels 11-50 (ig/dL. Evidence was mixed in indicating a shift
9 toward Th2 or Thl responses as the underlying mechanism. While recent studies did not
10 examine Pb-induced production of auto-antibodies, some provided indirect evidence by
11 indicating that Pb had the potential to induce formation of neo-antigens which in turn
12 could induce the formation of auto-antibodies. For example, Kasten-Jolly et al. (2010)
13 found that developmental Pb acetate exposure of BALB/c mice (100 (iM in drinking
14 water, GD8-PND21) upregulated genes for digestive and catabolizing enzymes, which
15 could lead to the generation of self-peptides, which combined with other Pb-induced
16 immune effects, had the potential to induce the generation of auto-antibodies. The
17 potential for auto-antibody generation also was indicated by the activation of neo-
18 antigen-specific T cells in adult BALB/c mice injected once with 25-50 (ig Pb chloride
19 (Carey et al., 2006). Evidence of Pb-associated autoimmune responses in humans is
20 limited to findings of higher levels of IgM and IgG auto-antibodies to neural proteins in
21 male battery-plant workers (n = 20, 56) with blood Pb level range 10-40 (ig/dL compared
22 with controls (n = 7, 15, blood Pb levels not reported) (El-Fawal et al.. 1999). Pb workers
23 and controls were matched by demographic and SES characteristics, but potential
24 confounding by other workplace exposures was not examined. Similar to findings in
25 Pb-exposed workers, modified neural proteins were found in CBA/J rats injected with
26 native protein altered by Pb acetate in vitro (Waterman et al.. 1994).
5.6.5.5 Tumors
27 Toxicological evidence indicates that high concentration Pb exposures directly promote
28 tumor formation or induce mutagenesis and genotoxicity (Section 5.10). and a study
29 provided evidence for involvement of the immune system. Kerkvliet and Baecher-
30 Steppan (1982) found that postnatal exposure of 6-8 week old male C57BL/6 mice to 130
31 and 1,300 ppm Pb acetate in drinking water for 10-12 weeks transiently enhanced
32 moloney sarcoma virus-induced tumor growth compared with control animals but did not
33 prevent subsequent tumor regression. The Pb-induced tumor growth was accompanied by
34 impaired macrophage phagocytosis (indicating suppressed Thl responses) but not
November 2012 5-468 Draft - Do Not Cite or Quote
-------
1 cytotoxicity. Cancer promotion is a relatively common outcome in chemical-induced
2 immunotoxicology, particularly when early life exposures are involved (Dietert. 2011).
5.6.6 Modes of Action for Pb Immune Effects
5.6.6.1 Cytokine Production
3 As referenced in preceding sections, cytokines are signaling molecules that affect
4 immune cell function. For example, IL-4 induces B cells into IgE-producing cells, and
5 IFN-y induces macrophage recruitment and antigen presenting activity. The
6 2006 Pb AQCD (U.S. EPA. 2006b) presented a large body of evidence that clearly
7 demonstrated that pre- and postnatal Pb exposure of animals such as rodents and chickens
8 suppressed the production of Thl cytokine IFN-y and/or increased production of Th2
9 cytokines such as IL-4 [Table 5-7 of the 2006 Pb AQCD (U.S. EPA. 2006g)1. The
10 combined evidence for Pb-induced cytokine changes in multiple cell types, including T
11 cells and macrophages, indicates a shift of acquired immunity responses away from Thl
12 responses and toward Th2 responses. In turn, the Thl to Th2 shift provides mode of
13 action support for downstream effects (Figure 5-34) such as inflammation, ROS
14 production, impaired macrophage function, decreased host resistance observed primarily
15 in toxicological studies, increased IgE production observed in both epidemiologic and
16 toxicological studies, and asthma and allergy observed in epidemiologic studies. Previous
17 toxicological studies found Pb to affect cytokine production via action on T cells and
18 macrophages, and a recent study provided new evidence that Th2 skewing may be
19 mediated via effects on dendritic cells.
20 Many studies found a shift to Th2 cytokine production in animals with long-term
21 (>4 weeks) dietary Pb exposure, and in some studies, the effects of prenatal exposure on
22 cytokine production persisted to the adult lifestage (Chen et al.. 2004; Miller et al.. 1998).
23 In the few studies that measured blood Pb levels shortly after cessation of Pb exposure
24 (gestational plus postnatal or postnatal only), higher IL-4 and/or lower IFN- y were found
25 in rodents with relevant blood Pb levels, means 6.75 and 17 (ig/dL (Chen et al.. 2004;
26 Dyatlov and Lawrence. 2002). Some studies found an increase in IL-4 or decrease in
27 IFN- y concomitantly with an increase in IgE (Heo et al.. 1997; 1996) or decrease in host
28 resistance (Fernandez-Cabezudo et al.. 2007) further supporting changes in cytokine
29 production as a mode of action for Pb-induced effects on downstream immune endpoints.
30 A recent study found a shift to Th2 cytokine production in mice over wide range of Pb
31 exposures, and provided evidence of effects at lower Pb exposures and a concentration-
32 dependent relationship. In this study, lifetime (gestation through adulthood) exposure of
November 2012 5-469 Draft - Do Not Cite or Quote
-------
1 Swiss mice to 0.06-400 ppm dietary Pb acetate produced blood Pb levels (upon
2 termination of exposure) of 1.23 to 61.48 (ig/dL (lavicoli et al.. 2006b). For IL-2 and
3 IL-4, nonlinear concentration-response relationships were found, with the largest
4 decrease and increase, respectively, found between animals with blood Pb levels of
5 0.8 (ig/dL (0.02 ppm Pb acetate, controls) and 1.23 (ig/dL. A linear concentration-
6 dependent decrease in IFN-y was observed in animals with blood Pb levels of
7 0.8-61.48 (ig/dL. Although examined in few in vivo studies, increases in IL-6 and IL-10,
8 also Th2 cytokines, were reported in juvenile and adult rodents exposed to Pb
9 gestationally or postnatally in drinking water [Table 5-7 of the 2006 Pb AQCD (U.S.
10 EPA. 2006g)1.
11 In vitro studies also reported a Pb-induced shift to production of Th2 cytokines. In
12 concordance with other indicators of Th2 skewing (i.e., suppressed DTH) in BALB/c
13 mice treated with Pb-exposed dendritic cells, Gao et al. (2007) observed that 10-day
14 25 (iM Pb chloride exposure lowered the ratio of IL-12:IL-10 production by dendritic
15 cells in vitro. Pb did not affect dendritic cell production of IL-6, IL-10, or TNF-a;
16 however, in co-cultures of Pb-treated dendritic cells and T cells, most results indicated
17 that dendritic cells stimulated T cells to produce Th2 cytokines. For example, although T
18 cell production of the Thl cytokine IL-2 increased, production of Th2 cytokines, IL-6
19 and IL-10 increased. Further, Pb-treated dendritic cells increased IL-4 production in
20 OVA-specific T cells, indicating that Pb affected the antigen presenting cell function of
21 dendritic cells. In another in vitro study, 24-hour Pb acetate exposures of 0.15 (ig/dL and
22 higher suppressed expression of Thl cytokines, IFN-y, IL-1(3, and TNF-a, and increased
23 secretion of Th2 cytokines, IL-5, IL-6, and IL-10 in cultures of human PMNs activated
24 with Salmonella enteritidis or with monoclonal antibodies of CD3, CD28, and CD40,
25 (Hemdan et al.. 2005).
26 Several toxicological studies found Pb-induced increases in the cytokine TNF-a, in some
27 cases, specifically from macrophages (Khan et al.. 2011; Cheng et al.. 2006; Flohe et al..
28 2002; Zelikoff et al.. 1993). This provides mode of action support for toxicological
29 evidence indicating Pb-induced decreases in resistance to bacterial infection since TNF-a
30 is produced primarily by activated macrophages, is increased in response to infection, and
31 induces inflammation. Among the in vivo studies, increases in TNF-a were found with
32 prenatal dietary Pb acetate exposure (250 ppm) of F344 rats (Chen et al.. 1999; Miller et
33 al.. 1998). postnatal Pb oxide air exposure (31 (ig/m3, 3 hours/day, 4 days) of rabbits
34 (Zelikoff et al.. 1993). and postnatal i.p. Pb acetate treatment (5.0 mg) of Swiss mice
35 (Dentener et al.. 1989). The effects of prenatal dietary Pb exposure were found to persist
36 to adulthood. In animals, the Pb-induced increases in TNF-a were accompanied by
37 functional changes in host responses such as decreased macrophage phagocytosis
38 (Zelikoff etal.. 1993). suppressed DTH (Miller etal.. 1998). and increased mortality to
November 2012 5-470 Draft - Do Not Cite or Quote
-------
1 E.coli endotoxin (Dentener et al.. 1989). Blood Pb levels of animals were infrequently
2 reported; however, Chen et al. (1999) found increased TNF-a in rats with embryonic
3 blood Pb levels of 149 (ig/dL. In addition to finding Pb-induced increases in TNF-a
4 (Khan etal.. 2011: Gao et al.. 2007: Cheng et al.. 2006: Flohe et al.. 2002: Krocova et al..
5 2000: Guo etal.. 1996a). some in vitro studies provided mechanistic explanation by
6 finding that Pb acetate or chloride (10-50 (iM, 1.5 hours-10 days) induced
7 phosphorylation of mitogen-activated protein kinase (MAPK) signaling molecules (Khan
8 etal.. 2011: Gao et al.. 2007: Cheng et al.. 2006). Further, Cheng et al. (2006) found that
9 blocking protein kinase C or MAPK reduced TNF-a production by macrophages in vitro,
10 which in turn, protected against Pb acetate + LPS-induced liver injury in A/J mice.
11 The few available epidemiologic studies that examined cytokines found higher
12 concurrent blood Pb levels in children and adults to be associated with higher Th2
13 cytokine and lower Thl cytokine levels in serum. The epidemiologic evidence overall
14 was based on cross-sectional analyses, which precludes identifying the temporal
15 sequence between Pb exposure and cytokine changes. Other limitations include the lack
16 of rigorous statistical analysis and limited consideration of potential confounding.
17 Because of these limitations, the epidemiologic evidence is not a primary consideration in
18 drawing conclusions about Pb-associated cytokine changes. However, it does not mitigate
19 the consistent toxicological evidence. Among children ages 9 months to 6 years in
20 Missouri recruited from a public assistance or Pb poisoning prevention program, Lutz et
21 al. (1999) found that 8 children with concurrent blood Pb levels 15-19 (ig/dL had 4-5 fold
22 higher serum levels of IL-4 (p = 0.08, Kruskal Wallis) and 3-fold higher IgE
23 (Section 5.6.3) than did 90 children with lower blood Pb levels. IL-4 levels in 9 children
24 with blood Pb levels 20-44 (ig/dL were lower than those in 90 children with blood Pb
25 levels <15 (ig/dL. The elevated IL-4 and IgE in children with blood Pb levels
26 15-19 (ig/dL were consistent with the action of IL-4 to activate B cells to induce class
27 switching to IgE. In another study of 214 children in grades 5 and 6 in Taiwan,
28 investigators compared cytokine levels not by blood Pb level groups but by potential for
29 Pb exposures due to age of home and location of residence (Hsiao etal.. 2011). Elevated
30 concurrent blood Pb levels were found only among 64 children living near an oil refinery,
31 in particular, among 34 children with known respiratory allergies (mean: 8.8 (ig/dL
32 versus 3.2-3.8 (ig/dL in urban and rural groups). Children with allergies near the oil
33 refinery also had the lowest serum levels of IFN-y (45-fold) and highest levels of IL-4 (6-
34 fold) (lower p <0.05 for comparisons with any subgroup). While the results suggested
35 that residence near the oil refinery contributed to differences in cytokine levels between
36 healthy and allergic children, they do not specify a contribution of Pb, other exposures or
37 co-occurring factors, or a combination of factors.
November 2012 5-471 Draft - Do Not Cite or Quote
-------
1 Evidence of association between blood Pb levels and cytokine levels in is unclear.
2 However, a larger study that considered potential confounding found an association.
3 Sufficient information was not provided to assess potential selection bias. Among 300
4 (93% male, mean age 24 years) healthy university students in Incheon, Korea, higher
5 concurrent blood Pb level was associated with higher serum levels of TNF-a and IL-6,
6 adjusting for age, BMI, and smoking status (Kim et al.. 2007). Associations were larger
7 in magnitude among the 147 males in the upper two quartiles of blood Pb levels,
8 2.51-10.47 (ig/dL. Associations with these blood Pb levels may reflect contributions of
9 higher past Pb exposures. A 1 (ig/dL higher blood Pb level was associated with a 0.75
10 (95% CI: 0.14, 1.36) pg/mL higher TNF-a and a 0.18 (95% CI: -0.02, 0.38) pg/mL higher
11 IL-6. The association between levels of blood Pb and plasma TNF-a was greater among
12 men who were GSTM1 null (n = 77) than men who were GSTM1 positive and men who
13 had the TNF-a GG genotype (n = 131) than men who had the GA or AA genotype. For
14 the association between blood Pb level and plasma IL-6, the effect estimate was slightly
15 elevated in TNF-a GG genotype but similar between GSTM1 genotypes. In this study,
16 there were multiple comparisons, but a consistent pattern of association was observed
17 across the immune endpoints examined. Subgroup analyses had fairly large sample sizes,
18 and some results had biological plausibility, but they are prone to higher probability of
19 findings by chance. Pb has been shown to increase ROS (Section 5.2.4). and cytokine
20 expression has been shown to be modulated by ROS-sensitive transcription factors. Thus,
21 it is biologically plausible that the null variant of GSTM1, which is associated with
22 reduced elimination of ROS, may be associated with increased cytokine levels. The
23 results for the TNF-a variant are difficult to interpret. The GG genotype is associated
24 with lower expression of TNF-a, but the literature is mixed with respect to which variant
25 increases risk of inflammation-related conditions.
26 A much smaller study of adults that did not consider potential confounding, did not report
27 quantitative results but only reported lack of statistically significant correlations between
28 concurrent blood Pb level and serum Th2 and Thl cytokine levels in men (n = 17 with
29 and 17 without allergy, ages 19-52 years, median blood Pb levels: -11 (ig/dL) (Boscolo
30 et al.. 1999) and women in Italy (n = 23 with and 25 without allergy, ages 19-49 years,
31 median blood Pb levels: 6.4 and 5.5 (ig/dL, respectively) (Boscolo et al., 2000).
32 Results from studies of occupationally-exposed adults also suggested that Pb exposure
33 may be associated with decreases in Thl cytokines and increases in Th2 cytokines (Di
34 Lorenzo et al.. 2007; Valentino et al.. 2007; Yiicesoy et al.. 1997a). Valentino et al.
35 (2007) had the most rigorous statistical methods comprising regression analyses with
36 adjustment for age, BMI, smoking status, and alcohol consumption status but not other
37 occupational exposures. Regression coefficients describing the concentration-response
38 functions were not reported; however, 44 male foundry workers in Italy (mean blood Pb
November 2012 5-472 Draft - Do Not Cite or Quote
-------
1 levels: 21.7 (ig/dL) and 14 pottery workers (mean blood Pb level: 9.7 (ig/dL, ages of all
2 workers 30-61 years) had higher plasma IL-10 (ANOVA, p <0.05) than did the 59
3 unexposed controls (mean blood Pb level: 3.9 (ig/dL, ages 25-61 years). Levels of Th2
4 cytokines IL-2, IL-6, and IL-10 also increased from the lowest to highest blood Pb group
5 (ANOVA, p >0.05). In contrast with most other studies, both exposed worker groups had
6 lower IL-4 levels compared with controls (ANOVA, p >0.05). In Yucesoy et al. (1997a).
7 serum levels of the Thl cytokines, IL-1(3 and IFN-y, were lower in 20 Pb-exposed
8 workers (mean blood Pb level: 59.4 (ig/dL, ages 19-49 years) than in the 12 age-matched
9 controls in Turkey (mean blood Pb level: 4.8 (ig/dL). Some (Di Lorenzo et al.. 2007;
10 Valentino et al.. 2007) but not all (Yucesoy et al.. 1997a) studies found higher serum
11 TNF-a in Pb-exposed workers. DiLorenzo et al. (2007) found a monotonic increase from
12 the 28 unexposed (blood Pb levels not reported, mean age: 48.2 years) to 17 intermediate
13 worker (9.1-29.4 (ig/dL) and 19 high worker (29.4-81.1 (ig/dL) blood Pb level groups
14 (mean age of workers: 45.3 years) in Italy. Results also indicated a potential interaction
15 between blood Pb level and smoking. Among current smokers (n = 9 to 20), a 12- to
16 16-fold difference in TNF-a levels was observed among blood Pb groups. Among
17 nonsmokers (n = 2 to 8), the differences were less than two fold.
18 In summary, the few epidemiologic studies indicate associations of higher concurrent
19 blood Pb level with higher levels of IL-4 and/or lower levels of IFN-y in children (Hsiao
20 et al.. 2011; Lutzetal.. 1999) and occupationally-exposed adults (Di Lorenzo et al..
21 2007; Valentino et al.. 2007; Yucesoy et al.. 1997a). Because quantitative results were
22 not reported in each study of nonoccupationally-exposed adults, implications of findings
23 are difficult to assess. Limitations of the epidemiologic evidence overall include the
24 cross-sectional design of studies and lack of rigorous statistical analysis that considered
25 potential confounding factors. Sufficient data were not reported to assess potential
26 selection bias. Because of the many limitations, the epidemiologic evidence alone is not
27 used to draw conclusions about Pb-associated cytokine changes. However, they are
28 useful in indicating the relevance of toxicological evidence to humans. Biological
29 plausibility for an effect of Pb on cytokine production is provided by a large body of
30 toxicological evidence that clearly demonstrates a Pb-induced shift to a Th2 phenotype
31 with increases in the Th2 cytokine IL-4 and decreases in the Thl cytokine IFN-y. Several
32 of these observations were made in juvenile and adult animals exposed prenatally or
33 postnatally via diet that resulted in blood Pb levels (upon cessation of Pb exposure)
34 relevant to humans, 1.23-17 (ig/dL (lavicoli et al.. 2006b; Chen et al.. 2004; Dyatlov and
35 Lawrence. 2002). While results were not uniform for other cytokines (i.e., Thl cytokine
36 IL-2), most available results pointed to increases in Th2 cytokines, specifically, IL-6 and
37 IL-10, in Pb-exposed animals. Several studies demonstrated Pb-induced increases in
38 TNF-a but in animals with high prenatal dietary or postnatal air Pb exposure (e.g.,
November 2012 5-473 Draft - Do Not Cite or Quote
-------
1 149 jig/dL) (Chen et al.. 1999; Miller et al.. 1998; Zelikoff etal. 1993). Gao et al. (2007)
2 described a role for dendritic cells in skewing T cells to Th2 cytokine production. In vitro
3 evidence indicates that Pb may induce increases in TNF-a via MAPK signaling pathways
4 (Khan etal.. 2011; Gao et al.. 2007; Cheng et al.. 2006). Toxicological evidence indicates
5 effects on cytokine production of prenatal and postnatal Pb exposures, whereas
6 epidemiologic studies examined only concurrent blood Pb level. Because concurrent
7 blood Pb level in children and adults reflects both previous and current Pb exposures,
8 associations with concurrent blood Pb level may reflect an effect of cumulative Pb
9 exposure. Overall, the consistent toxicological evidence for Pb-induced decreases in Thl
10 cytokines and increases in Th2 cytokines and pro-inflammatory cytokines such as TNF-a
11 provides clear mode of action support for the evidence indicating the effects of Pb on
12 both increases in IgE and inflammation and decreased host resistance.
5.6.6.2 Decreased Nitric Oxide
13 As described in the 2006 Pb AQCD (U.S. EPA. 2006b). key mode of action support for
14 the effects of Pb on impairing macrophage function and decreasing host defense was
15 provided by consistent toxicological findings for Pb-induced decreases in NO, which is
16 involved in the cytotoxic activity of macrophages in host defense processes [see 2006
17 Annex Table AX5.9.6 (U.S. EPA. 2006h)1. In adult rodents, decreases in NO from
18 macrophages were observed with short-term Pb acetate exposures (1 or 6 days) during
19 early gestation (BunnetaL 200 Ib: Lee etal.. 200 Ib) but not long-term exposures
20 occurring during the full gestational period (Bunn etal.. 200 Ic; Chen etal.. 1999; Miller
21 et al.. 1998). With short-term exposure, decreases in NO were found in Sprague-Dawley
22 rats with blood Pb level 4.5 (ig/dL in males and 5.3 (ig/dL in females measured 2 weeks
23 after Pb exposure in drinking water of dams was terminated (BunnetaL. 200 Ib) and in
24 chicks with blood Pb levels that did not exceed 11 (ig/dL but with Pb injected into eggs
25 embryos (Lee etal.. 200Ib).
26 The short-term in vivo findings are supported by several in vitro observations of
27 decreases in NO in macrophages and splenocytes induced by a wide range of Pb exposure
28 concentrations (0.625-5 (iM) and durations (2 hours-6 days) (Tarrer et al.. 2008; Mishra
29 et al.. 2006a: Krocova et al.. 2000; Chen etal.. 1997; Tian and Lawrence. 1996. 1995).
30 Farrer et al. (2008) further indicated that the mode of action for Pb (5 (iM) may involve a
31 decrease in inducible NO synthase function in myeloid cells without a change in its gene
32 expression. Additionally, Pb glutamate abrogated the myeloid cell (CD1 lb+)-mediated
33 suppression of CD4+ T cell proliferation, and exogenous NO restored suppression.
34 Together, these findings indicated that Pb may indirectly enhance T cell proliferation
35 through its effect on decreasing NO production. Combined with the observation that Pb
November 2012 5-474 Draft - Do Not Cite or Quote
-------
1 can alter antigen processing (Farreretal.. 2005) and, hence, the quality and magnitude of
2 the acquired immune response against pathogen exposure, evidence indicated that
3 multiple arms of host defense against infectious challenge can be compromised.
4 Diminished production of NO in innate immune cells such as macrophages could affect
5 other physiological systems (e.g., neurological, cardiovascular, endocrine) that require
6 NO signaling cascades.
7 Consistent with the toxicological evidence, cross-sectional studies found associations
8 between concurrent blood Pb level with lower NO in populations living near Pb sources
9 (Barbosa et al.. 2006c: Pineda-Zavaleta et al.. 2004). These studies did not provide
10 sufficient information on participation rates; however, examination of populations near
11 Pb sources could limit generalizability of findings. Additional limitations include the
12 cross-sectional design that does not permit determining the temporal sequence between
13 Pb exposure and NO suppression and the limited consideration for potential confounding.
14 Because of these limitations, epidemiologic evidence is not a major consideration in
15 drawing conclusions about the effects of Pb on NO. However, they are useful in that they
16 show Pb-associated decreases in NO in humans, similar to toxicological studies.
17 In a study of 65 children (ages 6-11 years) in Lagunera, Mexico, mean concurrent blood
18 Pb levels increased (7.02 to 20.6 to 30.38 (ig/dL) with increasing school proximity
19 (650-8,100 meters) to a Pb smelter (Pineda-Zavaleta et al.. 2004). With adjustment for
20 age and sex, a 1 (ig/dL higher blood Pb level was associated with a 0.00089 (95% CI:
21 -0.0017, -0.00005) nmol/(ig protein lower NO release from macrophages activated by
22 phytohemagglutinin (PHA). Because PHA activates macrophages indirectly through the
23 activation of lymphocytes, the results indicated that Pb suppressed T cell-mediated
24 macrophage activation. Blood Pb group comparisons indicated that associations were due
25 largely to the lower NO in the 23 children closest to the smelter who had blood Pb levels
26 10.31-47.49 (ig/dL. Though not described in detail, higher blood Pb level was not
27 associated with lower NO in girls.
28 Among 104 adults (ages 18-60 years) in Sao Paolo, Brazil residing near a closed battery
29 plant, Barbosa et al. (2006c) observed an association between higher concurrent blood Pb
30 level and lower plasma NO in the 69 adults (mean blood Pb level: 6.4 (ig/dL) with the TC
31 or CC eNOS genotype (r = 0.23, p = 0.048). The results are consistent with the reduced
32 promoter activity and potentially reduced gene expression of the TC/CC variants. Results
33 were not adjusted for potential confounding factors, but subjects were nonsmoking,
34 nonalcohol drinking with normal mean BMI and SEP. The exclusion criteria may further
35 limit the generalizability of findings. Because NO was measured in plasma, immune cells
36 could not be identified as the source of NO. In contrast, Valentino et al. (2007) found
37 similar plasma NO levels in 44 male foundry workers (mean blood Pb level: 21.7 (ig/dL),
November 2012 5-475 Draft - Do Not Cite or Quote
-------
1 14 male pottery workers (mean blood Pb level: 9.7 (ig/dL), and 59 male unexposed
2 workers of similar age (mean blood Pb level: 3.9 (ig/dL, ages 25-61 years). Quantitative
3 results were not reported, but blood Pb level was reported not to be correlated with NO.
4 Potential confounding factors, including other workplace exposures were not examined.
5 In summary, toxicological evidence indicates that short-term dietary Pb exposure early in
6 gestation but not long-term exposure for the full gestational period results in reduced NO
7 production by macrophages. Evidence consistently demonstrates Pb-induced decreases in
8 NO in cell cultures. The relevance of toxicological evidence is supported by observations
9 of an association between higher concurrent blood Pb level and lower release of NO from
10 macrophages of children in Mexico. The association in children was due largely to lower
11 NO in macrophages from children living near a Pb smelter with concurrent blood Pb
12 levels >10 (ig/dL, higher than those in most of the current U.S. population. This study
13 had limited consideration for potential confounding factors. However, the toxicological
14 evidence provides clear mode of action support for the effects of Pb on decreasing host
15 resistance given the role of NO in mediating cytotoxic activity of macrophages.
5.6.6.3 Increased Reactive Oxygen Species and Prostaglandins
16 ROS are released from macrophages during phagocytosis and are involved in killing
17 invading bacteria. ROS and PGE2 are important mediators of inflammation which can
18 result in local tissue damage (Figure 5-34). The roles of ROS and PGE2 in both host
19 defense and injury may explain some of the inconsistencies in the evidence as reported in
20 the 2006 Pb AQCD. In activated macrophages undergoing phagocytosis, high
21 concentration (10-1,000 (JVI, 15 minutes-20 hours) Pb chloride or acetate exposures were
22 found to reduce release of ROS (Hilbertz et al., 1986; Castranova et al.. 1980). consistent
23 with observations of Pb-induced decreased bacterial and viral resistance. In resting
24 macrophages, Hilbertz et al. (1986) found that Pb acetate induced an increase in ROS one
25 hour but not 20 hours after exposure, indicating a transient response. Chen et al. (1997)
26 also found a Pb-induced (4 (iM Pb-glutamate, 18 hours) decrease in ROS but did not
27 indicate the functional state of macrophages. Shabani and Rabbani (2000) found a
28 Pb-induced (240 (iM Pb nitrate, 3 hours) increased ROS from alveolar macrophages that
29 was linked to their apoptosis, also consistent with impaired host defense. Other studies
30 reported depletion of antioxidants such as glutathione and catalase in conjunction with
31 reduced macrophage function in Swiss mice treated with Pb nitrate by oral gavage (40
32 mg/kg/day, 30 days) (Lodi et al.. 2011) or increases in PGE2 and apoptosis in vitro with
33 0.01-10 (iM Pb nitrate (3 hours) (Chettv et al.. 2005). While several processes have been
34 proposed to explain the mechanisms of Pb-induced oxidative damage, the exact
35 combination of processes involved remains to be determined (Section 5.2.4).
November 2012 5-476 Draft - Do Not Cite or Quote
-------
1 In adult animals, Pb exposure increased ROS release from macrophages immediately
2 upon cessation of exposure (Baykov et al.. 1996; Zelikoff etal.. 1993) but not 9-10 weeks
3 after exposure (Miller et al.. 1998). consistent with in vitro findings. These Pb exposures
4 occurred through relevant routes of exposure, i.e., diet or air, but with high
5 concentrations, 31 (ig/m3 Pb oxide in air for 3 hours/day for 4 days in rabbits (Zelikoff et
6 al.. 1993) and 1.5 mg/kg Pb acetate in diet for 30 days in Swiss mice (Baykov et al..
7 1996). Neither study measured the blood Pb levels of animals.
8 Pb-associated increases in ROS also were found in macrophages of humans. However,
9 the findings are based on cross-sectional design, higher blood Pb levels (means: 7.02,
10 20.6, and 30.38 (ig/dL) than most of those in the current U.S. population, and limited
11 consideration for potential confounding. In addition to finding suppressed NO production
12 (Section 5.6.6.2). Pineda-Zavaleta et al. (2004) found a Pb-associated increase in ROS
13 production from macrophages in children in Mexico living in varying proximities to a Pb
14 smelter. With adjustment for age and sex, a 1 (ig/dL higher concurrent blood Pb level was
15 associated with a 0.00389 (95% CI: 0.00031, 0.00748) (imol/mg higher release of
16 superoxide anion from macrophages directly activated by IFN-y/LPS. The blood
17 Pb-associated superoxide anion release was larger from macrophages of males. Because
18 IFN-y directly activates macrophages, these results indicated that Pb stimulated cytokine-
19 induced macrophage activation. Blood Pb level was not associated with ROS from
20 neutrophils in a study of male Pb recycling workers (ages 19-45 years) in India. Despite
21 large differences in blood Pb levels between 30 Pb workers (mean: 106 (ig/dL) and 27
22 unexposed controls (mean: 4.5 (ig/dL), levels of ROS released from neutrophils
23 (indicators of respiratory burst) were similar between groups (Mishra et al.. 2006a).
24 Evidence does not clearly indicate that neutrophils are a major responding cell to Pb
25 exposure (Section 5.6.2.5).
26 PGE2 is produced from the metabolism of cell membrane phospholipids and may be
27 released by macrophages to modulate their function in a paracrine or autocrine manner.
28 Toxicological studies have found Pb-induced increases in PGE2 in macrophages with
29 high concentration Pb exposures. Increases in PGE2 were found in turkeys exposed to
30 Pb acetate in feed PND1-PND21 and with a mean blood Pb level of 42 (ig/dL (Knowles
31 and Donaldson. 1997). Dietary Pb acetate exposure (250-2,000 ppm) of chicks
32 PND1-PND19 resulted in an increase in serum arachidonic acid but not PGE2 or other
33 prostaglandins (Knowles and Donaldson. 1990). In vitro studies also used high Pb
34 exposure concentrations, >20 (iM Pb chloride (Flohe et al.. 2002; Lee and Battles. 1994).
35 A recent in vitro study with human neuroblastoma cells found increases in PGE2 with
36 lower Pb concentrations (0.01-1 (iM Pb acetate) than previously reported (Chetty et al..
37 2005).
November 2012 5-477 Draft - Do Not Cite or Quote
-------
1 In summary, ROS and PGE2 function in modulating macrophage function, aiding in
2 bacterial killing, and inducing tissue damage as part of an inflammatory response.
3 Consistent with these diverse roles, toxicological studies have found both Pb-associated
4 increases and decreases in ROS and PGE2. Consistent with toxicological findings, a
5 cross-sectional study found an association between higher concurrent blood Pb level
6 (>10 (ig/dL) higher ROS release from macrophages in children in Mexico that was
7 adjusted for potential cofounding by only age and sex. In animals, Pb-induced increases
8 in macrophage production of ROS and PGE2 have occurred concomitantly with
9 functional alterations such as impaired macrophage phagocytosis and apoptosis.
10 Although toxicological results were based on examination of high Pb concentrations, they
11 nonetheless provide clear evidence for modes of action underlying the effects of Pb on
12 reduced macrophage function and decreased host resistance.
5.6.6.4 Cellular Death (Apoptosis, Necrosis)
13 The 2006 Pb AQCD reported contrasting effects of Pb on the apoptosis of macrophages
14 in vitro. However, with recent studies in mice, evidence suggests that Pb exposure may
15 induce apoptosis or mediators of apoptosis in immune cells. Xu et al. (2008) found that 4-
16 week dietary exposure of juvenile ICR mice to Pb acetate (50-100 mg/kg) induced DNA
17 damage in peripheral blood lymphocytes, increased p53 and Bax expression in the liver,
18 but did not change Bel-2 expression (creating a Bax/Bcl-2 imbalance). Bax promotes
19 apoptosis, whereas Bel-2 inhibits apoptosis. Concomitant increases in indicators of
20 oxidative stress in liver homogenate suggested that oxidative stress mediated Pb-induced
21 apoptosis. In Swiss mice, Bishayi and Sengupta (2006) found splenic macrophages to
22 have elevated DNA fragmentation, a key event in apoptosis, but with i.p. Pb acetate
23 treatment (10 mg/kg). Consistent with in vivo findings, Gargioni et al. (2006) found 20
24 and 40 (iM Pb nitrate to induce cell death in mouse peritoneal macrophages in vitro with
25 a concomitant loss of cell membrane integrity, indicating that Pb primarily induced
26 macrophage necrosis or cell lysis. While evidence for Pb-induced apoptosis of immune
27 cells with routes of Pb exposure relevant to humans is sparse, the evidence suggests that
28 the induction of cell death may be a potential mode of action for the effects of Pb on
29 macrophage function and decreased host resistance.
November 2012 5-478 Draft - Do Not Cite or Quote
-------
5.6.7 Immune Effects of Pb within Mixtures
1 In the 2006 Pb AQCD (U.S. EPA. 2006b). the immune effects resulting from Pb within
2 metal mixtures were not well characterized; however some recent studies indicated that
3 immune effects may be observed with lower levels of Pb exposure when they occur in
4 conjunction with other metals. In Swiss mice treated with Pb acetate (10 mg/kg i.p., daily
5 for 15 days), As (0.5 mg/kg i.p., daily for 15 days), or both, Bishayi and Sengupta (2006)
6 reported a greater than additive effect of co-administered Pb and As on decreasing
7 bacterial resistance, decreasing macrophage myeloperoxidase release, and NO
8 production.
9 Epidemiologic studies have not widely examined interactions between Pb and other
10 metals. However, consistent with Bishayi and Sengupta (2006). Pineda-Zavaleta et al.
11 (2004) (Section 5.6.6.2) found interactions between Pb and As among 65 children in
12 Mexico ages 6-11 years. Contamination of drinking water by both Pb and As was a
13 concern in the study area; however, urinary As levels were higher in children who had
14 lower blood Pb levels. Higher urinary As was associated with lower NO release from
15 macrophages (similar to blood Pb). Higher As and Pb internal dose was associated with a
16 larger decrease in NO (p for interaction = 0.037) than was either metal alone. Higher
17 urinary As was associated with lower superoxide anion release (opposite direction of Pb).
18 However, higher Pb and As internal dose was associated with a larger increase in
19 superoxide anion (p for interaction = 0.042) than was blood Pb level alone. Due to the
20 high blood Pb in these children (means in three groups at varying distances from a Pb
21 smelter: 7, 20.6, 30.4 (ig/dL), it is not clear whether these relationships would apply to
22 the current U.S. population of children.
23 Institoris et al. (2006) found that Cd or Hg co-exposure potentiated the effects of Pb.
24 Lymph node weight decreased in 4 week-old Wistar rats exposed to 20 mg/kg Pb acetate
25 by drinking water plus another metal but not with Pb alone. In contrast, Fortier et al.
26 (2008) did not find metal co-exposure to increase the effects of Pb. Pb chloride
27 (7.5-20.7 (ig/dL) did not alter lymphocyte proliferation, monocyte phagocytosis, or NK
28 cell activity in human leukocytes. A mixture of 20.7 (ig/dL Pb chloride plus 12.0 (ig/dL
29 methylmercuric chloride (MeHgCl) decreased lymphocyte proliferation; however, these
30 effects were attributed to MeHgCl, which singly had a stronger suppressive effect. Other
31 toxicological studies found metal mixtures that included Pb to decrease antibody titers or
32 increase neutrophil counts (Jadhav et al.. 2007; Massadeh et al.. 2007) but did not test
33 each metal individually. The latter findings cannot be attributed to interactions between
34 Pb and other components within the mixture. Overall, several results indicated that
35 exposures to Pb-containing metal mixtures are associated with immune effects. However,
November 2012 5-479 Draft - Do Not Cite or Quote
-------
1 not all results showed that co-exposures to metals such as As, Cd, or Hg produce increase
2 the immune effects of Pb.
5.6.8 Summary and Causal Determination
3 The cumulative body of epidemiologic and toxicological evidence describes several
4 effects of Pb exposure on the immune system related to a shift from Thl- to Th2-type
5 responses, including an increase in atopic and inflammatory conditions and a decrease in
6 host resistance. Outcomes related to an increase in atopic and inflammatory conditions
7 include asthma, allergy, increased IgE, and mode of action endpoints such as selective
8 differentiation of Th2 cells, increased production of Th2 cytokines, B cell activation, and
9 inflammation. Outcomes related to decreased host resistance include enhanced
10 susceptibility to bacterial and viral infection, suppressed DTH, and those describing
11 mode of action, i.e., decreased production of Thl cytokines, reduced phagocyte function,
12 and increased inflammation. A small body of studies indicates the effects of Pb exposure
13 on autoimmunity. The sections that follow describe the evaluation of evidence for these
14 three groups of outcomes, decreased host resistance, increased atopic and inflammatory
15 conditions, and autoimmunity, with respect to causal relationships with Pb exposure
16 using the framework described in Table II of the Preamble. The application of the key
17 supporting evidence to the causal framework is summarized in Table 5-34.
5.6.8.1 Evidence for an Increase in Atopic and Inflammatory
Conditions
18 Collective epidemiologic and toxicological evidence indicates that a causal relationship is
19 likely to exist between Pb exposure and atopic and inflammatory conditions. This
20 relationship is supported by evidence for associations of blood Pb levels with asthma and
21 allergy in studies in children (Jedrychowski et al., 2011; Pugh Smith and Nriagu. 2011;
22 Joseph et al.. 2005). Pb-associated increases in IgE in children and animals, and evidence
23 describing modes of action including increases in Th2 cytokines and inflammation.
24 Recent studies on asthma and allergy expand upon the evidence presented in the
25 2006 Pb AQCD by providing additional evidence from prospective analyses, and by
26 better addressing uncertainties regarding potential confounding by factors such as SES,
27 smoking exposures, and residential allergen exposures (Table 5-34). Findings from
28 studies that prospectively ascertained outcomes increase confidence that associations are
29 not due to reverse causation (Jedrychowski et al.. 2011; Joseph et al.. 2005). In these
30 studies, the lack of selective participation and objective assessment of outcomes of
November 2012 5-480 Draft - Do Not Cite or Quote
-------
1 asthma and allergy through medical records and clinical testing, respectively, indicates
2 lack of biased reporting of asthma and allergy in children with higher blood Pb levels
3 (Section 5.6.5.2 and Table 5-34). Among children age 5 years in Poland, Jedrychowski et
4 al. (2011) found that a 1 (ig/dL increase in prenatal cord blood Pb level was associated
5 with an increased risk of allergic sensitization of 2.3 (95% CI: 1.1, 4.6). The magnitude
6 of risk did not differ with and without adjustment for maternal education or residential
7 allergen levels. An additional strength of this study was the adjustment for prenatal
8 cotinine levels and postnatal smoker in the home. A large study of 4,634 children in
9 Michigan ages 1-3 years found that compared with Caucasian children with blood Pb
10 levels <5 (ig/dL measured up to 12 months before asthma assessment, Caucasian children
11 with blood Pb levels > 5 (ig/dL had an increased risk of incident asthma of 2.7 (95% CI:
12 0.9, 8.1) (Joseph et al.. 2005). Adjustment was made for census block average income,
13 which may not adequately control for potential confounding by individual subject-level
14 SES.
15 Supporting evidence was provided by a cross-sectional study of 356 children ages 0-12
16 years in Michigan, which found that compared with children with concurrent blood Pb
17 levels <10 (ig/dL, children with concurrent blood Pb level > 10 (ig/dL had increased
18 parental report of an asthma diagnosis in the previous 12 months with an OR of 7.5 (95%
19 CI: 1.3, 42.9) (Pugh Smith and Nriagu, 2011). This study was cross-sectional and
20 produced an imprecise effect estimate; however, a strength of the study was the relatively
21 extensive consideration for potential confounding, including adjustment for family-level
22 income. As with Jedrychowski et al. (2011). Pugh Smith and Nriagu (2011) found an
23 association with adjustment for smoking exposures in the home plus other indicators of
24 housing exposures and condition (Table 5-34). The studies of asthma and allergy differed
25 in which and how potential confounding factors were considered, particularly SES. While
26 there is no single complete measure of SES, the various indicators used across these few
27 studies produces uncertainty regarding residual confounding. Residual confounding also
28 is possible by factors not examined. The examination of maternal education and exposure
29 to smoking or allergens in Jedrychowski et al. (2011) and family income, smoking,
30 housing conditions, pets, or pests in Pugh Smith and Nriagu (2011) increase confidence
31 in the associations observed for blood Pb levels. However, because evidence is limited to
32 a few populations, there is uncertainty regarding potential confounding by SES and other
33 exposures well characterized in the literature to be associated with asthma and allergy.
34 With respect to blood Pb levels associated with atopic and inflammatory conditions,
35 Joseph et al. (2005) found elevated incidence of asthma in Caucasian children with earlier
36 childhood blood Pb levels > 5 (ig/dL and in African American children with blood Pb
37 levels > 10 (ig/dL. Pugh Smith and Nriagu (2011) found higher asthma prevalence in
38 children with concurrent blood Pb levels > 10 (ig/dL. Jedrychowski et al. (2011) found
November 2012 5-481 Draft - Do Not Cite or Quote
-------
1 increased allergic sensitization in association with cord blood Pb levels that were low
2 (geometric mean: 1.16 (ig/dL) but that may have been affected by maternal higher past
3 Pb exposures mobilized from bone to blood during pregnancy.
4 Biological plausibility for the relationships found between blood Pb levels and asthma
5 and allergy in children is provided by evidence characterizing modes of action, namely, a
6 Pb-associated shift in production from Thl cytokines (e.g., IFN-y) to Th2 cytokines (e.g.,
7 IL-4) and increase in Th2-dependent IgE levels (Table 5-34). A majority of this evidence
8 was available in the 2006 Pb AQCD (U.S. EPA. 2006b). The shift from Thl to Th2
9 cytokine production in animals was found with prenatal or postnatal (4 weeks in
10 juveniles, 3 weeks or 8 weeks in adults) dietary Pb exposures. In the studies available in
11 humans (Table 5-34), higher concurrent blood Pb levels were associated with higher
12 serum IL-4 in children (Lutz et al.. 1999) and higher serum IL-6 in nonoccupationally-
13 exposed adults (adjusted for age, BMI, and current smoking status and additional
14 adjustment for income, physical activity, education, and history of inflammatory
15 conditions in the large NHANES analysis) (Songdej etal., 2010; Kim et al., 2007).
16 Because of the limitations in the small body of epidemiologic studies, i.e., the cross-
17 sectional design of studies and inconsistent consideration for potential confounding, the
18 epidemiologic evidence is a lesser consideration in drawing conclusions about
19 Pb-associated cytokine changes. However, epidemiologic evidence does not detract from
20 the clear toxicological evidence for Pb-induced increases in Th2 cytokine production.
21 Coherence for a shift from Thl to Th2 cytokine production is found in the in vitro
22 evidence for Pb-induced selective differentiation of naive T cells to a Th2 subtype (Heo
23 et al.. 1998; 1996; McCabe and Lawrence. 1991). A recent study in adult mice and in
24 vitro provided new evidence that Pb may promote the shift to Th2 responses by
25 increasing production of Th2 cytokines in dendritic cells, the major effector in antigen
26 response (Gao et al.. 2007).
27 Additional mode of action support is provided by associations observed between higher
28 concurrent blood Pb levels and higher serum IgE in several different populations of
29 children (Section 5.6.3. Table 5-34). While most studies found elevated IgE in groups of
30 children with concurrent blood Pb levels >10 (ig/dL, Karmaus et al. (2005) found higher
31 serum IgE in children ages 7-10 years in Germany with blood Pb levels 2.8-3.4 (ig/dL
32 compared with children with lower blood Pb levels. Some studies found increasing IgE
33 across increasing blood Pb groups, except in the highest group (Karmaus et al.. 2005;
34 Lutz et al.. 1999); however, a monotonic concentration-response relationship was not
35 found in a recent study of children in Egypt (Hegazy et al.. 2011). Lutz et al. (1999)
36 recruited children in Michigan from a public assistance program, and Karmaus et al.
37 (2005) recruited schoolchildren but excluded those from homes where more than 12
38 cigarettes were smoked per day. The nature of recruitment may limit generalizability of
November 2012 5-482 Draft - Do Not Cite or Quote
-------
1 findings. Sufficient information was not reported to assess biased participation by Pb
2 exposure and history of allergy or asthma. The limited consideration for potential
3 confounding comprised adjustment for age (Karmaus et al., 2005; Lutz et al., 1999).
4 smoking exposure, serum lipids, blood organochlorine levels, and previous infections
5 (Karmaus et al., 2005) but not SES or allergen exposure. Although these findings were
6 based on cross-sectional analyses and had limited consideration for potential
7 confounding, they were supported by similar findings in animals, which are not subject to
8 reverse causation and confounding bias. Despite clear evidence in animals overall, there
9 was some inconsistency for Pb-induced increases in IgE in animals with gestational or
10 gestational/lactational dietary Pb exposures that resulted in blood Pb levels 5-20 (ig/dL,
11 which are more relevant to humans (Chen et al.. 2004; Snyder et al.. 2000). Miller et al.
12 (1998) found elevated IgE in adult F344 rats after gestational Pb exposure via drinking
13 water of dams whose blood Pb levels peaked at 30-39 (ig/dL. There is lack of coherence
14 between the consistent results for IgE and the inconsistent findings for Pb-induced
15 activation of B cells, which differentiate into allergic antibody-producing cells
16 (Section 5.6.3). However, there is strong mode of action support in animals for
17 Pb-induced increases in IL-4, which stimulates differentiation of B cells.
18 Further support for the effects of Pb exposure on increasing risk of atopic and
19 inflammatory conditions is provided by evidence of Pb-associated inflammation
20 (Section 5.6.4 and Table 5-34). Coherence for this evidence is found with findings for
21 Pb-induced increases in IgE which primes basophils and mast cells to release pro-
22 inflammatory mediators. Pb-induced inflammation is clearly demonstrated by a large
23 toxicological evidence base for the effects of Pb exposure on inducing macrophages into
24 a hyperinflammatory state as characterized by enhanced production of TNF-a, PGE2, and
25 ROS. Inflammation was observed in rabbits exposed to Pb via air for 4 days (31 (ig/m3)
26 (Zelikoff et al.. 1993) and rodents exposed via diet (250 ppm drinking water during
27 gestation, 1.5 mg/kg food postnatally for 30 days) (Miller et al., 1998; Bavkov et al.,
28 1996). Consistent with previous toxicological evidence, a large analysis of adults
29 participating in NHANES found an association between concurrent blood Pb levels and
30 serum CRP, an indicator of systemic inflammation, in 4,278 men with adjustment for
31 age, BMI, income, physical activity, education, history of inflammatory conditions,
32 cardiovascular disease, diabetes, and smoking status (Songdei et al.. 2010). Because only
33 concurrent blood Pb levels were examined, there is uncertainty regarding the temporal
34 sequence between Pb exposure and inflammation and the magnitude, timing, frequency,
35 and duration of Pb exposures that contributed to the observed associations. Because of
36 the sparse epidemiologic evidence, it is a lesser consideration in drawing conclusions
37 regarding the effects of Pb exposure on inflammation.
November 2012 5-483 Draft - Do Not Cite or Quote
-------
1 With respect to important lifestages of Pb exposure, gestational Pb exposures, producing
2 blood Pb levels 8 and 20 (ig/dL, were found to affect endpoints such as IgE and/or
3 cytokine levels in juvenile and adult rodents (Chen et al.. 2004; Snvder et al.. 2000).
4 However, increases in Th2 cytokines also were found in adult animals with lifetime Pb
5 exposures beginning in gestation and producing blood Pb levels 1-12 (ig/dL (lavicoli et
6 al.. 2006b). The Pb exposure lifestage, magnitude, frequency, and duration associated
7 with atopic and inflammatory conditions are not well characterized in humans. Cord
8 blood Pb level was associated with allergic sensitization in children (Jedrychowski et al..
9 2011). whereas other studies of children and adults examined only concurrent blood Pb
10 levels. Neither toxicological nor epidemiologic evidence clearly identifies an individual
11 critical lifestage or duration of Pb exposure that is more strongly associated with atopic
12 and inflammatory conditions. In children and adults, concurrent blood Pb levels are
13 influenced by cumulative (from remodeling of bone) and recent Pb exposures. The
14 combined evidence indicates that gestational and cumulative postnatal Pb exposures may
15 influence atopic and inflammatory conditions.
16 In conclusion, prospective studies in a few populations of children indicate associations
17 of prenatal cord and earlier childhood blood Pb levels with asthma and allergy, with a
18 cross-sectional study providing supporting evidence with associations with concurrent
19 blood Pb level. Prospective design, lack of selective participation of subjects, and
20 objective assessment of outcomes reduce the likelihood of undue selection bias and
21 reverse causation. These few studies varied in their consideration for potential
22 confounding by SES and exposure to smoking or allergens. Thus, uncertainty remains
23 regarding residual confounding in associations observed between blood Pb levels and
24 asthma and allergy in children. The evidence for asthma and allergy is supported by
25 cross-sectional associations found between higher concurrent blood Pb levels in children
26 and higher IgE, an important mediator of asthma and allergy. The biological plausibility
27 for the effects of Pb on IgE is provided by consistent findings in animals with gestational
28 or gestational-lactational Pb exposures, with some evidence at blood Pb levels relevant to
29 humans. In epidemiologic studies, higher IgE and higher asthma prevalence were
30 examined and found in children with concurrent blood Pb levels >10 (ig/dL. Coherence
31 for the evidence of Pb-associated increases in asthma, allergy, and IgE is found with
32 evidence for most of the examined endpoints related to mode of action, i.e., Pb-induced
33 increases in Th2 cytokine production and inflammation in animals. Neither toxicological
34 nor epidemiologic evidence clearly identifies an individual critical lifestage or duration of
35 Pb exposure associated with atopic and inflammatory conditions but indicates that
36 gestational and cumulative postnatal Pb exposures may influence atopic and
37 inflammatory conditions. The strong toxicological evidence supporting modes of action
38 for a shift to a Th2 phenotype combined with the epidemiologic evidence for asthma and
39 allergy in a few populations with some uncertainty regarding potential confounding is
November 2012 5-484 Draft - Do Not Cite or Quote
-------
1 sufficient to conclude that a causal relationship is likely to exist between Pb exposures
2 and an increase in atopic and inflammatory conditions.
5.6.8.2 Evidence for Decreases in Host Resistance
3 Evidence indicates that a causal relationship is likely to exist between Pb exposure and
4 decreased host resistance based on consistent observations that relevant Pb exposures
5 decrease responses to antigens (i.e., suppresses DTH) and increase bacterial titers and
6 subsequent mortality in rodents (Table 5-34. Sections 5.6.2.3 and 5.6.5.1). A majority of
7 this evidence was available in the 2006 Pb AQCD (U.S. EPA. 2006b). The studies that
8 reported blood Pb levels demonstrated increased bacterial titers and mortality with adult-
9 only 16 week Pb exposure via drinking water in adult mice with Salmonella infection and
10 blood Pb level 20 (ig/dL (Fernandez-Cabezudo et al.. 2007) and with lactational
11 (PND1-PND22) Pb exposure in juvenile mice with Listeria infection and blood Pb level
12 25 (ig/dL (Dyatlov and Lawrence, 2002). DTH was suppressed in adult rats with blood
13 Pb levels 6 and 25 (ig/dL after gestational Pb exposure in drinking water (Chen et al..
14 2004; Bunnetal. 200la). While a few epidemiologic studies found higher prevalence of
15 respiratory infections in children with higher concurrent blood Pb levels (Karmaus et al..
16 2005; Rabinowitz et al.. 1990) and Pb-exposed workers (Ewers etal.. 1982). the
17 implications are limited by the lack of rigorous statistical analysis (i.e., regression) and
18 consideration for potential confounding. These limitations also apply to the recent cross-
19 sectional evidence of Pb-related increases in respiratory infections in children (Carreras
20 et al.. 2009) (Table 5-34). These limitations produce uncertainty about the effects of Pb
21 exposure on decreased host resistance in humans but do not detract from the consistent
22 evidence in animals.
23 The effects of Pb on decreased host resistance are well supported by evidence describing
24 underlying modes of action (Table 5-34). Evidence in animals indicates Pb-induced
25 functional impairment of macrophages, which phagocytize pathogens. Decreased
26 macrophage colony formation was found in rats after gestational Pb exposure (Bunn et
27 al.. 2001b). and decreased phagocytic activity was found in mice and turkeys after
28 lactational or 2-week juvenile Pb exposure (Knowles and Donaldson. 1997; Kowolenko
29 et al.. 1991). Additional coherence for Pb-induced decreased host resistance is found with
30 observations in animals that gestational Pb exposure suppressed macrophage production
31 of NO which is involved in bacteria killing (Section 5.6.6.2) and postnatal Pb exposure
32 (air for 4 days, food for 30 days) increased production of ROS and PGE2, which mediate
33 tissue damage (Section 5.6.6.3). Similarly, a cross-sectional epidemiologic study found a
34 smaller release of NO and larger release of superoxide anion from macrophages of
35 children with higher concurrent blood Pb levels (10.3-47.5 versus <10.3 (ig/dL) (Pineda-
November 2012 5-485 Draft - Do Not Cite or Quote
-------
1 Zavaleta et al.. 2004) after adjustment for age and sex. Because of the limited
2 consideration for potential confounding in this study and examination of higher blood Pb
3 levels than those in most of contemporary U.S. children, the results are a lesser
4 consideration in drawing conclusions about the effects of Pb on macrophages. However,
5 they do suggest the relevance of toxicological observations to humans. The killing
6 capability of macrophages is enhanced by the Thl cytokine IFN-y. Therefore, an effect of
7 Pb exposure on decreased host resistance is additionally supported by clear evidence in
8 animals for the effects of Pb exposure on suppressing IFN-y production (Section 5.6.6.1).
9 A recent study in mice indicated that Pb-induced suppression of DTH may be mediated
10 by a shift in production from Thl to Th2 cytokines specifically in dendritic cells (Gao et
11 al.. 2007).
12 Some evidence did not contribute strong support for the mode of action for Pb-induced
13 decreased host resistance. Pb-exposed workers were found to have reduced functionality
14 of neutrophils, which respond to bacterial infection (Table 5-34. Section 5.6.2.5) but
15 without consideration for potential confounding or analogous toxicological evidence.
16 Neither epidemiologic nor toxicological evidence clearly demonstrated an effect of Pb
17 exposure on NK cells, which respond to viral infection (Section 5.6.2.7).
18 With respect to important lifestages of Pb exposure, animal studies found that gestational
19 Pb exposures, producing blood Pb levels of 6 and 25 (ig/dL, resulted in decreases in Thl
20 cytokines, suppression of DTH, and greater susceptibility to bacterial infection (Chen et
21 al.. 2004; Bunnetal.. 200la). However, these effects related to decreased host resistance
22 in mice also were affected by postnatal lactational (Dyatlov and Lawrence. 2002). adult
23 long-term (>4 weeks) (Fernandez-Cabezudo et al.. 2007). and lifetime Pb exposures
24 beginning in gestation in adult mice (lavicoli et al.. 2006b) that produced blood Pb levels
25 1-25 (ig/dL. Thus, the animal toxicological evidence does not clearly identify a particular
26 lifestage of Pb exposure that is more strongly associated with decreased host resistance.
27 In conclusion, decreased host resistance is demonstrated by several toxicological
28 observations that dietary Pb exposure producing relevant blood Pb levels increased
29 susceptibility to bacterial infection and suppressed DTH in rodents and by the coherence
30 with evidence describing modes of action, including suppressed production of Thl
31 cytokines and decreased macrophage function in animals. These effects were found with
32 gestational, lactational, adult-only, and lifetime Pb exposures of animals. Cross-sectional
33 epidemiologic evidence indicates Pb-associated increases in respiratory infections but
34 limitations, including the lack of rigorous methodology and consideration for potential
35 confounding produce uncertainty in the epidemiologic evidence for decreased host
36 resistance in humans. The consistent toxicological evidence in animals but uncertainty in
37 the epidemiologic evidence for decreased host resistance in humans is sufficient to
November 2012 5-486 Draft - Do Not Cite or Quote
-------
1 conclude that a causal relationship is likely to exist between Pb exposure and decreased
2 host resistance.
5.6.8.3 Evidence for Autoimmunity
3 Toxicological evidence indicates the potential of Pb to increase autoimmunity, with a few
4 previous studies showing Pb-induced generation of auto-antibodies (Hudson et al., 2003;
5 Bunn et al.. 2000; El-Fawal et al.. 1999; Waterman et al.. 1994) and recent studies
6 providing indirect evidence by showing formation of neoantigens that could result in the
7 formation of auto-antibodies (Table 5-34). Several observations were made in animals
8 injected with Pb, which is a route of exposure with less relevance to humans. Higher
9 levels of auto-antibodies also were found in Pb-exposed battery workers; however,
10 implications are limited because of the high blood Pb levels (range: 10-40 (ig/dL) of
11 some of the workers and lack of consideration for potential confounding by several
12 factors, including other occupational exposures (El-Fawal et al., 1999). Because results
13 from available toxicological and epidemiologic studies do not sufficiently inform
14 Pb-induced generation of auto-antibodies with relevant Pb exposures, the evidence is
15 inadequate to determine if there is a causal relationship between Pb exposure and
16 autoimmunity.
November 2012 5-487 Draft - Do Not Cite or Quote
-------
Table 5-34 Summary of evidence supporting immune causal determinations.
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or
Blood Levels
Associated with
Effects0
Increase in Atopic and Inflammatory Conditions - Likely Causal
Associations
consistently
found in high-
quality
epidemiologic
studies with
relevant blood
Pb levels
Prospective studies indicate higher asthma and
allergy incidence in association with earlier
childhood or prenatal blood Pb levels in children
(ages 1-5 yr) in a few populations (U.S., Poland)
Joseph et al. (2005).
Jedrychowski et al. (2011).
Section 5.6.5.2
Children (ages 1-3
yr) with blood Pb
levels measured
earlier in childhood
>10 ug/dL
Prenatal (cord):
geometric mean
1.16 ug/dL
Some studies report high participation and/or
follow-up retention, not conditional on Pb
exposure or outcome.
Some studies objectively assessed outcomes
with clinical testing, medical records.
Adjustment or consideration for potential
confounding by SES, exposure to smoking,
and/or allergen exposure.
Heterogeneity in evaluation of potential
confounding among the few available studies
produces uncertainty regarding potential
confounding.
Supporting cross-sectional evidence in children
(ages 6 mo-10 yr) for increases in IgE but with
limited consideration for potential confounding
factors, particularly SES. Associations observed
in studies in U.S., Europe, Asia; insufficient
information to assess potential selection bias.
Evidence for C-R varies for IgE. Some studies
show increasing IgE across blood Pb groups,
except in highest group.
Another study did not show monotonic C-R
relationship.
Joseph etal. (2005).
Jedrychowski et al. (2011)
Jedrychowski et al. (2011),
Pugh Smith and Nriagu (2011)
Jedrychowski et al. (2011).
Pugh Smith and Nriagu (2011)
Children:
Karmauset al. (2005).
Hegazy et al. (2011).
Lutz et al. (1999).
Hon et al. (2010: 2009),
Sun et al. (2003)
Section 5.6.3
Karmauset al. (2005).
Lutz et al. (1999)
Hegazy et al. (2011
Groups (ages 6 mo
- 10yr) with
concurrent blood Pb
levels >10 ug/dL
Epidemiologic
evidence
supported by
toxicological
evidence at
relevant
exposures
Most animal studies show elevated IgE in
animals with prenatal and postnatal dietary Pb
exposures. Some inconsistency in animals with
relevant Pb concentrations.
Increase in IgE:
Snyderetal. (2000).
Miller et al. (1998)
No IgE increase in
Chen etal. (2004)
Also see Section 5.6.3
Increased IgE with
gestational-
lactational Pb
exposure, blood Pb
means 5, 20 ug/dL
Gestational Pb
exposure producing
maternal blood Pb
peak: 30-39 ug/dL
No increase with
gestational Pb
exposure, blood Pb
means 7-8 ug/dL
November 2012
5-488
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Evidence clearly
describes mode
of action
Stimulation of
Th2 phenotype
Key Supporting Evidence13 References'1
Extensive, consistent evidence of increased Section 5.6.6.1
production of Th2 cytokines (e.g., IL-4, IL-6, &
Pb Exposure or
Blood Levels
Associated with
Effects0
IL-10) in animals with prenatal and postnatal
(4 weeks juvenile, 3, 8 weeks adults) dietary Pb
exposures. Recent evidence for the role of
dendritic cells in mediating Th2 shift.
Limited available cross-sectional evidence in
children, adults. Epidemiologic evidence has
limited consideration for potential confounding. Is
not a major consideration in conclusions.
The few available in vitro studies indicate
activation Th2 cells from naTve T cells or over
Th1 cells.
Inflammation Extensive evidence for increased production of
TNF-a, IL-6, ROS, PGE2 by macrophages from
animal with prenatal and postnatal (dietary
4 weeks juvenile, dietary 3, 8 weeks adults, air
4 days adults) Pb exposure. Supported by in
vitro evidence.
Cross-sectional association observed in children
living near Pb source, adjusted for confounding
by age and sex but not other factors such as
SES. Cross-sectional evidence in adults in
NHANES that adjust for inflammatory conditions,
smoking and SES. Is not a major consideration
in conclusions.
B cell Inconsistent toxicological evidence in animals for
activation B cell activation by Pb exposure concentration
and duration and strain.
Inconsistent epidemiologic evidence for B cell
abundance, B cell activation not examined.
Table 5-7 of the 2006 Pb AQCD
(U.S. EPA. 2006g)
Children:
Lutz et al. (1999)
Adults:
Kim et al. (2007)
Section 5.6.6.1
Section 5.6.2.1
Sections 5.6.6.1 and 5.6.6.3
Children:
Pineda-Zavaleta et al. (2004)
Sections 5.6.6.2. and 5.6.6.3
Adults:
Songdej et al. (2010)
Sections 5.6.4
Section 5.6.3
Table 5-32 and Section 5.6.3
Children (ages 6 mo-
6yr):
Concurrent blood Pb
group range
15-19ug/dL
Adults:
Group range
2.5-10.5 ug/dL
Children (ages
6-11 yr):
Concurrent blood Pb
group
>10 ug/dL
Adults:
Concurrent group
>1.16 ug/dL
Decreases in Host Resistance - Likely Causal
Consistent
toxicological
evidence with
relevant
exposures
Available
epidemiologic
evidence is not
sufficiently
informative
The few studies with relevant dietary Pb
exposures demonstrate increased bacterial
infection, sickness behavior, and mortality in
mice. Similar observations in several other
studies with higher Pb exposures.
The few studies with relevant prenatal dietary Pb
exposures show suppressed DTH in rodents.
Similar observations in several other studies with
higher Pb exposures.
Epidemiologic studies found associations with
increased respiratory infections but limitations
include lack of consideration for potential
confounding, rigorous statistical analysis, or Pb
biomarker assessment, and/or ecological study
design
Dyatlov and Lawrence (2002).
Fernandez-Cabezudo et al.
(2007)
Section 5.6.5.1
Chenetal. (2004),
Bunn et al. (2001 a: 200 1c)
Section 5.6.2.3
Children:
Karmauset al. (2005),
Rabinowitz et al. (1990).
Carreras et al. (2009)
Pb-exposed workers:
Ewers (1 982)
Section 5.6.5.1
Blood Pb means
20 ug/dL after adult
1 6-week
Pb exposure,
25 ug/dL after
lactational
Pb exposure
Blood Pb means:
6.75, 25 ug/dL after
gestational Pb
exposure
Children (ages 7-10
yr):
Group with
concurrent blood Pb
>3.34 ug/dL,
Group with cord
blood Pb>10 ug/dL
Pb-exposed
workers:
Concurrent blood Pb
21-85 ug/dL
November 2012
5-489
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
References
Pb Exposure or
Blood Levels
Associated with
Effects0
Evidence clearly
describes mode
of action
Decreased
macrophage
function
Decreased
Th1 cytokine
(IFN-y)
production
Decreased macrophage colony formation in
animals with dietary prenatal and postnatal Pb
exposure; not widely examined.
Decreased macrophage phagocytosis in animals
and in cell culture, not widely examined.
Several studies demonstrate decreased NO
production by macrophages from animals with
prenatal and postnatal Pb exposure. Supported
by in vitro evidence.
Cross-sectional association of decreased NO in
macrophages of children living near Pb source
with higher concurrent blood Pb level, adjusted
for age and sex but not SES.
Inconsistent evidence in Pb-exposed workers
but for macrophage abundance, not function.
Consistent evidence from a large body of
lexicological studies with prenatal and postnatal
(4 weeks juvenile, 3, 8 weeks adults) dietary Pb
exposures of animals.
Section 5.6.2.4
Section 5.6.2.4
Section 5.6.6.2
Pineda-Zavaleta et al. (2004)
Section 5.6.6.2
Pinkerton et al. (1998).
Fischbein et al. (1993).
Conterato (In Press)
Section 5.6.6.1 and
Table 5-7 of the 2006 Pb AQCD
(U.S. EPA.2006g)
Children (ages
6-11 yr):
Group with
concurrent blood Pb
>10 ug/dL
Autoimmunity - Inadequate
Available
toxicological and
epidemiologic
evidence is not
sufficiently
informative
A study in rats shows generation of auto-
antibodies with relevant adult-only dietary Pb
exposure for 4 days. Several other studies have
Pb exposure concentrations and/or routes
(e.g., i.p.) with uncertain relevance to humans.
Rats:
EI-Fawaletal. (1 999)
Section 5.6.5.4
Rats:
Blood Pb level
range
11-50 ug/dL
Evidence for increased auto-antibodies in
Pb-exposed workers with high blood Pb levels
and limited consideration for potential
confounding, including other workplace
exposures.
Workers:
EI-Fawaletal. (1999)
Section 5.6.5.4
Workers:
Blood Pb level
range:
10-40ug/dL
Described in detail in Table II of the Preamble.
bDescribes the key evidence and references contributing the most heavily to causal determination. References to earlier sections
indicate where full body of evidence is described.
""Describes the blood Pb levels in children with which the evidence is substantiated and blood Pb levels in animals most relevant to
humans.
November 2012
5-490
Draft - Do Not Cite or Quote
-------
5.7 Hematological Effects
5.7.1 Introduction
1 The effects of Pb exposure on red blood cell function and heme synthesis have been
2 extensively studied over several decades in both human and animal studies. The 1978
3 National Ambient Air Quality Standard for Lead was set to protect blood Pb levels in
4 children from exceeding 30 (ig/dL as such levels were associated with impaired heme
5 synthesis, evidenced by accumulation of protoporphyrin in erythrocytes (U.S. EPA.
6 1978).
7 The 2006 Pb AQCD (U.S. EPA. 2006b) reported that Pb exposure significantly decreases
8 several hematological parameters including hemoglobin (Hb), hematocrit (Hct), mean
9 corpuscular volume (MCV), mean corpuscular hemoglobin (MCH), and mean
10 corpuscular hemoglobin concentration (MCHC). Further, the 2006 Pb AQCD reported
11 that Pb affects developing red blood cells (RBCs) in children and occupationally exposed
12 adults as noted by anemia observed with blood Pb >40 (ig/dL. Pb-induced anemia is
13 thought to occur due to decreased RBC life span and effects on Hb synthesis. The exact
14 mechanism for these effects is not known, although Pb-induced changes on iron uptake or
15 inhibition of enzymes in the heme synthetic pathway may be responsible. Once Pb enters
16 the cells, it is predominantly found in protein-bound form, with Hb and aminolevulinic
17 acid dehydratase (ALAD) both identified as targets.
18 Consistent with the majority of human evidence that high Pb blood levels
19 (i.e., >20 (ig/dL) are associated with decreased hematological indices, blood Pb levels
20 >100 (ig/dL were associated with decreased RBC survival in laboratory animals. Effects
21 on RBC membrane mobility were observed at blood Pb levels as low as 10 (ig/dL,
22 although the precise mechanisms underlying these effects of Pb are not known. Studies
23 conducted in animal and in vitro models provide evidence of multiple other effects on
24 RBC membranes, including altered microviscosity and fluidity, decreased sialic acid
25 content, decreased lamellar organization, decreased lipid resistance to oxidation (possibly
26 mediated by perturbations in RBC membrane lipid profiles), and increased permeability.
27 These alterations to RBC membranes may indicate potential modes of action by which Pb
28 induces RBC fragility, abnormal cellular function, RBC destruction, and ultimately
29 anemic conditions. Pb exposure also has been shown to result in increased activation of
30 RBC scramblase, an enzyme responsible for the expression of phosphatidylserine (PS) on
31 RBC membranes. This expression of PS decreases the life span of RBCs via phagocytosis
32 by macrophages. Pb exposure has been observed to alter the phosphorylation profiles of
November 2012 5-491 Draft - Do Not Cite or Quote
-------
1 membrane proteins, which may influence the activity of membrane enzymes and the
2 functioning of receptors and channels located on the membrane.
3 The 2006 Pb AQCD reported that Pb exposure affects heme synthesis in humans and
4 animals through the inhibition of multiple key enzymes, most notably ALAD, the enzyme
5 that catalyzes the second, rate-limiting step in heme biosynthesis (Figure 5-36 presents a
6 schematic representation of the heme biosynthetic pathway). The 2006 Pb AQCD (U.S.
7 EPA. 2006b) further reported that decreased RBC ALAD activity is the most sensitive
8 measure of human Pb exposure, in that measurement of ALAD activity is correlated with
9 blood Pb levels. Concentration-response changes in the ratio of activated/nonactivated
10 ALAD activity in avian RBCs were observed to be not dependent on the method of Pb
11 administration. The Pb-associated inhibition of the ALAD enzyme was consistently
12 observed in RBCs from multiple species, including birds, cynomolgous monkeys, and
13 humans. Pb was also observed to inhibit other enzymes responsible for heme
14 biosynthesis, including ferrochelatase, porphobilinogen (PEG) deaminase, and
15 coproporphyrinogen oxidase. Pb also potentially alters heme biosynthesis through
16 inhibition of transferrin (TF) endocytosis and iron transport.
17 Pb has been found to alter RBC energy metabolism through inhibition of enzymes
18 involved in anaerobic glycolysis and the pentose phosphate pathway. Pb was also found
19 to inhibit pyrimidine 5'-nucleotidase (P5N) activity, and the 2006 Pb AQCD indicated
20 that this might be another biomarker of Pb exposure. Inhibition of P5N results in an
21 intracellular increase in pyrimidine nucleotides leading to hemolysis and potentially
22 ultimately resulting in anemic conditions. The 2006 Pb AQCD indicated that
23 perturbations in RBC energy metabolism may be related to significant decreases in levels
24 of nucleotide pools, including nicotinamide adenine nucleotide (NAD), possibly due to
25 decreased NAD synthase activity, and nicotinamide adenine nucleotide phosphate
26 (NADP) accompanying significant increases in purine degradation products.
27 The 2006 Pb AQCD identified oxidative stress as an important potential mode of action
28 by which Pb exposure induced effects on RBCs. Increased lipid peroxidation and
29 inhibition of antioxidant enzymes in RBCs (e.g., superoxide dismutase [SOD], catalase
30 [CAT]) were observed following exposure to Pb.
31 The epidemiologic and toxicological studies published since the 2006 Pb AQCD, largely
32 support the reported Pb-associated effects on RBC function and heme synthesis.
33 Epidemiologic studies support previous observations that occupationally-exposed adults
34 with higher blood Pb levels than the current U.S. general population (>26 (ig/dL) have
35 decreased RBC numbers. However, a few epidemiological studies investigating
36 occupationally-exposed adults and pregnant women provide some evidence that more
37 relevant blood Pb levels, <10 (ig/dL, are associated with decreased RBC numbers,
November 2012 5-492 Draft - Do Not Cite or Quote
-------
1 possibly through decreased survival of the RBCs. Effects seen in children are largely
2 consistent with those observed in adults, and a number of toxicological studies support
3 findings observed in human populations. Recent epidemiologic and toxicological studies
4 also support previous findings that Pb exposure in adults, children, and laboratory
5 animals decreases ALAD activity, as well as the activity of other enzymes in the heme
6 biosynthetic pathway. Recent epidemiologic and toxicological studies expand upon the
7 evidence that Pb exposure results in oxidative stress in RBCs. Although the
8 epidemiologic studies included below are cross-sectional in study design, they do
9 improve upon earlier studies as more studies characterize the effects in children, and
10 investigate effects in populations with blood Pb levels more comparable to those in the
11 current U.S. population. Additionally, the associations observed in these cross-sectional
12 studies are supported by a large number of animal toxicology studies.
5.7.2 Red Blood Cell Function
13 As stated in the 2006 Pb AQCD (U.S. EPA. 2006b). Pb poisoning in children has been
14 associated with anemia. As of 2006, the mechanism for this was not clear, but it was
15 determined not to be due to iron deficiency, which can be found to occur independently
16 of Pb exposure. However, Zimmerman et al. (2006) found that blood Pb level differences
17 were statistically significant, and lower in non-anemic (or mildly-anemic) iron-
18 deficient 5- to 9-year-old children in India fed an iron-fortified diet for 30 weeks,
19 compared to 14 weeks (mean [range]: 8.1 [3.1-219] (ig/dL versus 12.1 [3.7-26.8] (ig/dL;
20 p <0.02); however, blood Pb levels were not significantly lower in children receiving the
21 no-iron diet for 30 weeks compared to 14 weeks (mean [range]: 10.2 [4.4-25.3] (ig/dL
22 versus 12.0 [3.8-25.5] (ig/dL). Although a number of epidemiologic studies found
23 decreases in RBCs and/or Hct levels associated with higher blood Pb levels, it is not
24 known whether this is due to reduced RBC survival or a decrease in RBC production.
25 Regardless, decreased RBC survival and hematopoiesis can be expected to occur
26 simultaneously, and any effect on RBC numbers is likely a combination of the two modes
27 of action.
5.7.2.1 Pb Uptake, Binding, and Transport into Red Blood Cells
28 The 2006 Pb AQCD (U.S. EPA. 2006b) reported that Pb uptake into human RBCs occurs
29 via passive anion transport mechanisms. Although Pb can passively cross the membrane
30 in both directions, little of the Pb is found to leave the cell after entry. Simons (1993b)
31 found that in vitro uptake of 203Pb (1-10 (iM) occurred via an anion exchanger while the
32 efflux occurred via a vanadate-sensitive pathway. After entry into the RBC, radioactive
November 2012 5-493 Draft - Do Not Cite or Quote
-------
1 Pb was found to partition with Hb at a ratio estimated to be about 6000:1 bound to
2 unbound (Simons. 1986). However, Bergdahl et al. (1997a) suggested that ALAD was
3 the primary Pb-binding protein and not Hb. The 2006 Pb AQCD also reported that the
4 majority (approximately 98%) of Pb accumulates in RBC cytoplasm bound to protein,
5 and only about 2% is found in the membrane. This is related to the high ratio of Pb in
6 RBCs compared to plasma Pb. Further information on Pb binding and transport in blood
7 can be found in the kinetics section of Chapter 4 (Section 4.2).
8 Although no recent studies were identified that examined transport of Pb into RBCs, Lind
9 et al. (2009) recently observed that several Zn ionophores (8-hydroxyquinoline
10 derivatives and Zn and Na pyrithione) were able to effectively transport Pb out of RBCs
11 into the extracellular space.
5.7.2.2 Red Blood Cell Survival, Mobility, and Membrane Integrity
12 A number of cross-sectional studies have investigated the effect of exposure to Pb on
13 various inter-connected and related hematological parameters in children and adults. As
14 these studies were cross-sectional in design, there is uncertainty regarding the
15 directionality of effects and the magnitude, timing, frequency, and duration of Pb
16 exposure that contributed to the observed associations. Additionally, unless explicitly
17 noted, potential confounding by subject characteristics and other workplace or residential
18 exposures was not accounted for in these studies. Adults and children exposed to Pb may
19 also have been co-exposed to other contaminants that can affect the hematological
20 system, and the potential for co-exposure was not assessed in most studies.
21 In an earlier cross-sectional study of children in Idaho (aged 1-9 years) with blood Pb
22 levels ranging from 11 to 165 (ig/dL (approximately 40% were >40 (ig/dL), a 10%
23 probability of anemia (Hct <35%) was predicted (in association with blood Pb levels of
24 ~20 (ig/dL [age 1 year], 50 (ig/dL [age 3 years] and 75 (ig/dL [age 5 years]) (Schwartz et
25 al.. 1990). More recent studies have also demonstrated adverse effects on hematological
26 parameters in children due to the Pb exposure. Ahamed et al. (2006) studied 39 male
27 urban adolescents in India who were separated into groups according to their blood Pb
28 level (Group 1: <10 (ig/dL [mean 7.4 (ig/dL], Group 2: >10 (ig/dL [mean 13.27 (ig/dL]).
29 Although the groups were similar in age (mean [SD]: 16.59 [0.91] versus 16.76 [0.90]
30 years, respectively), height, weight, and BMI (therefore, not considered to be potential
31 confounders), Group 2 had a significantly lower packed cell volume (PCV) compared to
32 Group 1. In a related study, Ahamed et al. (2007) investigated the relationship between
33 blood Pb level, anemia, and other hematological parameters in urban children in India
34 (n = 75). Children were split into two groups as above: Group 1 had blood Pb levels
November 2012 5-494 Draft - Do Not Cite or Quote
-------
1 <10 (ig/dL (mean [SD]: 6.89 [2.44] (ig/dL, n = 19), whereas Group 2 had blood Pb levels
2 >10 (ig/dL (mean [SD]: 21.86 [7.58] (ig/dL, n = 56). As with the earlier study, ages were
3 similar between the two groups: mean [SD]: 4.68 [1.49] and 4.11 [1.77] years,
4 respectively. Hb and Hct were significantly decreased in Group 2, compared to Group 1
5 after adjusting for age, sex, and area of residence. Children in Group 2 had an increased
6 odds ratio of anemia (OR: 2.87 [95% CI: 1.60, 2.87]) compared to Group 1 after
7 adjustment for age, sex, and area of residence.
8 In a cross-sectional study measuring blood Hb as the independent variable, blood Pb
9 levels were observed to decrease with increasing blood Hb. Riddell et al. (2007) found
10 that 21% of children, who were 6 months to 5 years of age living in the rural Philippines,
11 had concurrent blood Pb levels >10 (ig/dL (total population mean: 6.9 (ig/dL). After
12 controlling for potential confounding by age, sex, birth weight, and history of
13 breastfeeding, Hb levels were inversely related to blood Pb level, with a decrease of 3%
14 blood Pb associated with every 1 g/dL increase in Hb. Among children aged 6-36 months
15 (n = 222) living in Montevideo, Uruguay, 32.9% had blood Pb levels greater than
16 10 (ig/dL (population mean [SD]: 9.0 [6.0] (ig/dL) (Oueirolo etal.. 2010V The mean
17 [SD] Hb concentration was 10.5 [1.5] g/dL, and 44.1% of children were diagnosed as
18 anemic (Hb <10.5 g/dL). Blood Pb levels were significantly higher in anemic children
19 compared to non-anemic (mean [SD]: 10.4 [6.8] versus 7.9 [5.1] (ig/dL), and anemic
20 children were more likely to have elevated blood Pb after controlling for age and
21 mouthing behavior (OR = 1.9, 95% CI: [1.098, 3.350]). The likelihood of elevated blood
22 Pb was more pronounced in anemic children younger than 18 months (OR = 3.1, 95% CI:
23 [1.3,7.4]).
24 Similarly, in a cross-sectional study of 340 children (aged 1-5 years) from Karachi,
25 Pakistan, mildly-anemic and severely-anemic children (mean [SD] Hb levels: 8.9 [0.9]
26 and 7.4 [0.5] g/dL, respectively) had lower Hb levels but higher blood Pb levels
27 compared to non-anemic children (mean [SD] Hb: 12.1 [1.3] g/dL). Mean [SD] blood Pb
28 levels in the mildly-anemic, severely-anemic, and non-anemic children were 14.9 [0.81],
29 21.4 [2.7], and 7.9 [1.7] ng/dL, respectively (p <0.01) (Shah etal.. 2010). Additionally,
30 Hct, RBC count, and MCV were all decreased in anemic children versus non-anemic
31 children. Although statistical analyses were not reported, the levels of Hb, Hct, RBC
32 count, and MCV in anemic children all fell outside of the reported normal range for these
33 parameters, whereas the reported values in non-anemic children did not. Blood Pb level
34 was negatively correlated with Hb level in all groups, with the magnitude of negative
35 correlation increasing with increasing severity of anemia: r = -0.315 (non-anemic
36 children), -0.514 (mildly-anemic), and -0.685 (severely- anemic). In iron-deficient
37 anemic children (n = 23) from Denizli, Turkey, mean (SD) serum Pb levels were
38 statistically (p <0.05) increased compared to healthy children (n = 179): 0.013 (0.004)
November 2012 5-495 Draft - Do Not Cite or Quote
-------
1 versus 0.008 (0.001) (ig/dL, respectively (Turgut et al., 2007). The iron-deficient children
2 were observed to have decreased Hb, MCV, RBC, and ferritin compared to controls, but
3 increased RDW. In 140 children from southern Brazil aged 2-11 years, living within 25
4 km of a Pb smelter, blood Pb levels were not observed to differ between anemic and
5 non-anemic children (mean [SD]: 10.36 [6.8] versus 9.73 [5.8] (ig/dL, p = 0.98) (Rondo
6 et al.. 2006). However, blood Pb levels were significantly negatively correlated with Hb
7 in anemic children (r = -0.41, p = 0.01); this relationship was not observed in non-anemic
8 children (r = 0.018, p = 0.84).
9 In children aged 5-9 years (n = 189) without anemia living in Cartagena, Columbia, a
10 smaller percentage (4.7%) of children had blood Pb >10 (ig/dL (mean [SD]: 5.49
11 [0.23] (ig/dL). The only hematological parameters that fell outside of their reference
12 values were MCV and MCH, which were negatively correlated with blood Pb levels
13 (r = -0.159 [p = 0.029] and -0.171 [p = 0.019], respectively) (Olivero-Verbel et al.. 2007).
14 RBC count, which was not observed to differ from reference values, was positively
15 correlated with blood Pb level (r = 0.208, [p = 0.004]). In a group of 268 Lebanese
16 children, children aged 11-23 months with blood Pb levels >10 (ig/dL had increased
17 likelihood of having iron-dependent anemia and transferrin saturation (TF <12%)
18 compared to age-matched children with blood Pb levels <10 (ig/dL (OR = 4.59, 95% CI:
19 [1.51, 13.92]) (Muwakkit et al.. 2008). In children aged 24-35 months, higher blood Pb
20 level was not associated with increased likelihood of either effect. Huo et al. (2007)
21 found that children (less than 6 years of age) living near an area where electronic waste
22 was recycled in China had significantly higher mean blood Pb levels than did children in
23 the neighboring town with no waste recycling (15.3 versus 9.94 (ig/dL). However,
24 contrary to the findings above, no difference was detected in the mean Hb levels of the
25 children in the two towns (12.76 g/dL in children from the waste recycling town versus
26 12.35 g/dL in children from the town with no recycling).
27 In adult, occupationally exposed populations, decreased erythrocyte numbers and Hb
28 were observed in multiple, earlier cross-sectional studies investigating workers with
29 blood Pb levels >40 (ig/dL (Solliwav et al.. 1996; Horiguchi et al.. 1991; Poulos et al..
30 1986). However, a larger, longitudinal study (Hsiao etal. 2001) observed that
31 occupationally-exposed adults exhibited erythrocyte counts and Hct that were positively
32 associated with blood Pb levels. Most of the recent occupationally-exposed groups
33 represent populations highly exposed to Pb, with mean blood Pb levels ranging from 26-
34 74 (ig/dL. Although effects observed within these groups may not be generalizable to the
35 general population as a whole, they are useful in demonstrating consistent effects on a
36 number of hematological parameters, including Hb, MCV, MCH, MCHC, total RBCs,
37 and packed cell volume (PCV) (Khan etal.. 2008: Patil et al.. 2006a: Patil et al.. 2006b:
38 Karita et al.. 2005).
November 2012 5-496 Draft - Do Not Cite or Quote
-------
1 A few recent cross-sectional occupational studies did investigate the effect of moderate
2 occupational Pb exposure on hematological parameters. In gas station attendants in
3 Sarajevo (Bosnia and Herzegovina), workers (mean [SD] duration of exposure: 12.1 [9.1]
4 years) had significantly increased blood Pb levels (mean: 5.96 (ig/dL) in 2008, compared
5 to the same population that were previously examined in 2003 (mean: 4.07 (ig/dL; mean
6 [SD] duration of exposure: 10.4 [5.5]). Levels of MCH and MCHC were significantly
7 decreased when assessed in 2008, compared to the 2003 measurements, although RBC
8 numbers, Hb, Hct, and MCV were increased in 2008 compared to 2003. Positive
9 correlations were observed in all subjects between blood Pb and RBC count, Hb, and
10 MCH (r = 0.241, 0.201, and 0.213, respectively; p <0.05). No control group was included
11 in this study (Cabaravdic et al.. 2010). Ukaejiofo et al. (2009) studied the hematological
12 effects of Pb in 81 male subjects moderately exposed to Pb at three different
13 manufacturing companies in Nigeria for durations between six months and 20 years. The
14 exposed individuals had a mean blood Pb level of 7.00 (ig/dL compared to 3 (ig/dL in
15 controls drawn from industries not involved in Pb handling (control group I) and 2 (ig/dL
16 in controls drawn from the general population (control group II). Pb-exposed workers had
17 significantly reduced Hb and PCV levels and increased percentage of reticulocytes
18 compared to controls. Although the differences were statistically significant between the
19 exposed and control subjects, the study authors stated that the levels in the exposed
20 subjects were at the lower range of normal for Nigerians. The percent cell lysis did not
21 significantly differ between controls and exposed workers; however, when workers and
22 controls were stratified by age, there was a significant increase in cell lysis in workers
23 under age 30 compared to similarly aged controls in group II (p <0.01). Similarly,
24 stratification of subjects by duration of exposure revealed that MCHC was decreased in
25 exposed workers (6-60 months of exposure). Conterato et al. (In Press) investigated
26 hematological parameters in automotive painters exposed to Pb in Brazil. Exposed
27 painters had a mean [SEM] blood Pb concentration of 5.4 [0.4] (ig/dL compared to 1.5
28 [0.1] (ig/dL in controls. The mean [SEM] duration of exposure to Pb in painters was
29 133.9 [14.5] months, whereas the controls were not occupationally exposed to Pb.
30 Although differences in Hct, Hb concentration, and the number of RBCs were
31 significantly decreased in painters compared to controls, these differences were not
32 correlated with blood Pb levels; however, these parameters were correlated with blood
33 Cd2+ levels, which were also significantly elevated in painters compared to controls.
34 Taken together, the above occupational studies provide consistent evidence that high
35 (mean blood Pb >26 (ig/dL) occupational exposure to Pb reduces the number of RBCs in
36 circulation. Additionally, the Ukaejiofo et al. (2009) study suggests that blood Pb levels
37 below 10 (ig/dL (7.0 (ig/dL) may also result in decreased RBC survival. Although the
38 decrease in RBCs observed in highly exposed worker populations may be explained by
39 both decreased RBC survival and/or disruption of hematopoiesis, the observation of
November 2012 5-497 Draft - Do Not Cite or Quote
-------
1 increased reticulocytes in Ukaejiofo et al. (2009) seems to represent compensation for
2 decreased RBC survival due to Pb exposure.
3 In a non-occupational study, the associations between blood Pb levels, Ca2+, Fe, and Hb
4 were investigated in 55 pregnant Brazilian women (21.9% were 14-19 years old, 74.5%
5 were 20-34 years olds, and 3.6% were > 35 years old) (Zentner et al.. 2008). The
6 majority of women (across all age groups) had concurrent blood Pb levels <5 (ig/dL
7 (58.2%), although the mean blood Pb level was not reported; only 5.4% of women had
8 blood Pb levels >10 (ig/dL. The vast majority of the women (78.2%) were also observed
9 to have adequate levels of Hb (> 11 g/dL). In a multiple linear regression model, blood
10 Pb level was observed to be negatively associated with Hb ((3 = -0.359), when controlling
11 for age, BMI, income, energy intake, Ca2+ intake, vitamin C intake, and Fe intake.
12 The associations of blood Pb levels with hematological parameters observed in
13 epidemiologic studies are clearly supported by a number of animal toxicology studies
14 reporting blood Pb levels relevant to humans, i.e., <10 (ig/dL. Hb concentrations in
15 plasma (a marker of RBC hemolysis) was significantly increased in rats exposed to
16 Pb acetate (1,000 ppm in drinking water for 9 months; blood Pb level: 7.1 (ig/dL)
17 compared to controls (Baranowska-Bosiacka et al.. 2009). In a complementary in vitro
18 experiment, a concentration-dependent increase in the amount of hemolysis was observed
19 in human RBCs exposed to Pb at concentrations ranging from 0.1-100 (iM for 5-30
20 minutes. Hemolysis was increased even at the lowest concentration tested (i.e., 0.1 (iM).
21 Pb-induced hemolysis in these experiments may be due to inhibition of RBC
22 phosphoribosyltransferases (Section 5.7.2.5). In weanling rats (PND25 days, n = 10)
23 whose dams were exposed to Pb acetate in drinking water (2.84 mg/mL, approximating
24 mean [SD] daily exposures of 342.57 [28.11] and 744.47 [29.27] mg/kg [dam weight]
25 during gestation and lactation, respectively), blood Pb level was significantly elevated
26 compared to controls (mean [SE]: 69.8. [7.82] versus 0.54 [0.08] (ig/dL ). The only
27 hematological parameter affected by Pb exposure was Hct, which was decreased in
28 exposed rats (mean [SE]: 27.3 [0.5]%) versus controls (33.4 [0.3]%) (Molina et al.,
29 2011). In rats treated with 25 mg Pb/kg by oral gavage for 4 weeks, average plasma Pb
30 concentrations were 6.5 (ig/dL (9.6-fold higher than controls), and statistically significant
31 decreases in Hct, Hb, and RBCs were observed (Lee et al.. 2005). Effects on erythrocyte
32 survival were similar in mice treated with Pb nitrate (50 mg/kg via gavage for 40 days):
33 mean [SD] blood Pb levels were 1.72 [0.02] (ig/dL versus 0.09 [0.011] (ig/dL in control
34 mice, and exposed mice had significantly reduced total RBC counts, total leukocyte
35 counts, Hb, lymphocytes, and monocytes compared to controls (p <0.001) (Sharma et al..
36 2010b).
November 2012 5-498 Draft - Do Not Cite or Quote
-------
1 A number of toxicological studies also reported similar hematological effects, but did not
2 report final blood Pb concentrations. Rats exposed to Pb acetate (2 g/L in drinking water
3 for 30 days) had significantly decreased RBCs, Hb, PCV, MCH, and MCHC compared to
4 controls (p <0.05) (Simsek et al.. 2009). but not a disruption of hematopoiesis. Mice
5 exposed to Pb acetate (1 g/L in drinking water for 90 days, but not those exposed for 15
6 or 45 days), had significantly decreased RBC counts and Hct compared to controls
7 (p<0.05) (Marques et al., 2006). Spleen weights were observed to be increased relative to
8 body weight in animals exposed to Pb for 45 days. Mice injected daily with Pb acetate
9 (50 mg/kg subcutaneously) had significantly reduced Hb, MCV, MCH, and MCHC
10 compared to controls injected with 5% dextrose (Wang et al., 2010g).
11 Some toxicological studies found no evidence of hematological effects in animals
12 following exposure to Pb. Male rats exposed to Pb acetate in their drinking water for
13 4 weeks at concentrations ranging from 100-1,000 ppm had a concentration-dependent
14 increase in blood Pb levels (range: 6.57-22.39 (ig/dL) compared to controls (0.36 (ig/dL),
15 but there were no significant changes in any of the hematological parameters (complete
16 blood cell count performed) measured at the end of treatment (Lee et al.. 2006b). Slight,
17 statistically nonsignificant increases in PS expression on RBC membranes were also
18 observed. Similarly, exposure of male rats to 5,000 ppm Pb nitrate in drinking water
19 (blood Pb not reported) for three weeks had no affect on any measured hematological
20 parameter (Gautam and Flora. 2010). In vitro experiments with rat and human blood did
21 not demonstrate a significant increase in hemolysis after 4 hours of treatment with
22 Pb acetate at concentrations up to 10
23 Although Pb exposure has been consistently shown to shorten RBC life span and alter
24 RBC mobility, as of the 2006 Pb AQCD, the mechanism of this was not well understood.
25 While the mechanism is still not fully understood, there has been some indication for a
26 role of free Ca2+. Occupational studies investigating highly Pb-exposed worker
27 populations (mean blood Pb >28 (ig/dL) observed increased intracellular free Ca2+ levels
28 ([Ca2+]0 in RBCs, and decreased RBC membrane Ca2+/Mg2+ATPase activity in workers
29 compared with unexposed controls (Abam et al.. 2008; Quintanar-Escorza et al.. 2007).
30 [Ca2+]j levels were highly correlated with blood Pb levels even among unexposed control
31 populations with mean blood Pb levels of approximately 10 (ig/dL (9.9 ± 2 (ig/dL)
32 (Quintanar-Escorza et al., 2007). Changes in [Ca2+]j were associated with increased
33 fragility of the RBCs and dramatic morphological alterations, including the increased
34 presence of echinocytes (cells without normal biconcave shape) and crenocytes
35 (speculated cells) in Pb-exposed workers.
36 Similar to the associations observed in Quintanar-Escorza et al. (2007). [Ca2+]j increased
37 in a concentration-dependent manner when RBCs from healthy human volunteers were
November 2012 5-499 Draft - Do Not Cite or Quote
-------
1 exposed (in vitro) to 0.2 or 0.4 (iM Pb nitrate for 24 or 120 hours (0.4 (iM Pb nitrate
2 roughly approximates 10 (ig/dL Pb, although concentrations in exposure media are not
3 directly comparable to blood Pb levels) (Quintanar-Escorza et al., 2010). The increase in
4 [Ca2+]j levels was observed to be related to increased Ca2+ influx and decreased efflux. As
5 was observed among highly Pb-exposed workers, changes in [Ca2+]j were associated with
6 increased fragility of the RBCs and dramatic morphological alterations following
7 exposure to 0.4 (iM Pb. Similarly, Ciubar et al. (2007) found that RBC morphology was
8 disrupted, with > 50% RBCs having lost the typical discocytic morphology and
9 displaying moderate to severe echinocytosis following exposure to Pb nitrate
10 concentrations of 0.5 (iM or higher for 24 hours. Exposure of RBCs to higher
11 concentrations (concentrations not stated) of Pb nitrate resulted in cell shrinkage. In rats
12 exposed to 200 ppm Pb acetate via drinking water for three months (mean [SD] blood Pb
13 level: 40.63 [9.21] (ig/dL), the cholesterol/phospholipid ratio of RBC membranes was
14 increased, indicating that RBC membrane fluidity was decreased.
15 Khairullina et al. (2008) observed that the surface profiles of RBC membrane shadows
16 incubated with 0.5-10 (iM Pb acetate for three hours were much smoother than were
17 untreated RBC membranes when examined by atomic force microscopy. The authors
18 postulated that the observed smoothing in Pb-treated RBC membranes may be due to
19 clusterization of band 3 protein. Band 3 (anion exchanger 1 [AE1]), is a
20 chloride/bicarbonate (C1~/HCO3~) exchanger and is the most abundant protein in RBC
21 membranes. AE1 is integral in carbon dioxide (CO2) transport and linkage of the cellular
22 membrane to the underlying cytoskeleton (Akel et al.. 2007; Su et al.. 2007). The
23 observed smoothing of the RBC membrane may due to Pb interfering with how the
24 membrane attaches to the cytoskeletal structure of the RBC through perturbation of the
25 normal activity of AE1.
Eryptosis
26 Eryptosis is the suicidal death of RBCs. It is characterized by cell shrinkage, membrane
27 blebbing, and cell membrane phospholipid scrambling associated with PS exposure on
28 the cell membrane that leads to cell destruction via macrophages (Foller et al.. 2008;
29 Lang et al.. 2008). As previously reported in the 2006 Pb AQCD (U.S. EPA. 2006bX
30 Kempe et al. (2005) found that exposing human RBCs to Pb at concentrations ranging
31 from 0.3 (iM to 3 (iM caused increased activation of K+ channels that led to cell
32 shrinkage and scramblase activation. The activation of scramblase increased the exposure
33 to PS on the cell membrane, which causes an increase in the destruction of the RBCs by
34 macrophages.
November 2012 5-500 Draft - Do Not Cite or Quote
-------
1 Consistent observations were made in recent studies that included in vitro and in vivo
2 evidence. Shin et al. (2007) found that in vitro exposure of human RBCs to 1-5 (iM
3 Pb acetate increased PS expression in a time- and concentration-dependent manner. The
4 maximum mean [SE] increase in expression of PS was 26.8% [3.15] (compared to
5 deionized water), following exposure to 5 (iM Pb for four hours. Scramblase activity was
6 increased in Pb-exposed RBCs, and [Ca2+]l5 which regulates scramblase activation, was
7 also increased in exposed RBCs. Flippase, which translates PS exposure to inner
8 membranes, is inhibited by high levels of [Ca2+]j and was observed to exhibit reduced
9 activity following Pb exposure. The inhibition of flippase is additionally influenced by
10 the depletion of cellular adenosine triphosphate (ATP). ATP levels were decreased in a
11 concentration-dependent manner following exposure to Pb. To corroborate these findings
12 in vivo, Shin et al. (2007) treated male rats with Pb acetate (i.p. to 25, 50, or 100 mg/kg;
13 blood Pb not reported). Expression of PS was observed to increase in a concentration-
14 dependent manner at concentrations > 50 mg/kg, confirming the in vitro results. No
15 hemolysis or microvesicle formation was observed in the in vitro or in vivo experiments.
16 In a follow-up study, the same laboratory observed that in vitro exposure of human RBCs
17 to much lower concentrations of Pb acetate (0.1, 0.25, and 0.5 (JVI) also induced PS
18 expression. Most notably, exposure to 0.1 (iM Pb for 24 hours increased PS expression
19 on RBC membranes by approximately 20% (Jang et al., 2011). Accompanying the
20 increased expression of PS (associated with Pb exposure) was the presence of abnormal
21 echinocytic RBCs. Unlike the Shin et al. (2007) study described above, incubation of the
22 RBCs with low concentrations of Pb (0.1 (iM) induced the generation of microvesicles,
23 which also expressed PS on their membranes in this (Jang et al.. 2011) study. At 0.5 (iM,
24 Pb-exposed RBCs with externalized PS were observed to be targeted and engulfed by
25 differentiated macrophages. Similar ex vivo effects were observed in rat erythrocytes four
26 hours after oral exposure (0, 10 and 50 mg/kg) to Pb, although higher concentrations
27 were generally required. PS expression on the rat erythrocytes was also observed. To
28 corroborate these in vitro and ex vivo findings, rats were also exposed in vivo to 0, 50,
29 250, or 1,000 ppm Pb acetate in drinking water for 4 weeks. At 1,000 ppm, Hb and Hct
30 were significantly decreased relative to control, and liver and spleen weights were
31 increased. At the two highest doses, iron accumulation was observed in the spleen, a clear
32 sign of increased RBC clearance via phagocytosis.
November 2012 5-501 Draft - Do Not Cite or Quote
-------
1 Ciubar et al. (2007) also found that exposure to Pb nitrate (0.5-2 (iM) resulted in an
2 increase in PS exposure in RBCs and cell shrinkage, which the authors stated were
3 indicators of cell apoptosis. As reported above, Khairullina et al. (2008) observed
4 Pb-induced RBC membrane smoothing that may be due to alterations in AE1 activity.
5 Disruptions in AE1 activity may also result in enhanced PS exposure and premature cell
6 death. Akel et al. (2007) observed that in AE1"" knockout mice, Pb-induced PS exposure
7 was much greater than that in wild type mice. Decreased RBCs and increased
8 reticulocytes were also observed, an indication of high cell turnover.
5.7.2.3 Red Blood Cell Hematopoiesis
9 Erythropoietin (EPO) is a glycoprotein hormone excreted by the kidney to promote the
10 development of RBCs in the bone marrow. As reported in the 2006 Pb AQCD, analyses
11 of the cohort of children in Yugoslavia observed that EPO was increased in children aged
12 4.5 and 6.5 years of age living in a town near Pb sources (blood Pb levels >30 (ig/dL)
13 compared to children living in more distant town (blood Pb levels <10 (ig/dL), when
14 stratified by Hb concentrations (Graziano et al.. 2004; Factor-Litvak et al.. 1999; Factor-
15 Litvak et al., 1998). These differences were not observed in children aged 9.5 or 12 years.
16 With adjustment for Hb concentrations, blood Pb levels were observed to be significantly
17 associated with EPO levels at ages 4.5 and 6.5 years when considering all children
18 together. No significant association was observed at ages 9.5 and 12 years. Hb was not
19 observed to differ at any age between towns, thus possibly indicating that
20 hyperproduction of EPO is necessary to maintain Hb levels in young children living near
21 Pb sources. The authors postulated that increases in EPO in younger children reflect bone
22 marrow hyperactivity to counteract RBC destruction, whereas the lack of EPO elevation
23 in older children may reflect a transitional period where increasing renal and bone
24 marrow toxicity leads to decreases in EPO observed later in life, as observed in anemic,
25 pregnant women (Graziano et al.. 1991). Decreased EPO concentrations were also
26 observed in association with Pb exposure in adults in two cross-sectional studies cited in
27 the 2006 Pb AQCD (Osterode et al.. 1999; Romeo etal. 1996).
28 Consistent with findings that EPO is negatively associated with blood Pb levels in adults,
29 Sakata et al. (2007) observed that non-anemic tricycle taxi drivers (n=27) working in
30 Kathmandu, Nepal (blood Pb level: 6.4 (ig/dL) had significantly lower levels of EPO
31 (12.7 versus 18.8 mU/mL) compared to non-driver controls (blood Pb level: 2.4 (ig/dL).
32 In taxi drivers, there was an inverse relationship between the level of serum
33 erythropoietin and blood Pb level (r = -0.68, p <0.001). Blood Pb level was not associated
34 with any other hematological effects.
November 2012 5-502 Draft - Do Not Cite or Quote
-------
1 Recent toxicology studies of cytotoxicity and genotoxicity in RBC precursor cells
2 support the observations that Pb exposure disrupts normal hematopoiesis. Cytotoxicity
3 and genotoxicity in RBC precursor cells are strong indications of altered hematopoiesis in
4 bone marrow. Celik et al. (2005) observed that treatment of female rats with Pb acetate
5 (140, 250, or 500 mg/kg via gavage once per week for 10 weeks; blood Pb not reported)
6 resulted in decreased numbers of polychromatic RBCs (PCE) and increased numbers of
7 micronucleated PCEs, compared to controls (p <0.001). Alghazal et al. (2008b) exposed
8 male and female rats to 100 mg/L Pb acetate daily in drinking water for 125 days (blood
9 Pb not reported) and observed increases in micronucleated PCEs in female rats (p = 0.02)
10 but no significant reduction in the ratio of PCEs to normochromic RBCs (NCE). In male
11 rats, an increase in micronucleated PCEs was observed (p <0.001) along with a decrease
12 in the PCE/NCE ratio (p = 0.02). While the results from Alghazal et al. (2008b) indicate
13 that Pb is cytotoxic in male rats only, but is genotoxic in both sexes, results from Celik et
14 al. (2005) indicate that Pb is cytotoxic in female rats as well. Mice exposed to Pb acetate
15 (1 g/L in drinking water for 90 days; blood Pb not reported) had statistically significant
16 increases in micronucleated PCEs; a small, but statistically nonsignificant decrease in the
17 PCE/NCE ratio was also observed (Marques et al.. 2006).
5.7.2.4 Membrane Proteins
18 While there have been few studies, evidence included in the 2006 Pb AQCD indicated
19 there are effects of Pb on changes in RBC proteins. Huel et al. (2008) found that newborn
20 hair and cord blood Pb levels (mean [SD]: 1.22 [1.41] (ig/g and 3.54 [1.72] (ig/dL
21 respectively) were negatively associated with Ca2+ATPase activity in plasma membranes
22 of RBCs isolated from cord blood after controlling for gestational age and maternal Ca
23 pump activity. However, newborn hair Pb levels were more strongly associated with cord
24 Ca2+-pump activity than were cord blood Pb (p <0.0001 versus p <0.05). Maternal blood
25 Pb levels were not correlated with Ca2+-pump activity in maternal or newborn cord blood.
26 Pb-induced disruptions in Ca2+ homeostasis in RBCs can lead to cytotoxicity and
27 necrosis, and these effects may be representative of cellular dysfunction in other organ
28 systems.
29 In RBC membranes from Pb-exposed workers, Fukumoto et al. (1983) used
30 polyacrylamide electrophoresis analysis and found increased levels of polypeptides in
31 bands 2, 4, 6, and 7 and decreased levels of polypeptides in band 3. Apostoli et al. (1988)
32 found changes in RBC membrane polypeptides, including a significant decrease in band
33 3, in occupationally exposed workers with blood Pb levels greater than 50 (ig/dL.
34 Apostoli et al. (1988) suggested that band 3 may represent an anion channel protein,
35 whereas, Fukumoto et al. (1983) suggested that the changes in the RBC membrane
November 2012 5-503 Draft - Do Not Cite or Quote
-------
1 polypeptides may cause changes in membrane permeability. Exposure to Pb acetate at
2 concentrations above 0.1 (iM for 60 minutes has also been found to increase the
3 phosphorylation of proteins in human RBC membranes in vitro (Belloni-Olivi et al.,
4 1996). Phosphorylation did not occur in cells depleted of protein kinase C (PKC),
5 indicating a PKC-dependent mechanism.
5.7.2.5 Red Blood Cell Energy Metabolism Enzymes
6 RBCs use high energy purine nucleotides (i.e., ATP and guanine triphosphate [GTP]) to
7 support basic metabolic functions. In mature RBCs, these nucleotides are synthesized via
8 salvage reactions through either an adenine pathway, which requires adenine
9 phosphoribosyltransferase (APRT), or an adenosine pathway, which requires adenosine
10 kinase. The 2006 Pb AQCD (U.S. EPA. 2006b) reported that Pb significantly reduces the
11 nucleotide pool (including NAD and NADP, as well as increases purine degradation
12 products) resulting in altered RBC energetics. Since the 2006 Pb AQCD was published,
13 there have been few studies examining Pb effects on energy metabolism. Baranowska-
14 Bosiacka et al. (2009) examined the effects of Pb on RBC APRT and hypoxanthine-
15 guanine phosphoribosyltransferase (HPRT). In an in vitro experiment, APRT and HPRT
16 were measured in lysate of human RBCs after exposure to Pb at a concentration range
17 from 0.1 to 100 (iM for 5-30 minutes. Complementary in vivo tests measured APRT and
18 HPRT in RBC lysate from rats exposed to Pb acetate (1,000 ppm) in drinking water for
19 9 months. Both the in vivo and vitro studies found a significant decrease in both HPRT
20 and APRT levels. The levels in human RBCs were significantly decreased in vitro after
21 only 5 minutes of exposure to the 0.1 (iM concentration, but the decrease was also
22 concentration-dependent. However, the study authors considered the inhibition moderate
23 (30-35%) even with the highest Pb levels used in vitro. Shin et al. (2007) found a
24 concentration-dependent decrease in intracellular ATP in human RBCs in vitro with
25 significant decreases, found even with the lowest concentration (i.e., 1
5.7.2.6 Other Enzymes
26 The 2006 Pb AQCD (U.S. EPA. 2006b) reported that K+ permeability was increased by
27 Pb exposure due to altered sensitivity of the membrane Ca2+-binding site that caused
28 selective efflux of K+ ions from the RBC membrane. However, inhibition of the RBC
29 Na+/K+ATPase is more sensitive to Pb exposure than is the inhibition of
30 Ca2+/Mg2+ATPase. Few recent studies were found that examined the effects of Pb
31 exposure on other enzymes. Ekinci et al. (2007) tested the effects of Pb exposure on two
32 carbonic anhydrase isozymes (I and II) isolated from human RBCs. Carbonic anhydrases
November 2012 5-504 Draft - Do Not Cite or Quote
-------
1 are metalloproteins that use Zn to catalyze the equilibrium between CO2 and bicarbonate
2 in the cells of higher invertebrates. Although investigators found that Pb nitrate inhibited
3 both carbonic anhydrase isozymes in a concentration-dependent manner, the
4 concentrations used (i.e., 200-1,000 (iM) were above those that would be physiologically
5 relevant. Inhibition of isozyme I was noncompetitive, while the inhibition for isozyme II
6 was uncompetitive. Bitto et al. (2006) examined the mechanisms of action of Pb-induced
7 inhibition of P5N, an enzyme important in the pyrimidine salvage pathway that requires
8 Mn for normal activity. Pb was observed to bind directly to the active site of the enzyme
9 in a different position than the Mn, thus possibly resulting in improper protein folding
10 and inhibition of activity.
5.7.2.7 Red Blood Cell Oxidative Stress
11 It has been suggested that the Pb-associated decreases in ALAD activity result in
12 increased oxidative stress, owing to the buildup of ALA. ALA can act as an electron
13 donor in the formation of reactive oxygen species (ROS) (Nemsadze et al.. 2009;
14 Ahamed and Siddiqui. 2007). Many epidemiologic and toxicological studies have found
15 an association between the level of blood Pb and lipid peroxidation, antioxidant levels, or
16 indicators of ROS production. The same limitations regarding cross-sectional studies
17 listed in Section 5.7.2.2 (including uncertainty in directionality of effects and specific
18 information regarding exposure) apply to the epidemiologic studies investigating RBC
19 oxidative stress. Additionally, potential confounders and co-exposures were not
20 considered in the majority of these studies. However, in studies were confounders were
21 considered, they are explicitly delineated in the text.
Oxidative Stress, Lipid Peroxidation, and Antioxidant Enzymes
22 Malondialdehyde (MDA) is an end product of lipid peroxidation and is commonly
23 measured as an indicator of oxidative stress. Evidence of lipid peroxidation has been
24 observed in children moderately exposed to Pb. Ahamed et al.(2008; 2006. 2005)
25 investigated the relationship between blood Pb levels and antioxidant enzyme levels and
26 lipid peroxidation in children in India. In children (n = 62) aged 4-12 years in Lucknow,
27 India, children with mean blood Pb levels of 11.39 (SD: 1.39) (ig/dL had increased
28 measures of lipid peroxidation and decreased GSH levels compared to children with
29 mean blood Pb levels of 3.93 (SD: 0.61) or 7.11 (SD: 1.25) fig/dL (Ahamed et al.. 2005).
30 Catalase activity was decreased in children with a mean 7.11 (SD: 1.25) (ig/dL blood Pb
31 level, compared to children with mean 3.93 (SD: 0.61) (ig/dL blood Pb level.
32 Additionally, blood Pb levels were found to be significantly positively correlated with
November 2012 5-505 Draft - Do Not Cite or Quote
-------
1 MDA and CAT, and negatively correlated with GSH. In a similar study, Ahamed et al.
2 (2006) observed significantly higher levels of CAT and MDA in children with a mean
3 13.27 (ig/dL blood Pb level compared to children with a mean 7.40 (ig/dL blood Pb level;
4 with other characteristics such as age, height, wieght, and BMI not differing between the
5 two groups and thus, not considered as potential confounders. Examining all the study
6 subjects together, investigators found a correlation between blood Pb level and blood
7 MDA and RBC CAT levels, as well as an inverse relationship between ALAD activity
8 and MDA and CAT levels. Among Indian children with neurological disorders, blood Pb
9 levels were significantly increased compared to healthy control children (18.60 versus
10 10.37 (ig/dL respectively) (Ahamed et al., 2008). Potential confounding characteristics
11 such as age, sex, area of residence, and SES were not observed to be statistically different
12 between the two groups, and therefore, were not included in statistical analyses. In
13 addition, the following indicators of oxidative stress were elevated among case children:
14 increased blood MDA, RBC SOD and CAT levels, and decreased blood GSH levels. GPx
15 levels were similar between the two groups. Typical indicators of Pb exposure
16 (active/nonactive ALAD ratio) were found to be correlated with lipid peroxidation and
17 oxidative stress. Children aged 3-6 years old living near a steel refinery in China with
18 blood Pb levels > 10 (ig/dL also had a significant increase in plasma MDA compared to
19 children with blood Pb levels <10 (ig/dL. However, levels of RBC SOD, GSH, and GPx
20 were not different from those in controls (Jin et al., 2006).
21 Evidence of lipid peroxidation was also observed in occupational cohorts moderately
22 exposed to Pb. In auto repair apprentices in Turkey (mean [SD]: 16.8 [1.2] years of age,
23 3.8 [1.8] years duration of exposure) with minimum blood Pb levels of 7.9 (ig/dL
24 (Ergurhan-Ilhan et al.. 2008). increases in glutathione peroxidase (GPx) and MDA, as
25 well as decreases in a-tocopherol and (3-carotene were observed compared with controls
26 (mean [SD] age: 16.3 [1.0] years, mean blood Pb level: 2.6 (ig/dL). Decreases were
27 observed in SOD and CAT, but the results did not attain statistical significance.
28 Statistically significant alterations in measures of oxidative stress were also observed in
29 other occupationally exposed populations. SOD, glutathione (GSH), and CAT were
30 decreased; while oxidized GSH (i.e., GSSG) and thiobarbituric acid reactive species
31 (TEARS, expressed in terms of MDA) were increased in painters in India (mean [SD]
32 duration of exposure: 126.08 [49.53 months], mean blood Pb level: 21.92 (ig/dL,
33 compared to 3.06 (ig/dL in controls) (Mohammad et al., 2008). Glutathione-S-transferase,
34 GPx, and SOD were positively correlated with blood Pb levels (mean: 5.4 (ig/dL,
35 r = 0.34, 0.38, and 0.32, respectively; p <0.05) in automotive painters in Brazil
36 (Conterato et al.. In Press).
November 2012 5-506 Draft - Do Not Cite or Quote
-------
1 Numerous cross-sectional, occupational studies have also demonstrated increased lipid
2 peroxidation in highly-exposed worker populations (blood Pb levels ranging from 29.0 to
3 74.4 (ig/dL) (Kasperczyk et al.. 2009; Khan et al. 2008; Quintanar-Escorza et al.. 2007;
4 Patil et al.. 2006a: Patil et al.. 2006b). There was a correlation between MDA levels and
5 blood Pb levels, even in the unexposed workers who had lower (i.e., <12 (ig/dL) blood Pb
6 levels, although the magnitude of correlation in exposed workers was greater (Quintanar-
7 Escorza et al.. 2007). Increases in C-reactive protein and decreases in RBC SOD,
8 catalase, and plasma ceruloplasmin were also observed in these workers, further
9 indicating increased RBC oxidative stress due to higher Pb exposure.
10 Oral administration of Pb (25 mg/kg) to rats once a week (i.e., bolus gavage) for 4 weeks,
11 which produced a blood Pb level of about 6.5 (ig/dL, caused a significant increase in
12 RBC MDA levels (Lee et al.. 2005). Other indications of Pb-induced oxidative stress
13 included significant increases in RBC SOD and CAT levels accompanied by significant
14 decreases in GSH and GPx. Exposure of rats to Pb acetate (750 mg/kg in drinking water
15 for 11 weeks) resulted in decreased concentrations of plasma vitamin C, vitamin E,
16 nonprotein thiol, and RBC-GSH, with simultaneous increased activity of SOD and GPx
17 (Kharoubi et al.. 2008b). CAT activity was also slightly elevated in RBCs from the
18 Pb-exposed rats, but the increase failed to reach statistical significance. Exposure of male
19 rats to 5,000 ppm Pb nitrate in drinking water (blood Pb not reported) for three weeks
20 decreased GSH levels compared to that in controls (mean [SE]: 1.91 [0.02] versus 2.44
21 [0.09] mg/mL, respectively) (Gautam and Flora. 2010). SOD activity was significantly
22 decreased in rats injected with Pb acetate (15 mg/kg, i.p. for seven days, but not rats
23 injected with 5 mg/kg) (Berrahal et al.. 2007). GPx activity and MDA concentrations
24 were slightly elevated in the two exposed groups, but differences with the control
25 (15 mg Na acetate/kg) group failed to reach statistical significance. Effects on indices of
26 oxidative stress were also observed in in vitro studies: increased MDA and decreased
27 SOD and CAT in RBCs exposed to 2 (iM Pb (Ciubar et al.. 2007). decreased glutathione
28 reductase (GR) activity in human RBCs incubated with 5-18 (iM Pb (Coban et al.. 2007).
29 and decreased GSH and increased GSSG and lipid peroxidation in RBCs from healthy
30 volunteers (with no history of Pb exposure) incubated with 0.4 (iM Pb for 24-120 hours
31 (Quintanar-Escorza et al.. 2010).
Antioxidant Defense
32 In addition to the studies listed above that examined lipid peroxidation and oxidative
33 stress, there have been toxicological studies that indicate that the use of antioxidants and
34 free radical reactions is protective against Pb-induced RBC oxidative stress. Rats treated
35 with 500 ppm Pb acetate in drinking water for 15 or 30 days had a significant increase in
36 free RBC protoporphyrin and TEARS levels that was related to length of exposure and
November 2012 5-507 Draft - Do Not Cite or Quote
-------
1 blood Pb levels (Rendon-Ramirez et al. 2007). Vitamin E administration after exposure
2 to Pb significantly reduced the rat RBC TEARS levels and increased ALAD activity,
3 compared to exposure to Pb alone. Co-exposure to vitamin E and Pb simultaneously and
4 exposure to vitamin E before Pb exposure also prevented Pb-induced oxidative stress. In
5 vitro studies by Casado et al. (2007) found that Pb-induced hemolysis using blood from
6 non-occupationally exposed volunteers indicated that RBC membrane damage was
7 mediated via oxidative stress. The in vitro studies demonstrated a concentration- and
8 time-dependent formation in lipid peroxide that was inhibited with a number of
9 antioxidants, including desferrioxamine (iron chelator), trolox (chain breaking
10 antioxidant), and mannitol and Na formate (OH scavengers). Results suggested the role
11 of singlet oxygen in Pb-mediated membrane damage and hemolysis of exposed RBCs. In
12 rats exposed to 2,000 ppm Pb in drinking water for 5 weeks, MDA levels were
13 significantly increased, whereas vitamin E concentrations were significantly decreased
14 (Caylak et al., 2008). In the case of MDA, co-exposure to Pb and a number of sulfur-
15 containing antioxidants (e.g., L-methionine, N-acetylcysteine, and L-homocysteine)
16 reduced concentrations to a level not significantly different from that in controls, but
17 were significantly smaller than concentrations observed with Pb alone. Exposure to L-
18 methionine and N-acetylcysteine also reduced Pb-induced depletion of vitamin E.
5.7.2.8 Summary of Effects on RBC Survival and Function
19 In summary, Pb exposure has been shown to affect multiple hematological outcomes that
20 are related to RBC survival and function, as demonstrated in both cross-sectional
21 epidemiologic studies and toxicological studies. Pb exposure has been shown to decrease
22 RBC survival, either through direct effects on RBC membranes leading to increased
23 fragility, or through the induction of eryptosis and eventual phagocytosis by
24 macrophages. Limited evidence that Pb can negatively affect hematopoiesis is also
25 available. Consistent evidence also exists demonstrating that Pb exposure increases
26 oxidative stress in exposed adults and children, as well as in laboratory animals. The
27 epidemiologic studies demonstrating these effects are cross-sectional in design, therefore
28 there is some uncertainty regarding the direction of effects and the magnitude, timing,
29 frequency, and duration of Pb exposure that contributed to the observed observations.
30 Also, the majority of epidemiologic studies did not account for potential confounding,
31 although the effects observed in these studies are consistent with effects from studies that
32 did consider potential confounding. The coherence with effects observed in animal
33 toxicology studies supports the conclusion that Pb exposure affects both the survival and
34 function of RBCs.
November 2012 5-508 Draft - Do Not Cite or Quote
-------
5.7.3
Red Blood Cell Heme Metabolism
i
2
o
J
4
5
6
7
8
9
10
11
12
13
Pb exposure has been found to inhibit several enzymes involved in heme synthesis,
namely ALAD (a cytoplasmic enzyme catalyzing the second, rate-limiting, step of the
heme biosynthesis pathway), coproporphyrinogen oxidase (catalyses the sixth step in
heme biosynthesis converting coproporphyrinogen III into protoporphyrinogen IX), and
ferrochelatase (catalyses the terminal [eighth] step in heme synthesis converting
protoporphyrin IX into heme) (Figure 5-36). The observations of decreased Hb
(measured as total Hb, MCH, or MCHC) in occupationally-exposed adults (Ukaejiofo et
al.. 2009; Khan et al.. 2008; Patil et al.. 2006b: Karita et al.. 2005) and Pb-exposed
experimental animal models (Sharma et al.. 2010b: Baranowska-Bosiacka et al.. 2009;
Simsek et al.. 2009; Marques et al.. 2006; Lee et al.. 2005) and associations with blood
Pb levels in children (Oueirolo et al.. 2010; Shahet al.. 2010; Olivero-Verbel et al.. 2007;
Riddell et al.. 2007) are supporting lines of evidence for decreased heme synthesis due to
Pb exposure.
Cytosol
2x6-aminolevulpnic acid
1
ALA dehycfratase
(porpfiobitinagei
synthase
Porphobilinogen 1PBG)
3 | | P8G deomtnase
hydroxymethylbila
Uroporphyrin ogen
fit syinht-'tase
^> Uroporphyrinogen II
Copiopotphyrinogcn III
Uroporphyrinogen
decarboxylase
Note: Steps in the pathway potentially affected by Pb are indicated with curved arrows pointing to the affected enzyme, and the
directions of effects are represented by f and \, arrows.
Figure 5-36 Schematic representation of the enzymatic steps involved in the
heme synthetic pathway.
November 2012
5-509
Draft - Do Not Cite or Quote
-------
5.7.3.1 Red Blood Cell 5-Aminolevulinic Acid Dehydratase
1 Decreases in RBC 5-aminolevulinic acid dehydratase (ALAD) levels are strongly
2 associated with Pb exposure in humans; to such an extent that RBC ALAD activity is
3 used as a biomarker to assess Pb toxicity. Several epidemiologic studies published since
4 the 2006 Pb AQCD evaluated the relationship between Pb exposure, blood Pb levels and
5 ALAD activity in adults and children (see below). These studies were cross-sectional in
6 nature. This limits their utility in assessing the direction of effects and the magnitude,
7 timing, frequency, and duration of Pb exposure necessary to contribute to the observed
8 associations. In studies which considered potential confounders, those confounding
9 variables are listed in the test. However, potential confounding was not accounted for in
10 the majority of these studies.
11 Wang et al. (201 Of) found that, after controlling for sex, age, alcohol consumption and
12 smoking (adults only), there was also a concentration-dependent decrease in ALAD
13 activity in both children (4-13 years old) and adults (16-77 years old) (mean blood Pb
14 levels: 7.1 and 6.4 (ig/dL, respectively) in rural southwest China. Further, Wang et al.
15 (201 Of) observed that the relationship between blood Pb level and ALAD activity was
16 nonlinear and exponential, with larger decreases in ALAD activity occurring with blood
17 Pb levels >10 (ig/dL. No correlation was observed between urinary ALA levels and blood
18 Pb levels. Ahamed et al. (2006) studied male urban adolescents in India. The 39
19 adolescents were separated into two groups according to their blood Pb levels (Group 1:
20 <10 (ig/dL [mean 7.4 (ig/dL], Group 2: >10 (ig/dL [mean 13.27 (ig/dL]). Although
21 Groups 1 and 2 were similar in age (mean [SD]: 16.59 [0.91] versus 16.76 [0.90] years,
22 respectively), height, weight, and BMI (therefore not considered potential confounders),
23 Group 2 (with the higher blood Pb levels) had lower ALAD activity than did Group 1
24 (p <0.001). When all 39 adolescents were examined together, an inverse relationship was
25 found between blood Pb and ALAD activity. Similar decreases in ALAD activity were
26 observed in other populations of children from India (aged 4-12 and 1-7 years) with
27 elevated blood Pb levels (mean [SD]: 11.39 [1.39] and 21.86 [7.58] (ig/dL respectively)
28 compared to the two age ranges of the children with lower blood Pb levels (mean [SD]:
29 3.93 [0.61] and 6.89 [2.44] (ig/dL respectively) (Ahamed et al.. 2007: Ahamed et al..
30 2005). While Ahamed et al. (2005) did not address potential confounding, Ahamed et al.
31 (2007) observed decreases in ALAD activity after controlling for age, sex, and area of
32 residence. Decreases in ALAD activity were also observed in children 3-6 years of age
33 with Pb blood levels >10 (ig/dL, compared to children <10 (ig/dL (mean blood Pb
34 concentration for groups not reported) in northeastern China (Jin et al.. 2006).
35 As was seen with epidemiologic studies investigating Pb-associated deficits in
36 hematological parameters, most occupational studies investigating ALAD levels may not
November 2012 5-510 Draft - Do Not Cite or Quote
-------
1 be generalizable to the population as a whole; however, they are useful in demonstrating
2 consistent and negative effects of Pb exposure on the activity of this enzyme (Quintanar-
3 Escorzaetal.. 2007; Patil et al. 2006a: Patil et al.. 2006b: Ademuyiwa et al.. 2005b).
4 Occupationally-exposed adults had levels of inhibition of ALAD that were as great as
5 90% relative to control (Quintanar-Escorza et al.. 2007). There were few studies that
6 investigated Pb-associated decrements in ALAD levels among moderately-exposed
7 workers. Painters in India with a mean blood Pb level of 21.92 (ig/dL (mean [SD]
8 duration of exposure: 126.08 [49.53] months) had lower ALAD levels (p <0.01)
9 compared to controls whose mean blood Pb level was 3.06 (ig/dL (Mohammad et al..
10 2008). Stoleski et al. (2008) observed that workers in a Pb smelter in Macedonia (mean
11 [SD]: 16.4 [8.5] (ig/dL blood Pb; 18.8 [7.5] years employment) had lower ALAD activity
12 (p <0.001) and higher ALA levels (p <0.0005) compared to workers with no history of
13 exposure to Pb (mean [SD] blood Pb: 7.0 [5.4] (ig/dL). In automotive painters exposed to
14 Pb in Brazil (mean [SD]: 5.4 [0.4] (ig/dL blood Pb level; 133.9 [14.5] months duration of
15 exposure), the ALAD reactivation index was increased over that in controls, although
16 ALAD activity did not differ between groups (Conterato et al.. In Press). However,
17 ALAD activity was negatively correlated with blood Pb levels (r = -0.59, p <0.05) but not
18 blood Cd levels, whereas ALAD reactivation index was positively correlated with blood
19 levels of both metals (Pb: r = 0.84, p <0.05; Cd: r = 0.27, p <0.05). In a benchmark dose
20 (BMD)-based analysis (BMR = 5% using the hybrid approach and a 5% adversity cut-off
21 value), Murata et al. (2009) calculated the BMD and 95% lower confidence limit of the
22 BMD (BMDL) for decreased ALAD activity in RBCs of exposed Pb workers. The
23 calculated BMD and BMDL values for Pb blood levels of 2.7 and 2.3 (ig/dL,
24 respectively, were substantially lower than the BMDs (28.7-44.2 (ig/dL) and BMDLs
25 (19.4-29.6 (ig/dL) for decreased Hb, Hct, and RBC count in similarly exposed workers,
26 indicating decreases in ALAD activity can occur at blood Pb levels that do not decrease
27 RBC survival.
28 Decreased ALAD activity in response to Pb exposure has also been observed in
29 toxicological studies. Rats administered 500 ppm Pb acetate in drinking water for 15 or
30 30 days had decreased blood ALAD activity, which was related to duration of exposure
31 and blood Pb levels (Rendon-Ramirez et al.. 2007). Oral administration of Pb (25 mg/kg)
32 to rats once a week for 4 weeks achieved a blood Pb level of 6.5 (ig/dL, which was
33 associated with statistically significant decreases (approximately 50% lower than control
34 levels) in RBC ALAD activity (Lee et al.. 2005). Exposure of male Wistar rats to
35 5,000 ppm Pb acetate via drinking water for three weeks significantly decreased ALAD
36 activity by 72% (mean [SD]: 7.35 [0.35] versus controls: 26.14 [2.19] nmol/min/mL
37 RBCs [nanomoles of porphobilinogen (PEG) formed per minute, per 1 mL blood])
38 (Gautam and Flora. 2010).
November 2012 5-511 Draft - Do Not Cite or Quote
-------
5.7.3.2 Other Heme Metabolism Enzymes
1 The 2006 Pb AQCD (U.S. EPA. 2006b) indicated that Pb affects RBC PEG synthase
2 (Simons. 1995; Farant and Wigfield. 1990. 1987). PEG deaminase (Tomokuni and
3 Ichiba. 1990). and TF endocytosis and iron transport across membranes (Qian and
4 Morgan. 1990). all of which are directly or indirectly involved in heme synthesis.
5 Although there are no recent studies that examine the effect Pb has on the activities of
6 other heme metabolism enzymes, a number of studies investigated associations of blood
7 Pb level with concentrations of various intermediate products in the heme biosynthetic
8 pathway.
9 Pb intoxication has been shown to inhibit the function of ferrochelatase, the enzyme that
10 catalyzes the last (eighth) step in the heme biosynthetic pathway. Under normal
11 conditions, ferrochelatase incorporates ferrous iron (Fe2+) into protoporphyrin IX,
12 converting it into a heme molecule (Figure 5-36). However, Pb has been shown to inhibit
13 this insertion of Fe2+ into the protoporphyrin ring and instead, Zn is inserted into the ring
14 creating ZPP. A number of recent studies have shown that blood Pb level is significantly
15 associated with increased RBC ZPP levels in adults occupationally exposed to high levels
16 of Pb (blood Pb levels: 27-54 jig/dL) (Patil et al.. 2006b: Ademuyiwa et al.. 2005b).
17 workers exposed to moderate levels of Pb (blood Pb level = 21.92 (ig/dL) (Mohammad et
18 al.. 2008). children aged 1-21 years (blood Pb levels: 18-23 (ig/dL) (Counter et al.. 2009.
19 2008; Counter et al.. 2007). and animals exposed to 500 ppm Pb via drinking water for 15
20 or 30 days (Rendon-Ramirez et al.. 2007). Interestingly, Wang et al. (201 Of) found that in
21 children and adults living in a rural area of Southwest China, ZPP levels were negatively
22 correlated with blood Pb at blood Pb levels <10 (ig/dL and were only positively
23 correlated with blood Pb at higher blood Pb concentrations (i.e., >10 (ig/dL). The authors
24 suggested that this may be representative of ALAD activities at low blood Pb levels,
25 which contributes to lower ZPP levels. Scinicariello et al. (2007) performed a meta-
26 analysis and observed that Pb-exposed individuals who carried the ALAD2 allele had
27 slightly lower concentrations of blood ZPP levels compared to carriers of the ALAD1
28 allele (overall pooled standardized mean estimate: -0.09 [units not specified]; 95% CI:
29 -0.22, 0.03, p = 0.13).
5.7.3.3 Hematological Effects
30 In summary, Pb exposure has been shown in both cross-sectional epidemiologic studies
31 and toxicological studies to alter heme synthesis. Pb exposure has been shown to inhibit
32 the activities of two major enzymes in the heme biosynthetic pathway: ALAD and
33 ferrochelatase. Evidence for the inhibition of ALAD comes from direct measurements of
November 2012 5-512 Draft - Do Not Cite or Quote
-------
1 its activity in exposed human populations, whereas evidence for inhibition of
2 ferrochelatase comes from the observation of increased ZPP following exposure. Animal
3 toxicology and ecotoxicology studies provide evidence of coherent effects in animals.
4 The epidemiologic studies demonstrating these effects are cross-sectional in design,
5 therefore there is some uncertainty regarding the direction of effects and the magnitude,
6 timing, frequency, and duration of Pb exposure that contributed to the observed
7 observations. Also, the majority of epidemiologic studies did not account for potential
8 confounding, although the effects observed in these studies are consistent with effects
9 from studies that did account for confounding. The coherency of effects observed in
10 animal toxicology and ecotoxicology studies support the conclusion that Pb exposure
11 alters the synthesis of heme in RBCs.
5.7.4 Summary and Causal Determination
12 Recent toxicological and epidemiologic evidence substantiates evidence presented in the
13 2006 Pb AQCD that exposure to Pb affects hematological endpoints, and supports a
14 causal relationship between Pb exposure and decreased RBC survival and function and
15 altered heme synthesis. Outcomes related to decreased RBC survival and function
16 included alterations in multiple hematological parameters (e.g., Hb, Hct, PCV, MCV,
17 MCH), oxidative stress (altered antioxidant enzyme activities [SOD, CAT, GPx],
18 decreased cellular GSH, and increased lipid peroxidation), increased cytotoxicity in RBC
19 precursor cells, and mode of action endpoints such as decreased intracellular calcium
20 concentrations, decreased ATPase activity, and increased phosphatidylserine expression.
21 Outcomes related to altered heme synthesis included decreased activities of ALAD and
22 ferrochelatase, and decreased levels of Hb. The sections that follow describe the
23 evaluation of evidence for decreased red blood cell (RBC) survival and function and
24 heme synthesis, with respect to causal relationships with Pb exposure using the
25 framework described in Table II of the Preamble. The application of the key supporting
26 evidence to the causal framework is summarized in Table 5-35.
5.7.4.1 Evidence for Decreased RBC Survival and Function
27 The 2006 Pb AQCD (U.S. EPA. 2006b) reported that Pb exposure is associated with
28 multiple measures of decreased RBC survival and function. Epidemiologic evidence
29 included the observation of a 10% probability of anemia with blood Pb levels of
30 approximately 20 (ig/dL at age 1 year, and perturbed hematopoiesis in children and adults
31 at blood Pb levels below 40 (ig/dL. Oxidative stress was also identified by the
32 2006 Pb AQCD as a potential mode of action for Pb-induced effects in RBCs. A causal
November 2012 5-513 Draft - Do Not Cite or Quote
-------
1 relationship between Pb exposure and decreased RBC survival and function is strongly
2 supported by the available, recent toxicological and epidemiological data. Among the
3 strongest evidence for Pb-induced decreases in RBC survival and function is the
4 consistent observation of alterations in hematological parameters (e.g., Hb, Hct, PCV,
5 MCV, MCH), oxidative stress (altered antioxidant enzyme activities [SOD, CAT, GPx],
6 decreased cellular GSH, and increased lipid peroxidation), and increased cytotoxicity in
7 RBC precursor cells in rodents exposed to various forms of Pb via drinking water (Jang
8 et al.. 2011; Molina et al. 2011; Gautam and Flora. 2010; Baranowska-Bosiacka et al..
9 2009; Simsek et al.. 2009; Alghazal et al.. 2008b: Kharoubi et al.. 2008b: Marques et al..
10 2006). Some of these effects have been observed in toxicological studies reporting blood
11 Pb levels <10 (ig/dL, and therefore occur at blood Pb levels that are relevant to humans.
12 These effects at relevant blood Pb levels were found primarily in adult animals with Pb
13 exposure durations of 4 weeks to 9 months. Although not as representative of potential
14 human exposure pathways as exposure via drinking water, numerous toxicological
15 studies utilizing oral gavage have also observed effects on hematological parameters,
16 oxidative stress, and hematopoiesis (Sharma et al., 2010b; Celik etal.. 2005; Lee et al..
17 2005). The animal toxicological evidence for decreased RBC survival and function is
18 particularly important to the weight of evidence as it establishes clear temporality of
19 exposure to Pb and induction of effects on red blood cells.
20 Associations between increased Pb blood levels and decreased RBC survival and
21 function, are also evident in diverse populations of human adults and children. Cross-
22 sectional studies in children measuring concurrent blood Pb levels are consistent
23 regarding effects on hematological parameters (Oueirolo et al.. 2010; Shah et al.. 2010;
24 Ahamed et al.. 2007; Huo et al.. 2007; Olivero-Verbel et al.. 2007; Riddell et al.. 2007;
25 Turgut et al.. 2007; Ahamed et al.. 2006; Jin et al.. 2006; Rondo et al.. 2006).
26 Associations between altered indices of RBC oxidative stress and blood Pb levels were
27 also seen in adolescents and children (Ahamed et al.. 2008; Ahamed et al.. 2006; Jin et
28 al.. 2006). The blood Pb levels observed in cross-sectional studies of children tended to
29 be lower than those observed in adult populations (see below), with the majority of
30 studies in children (ages 5 months to 5 years old) reporting mean blood Pb levels
31 <15 (ig/dL (range: 6.9 - 21.86 (ig/dL). The difference in blood Pb levels may reflect the
32 comparatively shorter duration and lower magnitude of Pb exposure experienced by
33 children compared to adults.
34 For adult populations, the largest body of evidence consists of occupationally-exposed
35 workers in which measures of RBC survival (e.g., Hb, Hct, PCV, MCV, MCH) are
36 altered when compared with unexposed control populations in cross-sectional studies
37 (Cabaravdic et al.. 2010; Ukaeiiofo et al.. 2009; Khan et al.. 2008; Patil et al.. 2006a;
38 Patil et al.. 2006b; Karitaet al.. 2005; Conterato et al.. In Press). Only one
November 2012 5-514 Draft - Do Not Cite or Quote
-------
1 non-occupational study was found investigating the association of Pb with hematological
2 parameters; in pregnant women, concurrent blood Pb levels were found to be negatively
3 correlated with Hb concentrations. Cross-sectional studies have also observed consistent
4 increases in lipid peroxidation in occupationally-exposed adult populations (Ergurhan-
5 Ilhan et al.. 2008; Khan et al.. 2008; Mohammad et al.. 2008; Quintanar-Escorza et al..
6 2007; Patil et al.. 2006a: Patil et al.. 2006b). and have observed changes in oxidative
7 stress parameters, including lowered activities of antioxidant enzymes such as SOD, GR,
8 and CAT. Recent evidence of disrupted hematopoiesis, including the observation of
9 decreased serum EPO in occupationally-exposed adults with a mean blood Pb level of
10 6.4 (ig/dL (Sakata et al., 2007). was consistent with previous findings of decreased EPO
11 in exposed adults reported in the 2006 Pb AQCD. Although the mean blood Pb level in
12 most occupationally exposed populations was >20 (ig/dL, multiple studies observed
13 adverse effects in occupationally-exposed populations with mean blood Pb levels
14 <10 (ig/dL, including significant correlations PCV (7 (ig/dL), significant correlations
15 between RBC distribution width and MCHC (5.4 (ig/dL), and decreased EPO
16 (6.4 (ig/dL). Any differences in the effects on specific hematological and oxidative stress
17 parameters between adult populations and children may reflect differences in exposure
18 durations or patterns of exposure, although there is greater uncertainty regarding the
19 timing and duration of exposure associated with these effects in adults.
20 The evidence for Pb-associated decrements in RBC function and survival in adults and
21 children comes from cross-sectional studies measuring concurrent blood Pb levels, and
22 thus, the temporality of effects and the timing and duration of exposure associated with
23 altered RBC survival and function in RBCs is unclear. This uncertainty is greatest in
24 adults and older children as concurrent blood Pb levels also reflect higher past Pb
25 exposures. Additional limitations of the epidemiologic database include the general lack
26 of controlling for potential confounders or other possible co-exposures to contaminants
27 that can affect the hematological system. Although most studies did not control for
28 potential confounders, a few studies investigating effects in children did adjust for
29 potential confounders such as age, sex, area of residence, breastfeeding, mouthing
30 behavior, family structure, and SES-related variables, and still observed negative effects
31 on RBC survival and function. However, no studies controlled for nutritional status,
32 including iron intake. Further, while the epidemiologic database may be limited for the
33 above reasons, the findings in these studies demonstrated coherence with findings from
34 multiple toxicological studies that either reported blood Pb levels that are relevant to
35 humans, i.e., <10 (ig/dL (drinking water and gavage studies) or utilized a relevant route
36 of exposure (drinking water), and reported clear evidence for decreased RBC survival
37 and function.
November 2012 5-515 Draft - Do Not Cite or Quote
-------
1 The causal relationship between Pb exposure and decreased RBC survival and function is
2 further supported by epidemiologic and toxicological evidence characterizing mode of
3 action and biological plausibility. Pb was shown to reduce Ca2+ATPase and
4 Ca2+/Mg2+ATPase activities in RBC membranes, which leads to an increase in RBC
5 [Ca2+]l5 increased membrane fragility, and abnormal morphological changes in studies of
6 occupationally exposed adults (Quintanar-Escorza et al.. 2007) and in in vitro studies
7 (Quintanar-Escorza et al., 2010; Ciubar et al., 2007). Heul et al. (2008) observed a
8 reduction in plasma membrane Ca2+ATPase pump activity in newborn children's RBC
9 membranes in association with a concurrent group mean newborn cord blood Pb level of
10 3.54 (ig/dL. Pb exposure has also been observed to increase PS expression on RBC
11 membranes, leading to cell shrinkage, erythropoiesis, and destruction of the RBCs by
12 macrophages (Jang et al.. 2011; Ciubar etal.. 2007; Shin et al.. 2007).
13 Experimental animal studies demonstrate that Pb exposures via drinking water and
14 gavage, resulting in blood Pb levels relevant to humans, alter several hematological
15 parameters, increase measures of oxidative stress, and increase cytotoxicity in RBC
16 precursor cells. These effects were found primarily in adult animals with Pb exposure
17 durations of 4 weeks to 9 months. Support for these findings is provided by biologically
18 plausible modes of action, including decreased intracellular calcium concentrations,
19 decreased ATPase activity, and increased phosphatidylserine expression. Epidemiologic
20 studies demonstrate evidence in both adults and children that concurrent blood Pb levels
21 are associated with altered hematological endpoints and increased measures of oxidative
22 stress, and altered hematopoiesis. However, the majority of these studies are limited by
23 the lack of rigorous methodology and consideration for potential confounding. While
24 some studies in children did control for or considered potential confounding and effects
25 in adults and children are coherent with effects observed in exposed animals, there
26 remains some uncertainty regarding the evidence for altered RBC survival and function
27 in human populations. Because epidemiologic evidence is limited to associations with
28 concurrent blood Pb levels, there is uncertainty regarding the timing, duration,
29 magnitude, and frequency of Pb exposure associated with decreased RBC survival and
30 function. Collectively, the strong evidence from toxicological studies that is supported by
31 findings from mode of action and epidemiologic studies is sufficient to conclude that
32 there is a causal relationship between Pb exposures and decreased RBC survival and
33 function.
November 2012 5-516 Draft - Do Not Cite or Quote
-------
5.7.4.2 Evidence for Altered Heme Synthesis
1 The 2006 Pb AQCD (U.S. EPA. 2006b) reported that Pb exposure affects heme synthesis
2 in humans and animals through the inhibition of multiple key enzymes in the heme
3 biosynthetic pathway, including ALAD and ferrochelatase. A causal relationship between
4 Pb exposure and altered heme synthesis is strongly supported by the available
5 toxicological, ecotoxicological, and epidemiologic data (Table 5-35). The greatest weight
6 of evidence for Pb-induced alterations in heme synthesis lies primarily in the
7 toxicological and ecotoxicological literature. A small, but coherent, body of recent
8 toxicological evidence demonstrates decreased ALAD activity (Gautam and Flora. 2010;
9 Lee et al.. 2005) and ferrochelatase (Rendon-Ramirez et al.. 2007) in adult rats exposed
10 to Pb via drinking water and oral gavage for 3-4 weeks. Lee et al. (2005) observed effects
11 on ALAD activity at mean blood Pb levels of 6.5 (ig/dL after Pb administration by oral
12 gavage once per week for 4 weeks. Evidence from previous studies cited in the
13 2006 Pb AQCD consistently observed Pb-induced ALAD inhibition in multiple species,
14 including birds, primates, and humans, further supporting the causal association between
15 Pb exposure and altered heme synthesis.
16 Similar to the earlier and more recent toxicological studies that demonstrate an
17 association between Pb exposure and hematological effects in humans and laboratory
18 animals, the ecological literature has consistently reported on hematological responses in
19 aquatic and terrestrial invertebrates and vertebrates (Sections 7.3.12.5. 7.4.12.5. and
20 7.4.21.5). The most consistently observed effect in metal impacted environments is
21 decreased RBC ALAD activity. This effect has been observed across a wide range of
22 taxa, including bivalves, fish, amphibians, birds, and mammals. More limited evidence
23 exists regarding deleterious effects of Pb exposure on serum enzyme levels and white
24 blood cell counts in birds and mammals.
25 Consistent associations between increased Pb blood levels and decreased activity of
26 multiple enzymes involved in the heme synthetic pathway have also been observed in
27 diverse populations of adults and children. The strongest evidence for altered heme
28 synthesis in adults and children come from cross-sectional epidemiological studies
29 measuring concurrent blood Pb and reporting decreases in RBC ALAD levels and
30 activity (Wang et al.. 2010f; Mohammad et al.. 2008; Ahamed et al.. 2007; Quintanar-
31 Escorza et al.. 2007; Ahamed et al.. 2006; Patil et al.. 2006a; Patil et al.. 2006b;
32 Ademuyiwa et al.. 2005b; Ahamed et al.. 2005; Conterato et al.. In Press). In addition to
33 ALAD inhibition, recent studies have also shown that Pb exposure inhibits the activity of
34 ferrochelatase, leading to increased RBC ZPP levels in children and occupationally-
35 exposed adults (Counter et al.. 2009. 2008; Mohammad et al.. 2008; Counter et al.. 2007;
36 Patil et al.. 2006b; Ademuyiwa et al.. 2005b). Although the mean blood Pb levels in most
November 2012 5-517 Draft - Do Not Cite or Quote
-------
1 of the studies investigating these effects in adults and children were >20 (ig/dL, two
2 studies did observe adverse effects in populations with mean blood Pb levels <10 (ig/dL:
3 increased ALAD reactivation index in exposed painters (5.4 (ig/dL), and statistically
4 significant, positive associations between ALAD and blood Pb level in children and the
5 elderly (7.1 and 6.4 (ig/dL, respectively).
6 The cross-sectional nature of the above epidemiologic studies in adults and children, and
7 the measurement of concurrent blood Pb, introduces some uncertainty regarding the
8 temporality of effects and the timing and duration of exposure associations with altered
9 heme synthesis. Although most studies did not control for potential confounders, a few
10 studies investigating effects in children, and one study investigating effects in adults, did
11 adjust for confounders such as age, sex, urban/rural residence, height, weight, BMI,
12 smoking status, and alcohol use, and still observed negative effects on heme synthesis.
13 However, no studies controlled for nutritional status, including iron intake. Further, while
14 the epidemiologic database may be limited for the above reasons, the findings in these
15 studies demonstrated coherence with findings from multiple toxicological and
16 ecotoxicological studies.
17 The causal relationship between Pb exposure and altered heme synthesis is further
18 supported by cross-sectional studies observing decreased Hb (measured as total Hb,
19 MCH, or MCHC) in occupationally-exposed adults (Ukaejiofo et al.. 2009; Khan et al..
20 2008; Patil et al.. 2006b: Karita et al.. 2005) and in children (Queirolo etal. 2010; Shah
21 etal.. 2010; Olivero-Verbel et al.. 2007; Riddell et al.. 2007). Several recent toxicological
22 studies also observed decreased Hb levels in laboratory animals exposed to Pb (Sharma et
23 al..2010b: Baranowska-Bosiacka et al.. 2009: Simsek et al.. 2009: Marques et al.. 2006:
24 Lee et al.. 2005). Decreased Hb levels are a direct indicator of decreased heme synthesis
25 due to Pb exposure.
26 In summary, altered heme synthesis is demonstrated by a small, but coherent, body of
27 studies in adult animals reporting that Pb exposures via drinking water and gavage
28 (resulting in blood Pb levels relevant to humans) for 15 days to 9 months decreased
29 ALAD and ferrochelatase activities. Supporting this toxicological evidence is a larger
30 body of ecotoxicological studies that demonstrate decreased ALAD activity across a wide
31 range of taxa exposed to Pb. Epidemiologic studies demonstrate evidence in both adults
32 and children that concurrent blood Pb levels are associated with decreased ALAD and
33 ferrochelatase activities. However, the majority of these studies are limited by the lack of
34 rigorous methodology and consideration for potential confounding. While some studies
35 in children did control for or considered potential confounding and effects in adults and
36 children are coherent with effects observed in exposed animals, there remains some
37 uncertainty regarding the evidence for altered heme synthesis in human populations.
November 2012 5-518 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
Because epidemiologic evidence is limited to associations with concurrent blood Pb
levels, there is uncertainty regarding the timing, duration, magnitude, and frequency of
Pb exposure associated with decreased RBC survival and function. Evidence for altered
heme synthesis is also provided by a large body of toxicological and epidemiologic
studies that report decreased Hb concentrations due to Pb exposure. Collectively, the
strong evidence from toxicological and ecotoxicological studies, which is supported by
findings from epidemiologic studies, is sufficient to conclude that there is a causal
relationship between Pb exposures and altered heme synthesis.
Table 5-35 Summary of evidence supporting RBC survival and heme synthesis
causal determinations.
Attribute in
Causal
Framework3 Key Supporting Evidence13 Recent References'3
Pb Exposure or
Blood Pb Levels
Associated with
Effects0
Decreased RBC Survival and Function: Causal
Consistent Large body of studies with
toxicological consistent findings for decreased
evidence with RBC survival and function
relevant (decreased Hb, Hct, PVC,
exposures increased eryptosis, decreased
hematopoiesis, increased oxidative
stress) in rodents with relevant
concentrations of Pb and routes of
exposure
Baranowska-Bosiacka et al.
(2009).
Lee et al. (2005).
Sharma et al. (2010b).
Simsek et al. (2009),
Marques et al. (2006),
Molina et al. (2011),
Jang et al (2011),
Celik et al. (2005).
Alghazal et al. (2008b).
Kharoubi et al. (2008b).
Gautam and Flora (2010)
Rodents:
Blood Pb level:
1.7—7.1 ug/dL
Exposures:
Drinking water
50-2,000 ppm,
21—270 days as
adults
Oral gavage
25—500 mg/kg,
28—70 days
Associations Cross-sectional studies that
consistently considered potential confounding
found in factors found blood Pb-associated
multiple decreases in Hb, increases in
epidemiologic anemia prevalence, increased
studies with oxidative stress in children ages 6
relevant blood mo-5 yr
Pb levels
Association with Hb found in
children with concurrent blood Pb
levels with consideration for
potential confounding by age, sex,
mouthing behavior, anemia, dairy
product consumption, maternal
age, education, employment,
marital status, family structure,
SES-related variables
Other studies of Hb, oxidative
stress adjusted for factors such as
age, sex, birthweight, breastfeeding
history, urban/rural residence
Riddell et al. (2007),
Queirolo et al. (2010),
Ahamed et al. (2007),
Ahamed et al. (2008)
Queirolo et al. (2010)
Riddell et al. (2007),
Ahamed et al. (2008),
Ahamed et al. (2007)
Children: majority of
concurrent blood Pb
levels <15 ug/dL
November 2012
5-519
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
Recent References
Pb Exposure or
Blood Pb Levels
Associated with
Effects0
Consistent evidence in large body
of cross-sectional studies without
consideration for potential
confounding in occupationally-
exposed adults and in children of
associations of blood Pb levels with
decreases RBC survival, interferes
with hematopoiesis, and increases
oxidative stress
Karita et al. (2005). Khan et al.
(2008), Patil et al. (2006a), Patil
et al. (2006b), Ukaejiofo et al.
(2009), Conterato et al. (]n
Press), Cabaravdic et al. (2010),
Ergurhan-llhan et al. (2008).
Mohammad et al. (2008).
Quintanar-Escorza et al. (2007).
Sakata et al. (2007). Riddell et al.
(2007), Queirolo et al. (2010),
Olivero-Verbel et al. (2007),
Ahamed et al. (2006), Ahamed et
al. (2007), Ahamed et al. (2008),
Turgut et al. (2007). Huo et al.
(2007). Shah et al. (2010).
Rondo et al. (2006). Jin et al.
(2006)
Adults (occupational
exposures): majority of
blood Pb levels
>20 ug/dL, some
studies observed
effects in the range of
5-7 ug/dl_
Evidence
clearly
describes
Mode of Action
Altered RBC
membrane
ion transport
Phosphatidyl
serine (PS)
expression
Evidence of increased [Ca2+]i and
decreased Ca2+/Mg2+ATPase
activity in the RBCs of exposed
workers. [Ca2+]i levels highly
correlated with blood Pb even
among unexposed controls.
[Ca2+]i levels increased in RBCs
from healthy volunteers when
exposed in vitro to Pb
[Ca2+]i associated with increased
RBC fragility and alterations in
RBC morphology
Consistent evidence from in vivo
and in vitro studies that Pb
exposure increases PS expression
on RBC membranes via modulation
of [Ca2+]i concentrations. Increased
PS expression leads to eryptosis
and phagocytosis by macrophages
See Section 5.7.2.2
November 2012
5-520
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Key Supporting Evidence
Recent References
Pb Exposure or
Blood Pb Levels
Associated with
Effects0
Altered Heme Synthesis: Causal
Consistent A small, but coherent toxicology
toxicological database indicates decreased
evidence with heme synthesis in rodents with
relevant relevant Pb concentrations and
exposures routes of exposure
Rendon-Ramirez et al. (2007),
Lee et al. (2005),
Gautam and Flora (2010)
Blood Pb levels
6.5 ug/dL
Exposures: 500-
5,000 ppm Drinking
water, 15-30 days as
adults
Consistent Pb-induced decreased ALAD
ecotoxicologica activity observed across many taxa
I evidence (bivalves, fish, amphibians, birds,
and mammals) in multiple
ecotoxicity studies
Birds:
Berglund et al. (2010).
Gomez-Ramirez et al. (2011).
Hansen et al. (2011 a),
Martinez-Haro et al. (2011)
Freshwater Invertebrates:
Aisemberg et al. (2005)
Fish:
Schmitt et al. (2005),
Schmitt et al. (2007b),
Heier et al. (2009).
Bivalves:
Kalman et al. (2008).
Company et al. (2011).
Birds:
6->100ug/dL
Freshwater
Invertebrates
(48-h exposure in
aquaria)
0.2-300 ug/g wet
tissue
Fish:
6-14 ug/g
(gill or liver
concentrations)
Bivalves:
0.38-3.50 ug/g dry
weight
Associations
found in
epidemiologic
studies with
relevant blood
Pb levels
Cross-sectional studies that Ahamed et al. (2006),
considered potential confounding Ahamed et al. (2007)
by age, sex, urban/rural residence,
height, weight, BMI found
associations with lower ALAD and
ferrochelatase activities in children.
Concurrent blood Pb level Wang et al. (201 Of)
associated with lower ALAD and
higher ZPP in adults with
consideration for potential
confounding by age, sex, smoking
status, and alcohol use.
Adults (occupational
exposure) and
children: Majority of
concurrent blood Pb
levels >20 ug/dL,
Two studies observed
associations in the
range of concurrent
blood Pb levels 5—
7 ug/dL.
T~ty' *" •— •
Associations found in several
studies, mostly in occupationally-
exposed adults, that did not
consider potential confounding
Children:
Ahamed et al. (2005)
Occupational:
Ademuyiwa et al. (2005b).
Mohammad et al. (2008).
Patil et al. (2006a). (2006b).
Quintanar-Escorza et al. (2007),
Conterato et al. (In Press)
November 2012
5-521
Draft - Do Not Cite or Quote
-------
Attribute in
Causal
Framework3
Epidemiologic
and
toxicological
evidence for
altered heme
synthesis
supported by
consistent
evidence of
decreased Hb,
a direct marker
for decreased
heme
synthesis.
Key Supporting Evidence13
Consistent evidence in animals for
decreases in Hb with relevant Pb
exposures.
Association found in children with
concurrent blood Pb levels with
consideration for potential
confounding by age, sex, mouthing
behavior, anemia, dairy product
consumption, maternal age,
Recent References'3
Animals:
Baranowska-Bosiacka et al.
(2009),
Lee et al. (2005),
Marques et al. (2006),
Sharma et al. (2010b),
Simsek et al. (2009)
Queirolo et al. (2010)
Pb Exposure or
Blood Pb Levels
Associated with
Effects0
Adult animals: Blood
Pb levels 1.7-
7.1 ug/dL after 15 day-
9 month Pb exposure
Children: Majority of
concurrent blood Pb
<15ug/dL
status, family structure, SES-
related variables
Other studies in children had
limited or no consideration for
potential confounding.
Associations found in adults and,
as well as coherent findings in
animal toxicological studies, for
decreased Hb.
Shah et al. (2010),
Olivero-Verbel et al. (2007),
Riddell et al. (2007)
Adults: Karita et al. (2005).
Khan et al. (2008).
Patil et al. (2006b).
Ukaejiofo et al. (2009)
Adults (occupational
exposure): Majority of
blood Pb >20 ug/dl_
aDescribed in detail in Table II of the Preamble.
Describes the key evidence and references contributing most heavily to causal determination. Also noted are the
sections where full body of evidence is described.
cDescribes the blood Pb levels in humans with which the evidence is substantiated and blood Pb levels in animals
most relevant to humans.
November 2012
5-522
Draft - Do Not Cite or Quote
-------
5.8 Reproductive and Developmental Effects
1 The effect of Pb on reproductive and developmental outcomes has been of interest for
2 years, starting in cohorts of occupationally-exposed individuals. More recently,
3 researchers have begun to focus on reproductive and developmental effects in
4 populations without occupational exposures, with more environmentally-relevant levels
5 of Pb exposure. The toxicological and epidemiologic literature on reproductive effects of
6 Pb include research on female and male reproductive function such as hormone levels,
7 fertility, spontaneous abortions, effects on sperm, estrus, and effects on reproductive
8 organs. Evaluation of effects on the developing organism includes effects on puberty,
9 postnatal growth, and effects on the development of the teeth, sensory organs, and other
10 systems. Research on birth outcomes includes birth defects, infant mortality, preterm
11 birth, and low birth weight. A few studies of pregnancy-induced hypertension and
12 eclampsia have been conducted and are reported on in the section on hypertension
13 (Section 5.4.2.1). Briefly, the relatively small number of studies found consistently
14 positive associations between blood Pb levels and pregnancy-induced hypertension.
15 Biomarkers of Pb exposure, including blood Pb and bone Pb, are used in the
16 epidemiologic studies reviewed in this section. Bone Pb typically indicates cumulative
17 exposure to Pb, whereas, blood Pb may indicate more recent exposure. However, Pb can
18 also be remobilized from the bone during times of active bone remodeling, such as
19 pregnancy or lactation. Toxicological studies typically report exposure using blood Pb.
20 More detailed discussion of these measures and Pb transfer via umbilical cord blood Pb
21 across the placenta, and via lactation is given in Section 4.2.2.4 on Pb Toxicokinetics.
22 Overall, the recent literature on reproductive effects of Pb exposure continues to support
23 associations reported in earlier Pb AQCDs between Pb exposure and effects on various
24 parameters of sperm (function, motility, count, integrity, histology). The toxicological
25 and epidemiologic literature of developmental effects of Pb exposure also indicates that
26 Pb exposure is associated with delayed onset of puberty in both males and females.
27 Associations between Pb exposure and other reproductive and developmental effects
28 have less consistent findings. The recent information from epidemiologic and
29 toxicological studies is integrated with conclusions from previous Pb AQCDs below.
5.8.1 Effects on Development
30 The 2006 Pb AQCD (U.S. EPA. 2006b) reported Pb-associated developmental effects on
31 teeth, sensory organs, the GI system, the liver, and postnatal growth as well as delayed
32 puberty (U.S. EPA, 2006b). There was recognition that Pb is transferred across the
33 placenta and through the breast milk, contributing to exposure during development. The
November 2012 5-523 Draft - Do Not Cite or Quote
-------
1 2006 Pb AQCD reported delayed puberty in both male and female populations in animal
2 toxicology studies showing associations with dam blood Pb levels of ~40 (ig/dL and pup
3 blood Pb levels of 26 (ig/dL. The research reported in this ISA continues to find delayed
4 puberty with Pb exposure at even lower Pb doses in animal toxicology studies as is
5 detailed below. Mechanistic understanding of delayed puberty is also reported in this
6 ISA. Lower dose Pb exposure studies in animal toxicology are also reported in studies of
7 sensory organ function and postnatal growth in this ISA. Studies included in this ISA
8 expand upon evidence reported in previous Pb AQCDs for the aforementioned systems
9 sensitive to developmental effects with recent studies showing effects at lower doses of
10 Pb.
5.8.1.1 Effects on Puberty among Females
11 Recent toxicological studies of rodents have examined the effects of Pb on pubertal and
12 reproductive organ development and on biomarkers of pubertal development among
13 females. There have also been recent epidemiologic studies examining associations
14 between blood Pb levels and onset of puberty among girls, which are summarized in
15 Table 5-36 and in the text below. All of the epidemiologic studies examined concurrently
16 measured blood Pb and puberty and are reported below. Additionally, while there was a
17 longitudinal investigation by Naicker et al. (2010). who followed girls to determine their
18 age of menarche, blood Pb levels were measured once at 13 years of age.
November 2012 5-524 Draft - Do Not Cite or Quote
-------
Table 5-36 Summary of recent epidemiologic studies of associations between Pb levels and puberty for
females.
Reference
(Studies are
presented
in order of
first
appearance
in the text of Study Location
this section) and Years
Wu et al. U.S.A.
(2003b) 1988-1994
Selevanetal. U.S.A.
(2003) 1988-1994
Out- Methodological
come Study Population Details
Tanner Girls ages 8-16 from the Cross-sectional
staging and NHANES III study study using
age at logistic regression
menarche with weighting
N=1706
Tanner Girls ages 8-18 from the Cross-sectional
staging and NHANES III study study using ordinal
age at N =600 logistic regression
menarche ' ,NHwhlle ° and Cox
NNHbiack-805 proportional
NMexican-American=781 hazards
Mean Pb
Pb (SD)
Biomarker in ug/dL
Blood Pb 2.5 (2.2)
Weighted
proportion of
the sample
with blood Pb
5.0-21 .7: 5.9%
Blood Pb Geometric
mean
NHWhites: 1.4
NHBIacks: 2.1
Mexican-
Americans: 1.7
Adjusted
Effect
Estimates
OR (95% Cl)
Breast
development
0.7-2.0 ug/dL:
1 .00 (Ref)
2.1-4.9 ug/dL:
1.51 (0.90,2.53)
5.0-21 .7 ug/dL:
1.20(0.51, 2.85)
Pubic hair
development
0.7-2.0 ug/dL:
1 .00 (Ref)
2.1-4.9 ug/dL:
0.48 (0.25, 0.92)
5.0-21 .7 ug/dL:
0.27 (0.08, 0.93)
Menarche
0.7-2.0 ug/dL:
1.00 (Ref)
2.1-4.9 ug/dL:
0.42(0.18,0.97)
5.0-21. 7 ug/dL:
0.19(0.08, 0.43)
OR (95% Cl)
Breast
development
NH Whites:
1 ug/dL: 1.00
Potential
Confounders
Adjusted for in
Analysis
Race/ethnicity,
age, family size,
residence,
poverty income,
ratio, BMI
For breast
development:
Age, age2, height,
BMI, family
income, ever
smoked>100
cigarettes, dietary
November 2012
5-525
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented
in order of
first
appearance
in the text of Study Location
this section) and Years
Mean Pb
Out- Methodological Pb (SD)
come Study Population Details Biomarker in ug/dL
Blood Pb
levels>5|jg/dL:
NHWhites:
2.7%
NHBIacks:
11.6%
Mexican-
Americans:
12.8%
Blood Pb
levels
>10|jg/dL:
NHWhites:
0.3%
NHBIacks:
1.6%
Mexican-
Americans:
2.3%
Adjusted
Effect
Estimates
(Ref)
3 |jg/dL: 0.82
(0.47, 1 .42)
NH Blacks:
1 |jg/dL: 1 .00
(Ref)
3 |jg/dL: 0.64
(0.42, 0.97)
Mexican
Americans:
1 |jg/dL: 1 .00
(Ref)
3 |jg/dL: 0.76
(0.63,0.91)
Pubic hair
development
NH Whites:
1 ug/dL: 1 .00
(Ref)
3 ug/dL: 0.75
(0.37,1.51)
NH Blacks:
1 ug/dL: 1 .00
/D _ f\
(Ref)
3 ug/dL: 0.62
(0.41 , 0.96)
Mexican
Americans:
1 ug/dL: 1 .00
(Ref)
3 ug/dL: 0.70
(0.54,0.91)
HR (95% Cl)
'included only
girls 8-1 6
Age at menarche
NH Whites:
1 ug/dL: 1 .00
Potential
Confounders
Adjusted for in
Analysis
Fe, dietary
vitamin C, dietary
Ca .
For pubic hair
development:
Age, age2, height,
family income,
ever smoked>100
cigarettes,
anemia, dietary
Fe, dietary
vitamin C
For age at
menarche:
Height, BMI,
family income,
anemia, dietary
Ca2+.
Considered in all
models: age,
smoking, dietary
Ca , dietary Fe,
dietary vitamin C,
dietary total fat,
anemia, urban
residence, family
income
November 2012
5-526
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented
in order of _ . .. .
first Potential
appearance Mean Pb Adjusted n(:onf?"Jnder?
in the text of Study Location Out- Methodological Pb (SD) Effect Adjusted for in
this section) and Years come Study Population Details Biomarker in ug/dL Estimates Analysis
(Ref)
3 |jg/dL: 0.74
(0.55, 1.002)
NH Blacks:
1 |jg/dL: 1.00
(Ref)
3 |jg/dL: 0.78
(0.63, 0.98)
Mexican
Americans:
1 |jg/dL: 1.00
(Ref)
3 |jg/dL: 0.90
(0.73,1.11)
November 2012 5-527 Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented
in order of
first
appearance
in the text of Study Location
this section) and Years
Gollenberg et U.S.A.
al. (2010) 1988-1994
Potential
Mean Pb Adjusted n(:onf?"Jnder?
Out- Methodological Pb (SD) Effect Adjusted for in
come Study Population Details Biomarker in ug/dL Estimates Analysis
Luteinizing Girls ages 6-11 from the Cross-sectional Blood Pb Median 2.5 OR (95% Cl) for Age,
hormone NHANES III study study using survey (range 0.07, exceeding race/ethnicity,
(LH) and logistic regression 29.4) pubertal inhibin B BMI, census
inhibin B blood Pb cutoff (>35pg/mL) region, poverty -
>10ug/dL:5% <1 ug/dL: 1.00 income ratio
(Ref)
1-4.9ug/dL: 0.38
(0.12, 1.15)
> 5 ug/dL: 0.26
(0.11,0.60)
OR (95% Cl) for
exceeding
pubertal LH cutoff
(>0.4mlU/mL)
<1 ug/dL: 1.00
(Ref)
1 -4.9 ug/dL: 0.98
(0.48, 1.99)
> 5 ug/dL: 0.83
(0.37, 1.87)
*Note: a
sensitivity
analysis including
only those with
blood Pb
<10 ug/dLhad
similar results but
ORs were slightly
attenuated
November 2012
5-528
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented
in order of
first
appearance
in the text of Study Location Out-
this section) and Years come
Denham et Akwesasne Mohawk Age at
a 1. (2005) Nation (boundaries of menarche
New York, Ontario,
and Quebec
NS
Potential
Mean Pb Adjusted ,£?nfou"ders
Methodological Pb (SD) Effect Adjusted for in
Study Population Details Biomarker in ug/dL Estimates Analysis
10- to 16.9-yr-old girls in Cross-sectional Blood Pb 0.49(0.905) Coefficients for Age, SES, BMI
the Akwesasne study using probit binary logistic
community and logistic . regression
regression Median: 1 .2 predicting
menarche with Pb
N=138 centered at the
mean:
log blood Pb
-1 .29 (p-value
0.01)
log blood Pb -
squared: -1.01 (p-
value 0.08)
Non-linear
relationship
observed and Pb
below the mean
did not appear to
affect the odds of
menarche.
Increasing blood
Pb from 0.49 to
0.98 ug/dL
decreased the
odds of menarche
attainment by
72%
November 2012
5-529
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented
in order of
first
appearance
in the text of Study Location
this section) and Years
Naicker et al. Johannesburg/Soweto,
(2010) South Africa
Born in 1990
Den Hond et Flanders
al. (2011) 2003-2004
Out-
come
Self-
reported
Tanner
staging at
age 13 and
age at
menarche
Tanner
staging, age
at
menarche,
regular
menses
Study Population
Girls of blacker mixed
ancestry who were
enrolled in the Birth to
Twenty (Bt20) cohort
(born in 1990) that lived
in Johannesburg/Soweto
for at least 6 mo after
birth
N=682
Girls ages 14 and 15, in
their 3rd year of
secondary education and
living in the same study
areas for at least 5 years
N=792
Mean Pb
Methodological Pb (SD)
Details Biomarker in ug/dL
Cross-sectional Blood Pb at 4.9 (1 .9)
and longitudinal 1 Syr of age blood Pb
study using ,eve|s
logistic regression >10ug/dL
1%
Cross-sectional Blood Pb Median: 1 .81
study using 10th
logistic regression percentile:
0.88
90th
percentile:
3.81
Adjusted
Effect
Estimates
OR (95% Cl)
Delay in breast
development at
age 13
<5 ug/dL: 1 .00
(Ref)
> 5 ug/dL: 2.34
(1 .45, 3.79)
Delay in pubic
hair development
at age 13
<5 ug/dL: 1 .00
(Ref)
> 5 ug/dL: 1 .81
(1.15, 2.84)
Delay in
attainment of
menarche at age
13
<5 ug/dL: 1 .00
(Ref)
> 5 ug/dL: 2.01
(1 .38, 2.94)
OR (95% Cl) for
pubic hair
development with
doubling of
exposure
0.65 (0.45, 0.93)
'Association was
no longer
statistically
significant when
PCB marker
included in the
model
No association
between Pb and
breast
development
(results not given)
Potential
Confounders
Adjusted for in
Analysis
BMI
Age, BMI,
smoking, oral
contraceptive use
Considered but
did not include:
food intake and
lifestyle
parameters
November 2012
5-530
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented
in order of
first
appearance
in the text of Study Location
this section) and Years
Tomoum et Cairo, Egypt
al. (2010) 2007
Wolff et al. New York City, NY
(2008): Wolf 1 gg^ 997
and Daley
(2007)
Out-
come
Hormones
and pubertal
development
Pubertal
stages
defined
using
standard
drawings
Study Population
Healthy children aged
10-13 yr; seeking
treatment for minor
health problems and
living in one of two
designated areas (one
with high-risk for Pb
contamination and one
with no Pb source)
N=20
9-yr old girls from the
study hospital and
nearby pediatric offices
N=192
Methodological
Details
Cross-sectional
study using Chi-
square
Cross-sectional
study using
Poisson
multivariate
regression with
robust error
variance
Mean Pb
Pb (SD)
Biomarker in ug/dL
Blood Pb NS for girls
only
(combined
with boys in
the study the
mean was
9.46 [3.08])
Blood Pb Median: 2.4
Adjusted
Effect
Estimates
Breast
Development
<10 ug/dL:
Stage 2: 36.4%
Stage 3: 63.6%
> 10 ug/dL:
Stage 2: 100%
Stage 3: 0%
Chi-square p-
value<0.01
Pubic Hair
Development
<10 ug/dL:
Stage 2: 36.4%
Stage 3: 63.6%
>10ug/dL:
Stage 2: 77.8%
Stage 3: 22.2%
Chi-square p-
value>0.05
'Quantitative
results for
hormones not
provided
PR (95% Cl) (unit
not given,
assume results
are per 1 ug/dL)
Breast stage:
1.01 (0.79, 1.30)
Pubic hair stage:
1.25(0.83, 1.88)
Potential
Confounders
Adjusted for in
Analysis
None
For breast
development:
Age, BMI, race
For hair stage:
Height, private
clinic, race
November 2012
5-531
Draft - Do Not Cite or Quote
-------
1 Multiple studies have been performed examining blood Pb levels and puberty using
2 NHANES III data (Gollenbera etal.. 2010: Selevan etal. 2003: Wu et al.. 2003b). A
3 study that included girls aged 8-16 years reported an association between increased blood
4 Pb and delayed attainment of menarche and pubic hair development, but not for breast
5 development (Wu et al., 2003b). The associations were observed even at blood Pb levels
6 of 2.1-4.9 (ig/dL compared to girls with blood Pb levels <2.1 (ig/dL. Another NHANES
7 III study included girls 8-18 years of age and reported the results stratified by race
8 (Selevan et al.. 2003). This study also included many important potential confounders,
9 such as nutritional information. Higher blood Pb levels were associated with lower
10 Tanner stage of breast and pubic hair development and later age at menarche among
11 African Americans and with lower stage of breast and pubic hair development among
12 Mexican Americans. For whites, the associations were in the same directions, but none
13 reached statistical significance. In a study of girls aged 6-11 years old from NHANES III
14 data, higher blood Pb levels were associated with lower inhibin B, a protein that inhibits
15 FSH production, but no association was observed for LH. (Gollenberg et al.. 2010). The
16 inverse association between blood Pb and inhibin B was greater among girls with iron
17 deficiency compared to those with high Pb but sufficient iron levels. Inhibin B and LH
18 were chosen for this study because, as the authors indicated, these hormones are,
19 "believed to be relevant for younger girls... near the onset of puberty and... serve as
20 markers for hypothalamic-pituitary-gonadal functioning."
21 A study of girls aged 10-16.9 years of age in the Akwesasne Mohawk Nation reported a
22 nonlinear association between higher blood Pb and greater age at menarche (Denham et
23 al., 2005). No association was observed below blood Pb of 0.49 (ig/dL in a nonlinear
24 model of the Pb-menarche relationship. A study conducted in South Africa reported an
25 association between increased blood Pb levels and older age at first menarche and
26 pubertal development (Naicker et al.. 2010). Another study reporting on girls with low
27 blood Pb concentrations observed an association between higher blood Pb and less pubic
28 hair development but not breast development (DenHond E. 2011). The association was
29 no longer statistically significant when a marker for polychlorinated biphenyl exposure
30 was included in the model. A study among girls aged 10-13 years (median: 12 years)
31 reported lower levels of FSH and LH levels in the group with blood Pb of at least
32 10 (ig/dL compared to the group with blood Pb less than 10 (ig/dL (Tomoum et al..
33 2010). In addition, there were some indications of lower Tanner stages of breast
34 development associated with Pb levels of at least 10 (ig/dL, but this relationship was not
35 present for stages of pubic hair development and there was no control for potential
36 confounders. A study performed in NYC among 9-year old girls reported no association
37 between Pb levels and pubertal development (Wolff etal.. 2008). but this age group may
38 be too young to study when investigating delayed puberty as the outcome.
November 2012 5-532 Draft - Do Not Cite or Quote
-------
Puberty; Neonate/adult; Mouse;
Female; lavicoli et al. (2006)
Neurotransmitter; Adult; Mouse;
Both; Leasure et al. (2008)
Physical development; Adult; Mouse;
Male; Leasure et al. (2008)
Eye; Ad lilt; Rat; Both; Fox et al. (2008)
Redox-oxidative stress; Adult; Rat;
Male; Nava-Hernandez et al. (2009)
Sperm; Adult; Rabbit; Male; Moorman
etal. (1998)
Neurobehavioral; Adult; Mouse; Male;
Leasure et al. (2008)
Hematological parameters; Adult; Rat;
Both; Teijon et al. (2006)
Histology; Adult; Rat; Both; Teijon et
al. (2006)
Biomarkers; Adult; Rat; Both; Teijon et
al, (2006)
Physical development; Adult; Rat;
Both; Teijon et al. (2006)
J
^
<
o Highest Concentration
* Lowest Cone, with Response
A Highest Cone, with No Response
o Lowest Concentration
«o
i An
fc— *
10 100
Blood Lead Level (|jg/dL)
1000
Note: This figure illustrates reproductive and developmental effects associated with Pb exposure in studies that examined multiple
exposure concentrations. Dosimetric representation reported by blood Pb level. (Studies are described in Table 5-37).
Figure 5-37 Toxicological concentration-response array for reproductive and
developmental effects of Pb.
November 2012
5-533
Draft - Do Not Cite or Quote
-------
Table 5-37 Toxicological concentration-response array summary for
reproductive and developmental effects of Pb presented in
Figure 5-37.
Reference
lavicoli et al. (2006a)
Leasure et al. (2008)
Fox et al. (2008)
Nava-Hernandez et al. (2009)
Moorman et al. (1998)
Teijon et al. (2006)
Fox et al. (2008)
Nava-Hernandez et al. (2009)
Moorman et al. (1998)
Teijon et al. (2006)
Fox et al. (2008)
Blood Pb level with Effect (|jg/dL)
8&13
10&42
10, 24 & 42
10&42
12
19.5
25-130
40 & 100
40 & 100
40 & 100
100
12
19.5
25-130
40 & 100
40 & 100
40 & 100
100
12
Altered Outcome
Delayed onset female puberty
Neurotransmitter, Dopamine homeostasis
Physical Development, Adult obesity
(males)
Aberrant response to amphetamine
Retinal aberrations
Sperm affected via redox imbalance
Semen quality affected
Hematology
Histology-Offspring renal & hepatic
Biomarker-Offspring renal function
Physical development: birth weight
Retinal aberrations
Sperm affected via redox imbalance
Semen quality affected
Hematology
Histology-Offspring renal & hepatic
Biomarker-Offspring renal function
Physical development: birth weight
Retinal aberrations
1
2
o
3
4
5
6
7
8
9
10
11
12
13
Earlier studies showed that prenatal and lactational exposures to Pb can cause a delay in
the onset of female puberty in rodents. Recent studies corroborate these findings and
show that puberty onset is one of the more sensitive markers of effects of Pb exposure as
is demonstrated in the exposure response array (Figure 5-37 and Table 5-37; including
outcomes described in sections that follow). Dumitrescu et al. (2008c) exposed adult
Wistar female rats to varying doses of Pb acetate (50-150 ppb) in drinking water for
3 months before mating and during pregnancy. Vaginal opening, an indicator of sexual
maturation, was statistically significantly delayed in pups from all Pb treated groups
when compared to pups from non-treated dams. The age at vaginal opening in female
pups from the Pb treated groups increased, in a concentration-dependent manner, from
39 days to 43-47 days. The authors also observed a correlation between body weight and
age at vaginal opening meaning that as body weight decreased the age at vaginal opening
increased. This effect also exhibited a concentration-dependent relationship.
November 2012
5-534
Draft - Do Not Cite or Quote
-------
1 In another recent study, lavicoli et al. (2006a) reported a statistically significant delay in
2 several indicators of sexual maturity in offspring (Swiss mice, FI generation) born to
3 dams that ingested 3.5-40 ppm Pb in their daily diet; offspring had continuous dietary
4 exposure until the termination of the experiment at puberty. Maternal ingestion of Pb at
5 the various doses resulted in female pup blood Pb levels of 3.5-13 (ig/dL. For all diet
6 groups in this range (3.5-13 (ig/dL), there was a delay in age at vaginal opening, age of
7 first estrus, age of vaginal plug formation, and age of first parturition when compared to
8 the group at background Pb concentration (2 (ig/dL). A novel finding in the lavicoli study
9 was that very low dose Pb (blood Pb of 0.7 (ig/dL, food concentration of 0.02 ppm
10 continuous through gestation, lactation and until the termination of the experiment)
11 induced statistically significant acceleration of markers of sexual maturation in female
12 offspring versus background Pb level animals (blood Pb of 2 (ig/dL). There were
13 statistically significant increases in time of vaginal opening (30% earlier), first estrous,
14 first vaginal plug formation, and first parturition at the very low Pb exposure versus
15 2 (ig/dL animals. Thus, the timing of puberty is delayed in a concentration-dependent
16 fashion with very low dose Pb having a statistically significant earlier onset of puberty
17 than the background Pb animals (2 (ig/dL). Also, the animals exposed to the higher dose
18 of Pb (blood Pb up to 13 (ig/dL) had statistically significant delays in onset of puberty
19 when compared to the other dose groups.
20 In addition, Pb-induced shifts in sexual maturity were observed in the subsequent
21 generation (F2 generation) across that dose range. These F2 animals continued to be
22 exposed to same concentrations of Pb over multiple generations through the diet. Results
23 in the F2 generation closely resembled those of the FI generation, as both generations
24 received Pb exposure. The authors concluded that a modest elevation in blood Pb level
25 (13 (ig/dL) over background (2-3 (ig/dL) can result in a profound delay in the onset of
26 puberty (15-20%). In the F2 generation, reduction in blood Pb (0.7 (ig/dL) below
27 background (2-3 (ig/dL) was associated with an earlier onset of sexual maturity (30%
28 increase) above background.
29 In the 2006 Pb AQCD (U.S. EPA. 2006b). it was reported that a statistically significant
30 reduction in the circulating levels of insulin-like growth factor 1 (IGF-1), LH, and
31 estradiol (E2) was associated with Pb-induced delayed puberty in Fischer 344 pups.
32 Subsequently, Pine et al. (2006) evaluated whether IGF-1 replacement could reverse the
33 effects of Pb on delayed female puberty onset. The authors reported that offspring from
34 dams exposed to Pb during gestation and lactation (daily oral gavage of dam with 1.0 mL
35 solution of Pb acetate 12 mg/mL; mean maternal blood Pb level 40 (ig/dL) exhibited a
36 marked increase in LH and luteinizing hormone releasing hormone (LHRH) secretion
37 after IGF-1 administration (200 ng3/(iL i.p. injection twice daily from PND23 until the
38 appearance of vaginal opening which appears in control animals at -PND40) resulting in
November 2012 5-535 Draft - Do Not Cite or Quote
-------
1 restored timing of vaginal to that of control animals. It should be noted that, IGF-1
2 replacement in Pb-exposed animals did not cause advanced puberty over non-Pb-exposed
3 controls. The results of this study provide support to the theory that Pb-induced delayed
4 onset of puberty may be due to disruption of pulsatile release of sex hormones (U.S.
5 EPA. 2006b) and not necessarily due to a direct toxic effect on the hypothalamic-
6 pituitary-gonadal axis (Salawu et al.. 2009). and IGF-1 may play a prominent role in the
7 process.
8 In summary, epidemiologic studies consistently show an association between higher
9 concurrent blood Pb and delayed pubertal development in girls. This association is
10 apparent even at low blood Pb levels. Most of the studies had good sample sizes and
11 controlled for some potential confounders. Nutritional information was rarely controlled
12 for although this could be important, especially in populations where malnutrition is
13 prevalent. These epidemiologic studies are cross-sectional, which does not allow for the
14 study of temporality between Pb levels and pubertal onset nor does it consider the
15 influence of past blood Pb levels. New evidence from the toxicology literature continues
16 to indicate Pb-induced delays in the onset of puberty. Further, the biological plausibility
17 of delayed puberty is expanded with the toxicological literature that shows this pathway
18 is mediated by IGF-1.
5.8.1.2 Effects on Puberty among Males
19 Recent epidemiologic studies examining the association between blood Pb and onset of
20 puberty in males are summarized in Table 5-38. The majority of studies used concurrent
21 measures of blood Pb and puberty (DenHond E. 2011; Tomoum et al.. 2010; Hauser et
22 al.. 2008). but Williams et al. (2010) performed a longitudinal analysis of blood Pb levels
23 measured at ages 8-9 years and pubertal onset, following the participants for 3 years.
24 Little epidemiologic information was available regarding pubertal onset in the
25 2006 Pb AQCD (U.S. EPA. 2006b).
November 2012 5-536 Draft - Do Not Cite or Quote
-------
Table 5-38 Summary of recent epidemiologic studies of associations between Pb levels and puberty for males.
Reference
(Studies are
presented in
order of first
appearance in
the text of this Study
section) Location Outcome
Mauser etal. Chapaevsk, Pubertal
(2008) Russia stages
2003-2005 defined using
standard
drawings
Pb Adjusted _ . .. , _ ,
Methodological Bio- Mean Pb (SD) Effect Potential Confounders
Study Population Details marker in ug/dL Estimates Adjusted for in Analysis
Healthy boys aged 8-9 Cross-sectional Blood Median: 3 (IQR OR (95% Cl) Gestational age, height, BMI,
study using Pb 2-5) Pubertal onset age at exam
M-489 multivariable blood Pb based on
IOglStlCregreSSIOn >10ug/dL3% testicular volume considered but did not
<5 ug/dL: 1.00 include: parental education,
(Ref) household income
> 5 ug/dL: 0.83
(0.43, 1 .59)
*after adjustment
for
macronutrients,
the OR (95% Cl)
became 0.66
(0.44, 1 .00)
Genital
development
<5 ug/dL: 1.00
(Ref)
> 5 ug/dL: 0.57
(0.34, 0.95)
*after adjustment
for
macronutrients,
the OR (95% Cl)
became 0.52
(0.31 , 0.88)
Pubic hair
development
<5 ug/dL: 1.00
(Ref)
> 5 ug/dL: 0.74
(0.34, 1 .60)
November 2012
5-537
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this Study
section) Location Outcome
Williams et al. Chapaevsk, Pubertal
(2010) Russia stages
2003-2008 defined using
standard
drawings
Tomoum et al. Cairo, Hormones
(2010) Egypt and pubertal
2007 development
Study Population
Healthy boys aged 8-9
at enrollment who had
3 annual follow-up
evaluations
N=481
Healthy children aged
10-1 3 seeking
treatment for minor
health problems and
living in one of two
designated areas (one
with high-risk for Pb
contamination and
one with no Pb
source)
N=21
Pb
Methodological Bio- Mean Pb (SD)
Details marker in ug/dL
Longitudinal cohort Blood Median: 3 (IQR
using Cox Pb at 2-5)
proportional ages B,ood pb |eve|
hazards 8-9 >lOug/dL:3%
Cross-sectional Blood NS for boys only
study using Chi- Pb (combined with
square girls in the study
the mean was
9.46 [3.08])
Adjusted
Effect
Estimates
HR (95% Cl)
Pubertal onset
based on
testicular volume
<5 ug/dL: 1 .00
(Ref)
> 5 ug/dL: 0.73
(0.55, 0.97)
Genital
development
<5 ug/dL: 1.00
CRpn
\,na\)
> 5 ug/dL: 0.76
(0.59, 0.98)
Pubic hair
development
<5 ug/dL: 1.00
(Ref)
> 5 ug/dL: 0.69
(0.44, 1 .07)
Testicular size
<10 ug/dL:
Stage 1 : 0%
Stage 2: 44.4%
Stage 3: 55.6%
> 10 ug/dL:
Stage 1 : 33.3%
Stage 2: 66.7%
Stage 3: 0%
Chi-square p-
value<0.01
Pubic Hair
Development
<10 ug/dL:
Stage 1 : 0%
Stage 2: 55.6%
Potential Confounders
Adjusted for in Analysis
Birthweight, gestational age,
energy intake, proportion of fat
consumption, proportion of
protein consumption, maternal
alcohol consumption during
pregnancy, height at study
entry, BMI at study entry,
household income, parental
education
NOTE: exclusion of BMI and
height, in case they were part
of the causal pathway,
resulted in very similar
estimates
Considered but not included:
parity, maternal or household
smoking during pregnancy,
maternal age at birth
None
November 2012
5-538
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this Study
section) Location
Outcome Study Population
Methodological
Details
Pb
Bio-
marker
Mean Pb (SD)
in ug/dL
Ad justed
Effect
Estimates
Potential Confounders
Adjusted for in Analysis
Stage 3: 44.4%
> 10ug/dL:
Stage 1: 33.3%
Stage 2: 66.7%
Stage 3: 0%
Chi-square p-
value<0.05
Penile staging
<10ug/dL:
Stage 1: 11.1%
Stage 2: 44.4%
Stage 3: 44.4%
> 10ug/dL:
Stage 1: 58.3%
Stage 2: 41.7%
Stage 3: 0%
Chi-square p-
value<0.05
Mean
testosterone level
<10ug/dL:
4.72 (SD 1.52)
> 10ug/dL:
1.84 (SD 1.04)
•Quantitative
results for LH and
FSH not provided
November 2012
5-539
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Den Hond et al.
(2011)
Study
Location Outcome Study Population
Flanders Tanner Boys ages 14 and 15,
2003-2004 stagir|g and in their 3rd year of
gynecomastia secondary education
and living in the same
study areas for at
least 5 years
N=887
Methodological
Details
Cross-sectional
study using logistic
regression
Pb
Bio- Mean Pb (SD)
marker in ug/dL
Blood Median: 2.50
Pb 10thpercentile:
1.20
90th percentile:
5.12
Adjusted
Effect
Estimates
OR (95% Cl) for
gynecomastia
with doubling of
exposure
1.84(1.11, 3.05)
No association
between Pb and
pubic hair or
genital
development
(results not given)
Potential Confounders
Adjusted for in Analysis
Parental education, age, BMI,
smoking status
Considered but not included:
food intake, lifestyle
parameters
NOTE: results were the same
when hexachlorobenzene was
included in the model
November 2012
5-540
Draft - Do Not Cite or Quote
-------
1 Studies were performed among a cohort of Russian boys enrolled between ages 8-9 years
2 ("Williams et al.. 2010; Hauser et al.. 2008). The area where these studies were performed
3 had various environmental contaminants, such as dioxin, polychlorinated biphenyls, and
4 other metals, present but these were not included in the analyses (although preliminary
5 analyses found no correlation between blood Pb levels and serum dioxin levels). Both the
6 cross-sectional study (Hauser et al.. 2008) and the prospective study with annual follow-
7 ups (Williams et al.. 2010) demonstrated an association; higher blood Pb levels at
8 8-9 years of age was associated with later onset of puberty. In a study of boys in Egypt,
9 boys with higher blood Pb had delayed pubertal onset compared to those with lower
10 levels (Tomoum et al., 2010). In addition, compared to the low blood Pb group, those
11 boys with higher blood Pb had lower testosterone, FSH, and LH levels but there was no
12 control for confounding. A study in Flanders reported no associations between blood Pb
13 concentration and pubertal development among 14- and 15-year old boys (Den Hond E.
14 2011). However, higher blood Pb levels were associated with an increased odds of
15 gynecomastia.
16 No recent toxicological studies address Pb-induced male sexual maturation and
17 development, but earlier studies do provide support to findings in epidemiologic cohorts.
18 Pb exposure resulted in delayed sexual maturity as measured by prostate weight in male
19 Sprague-Dawley pups at PND35. These pups were exposed chronically to 1,500 or
20 4,500 ppm Pb acetate in dam or their own drinking water from GD5 until PND85 and had
21 blood Pb ranges from low to high of 88-196 and 120-379 ug/dL, respectively (Ronis et
22 al.. 1998b). Cynomolgus monkeys exposed to Pb over a lifetime (an oral capsule of
23 1,500 (ig/kg body weight/day for 10 years, blood Pb levels ranging from 30-60 ug/dL)
24 had altered pituitary and Sertoli cell function along with decreases in inhibin/FSH ratio
25 and reduced gonadotropin-releasing hormone (GnRH) stimulation of LH release in
26 adulthood (Foster et al.. 1993). all indicators that are important in proper sexual
27 maturation. Further mechanistic understanding of the effect of Pb can be gleaned from
28 studies in adult male Wistar rats exposed to Pb for 1 month (starting at PND56, 1,000 or
29 3,000 ppm Pb acetate in drinking water, respective blood Pb levels of 34 or 60 ug/dL)
30 that showed significant decreases in FSH, ventral prostate weight and serum testosterone
31 but no change in serum LH (Sokol et al., 1985). These Pb-exposed adult male rats
32 (3,000 ppm Pb acetate in drinking water starting at PND56 for 30 days) demonstrated an
33 impaired pituitary release of LH in response to challenge of the hypothalamic-pituitary-
34 adrenal (HPA) axis with the opiate antagonist naloxone, an enhanced release of LH from
35 the pituitary in response to direct stimulation of the pituitary with luteinizing hormone-
36 releasing hormone (LHRH), an enhanced response to human chorionic gonadotropin
37 (hCG) by the testes, increased pituitary LH stores, and increased GnRH mRNA levels in
38 the hypothalamus (Klein etal. 1994; Sokol. 1987). Thus, Pb likely interferes with the
39 male HPA axis, contributing to its reproductive toxicity.
November 2012 5-541 Draft - Do Not Cite or Quote
-------
1 In summary, recent epidemiologic studies have demonstrated an inverse effect of Pb on
2 pubertal development among boys at low concurrent blood Pb levels. These studies were
3 mostly cross-sectional, but associations were observed between Pb levels and delayed
4 puberty in a longitudinal study as well (Williams et al. 2010). The larger studies
5 controlled for some potential confounders, with a few studies at least considering the
6 inclusion of dietary factors, which may be an important confounder, especially in
7 populations with high prevalence of malnutrition. Some populations, such as the Russian
8 boys cohorts, had potential exposures to dioxins and polychlorinated biphenyls, but these
9 were not considered in the analyses. No recent toxicological studies were found that
10 addressed the effect of Pb on male sexual development and maturation; however, the
11 2006 Pb AQCD (U.S. EPA. 2006b) supported earlier findings that Pb exposure may
12 result in delayed onset of male puberty and altered reproductive function later in life in
13 experimental animals.
5.8.1.3 Effects on Postnatal Stature and Body Weight
14 Findings from previous toxicological studies of rodents and primates have demonstrated
15 Pb induced impairment of postnatal growth (U.S. EPA. 2006b). Little epidemiologic
16 evidence was available in the 2006 Pb AQCD on postnatal growth. Several recent
17 epidemiologic studies examining the association of various biomarkers of Pb exposure
18 with stature and body weight have been conducted and the evidence reported is mixed.
November 2012 5-542 Draft - Do Not Cite or Quote
-------
Table 5-39
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Afeiche et al.
(2011)
Schelletal.
(2009)
Summary of recent epidemiologic
Study
Location Study Methodological
and Years Population Details
Mexico City, n=523 boys Longitudinal cohort
Mexico n-477 nirls Usin9 varying
Children born 9 coefficient models
between witn random effects
1994 and
2005
Albany, New n=244 Longitudinal cohort
York study using
1986-1992 multivariate
and 1992-' regression
1998
studies of associations between Pb levels and postnatal growth.
Pb Bio-
marker
Maternal bone Pb
1 month
postpartum
Maternal blood Pb
during second
trimester, third
trimester, and
delivery; Infant
blood Pb at
delivery,
6 months, and
12 months
Mean Pb (SD)
Patella: 10.4
(11.8)ug/g
Maternal blood Pb
during second
trimester 2. 8
(2.6) ug/dL,
maternal blood Pb
during third
trimester: 2.6
(2.2) ug/dL,
maternal blood Pb
at delivery: 2.8
(2.4) ug/dL
Infant blood Pb at
delivery: 2.3
(2.7) ug/dL, infant
blood Pb at
6 months: 3.2
(3.3) ug/dL, and
infant blood Pb at
1 2 months' 6 3
(4.8) ug/dL
Adjusted Effect
Estimates
Change in weight at 5
years of age (g) per 1 SD
increase in maternal bone
Pb (95% Cl):
Girls: -171 .6 (-275.2, -68.0)
Boys: -35.0 (-132.4, 62.3)
P (p-value) for maternal
second trimester Pb
Length for age:
6 month: 0.1 49 (0.05)
12 month: 0.073(0.38)
Weight for age:
6 month: 0.01 3 (0.89)
12 month: 0.124(0.25)
Weight for length:
6 month: -0.1 58 (0.1 6)
12 month: 0.084(0.45)
Head circumference for
age:
6 month: -0.242 (0.01)
12 month: -0.220 (0.05)
Upper arm circumference
for age:
12 month: -0.1 32 (0.25)
Note: When examining
second trimester maternal
Pb a 3 ug/dL, associations
were observed for 6 mo
weight for age, 6 mo weight
for length, 6 and 12 mo
head circumference, and
12 mo upper arm
circumference for age
Potential Confounders
Adjusted for in Analysis
Cohort, maternal age, calf
circumference, height,
education, number of
pregnancies, brea+st feeding
for 6 months, Ca + treatment,
child's gestational age at
birth, height, repeated
measures of concurrent child
blood Pb
Infant sex, infant birth
weight, infant nutrition,
maternal age, marital status,
employment, race, height,
parity, second trimester
smoking, and education.
November 2012
5-543
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in Study
the text of this Location Study Methodological Pb Bio-
section) and Years Population Details marker Mean Pb (SD)
Lamb et al. Kosovo, n=309 Longitudinal cohort Maternal blood Pb Pristina: 5.60
(2008) Yugoslavia mother child study using linear measured mid- (1.99)ug/dL
1985-1986 Pairs regression pregnancy Mitrovica- 20 56
(7.38) ug/dL
Adjusted Effect
Estimates
Regression coefficients
relating maternal blood Pb:
To Height (95% Cl):
Pristina
1 yr: -0.61 (-2.24, 1 .03)
4 yr: 0.79 (-1.71, 3.29)
6.5 yr: 0.1 5 (-2.43, 2.74)
10yr: -0.09 (-3.69, 3.52)
Mitrovica
1 yr: -0.30 (-2.55, 1 .96)
4 yr: -0.72 (-3.26, 1 .82)
6.5 yr: -1 .87 (-4.38, 0.64)
10yr: -2.87 (-6.21, 0.47)
To BMI (95% Cl):
Pristina
1 yr: 0.61 (-0.28, 1 .50)
4 yr: 0.1 7 (-0.67, 1.00)
6.5 yr: 0.61 (-0.09, 1.30)
10yr: -0.49 (-1.45, 0.46)
Mitrovica
1 yr: 0.23 (-0.84, 1 .30)
4 yr: 0.1 6 (-0.66, 0.98)
6.5 yr: -0.12 (-0.90, 0.66)
10yr: 1.31 (-0.95,3.57)
Potential Confounders
Adjusted for in Analysis
Infant sex, ethnicity, parity,
maternal height or maternal
BMI, maternal education,
gestational age at delivery,
gestational age at blood
sample, HOMES score
November 2012
5-544
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Ignasiak et al.
(2006)
Mauser et al.
(2008)
Little et al.
(2009)
Study
Location
and Years
South-
western
Poland
1995
(Industrial
area with Cu
smelters and
refineries)
Chapaevsk,
Russia
May 2003 -
May 2005
Dallas, Texas
1 980-1 989
and 2002
Study
Population
school
children 7-15
years
n=463 boys
n= 436 girls
n=489 boys
8-9 yrs old
n=196
(1980s)
n=169
(2002)
2-1 2 yrs old
Methodological Pb Bio-
Details marker Mean Pb (SD)
Cross-sectional Concurrent blood 7.7 (3.5) ug/dL
study using stepwise Pb
multiple regression
analysis
Cross-sectional Concurrent blood 3 (2-5) ug/dL
study using multiple Pb Median (25.75
linear regression percentile)
Cross-sectional Concurrent blood 1980s: 23.6 (1.3
study using Pb SE) ug/dL
MANOVA, 2002' 1 6 (0 2
MANCOVA.and SE) ug/dL
regression models
Adjusted Effect
Estimates
Estimated decrement per
10 ug/dL increase in blood
Pb (p-value)
Weight:
Boys: 2.8 kg (0.002)
Girls: 3.5 kg (0.007)
Height:
Boys: 3.2cm (0.10)
Girls: (0.001)4.0 cm
Trunk length:
Boys: 1 .2 cm (0.02)
Girls: 1.1 cm (>0.01)
Leg length:
Boys: 2.1 cm (0.002)
Girls: 2.9 cm (0.0001)
Arm length:
Boys: 1.8cm (0.0001)
Girls: 1 .9 cm (0.008)
Regression coefficient
(95% Cl)
Height (cm): -1 .439 (-2,25,
-0.63)
Weight (kg): -0.761 (-1 .54,
0.02)
BMI: -0.107 (-0.44, 0.23)
Changes in mean scaled
measure per 10ug/dL Pb
increase (95%CI):
Height (cm): -2.1 (-1.9,
-2.3)
Weight (kg): -1.9 (-1 .7, -2.1)
BMI (kg/m2): -0.5 (-0.4,
-0.7)
Potential Confounders
Adjusted for in Analysis
Age, age2, education
Birth weight, gestational age,
age at exam
Age, age2, sex and cohort
effect
November 2012
5-545
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Min et al.
(2QQ8b)
Study
Location
and Years
Seoul, South
Korea
Date(s) not
specified
Study
Population
n=62 boys
n= 46 girls
5-13yrs
Methodological Pb Bio-
Details marker Mean Pb (SD)
Cross-sectional Concurrent blood 2.4 (0.7) ug/dL
study using multiple Pb
linear regression
Adjusted Effect
Estimates
Linear model estimate (SE;
P)
Height: -1 .449 (0.639;
p=0.026)
Total arm length: -1 .804
(0.702;p=0.012)
Body weight: -0.646(0.718;
p=0.370)
BMI: -0.006 (0.272;
p=0.982)
Potential
Adjusted
Confounders
for in Analysis
Age, sex, and father's
education
Sanna and Sardinia, n=825 Cross-sectional Pb in hair
Vallascas (2011) Italy children study using multiple
Data 11-14yrsold regression analysis
collected in
1 998, 2002
and 2007
1998: 5.84
(6.56) ug/g
2002: 1 .49
(1.72) ug/g
2007: 0.78
(0.93) ug/g
Height Age, sex
1998: plog Pb=-0.121
(p=0.0021)
2002: p log Pb= -0.1 15
(p=0.0349)
2007: plog Pb= 0.011
(p=0.8665)
Sitting Height
1998:plogPb=-0.117
(p=0.0017)
2002: p log Pb=-0.036
(p=0.5149)
2007: p log Pb=0.028
(p=0.6633)
ELL
1998:plogPb=-0.103
(p=0.0209)
2002: plog Pb=-0.164
(p=0.0057)
2007: p log Pb=-0.008
(p=0.9058)
November 2012
5-546
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Zailina et al.
(2008)
Tomoum et al.
(2010)
Study
Location Study Methodological Pb Bio-
and Years Population Details marker Mean Pb (SD)
Kuala n=269 Cross-sectional Concurrent blood Industrial:
Lumpur, children study using Pb 3.75 ug/dL
Malaysia 6.5-8.5 yrs correlations Urban: 3.56 ug/dL
n=169 urban
n=100
industrial
Cairo, Egypt n=41 boys Cross-sectional Concurrent blood 9.46 (3.08) ug/dL
Jan-Jun and 9irls study using t-test or Pb
2007 10-13 yrs old Mann-Whitney U-
' test
Adjusted Effect Potential Confounders
Estimates Adjusted for in Analysis
Correlation with blood Pb: N/A
Height for age:
Urban: -0.095 (p=0.21 9)
Industrial: -0.037 (p=0.71 6)
Weight for age:
Urban: 0.01 9 (p=0.806)
Industrial: -0.063 (p=0.535)
Weight for height:
Urban: 0.1 36 (p=0.079)
Industrial: -0.069 (p=0.493)
Left arm circumference:
Urban: 0.041 (p=0.595)
Industrial: -0.055 (p=0.587)
Percentage of the median N/A
(SD):
Pb<10ug/dL
Weight:
Boys: 127.56(16.26)
Girls: 114.8(10.8)
Height:
Boys: 98.06 (3.1 9)
Girls: 96.75 (2.91)
Pb> 10 ug/dL
Weight:
Boys: 122.0(16.71)
Girls: 123.11 (12.52)
Height:
Boys: 99.5 (5.04)
Girls: 100.33 (4.53)
p-value for all comparisons
>0.05
November 2012
5-547
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Olivero-Verbel et
al. (2007)
Study
Location
and Years
Cartegena,
Columbia
Jun-Aug
2004
Study
Population
n=189
children 5-9
yrs old
Methodological
Details
Cross-sectional
study using
Spearman
correlations
Pb Bio-
marker Mean Pb (SD)
Concurrent blood 5.49 (0.23) ug/dL
Pb
Adjusted Effect
Estimates
Spearman correlation
coefficient (p-value)
between blood Pb and
body size: -0.224 (0.002)
Potential Confounders
Adjusted for in Analysis
weight: -0.126(0.087)
*no significance in partial
correlation between blood
Pb and size when
controlled forage:
-0.096(0.189)
Guiyu, China
Chendian,
China
Jan-Feb
2008
n=303
3-7 yrs old
Cross-sectional
study using sample
t-tests
Concurrent blood
Pb
Guiyu: 13.2
(4.0-48.5) ug/dL
Chendian: 8.2
(0-21.3) ug/dL
Median (range)
Mean chest circumference
among girls:
<10ug/dL: 50.31 +1-3.22
cm
> 10 ug/dL: 49.03+/-2.27
cm
(p-value <0.05)
Mean chest circumference
among children >6 years
old
<10 ug/dL: 51.70+/-3.35
cm
> 10 ug/dL: 52.87+/-
2.49 cm
(p-value <0.05)
Mean head circumference
among children >6 years
old
<10 ug/dL: 48.71 +/-1.66
cm
> 10 ug/dL: 50.04+/-1.29
cm
(p-value <0.01)
November 2012
5-548
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Mahram et al.
(2007)
Study
Location
and Years
Zanjan
province,,
Iran
Date(s) not
specified
Study
Population
n=42 boys
n~ 39 girls
n-45 cases
n-36
controls
7-11 yrs
Methodological Pb Bio-
Details marker
Case-control study Concurrent blood
using t-tests Pb
Mean Pb (SD)
Area with Pb
smelters: 37.0
(24.7) ug/dL
Area without Pb
smelters: 15.6
(13.4)ug/dL
Adjusted Effect
Estimates
Comparison of control and
study groups Height,
standardized for age: p-
value 0.52
Weight, standardized for
age: p-value 0.8
Potential Confounders
Adjusted for in Analysis
N/A
*Estimated Lower Limb Length
November 2012
5-549
Draft - Do Not Cite or Quote
-------
1 Results from recent epidemiologic studies of postnatal growth are summarized in Table
2 5-39. Longitudinal epidemiologic studies have had inconsistent findings regarding the
3 association between Pb levels and post-natal growth. Afeiche et al. (2011) conducted a
4 longitudinal study of children in Mexico City, born between 1994 and 2005. Maternal
5 bone Pb during pregnancy was associated with a statistically significant decrease in
6 weight at age 5 years in girls but not in boys. The findings were robust to additional
7 adjustment for child's concurrent blood Pb level. A study in New York reported an
8 inverse association between maternal blood Pb during the second trimester of pregnancy
9 and various measures of growth, especially among those mothers with blood Pb levels of
10 at least 3 (ig/dL (Schell et al., 2009). These associations did not persist for those with
11 maternal blood Pb levels less than 3 (ig/dL. Among infants, 6 month blood Pb levels were
12 not associated with measures of growth at 12 months. In comparisons of changes in blood
13 Pb levels over time, high maternal blood Pb combined with low 12 month blood Pb
14 among infants (indicating a decrease in blood Pb over time) resulted in the greatest
15 growth, even compared to those with both low or both high maternal and infant blood Pb
16 measures. In a prospective study of 309 mother-child pairs from Yugoslavia, the
17 relationship between maternal blood Pb measured mid-pregnancy and attained height in
18 children was investigated in those living in a highly exposed town with a smelter and
19 battery plant and those living in a relatively lower exposed town (Lamb et al.. 2008). In
20 multivariate adjusted regression models, neither attained height (at birth, 1, 4, 6.6, or
21 10 years age) nor rate of height change per month (at birth-1 year, 1-4 years, 4-6.5 years,
22 6.5-10 years age) was associated in a consistent direction with maternal pregnancy blood
23 Pb levels in either the industrial or less exposed town. Weight was also not associated
24 with maternal blood Pb in this study.
25 Multiple cross-sectional studies reported an association between Pb levels and impaired
26 growth. Ignasiak et al. (2006) studied school children aged 7-15 years living close to Cu
27 smelters and refineries in Poland to assess the impact of Pb exposure on their growth
28 status. There was a statistically significant linear relationship between concurrent blood
29 Pb and reduced weight, height, trunk, leg and arm lengths. This decrease in height was
30 more influenced by decreases in leg length than trunk length. These results also indicated
31 that there was attenuation in osteoblast activity associated with higher blood Pb levels,
32 consistent with animal toxicological studies (Long etal. 1990). Hauser et al. (2008)
33 investigated the relationship between blood Pb and height in boys living in Chapaevsk,
34 Russia, an area contaminated with multiple pollutants including dioxins and metals. In a
35 multivariate adjusted regression analysis, height significantly decreased with increasing
36 blood Pb. Statistically nonsignificant decreases in weight and BMI were also observed.
37 The association of blood Pb with height, weight, and BMI was examined among two
38 cohorts of children living near Pb smelters in Texas (Little et al.. 2009). The first cohort
39 included children 2-12 years old in 1980 and the second cohort included children of the
November 2012 5-550 Draft - Do Not Cite or Quote
-------
1 same age in 2002 when blood Pb levels were substantially lower. Decreases in height,
2 weight, and BMI with increasing blood Pb levels were observed among children in both
3 cohorts and increases in height and weight were observed comparing children from the
4 2002 cohort to those from the 1980 cohort. In a study with Korean children, Min et al.
5 (2008b) observed that height and total arm length decreased significantly with increasing
6 blood Pb in multivariate adjusted regression models. A statistically nonsignificant
7 decrease in body weight was observed with increasing blood Pb while no effect on BMI
8 was reported. In a study of children in Sardinia Italy, Sanna and Vallascas (2011)
9 measured Pb in hair at three points in time (1997, 2002, and 2007) and reported cross-
10 sectional results from regression analyses for each of these time periods. Pb in hair
11 decreased over time and significant associations of Pb in hair with height were observed
12 only in earlier time periods when hair Pb levels were relatively high. However, Pb in hair
13 samples is not a well-characterized biomarker (see Chapter_4 and Section 4.3.4.2).
14 Contrary to the results summarized above, several cross-sectional studies do not observe
15 associations between blood Pb levels and impaired growth. In a study with a similar
16 design, Zailina et al. (2008) studied the relationship of blood Pb and height in 7 year-old
17 Malaysian school children comparing those attending two schools in an urban setting to
18 those attending a school near an industrial area. After adjustment for age no statistically
19 significant associations between concurrent blood Pb and physical development were
20 observed. Tomoum et al. (2010) investigated the association between blood Pb and height
21 in pubertal children in Cairo, Egypt. Neither boys nor girls with concurrent blood Pb
22 levels >10 (ig/dL differed significantly in height or weight when compared to those with
23 blood Pb <10 (ig/dL. In a simple correlation analysis of children aged 5-9 years in
24 Colombia, Olivero-Verbel et al. (2007) reported that concurrent blood Pb levels were
25 negatively associated with body size (r = -0.224, p <0.002). However, when a partial
26 correlation analysis was performed controlling for age, the association between blood Pb
27 and body size was no longer statistically significant. In a study of school children in
28 China, chest and head circumference were found to differ between high (>10(ig/dL) and
29 low concurrent blood Pb level groups; however, the direction of the difference was not
30 consistent (Liu et al.. 20 lib). Among girls, in comparison of those with high and low
31 blood Pb levels, a reduction in head circumference was observed. Among children greater
32 than 6 years of age, those with higher blood Pb levels were reported to have greater head
33 and chest circumferences. In a study of children aged 7-11 years and living in an area of
34 Iran with or without Pb smelters, age-standardized weight and height did not vary by
35 study area (Mahram et al.. 2007).
36 Evidence from previous toxicological studies has shown an association between
37 gestational Pb exposure and impaired postnatal growth (U.S. EPA. 2006b). Recent
38 toxicological studies report significant changes in postnatal or adult body weight after Pb
November 2012 5-551 Draft - Do Not Cite or Quote
-------
1 exposure during different developmental windows. Masso-Gonzalez and Antonio-Garcia
2 (2009) found Pb-induced decreased body weights at weaning (PND21) in rat pups from
3 dams exposed to Pb during pregnancy and lactation (drinking water, 300 mg/L). Blood
4 Pb level in the control group was 1.43 ug/dL, in the Pb group it was 22.8 ug/dL. Dong et
5 al. (2009) reported decreased body weight in adult Kunming mice after exposure to
6 6,000 ppm Pb acetate in drinking water for 8 weeks. In contrast, Leasure et al. (2008)
7 reported a statistically significant inverse relationship between Pb exposure and body
8 weight for male mice exposed to lower (27 ppm), moderate (55 ppm) and higher levels
9 (109 ppm) levels of Pb during gestation and lactation (dam drinking water, 2 weeks
10 before mating, through gestation and to PND10) with those exposed to the lowest dose
11 having the highest adult body weight among the overweight Pb-exposed animals. Male
12 mice exposed to the lower and higher Pb concentrations during gestation were 26% and
13 13% heavier than were controls at 1 year of age, respectively. In this study, dams were
14 administered 27 ppm (low), 55 ppm (moderate), and 109 ppm (high) Pb in drinking water
15 beginning which resulted in respective blood Pb levels from 10 ug/dL or less in the
16 low-exposure offspring to 42 ug/dL in the high-exposure offspring at PND10. Leasure et
17 al. (2008) also exposed a separate group of mice to Pb only during the postnatal period
18 (PNDO-PND21, lactation only exposure) and mice exposed to the same aforementioned
19 low or high dose of Pb did not exhibit a difference in body weight when compared to
20 control offspring. Wang et al. (2009e) observed a statistically significant decrease in fetal
21 body weight and body length of Wistar rats at GD20 after maternal exposure to 250 ppm
22 Pb acetate during gestation days 1-10, 11-20, or 1-20. Also, associations were reported
23 between elevated maternal blood Pb levels (0.6, 1.3, or 1.74 (iM, respectively or -12.4,
24 26.9, or 36.0 (ig/dL, respectively) compared to control (0.04 (iM or -0.83 (ig/dL)
25 decreased pup body length, and placental weight in Wistar rats at GD20. The greatest
26 decrease in fetal body weight and length was observed in the group exposed to Pb during
27 gestation days 1-20 followed by the group exposed to Pb during gestation days 11-20.
28 Teijon et al. (2006) observed reductions in birthweight of litters administered 200 ppm or
29 400 ppm Pb acetate in drinking water (Wistar rats, Pb to dams from GD1 through
30 lactation to 1 and 3 months postweaning to pups), but found that this effect did not persist
31 in the postnatal growth of the rats.
32 Notably, previous toxicological studies observed reductions in postnatal weight as well as
33 birth weight after exposure to Pb, albeit often at higher concentrations of Pb exposure.
34 Ronis et al. (2001; 1998a: 1998b: 1996) have published a series of papers exposing rats to
35 Pb over different developmental windows, showing associations between Pb exposure
36 and deficits in growth. Sprague-Dawley rats with lifetime Pb exposure to 6,000 ppm
37 Pb acetate in drinking water (gestational-termination of experiment Pb exposure,
38 maximum blood Pb of 316 (ig/dL in males and 264 (ig/dL in females) had sex-
39 independent pre-pubertal growth suppression, male-specific suppression of pubertal
November 2012 5-552 Draft - Do Not Cite or Quote
-------
1 growth and loss of growth effects postnatally but still maintained an overall decreased
2 body size out to PND60 due to earlier deficits. In a follow up study using the same
3 exposure duration with a dose of 4,500 ppm Pb acetate (resulting in blood Pb of
4 263 (ig/dL at PND85) yielded the same results (Ronis etal. 1996) with mechanistic
5 insight showing decrements in insulin-like growth factor 1 (IGF1) accompanying the
6 decreases in growth rates.
7 In summary, the body of toxicological literature on postnatal growth with Pb exposure
8 indicates that Pb exposure can induce decrements in both height/body length and BW that
9 may be persistent and differ by sex. However, findings from epidemiologic studies of
10 postnatal growth are not consistent. Many of these studies were limited by their cross-
11 sectional design. A few studies used longitudinal cohorts and controlled for multiple
12 potential confounders, such as parity, but the results of these studies are inconsistent.
13 Animal toxicology studies give insight to mechanistic changes that may contribute to this
14 Pb-induced decrement and to the windows of exposure that may contribute greatest to
15 these decrements.
5.8.2 Toxicological Studies of Other Developmental Effects
5.8.2.1 Developmental Effects on Blood and Liver
16 The 1986 and 2006 Pb AQCDs [(U.S. EPA. 1986b) and (U.S. EPA. 2006b)1 reported
17 studies that suggest Pb may alter hematopoietic and hepatic function during development.
18 Some recent studies provide evidence that support these findings; however recent results
19 are not consistent among the studies.
20 Masso et al. (2007) reported a decrease in liver weights of pups born to dams that
21 consumed 300 mg/L Pb in drinking water during gestation and lactation. They also
22 reported an increase in the number of erythrocytes; however the erythrocyte size was
23 diminished by 62%. Pb produced microcytic anemia as evidenced by decreased
24 hemoglobin content and hematocrit values without changes in mean corpuscular
25 hemoglobin (MCH) concentration. Alkaline phosphatase (ALP) activity, CAT activity, or
26 thiobarbituric acid reactive substances (TEARS) production did not change in pups at
27 postnatal 0, but increased statistically significantly by PND21 indicating reactive oxygen
28 generation. No change in acid phosphatase (ACP) activity was observed in the livers of
29 pups at PNDO or PND21.
November 2012 5-553 Draft - Do Not Cite or Quote
-------
1 Masso-Gonzalez and Antonia-Garcia (2009) reported normochromic and microcytic
2 anemia and a significant decrease in hematocrit values and blood 5-aminolevulinic acid
3 dehydratase (ALAD) activity (90% reduction) in pups from dams administered 300 mg/L
4 Pb acetate in drinking water during gestation. The authors also reported that erythrocyte
5 osmotic fragility was four times greater in Pb-exposed pups than in control pups.
6 Masso-Gonzalez and Antonia-Garcia (2009) reported increases in TEARS and CAT
7 activity in the liver after Pb exposure. Intoxication with Pb also resulted in decreased
8 liver protein concentrations and Mn-dependent SOD activity. Abnormalities in liver
9 function were further exemplified by increases in liver concentrations of ALP and ACP.
10 Teijon et al. (2006) observed that gestational exposure to Pb caused a decrease in
11 erythrocytes, hemoglobin, and MCH at weaning; however, by 1 and 3 months
12 postweaning, these parameters had returned to normal values. The authors observed a
13 slight increase in serum ALP, alanine aminotransferase (ALT), and aspartate
14 aminotransferase (AST) levels after Pb exposure in the absence of liver histological
15 changes.
16 Pb-induced effects on SOD activity in the liver of fetuses after Pb intoxication was
17 supported by a study by Uzbekov et al. (2007). The authors reported an initial increase in
18 SOD activity in livers of pups exposed to 0.3 mg/L and 3.0 mg/L Pb nitrate in drinking
19 water during gestation for 1 month (mean daily consumption 27 (ig/kg). In contrast, long-
20 term exposure (5 months) to the same concentrations of Pb nitrate concentration during
21 gestation resulted in decreased hepatic SOD activity.
22 Effects on hepatic Phase I and Phase II enzymes after early developmental exposure of
23 offspring to Pb during gestation and lactation was evaluated by Pillai et al. (2009). In the
24 study, pregnant Charles Foster rats were administered 0.05 mg/kg body weight Pb
25 subcutaneously throughout gestation until PND21. Pups were evaluated on PND56.
26 Results of the study show that Phase I xenobiotic-metabolizing enzymes (NADPH- and
27 NADH cytochrome c reductase) and Phase II xenobiotic- and steroid-metabolizing
28 enzymes (5-glutamyl transpeptidase, UDPGT, glutathione-s-transferase, and 17(3-
29 hydroxysteroid oxidoreductase) were reduced in both male and female pups by PND56.
30 Only inhibition in glutathione-s-transferase and 17(3-hydroxysteroid oxidoreductase
31 activities demonstrated a sex-specific pattern (glutathione-s-transferase inhibition in
32 males; 17|3-hydroxysteroid oxidoreductase inhibition greater in females). Observed
33 Pb-induced histological changes included massive fatty degeneration in hepatocytes,
34 large vacuoles in cytoplasm, appearance of pyknotic nuclei, and infiltration of
35 lymphocytes in the liver. Activities of antioxidant enzymes (SOD, CAT, glutathione
36 peroxidase, and glutathione reductase) were also reduced after Pb intoxication.
37 Alterations in biochemical parameters included decreased DNA, RNA, and cholesterol
November 2012 5-554 Draft - Do Not Cite or Quote
-------
1 content, although it was not clear whether these changes were related to genetic
2 expression of xenobiotic-metabolizing enzymes or changes in steroid hormone
3 homeostasis.
5.8.2.2 Developmental Effects on Skin
4 The 2006 Pb AQCD (U.S. EPA. 2006b) reported a study that demonstrated Pb-induced
5 abnormalities in skin development. No current studies were identified that addressed
6 Pb-induced skin alterations.
5.8.2.3 Developmental Effects on the Retina
7 The 2006 Pb AQCD (U.S. EPA. 2006b) concluded that Pb exposure during early
8 postnatal development (resulting in blood Pb levels ~20 (ig/dL) impaired retinal
9 development in female Long-Evans hooded rats. A more recent study (Fox et al., 2008)
10 exposed female Long-Evans hooded rats to low (27 ppm), moderate (55 ppm), and high
11 (109 ppm) levels of Pb acetate in drinking water beginning 2 weeks before mating,
12 throughout gestation, and until PND10. Blood Pb levels measured in these pups on
13 postnatal days 0-10 were 10-12 (ig/dL (low), 21-24 (ig/dL (moderate), and 40-46 (ig/dL
14 (high). Results of the study demonstrated supernormal persistent rod photoreceptor-
15 mediated (scotopic) electroretinograms (ERGs) [(Fox et al.. 2008). and Table 5-131 in
16 adult rats similar to ERG findings in male and female children in association with
17 maternal first trimester blood Pb levels 10.5-32 (ig/dL [(Rothenberg et al., 2002b), and
18 Table 5-131. In rats, low- and moderate-levels of Pb increased neurogenesis of rod
19 photoreceptors and rod bipolar cells without affecting Miiller glial cells and statistically
20 significantly increased the number of rods in central and peripheral retina. High-level Pb
21 exposure (109 ppm) statistically significantly decreased the number of rods in central and
22 peripheral retina. Pb-exposure induced concentration-dependent decreases in adult rat
23 retinal dopamine synthesis and utilization/release.
5.8.2.4 Developmental Effects on Teeth
24 Pb has been associated with multiple health effects including dental caries, however,
25 there is very limited information available on the temporal and spatial incorporation of Pb
26 in dental tissue (Arora et al. 2005). Arora et al. (2005) demonstrated that Wistar rat pups
27 exposed to Pb during gestation and lactation (40 mg/L of Pb nitrate in drinking water of
28 pregnant dams) had higher concentrations of Pb on the surface of enamel and in the
November 2012 5-555 Draft - Do Not Cite or Quote
-------
1 dentine immediately adjacent to the pulp. The authors concluded that additional research
2 is needed on the intracellular uptake of Pb during tooth development to fully understand
3 the spatial distribution of Pb in teeth.
5.8.3 Effects on Birth Outcomes
4 The 2006 Pb AQCD reported on multiple studies of adverse birth outcomes such as, fetal
5 mortality, birth defects, preterm birth, and low birth weight/fetal growth (U.S. EPA.
6 2006b). The toxicological studies reviewed in the 2006 Pb AQCD concluded that Pb
7 exposure can increase fetal mortality and produce sublethal effects, smaller litters, and
8 fewer implantation sites. Epidemiologic studies using occupational histories reported the
9 possibility of small associations between increased Pb exposure and birth defects, and
10 toxicological studies demonstrated associations between exposure to high doses of Pb
11 and increased incidences of teratogenic effects in experimental animals. Epidemiologic
12 studies on preterm birth and low birth weight/fetal growth included in the
13 2006 Pb AQCD reported inconsistent findings. Evidence from previous toxicological
14 studies has shown an association between gestational Pb exposure and reduced birth
15 weight and decreased litter size or number of pups. Continued research on adverse birth
16 outcomes is described in the sections that follow.
5.8.3.1 Infant Mortality and Embryogenesis
17 No recent epidemiologic or toxicological studies have reported on the relationship
18 between Pb levels and infant mortality. The 2006 Pb AQCD (U.S. EPA. 2006b)
19 concluded that Pb exposure can increase fetal mortality and produce sublethal effects
20 (disrupt growth and development) in offspring of Pb exposed dams at concentrations that
21 do not result in clinical toxicity to the dams by disrupting implantation and pregnancy,
22 particularly at the blastocyst stage of development. In rodent studies gestational exposure
23 to Pb (blood Pb levels 32 to >70 ug/dL) resulted in smaller litters and fewer implantation
24 sites and in non-human primates pre- and perinatal mortality was reported in squirrel
25 monkeys exposed to Pb (mean dam blood Pb level of 54 ug/dL) in the last two-thirds of
26 gestation (U.S. EPA. 2006b). There is substantial evidence to show that there is no
27 apparent maternal-fetal barrier to Pb and it can easily cross the placenta and accumulate
28 in fetal tissue during gestation (Pillai et al., 2009; Wang et al., 2009e; Uzbekov et al.,
29 2007).
November 2012 5-556 Draft - Do Not Cite or Quote
-------
5.8.3.2 Birth Defects
1 The 2006 Pb AQCD (U.S. EPA. 2006b) reported the possibility of small associations
2 between high Pb exposure and birth defects, but many of the epidemiologic studies used
3 occupational histories instead of actual measures of blood Pb levels. Among the studies
4 included in the 2006 Pb AQCD, a couple reported associations between parental
5 exposure to Pb and neural tube defects (Irgens et al.. 1998; Bound et al.. 1997). Recent
6 studies also examined indicators of Pb exposure and neural tube defects (Table 5-40). No
7 other recent epidemiologic studies of Pb exposure and birth defects were identified in the
8 literature. No recent toxicological studies were found that investigated Pb-induced
9 changes in morphology, teratology effects, or skeletal malformations of developing
10 fetuses as a result of maternal Pb exposure; however, in the 2006 Pb AQCD toxicological
11 studies demonstrated associations between exposure to high doses of Pb and increased
12 incidences of teratogenic effect in experimental animals.
November 2012 5-557 Draft - Do Not Cite or Quote
-------
Table 5-40 Summary of recent epidemiologic studies of associations between
Reference
(Presented
in order of
appearance
in the text)
Zeyrek et
al. (2009)
Brender et
al. (2006)
Huang et
al. (201 1b)
Pb levels and
Study Study
Location Population
Turkey Infants with
NS gestational
age of at
least
20 weeks
NNTD=74
NControls=70
Texas Infants of
1995-2000 Mexican-
American
women
NNTD=184
Ncontrols=225
China Live and
2002-2004 sti" birtns of
women
living in the
study area
(villages in
the Lvliang
region of
Shanxi
province)
N=112
villages
neural tube
Methodological
Details
Case-control
study using
Student's t-test
and Mann-
Whitney U-test
Case-control
study using
logistic
regression
Ecologic
defects.
Pb Biomarkers
and Exposure
Measurement
Maternal and
umbilical cord
blood Pb taken
0.5h afterbirth
Maternal blood
Pb
taken 5-6 weeks
post-partum
2 soil samples
from each
village
Mean Pb
(SD)
in ug/dL
Cases:
Maternal:
15.5(15.0)
Umbilical
cord' 182
(17.8)
Controls:
Maternal:
12.5(12.7)
Umbilical
cord: 16.5
(16.1)
Cases: 2.4
(1.9)
Controls: 2.5
(1.6)
56.14ug/g
(11.43ug/g)
Adjusted
Effect
Estimates
P-values for
differences of
Student's t-test
or Mann-
Whitney U test
(dependent on
distribution)
were 0.35 for
maternal blood
Pb and 0.63 for
umbilical cord
blood Pb
OR (95% Cl):
Blood
Pb<6.0 ug/dL:
1 .0 (Ref)
Blood
Pb> 6.0 ug/dL:
1.5(0.6, 4.3)
N/A
Potential
Confounders
Adjusted for
in Analysis
N/A
Inclusion of
breast feeding
in the model
changed the
OR (95% Cl)
to 3 8 (0 8
19.5)
1
2
o
J
4
5
6
7
8
9
10
11
12
13
Among the recent epidemiologic studies (described in Table 5-40). a study of women in
Turkey detected no difference between the blood Pb of mothers or the umbilical cord
blood Pb of the newborns for healthy infants compared with infants with neural tube
defects (cases of spina bifida occulta were excluded, but other forms of spina bifida were
included) (Zevrek et al.. 2009). Brender et al. (2006) performed a study of Mexican-
American women living in Texas. Measurements were taken 5-6 weeks postpartum,
which is a limitation of this study because the blood Pb levels may be different from
those during the developmental period of gestation. The OR comparing women with at
least 6 (ig/dL blood Pb to those with less than 6 (ig/dL blood Pb was 1.5 (95% Cl: 0.6,
4.3). This increased after adjusting for breast feeding, although this variable was not a
confounder because it cannot be associated with neural tube defects. For these women,
neither occupational exposure to Pb nor proximity of residence to a facility with Pb air
emissions at the time of conception was associated with increased odds of neural tube
November 2012
5-558
Draft - Do Not Cite or Quote
-------
1 defects. A study with an ecologic design was performed in China and did not use
2 individual-level biomarkers to determine Pb levels (Huang et al.. 201 Ib). A positive
3 association between Pb levels in soil samples and neural tube defects was reported.
4 Exposure to multiple other trace elements also demonstrated a positive association but no
5 control for co-exposures was included in the models for Pb.
6 In summary, previous studies included in the 2006 Pb AQCD observed associations
7 between Pb and neural tube defects but were limited due to the lack of biologically
8 measured Pb [Pb was measured in drinking water (Bound etal.. 1997) and estimated
9 from occupational reports drgens et al.. 1998)1. A recent ecologic study reported an
10 association between Pb in the soil and neural tube defects but was also limited by its lack
11 of biological samples, as well as a lack of individual-level data and the prevalence of
12 several other metals (Huang et al.. 201 Ib). Other recent epidemiologic studies of
13 maternal blood Pb levels and neural tube defects observed no statistically significant
14 associations (Zeyrek et al.. 2009; Brender et al.. 2006). These studies also have
15 limitations, including the timing of Pb measurements and lack of control for potential
16 confounders.
5.8.3.3 Preterm Birth
17 Epidemiologic studies on preterm birth included in the 2006 Pb AQCD (U.S. EPA.
18 2006b) reported inconsistent findings regarding the relationship between Pb and
19 gestational age. Recent studies examined this potential association and again mixed
20 results were reported (Table 5-41). Of these studies, the ones that categorized births as
21 preterm or term all defined preterm birth as less than 37 weeks of gestation. One
22 limitation to note for these studies is that if Pb affects spontaneous abortion and length of
23 gestation via a similar pathway, then the studies that only collect data at delivery and not
24 at earlier stages of pregnancy would be biased toward the null.
November 2012 5-559 Draft - Do Not Cite or Quote
-------
Table 5-41 Summary of recent epidemiologic studies of associations between Pb levels and preterm birth.
Reference
Presented in
order of
appearance
in the text)
Jelliffe-
Pawlowski et
al. (2006)
Vigeh et al.
Cantonwine
et al. (2010a)
Study
Location Outcome Study Population
California Preterm birth Singleton births to
1995-2002 (<37 . . . . . non-smoking mothers
completed week) with blood Pb
measures during
pregnancy from
either the California
Childhood Lead
Poisoning Prevention
Dl Ctl 1UI 1 Ul LI 1C
California
Occupational Lead
Poisoning Prevention
Program
Npreterm birth=30
Nterm birth=232
Tehran, Preterm birth Singleton births from
Iran (20-37 week) non-smoking, non-
2006 obese mothers aged
16-35 and referred
for prenatal care
during the
8th-12th week of
gestation
Npreterm birth~44
Nterm birth=304
Mexico Preterm birth Births to mothers with
City (<37wk), at least 1 blood Pb
1997-1999 Gestational age measurement during
pregnancy and no
chronic diseases
requiring medication
Methodological
Details
Longitudinal
cohort study using
logistic regression
Longitudinal
cohort study using
logistic regression
Longitudinal
cohort study using
linear regression
Pb Biomarkers or Mean Pb
Exposure (SD)
Measurement in ug/dL
Maximum maternal blood a 10 ug/dL:
Pb during pregnancy 30.9%
Maternal blood Pb at 3.8 (2.0)
8-12 weeks gestation
Maternal blood Pb during Blood Pb
pregnancy visit gt
<20wks
pregnant 7.2
(5 2)
\<->.*-i
Visit at
Adjusted
Effect
Estimates
Odd Ratios:
< 5 ug/dL: 1 .00
(Ref)
V *<-'y
6-9 ug/dL: 0.8
(0.1 , 6.4)
10-19 ug/dL: 1.1
(0.2, 5.2)
20-39 ug/dL: 4.5
(1 .8, 10.9)
> 40 ug/dL: 4.7
(1.1, 19.9)
<10 ug/dL: 1.00
(Ref)
> 10ug/dL: 3.2
(1.2,7.4)
Mean blood Pb
(SD):
Preterm birth:
4.52 (1 .63)
Term birth:
3.72 (2.03)
p-value for
difference: <0.05
OR (95% Cl)
1.41 (1.08, 1.84)
(unit not given,
assume per
1 ug/dL)
Linear regression
P (95% Cl) per
SD increase in
centered log-Pb
concentration
Blood Pb
Potential
Confounders
Adjusted for in
Analysis
In the 10ug/dL
model: race,
insurance, maternal
age, parity, infant
sex, low birthweight
Age, infant sex,
education, passive
smoking exposure,
pregnancy weight
gain, parity,
hematocrit, type of
delivery
Infant sex, maternal
age, maternal
education, history
of adverse birth
outcomes, cigarette
smoking, parity
November 2012
5-560
Draft - Do Not Cite or Quote
-------
Reference
Presented in
order of
appearance Study
in the text) Location Outcome
Pb Biomarkers or Mean Pb
Methodological Exposure (SD)
Study Population Details Measurement in ug/dL
20-28 weeks
Npre«ermbirth=22 pregnant 6.3
N,ermbirth=213 ^ g(
>28 weeks
pregnant 6.8
(4.5)
Plasma Pb
Visit at
<20wks
pregnant
0.17(0.16)
Visit at
20-28 weeks
pregnant
0.13(0.10)
Visit at
>28 weeks
pregnant
0.16(0.26)
Potential
Adjusted 9°Pfc?J"1clers-
Effect Adjusted for in
Estimates Analysis
Visit at <20wks:
-2.76 (-5.21,
-0.31)
Visit at
20-28 weeks:
-1.77 (-3.39,
-0.15)
Visit at
>28 weeks: -0.47
(-1 .78, 0.84)
Average: -1.49
(-3.63, 0.64)
Plasma Pb
Visit at <20wks:
-2.38 (-4.97,
0.21)
Visit at
20-28 weeks:
-1.34 (-2.98,
0.29)
Visit at
>28 weeks: -1 .28
(-2.63, 0.06)
Average: -0.28
(-2.81, 2.25)
Plasma-to-blood
Pb ratio
Visit at <20wks:
-3.23 (-6.01 ,
-0.44)
Visit at
20-28 weeks:
-1.41 (-3.10,
0.29)
Visit at
>28 weeks: -1 .30
(-2.67, 0.07)
Average: -1 .27
(-3.89, 1 .35)
Cord blood Pb
-0.68 (-2.37,
1.00)
November 2012
5-561
Draft - Do Not Cite or Quote
-------
Reference
Presented in
order of
appearance
in the text)
Zhuetal.
(2010)
Chen et al.
(2006a)
Study
Location Outcome Study Population
New York Preterm birth Singleton births to
2003-2005 (<37 , , . .. m°'h.f.rs ^1 5-48
completed week) with blood Pb
measures before or
on the date of
delivery and blood Pb
measuring <10 ug/dL
Npreterm birth=351 9
Nterm birth=39,769
Taiwan Preterm birth Infants born to at
1993-1997 (<37 week) least one parent who
was part of the
Program to Reduce
Exposure by
Surveillance System
- Blood Lead Levels
cohort that monitored
workers
occupationally
exposed to Pb
Npreterm birth=74
N,ermbirth=1537
*738 births had
maternal Pb
information and 967
had paternal Pb
information
Methodological
Details
Retrospective
cohort study using
logistic regression
with fractional
polynomials
Occupational
cohort study using
regression models
Pb Biomarkers or
Exposure
Measurement
Maternal blood Pb
Maternal blood Pb during
pregnancy (or if that
wasn't available, the 1
year prior to fertilization)
and/or paternal blood Pb
during spermatogenesis
(the 64 days before
fertilization, or if that
wasn't available, the 1
year prior to
spermatogenesis)
Mean Pb
(SD)
in ug/dL
2.1
Maternal
blood Pb
10.1 (10.4)
Paternal
blood Pb
12.9(13.8)
Adjusted
Effect
Estimates
Odd Ratios:
<1.0ug/dL: 1.00
(Ref)
V *cly
1.1-2.0ug/dL:
1.03 (0.93, 1.13)
2.1-3.0ug/dL:
1.01 (0.92, 1.10)
3.1-9.9ug/dL:
1 .04 (0.89, 1 .22)
Risk Ratios
Maternal blood
Pb
<10 ug/dL: 1.00
10-19 ug/dL:
1 .97 (0.92, 3.86)
>20ug/dL: 1.86
(0.68, 4.28)
Paternal blood
Pb
<10 ug/dL: 1.00
10-19 ug/dL:
1.17 (0.53, 2.32)
> 20 ug/dL: 0.55
(0.19, 1.28)
Potential
Confounders
Adjusted for in
Analysis
Timing of Pb test,
maternal age, race,
Hispanic ethnicity,
smoking status,
drug abuse, marital
status, special
financial program
participation, parity,
infant sex
Parental age,
parental education,
parity, infant sex
November 2012
5-562
Draft - Do Not Cite or Quote
-------
Reference
Presented in
order of
appearance
in the text)
Patel and
Prabhu
(2009)
Jones et al.
(2010)
Wells et al.
(201 la)
Study
Location Outcome Study Population
Nagpur, Gestational age Consecutive births at
India the study hospital
NS
N=205 (mean
gestational age 39
+1-2 weeks)
Tennessee Gestational Age: Singleton births
2006 preterm (<37wk) > 27 week gestation
, term from mothers aged
(37-40 week), 1 6-45 living in the
post-term Shelby County area
(>40 week) for at least 5 mo
during pregnancy
Npreterm birth=10
Nterm birth=81
Nposttermbirth = 11
Baltimore, Gestational age Singleton births from
MD the Baltimore
2004-2005 Tracking Health
Related to
Environmental
Exposures (THREE)
study
Nnraf-arm hir+h=39
preterm birth ^^
Nterm birth=261
Pb Biomarkers or
Methodological Exposure
Details Measurement
Cross-sectional Umbilical cord blood Pb
study using linear
regression
Cross-sectional Umbilical cord blood Pb
study comparing
across geometric
means (test not
specified)
Cross-sectional Umbilical cord Pb
study using
multivariable linear
regression
Mean Pb
(SD)
in ug/dL
Umbilical
cord blood
Pb: 4.7
(12.1)
2.4 (4.3)
Geometric
mean: 1 .3
0.84 (95%:
Cl 0.72,
0.96)
a 5 ug/dL:
0.7%
Adjusted
Effect
Estimates
>5 ug/dL: mean
gestational age
38 weeks
< 5 ug/dL: mean
gestational age
39 weeks
Linear
regression:
gestational age
decreased
1 week with
every 1 ug/dL
increase in
umbilical cord
blood Pb (exact
values and 95%
Cl: not given)
Geometric Mean:
Preterm birth: 1.4
Term birth: 1.2
Post-term birth:
1.3
p-value for
difference: >0.10
Ratio for Pb
concentration per
10 days of
gestation: 0.99
(0.93, 1.06)
Potential
Confounders
Adjusted for in
Analysis
Not specified
None
Maternal age, race,
insurance, pre-
pregnancy BMI,
smoking status,
gestational age,
birthweight,
average year of
neighborhood
home construction
November 2012
5-563
Draft - Do Not Cite or Quote
-------
Reference
Presented in
order of
appearance
in the text)
Study
Location
Outcome
Study Population
Methodological
Details
Pb Biomarkers or
Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted
Effect
Estimates
Potential
Confounders
Adjusted for in
Analysis
Berkowitz et
al. (2006)
Shoshone
County,
Idaho
1970-1981
Preterm birth
(<37wk)
Singleton births
with 28-45 week
gestation
Npreterm birth=7843
N,ermbirth= 162,035
Cohort study
using logistic
regression
Three time periods of two
locations (unexposed
and exposed/near
smelter): pre-fire, "high-
exposure period" (when
a fire happened at the
smelter and resulted in
damages leading to high
air Pb concentrations for
6 mo), and "post-fire"
During the
time of the
fire,
estimates of
Pbin
ambient air
were as high
as 30 ug/m
OR (90% Cl)
(unexposed
location is
referent group):
Pre-fire 0.93
(0.67, 1.28)
High exposure
0.68(0.34, 1.35)
Post-fire 1.17
(0.95, 1.45)
Maternal age, infant
sex, first birth,
previous
miscarriage or
abortion
Orun et al.
(2011)
Turkey
NS
Preterm birth
(<37 week)
Births to mothers not
occupationally
exposed to toxic
metals and living in a
suburban but non-
industrial area
Cohort study
using Mann-
Whitney U-test
Breast milk 2 months
post-partum
Median:
20.6 ug/L
>WHO limit
(5 ug/L):
Median Pb (IQR)
>37 week:20.6
(11.2, 29.2) ug/L
< 37 week: 20.4
(14.4, 27.9) ug/L
None
87%
Npreterm birth" 17
Nterm birth=127
p-value for
Mann-Whitney U
test: > 0.05
November 2012
5-564
Draft - Do Not Cite or Quote
-------
1 In a study taking place in California, women with information on blood Pb levels during
2 pregnancy based on their participation in a surveillance program (reason for participation
3 in the surveillance program was unknown but the authors speculate it was likely because
4 of potential Pb exposure due to occupational or environmental exposures or a family
5 member was identified as exposed to Pb) were matched with the birth certificates of their
6 infants (Jelliffe-Pawlowski et al.. 2006). Almost 70% of women had maximum blood Pb
7 measurements <10 (ig/dL with the majority being <5 (ig/dL. Preterm birth was associated
8 with higher blood Pb when comparing women with maximum pregnancy blood Pb levels
9 > 10 (ig/dL to women with blood Pb levels <10 (ig/dL in adjusted analyses. In analyses
10 of maximum Pb levels further refined into additional categories, the odds of preterm birth
11 were elevated among women with maximum blood Pb measurement > 20 (ig/dL
12 compared with women with maximum blood Pb levels < 5 (ig/dL. A study in Iran also
13 reported higher maternal blood Pb for preterm births than for term births fVigeh et al..
14 2011). The women in this study had lower blood Pb levels than did those observed in the
15 Jelliffe-Pawlowski et al. (2006). Higher maternal blood Pb level was associated with
16 higher odds of preterm birth. Another study examining blood Pb and gestational age
17 among women with lower blood Pb levels reported an inverse association between
18 maternal blood Pb concentration and gestational age, especially for blood Pb levels early
19 in pregnancy (Cantonwine et al.. 2010a). However, a study conducted in New York
20 among women with lower blood Pb levels (inclusion criteria mandated that blood Pb
21 levels be less than 10 (ig/dL), no association was observed between blood Pb levels and
22 preterm birth (Zhu et al.. 2010). Similarly, a study of maternal and paternal blood Pb
23 concentrations reported no association between maternal or paternal blood Pb levels and
24 preterm birth (Chen et al.. 2006a).
25 In another study, measurements of umbilical cord blood were taken after birth at a
26 hospital in Nagpur, India (Patel and Prabhu. 2009). A sample of women had their blood
27 Pb measured and among this sample, maternal blood Pb was correlated with the umbilical
28 cord Pb levels. Mean gestational age differed between infants with >5 (ig/dL cord blood
29 Pb and infants with < 5 (ig/dL cord blood Pb. In a linear regression model, gestational
30 age was found to decrease with increasing umbilical cord Pb levels. A study of women in
31 Tennessee consisted primarily of African American women living in an urban setting
32 (Jones et al.. 2010). The mean level of umbilical cord blood Pb was slightly higher
33 among infants born preterm but the difference was not statistically significant. Using
34 umbilical cord blood Pb measures, a study reported no association between cord blood Pb
35 levels and gestational age. The concentrations of cord blood Pb among study participants
36 were overall low (99.3% had umbilical cord blood Pb < 5 (ig/dL) (Wells et al.. 201 la).
November 2012 5-565 Draft - Do Not Cite or Quote
-------
1 A study of preterm birth included women living in two different residential areas over
2 three different time periods (Berkowitz et al.. 2006). One residential area had consistently
3 lower exposures but the other had a period of high Pb emissions due to damage at a local
4 factory (Pb measured in ambient air was up to 30 (ig/m3). Preterm birth rates were
5 examined during three time periods: before, during, and after the time of higher Pb
6 exposure. No association was observed between women living in the high exposure area
7 compared to those in the low exposure area during any of the exposure time periods, but
8 the number of preterm infants born during the period of higher exposure was small.
9 A study of breast milk in the second month postpartum reported no difference in breast
10 milk Pb levels for those infants born preterm or term; however, a limitation of this study
11 is that Pb levels were not measured until two months after the birth (Oriin et al., 2011).
12 In summary, as in the 2006 Pb AQCD, (U.S. EPA. 2006b) recent epidemiologic studies
13 report inconsistent findings for a relationship between indicators of Pb exposure and
14 preterm birth. No patterns were apparent within type of exposure measurement or Pb
15 level. Many of these studies are limited by the small number of preterm births and their
16 cross-sectional design (i.e., studies of umbilical cord blood may not adequately
17 characterized blood Pb levels earlier in pregnancy).A few studies utilized a longitudinal
18 cohort design (Vigeh etal.. 2011; Cantonwine etal.. 2010a; Chen et al.. 2006a: Jelliffe-
19 Pawlowski et al.. 2006). and although results among these studies were mixed some did
20 report an association between maternal blood Pb during pregnancy and preterm birth.
21 Most studies controlled for important confounders, such as maternal age and smoking.
5.8.3.4 Low Birth Weight/Fetal Growth
22 The 2006 Pb AQCD reported inconsistent epidemiologic study results for the
23 associations between Pb and birth weight/fetal growth but concluded that there could be a
24 small effect of Pb exposure on birth weight and fetal growth (U.S. EPA. 2006b). Since
25 then, multiple epidemiologic studies on the relationship between Pb exposure and birth
26 weight and fetal growth have been published using various measures of exposure, such as
27 air levels, umbilical cord blood, and maternal blood and bone. These studies are
28 summarized in Table 5-42 below (organized in the text and table by the type of Pb
29 measurement and then by study design). Additionally, there have been a few recent
30 toxicological studies evaluating the effect of Pb exposure during gestation on birth
31 weight.
November 2012 5-566 Draft - Do Not Cite or Quote
-------
Table 5-42 Summary of recent epidemiologic studies of associations between Pb levels and low birth weight
and fetal growth.
Reference
(Presented
in order of
appearance
in the text)
Gundacker
et al. (201 0)
Zhuetal.
(2010)
Study
Location Outcome
Vienna, Birth length,
Austria birth weight,
2005 nead
circumference
New York Birth weight,
2003-2005 sma" for
gestational age
(birth weight for
gestational age
<10th percentile
based on
national birth
weight by
gestational week
from weeks
25-42
Study
population
Infants of women
recruited during their
second trimester
N=53
Singleton births to
mothers aged 15-49
with blood Pb
measures before or
on the date of
delivery and blood
Pb measuring
<10 ug/dL
NLBw=2744
N. ~.,=40 544
normal BW ^w,^-r-r
NSGA=4092
Methodological
Details
Cohort study
using categorical
regression
Retrospective
cohort using linear
regression with
factional
polynomials for
birth weight and
logistic regression
with fractional
polynomials for
SGA
Pb Biomarkers
and Exposure
Measurement
Maternal blood Pb
between week 34-38
of gestation, whole
placentas and
umbilical cord Pb
shortly after birth,
meconium samples
in first five days after
birth
Maternal blood Pb
before or at delivery
Mean Pb
(SD)
in ug/dL
Median
(IQR):
Maternal
blood Pb'
2.5
(1.8,3.5)
Umbilical
cord blood
Pb:1.3
(0.8, 2.4)
Placenta
Pb:
25.8 ug/kg
(21.0,
36.8 ug/kg)
Meconium
Pb:
1 5.5 ug/kg
(9.8,
O~7 O i I/N l\fit\
27.9 ug/kg)
2.1
Adjusted Effect
Estimates
Regression
coefficients (units
not given, assume
results are per
10 ug/dLor
1 ug/kg)
Birth length:
Placenta Pb:
0.599
(SE0.154, p-value
<0.001)
Meconium Pb:
-0.385
(SE 0.157, p-value
0.012)
Birth weight:
Placenta Pb:
0.658
(SE 0.136, p-value
<0.001)
Maternal blood
Pb:
-0.262
(SE0.131, p-value
0.058)
Difference in
birthweight in
grams:
0 ug/dL: Ref
1 ug/dL: -27.4
(-37.8, -17.1)
2 ug/dL: -38.8
(-53.4, -24.1)
3 ug/dL: -47.5
(-65.4, -29.6)
4 ug/dL: -54.8
(-75.5, -34.2)
Potential
Confounders
Adjusted for in
Analysis
Model for birth length:
placenta Pb, gestational
age, meconium Pb
Model for birth weight:
gestational age,
placenta Pb, maternal
blood Pb
Model for birth weight:
Timing of Pb test,
maternal age, race,
Hispanic ethnicity,
education, smoking
status, alcohol use, drug
abuse, marital status,
financial assistance
program participation,
parity, infant sex
Model for SGA: Timing
of Pb test, maternal age,
November 2012
5-567
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Study
Location
Outcome
Pb Biomarkers Mean Pb
Study Methodological and Exposure (SD)
population Details Measurement in ug/dL
Adjusted Effect
Estimates
Potential
Confounders
Adjusted for in
Analysis
NnoSGA=39,084
5 ug/dL:-61.3
(-84.4, -38.2)
6 ug/dL: -67.2
(-92.5, -41.8)
7 ug/dL: -72.5
(-99.9, -45.2)
8 ug/dL: -77.6
(-106.8,-48.3)
9 ug/dL: -82.3
(-113.3,-51.2)
10 ug/dL: -86.7
(-119.4,-54.0)
After exclusion of
blood Pb
<1 ug/dL, a
1 ug/dL increase
in blood Pb was
associated with a
7.0 g decrease in
birthweight
Odd Ratios for
small for
gestational age:
< 1.0 ug/dL: 1.00
(Ref)
1.1-2.0 ug/dL:
1.07(0.98,1.17)
2.1-3.0ug/dL:
1.06(0.98, 1.16)
3.1-9.9 ug/dL:
1.07 (0.93, 1.23)
race, education,
smoking status, drug
abuse, marital status,
financial assistance
program participation,
parity, infant sex
November 2012
5-568
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Chen et al.
(2006a)
Study
Location Outcome
Taiwan Low birth weight
1993-1997 (<2,500g),
small for
gestational age
(birth weight
< 10th percentile
of sex- and
gestational week
weights for
singletons in
1993-1996)
Study
population
Infants born to at
least one parent
who was part of the
Program to Reduce
Exposure by
Surveillance System
- Blood Lead Levels
cohort that
monitored workers
occupationally
exposed to Pb
NLBW=72
Nnormal BW/I ,539 =
NSGA=135
NnoSGA=1,476
*738 births had
maternal Pb
information and 967
had paternal Pb
information
Pb Biomarkers Mean Pb
Methodological and Exposure (SD)
Details Measurement in ug/dL
Occupational Maternal blood Pb Maternal
cohort study using during pregnancy (or blood Pb
regression models if that wasn't 10.1(10.4)
available, the 1 year
prior to fertilization)
and/or paternal blood Paternal
Pb during blood pb
spermatogenesis 12.9(13.8)
(the 64 days before
fertilization, or if that
wasn't available, the
1 year prior to
spermatogenesis)
Adjusted Effect
Estimates
Risk Ratios
Low birth weight
Maternal blood Pb
<10 ug/dL: 1.00
(Ref)
10-19ug/dL:2.22
(1 .06, 4.26)
> 20 ug/dL: 1 .83
(0.67, 4.20)
Paternal blood Pb
<10ug/dL:1.00
(Ref)
10-19ug/dL:0.83
(0.34, 1 .75)
> 20ug/dL: 0.42
(0.12, 1 .06)
SGA
Maternal blood Pb
<10 ug/dL: 1.00
(Ref)
10-19ug/dL:1.62
(0.91 , 2.75)
>20ug/dL:2.15
(1.15,3.83)
Paternal blood Pb
<10ug/dL:1.00
(Ref)
10-19 ug/dL:0.94
(0.49, 1 .66)
> 20 ug/dL: 0.94
(0.51 , 1 .62)
Potential
Confounders
Adjusted for in
Analysis
Low birth weight models:
parental age, parental
education, infant sex,
parity
SGA models: parental
age, parental education
November 2012
5-569
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Jelliffe-
Pawlowski
et al. (2006)
Study
Location Outcome
California Low birth weight
1995-2002 (<2,500g)
Small for
gestational age
(birth weight for
gestational age
<10th percentile
of race- and
gender- specific
norms
Pb Biomarkers
Study Methodological and Exposure
population Details Measurement
Singleton births to Longitudinal Maximum maternal
non-smoking cohort study using blood Pb during
mothers with blood logistic regression pregnancy
Pb measures during
pregnancy from
either the California
Childhood Lead
Poisoning
Prevention Branch
or the California
Occupational Lead
Poisoning
Prevention Program
and matched to birth
records
NLBw=9
Nnormal BW=253
NsGA=17
NnoSGA=245
Mean Pb
(SD) Adjusted Effect
in ug/dL Estimates
>10ug/dL: Odd Ratios:
30.9% Low bjrth weight
<5 ug/dL:
1.00(Ref)
6-9 ug/dL:
10-19 ug/dL:
2.7(0.5,14.8)
20-39 ug/dL:
1.5(0.3,7.7)
> 40 ug/dL:
—
<10 ug/dL:
1.00(Ref)
> 10 ug/dL:
3.6 (0.3, 40.0)
Small for
gestational age
<5 ug/dL:
1.00(Ref)
6-9 ug/dL:
10-19ug/dL:
2.3(0.6,9.2)
20-39 ug/dL:
2.1 (0.7,6.7)
> 40 ug/dL:
<10 ug/dL:
1.00(Ref)
> 10 ug/dL:
4.2(1.3,13.9)
Potential
Confounders
Adjusted for in
Analysis
Adjusted for in 10 ug/dL
model for birth weight:
preterm birth, race,
insurance, parity,
maternal age, infant sex
Adjusted for in 10 ug/dL
model for SGA:
insurance, parity,
maternal age
November 2012
5-570
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Lambet al.
(2008)
Iranpouret
al. (2007)
Study Study
Location Outcome population
Mitrovica Height and BMI Participants of the
and at birth Yugoslavia Study of
Pristina, Environmental Lead
Yugoslavia Exposure,
1985-1986 Pregnancy
Outcomes, and
Childhood
Development
n=292
Isfahan, Low birth weight Full-term infants
Iran (<2,500g, born at a hospital
2005 >37wk) affiliated with
Isfahan University
NLBW32
N normal BW=34
Pb Biomarkers
Methodological and Exposure
Details Measurement
Population-based Mid-pregnancy blood
prospective cohort Pb
study using linear
regression
Cross-sectional Umbilical cord and
study using t-tests maternal blood Pb
and Spearman's within 12 h of
correlations delivery
Mean Pb
(SD)
in ug/dL
Mitrovica:
20.56 (7.38)
Pristina'
5.60 (1 .99)
Maternal
blood Pb:
Cases: 12.5
(2.0)
Controls:
13.5(2.7)
Umbilical
cord blood
Pb:
Cases: 10.7
(1.7)
Controls:
11 3 (1 9)
\ /
Adjusted Effect
Estimates
Regression
Coefficients (95%
Cl) for 1 ug/dL
increase in Pb:
BMI
Mitrovica:
-0.1 8 (-0.69, 0.33)
Pristina:
-0.1 4 (-0.69, 0.42)
Height
Mitrovica:
0.43 (-0.83, 1 .69)
Pristina:
0.35 (-0.64, 1 .34)
P-values fort-
tests:
Maternal blood
Pb: 0.07
Umbilical cord
blood Pb:
0.20
P-values for
correlations:
Maternal blood Pb
and Birth weight:
Low birth weight:
0.17
Normal birth
weight: 0.3
P-values for
correlations:
Umbilical cord
blood Pb and birth
weight:
Low birth weight:
0.84
Normal birth
weight: 0.26
Potential
Confounders
Adjusted for in
Analysis
Infant sex, ethnicity,
parity, maternal height or
BMI, maternal
education, gestational
age at blood sample,
gestational age at birth,
quality of home
environment
None
November 2012
5-571
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance Study
in the text) Location Outcome
Kordasetal. Mexico Head
(2009) City, circumference,
Mexico birth weight,
1994-1995 birth length
Afeiche et Mexico Birth weight
al. (2011) City
1 994-2005
Study
population
Infants of mothers
receiving antenatal
care at hospitals
serving low-to-
middle income
populations (cross-
sectional study of
baseline info from
Ca supplementation
trial)
N=474
Term, singleton
births, at least 2,500
grams enrolled in
one of three birth
cohorts recruited for
other longitudinal
studies
N=1,000
Pb Biomarkers
Methodological and Exposure
Details Measurement
Cross-sectional Umbilical cord and
study using linear maternal blood Pb
regression within 12 h of
delivery; maternal
tibia Pb 1 month
post-pa rtum
Cross-sectional Maternal patella and
study using tibia Pb measured at
varying coefficient 1 month postpartum
models with
random effects
Mean Pb
(SD)
in ug/dL
Maternal
tibia Pb:
9.9 ug/g
(9.8 ug/g)
Maternal
blood Pb
> 10ug/dL:
27%
Umbilical
cord blood
Pb
> 10ug/dL:
13.7%
Patella Pb
10.4
(11. 8) ug/g
Tibia Pb 8.7
(9.7) ug/g
Adjusted Effect
Estimates
Regression
coefficients (SE)
for each 1 ug/g
increase in tibia
Pb:
Birth weight: -4.9
C1 81
V ' -°f
Birth length: -0.02
(0.01)
Head
circumference:
-0.01 (0.01;
p-value <0.05)
Women with 4th
quartile tibia Pb
(15.6-76.5 ug/g)
delivered infants
1 40 g less than
women with tibia
Pb in the lowest
quartile
P (95% Cl) for 1
SD increase in
maternal patella
Pb
Girls:
-45.7 (-131 .7,
40.2)
Boys:
72.3 (-9.8, 154.4)
No association for
birth weight and
tibia Pb among
girls. A positive
association was
observed for tibia
Pb and birth
weight among
boys, (results not
given)
Potential
Confounders
Adjusted for in
Analysis
Maternal age, pre-
pregnancy BMI,
maternal height,
education, parity, marital
status, ever smoker,
postpartum calf
circumference,
gestational age, infant
sex
Birth cohort, maternal
age, maternal calf
circumference, maternal
height, education, parity,
breast feeding,
Ca + treatment group
assignment, gestational
age, height at birth,
repeated concurrent
child blood Pb measures
November 2012
5-572
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Cantonwine
etal.
(201 Ob)
Wells et al.
(2011 a)
Al-Saleh et
al. (2008b)
Study
Location Outcome
Mexico Birthweight
City
1994-1995
Baltimore, Birth weight
MD
2004-2005
Saudi Head
Arabia circumference
2004
Study
population
Infants who were
part of a clinical trial
to assess maternal
Ca2+-
supplementation on
bone Pb
mobilization during
lactation
N=533
Singleton births from
the Baltimore
Tracking Health
Related to
Environmental
Exposures (THREE)
study
Ni_Bw=33
Nnormal BW=267
Infants with a
gestational age of at
least 34 weeks born
to healthy mothers
aged 1 7-46 years
and non-
occupationally
exposed to Pb
N=653
Methodological
Details
Cross-sectional
study using linear
regression
Cross-sectional
study using
multivariable
linear regression
Cross-sectional
study using linear
regression
Pb Biomarkers
and Exposure
Measurement
Umbilical cord blood
Pb
Maternal tibia and
patella Pb one month
after delivery
Umbilical cord Pb
Umbilical cord blood
Pb
Mean Pb
(SD)
in ug/dL
Umbilical
cord blood
Pb varied by
genotype
from 6.3 to
6.9
Umbilical
cord blood
Pb
£ 10 ug/dL'
12.6%
0.84 (95%:
Cl 0.72,
0.96)
> 5 ug/dL:
0.7%
2.210
(1.691)
Umbilical
cord blood
Pb
>10 ug/dL:
1 .23%
Adjusted Effect
Estimates
Regression
models
P (95% Cl)
Umbilical cord
blood Pb: -31.1
(-105.4, 43.3)
Maternal tibia Pb
Overall: -4.4 (-7.9,
-0.9)
<1-4.1 ug/g: Ref
4.1-9.2 ug/g: 17.2
(-75.6, 110.1)
9.2-1 5.4 ug/g:
-19.1 (-112.1,
73.9)
15.4-43.2 ug/g:
-95.4 (-189.9, -0.8)
Ratio for Pb
concentration per
100g birth weight:
1 .01 (0.99, 1 .02)
Regression
models for those
above the 75th
percentile of cord
blood Pb levels
P (SE) per unit of
log-transformed
Pb
-0.158(0.718), p-
value: 0.036
Potential
Confounders
Adjusted for in
Analysis
Maternal age, education,
infant sex, maternal arm
circumference,
gestational age, smoking
status during pregnancy,
marital status, maternal
hemoglobin first month
postpartum, parity
Maternal age, race,
insurance, pre-
pregnancy weight,
smoking status,
gestational length, birth
weight, average year of
neighborhood home
construction
BMI, gestational age
Considered but not
included: prenatal
supplements, location
of residence
November 2012
5-573
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Atabeketal.
(2007)
Llanos and
Ronco
(2009)
Zentner et
al. (2006)
Janjua et al.
(2009)
Study
Location Outcome
Turkey Birth weight,
NS birth length,
head
circumference,
mid-arm
circumference
Santiago, Fetal growth
Chile restriction
NS (1 ,000-2,500g)
"note normal
birth weights
were >3,000g
Santo Birth weight and
Amaro, length
Brazil
2002
Karachi, Low birth weight
Pakistan (< 2,500g)
2005
Study
population
Term, singleton
infants born to
healthy mothers
living in urban areas
and assumed to
have high Pb
concentrations
N-E.A
— \Jt
Term births
(37-40 weeks) from
non-smoking
mothers
N -9D
'^growth restricted ^^*
N normal BW=20
Singleton births with
maternal residence
within 5 km of Pb
smelter
N=55
Infants of randomly
selected women
who planned to
deliver between
37-42 week
NLBw=100
Nnormal BW=440
Pb Biomarkers
Methodological and Exposure
Details Measurement
Cross-sectional Umbilical cord blood
study using linear Pb
regression
Cross-sectional Placenta Pb
study using Mann-
Whitney U-test
Cross-sectional Umbilical cord blood
study using linear Pb from delivery
regression
Cross-sectional Umbilical cord blood
study using Pb
binomial
regression
Mean Pb
(SD)
in ug/dL
14.4(8.9)
Umbilical
cord blood
Pb>
10 ug/dL:
53.7%
Umbilical
cord blood
Pb
> 25 ug/dL:
9.2%
Fetal growth
restricted:
0.21 ug/g
(0.04 ug/g)
Controls:
0.04 ug/g
(0.009 ug/g)
Umbilical
cord blood
Pb:
3.9 (3.6)
Umbilical
cord blood
Pb:
10.8(0.2)
Adjusted Effect
Estimates
Regression
models
P (p-value)
Birth weight:
-0.81 (0.01)
Birth length:
0.41 (0.05)
Mid-arm
circumference:
0.30 (0.05)
P-value for Mann-
Whitney U-test
<0.01
Linear regression
coefficient with
umbilical cord
blood Pb as the
dependent
variable in model
with only length
and weight (unit
not given, assume
per 1 ug/dL):
Length -0.46 (p-
value 0.003)
and Weight -0.275
(0.048)
(i.e., in this study,
Pb is assessed as
the outcome)
Prevalence ratio:
<10 ug/dL:
1 .00 (Ref)
a 10 ug/dL:
0.82
(0.57, 1.17)
Potential
Confounders
Adjusted for in
Analysis
Age, sex
Note: inclusion of SES
did not change the
results
None
No other variables
besides length and
weight were included in
the model
None
November 2012
5-574
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Jones et al.
(2010)
Orun et al.
(2011)
Study Study
Location Outcome population
Tennessee Low birth weight Singleton births
2006 (<2.500g) > 27 weeks
gestation from
mothers aged 16-45
living in the Shelby
County area for at
least 5 mo during
pregnancy
NLBw=10
N normal BW=92
Turkey Birth weight and Births to mothers
NS head not occupationally
circumference exposed to toxic
metals and living in
a suburban but non-
industrial area
NLBw=9
NnormalBW=135
Pb Biomarkers
Methodological and Exposure
Details Measurement
Cross-sectional Umbilical cord blood
study comparing Pb
across geometric
means (test not
specified)
Cohort study Breast milk 2 months
using Pearson post-partum
correlation
coefficients
Mean Pb
(SD)
in ug/dL
2.4 (4.3)
Geometric
mean' 1 3
Median:
20.6 ug/L
>WHO limit
(5 ug/L):
87%
Median
(IQR)
<2500g:
20.4 (8.5,
27.1) ug/L
> 2500g:
20.6(11.8,
29.5) ug/L
Potential
Confounders
Adjusted Effect Adjusted for in
Estimates Analysis
Geometric Mean: None
Low birth weight:
1 .2
Normal birth
weight: 1.3
p-value for
difference: >0.10
Correlations for None
breast milk Pb and
z-scores of head
circumference
Girls: 0.087
Boys: 0.029
Correlations for
breast milk Pb and
z-scores of birth
weight
Girls: 0.097
Boys: 0.045
*AII p-values for
correlations>0.05
November 2012
5-575
Draft - Do Not Cite or Quote
-------
Reference
(Presented
in order of
appearance
in the text)
Williams et
al. (2007)
Berkowitz et
al. (2006)
Study
Location Outcome
Tennessee Birth weight
2002
Idaho Low birth weight
1970-1981 (<2,500gand
> 37 week)
Small for
gestational age
(birth weight
£ 5th percentile
of sex- and
gestational week
weights for
singletons in
Idaho)
Study
population
Infants from
singleton births or
the firstborn infant in
a set of multiples
N=not specified
Singleton infants
with 28-45 week
gestation
NLBW=4297
NnormalBW= 162,035
NSGA=7020
NnoSGA=1 62,035
Pb Biomarkers
Methodological and Exposure
Details Measurement
Longitudinal Air Pb levels during
cohort study using first trimester of
hierarchical linear pregnancy
models
Cohort study Three time periods of
using logistic two locations
regression (unexposed and
exposed/near
smelter): pre-fire,
"high-exposure
period" (when a fire
happened at the
smelter and resulted
in damages leading
to high air Pb
concentrations for 6
mo), and "post-fire"
Mean Pb
(SD)
in ug/dL
0.12 ug/m3
(0.04 ug/m3)
During the
time of the
fire,
estimates of
Pbin
ambient air
were as high
as 30 ug/nr
Adjusted Effect
Estimates
p-value for
multilevel
regression of Pb
with birth weight:
0.002
Increase of Pb
from 0 to 0.04
relates to a 38g
decrease in birth
weight
Increase of Pb
from 0 to 0.13
(maximum) relates
to a 1 24g
decrease in birth
weight
Term Low birth
weight:
OR (90% Cl)
(unexposed
location is referent
group):
Pre-fire: 0.81
(0.55, 1 .20)
High exposure:
2.39(1.57,3.64)
Post-fire: 1.28
(0.95, 1 .74)
Small for
gestational age:
OR (90% Cl)
(unexposed
location is referent
group):
Pre-fire: 0.98
(0.73, 1 .32)
High exposure:
1.92(1.33,2.76)
Post-fire: 1.32
(1.05,1.67)
Potential
Confounders
Adjusted for in
Analysis
Previous preterm birth,
previous birth>4000g,
pregnancy-induced
hypertension, chronic
hypertension,
oligohydramios, other
maternal risk factors,
education,
cigarettes/day, black
race, Hispanic ethnicity,
other race/ethnicity,
plurality, infant sex, first
trimester SO2, within
5km of an air monitor,
poverty, interaction of
poverty and other
maternal risk factors,
percentage of previous
pregnancies that
resulted in non-live
births
Maternal age, infant sex,
first birth, previous
miscarriage or abortion
November 2012
5-576
Draft - Do Not Cite or Quote
-------
1 Multiple studies were conducted that examined the association between maternal blood
2 Pb and birth weight/fetal growth. A study in Vienna, Austria reported an inverse
3 association between maternal blood Pb levels and birth weight but no associations for
4 birth length or head circumference (Gundacker et al.. 2010). Similarly, increased
5 maternal blood Pb was associated with decreased birth weight among infants in a study
6 performed in New York (Zhu et al.. 2010). No association was observed between
7 maternal blood Pb levels and SGA. A study in Taiwan examined both maternal and
8 paternal blood Pb levels among those occupationally exposed to Pb and their associations
9 with birth weight and SGA (Chen et al.. 2006a). Paternal blood Pb levels were not
10 associated with increased risk of low birth weight or SGA. Higher maternal blood Pb
11 concentration was associated with higher risk of low birth weight and SGA, although not
12 all of the associations were statistically significant. There were small numbers of infants
13 with low birth weight or SGA, especially at the highest blood Pb levels (> 20 (ig/dL). In
14 California, blood Pb measurements of women during pregnancy were matched with the
15 corresponding birth certificates (Jelliffe-Pawlowski et al.. 2006). The adjusted OR for
16 low birth weight that compared women with blood Pb levels > 10 (ig/dL to women with
17 levels <10 (ig/dL was elevated. However, it was difficult to draw conclusions about the
18 relationship between blood Pb and birth weight due to small numbers (n = 9 for low birth
19 weight) and the subsequently wide 95% CI. An association was detected for increased
20 blood Pb and having an infant who was small for his/her gestational age (SGA). Women
21 residing in two different towns in Yugoslavia (one with a Pb smelter and one without a
22 Pb smelter) were recruited during their first prenatal visit (Lamb et al., 2008) (study
23 based on previous work by Factor-Litvak et al. (1991)). The mid-pregnancy blood Pb
24 levels were greater in women from the town with a Pb smelter. No association was
25 reported between maternal blood Pb and height or BMI at birth for the infants of these
26 women despite the differences in maternal blood Pb between the two towns. A study of
27 term births in Iran reported no difference in blood Pb levels of women giving birth to a
28 normal weight infant and women giving birth to an infant with low birth weight (Iranpour
29 et al.. 2007).
30 A study examining the association between Pb biomarker levels and birth weight used
31 tibia bone measurements one month post-partum from mothers living in Mexico City
32 (Kordas et al.. 2009). Tibia Pb levels were inversely associated with birth weight but not
33 with birth length. This association between Pb and birth weight was not modified by
34 maternal folate consumption or maternal or infant MTHFR genotype, although the
35 association between tibia Pb levels and birth weight was greater in magnitude among
36 women with certain MTHFR SNPs (statistical tests not reported). Another study in
37 Mexico City reported no association between maternal tibia Pb levels and birth weight
38 among girls but reported a positive association for boys (Afeiche et al.. 2011). No
39 associations were observed with maternal patella Pb concentration, although among boys,
November 2012 5-577 Draft - Do Not Cite or Quote
-------
1 the relationship was positive but not statistically significant. One of the cohorts used by
2 Afeiche et al. (2011) was also evaluated in another study (Cantonwine et al.. 2010b). An
3 inverse association was observed between tibia Pb and birth weight, especially at higher
4 levels (over 15.4 (ig/dL). This association was stronger among those mothers with
5 variants of the hemochromatosis iron gene (HFE).
6 Multiple studies examined the relationship between Pb level and birth weight using Pb
7 measured from the placenta or umbilical cord. A study performed in Baltimore, MD
8 reported no association between umbilical cord blood Pb concentration and birth weight
9 (Wells etal.. 201 la). This study had low blood Pb levels, with only 0.7% of participants
10 having umbilical cord blood Pb measuring >5 (ig/dL. In Saudi Arabia, a study was
11 conducted among non-occupationally exposed women (Al-Saleh et al., 2008b). Umbilical
12 cord blood Pb concentrations were low and an association was observed between
13 umbilical cord Pb and head circumference. A study with high Pb concentrations in
14 umbilical cord blood reported an inverse association between Pb levels and birth weight
15 (Atabek et al., 2007). However, no correlation was detected in an analysis restricted to
16 umbilical cord Pb less than 10 (ig/dL. No associations with other measures of growth,
17 such as birth length and mid-arm circumference, were detected. Researchers in Chile
18 collected the placentas from term births and compared the Pb levels for those born with
19 normal birth weights to those with low birth weights (Llanos and Ronco. 2009). Pb levels
20 were greater in the placentas of infants with low birth weights. In addition, the authors
21 note that 3 low birth weight infants had extremely high Pb levels in the placentas
22 (>1.5 (ig/g) and were excluded from these analyses. A study in Brazil examined Pb levels
23 in umbilical cord blood from term births of women residing within 5 km of a Pb smelter
24 (Zentner et al.. 2006). The cord blood Pb level was found to be inversely correlated with
25 length and weight of the infants. Another study recruited women in Pakistan (Janjua et
26 al.. 2009). Umbilical cord blood Pb levels were not associated with low birth weight. The
27 study by Iranpour et al. (2007) discussed above investigated the association with
28 umbilical cord blood Pb levels in addition to their examination of maternal whole blood
29 Pb. They again report no difference in levels between term infants of normal and low
30 birth weight. A study comparing geometric mean umbilical cord blood Pb levels reported
31 no difference in the levels for normal and low birth weight infants born to women living
32 primarily in urban areas of Memphis, TN (Jones etal.. 2010). A study previously
33 mentioned that observed an inverse association between maternal tibia Pb and birth
34 weight in Mexico City reported no association between umbilical cord blood Pb
35 concentration and birth weight (Cantonwine et al., 2010b). Finally, a study in Vienna
36 measured Pb in the placenta (Gundacker et al.. 2010). A positive correlation was
37 observed between placenta Pb and birth length and weight; however, in the same study,
38 maternal blood Pb was inversely related to birth weight.
November 2012 5-578 Draft - Do Not Cite or Quote
-------
1 A study performed in Turkey examined the relationship between Pb levels in breast milk
2 two months postpartum and size at birth (Orim et al.. 2011). No association was observed
3 between breast milk Pb concentration and birth weight or head circumference.
4 A few studies examined air exposures and reported inverse associations between air Pb
5 concentrations and birth weight. However, a limitation of these studies is the difficulty in
6 assessing if the measured concentrations represent population exposures (see
7 Section 3.5.3). Williams et al. (2007) examined Pb concentrations in the air during the
8 first trimester. The purpose of their study was to demonstrate the use of hierarchical
9 linear models and they used the example of air pollution and birth weight in Tennessee.
10 The model results showed an association between ambient Pb concentration and birth
11 weight, with an estimated decrease in birth weight of 38 grams for every 0.04 (ig/m3
12 (i.e., one standard deviation) increase in Pb concentration. Another study of air Pb levels
13 was conducted in Idaho and included two areas over three time periods. One study area
14 was affected by damage to a local factory that led to high Pb emissions during one of the
15 time periods under study (Berkowitz et al.. 2006). During the time of the fire, estimates
16 of Pb in ambient air were as high as 30 (ig/m3. Mean birth weight forterm births was
17 decreased among infants born to women living in the high exposure area during the
18 period of high exposure compared to those living in the lower exposure area. The
19 difference in birth weight of term births remained, but was reduced, between the two
20 areas during the time period after the exposure ended. During the period of higher
21 exposure, the odds of low birth weight among term births was increased among those
22 living in the higher exposed area compared to those in the lower exposed area, but the
23 odds were not different between the two study areas during the time periods before or
24 after the high level of exposure. An increase in SGA infants (defined as infants with
25 weights less than or equal to the lowest 5th percentile of birth weight for their sex and
26 age) was also associated with living in the higher exposed area during the time period of
27 higher exposure. The odds of SGA infants decreased during the time period after the
28 exposure but the odds were still elevated compared to those residing in the lower exposed
29 area.
30 Evidence from previous toxicological studies has shown an association between
31 gestational Pb exposure and reduced birth weight (U.S. EPA. 2006b). More recent studies
32 have reported conflicting results. Wang et al. (2009e) demonstrated a statistically
33 significant decrease in fetal body weight and body length of Wistar rats after maternal
34 exposure to 250 ppm Pb acetate during gestation days 1-10, 11-20, or 1-20. The greatest
35 decrease in fetal body weight and length was observed in the group exposed to Pb during
36 gestation days 1-20 followed by the group exposed to Pb during gestation days 11-20.
37 Teijon et al. (2006) observed that when pregnant dams were administered 200 ppm or
38 400 ppm Pb acetate in drinking water, litter weight was significantly decreased (400 ppm
November 2012 5-579 Draft - Do Not Cite or Quote
-------
1 Pb only) versus controls due to significant decrements in female pup birth weight; male
2 birth weight was unaffected. The results of these studies indicate that as Pb exposure
3 increases, the body weight of exposed offspring decreases. Masso-Gonzalez and
4 Antonia-Garcia (2009) also observed an 8-20% decrease in body weight of pups from rat
5 dams given 300 mg/L Pb acetate in drinking water (exposure during gestation and
6 lactation resulting in mean blood Pb level of 22.8 (ig/dL), but no changes in body length
7 were reported.
8 In summary, associations were observed between Pb and low birth weight in
9 epidemiologic studies of maternal bone Pb and studies of Pb air exposures and birth
10 weight. The associations were less consistent when using maternal blood Pb or umbilical
11 cord and placenta Pb as the exposure measurement although some studies did
12 demonstrate associations. Epidemiologic studies of Pb and fetal growth face multiple
13 limitations. One limitation is the cross-sectional nature of many studies. These do not
14 allow an understanding of the temporality for Pb and fetal growth. In addition, some
15 studies suffer from small sample size. The studies of air Pb levels and birth weight
16 demonstrate positive associations but are limited in that individual exposure levels are
17 unknown. Also, many of the studies controlled for important confounders, such as parity
18 and gestational age, but adjustment in some studies was lacking. Previous toxicological
19 studies observed an association between gestational Pb exposure and reduced birth
20 weight with moderate to high dose Pb.
5.8.4 Effects on Male Reproductive Function
21 The 2006 Pb AQCD (U.S. EPA. 2006b) reported on male Pb exposure or biomarker
22 levels and reproductive functions in males as measured by sperm
23 count/motility/morphology, time to pregnancy, reproductive history, and chromosomal
24 aberrations. Despite limitations, most of the studies found slight associations between
25 high blood Pb levels (i.e., > 45 (ig/dL) and reduced male fecundity or fertility (U.S. EPA.
26 2006b). Evidence reviewed in the 1986 Pb AQCD (U.S. EPA. 1986a) also demonstrated
27 that Pb exposure affects male reproductive function in humans and experimental animals.
28 Recently published research has continued to support an association between Pb and
29 sperm/semen production, quality, and function. Studies of Pb and male reproductive
30 function are described in the sections below.
November 2012 5-580 Draft - Do Not Cite or Quote
-------
5.8.4.1 Effects on Sperm/Semen Production, Quality, and
Function
1 Multiple epidemiologic and toxicological studies have examined the relationship between
2 Pb and sperm and semen production, quality, and function. These studies are summarized
3 in the text below. In addition, recent epidemiologic studies are included in Table 5-43.
4 All epidemiologic studies were cross-sectional with concurrent measurements of Pb
5 levels in biological samples and sperm-related outcomes.
November 2012 5-581 Draft - Do Not Cite or Quote
-------
Table 5-43 Summary of recent epidemiologic studies of associations between Pb levels and effects on sperm
and semen.
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Hsuetal.
(2009b)
Study
Location
and
Years
Taiwan
NS
Study Population
Men working at a
battery plant
N=80
Methodological
Details
Occupational cohort
study (cross-sectional)
using ANOVA and
linear regression
Pb Biomarker
or Exposure
Measurement
Blood Pb
Categorized into 3
groups:
<25 ug/dL,
25-45 ug/dL,
>45 ug/dL
Mean Pb
(SD)
in ug/dL
40.2
Adjusted Effect Estimates
p-values for difference across the
three groups were <0.05 for:
sperm head abnormalities, sperm
neck abnormalities, sperm
chromatin structure assay
(aT, COMPaT)
p-values for difference across the
Potential
Confounders
Adjusted for in
Analysis
Smoking status
three groups were >0.05 for:
semen volume, sperm count,
motility, sperm tail abnormalities,
sperm immaturity, computer-
assisted semen analysis, % sperm
with ROS production
Coefficients for regression
analysis with blood Pb:
Morphologic abnormality 0.271 (p-
value O.0001)
Head abnormality 0.237 (p-value
0.0002)
aT 1.468 (p-value 0.011)
COMPaT 0.233 (p-value 0.21)
Kasperczyk et Poland
al. (2008) NS
Healthy, non-smoking,
fertile men that worked
at the Zn and Pb
Metalworks
NControls=14
' *low exposure"*^
l^high exposure"*^*?
Occupational cohort
study (cross-sectional)
using Kruskal-Wallis
ANOVA and
Spearman's coefficient
for non-parametric
correlation
Blood Pb; seminal
fluid Pb
Categorized as:
high exposure
workers
(blood Pb
40-81 ug/dL),
low exposed
workers
(blood Pb
Blood Pb
High
exposure
workers:
53.1 (2.05)
Low
exposure
workers:
34.7 (0.83)
Controls:
8.47 (0.54)
Mean (SE) None
Sperm volume (ml)
Controls:
2.94 (0.32)
Low exposure:
2.89 (0.22)
High exposure:
2.98 (0.22)
(p-value for ANOVA: 0.993)
November 2012
5-582
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Study
Location
and
Years Study Population
Methodological
Details
Pb Biomarker
or Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted Effect Estimates
Potential
Confounders
Adjusted for in
Analysis
25-40 ug/dL),
and controls
(office workers
with no history of
occupational Pb
exposure)
Seminal
plasma Pb
High
exposure
workers:
2.02 (0.23)
Low
exposure
workers:
2.06 (0.40)
Controls:
1.73(0.16)
Sperm cell count (mln/mL)
Controls:
43.1 (7.0)
Low exposure:
44.6(10.1)
High exposure:
42.2 (5.86)
(p-value forANOVA: 0.400)
Normal morphology (%)
Controls:
63.3 (2.7)
Low exposure:
57.3 (2.5)
High exposure:
58.4(2.1)
(p-value forANOVA: 0.266)
Progressively motile sperm
after 1 h (%)
Controls:
16.4(3.2)
Low exposure:
14.8(2.6)
High exposure:
10.5(1.9)
(p-value forANOVA: 0.217)
Motile sperm after 24 h (%)
Controls:
4.4(1.8)
Low exposure:
7.3(1.7)
High exposure:
3.1 (0.8)
(p-value forANOVA: 0.188)
p-value for correlation between
blood Pb and sperm cell motility
after! h: 0.011
November 2012
5-583
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Naha and
Manna (2007)
Naha and
Chowdhury
(2006)
Study
Location
and
Years Study Population
Bangalore, Non-occupationally
India exposed controls and
NS occupationally exposed
workers
NControls=50
'^low exposure"^"-"
N-9D
nigh exposure *-**
Kolkata, Men aged 31 -45 that
India were non-
NS occupationally exposed
controls and
occupationally exposed
workers)
NControls=50
'^low exposure"^"-"
N-^n
nigh exposure ^u
Methodological
Details
Occupational cohort
study using ANOVA,
Student's t-test, and
Scheffe's F-test
Occupational cohort
study using ANOVA,
Student's t-test, and
Scheffe's F-test
Pb Biomarker
or Exposure
Measurement
Categorized by
work history as
controls, low
exposure (7-10 yr
of exposure for 8
h/day) and high
exposure (>10 yr
of exposure for 8
h/day)
Categorized by
work history as
controls, low
exposure (7-1 0 yr
of exposure for 8
h/day) and high
exposure (>10yr
of exposure for 8
h/day)
Mean Pb
(SD)
in ug/dL
Blood Pb
measurement
Controls
10.25(2.26)
Low
exposure
50.29 (3.45)
High
exposure
68.26 (2.49)
Semen Pb
measurement
Controls 2.99
(0.76)
Low
exposure
15.85(1.95)
High
exposure
25.30 (2.28)
Blood Pb
measurement
Controls
13.62 (2.45)
Low
exposure
48.29 (4.91)
High
exposure
77.22 (1 .25)
Semen Pb
measurement
Controls 3.99
(1.36)
Low
exposure
10.85(0.75)
High
exposure
18.30(2.08)
Potential
Confounders
Adjusted for in
Adjusted Effect Estimates Analysis
p-values for difference across the None
three groups for mean values of
semen profiles were <0.01 for:
liquefaction time, seminal volume,
sperm count, sperm DNA hyploidy,
sperm morphological abnormality,
sperm motility, sperm ATPase
activity, seminal plasma fructose,
seminal plasma total protein,
seminal plasma free amino acid,
seminal plasma cholesterol
p-values for difference across the None
three groups for mean values of
semen profiles were <0.01 for:
sperm count, sperm protein,
sperm DNA hyploidy, sperm DNA,
sperm RNA, sperm viability, sperm
membrane lipid peroxidation,
seminal plasma total ascorbate,
seminal plasma DHAA, sperm
ATPase activity, sperm motility,
sperm velocity, seminal plasma
fructose
November 2012
5-584
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Study
Location
and
Years Study Population
Methodological
Details
Pb Biomarker
or Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted Effect Estimates
Potential
Confounders
Adjusted for in
Analysis
Telisman et al. Croatia Men aged 19-55, never
(2007) 2002-2005 occupationally exposed
to metals and going to a
clinic for infertility
examination or for
semen donation to be
used for artificial
insemination
N=240
Cross-sectional study
using linear multiple
regression
Blood Pb
Median: 4.92 Standardized regression
(range coefficients for log blood Pb (units
1.13-14.91) not given)
Immature sperm: 0.13 (p-value
<0.07)
Pathologic sperm: 0.31 (p-value
<0.0002)
Wide sperm: 0.32 (p-value
O.0001)
Round sperm: 0.16 (p-value
<0.03)
Cd, Cu, Zn, Se,
age, smoking
status, alcohol use
Coefficients and p-values not
given if not statistically significant:
semen volume, sperm
concentration, slow sperm, short
sperm, thin sperm, amorph sperm
November 2012
5-585
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Study
Location
and
Years Study Population
Methodological
Details
Pb Biomarker
or Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted Effect Estimates
Potential
Confounders
Adjusted for in
Analysis
Meeker et al. Michigan Men aged 18-55 going
(2008) NS to infertility clinics
(distinction not made
between clinic visits for
male or female fertility
issues)
N=219
Cross-sectional study
using multiple logistic
regression
Blood Pb
Median: 1.50 OR (95% Cl) for having below
(IQR 1.10, reference-level semen parameters
2-°°) Concentration
Istquartile: 1.00 (ref)
2nd quartile: 0.88 (0.32, 2.44)
3rd quartile: 2.58 (0.86, 7.73)
4th quartile: 1.16(0.37,3.60)
Age, smoking
status
Models with
multiple metals
included: smoking
status, Mo, Mn,
Cd, and Hg
Motility
Istquartile: 1.00 (ref)
2nd quartile: 1.04(0.43,2.53)
3rd quartile: 1.95(0.70,5.46)
4th quartile: 1.66(0.64,4.29)
Considered but did
not include: BMI,
race
Morphology
Istquartile: 1.00 (ref)
2nd quartile: 0.83 (0.37, 1.87)
3rd quartile: 1.41 (0.54,3.67)
4th quartile: 1.18(0.50,2.79)
Models with adjustment for
multiple metals
Concentration
Istquartile: 1.00 (ref)
2nd quartile: 0.89 (1.57, 2.89)
3rd quartile: 3.94(1.15, 13.6)
4th quartile: 2.48(0.59, 10.4)
November 2012
5-586
Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Slivkova et al.
(2009)
Study
Location
and
Years
NS
Study Population
Men aged 22-48
undergoing semen
analysis at an infertility
clinic
Methodological
Details
Cross-sectional study
using correlation
Pb Biomarker
or Exposure
Measurement
Semen Pb
Mean Pb
(SD)
in ug/dL
1 .49 mg/kg
(0.40 mg/kg)
Adjusted Effect Estimates
Correlation between Pb and
flagellum ball : -0.39 (p-value not
given)
Potential
Confounders
Adjusted for in
Analysis
None
'correlations not given for any
N=47 other sperm pathological changes
(therefore assume not statistically
significant): broken flagellum,
separated flagellum, separated
flagellum, small heads, retention
of cytoplasmic drop, other
pathological spermatozoa, large
heads, acrosomal changes, and
knob twisted flagellum
November 2012 5-587 Draft - Do Not Cite or Quote
-------
Reference
(Studies are
presented in
order of first
appearance in
the text of this
section)
Mendiola et al.
(2011)
Study
Location
and Methodological
Years Study Population Details
Spain Men attending infertility Case-control study
2005-2007 clinics and classified as using multiple linear
either normal sperm regression
(controls) or oligo-
astheno-
teratozoospermia
(cases) based on WHO
semen quality criteria
NControls=31
NCases=30
Pb Biomarker Mean Pb
or Exposure (SD)
Measurement in ug/dL
Seminal plasma Seminal
Pb plasma: 2.90
OQR2.70,
Whols blood*
9.50 (IQR
7.50, 11.90)
Blood
plasma: 2.90
(IQR 2.70,
3.10)
Cases:
Seminal
plasma* 3 0
(0.3)
Whole blood:
9.8 (2.3)
Blood
plasma: 2.9
(0.2)
Controls:
Seminal
plasma: 2.9
(0.3)
Whole blood:
9.7 (2.3)
Blood
plasma: 2.9
(0.3)
Potential
Confounders
Adjusted for in
Adjusted Effect Estimates Analysis
P (95% Cl) Age, BMI, number
Sperm concentration of cigarettes/day
Seminal plasma: -1 .0 (-3.1 , 2.3)
Whole blood: -0.2 (-1.7, 1.6)
Blood plasma: 0.08 (-4.1 , 5.2)
% Immotile sperm
Seminal plasma: 1 .5 (0.37, 1 .9)
Whole blood: 0.05 (-0.32, 0.43)
Blood plasma: -0.49 (-1 .8, 0.62)
% Morphologically normal sperm
Seminal plasma: -0.54 (-3.1 , 2.0)
Whole blood: -0.31 (-1.5, 0.89)
Blood plasma: -0.08 (-3.5, 3.4)
"Units not given (assume 1 ug/dL)
Note: No correlation in Pb levels
among bloods or seminal plasma.
There was correlation between Pb
and other metals (Cd and Hg)
within each body fluid. Other
metals were not controlled for in
models
November 2012
5-588
Draft - Do Not Cite or Quote
-------
1 International epidemiologic studies of men occupationally exposed to Pb have reported
2 on associations between Pb exposure or biomarker levels and sperm count and quality
3 and semen quality. In most of these occupational studies, mean blood Pb levels over
4 40 (ig/dL have been reported for individuals occupationally exposed to Pb. In addition,
5 they did not control for other potential occupational exposures. A study performed in
6 Taiwan among men with high levels of blood Pb reported that men with higher blood Pb
7 levels had increased sperm head abnormalities, increased sperm DNA denaturation, and
8 increased sensitivity to denaturation compared to men with lower blood Pb levels (Hsu et
9 al.. 2009b). No difference was detected between three Pb exposure groups and semen
10 volume, sperm count, motility, velocity, and reactive oxygen species production. A
11 similar study in Poland included employees exposed to Pb and compared them with a
12 group of male office workers (Kasperczyk et al., 2008). Pb levels measured in seminal
13 fluid were slightly higher among those in the exposed groups although they were not
14 statistically different from the levels in the control group. No difference was observed for
15 semen volume, sperm count, or sperm morphology among the groups. Sperm motility
16 was lower in the highest exposure group compared to both the control and moderate
17 exposure groups. Lipid peroxidation, which can induce tissue damage in sperm via
18 reactive oxygen species, was greater in the highest exposure group compared to the
19 controls. Studies performed in India (Naha and Manna. 2007; Naha and Chowdhury.
20 2006) reported that men in the highest exposure group (men working in battery or paint
21 manufacturing plants for 10-15 years for 8 hours/day) had mean blood Pb levels of
22 77.22 (ig/dL (Naha and Chowdhury. 2006) and 68.26 (ig/dL (Naha and Manna. 2007).
23 Control groups in these studies (those without occupational Pb exposure) had mean blood
24 Pb levels below 15 (ig/dL. Increases in levels of Pb in semen were also noted across
25 exposure groups. Both studies report decreases in sperm count and in sperm velocity and
26 motility with increasing Pb exposure. Higher Pb exposure was also associated with
27 greater hyploidy of sperm DNA and morphologic abnormalities (Naha and Manna. 2007;
28 Naha and Chowdhury. 2006). Decreased viability and increased lipid peroxidation were
29 detected (Naha and Chowdhury. 2006).
30 A few studies examined blood or seminal plasma Pb levels and semen quality of men at
31 infertility clinics (Mendiolaet al., 2011; SlivkovaetaL 2009; Meeker etal., 2008)
32 (Telisman et al.. 2007). In general, these men had lower levels of Pb biomarkers than
33 men who were occupationally exposed, but the studies are limited by the strong
34 possibility of selection bias related to the recruitment of men attending infertility clinics.
35 A study performed in Croatia recruited men attending a clinic for infertility examination
36 or to donate semen for use in artificial inseminations, who had never been occupationally
37 exposed to metals and therefore had lower blood Pb levels than the occupational studies
38 (although leaded gasoline was still sold during the study period) (Telisman et al.. 2007).
39 Increased blood Pb was associated with increased percentages of pathologic sperm, wide
November 2012 5-589 Draft - Do Not Cite or Quote
-------
1 sperm, and round sperm. There was also a slight increase in immature sperm although it
2 was not statistically significant. Similar results were seen when other biomarkers for Pb
3 (erythrocyte protoporphyrin and 5-aminolevulinic acid dehydratase [ALAD]) were used
4 instead. This study controlled for multiple potential confounders, including other metals.
5 Meeker et al. (2008) detected no associations between higher blood Pb and semen
6 concentration, morphology, or motility (although a slight positive trend was observed
7 between higher Pb levels and motility in unadjusted models). In models that include
8 multiple metals, blood Pb was associated with being below the WHO limit of sperm
9 concentration levels (less than 20 million sperm/mL), although the 95% CI: was wide for
10 the 4th quartile of Pb levels and included the null. The precision of estimates in this study
11 was extremely low. Slivkova et al. (2009) reported a negative correlation between semen
12 Pb and pathological changes in sperm (specifically, flagellum ball), but no correlations
13 were observed for other alterations in the sperm. Another study reported a positive
14 association between seminal plasma Pb concentration and percentage of immotile sperm,
15 but this analysis did not adjust for exposure to other metals reported to be correlated with
16 Pb concentration in the seminal plasma (Mendiola et al.. 2011). No association was
17 observed for seminal plasma Pb concentration and sperm concentration or percentage of
18 morphologically normal sperm. Additionally, neither Pb levels in whole blood nor
19 plasma were associated with sperm concentration, percentage of immotile sperm, or
20 percentage of morphologically normal sperm.
21 Extensive evidence in the toxicological literature demonstrates that Pb exposure is
22 detrimental to the quality and overall health of testicular germ cells, affecting sperm and
23 semen quality and production. Earlier Pb AQCDs contained studies of Pb-induced
24 decreased sperm counts, decreased sperm production rate, and dose-dependent
25 suppression of spermatogenesis in adult rodents with 30 day drinking water Pb exposure
26 lYSokol and Berman. 1991). blood Pb level 35 and 37 (ig/dL; (Sokoletal.. 1985). blood
27 Pb level 34 (ig/dL; (Sokol. 1989). blood Pb levels <43 (ig/dL]. Chronic Pb exposure
28 (15 weeks) in adult male rabbits, resulting in blood Pb of 24 (ig/dL, induced statistically
29 significant decrements in semen quality and greater testicular pathology (Moorman et al..
30 1998) with dosing by subcutaneous injection, loading dose of 0.2-3.85 mg/kg BW
31 Monday (M), Wednesday (W) and Fridays (F) weeks 6-10, followed by maintenance
32 dose of 0.13-2.0 mg/kg BW Pb acetate MWF over weeks 11-20 of the study. The
33 2006 Pb AQCD also cited studies in which sperm from Pb exposed rats yielded lower
34 rates of fertilization when used for in vitro fertilization of eggs harvested from unexposed
35 females [(Sokoletal.. 1994). blood Pb level 33-46 (ig/dL].
36 Recent studies corroborate earlier findings that Pb alters sperm parameters such as sperm
37 count, viability, motility, and morphology. Anjum et al. (2010) exposed 50 day old male
38 albino Wistar/NIN rats to Pb acetate (273 or 819 mg/L in drinking water, 500 or
November 2012 5-590 Draft - Do Not Cite or Quote
-------
1 1,500 ppm, respectively) for 45 days. Affected endpoints included reduced epididymal
2 sperm count, motile sperm, and viable sperm, indicating decreased sperm production and
3 quality. Anjum did not report blood Pb values. Wistar/NIN rats (1,500 ppm Pb acetate in
4 drinking water for 70 days) supplemented with the herb Centella asiatica (Sainath et al..
5 2011) had significant attenuation of the Pb-induced changes observed by Anjum et al.
6 (2010). Pillai et al. (2012) found gestational and lactational treatment with Pb acetate in
7 Charles Foster rats (subcutaneous injection of 0.05 mg/kg BW/day) induced effects on
8 sperm in adults (PND65) including significant decreases in testicular sperm count,
9 epididymal sperm count, and sperm motility. These findings are consistent with
10 Pb-associated effects on sperm and male reproductive organs in wildlife from the
11 ecological literature including deer, Asian earthworms, rainbow trout, marine worms, and
12 the fathead minnow (see Sections 7.3.12.1 and 7.4.12.1 from the ecology terrestrial and
13 aquatic reproduction sperm sections).
14 Pb exposure has been shown to affect the male reproductive organs, as is seen with
15 histological or morphological changes. Studies included in previous Pb AQCDs showed
16 that histological and ultrastructural damage to the testes or seminiferous tubules was seen
17 in non-human primates with chronic oral Pb exposure (daily Pb exposure, gelatin
18 capsule; control plus 3 treatment groups: (1) infancy exposure group [PNDO-PND400,
19 resulting in maximum blood Pb level of 36 ug/dL], (2) post-infancy exposure group
20 [PND300 up to 10 years of age, resulting in maximum blood Pb level of 33 ug/dL], and
21 (3) lifetime exposure group [PNDO up to 10 years of age, resulting in maximum blood Pb
22 level of 32 ug/dL]) (Foster et al.. 1998; Singh etal.. 1993a). Rodent studies using i.p.
23 injections of Pb also showed ultrastructural damage to structures involved in
24 spermatogenesis (blood Pb level after i.p. injection treatment for 16 days: 7.4 ug/dL)
25 (Murthy et al.. 1995). More recently, Salawu et al. (2009) observed a decrease in absolute
26 testicular weight after Pb exposure (adult male SD rats, 10,000 ppm Pb acetate in
27 drinking water for 8 weeks). Anjum et al. (2010) reported decreased testicular and
28 epididymal weights of male rats exposed to Pb acetate (500 or 1,500 ppm Pb acetate in
29 drinking water for 45 days) which were significantly attenuated with Pb co-exposure to
30 the herb Centella asiatica (Sainath et al.. 2011). Pb induced morphological abnormalities
31 in sperm in a concentration-dependent manner (Allouche et al.. 2009; Oliveiraet al..
32 2009: Salawu et al.. 2009: Shan et al.. 2009: Tapisso et al.. 2009: Massanvi et al.. 2007:
33 Wang et al.. 2006a). Sperm abnormalities reported after Pb exposures were amorphous
34 sperm head, abnormal tail, and abnormal neck. Dong et al. (2009) reported decreased
35 epididymis and body weights in mice after an eight-week exposure to 6,000 ppm
36 Pb acetate in drinking water (adult male Kunming mice, 8-week exposure). Oliveira et al.
37 (2009) observed a negative correlation between Pb dose and intact acrosomes (8 week
38 old ICR-CD1 mice, subcutaneous injection of 74 and 100 mg PbCl2/kg body weight for
39 four consecutive days). Rubio et al. (2006) (adult mice treated with i.p. injections of 8, 16
November 2012 5-591 Draft - Do Not Cite or Quote
-------
1 or 24 mg/kg of Pb acetate for 35 days), Biswas and Ghosh (2006) (adult male Wistar rats,
2 i.p. injection of 8 mg/kg body weight Pb acetate for 21 days). Rubio et al. (2006) and
3 Biswas and Ghosh (2006) also observed a Pb-induced decrease in seminal vesicle and
4 ventral prostate weights. Rubio et al. (2006) reported that Pb acetate, in an exposure
5 concentration-dependent manner (8-24 mg/kg body weight), reduced the length of certain
6 stages of the spermatogenic cycle of rat seminiferous tubules and thus affected
7 spermatogenesis. Oral Pb acetate exposure (25 mg/kg bw in drinking water for 3 months,
8 resulting in blood Pb level of 5.3 (ig/dL) to adult male albino rats produced significant
9 histological seminiferous tubule damage (epithelium, spermatocytes, acrosomes) that was
10 attenuated with ascorbic acid treatment (Pb exposure + 100 mg/kg bw/day ascorbic acid,
11 resulting in blood Pb level of 4.7 (ig/dL) (El Shafai et al.. 2011). However, the majority
12 of studies did not observe a statistically significant difference in body weight or
13 reproductive organ weights after Pb exposure at the doses used in the studies. Not all of
14 the aforementioned studies observed changes in every parameter. This may be due to the
15 use of different strains or species, chemical form of the Pb compound administered,
16 dosage schedule, duration of exposure, and age of animals at the time of the study
17 (Oliveira et al.. 2009).Data from recent studies suggested that the generation of reactive
18 oxygen species (ROS) in the male reproductive tissues, which can then affect antioxidant
19 defense systems of cells (Pandya et al.. 2010) (adult male Charles Foster rats, Pb acetate
20 0.025 mg/kg body weight/day i.p. for 8 weeks) contributes to the MOA of Pb damage to
21 the male reproductive organs and sperm or semen. Salawu et al. (2009) observed a
22 statistically significant increase in malondialdehyde (MDA, oxidative stress marker) and
23 a significant decrease in the activity of antioxidant enzymes superoxide dismutase (SOD)
24 and catalase (CAT) in plasma and testes of adult male Sprague Dawley rats after
25 administration of 10,000 ppm Pb acetate in drinking water for 8 weeks. Supplementation
26 with tomato paste (used as a source of antioxidants) reduced Pb-induced ROS production
27 and prevented the Pb-induced increase in MDA formation and decrease in SOD and CAT
28 activity. Furthermore, co-treatment of Pb with substances that are known to have
29 antioxidant properties [i.e., tomato paste, Maca (Lepidium meyenii), and ascorbic acid]
30 prevented the Pb-induced reduction in sperm count, sperm motility, and sperm viability
31 (Salawu et al.. 2009: Shan et al.. 2009: Madhavi et al.. 2007: Rubio et al.. 2006: Wang et
32 al.. 2006a).
33 Recent studies continue to demonstrate that Pb may be directly toxic to mature
34 spermatozoa (adult Algerian mice, Pb acetate 21.5 mg/kg BW/every other day i.p. for 11
35 or 21 days) (Tapisso et al., 2009; Hernandez-Ochoa et al.. 2006) (adult NMRI mice,
36 6,000 ppm Pb chloride in drinking water for 16 weeks) as well as primary spermatocytes
37 (adult male Wistar rats were treated with drinking water containing 250mg/L Pb acetate
38 for) (Nava-Hernandez et al.. 2009: Rafique et al.. 2009) (adult albino rats, 10 mg/kg BW
39 Pb chloride i.p. once daily for 8 weeks). Nava-Hernandez et al. (2009) exposed two
November 2012 5-592 Draft - Do Not Cite or Quote
-------
1 groups of adult male rodent to Pb via drinking water (LI and L2, 250mg/L or 500 mg/L
2 Pb acetate starting at PND60 for 90 days). They found significant increases in spermatid
3 DNA damage with Pb exposure. In their study, all Pb-treated animals had blood Pb levels
4 statistically significantly higher than controls (Ll:19.54 (ig/dL and L2:21.90 (ig/dL); no
5 statistically significant difference in blood Pb levels existed between the two Pb exposure
6 groups likely because the L2 group drank less water than did the LI group. Piao et al.
7 (2007) reported that Pb exposure caused DNA damage to sperm; the Pb exposed group
8 had a blood Pb of 67 (ig/1. Piao et al. (2007) also examined the effect of Zn
9 supplementation on Pb-induced sperm aberrations and found that the proportion of
10 abnormal sperm was statistically significantly higher in the Pb group and the Pb+Zn
11 group than in controls (25 mg/kg Pb acetate i.p., 4 mg/kg Zn acetate i.p., both Pb acetate
12 and Zn acetate, once every two days, for 2 weeks). However, the proportion of abnormal
13 sperm in Pb+Zn group was statistically significantly lower than in Pb alone group.
14 Hernandez-Ochoa et al. (2006) reported that Pb reaches the sperm nucleus in the
15 epididymis of mice chronically exposed (16 weeks in adult animals) to Pb (resulting in
16 mean blood Pb of 75.6 (ig/dL) by binding to nuclear sulfhydryl groups from the
17 DNA-protamine complex, increasing sperm chromatin condensation, and thereby
18 interfering with the sperm maturation process without altering sperm quality parameters.
19 Tapisso et al. (2009) observed a statistically significant increase in the number of
20 micronuclei and frequency of sister chromatid exchange with increasing treatment
21 duration in adult male mice administered 21.5 mg/kg body weight Pb acetate by i.p.
22 injection (adult Algerian mice, Pb acetate 21.5 mg/kg BW/every other day for 11 or
23 21 days). Nava-Hernandez (2009) reported a concentration-dependent increase in DNA
24 damage in rat primary spermatocytes after a 13-week exposure period to Pb acetate in
25 drinking water (resulting in mean blood Pb levels between 19.5 and 21.9 (ig/dL). Rafique
26 et al. (2009) reported degenerative changes from pyknosis to apoptosis in primary
27 spermatocytes (adult albino rats, 10 mg/kg BW Pb chloride i.p. once daily for 8 weeks).
28 Hepatic expression of spermatogenic genes was transiently down-regulated in 8 week old
29 male Wistar-Kyoto (WKY) rats in response to Pb nitrate (100 (imol single i.v. injection)
30 3 hours after injection and recovered to baseline by 12 hours (Nemoto et al.. 2011); this
31 effect was not seen in the stroke-prone spontaneously hypertensive rats, which are from a
32 WKY background, or in Sprague-Dawley rats, demonstrating strain specificity.
33 Pb-induced apoptosis in germ cells within the seminiferous tubules is another suggested
34 mechanism by which Pb exerts its toxic effects on sperm production and function (Wang
35 et al.. 2006a) (Kunming mice, 2,000 ppm Pb acetate in drinking water for 14-42 days).
36 Dong et al. (2009) reported a exposure concentration-related increase in apoptosis in
37 spermatogonia and spermatocytes of Kunming mice after exposure to 1,500-6,000 ppm
38 Pb acetate in drinking water. Pb-induced testicular germ cell apoptosis was associated
39 with up-regulation of genes involved in the signal pathway of MAPK and death receptor
November 2012 5-593 Draft - Do Not Cite or Quote
-------
1 signaling pathway of FAS. For instance, up-regulation of K-ras and Fas expressions was
2 concomitant with activation of c-fos and active caspase-3 proteins. Wang et al. (2006a)
3 observed an exposure concentration-dependent increase in the expression of apoptotic
4 markers TGF(31 and caspase-3 in spermatogenic cells, Sertoli cells, and Leydig cells.
5 Shan et al. (2009) (20 mg/kg BW intragastric Pb acetate for 6 weeks) also reported a
6 statistically significant increase in mRNA expression and protein levels of Fas, Fas-L and
7 caspase-3 after Pb exposure. Supplementation with ascorbic acid inhibited or reduced the
8 Pb-induced apoptosis in germ cells and protected testicular structure and function (El
9 ShafaietaL 2011; Shan et al.. 2009; Wang et al.. 2006a) suggesting ROS generation is a
10 major contributing factor in decreased male fertility observed after chronic Pb exposure.
11 Similar to the results summarized in previous Pb AQCDs, recent epidemiologic and
12 toxicological studies indicate that Pb exposure has effects on sperm, semen, and male
13 reproductive organs. Consistent toxicological evidence from multiple labs with multiple
14 species of animals showed decrements in sperm or semen quality with Pb exposure
15 including decreased sperm counts, decreased sperm production rate, and a dose-
16 dependent suppression of spermatogenesis. Histological damage to rodent sperm and
17 ultrastructural damage to rodent and non-human primate seminiferous tubules has been
18 reported. Sperm from Pb-exposed male rodents used for in vitro fertilization of eggs from
19 unexposed females yielded a lower rate of fertilization. Also, direct effects of Pb on
20 rodent sperm DNA have been reported in rodents with drinking water exposure. The
21 toxicological findings cross species and are seen in wildlife including deer, earthworms,
22 rainbow trout, marine worms and fathead minnow. In studies of men exposed to Pb in
23 occupational settings, associations were observed between blood Pb levels of at least
24 25 (ig/dL and sperm count and quality. Multiple epidemiologic studies of occupational
25 cohorts included control populations with high blood Pb levels (close to or greater than
26 10 (ig/dL), which makes identification of effects at lower levels difficult. Occupational
27 studies had limited consideration for potential confounding factors, such as other
28 workplace exposures. An epidemiologic study of men attending a clinic for purposes of
29 infertility exam or semen donation demonstrated an inverse relationship between Pb
30 levels and sperm and semen quality (Telisman et al.. 2007). This study also controlled for
31 other metals in the analyses. Other studies of men at infertility clinics had greater
32 imprecision in their estimates, less control for confounding (such as other metals), and/or
33 small sample sizes. Additionally, studies limited to men at infertility clinics may suffer
34 from selection bias and are not generalizable. Future epidemiologic studies are warranted
35 to determine whether this association is observed at lower Pb levels.
November 2012 5-594 Draft - Do Not Cite or Quote
-------
5.8.4.2 Effects on Hormone Levels
1 The 2006 Pb AQCD (U.S. EPA. 2006b) provided evidence that Pb acts as an endocrine
2 disrupter in males at various points along the hypothalamic-pituitary-gonadal axis. The
3 2006 Pb AQCD also reported inconsistencies in the effects of Pb exposure on circulating
4 testosterone levels. Recent epidemiologic and toxicological studies are reported below.
5 Epidemiologic studies are summarized in Table 5-44. Epidemiologic studies were cross-
6 sectional; biological samples used for the measurement of Pb were measured
7 concurrently with hormone levels. One study estimated cumulative blood Pb.
November 2012 5-595 Draft - Do Not Cite or Quote
-------
Table 5-44 Summary of recent epidemiologic studies of associations between Pb levels and hormones for
males.
Reference
(Presented in
order of
appearance in
the text)
Telisman et al.
(2007)
Meeker et al.
(2010)
Study
Location Pb Biomarker
and Methodological or Exposure
Years Outcome Study Population Details Measurement
Croatia FSH, LH, Men aged 19-55, never Cross-sectional Blood Pb
2002-2005 testosterone, occupationally exposed study using linear
estradiol, to metals and going to multiple
prolactin a clinic for infertility regression
examination or for
semen donation to be
used for artificial
insemination
N=240
Michigan FSH, LH, Men aged Cross-sectional Blood Pb
NS inhibin B, 18-55 going to infertility study using
testosterone, clinics (distinction not multiple linear
SHBG, FAI, made between clinic regression
testosterone visits for male or
/LH female fertility issues)
N=219
Mean Pb
(SD) Adjusted Effect
in ug/dL Estimates
Median: 4.92 Standardized regression
(range coefficients for log blood Pb
1.13-14.91) (units not given)
T6stost6ron6 '
0.21
(p-value <0.003)
Estradiol:
0.22
(p-value <0.0008)
Prolactin'
-0.18
(p-value <0.007)
Note: Coefficients and p-
values not given if not
statistically significant (LH,
FSH)
Median: Regression coefficients
1 .50 (95% Cl)
(IQR1.10, FSH
2-°°) Istquartile:
O(ref)
2nd quartile:
0.13
(-0.10, 0.37)
3rd quartile:
0.10
(-0.15, 0.35)
4th quartile:
0.07
(-0.18,0.31)
LH
1st quartile:
O(ref)
2nd quartile:
0.004
(-0.20, 0.21)
Potential
Confounders
Adjusted for
in Analysis
Age, smoking
status, alcohol
use, Cd, Cu,
Zn, Se
An interaction
between Pb
and Cd was
included in
models for
testosterone
Age, BMI,
current
smoking
Considered but
did not include:
race, income,
season
November 2012
5-596
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in
the text)
Study
Location
and
Years Outcome
Study Population
Methodological
Details
Pb Biomarker
or Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted Effect
Estimates
Potential
Confounders
Adjusted for
in Analysis
3rd quartile:
0.13
(-0.09, 0.35)
4th quartile:
0.88
(-0.14,0.29)
Inhibin B
1st quartile:
O(ref)
2nd quartile:
-6.45
(-27.2, 14.3)
3rd quartile:
-4.62
(-26.6,17.4)
4th quartile:
-7.79
(-29.0,13.4)
Testosterone
1st quartile:
O(ref)
2nd quartile:
28.6
(-6.82,64.1)
3rd quartile:
15.8
(-21.8, 53.3)
4th quartile:
39.9
(3.32, 76.4)
SHBG
1st quartile:
O(ref)
2nd quartile:
-0.01
(-0.16,0.15)
3rd quartile:
0.04
(-0.12,0.21)
4th quartile:
0.07
(-0.10,0.23)
November 2012
5-597
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in
the text)
Study
Location
and
Years Outcome
Study Population
Methodological
Details
Pb Biomarker
or Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted Effect
Estimates
Potential
Confounders
Adjusted for
in Analysis
FAI
1st quartile:
O(ref)
2nd quartile:
0.8
(-0.04, 0.20)
3rd quartile:
0.03
(-0.10,0.17)
4th quartile:
0.08
(-0.05,0.21)
Testosterone /LH
1st quartile: 1.00 (ref)
2nd quartile:
0.07
(-0.16,0.30)
3rd quartile:
-0.05
(-0.29,0.19)
4th quartile:
0.07
(-0.17,0.31)
November 2012
5-598
Draft - Do Not Cite or Quote
-------
Reference
(Presented in Study
order of Location
appearance in and
the text) Years Outcome
Mendiola et al. Spain FSH, LH,
(2011) 2005-2007 testosterone
Pb Biomarker
Methodological or Exposure
Study Population Details Measurement
Men attending infertility Case-control Seminal plasma
clinics and classified as study using Pb
either normal sperm multiple linear Blood Pb
(controls) or oligo- regression
astheno-
teratozoospermia
(cases) based on WHO
semen quality criteria
NCases=30
N-^1
controls"0 '
Mean Pb
(SD)
in ug/dL
Seminal
plasma: 2.90
(IQR 2.70,
3.20)
Whole blood:
9.50 (IQR
7.50,11.90)
Blood
plasma: 2.90
(IQR 2.70,
3.10)
Cases:
Seminal
plasma' 3 0
(0.3)
Whole blood:
9.8 (2.3)
Blood
plasma: 2.9
(0.2)
Controls:
Seminal
plasma: 2.9
(0.3)
Whole blood:
9.7 (2.3)
Blood
plasma: 2.9
(0.3)
Adjusted Effect
Estimates
Linear regression p (95%
Cl)
FSH
Seminal plasma:
0.05
(-0.24, 0.39)
Whole blood:
0 04
(-0.03, 0.04)
Blood plasma:
-0.20
(-0.64, 0.25)
LH
Seminal plasma:
0.14
(-0.13,0.41)
Whole blood:
0 05
(-0.05, 0.07)
Blood plasma:
-0.07
(-0.49,0.31)
Testosterone
Seminal plasma:
0 11
(-0.10,0.31)
Whole blood:
0 01
(-0.05, 0.02)
Blood plasma:
-0.12
(-0.40,0.14)
Potential
Confounders
Adjusted for
in Analysis
Age, BMI,
number of
cigarettes/day
"Units not given (assume
1 ug/dL)
November 2012
5-599
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in
the text)
Hsieh et al.
(2009a)
Study
Location
and
Years Outcome
Taiwan FSH, LH,
1991-NS testosterone,
mhibm B
Study Population
Workers at a Pb-acid
battery factory with
annual blood Pb
measures
N=181
Methodological
Details
Longitudinal
occupational
cohort study using
multivariate linear
regression
Pb Biomarker
or Exposure
Measurement
Current blood Pb,
cumulative blood
Pb, time-
weighted
cumulative blood
Pb
Mean Pb
(SD)
in ug/dL
Current blood
Pb:
<10 ug/dL:
11.6%
>40 ug/dL:
17.1%
Adjusted Effect
Estimates
P from linear regression
Inhibin B
Current blood Pb:
0.40 (p-value 0.40)
Cumulative blood Pb:
0.05 (p-value 0.02)
Time-weighted cumulative
blood Pb:
1 .33 (p-value 0.007)
Potential
Confounders
Adjusted for
in Analysis
LH, FSH,
testosterone,
age, smoking
status, alcohol
use, BMI
Pearson's correlations
detected no correlations
between current blood Pb
levels and FSH, LH, or
testosterone. Cumulative
blood Pb levels were
correlated with FSH and
LH, but not testosterone.
Time-weighted cumulative
blood Pb levels were
correlated with LH, but not
FSH or testosterone.
November 2012
5-600
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in
the text)
Study
Location
and
Years
Outcome Study Population
Methodological
Details
Pb Biomarker
or Exposure
Measurement
Mean Pb
(SD)
in ug/dL
Adjusted Effect
Estimates
Potential
Confounders
Adjusted for
in Analysis
Naha and
Manna (2007)
Bangalore,
India
NS
FSH, LH,
testosterone
Non-occupationally
exposed controls and
occupationally exposed
workers
NControls=50
NIC
Occupational
cohort study using
ANOVA, Student's
t-test, and
Scheffe's F-test
''high exposure'
>sure=30
.=20
Categorized by
work history as
controls, low
exposure (7-10 yr
of exposure for 8
h/day) and high
exposure (>10yr
of exposure for 8
h/day)
Blood Pb
measurement
Controls
10.25(2.26)
Low
exposure
50.29 (3.45)
High
exposure
68.26 (2.49)
Semen Pb
measurement
Controls 2.99
(0.76)
Low
exposure
15.85(1.95)
High
exposure
25.30 (2.28)
Mean FSH (SD)
Control:
2.69(1.22)
Low exposure:
2.58 (1.94)
High exposure:
2.16(0.99)
p-values for difference
>0.05
Mean LH (SD)
Control:
5.14(2.35)
Low exposure:
4.27 (2.52)
High exposure:
3.9 (1.69)
p-values for difference
>0.05
Mean Testosterone (SD)
Control:
5.24 (2.40)
Low exposure:
4.83(1.21)
High exposure:
4.59(1.27)
p-values for difference
>0.05
None
November 2012
5-601
Draft - Do Not Cite or Quote
-------
1 Hormone levels were measured in a few recent epidemiologic studies. In a study of men
2 non-occupationally exposed to Pb in Croatia, increased blood Pb level was associated
3 with increasing serum testosterone and estradiol but decreasing serum prolactin
4 (Telisman et al.. 2007). In addition, the analysis of an interaction term for blood Pb and
5 blood Cd levels demonstrated a synergistic effect on increasing serum testosterone levels.
6 No association was observed between blood Pb and FSH or LH. This study controlled for
7 multiple potential confounders, including other metals. Among men recruited from
8 infertility clinics in Michigan, median blood Pb levels were much lower than those
9 observed in the other studies of Pb and hormone levels among men (Meeker et al., 2010).
10 No association was detected between blood Pb and levels of FSH, LH, inhibin B, sex
11 hormone-binding globulin (SHBG), free androgen index (FAI), or a measure of Leydig
12 cell function (T/LH). A positive association between the highest quartile of blood Pb and
13 testosterone was present, but this association did not persist when other metals were
14 included in the model. Similarly, another study of men recruited from infertility clinics
15 observed no association between Pb concentrations from seminal plasma, whole blood, or
16 blood plasma and FSH, LH, or testosterone (Mendiola et al.. 2011).
17 A study of occupationally-exposed men in Taiwan reported an association between
18 measures of cumulative blood Pb levels and inhibin B levels, but no association was
19 detected when using current blood Pb levels (Hsieh et al., 2009a). A correlation between
20 cumulative blood Pb measures and LH levels was detected but correlations were not
21 present when examining FSH or testosterone levels. No correlations were apparent
22 between FSH, LH, or testosterone and current blood Pb levels. Another study of men
23 with high blood Pb levels reported no difference in serum FSH, LH, and testosterone
24 among the three groups (controls: mean blood Pb 10.25 (ig/dL, low exposure: mean
25 blood Pb 50.29 (ig/dL, high exposure: mean blood Pb 68.26 (ig/dL) (Naha and Manna.
26 2007). This study did not assess any potential confounding factors.
27 In a recent toxicological study, Rubio et al. (2006) observed a decrease in testosterone
28 levels in Pb acetate-treated rats in a exposure concentration-related fashion (8-24 mg/kg
29 body weight), and this decrease correlated with reduced lengths of spermatogenic cycle
30 stages VII-VIII (spermiation) and IX-XI (onset of spermatogenesis). Anjum et al. (2010).
31 who dosed 50 day old male rats with 273 or 819 mg/L Pb acetate in drinking water (500
32 or 1,500 ppm, respectively; blood Pb not reported), found significant decreases in serum
33 testosterone and testicular 3(3-HSD and 17(3-HSD levels in Pb-exposed animals versus
34 controls. Pandya et al. (2010) reported altered hepatic steroidogenic enzyme activity.
35 Pillai et al. (2012) found gestational and lactational exposure to Pb acetate in Charles
36 Foster rats (subcutaneous injection of 0.05 mg/kg BW/day, blood Pb not reported)
37 induced significant decreases in testicular 17(3-HSD and serum testosterone. Biswas and
38 Ghosh (2006) reported a Pb-induced decrease in serum testosterone and gonadotropins
November 2012 5-602 Draft - Do Not Cite or Quote
-------
1 (FSH, LH) with inhibition of spermatogenesis, however, there was a statistically
2 significant increase in adrenal steroidogenic enzyme, A5-3(3-HSD, activity and serum
3 corticosterone levels indicating disruption of the adrenocortical process. Exposure
4 concentration-dependent decreases in serum testosterone were reported in Pb-exposed
5 male rats (Anjum et al., 2010). In contrast, Salawu et al. (2009) did not observe a
6 decrease in serum testosterone between control animals and animals administered
7 10,000 ppm Pb acetate in drinking water for 8 weeks. Allouche et al. (2009) not only did
8 not observe any statistically significant changes in serum FSH or LH, but reported an
9 increase in serum testosterone levels after 500-3,000 ppm Pb acetate treatment in
10 drinking water (only statistically significant in animals administered 500 ppm Pb acetate).
11 The results of these recent studies further support the theory that compensatory
12 mechanisms in the hypothalamic-pituitary-gonadal axis may allow for the adaptation of
13 exposed animals to the toxic endocrine effects of Pb (Rubio et al.. 2006; U.S. EPA.
14 2006b).
15 Overall, recent epidemiologic and toxicological studies report mixed findings regarding
16 hormone aberrations in males associated with Pb exposure or Pb biomarker levels. These
17 results are similar to those from the 2006 Pb AQCD (U.S. EPA. 2006b)on the effects of
18 Pb exposure on circulating testosterone levels. Epidemiologic studies are limited by their
19 sample populations, often occupational cohorts or men at infertility clinics, which may
20 not be generalizable. Occupational cohorts may have other exposures that confound the
21 associations, and studies at infertility clinics are subject to selection bias. A few of the
22 recent epidemiologic studies include important confounding factors, such as smoking, but
23 other factors, such as exposure to other metals, were often absent. Additionally, most
24 studies examine concurrent Pb and hormone levels which may not reflect changes
25 resulting from long-term exposures, as demonstrated by the longitudinal occupational
26 cohort study. Further, in cross-sectional studies, the temporality of effects cannot be
27 established.
5.8.4.3 Fertility
28 Epidemiologic studies have been performed comparing Pb and infertility in men. The
29 SMART study is a longitudinal study that examined the success of IVF treatment for
30 women and their partners starting their first round of treatment (Bloom et al.. 20 lib;
31 Bloom et al.. 2010). A small number of the male partners participated (n=16). Their mean
32 (SD) blood Pb level was 1.50 (0.80) (ig/dL. Higher blood Pb levels were associated with
33 greater oocyte fertilization (OR 1.08 [95% CI: 0.97, 1.21] per 1 (ig/dL increase in blood
34 Pb when adjusted for Cd, Hg, age, cigarette smoking, race/ethnicity, and creatinine),
35 which is not the expected direction (Bloom et al.. 2010). However, higher blood Pb was
November 2012 5-603 Draft - Do Not Cite or Quote
-------
1 associated with lower embryo cell number, a predictor of IVF success, and with higher
2 embryo fragmentation score, an inverse predictor of IVF success (OR for embryo cell
3 number: 0.58 [95% CI: 0.37, 0.91]; OR for embryo fragmentation score: 1.47 [95% CI:
4 1.11, 1.94] per 1 (ig/dL, controlled for age, race/ethnicity, cigarette smoking, creatinine,
5 Cd, and Hg, plus day of embryo transfer for embryo cell number) (Bloom et al.. 20 lib).
6 A case-control study conducted in Turkey reported that blood and seminal plasma Pb
7 levels were different in fertile (n=45; mean [SD] blood Pb: 23.16 [5.59] (ig/dL) and
8 infertile men (n=50; mean [SD] blood Pb: 36.82 [12.30] jig/dL) (p <0.001, ANOVA)
9 (Kiziler et al., 2007). There was no control for potential confounding factors although the
10 relationship persisted when limited to non-smokers. Another case-control study examined
11 occupational Pb exposure (determined by self-report of occupational exposure in the
12 past month) and detected no difference in reported exposure for infertile (n=650) versus
13 fertile men (n=698) (unadjusted OR 0.95 [95% CI: 0.6, 1.6]) (Gracia et al.. 2005). Blood
14 Pb was not measured but approximately 5.0% of infertile men and 5.3% fertile men
15 reported occupational exposure to Pb. A limitation present in these studies is that the
16 cases included are men who are seeking help at fertility clinics; the study populations are
17 not a sample of the general population regarding fertility. The results could be biased due
18 to the recruitment of individuals going to an infertility clinic, who may be different than
19 individuals suffering from infertility without knowing it or without going to a clinic.
20 Recent animal toxicology studies assessed paternal-mediated reproductive fitness by
21 examining the reproductive success of Pb-exposed males with non-exposed control
22 females. Anjum et al. (2010) found that adult male rats who were exposed to 273 or
23 819 mg/L Pb acetate in drinking water (500 or 1,500 ppm, respectively; blood Pb not
24 reported) spent a significantly longer time copulating than did their control littermates.
25 The Pb-exposed males were less successful copulators with only 73% of the 0.05%
26 Pb acetate exposed males, and 53% of the 1,500 ppm exposed males generating
27 copulatory plugs in the unexposed female mates. While the number of pregnant females
28 did not significantly differ from controls, Pb exposed males contributed to the formation
29 of significantly fewer implantations/dam, and significantly fewer fetuses/dam.
30 Pb-exposed males were able to sire offspring, but produced fewer offspring per litter. In a
31 group of males rats with co-exposure to Pb and the herb Centella asiatica, these
32 reproductive decrements were attenuated relative to rats exposed to Pb alone (adult albino
33 male rats, 1,500 ppm Pb acetate in drinking water for 70 days) (Sainath et al., 2011).
November 2012 5-604 Draft - Do Not Cite or Quote
-------
1 Overall, large, well-conducted epidemiologic studies of Pb exposure and fertility in males
2 are lacking. The few available studies reported inconsistent findings. Toxicological
3 studies demonstrated paternal associated subfecundity (fewer pups sired per pregnancy)
4 with altered mating behavior (longer time spent copulating), albeit in studies with no
5 blood Pb levels reported. Supplementation with antioxidants in a separate study showed
6 restoration of this subfecundity, possibly contributing MOA support to this decreased
7 fertility in male rodents.
5.8.4.4 Effects on Morphology and Histology of Male Sex Organs
8 Recent toxicological studies further support historical findings that showed an association
9 between Pb exposure and changes in the sex organs as well as germ cells. Histological
10 changes of testes in Pb nitrate-treated adult animals (a single i.p. dose of 12.5, 25, or
11 50 mg/kg of BW and were sacrificed 48 hours later) included seminiferous tubule
12 atrophy, Sertoli cell and Leydig cell shrinkage with pyknotic nuclei (Shan et al.. 2009;
13 Wang et al.. 2006a). dilatation of blood capillaries in the interstitium, undulation of basal
14 membrane, and occurrence of empty spaces in seminiferous epithelium (adult male
15 Wistar rats, single i.p. dose of 12, 25 or 50 mg/kg BW Pb acetate) (Massanyi et al..
16 2007). Pillai et al. (2010) found gestational and lactational exposure to Pb acetate in
17 Charles Foster rats (subcutaneous injection of 0.05 mg/kg BW/day) induced significant
18 decreases in absolute organ weight (testes and epididymis) and significant decreases in
19 relative epididymal weight. Anjum et al. (2010). who exposed 50 day old male albino
20 Wistar/NIN rats to Pb acetate (273 or 819 mg/L in drinking water, 500 or 1,500 ppm,
21 respectively, blood Pb levels not reported) for 45 days, reported significant decreases in
22 relative reproductive organ weight (epididymis, testis, vas deferens, and seminal vesicle)
23 in Pb-exposed animals.
5.8.4.5 Summary of Effects on Male Reproductive Function
24 Evidence of associations between Pb exposure and male reproductive function vary by
25 outcome. The strongest evidence of an association is the relationship observed between
26 Pb and negative effects on sperm and semen in both recent epidemiologic and
27 toxicological studies and studies reviewed in previous Pb AQCDs. Decrements in sperm
28 count, sperm production rate and semen quality were reported in animal toxicological
29 studies in rodents with drinking water Pb exposure rodents [(Sokol and Berman. 1991;
30 Sokol et al.. 1985). (blood Pb level 34-37 (ig/dL)] and rabbits exposed to subcutaneous
31 Pb [blood Pb levels of 25 (ig/dL, (Moorman et al.. 1998)]. Rodents exposed to Pb had
32 direct effects of Pb on sperm DNA, i.e., elevated levels of DNA damage [(Nava-
November 2012 5-605 Draft - Do Not Cite or Quote
-------
1 Hernandez et al.. 2009). blood Pb levels 19 and 22 (ig/dL]. Histological or ultrastructural
2 damage to the male reproductive organs were reported in studies from rodents [(El Shafai
3 et al.. 2011). blood Pb level 5.1 (ig/dL] and non-human primates [(Singh et al.. 1993a).
4 blood Pb level 43 (ig/dL]. Subfecundity has been reported in unexposed females mated to
5 Pb exposed males (decreased number of pups born/litter). Also, sperm from Pb-exposed
6 rats (blood Pb 33 to 46 ug/dL) used for in vitro fertilization of eggs harvested from
7 unexposed females yielded lower rates of fertilization (Sokol et al.. 1994). Many of the
8 epidemiologic studies included occupational cohorts, which had high blood Pb levels
9 (> 25(ig/dL), or men attending infertility clinics, which have potential selection bias.
10 Additionally, control for confounding factors, such as other workplace exposures, was
11 not often performed. A study of men (who were attending a clinic for an infertility exam
12 or to donate semen for use in artificial insemination) did control for multiple factors,
13 including other metals and smoking status, and reported an association between blood Pb
14 levels and some indicators of poor sperm quality (Telisman et al.. 2007). Recent
15 toxicological studies also reported an association between Pb exposure and decreases in
16 reproductive organ weight, organ histological changes in the testes and germ cells. Male
17 rats exposed to Pb also showed subfecundity, in that they produced smaller litters when
18 mated with unexposed females (Anjum et al.. 2010). Further coherence for these findings
19 in laboratory animal models is found in with findings in the ecological literature for the
20 effects of Pb exposure on reduced fecundity in terrestrial and aquatic animal species
21 (Sections 7.4.5.2. 7.3.4.2. and 7.4.5.3). Similar to the 2006 Pb AQCD (U.S. EPA. 2006b).
22 recent epidemiologic and toxicological studies reported inconsistent results regarding
23 hormone aberrations associated with Pb exposure. Mixed findings were also apparent
24 among epidemiologic studies of fertility among men.
5.8.5 Effects on Female Reproductive Function
25 The epidemiologic studies on Pb and female reproductive function presented in the
26 2006 Pb AQCD (U.S. EPA. 2006b) provided little evidence for an association between
27 Pb biomarkers and effects on female reproduction and fertility. However, the 1986 and
28 2006 Pb AQCDs (U.S. EPA. 2006b. 1986a) reported toxicological findings that Pb
29 exposure was associated with effects on female reproductive function that can be
30 classified as alterations in female sexual maturation, effects on fertility and menstrual
31 cycle, endocrine disruption, and changes in morphology or histology of female
32 reproductive organs including the placenta. Since the 2006 Pb AQCD, many
33 epidemiologic studies have been published regarding Pb biomarker levels in women and
34 reproductive effects. In addition, recent toxicological studies add further knowledge of
35 Pb-related effects on the female reproductive system.
November 2012 5-606 Draft - Do Not Cite or Quote
-------
5.8.5.1 Effects on Female Sex Endocrine System and Estrus
Cycle
1 Multiple epidemiologic studies have examined the association between blood Pb levels
2 and hormone levels and the estrus cycle. Epidemiologic studies (characterized in Table
3 5-45) were cross-sectional in design, analyzing measures of Pb and hormones that were
4 collected either concurrently or close in time. These studies support the toxicological
5 findings, which are the major body of evidence on endocrine effects of Pb.
November 2012 5-607 Draft - Do Not Cite or Quote
-------
Table 5-45 Summary of recent epidemiologic studies of associations between Pb levels and hormones for
females.
Reference
(Presented in
order
appearance in
the text)
Study,
Location,
and
Years
Outcome Study Population
Mean
Methodological Pb Pb (SD) Adjusted Effect
Details Biomarker in ug/dL Estimates
Potential Confounders
Adjusted for in
Analysis
U.S.
1988-1994
FSH, LH Women aged 35-60 from
the NHANES III study
N=3375
Cross-sectional Blood Pb 2.8 Linear regression
study using linear slope (95% Cl)
regression for log-transformed
Pb
FSH:
Post-menopausal
22.2(13.5, 30.8)
Pregnant
0.1 (-0.1,0.3)
Menstruating at
time of exam
2.1 (-2.1,6.3)
Both ovaries
removed
32.6(10.1, 55.1)
Birth control pills
being used
-6.3 (-10.0,-2.5)
Pre-menopausal
8.3(3.8, 12.7)
LH:
Post-menopausal
6.2 (3.0, 9.5)
Pregnant
-0.8 (-1.9, 0.4)
Menstruating at
time of exam
-0.3 (-1.8, 1.3)
Both ovaries
removed
10.0(1.1, 18.9)
Birth control pills
being used:
-0.6 (-2.9, 1.6)
Pre-menopausal
1.7 (-0.6, 4.1)
Age, total bone mineral
density, serum cotinine,
alcohol use, current breast
feeding, hysterectomy, one
ovary removed, Norplant
use, radiation or
chemotherapy, hormone pill
use, vaginal cream use,
hormone patch use
November 2012
5-608
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order
appearance in
the text)
Pollack et al.
(2011)
Study,
Location,
and
Years Outcome Study Population
Buffalo, FSH, Healthy, premenopausal
NY estradiol, LH, women aged 18-44 with
onnc 9m7 progesterone, menstrual cycle length of
and cycle 21-35 days, BMI of 18-35
length kg/m , not recently using
birth control, not planning
to become pregnant, and
not breast feeding
N=252
Mean
Methodological Pb Pb (SD) Adjusted Effect
Details Biomarker in ug/dL Estimates
Longitudinal Blood Pb 0.93 Mean % Estradiol
cohort using |op. 0.30-0.72 ug/dL:
nonlinear mixed 0 68' 1 2rj Ref
models with ' ' ' 0.73-1.10 ug/dL:
harmonic terms a 9 c 1 9 IRK^
andwei9hted ?~(«™ 1
linear mixed 1.11-6.20 ug/dL:
models 4.7 (-4.7, 15.2)
Amplitude
Estradiol
0.30-0.72 ug/dL:
Ref
0.73-1. 10ug/dL:
-0.01
(-0.06, 0.04)
1.11-6.20 ug/dL:
-0.02
(-0.7, 0.03)
Phase Shift
Estradiol
0.30-0.72 ug/dL:
Ref
0.73-1.10 ug/dL:
-0.09 (-0.24, 0.05)
1. 11-6.20 ug/dL:
0.1 4 (-0.01, 0.29)
Mean % FSH
0.30-0.72 ug/dL:
Ref
0.73-1. 10ug/dL:
8.0 (-0.9, 17.7)
1.11-6.20 ug/dL:
3.6 (-5.3, 13.3)
Amplitude FSH
0.30-0.72 ug/dL:
Ref
0.73-1.10 ug/dL:
-0.01 (-0.03, 0.02)
1. 11-6.20 ug/dL:
-0.02 (-0.04, 0.01)
Phase Shift FSH
0.30-0.72 ug/dL:
Ref
Potential Confounders
Adjusted for in
Analysis
Age, BMI, race
Also examined, but did not
include: smoking, income,
education, physical activity,
parity, dietary Fe, fish
consumption, shellfish
consumption, vegetable
consumption, total calories
0.73-1.10ug/dL:
November 2012
5-609
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order
appearance in
the text)
Study,
Location,
and
Years
Outcome Study Population
Methodological
Details
Pb
Biomarker
Mean
Pb (SD)
in ug/dL
Adjusted Effect
Estimates
Potential Confounders
Adjusted for in
Analysis
-0.06 (-0.25, 0.12)
1.11-6.20 |jg/dL:
-0.02 (-0.21, 0.18)
Mean % LH
0.30-0.72 |jg/dL:
Ref
0.73-1.10|jg/dL:
5.1 (-5.1,16.4)
1.11-6.20 |jg/dL:
-0.5 (-10.5, 10.7)
Amplitude LH
0.30-0.72 |jg/dL:
Ref
0.73-1.10 |jg/dL:
-0.01 (-0.03, 0.02)
1.11-6.20 |jg/dL:
-0.02 (-0.04, 0.01)
Phase Shift LH
0.30-0.72 |jg/dL:
Ref
0.73-1.10|jg/dL:
-0.16 (-0.36, 0.03)
1.11-6.20 |jg/dL:
-0.11 (-0.32, 0.10)
Mean %
Progesterone
0.30-0.72 |jg/dL:
Ref
0.73-1.10|jg/dL:
7.5(0.1,15.4)
1.11-6.20 |jg/dL:
6.8 (-0.8, 14.9)
Amplitude
Progesterone
0.30-0.72 |jg/dL:
Ref
0.73-1.10|jg/dL:
0.07(0.01,0.15)
1.11-6.20 |jg/dL:
-0.06 (-0.13, 0.01)
Phase Shift
Progesterone
0.30-0.72 |jg/dL:
November 2012
5-610
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order
appearance in
the text)
Study,
Location,
and
Years
Outcome Study Population
Methodological
Details
Pb
Biomarker
Mean
Pb (SD)
in ug/dL
Adjusted Effect
Estimates
Potential Confounders
Adjusted for in
Analysis
Ref
0.73-1.10|jg/dL:
0.04 (-0.06, 0.15)
1.11-6.20 |jg/dL:
0.15(0.05, 0.26)
Linear models
P (95% Cl)
Estradiol
0.03 (-0.05, 0.11)
FSH
-0.01 (-0.07, 0.06)
LH
0.02 (-0.06, 0.10)
Progesterone
0.06 (-0.04, 0.17)
OR (95% Cl) for
anovulation per
1 |jg/dL
1.20(0.62, 2.34)
November 2012
5-611
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order
appearance in
the text)
Jackson et al.
(2011)
Chang et al.
(2006)
Study,
Location,
and
Years Outcome
Buffalo, FSH,
NY estradiol, LH,
2005-2007 progesterone,
and cycle
length
Kaohsiung Estradiol
City,
Taiwan
1999
2000-2001
Study Population
Healthy, pre-menopausal
women aged 1 8-44 with
menstrual cycle length of
21 -35 days, BMI of 18-35
kg/m , not recently using
birth control, not planning
to become pregnant, and
not breast feeding
N=252
Women receiving care at a
infertility clinic in
2000-2001 or delivering a
normal infant at a nearby
medical center in 1 999
N=147
Methodological
Details
Longitudinal
cohort study using
linear regression
and logistic
regression
Case-control
study using
multivariate linear
regression
Mean
Pb Pb (SD) Adjusted Effect
Biomarker in ug/dL Estimates
Blood Pb Median: Adjusted percent
0.87 change (95% Cl)
IQR. in serum hormone
0 68 1 20 level for Cnan9e in
' ' blood Pb
FSH'
-2.5 (-11. 2, 7.0)
Estradiol:
4.9 (-5.0, 15.9)
LH:
2.5 (-12.3, 19.9)
Progesterone:
4.6 (-12.2, 24.6)
Cycle length:
0.2 (-2.8, 3.3)
OR (95% Cl) per
unit Pb
<25 day vs.
25-35 day cycle
length:
0.9 (0.4, 2.3)
>35 day vs.
25-35 day cycle
length:
0.5(0.1, 1.9)
Blood Pb 3.12 Linear regression
(0.19) p(SE)forPb
1.18(0.60)
p-value: 0.049
Potential Confounders
Adjusted for in
Analysis
Cd, Hg, age, race /ethnicity
Not specified
November 2012
5-612
Draft - Do Not Cite or Quote
-------
1 An epidemiologic study using the NHANES III data and including women aged
2 35-60 years old examined the relationship between blood Pb levels (mean 2.8 (ig/dL) and
3 serum follicle stimulating hormone (FSH) and luteinizing hormone (LH) (Krieg. 2007).
4 Deviation from normal FSH and LH levels may indicate endocrine disruption related to
5 ovary functioning. Researchers found that higher blood Pb levels were associated with
6 higher levels of serum FSH and LH among both postmenopausal women and women
7 with both ovaries removed. There was also a trend of increasing serum FSH with blood
8 Pb levels for pre-menopausal women who were not menstruating at the time of the exam
9 or pregnant, although the association was not statistically significant for LH. A limitation
10 of this portion of the study is that FSH and LH were measured without attention to day of
11 a woman's menstrual cycle and LH and FSH are known to vary throughout the cycle of
12 non-menopausal, cycling women who are not taking birth control pills. Higher blood Pb
13 levels were associated with lower levels of serum FSH among women taking birth
14 control pills. The inverse association was also present for LH, but it was not statistically
15 significant. No associations between blood Pb and FSH or LH were apparent for women
16 who were menstruating at the time of the exam or were pregnant. Further analysis
17 indicated that the lowest level of blood Pb for which a statistically significant association
18 between blood Pb and FSH could be observed was 1.7 (ig/dL among women with their
19 ovaries removed. For LH, the lowest level of blood Pb for which a statistically significant
20 association between blood Pb and LH could be observed was 2.8 (ig/dL among
21 postmenopausal women. Associations between hormones and blood Pb level were also
22 investigated using the BioCycle study cohort (Jackson et al., 2011; Pollack et al., 2011).
23 These women were premenopausal with normal cycles and not on birth control. Blood Pb
24 was measured at enrollment and hormones were measured multiple times throughout the
25 menstrual cycle. Neither study detected an association between unit change in blood Pb
26 and hormone levels. However, when examining tertiles of Pb, women in the highest
27 tertile blood Pb (1.11-6.20 (ig/dL) had higher mean progesterone and longer length of a
28 phase shift compared to women in the lowest tertile (0.30-0.72 (ig/dL) (Pollack et al..
29 2011). Other associations were observed but were not statistically significant (Pollack et
30 al., 2011). No associations were detected for anovulation (Pollack et al., 2011) or for
31 cycle length (Jackson et al.. 2011). Another epidemiologic study was performed in
32 Kaohsiung City, Taiwan among two groups of women aged 23-44 years: those who were
33 seeking help at a fertility clinic after one year of trying to conceive, and those who had
34 previously delivered an infant and were identified from medical records of a postpartum
35 care unit (Chang et al.. 2006). The mean (SD) concurrent blood Pb level in this study was
36 3.12 (0.19) (ig/dL. The study reported apositive association between blood Pb levels and
37 serum estradiol concentrations during the early follicular phase, which reflects ovary
38 activity.
November 2012 5-613 Draft - Do Not Cite or Quote
-------
1 The effect of Pb exposure on the female endocrine system was demonstrated in
2 toxicological studies reviewed in the 1986 and 2006 Pb AQCD (U.S. EPA. 2006b.
3 1986a). However, the mechanism by which Pb affects the endocrine system has not been
4 fully elucidated. Several recent articles continue to demonstrate that Pb alters the
5 concentration of circulating hormones in female experimental animals. As mentioned in
6 the previous AQCD, Pine et al. (2006) observed that maternal Pb exposure (during
7 gestation and lactation) caused a decrease in basal LH levels in pre-pubertal female
8 Fischer 344 rat pups as compared to control, non-Pb exposed pups. Dumitrescu et al.
9 (2008a) observed alteration of hormone levels in adult female Wistar rats after ingesting
10 Pb acetate (50, 100, 150 ppb) in drinking water for 6 months; measurements were made
11 during the pro-estrous stage of the estrous cycle to allow for consistent timing for
12 comparison of cyclic hormonal variation. The authors reported decreases in FSH,
13 estradiol, and progesterone levels with increases in LH and testosterone levels.
14 Nampoothiri and Gupta (2008) administered Pb acetate at a concentration that did not
15 affect reproductive performance, implantation or pregnancy outcome (0.05 mg/kg body
16 weight) to Charles Foster female rats 5 days before mating and during the gestational
17 period. They observed a decrease in steroidogenic enzymes, 3(3- hydroxysteroid
18 dehydrogenase (HSD) and 17J3-HSD, activity in reproductive organs, as well as a
19 decrease in steroid hormones (progesterone and estradiol), suggesting that chronic
20 exposure to low levels of Pb may affect reproductive function of mothers and their
21 offspring. Similarly, Pillai et al. (2010) reported impaired ovarian steroidogenesis in
22 Charles Foster adult female rats (PND56) from dams exposed gestationally and
23 lactationally to Pb acetate (subcutaneous daily injections of 0.05 (ig/kg BW). Pillai
24 observed a decrease in steroidogenic enzymes, 3(3-HSD and 17(3-HSD, but saw no
25 changes in ovarian steroidogenic acute regulatory protein (StAR) or CYP11 mRNA
26 levels indicating Pb-induced inhibition of ovarian steroidogenesis.
27 Kolesarova et al. (2010) conducted an in vitro study to examine the secretory activity of
28 porcine ovarian granulose cells after Pb administration for 18 hours. The results of the
29 study showed that Pb acetate concentrations of 0.046 mg/mL and 0.063 mg/mL
30 statistically significantly inhibited insulin-like growth factor-1 (IGF-1) release, but
31 concentrations of 0.25 mg/mL and 0.5 mg/mL did not influence IGF-1 release.
32 Progesterone release was not affected by Pb treatment; however, Pb caused a reduction in
33 LH and FSH binding in granulose cells and increased apoptosis as evidenced by
34 increased expression of caspase-3 and cyclin Bl, suggesting a Pb-induced alteration in
35 the pathways of proliferation and apoptosis of porcine ovarian granulose cells. Decreased
36 gonadotropin binding was also observed in rats after Pb exposure subcutaneously
37 administered Pb (0.05 mg/kg body weight daily before mating and during pregnancy)
38 with a resulting blood Pb of 2.49 (ig/mL) (Nampoothiri and Gupta. 2006).
November 2012 5-614 Draft - Do Not Cite or Quote
-------
1 No recent toxicological studies were found that examined Pb-induced effects on the
2 estrus cycle.
3 Overall, toxicological studies report alterations in hormone levels related to blood Pb
4 concentration. Similarly, epidemiologic studies reported associations between
5 concurrent/closely timed blood Pb levels and hormone levels in female adults. Although
6 Pb-associated changes in hormone levels are observed, there are discrepancies and the
7 hormones examined vary by study. One explanation for the inconsistent findings is that
8 changes could vary based on current hormonal and reproductive status of the participants.
9 Also, the covariates included in statistical models as potential confounders varied among
10 studies, which could contribute to between study heterogeneity. This is also a limitation
11 of the epidemiologic studies; not all of the studies investigated important confounders,
12 such as other metal exposures or smoking. Additionally, the cross-sectional design of
13 these studies leaves uncertainty regarding Pb exposure timing, duration, and frequency
14 that contributed to the observed associations.
5.8.5.2 Effects on Fertility
15 Previous studies indicated that Pb exposure does not produce total sterility, but it can
16 disrupt female fertility (U.S. EPA, 2006b). Recent epidemiologic studies and studies in
17 experimental animals have inconsistent results. The epidemiologic studies are
18 summarized in Table 5-46. Most of these studies examining biological measures of Pb
19 collected at or during the period of possible fertilization or start of fertility treatment,
20 although Bloom et al. (2011 a) measured blood Pb at baseline and followed women for at
21 least 12 menstrual cycles (or until pregnancy).
November 2012 5-615 Draft - Do Not Cite or Quote
-------
Table 5-46 Summary of recent epidemiologic studies of associations between Pb levels and
Reference
(Presented in
order of
appearance in
the text)
Bloom et al.
(2011 a)
Chang et al.
(2006)
Study
Location,
and
Years Outcome
New York Achieving
1996-1997 pregnancy
Kaohsiung Infertility
City,
Taiwan
1999,
2000-2001
Study Population
Women who were aged
18-34 years, were
previously part of a study
about fish consumption,
and were not currently
pregnant and were
followed for 12 menstrual
cycles or until pregnant
N=80
Women receiving care at
a infertility clinic in
2000-2001 or delivering
a normal infant at a
nearby medical center in
1999
N:
Cases
=64
N:
Controls
=83
Methodological
Details
Longitudinal
cohort using Cox
proportional
hazards
Case-control
study using
unconditional
logistic regression
Mean Pb
Pb (SD)
Biomarker in ug/dL
Blood Pb at No positive
baseline pregnancy
test: 1 .55
(0.16)
Positive
pregnancy
test: 1 .54
(0.12)
Blood Pb 3.12(0.19)
Adjusted
Effect
Estimates
P (95% Cl)
-0.031
(-1 .066, 1 .004)
per 0.6 ug/dL
OR (95% Cl)
Infertility
< 2.5 ug/dL:
1.00
(Referent group)
>2.5 ug/dL:
2.94
(1.18,7.34)
fertility for females.
Potential Confounders
Adjusted for in Analysis
Baseline As, baseline Cd,
baseline Mg, baseline Ni,
baseline Se, baseline Zn, total
serum lipids, age, parity,
frequency of intercourse
during fertility window, alcohol
use, cigarette use
Age, BMI, active smoking, use
of Western medicine
Considered but did not
include: irregular
menstruation, age at first
menses, marital status,
passive smoking,
contraceptive drugs
November 2012
5-616
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in
the text)
Bloom et al.
(2010)
Bloom et al.
(2011b)
Study
Location,
and
Years Outcome
California Oocyte
2007-2008 maturity,
oocyte
fertilization
California Embryo cell
2007-2008 number,
embryo
fragmentation
score
Study Population
Women who were part of
the Study of Metals and
Assisted Reproductive
Technologies (SMART):
women referred to the
Center for Reproductive
Health of UCSF for
infertility treatment and
their first IVF procedure
N=15
Women who were part of
the Study of Metals and
Assisted Reproductive
Technologies (SMART)
(women referred to the
Center for Reproductive
Health of UCSF for
infertility treatment and
their first IVF procedure)
and who generated
embryos
N=24
Mean Pb
Methodological Pb (SD)
Details Biomarker in ug/dL
Longitudinal Blood Pb at 0.82 (0.32)
cohort using the time of
multivariable log- oocyte
binomial retrieval
regression
Longitudinal Blood Pb at 0.83 (0.30)
cohort using the time of
logistic regression oocyte
retrieval
Adjusted
Effect
Estimates
RR per 1 ug/dL
Oocyte
maturity
(determined by
Metaphase II
arrest):
0.54 (0.31 , 0.93)
0.25 (0.03,
2.50)*
Oocyte
fertilization:
0.97 (0.66, 1 .43)
1 .09 (0.72,
1 .65)*
OR per 1 ug/dL*
Embryo cell
number:
0.25 (0.07, 0.86)
Embryo
fragmentation
score:
1 .71 (0.45, 6.56)
Potential Confounders
Adjusted for in Analysis
Age, cigarette smoking,
race/ethnicity
*Also, controlled for Cd.
Age, race/ethnicity, cigarette
smoking, urine creatinine
Additionally included for
embryo cell number: day of
embryo transfer
*Also controlling for Hg and
Cd
November 2012
5-617
Draft - Do Not Cite or Quote
-------
Reference
(Presented in Study.
order of Location,
appearance in ar|d
the text) Years Outcome
Al-Saleh et al. Riyadh, Achieving
(2008a) Saudi pregnancy
Arabia and/or
2002-2003 fertilization
Study Population
Women aged 1 9-50
undergoing IVF
N.
.
pregnancy
_or\o
-ZUo
N:
No pregnancy
=321
N:
fertility
=556
N:
No fertility
=63
Mean Pb
Methodological Pb (SD)
Details Biomarker in ug/dL
Longitudinal Blood Pb Blood Pb:
cohort using Follicular 3-34 <2-24)
logistic regression fluid pb B|ood pb
levels
>10 ug/dL:
1.7%
Follicular
fluid:
0.68(1.82)
Adjusted
Effect
Estimates
OR (95% Cl)
(unit not given,
assume results
are per 1 ug/dL)
Pregnancy
Blood Pb:
0.55
(0.23, 1.31)
Follicular fluid
Pb:
1.36
(0.91 , 2.02)
Fertilization
Blood Pb:
0.30
(0.08, 1 .03)
Follicular fluid
Pb:
1.45
(0.69, 3.02)
Note: In a
reduced
adjusted model
for fertilization,
the OR for blood
Pb was 0.38
(0.14,0.99)
Potential Confounders
Adjusted for in Analysis
Age, husband's age, BMI,
location and duration at that
location, previous location and
duration at that location, age
at first menses, number
of days of menstrual cycle,
education, work status,
husband's education, family
income, husband's smoking
status, blood and follicular Cd
and Hg, follicular cotinine
Also included for pregnancy
as outcome: coffee
consumption, tea
consumption, caffeinated soft
drink consumption,
November 2012
5-618
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in
the text)
Study
Location,
and
Years Outcome
Study Population
Methodological
Details
Mean Pb
Pb (SD)
Biomarker in ug/dL
Adjusted
Effect
Estimates
Potential Confounders
Adjusted for in Analysis
Silberstein et
al. (2006)
Providence, Achieving
Rl pregnancy
NS
Women undergoing IVF
at the study hospital
N.
.
pregnancy
=4
N:
No oreanancv
Longitudinal Follicular
cohort study using fluid Pb
Mann-Whitney
U-test
Not given
quantitatively
From a figure
in the paper:
Median Pb in
follicular fluid
of pregnant
women: -1.3
P-value for
difference in
medians by
Mann-Whitney
U test: 0.0059
*Note, This
study only
None
=5
Median Pb in
follicular fluid
of non-
pregnant
women: -2.2
included 9
women
November 2012
5-619
Draft - Do Not Cite or Quote
-------
1 A prospective cohort study enrolled women who previously participated in a study offish
2 consumption for a length of up to 12 menstrual cycles and investigated the relationship
3 between blood Pb levels at baseline and having a positive pregnancy test at some point
4 during the next 12 menstrual cycles (Bloom etal. 201 la). No association was observed
5 between blood Pb and achieving pregnancy.
6 Among women aged 23-44 years, a difference in blood Pb was reported between women
7 who were seeking help at a fertility clinic after one year of trying to conceive and women
8 who had previously delivered an infant and were identified from medical records of a
9 postpartum care unit at a medical center (Chang et al.. 2006). Higher odds of infertility
10 were observed when comparing women with blood Pb levels >2.5 (ig/dL to those with
11 blood Pb levels < 2.5 (ig/dL although this study is limited by its case-control design.
12 Epidemiologic studies have also examined women having difficulty conceiving by
13 performing studies among patients of fertility clinics or undergoing in vitro fertilization
14 (IVF). The Study of Metals and Assisted Reproductive Technologies (SMART) enrolled
15 women undergoing their first round of IVF and investigated multiple steps before
16 pregnancy as the outcomes (Bloom et al., 20 lib; Bloom etal.. 2010). Higher blood Pb
17 levels were associated with lower oocyte maturity although the lack of power made
18 interpretation of models controlling for Cd difficult. No association was observed
19 between blood Pb and oocyte fertilization (Bloom et al.. 2010). In the examination of
20 markers of IVF success, inconsistent results were observed. Embryo cell number was
21 lower in association with higher blood Pb levels but no association was observed for
22 embryo fragmentation score (Bloom etal.. 20 lib). Another study examining fertility
23 reported on women in Saudi Arabia aged 19-50 years who were undergoing IVF
24 treatment (Al-Saleh et al., 2008a). Women were categorized as having achieved a
25 pregnancy versus not having achieved a pregnancy and achieved fertilization versus not
26 achieving fertilization. The majority of women had follicular Pb levels that were below
27 the limit of detection, whereas less than 2% of women had blood Pb levels below the
28 limit of detection. In addition, less than 2% of women had blood Pb levels that were
29 above 10 (ig/dL. Follicular Pb levels were not correlated with the blood Pb. No
30 association was observed between blood or follicular Pb and pregnancy outcomes in
31 either crude or adjusted models. An association was not detected between follicular Pb
32 and fertilization, but higher blood Pb was associated with lower rates of fertilization.
33 Finally, a study that included nine women undergoing IVF treatment in Rhode Island
34 (Silberstein et al., 2006) found that median follicular Pb levels in women who achieved
35 pregnancy were lower than the follicular Pb levels among nonpregnant women.
November 2012 5-620 Draft - Do Not Cite or Quote
-------
1 Overall, these epidemiologic studies examine a variety of fertility-related endpoints and
2 although some studies demonstrate an association between higher Pb levels and
3 fertility/pregnancy, as a whole the results are inconsistent across studies. One limitation
4 present in most of these studies is that the participants are women who are seeking help
5 for fertility problems. The participants are not samples of the general population and
6 therefore cannot be generalized to all women of childbearing age. This may also have
7 introduced substantial selection bias into the study.
8 Animal toxicology studies following female fertility looked at various outcomes. Several
9 studies observed a decrease in litter size when females were exposed to Pb before mating
10 or during pregnancy (Dumitrescu et al.. 2008c: lavicoli et al.. 2006a: Teijon et al.. 2006).
11 Pups in a study by Teijon et al. (2006) receiving 400 ppm Pb acetate in drinking water
12 had blood Pb of 97 (ig Pb/dL blood at 1 week post-weaning and 18.2 (ig Pb/dL blood at
13 2 week post-weaning. Dumitrescu et al. (2008c) observed a modification in sex ratio of
14 pups born to dams exposed to Pb before mating and during the entirety of pregnancy. As
15 the dose of Pb increased, the number of females per litter also increased (i.e., 1 male to
16 0.8 female in non-Pb exposed group; 1 male to 0.66 female in 50 ppb Pb acetate group; 1
17 male to 2.25 females in 100 ppb group; and 1 male to 2.5 females in 150 ppb group).
18 These results are not consistent with earlier results of Ronis et al. (1998b). who did not
19 observe differences in sex ratio dams and offspring were exposed only during pregnancy.
20 Thus, Pb exposure in animal studies during or before pregnancy have shown effects on
21 litter size and mixed effects on sex ratio.
22 Nandi et al. (2010) demonstrated a concentration-dependent decline in viability rate,
23 maturation, fertilization, and cleavage rates of buffalo oocytes cultured in medium
24 containing 1-10 (ig/mL Pb acetate (24 hour culture). Karaca and §im§ek (2007) observed
25 an increase in the number of mast cells in ovary tissue after Pb exposure (2,000 (ig/mL in
26 drinking water for 6 weeks prior to estrous monitoring then for 1 additional month during
27 which estrous cyclicity was monitored) suggesting that Pb may stimulate an
28 inflammatory response in the ovaries which may contribute to Pb-induced female
29 infertility.
30 In contrast, Nampoothiri and Gupta (2008) did not observe any statistically significant
31 change in fertility rate or litter size in female rats subcutaneously administered Pb
32 (0.05 mg/kg body weight daily before mating and during pregnancy) with a resulting
33 blood Pb of 2.49 (ig/mL. Although reproductive performance was not affected in this
34 study, the authors did report an alteration in implantation enzymes. Cathepsin-D activity
35 decreased and alkaline phosphatase activity increased after Pb exposure.
November 2012 5-621 Draft - Do Not Cite or Quote
-------
1 In summary, recent epidemiologic and toxicological studies on the effect of Pb on
2 fertility outcomes have generated inconsistent results. Most of the epidemiologic studies
3 are limited by their small sample sizes and selection bias and lack of generalizability due
4 to a focus on women seeking help for infertility. Most of the studies control for multiple
5 potential confounders, such as smoking status and age. The studies from the toxicological
6 literature show that Pb exposure to females affects litter size (decreased litter size), sex
7 ratio (ratio of male to female offspring in a litter) and ovarian viability, albeit often at
8 higher dose of Pb. However, the bulk of the evidence including the current and historical
9 Pb literature (U.S. EPA. 2006b) indicate that increased Pb exposure may decrease female
10 fertility.
5.8.5.3 Ovaries, Embryo Development, Placental function, and
Spontaneous Abortions
11 The 2006 Pb AQCD included studies of Pb exposure among men and women and their
12 associations with spontaneous abortions. The 2006 Pb AQCD concluded that overall
13 there was little evidence to support an association between Pb exposure among women
14 and spontaneous abortion (U.S. EPA. 2006b). Most of the studies examined in the
15 2006 Pb AQCD assigned exposure based on living near a smelter or working in
16 occupations that often result in Pb exposure and the results of these studies were
17 inconsistent. Little evidence was available in the 2006 Pb AQCD to suggest an
18 association with paternal Pb levels, and no recent studies have been performed to
19 examine paternal Pb levels and spontaneous abortion. Since the 2006 Pb AQCD, multiple
20 epidemiologic studies have been published that examine Pb levels in women and their
21 possible association with spontaneous abortion. Table 5-47 provides information on these
22 longitudinal and cross-sectional studies. Additionally, toxicological studies have studied
23 the effects of Pb on fetal loss and the contribution of the ovaries and placenta to fetal loss.
November 2012 5-622 Draft - Do Not Cite or Quote
-------
Table 5-47 Summary of recent epidemiologic studies of associations between Pb levels and
spontaneous abortions.
Reference
(Presented in
order of
appearance in the
text)
Vigeh et al.
(2010)
Study
Location
Tehran,
Iran
2006-2008
Outcome
Pregnancy
ended before
20 weeks of
gestation
Study population
Women who were non-
smokers, non-obese, had
no chronic health
conditions, had their last
menstrual period less than
1 2 weeks prior, and were
pregnant with a singleton
infant
N:
Methodological
Details
Longitudinal
cohort study using
t-test and logistic
regression
Pb
Biomarker
Maternal blood
Pb
during weeks
8-1 2 of
pregnancy
Mean Pb
(SD)
in ug/dL
3.8 (2.0)
Spontaneous
abortion'
3.51 (1.42)
Non-
spontaneous
abortion'
3.83 (1 .99)
Adjusted Effect
Estimates
T-test for difference in
mean values: 0.41
OR:
0.331 (95% Cl: 0.011,
10.096) for an
increase in log-
transformed blood Pb
(units not given,
assume 1 ug/dL)
Potential
Confounders
Adjusted for
in Analysis
Age, parity,
hematocrit,
passive
cigarette
smoking
exposure
a 1 spontaneous abortions
=15
N:
No spontaneous abortions
=336
Yin et a I. (2008) Shanxi Anembryonic Women age 25-35 yr old
Province, pregnancy and at 8-1 2 weeks of
China gestation at study entry;
2004-2006 cases were anembryonic
pregnancies and controls
were normal pregnancies
that ended in a live birth
between 37-42 weeks
N:
Cases
=40
N:
Controls
=40
Case-control Maternal blood
study using t-test Pb after
miscarriage for
cases and at
study
enrollment for
controls
Cases:
5.3 (95% Cl:
5.2, 5.9)
Controls:
4.5 (95% Cl:
37, 5.0)
Comparisons None
between log-
transformed blood Pb
levels of cases and
controls performed via
Student's t-test had a
p-value of 0.03
November 2012
5-623
Draft - Do Not Cite or Quote
-------
Reference
(Presented in
order of
appearance in the
text)
Lamadrid-
Figueroa et al.
(2007)
Gundacker et al.
(2010)
Study
Location Outcome
Mexico Previous
City, miscarriage
Mexico
1997-1999,
2001 -2004
Vienna, Previous
Austria miscarriage
2005
Study population
Women who had a
previous pregnancy and
were currently pregnant
with gestational age of
£ 14 weeks
N:
> 1 previous miscarriages
=71
N:
No previous miscarriages
=136
Women recruited during the
second trimester of
pregnancy
N:
a 1 previous miscarriages
=8
N:
No previous miscarriages
=22
Methodological
Details
Cross-sectional
study using
Poisson
regression
Cross-sectional
study using non-
parametric tests
Pb
Biomarker
Maternal and
umbilical cord
blood Pb,
maternal bone
Pb
Whole
placentas
shortly after
hirf h
DIIIM
Mean Pb
(SD)
in ug/dL
Overall:
Blood Pb: 6.2
(4.5)
Plasma Pb:
0.014(0.013)
Cases:
Blood Pb: 5.8
(3.4)
Plasma Pb:
0.014(0.013)
Controls:
Blood Pb: 6.5
(4.9)
Plasma Pb:
0.013(0.013)
Median
(IQR):
25.8(21.0,
36.8)
Adjusted Effect
Estimates
Categorized Plasma
Blood Pb ratio:
Isttertile:
1 .00 (Ref)
2nd tertile:
1.16(p-value0.61)
3rd tertile:
1.90(p-value0.015)
IRR (95%CI) Per 1
SD increase: Plasma
Pb
1.12(p-value0.22)
Blood Pb
0.93 (p-value 0.56)
Plasma/Blood Pb ratio
1.1 8 (p-value 0.02)
Patella Pb
1.1 5 (p-value 0.39)
Tibia Pb
1 .07 (p-value 0.56)
Median Placenta Pb:
Women who had not
previously miscarried:
27 ug/kg
Women who had
previously miscarried:
39 ug/kg
(p-value for
difference: 0.039)
Potential
Confounders
Adjusted for
in Analysis
Age, education
N/A
November 2012
5-624
Draft - Do Not Cite or Quote
-------
1 A longitudinal study examining spontaneous abortions occurring early in the pregnancy
2 was conducted in Iran (Vigeh et al.. 2010). Mean blood Pb concentrations, measured at
3 8-12 weeks of pregnancy, were similar in women who did and did not have spontaneous
4 abortions. Higher blood Pb levels were not associated with greater odds of spontaneous
5 abortions before 20 weeks of pregnancy. Yin et al. (2008) performed a study in the
6 Shanxi Province of China to examine if plasma Pb levels were associated with
7 anembryonic pregnancies (spontaneous abortions during the first trimester, which
8 account for 15% of all spontaneous abortions). Women were enrolled at 8-12 weeks of
9 gestation. Women who delivered a term pregnancy had mean plasma Pb levels that were
10 lower than those of women who had an anembryonic pregnancy (plasma Pb measured at
11 the time of miscarriage for cases and at 8-12 weeks for controls). Of note, among cases
12 plasma Pb level was inversely correlated with folate and vitamin B12, but this correlation
13 was not observed among those who delivered at term; no models examining plasma Pb
14 levels adjusted for nutrient status. A study in Mexico City examined a group of pregnant
15 women (maximum gestational period at enrollment was 14 weeks) who had previously
16 been pregnant and either given birth or had a spontaneous abortion (Lamadrid-Figueroa
17 et al.. 2007). Women in the highest tertile of plasma/blood Pb ratio had higher rates of
18 previous spontaneous abortions than did women in the lowest tertile. The authors state
19 that the plasma/whole blood ratio represents the bioavailability of Pb, which is capable of
20 crossing the placental barrier for a given blood concentration. No association was
21 observed when examining the relationship between Pb and spontaneous abortions using
22 whole blood, plasma, or bone Pb alone. Similarly, a study of placental Pb levels among
23 pregnant women in Austria observed higher placenta Pb levels among women who had
24 miscarried a previous pregnancy compared to women who had not miscarried a previous
25 pregnancy (Gundacker et al.. 2010). It is important to note that the number of women
26 included in the study was small (only 8 women reported previously having a miscarriage)
27 In toxicological studies, isolated embryo cultures are often used to understand the
28 mechanisms responsible for aberrant embryo development as it may contribute to
29 teratogenesis, fetal loss or negative postnatal pup outcomes. Nandi et al. (2010)
30 demonstrated an exposure concentration-dependent decline in embryo development of
31 fertilized buffalo oocytes cultured for 24 hours in medium containing 0.05-10 (ig/mL
32 Pb acetate as evidenced by reduced morula/blastocyst yield and increased four-to eight-
33 cell arrest, embryo degeneration, and asynchronous division. This study provides
34 evidence of the negative effect of Pb on embryo development and contributes
35 mechanistic understanding to Pb-dependent pregnancy loss.
36 A possible explanation for reduced fertility and impaired female reproductive success as
37 a result of Pb exposure is changes in morphology or histology in female sex organs and
38 the placenta (Dumitrescu et al.. 2007; U.S. EPA. 2006b). Wang et al. (2009e) observed
November 2012 5-625 Draft - Do Not Cite or Quote
-------
1 that elevated maternal blood Pb (0.6-1.74 (JVI, -12.4-36.0 (ig/dL) compared to control
2 (0.04 (JVI, -0.83 (ig/dL) were associated with decreased fetal body weight, pup body
3 length, and placental weight in Wistar rats. The authors reported that placentae from
4 Pb-exposed groups showed concentration-dependent increasing pathology of
5 cytoarchitecture and cytoplasmic organelles. The authors also reported a positive
6 expression of NF-KB, a transcription factor that controls the expression of genes involved
7 in immune responses, apoptosis, and cell cycle, in the cytotrophoblasts, decidual cells,
8 and small vascular endothelial cells in rat placenta under a low-level Pb exposure
9 condition which correlated with low blood Pb levels.
10 Pb-exposed (273 mg/L or 819 mg/L in drinking water, 500 or 1,500 ppm Pb acetate,
11 respectively) male rats from Anjum et al. (2010) that had an exposure concentration-
12 dependent decreases in serum testosterone, decreased male reproductive organ weight
13 and decreased sperm were mated to untreated females. These untreated dams bred to the
14 Pb exposed males had male related exposure concentration-dependent decreased
15 implantation rate and higher pre- and post-implantation loss, indicating paternally
16 mediated fetal loss. The magnitude of these effects in dams was dependent on the
17 concentration of Pb exposure in their male mating partners.
18 As observed in sperm cells, Pb stimulates changes in antioxidant enzyme activity in rat
19 ovaries indicating that oxidative stress may be a contributing factor in Pb-induced ovarian
20 dysfunction. Nampoothiri et al. (2007) observed a reduction in SOD activity and an
21 increase in CAT activity along with a decrease in glutathione content and an increase in
22 lipid peroxidation in rat granulosa cells after 15 days of Pb treatment (subcutaneously
23 administered Pb (0.05 mg/kg body weight daily before mating and during pregnancy)
24 with a resulting blood Pb of 2.49 (ig/mL).
25 Previous studies demonstrated that Pb accumulates in the ovaries and causes histological
26 changes, thus contributing to Pb-induced effects on female fertility (U.S. EPA. 2006b). In
27 support of historical studies, recent studies demonstrate Pb-induced histological changes
28 in ovarian cells of pigs (Kolesarova et al.. 2010) and rats (Nampoothiri et al.. 2007;
29 Nampoothiri and Gupta, 2006). Kolesarova et al. (2010) observed a reduction of the
30 monolayer of granulosa cells after Pb addition (0.5 mg/mL, 18 hours culture).
31 Nampoothiri and Gupta (2006) reported that Pb exposure caused a decrease in cholesterol
32 and total phospholipid content in the membranes of granulosa cells which resulted in
33 increased membrane fluidity (subcutaneously administered Pb, 0.05 mg/kg body weight
34 daily before mating and during pregnancy with a resulting blood Pb of 2.49 (ig/mL).
35 Overall, the recent studies support the conclusions of the 2006 Pb AQCD (U.S. EPA.
36 2006b) that there is mixed evidence among epidemiologic studies to suggest an
37 association between Pb and spontaneous abortions. It is important to note that studies of
November 2012 5-626 Draft - Do Not Cite or Quote
-------
1 spontaneous abortions are difficult to conduct. The majority of spontaneous abortions are
2 during the first trimester, which makes them difficult to capture. Women may miscarry
3 before being enrolled in a study and many women may not have known they were
4 pregnant when they miscarried. This limits the ability to detect subtle effects, especially
5 if higher Pb levels do lead to increased risk of early spontaneous abortions. In addition,
6 some studies are limited by their retrospective examination of current Pb levels in
7 relation to previous miscarriages. Sample size is another limitation of the available
8 epidemiologic studies. The epidemiologic studies also had little control for potential
9 confounding factors, with some studies including no potential confounders in their
10 analyses. Toxicological data provide mechanistic understanding of the contribution of Pb
11 exposure to spontaneous abortions. These laboratory data show that Pb exposure
12 impaired placental function, induced oxidative stress and histological changes in the
13 ovaries, and affected embryo development. The toxicological and epidemiologic data
14 provide inconsistent findings for the role of Pb in spontaneous abortions.
5.8.5.4 Effects on Breast Milk
15 Experiments in laboratory animals have shown that dietary manipulation of maternal
16 fatty acid (FA) levels in diet can worsen Pb-related behavioral effects of offspring after
17 lactational Pb exposure (Lim et al., 2005). To determine if components of dam milk
18 contributed to this change, dam milk fatty acids were altered via diet. Diets deficient in
19 n-3 fatty acids can lead to a deficiency of DHA, which is essential for proper nervous
20 system development. Lim et al. (2005) found that dam Pb exposure (Long-Evans rats,
21 2,000 ppm Pb acetate trihydrate/BW) during lactation (PNDO-PND21) led to a decrement
22 in non-essential fatty acids in the maternal organs at PND25 (mean [SD] blood Pb levels
23 in dams: 308 [56] (ig/dL). In animals with a diet deficient in n-3 FAs, there was a Pb-diet
24 interaction with a specific size PUFA (i.e., a 20-carbon n-6 PUFA). In general, Pb
25 exposure caused a decrement in shorter chain monounsaturated and saturated FAs in
26 maternal organs.
27 Dietary supplementation with calcium can be an especially important contributor to Pb
28 mobilization during periods of high calcium demand including pregnancy/lactation. For
29 example, mothers with elevated blood Pb levels given calcium phosphate and ascorbic
30 acid supplementation during lactation had a 90% decrease in placental Pb content and a
31 15% decrease in the concentration of Pb in breast milk (Altmann et al.. 1981) versus the
32 control group that did not receive dietary treatment. Another study (Gulson et al., 2004a)
33 has shown that calcium supplementation during the lactation is less beneficial in
34 modulating maternal blood Pb levels (mean blood Pb at first sampling was 2.4 (ig/dL);
35 the Gulson cohort (Gulson et al.. 2004a) was limited by power (n=10 women). In a
November 2012 5-627 Draft - Do Not Cite or Quote
-------
1 cohort of women from Mexico City, daily calcium supplementation during lactation
2 reduced maternal blood Pb by 15-20% and Pb in breast milk by 5-10% (Ettinger et al..
3 2004a). Another study by the same investigators showed that using calcium supplements
4 daily during pregnancy also reduced blood Pb levels during pregnancy (Ettinger etal..
5 2009) with the effect strongest in women with higher biomarkers of Pb exposure
6 (elevated baseline bone Pb or >5 (ig/dL blood Pb) or in women with higher Pb exposure
7 (self-reported use of Pb-glazed ceramics). Thus, dietary modulation with calcium
8 supplementation during pregnancy and lactation may decrease the amount of Pb to which
9 the developing fetus of infant is exposed. The evidence for this seems especially strong
10 for protection during pregnancy and more mixed for protective effects of calcium during
11 lactation.
5.8.5.5 Summary of Effects on Female Reproductive Function
12 In summary, Pb exposure was found to affect female reproductive function as
13 demonstrated by both epidemiologic and toxicological studies. Some evidence is also
14 available regarding blood Pb levels and altered hormone levels in adults, but varied
15 among studies. The differences may have been due to the different hormones examined
16 and the different timing in the menstrual and life cycles of the women. Although studies
17 reported inconsistent findings for the association between Pb and fertility, there is some
18 evidence of a potential relationship. Adjustment for potential confounders varies from
19 study to study, with some potentially important confounders, such as BMI, not included
20 in all studies. Also, many epidemiologic studies are limited by small samples sizes and
21 are generally of women attending infertility clinics, which presents the possibility of
22 selection bias and lack of generalizability. Toxicological studies found effects on female
23 reproductive function after prenatal or early postnatal exposures. Further coherence for
24 these findings in laboratory animal models is found in with findings in the ecological
25 literature for the effects of Pb exposure on reduced fecundity in terrestrial and aquatic
26 animal species (Sections 7.4.5.2. 7.3.4.2. 7.3.4.3. and 7.4.5.3). Although epidemiologic
27 and toxicological studies provide information on different exposure periods, both types of
28 studies support the conclusion that Pb affects at least some aspects of female reproductive
29 function.
November 2012 5-628 Draft - Do Not Cite or Quote
-------
5.8.6 Summary and Causal Determination
1 Many epidemiologic and toxicological studies of the effects of Pb on reproductive and
2 developmental outcomes have been conducted since the 2006 Pb AQCD. The evaluation
3 of causal relationships with Pb exposure focuses on four areas: developmental effects,
4 birth outcomes, reproductive function among males, and reproductive function among
5 females. The sections that follow describe the evaluation of evidence for these outcomes
6 with respect to causal relationships with Pb exposure using the framework described in
7 Table II of the Preamble. The application of the key supporting evidence to the causal
8 framework is summarized in Table 5-48.
5.8.6.1 Effects on Development
9 The 2006 Pb AQCD (U.S. EPA. 2006b) reported Pb-associated effects on development in
10 toxicological studies. Multiple recent epidemiologic studies of Pb and puberty have
11 shown associations between concurrent blood Pb levels and delayed pubertal onset for
12 girls and boys. Delayed puberty has been linked to decreased peak bone mass and
13 increased risk of osteoporotic fractures (Gilsanz etal., 2011; Naves et al., 2005). In cross-
14 sectional epidemiologic studies of girls (ages 6-18 years) with mean and/or median
15 concurrent blood Pb levels less than 5 (ig/dL consistent associations with delayed
16 pubertal onset (measured by age at menarche, pubic hair development, and breast
17 development) were observed. In boys (ages 8-15 years), fewer epidemiologic studies
18 were conducted but associations were observed, including associations among boys in a
19 longitudinal study. These associations are consistently observed in populations with
20 concurrent blood Pb levels <10(ig/dL. Potential confounders considered in the
21 epidemiologic studies of both boys and girls that performed regression analyses varied.
22 Most studies controlled for age and BMI. Other variables, such as measures of diet, SES,
23 and race/ethnicity, were included in some of the studies. Adjustment for nutritional
24 factors was done less often and this could be an important confounder. A study using a
25 cohort of girls from the NHANES analysis controlled for various dietary factors as well
26 as other potential confounders and reported an association between increased concurrent
27 blood Pb levels and delayed pubertal onset (Selevan et al.. 2003). A limitation across
28 most of the epidemiologic studies of blood Pb levels and delayed puberty is their cross-
29 sectional design, which does not allow for an understanding of temporality. There is
30 uncertainty with regard to the exposure frequency, timing, duration, and level that
31 contributed to the associations observed in these studies.
32 Recent toxicological studies show that pubertal onset is one of the more sensitive markers
33 of Pb exposure with effects observed after gestational exposures leading to blood Pb
November 2012 5-629 Draft - Do Not Cite or Quote
-------
1 levels in the female pup of 3.5-13 (ig/dL (lavicoli et al.. 2006a; lavicoli et al. 2004).
2 Toxicological studies have reported delayed male sexual maturity as measured with sex
3 organ weight, among other outcomes, seeing significant decrements at blood Pb levels of
4 34 (ig/dL (Sokol et al. 1985). Thus, data from the toxicological literature and from
5 epidemiologic findings demonstrate that puberty onset in both males and females is
6 delayed with Pb exposure.
7 Findings from epidemiologic studies of postnatal growth are inconsistent and findings
8 from the toxicological literature are mixed with recent growth findings showing adult
9 onset obesity. Toxicological studies demonstrated that the effects of Pb exposure during
10 early development include impairment of retinal development and alterations in the
11 developing hematopoietic and hepatic systems. Affected developmental outcomes with
12 Pb exposure also included effects on the eyes and teeth.
13 The collective body of evidence integrated across epidemiologic and toxicological
14 studies, based on the findings of delayed pubertal onset among males and females, is
15 sufficient to conclude that there is a causal relationship between Pb exposure and
16 developmental effects.
5.8.6.2 Effects on Birth Outcomes
17 Overall, results of pregnancy outcomes were similar to those of the 2006 Pb AQCD;
18 (U.S. EPA. 2006b) inconsistent evidence of a relationship with Pb was available for
19 preterm birth. The 2006 Pb AQCD included a few studies that reported potential
20 associations between Pb and neural tube defects, but the recent epidemiologic studies
21 found no association. Some associations were observed between Pb and low birth weight
22 when epidemiologic studies used measures of postpartum maternal bone Pb or air
23 exposures. The associations were less consistent when using maternal blood Pb measured
24 during pregnancy or at delivery or umbilical cord and placenta Pb (maternal blood Pb or
25 umbilical cord and placenta Pb were the biomarkers most commonly used in studies of
26 low birth weight) but some associations between increased Pb levels and decreased low
27 birth weight/fetal growth were observed. The effects of Pb exposure during gestation in
28 animal toxicological studies included mixed findings with some studies showing
29 reduction in litter size, implantation, and birth weight, and some showing no effect.
30 Based on the mix of inconsistent results of studies on various birth outcomes but some
31 associations observed in well-conducted epidemiologic studies of preterm birth and low
32 birth weight/fetal growth, the evidence is suggestive of a relationship between Pb
33 exposure and birth outcomes.
November 2012 5-630 Draft - Do Not Cite or Quote
-------
5.8.6.3 Effects on Male Reproductive Function
1 Toxicological evidence and supporting epidemiologic evidence indicate that a causal
2 relationship exists between Pb exposure and effects on male reproductive function. Key
3 evidence is provided by toxicological studies in rodents, non-human primates, and rabbits
4 showing detrimental effects on semen quality, sperm and fecundity/fertility with
5 supporting evidence in epidemiologic studies of associations between Pb exposure and
6 detrimental effects on sperm. This is consistent with studies reported in the
7 2006 Pb AQCD (U.S. EPA. 2006b).
8 Toxicological studies with relevant Pb exposure routes reported effects on rodent sperm
9 quality and sperm production rate [(Sokol and Berman. 1991; Sokol etal.. 1985). blood
10 Pb level 34-37 (ig/dL], sperm DNA damage lYNava-Hernandez et al.. 2009). blood Pb
11 levels 19 and 22 jig/dL], and histological or ultrastructural damage to the male
12 reproductive organs in studies from rodents [(El Shafai et al.. 2011). blood Pb level
13 5.1 (ig/dL] and non-human primates [(Cullen etal.. 1993). blood Pb level 43 (ig/dL].
14 These effects were found in animals exposed to Pb during peripuberty or adults for 1
15 week to 3 months. The toxicological studies reported an association between Pb exposure
16 and decreases in reproductive organ weight and organ histological changes in the testes
17 and germ cells. Subfecundity (decreased number of pups born/litter) was reported in
18 unexposed females mated to Pb exposed males. Also, sperm from Pb-exposed rats (blood
19 Pb level: 33 to 46 ug/dL) used for in vitro fertilization of eggs harvested from unexposed
20 females yielded lower rates of fertilization. (Sokol etal.. 1994). Supporting evidence was
21 provided by decrements in sperm quality from rabbits administered Pb subcutaneously
22 (blood Pb levels of 25 (ig/dL) (Moorman et al.. 1998).
23 The detrimental effects of Pb on sperm were observed in epidemiologic studies with
24 concurrent blood Pb levels of 25 (ig/dL and greater among men occupationally exposed
25 (Hsu et al.. 2009b: Kasperczyk et al.. 2008; Naha and Manna. 2007; Nahaand
26 Chowdhury. 2006). The epidemiologic studies were limited due to these high exposure
27 levels among the occupational cohorts and the lack of consideration for potential
28 confounding factors, including other occupational exposures. Studies among men with
29 lower Pb levels were limited to infertility clinic studies, which may be a biased sample
30 and lack generalizability. However, a well-conducted epidemiologic study that enrolled
31 men going to a clinic for either infertility issues or to make a semen donation and
32 controlled for other metals as well as smoking reported a positive association with
33 various detrimental effects in sperm (Telisman et al.. 2007). The median concurrent
34 blood Pb levels in this study were 4.92 (ig/dL (range: 1.13-14.91). A similar study
35 (Meeker et al.. 2008) also reported possible associations between concurrent blood Pb
November 2012 5-631 Draft - Do Not Cite or Quote
-------
1 and various semen parameters, but the results were extremely imprecise, making it
2 difficult to draw conclusions.
3 Similar to the 2006 Pb AQCD (U.S. EPA. 2006b). recent epidemiologic and toxicological
4 studies reported inconsistent results regarding hormone aberrations associated with Pb
5 exposure. Due to the complexity of the reproductive system, uncertainty exists as to
6 whether Pb exerts its toxic effects on the reproductive system by affecting the
7 responsiveness of the hypothalamic-pituitary-gonad axis, by suppressing circulating
8 hormone levels or by some other pathway. Mixed findings were also apparent among
9 epidemiologic studies of fertility among men.
10 More recent toxicological studies suggest that oxidative stress is a major contributor to
11 the effects of Pb exposure on the male reproductive system, providing mode of action
12 support. The effects of ROS may involve interference with cellular defense systems
13 leading to increased lipid peroxidation and free radical attack on lipids, proteins, and
14 DNA. Several recent studies showed that Pb induced an increased generation of ROS
15 within the male sex organs, and germ cell injury, as evidenced by aberrant germ cell
16 structure and function. Co-administration of Pb with various antioxidant compounds
17 either eliminated Pb-induced injury or greatly attenuated its effects. In addition, many
18 studies that observed increased oxidative stress also observed increased apoptosis which
19 is likely a critical underlying mechanism in Pb-induced germ cell DNA damage and
20 dysfunction.
21 Based on the consistency and coherence of findings for the detrimental effects of Pb
22 exposure on sperm and semen in the toxicological literature, the support from
23 epidemiologic studies, and biological plausibility provided by mode of action evidence,
24 the evidence is sufficient to conclude that there is a causal relationship between Pb
25 exposures and male reproductive function.
5.8.6.4 Effects on Female Reproductive Function
26 Epidemiologic and toxicological studies of reproductive function among females
27 investigated whether Pb biomarker levels were associated with hormone levels, fertility,
28 estrus cycle changes, and morphology or histology of female reproductive organs
29 including the placenta. Toxicological studies reported in the 2006 Pb AQCD (U.S. EPA.
30 2006b) reported associations between Pb exposure and female reproductive function,
31 although little evidence was provided by epidemiologic studies. Some epidemiologic
32 studies have shown associations with concurrent blood Pb levels and altered hormone
33 levels in adults, but varied among studies, likely due to the different hormones examined
34 and the different timing in the menstrual and life cycles. There is some evidence of a
November 2012 5-632 Draft - Do Not Cite or Quote
-------
1
2
3
4
5
6
7
8
9
10
11
12
13
potential relationship between Pb exposure and female fertility, but findings are mixed.
The majority of the epidemiologic studies are cross-sectional, and adjustment for
potential confounders varies from study to study, with some potentially important
confounders, such as BMI, not included in all studies. Also, most of the studies have
small samples sizes and are generally of women attending infertility clinics.
Toxicological study design often employs prenatal or early postnatal Pb exposures with
Pb contributing to placental pathology and inflammation, decreased ovarian antioxidant
capacity, altered ovarian steriodogenesis and aberrant gestational hormone levels.
Although epidemiologic and toxicological studies provide information on different
exposure periods, both types of studies support the conclusion that Pb possibly affects at
least some aspects of female reproductive function. Overall, the relationship observed
with female reproductive outcomes is sufficient to conclude that there is a suggestive
relationship between Pb exposure and female reproductive function.
Table 5-48 Summary of evidence supporting reproductive and developmental
causal determinations.
Attribute in Causal
Framework3
Key Supporting Evidence
Recent References
Pb Exposure or
Blood Pb Levels
Associated with Effects0
Effects on Development - Causal
Delayed Puberty Onset
Consistent Consistent evidence in multiple large
associations cross-sectional epidemiologic studies for
between higher females and males plus a longitudinal
blood Pb levels in study for males. Most of these studies
high-quality have large sample sizes and controlled for
epidemiologic potential confounding by covariates such
studies as age, BMI, and education/SES.
Tomoum et al. (2010).
Mauser et al. (2008),
Williams et al. (2010).
Denham et al .(2005).
Naickeretal. (2010).
Wu et al. (2003b).
Gollenberg et al. (2010)
Sections
5.8.1.1 and 5.8.1.2
Concurrent blood Pb levels:
<10 ug/dL
Epidemiologic
evidence supported
by consistent
toxicological findings
with relevant Pb
exposure
Evidence clearly
describes Mode of
Action.
A large study using females aged 8-18
years from the NHANES III study also
controlled for various dietary factors and
reported associations between blood Pb
levels and delayed puberty onset
Consistent toxicological evidence from
multiple laboratories of delayed male and
female puberty onset with Pb exposure via
diet or oral gavage in rodents
Toxicological evidence describes HPG axis
dysfunction providing mechanism of action
support for delayed puberty findings. MOA
further supported by IGF-1 changes
contributing to Pb-induced delay in puberty
onset.
Selevan et al. (2003)
Dumitrescu et al. (2008c).
lavicoli et al. (2006a).
Pine et al. (2006)
Sections
5.8.1.1 and 5.8. 1.2
Section 5.8.1.1
Blood Pb level after dietary
exposure from gestation to
estrus:
3.5-13ug/dL
November 2012
5-633
Draft - Do Not Cite or Quote
-------
Attribute in Causal
Framework3
Key Supporting Evidence
Recent References
Pb Exposure or
Blood Pb Levels
Associated with Effects0
Postnatal Growth
Available
toxicological studies
reported mixed
findings of effects of
Pb on postnatal
growth
There are mixed findings in the
toxicological literature on Pb exposure and
postnatal growth with some studies
showing stunted growth in animals
exposed to Pb and some showing no
effects.
Recent toxicological evidence of effect of
Pb on postnatal growth: obesity in adult
male offspring after gestational +
lactational Pb exposure
Section 5.8.1.3
Available
epidemiologic
findings of
associations
between higher
blood Pb levels and
postnatal growth are
inconsistent
Multiple studies, mostly cross-sectional, for
children of varying ages have reported
inconsistent results for the association
between blood Pb levels and various
measures of growth
Section 5.8.1.3
Impaired Organ Systems
Consistent
toxicological
findings of effects
on sensory organ
systems, bone,
teeth, and Gl
system but not
always at relevant
Pb exposure levels.
Relevant gestational and
exposure of rats resulted
lactational Pb Fox et al. (2008)
in retinal ERG
aberrations and increased retinal cell layer
thickness.
Blood Pb level after
gestational-lactational
exposure:
10-12 ug/dL
Effects on Birth Outcomes - Suggestive
Inconsistent findings
in epidemiologic
studies of various
birth outcomes
Inconsistent findings for studies for birth
defects, preterm birth, and low birth
weight/fetal growth.
A few well-conducted epidemiologic studies
of preterm birth and low birth weight/fetal
growth using measures of maternal blood
Pb at the time of pregnancy reported
associations.
See Section 5.8.3 (and all
subsections): 5.8.3.1.
5.8.3.2. 5.8.3.3. 5.8.3.4
Jelliffe-Pawlowski et al.
(2006).
Vigehetal.(2011).
Zhu et al. (2010).
Chenetal. (2006a).
Gundacker et al. (2010)
Maternal pregnancy blood
Pb levels:
>10 ug/dL
Inconsistent findings
in toxicological
literature for birth
outcomes
The toxicological literature reported mixed
findings with some studies showing smaller
litter size (fewer pups born) or decreased
birth weight with Pb exposure and some
studies showing no effect.
See Section 5.8.3.4 and
5.8.3.1
November 2012
5-634
Draft - Do Not Cite or Quote
-------
Attribute in Causal
Framework3
Key Supporting Evidence
Recent References
Pb Exposure or
Blood Pb Levels
Associated with Effects0
Effects on Male Reproductive Function - Causal
High-quality and
consistent findings in
toxicological studies of
detrimental effects of
Pb exposure on
sperm or semen in
multiple species.
Decreased sperm counts, decreased
sperm production rate, dose-dependent
suppression of spermatiogenesis in
rodents with drinking water Pb exposure.
All results:
Section 5.8.4.1
Sokoletal. (1985)
Sokol and Berman (1991)
Blood Pb level after adult
drinking water exposure for
30 days: 34 ug/dL
Blood Pb level after
peripubertal or adult drinking
water exposure for 30 days:
35 and 37 ug/dL.
Ultrastructural damage and histological to
non-human primates testis and
seminiferous tubules.
Histologic damage to rodent seminiferous
tubules including spermatids and
developing sperm.
Ultrastructural abnormalities to rat
spermatogenesis.
Direct effects on rodent sperm DMA after
drinking water Pb exposure.
Sperm from Pb exposed rats used for in
vitro fertilization of eggs harvested from
unexposed females yielded lower rates of
fertilization.
Semen and sperm quality in rabbits with
subcutaneous Pb treatment; Ultrastructural
damage to spermatids with i.p. injection of
Pb.
Findings of detrimental effects of Pb
exposure on sperm from multiple species
(Deer, Asian earthworm, rainbow trout,
marine worm, H. elegans,Fathead
minnow)
Singh et al. (1993a)
Foster etal. (1998)
El Shafai et al. (2011)
Murthv etal.(1995)
Nava-Hernandez et al.
(2009)
Sokol et al.(1994)
Moorman etal.(1998).
See Ecological Effects;
Sections 7.4.12.1 and
7.4.21.1)
Maximum blood Pb levels
after daily oral Pb exposure
(gelatin capsule) during
infancy, post infancy, or over
a lifetime (up to 10yr):
32 to 36 ug/dL
Blood Pb level after adult
exposure (oral gavage) for 3
months: 5.31 ug/dL
Blood Pb level after i.p.
injection for 16 days:
7.4 ug/dL
Blood Pb level after adult
exposure for 13 weeks:
19 and 22 ug/dL
Blood Pb level after adult
exposure for 14-60 days:
33-46 ug/dL
Blood Pb level after adult
exposure for 15 weeks:
16-24ug/dL
Toxicological evidence
is supported by
consistent findings in
epidemiologic studies
of associations
between higher blood
Pb levels and
decrements in sperm
count and quality in
occupational cohorts
Consistent evidence from studies of
occupational cohorts with high blood Pb
levels. Results from occupational cohorts
may have been confounded by other
workplace exposures, which were not
adjusted for in the epidemiologic studies.
Potential confounding by smoking was
considered in one study.
Results less consistent at lower blood Pb
level. A well-conducted epidemiologic
study at an infertility clinic reported
associations between detrimental effects
in sperm and blood Pb levels after
controlling for smoking and other metal
exposure. A similar study also reported
some elevated effect estimates but the
results were too imprecise to draw
definitive conclusions.
Naha and Manna (2007).
Naha and Chowdhury
(2006). Hsu et al.
(2009b). Kasperczyk et al.
(2008)
Telisman etal. (2007),
Meeker etal. (2008)
Concurrent blood Pb levels:
> 25 ug/dL
Concurrent blood Pb level:
<10ug/dL
November 2012
5-635
Draft - Do Not Cite or Quote
-------
Attribute in Causal
Framework3
Key Supporting Evidence
Recent References
Pb Exposure or
Blood Pb Levels
Associated with Effects0
Evidence describes
mode of action for
effects on sperm.
Inconsistent findings
of associations
between Pb levels
and hormone levels in
epidemiologic studies;
few studies available
Lack of large, well-
conducted
epidemiologic studies
examining
associations between
Pb levels and fertility
Limited toxicological
findings of Pb
exposure inducing
effects on fertility
Consistent MOA evidence in reproductive
organs of Pb-exposed male animals of
increased apoptosis, decreased
antioxidant activity (SOD and CAT), and
increased oxidative stress (MDA).
There are a small number of studies
examining hormone levels and the results
are inconsistent.
The few epidemiologic studies examining
this outcome generally have small
samples sizes and are drawn from men
attending infertility clinics.
Paternal Pb exposure resulted in less
successful copulation, fewer implantations,
and longer periods of time copulating for
successful matings. Unexposed females
with Pb-exposed male partners did not
have fewer pregnancies, but did produce
smaller litters.
Sections 5. 8. 4.1 and
5.8.4.2
Telisman et al. (2007).
Naha and Manna (2007).
Hsiehetal. (2009a),
Meeker etal. (2010),
Mendiola et al. (2011)
Section 5.8.4.2
Kizileretal. (2007).
Bloom etal. (201 1b).
Bloom etal. (2010).
Gracia et al. (2005)
Section 5.8.4.3
Anjum et al. (2010) 45-day exposure of adult
Sainatha et al. (2011) male rats to 500 or
Pace et al. (2005) 1 ,500 ppm Pb acetate
Section 5.8.4.3 exposure in drinking water,
followed by behavioral
mating studies with
unexposed females.
Effects on Female Reproductive Function - Suggestive
Epidemiologic studies
of Pb levels and
hormones
demonstrate
associations but are
inconsistent overall
Evidence in some high-quality cross-
sectional epidemiologic studies
demonstrates associations with hormone
levels but results are mixed and vary by
hormone examined and timing in a
woman's menstrual and life cycles. In
addition, the potential confounders vary
from study to study, with some potentially
important confounders, such as BMI, not
included in all studies.
Jackson et al. (2011).
Pollack etal. (2011).
Chang et al. (2006).
Krieg (2007)
Section 5.8.5.1
Concurrent mean blood
Pb levels:
<5 ug/dL
Lack of large, well-
conducted
epidemiologic studies
examining
associations between
Pb levels and fertility
Epidemiologic studies of this association
are limited by the small sample sizes
included in those studies. In addition, most
of the study populations were drawn from
women undergoing IVF and/or attending
infertility clinics.
Section 5.8.5.2
Toxicological studies
of Pb and effects on
female reproduction
demonstrate effects in
some studies.
Evidence in the toxicological literature of
Pb contributing to placental pathology and
inflammation, decreased ovarian
antioxidant capacity, altered ovarian
steriodogenesis and aberrant gestational
hormone levels.
Section 5.8.5.1 and
5.8.5.3
"Described in detail in Table II of the Preamble.
bDescribes the key evidence and references contributing the most heavily to causal determination. References to earlier sections
indicate where full body of evidence is described.
""Describes the blood Pb level in humans with which evidence is substantiated or the blood Pb levels or Pb exposure
concentrations in animals relevant to humans.
November 2012
5-636
Draft - Do Not Cite or Quote
-------
5.9 Effects on Other Organ Systems
1 The 2006 Pb AQCD described limited evidence for the effects of Pb exposure on various
2 organ systems including the liver, GI tract, endocrine system, bone and teeth, eyes, and
3 respiratory tract. These lines of evidence largely are supported by recent toxicological
4 and epidemiologic studies, although the collective evidence remains relatively limited in
5 terms of the quantity and design of studies and/or the populations examined.
5.9.1 Effects on the Hepatic System
6 Hepatotoxic effects of Pb exposure indicated in various animal models and/or human
7 populations include alterations in hepatic metabolism, hepatic cell proliferation, changes
8 in cholesterol metabolism, as well as oxidative stress-related injury.
5.9.1.1 Summary of Key Findings of the Effects on the Hepatic
System (2006 Pb AQCD)
9 The 2006 Pb AQCD (U.S. EPA. 2006b) stated that the experimental animal database
10 indicated hepatotoxic effects, including liver hyperplasia, at very high dose Pb exposures.
11 Other effects noted in the liver following exposure to Pb included altered cholesterol
12 synthesis, DNA synthesis, and glucose-6-phosphotase dehydrogenase (G6DP) activity.
13 The 2006 Pb AQCD reported that cytochrome (CYP) P450 levels decreased following
14 single doses of Pb nitrate. Induced and constitutive expression of microsomal CYP1A1
15 and CYP1A2 was inhibited by Pb exposure. Inhibition of these (Phase I) xenobiotic
16 metabolizing enzymes was accompanied by an increase in Phase II enzymes following
17 exposure to Pb nitrate and other Pb compounds, suggesting that Pb is capable of inducing
18 a biochemical phenotype similar to hepatic nodules. Studies related to Pb-induced hepatic
19 hyperplasia suggested alterations in the gluconeogenic mechanism, DNA
20 hypomethylation, changes in proto-oncogene expression, as well as cholesterol synthesis.
21 Cholesterol metabolism changes following exposure to Pb were reportedly mediated by
22 induction of several enzymes related to cholesterol metabolism as well as a decrease in
23 the cholesterol catabolizing enzyme, 7 a-hydroxylase. Tumor necrosis factor alpha
24 (TNF-a) was reported to be one of the major mitogenic signals that mediated Pb nitrate-
25 induced hepatic hyperplasia, based on findings showing that inhibitors blocking TNF-a
26 activity also blocked Pb-induced hyperplasia. Other Pb-related effects presented in the
27 2006 Pb AQCD included liver cell apoptosis mediated by Kupffer cell derived signals
28 and Pb-induced oxidative stress in vitro cell cultures. The 2006 Pb AQCD further
29 suggested that alterations in liver heme metabolism may involve changes in
November 2012 5-637 Draft - Do Not Cite or Quote
-------
1 5-aminolevulinic acid dehydrogenase (ALAD) activity, porphyrin metabolism, transferrin
2 gene expression and changes in iron metabolism.
3 With regard to human studies, the 2006 Pb AQCD stated that increases in serum liver
4 enzymes suggest that Pb exposure results in nonspecific liver injury in occupationally-
5 exposed adults. However, studies did not adjust for potential confounding factors,
6 including other occupational exposures or establish explicit associations between Pb
7 exposure and hepatic injury (i.e., observation of histopathological effects). In addition,
8 similar to effects were noted in animal studies, and decreased cytochrome P450 activity
9 was associated with higher blood Pb levels in a few studies of children or adults (drawn
10 from the general population). The 2006 Pb AQCD reported that hepatic effects in humans
11 were associated only with high blood Pb levels, i.e., >30 (ig/dL.
5.9.1.2 Recent Epidemiologic Studies
12 A few epidemiologic studies examined antioxidant status and oxidative stress effects, as
13 measured by liver biochemical parameters, associated with occupational exposure to Pb.
14 However, all of these occupationally-exposed cohorts represented populations highly
15 exposed to Pb, with mean or median blood Pb levels ranging from 29 to 53 (ig/dL.
16 Although the hepatic effects observed within these cohorts may not be generalizable to
17 the general population as a whole, they are useful in demonstrating consistent effects on a
18 number of liver outcomes, including altered liver function (i.e., changes in the level of
19 liver function enzymes), oxidative stress, and antioxidant status (Can et al.. 2008; Khan et
20 al., 2008; Patil et al., 2007). However, these studies were cross-sectional in design with
21 concurrent blood Pb measurement. Thus, there is uncertainty regarding the directionality
22 of effects and the magnitude, timing, frequency, and duration of Pb exposure that
23 contributed to the observed associations. Further, analyses did not consider potential
24 confounding by factors such as age, diet, BMI, smoking, or other occupational exposures.
25 In spray painters from Kolhapur City in western Maharashtra, India, exposed to Pb for
26 >6 hours/day for 2 to 20 years examined by Patil et al. (2007). mean concurrent (SD)
27 blood Pb levels in 30 workers (mean [SD]: 22.32 [8.87] (ig/dL) were significantly higher
28 (p <0.001, t-test) than those in the 35 concurrent controls (mean [SD]: 12.52
29 [4.08] (ig/dL), who had no history of Pb exposure and lived in rural areas. Levels of liver
30 function enzymes, including the two serum transaminase enzymes SGOT (also known as
31 AST; serum glutamic oxaloacetic transaminase/aspartate aminotransferase) and SGPT
32 (also known as ALT; serum glutamic pyruvic transaminase/alanine aminotransferase),
33 were increased in spray painters compared to those in controls, whereas total serum
34 protein levels were decreased compared to controls (p <0.01, t-test). In another
November 2012 5-638 Draft - Do Not Cite or Quote
-------
1 occupational study, Conterato et al. (In Press) investigated liver function parameters in
2 automotive painters exposed to Pb in Brazil. Mean (SD) concurrent blood Pb levels were
3 5.4 (0.4) (ig/dL in the 50 exposed painters and 1.5 (0.1) (ig/dL in the 36 unexposed
4 controls. The mean (SD) duration of exposure to Pb in painters was 133.9 (14.5) months.
5 In exposed workers, the levels of AST, but not y-glutamyltransferase, were increased
6 approximately 2-fold compared to levels in controls (p <0.05). The activity of AST was
7 positively correlated with blood Pb levels (r = 0.26, p <0.05). The authors suggested that
8 confounding exposures to toxic constituents of the paints regularly used by painters, and
9 not Pb, may be the etiological cause of decrements in AST function as these effects were
10 not also seen in battery workers with much higher blood Pb levels (49.8 (ig/dL)
11 (Conterato et al.. In Press). Co-exposure to other environmental contaminants may also
12 explain the effects that were previously reported in occupationally-exposed spray-
13 painters in Patil et al. (2007).
5.9.1.3 Recent lexicological Studies
Hepatic Metabolism
14 As stated in the 2006 Pb AQCD (U.S. EPA. 2006b). acute (e.g., single dose) treatment of
15 rodents with Pb nitrate and other Pb compounds was found to result in a decrease in
16 Phase I enzymes and a simultaneous increase in Phase II enzymes. The conclusions
17 presented in the 2006 Pb AQCD were also reviewed by Mudipalli (2007).
18 Recent studies found changes in biochemical parameters, suggestive of liver damage, in
19 animals exposed to Pb; however, the relevance to humans is uncertain because of the
20 high blood Pb levels used in animal studies and the exposure routes of Pb administration.
21 Undernourished male Wistar rats (fed low-protein diet without mineral supplements)
22 exposed to 500 ppm Pb acetate in drinking water over a 10 month period had decreases in
23 serum protein and albumin levels as well as increases in AST, ALT, serum alkaline
24 phosphatase (ALP), and gamma glutamyl transpeptidase (GGT) levels (Herman et al..
25 2009). In Pb-treated animals, the blood Pb levels steadily increased throughout the initial
26 portion of the study period, reaching a maximum of approximately 30 (ig/dL after
27 2 months. After this time, blood Pb levels rapidly increased to approximately 110 (ig/dL
28 by six months time, and remained at this level until the termination of exposure at
29 10 months. Similar biochemical changes were not observed in animals treated with
30 Pb acetate maintained on protein-adequate, mineral rich diet.
31 Similarly, mice gavaged with Pb nitrate (50 mg/kg for 40 days) also demonstrated
32 increased activities of AST, ALT, ALP, and acid phosphatase (ACP) compared to
November 2012 5-639 Draft - Do Not Cite or Quote
-------
1 controls (Sharma et al., 2010a). Upadhyay et al. (2009) reported that treatment of
2 Sprague-Dawley rats with Pb acetate (35 mg/kg via i.p. injection for 3 days, blood Pb not
3 reported) significantly increased the activities of ALT, AST, serum ALP, and acid
4 phosphatase over those in controls but decreased liver ALP activity. Concomitant
5 treatment with Zn and varying levels of vitamin C were observed to ameliorate the toxic
6 effects of Pb. The serum activities of glutamic pyruvic transaminase (GPT) and lactate
7 dehydrogenase (LDH) were similarly significantly increased over those in controls in
8 mice subcutaneously injected with Pb acetate (50 mg/kg daily for 15 days, blood Pb not
9 reported) (Wang etal. 2010g). Swarup et al. (2007) investigated serum biochemical
10 changes in cows living in Pb-contaminated environments. Serum levels of ALT, AST,
11 ALP, total protein, albumin, globulin, and A/G ratio were significantly altered in cows
12 living near Pb-Zn smelters (mean [SD] blood Pb level: 86 [6] (ig/dL) compared to control
13 cows (mean [SD] blood Pb level: 7 [1] (ig/dL). Significant positive correlations were
14 found between blood Pb level and ALT and AST, whereas a negative correlation was
15 observed between blood Pb level and total lipids, protein, and albumin.
16 Pillai et al. (2009) investigated hepatic Phase I and II enzymes in male and female rats
17 born to dams that were treated with Pb acetate (50 (ig/kg, via s.c. injection daily
18 throughout gestation and continuing until PND21). Thus, the offspring of treated dams
19 were exposed to Pb via placental and lactational transfer. The female and male pups were
20 then allowed to reach sexual maturity (PND55-PND56) to assess continuing exposure to
21 bioaccumulated Pb. The activities of hepatic Phase I enzymes NADPH- and NADH-
22 cytochrome c reductase were significantly reduced in Pb-exposed male and female rats
23 on PND56 (blood Pb not reported), compared to controls. In rats treated with 25 (ig/kg Pb
24 and Cd, the effect on Phase I enzymes was increased. Pb treatment additionally decreased
25 the activities of Phase II enzymes uridine diphosphate-glucoronyl transferase and GST in
26 males and females, but no effect was observed on GGT or 17|3-hydroxysteroid
27 oxidoreductase. Additionally, no effect was observed on serum glutamate pyruvate
28 dehydrogenase or ALP activities in Pb-treated males or females. Histological
29 observations demonstrated fatty degeneration of the liver, vacuolization, and pycnotic
30 nuclei, indicating general hepatotoxicity following Pb treatment in both male and female
31 rats.
32 In a similar study, Teijon et al. (2006) exposed Wistar rats to Pb acetate (200 or 400 ppm
33 drinking water) throughout gestation, lactation, and 3 months postweaning, or only
34 1 month postweaning. In the animals exposed continuously throughout gestation and
35 lactation, the concentrations of Pb in the liver were elevated in the 200- and 400-ppm
36 groups 1 and 3 months postweaning. Liver concentrations of Pb were greater in the
37 200 ppm animals compared to the 400 ppm animals at one month postweaning (mean
38 [SE]: 1.19 [0.30] (ig Pb/g tissue versus 0.76 [0.06] ng Pb/g tissue, respectively), but were
November 2012 5-640 Draft - Do Not Cite or Quote
-------
1 similar between the 2 dosing regimens (200 ppm versus 400 ppm) at 3 months
2 postweaning (mean [SE]: 0.54 [0.06] versus 0.55 [0.07] \ig Pb/g tissue, respectively).
3 ALP activity was increased at 2 weeks postweaning in animals continuously exposed to
4 Pb throughout gestation and lactation, whereas ALT activity was decreased only at 2 and
5 3 months postweaning. In animals exposed for 1 month postweaning alone, only serum
6 ALP activity was significantly increased, although not in a concentration-dependent
7 manner. ALT and AST activities did not show significant changes.
8 Cheng et al. (2006) studied the mechanism of Pb effects on bacterial lipopolysaccharide
9 (LPS)-induced TNF-a expression. A/J mice were injected with Pb acetate (100 (imol/kg
10 via i.p.), with or without LPS (5 mg/kg). Pb alone did not affect AST or ALT activity or
11 the level of TNF-a in the serum of the mice. In comparison, treatment of mice with low
12 doses of Pb and LPS together caused a statistically significant increase in TNF-a
13 induction as well as enhanced liver injury, suggesting that Pb potentiated LPS-induced
14 inflammation. In a complementary in vitro experiment, co-exposure of Pb and LPS
15 stimulated the phosphorylation of p42/44 mitogen-activated protein kinase (MAPK) and
16 increased TNF-a expression in mouse whole blood cells, peritoneal macrophages, and
17 RAW264.7 cells (a macrophage cell line). These results indicated that Pb increased LPS-
18 induced TNF-a levels via the protein kinase C (PKC)/MAPK pathway in
19 monocytes/macrophages rather than hepatocytes. Similarly, Pb chloride potentiated
20 bovine serum albumin (BSA)-induced inflammation in the livers of mice subcutaneously
21 injected with Pb (Saet al.. 2012).
Lipid Metabolism
22 Several recent toxicological studies indicated Pb-induced impaired lipid metabolism, as
23 evidenced by increases in liver cholesterol. There was some evidence in animals exposed
24 to Pb in diet, albeit at relatively high exposure concentrations or measured blood Pb.
25 Ademuyiwa et al. (2009) reported that male albino Sprague Dawley rats exposed to 200,
26 300 and 400 ppm Pb acetate in drinking water had mean (SD) blood Pb levels of 40.63
27 (9.21), 61.44 (4.63), and 39.00 (7.90) (ig/dL, respectively. Animals exposed to 200 ppm
28 Pb had mean (SD) liver Pb concentrations of 10.04 (1.14) (ig/g, compared to 3.24
29 (1-19) (ig/g and 2.41 (0.31) (ig/g in animals exposed to 300 or 400 ppm Pb, respectively.
30 Animals exposed to Pb exhibited increased hepatic cholesterogenesis at all doses tested
31 compared to controls. Additionally, a decrease in triglyceride levels was observed at 300
32 and 400 ppm Pb; a decrease in phospholipid levels was observed at 400 ppm Pb. The
33 authors also reported positive correlations between tissue cholesterol and phospholipids
34 and Pb accumulation in liver across all doses. In contrast, the association between tissue
35 triglyceride levels and Pb accumulation was negative. In related studies, Khotimchenko
36 and Kolenchenko (2007) reported that adult male albino rats treated with Pb acetate
November 2012 5-641 Draft - Do Not Cite or Quote
-------
1 (100 mg/kg for 14 days, blood Pb not reported) exhibited disorders in lipid metabolism
2 that were accompanied by increased levels of total cholesterol and triglycerides in the
3 liver tissue. Sharma et al. (2010a) reported increased liver cholesterol in mice gavaged
4 with Pb nitrate, 50 mg/kg for 40 days. Pillai et al. (2009) observed decreases in total liver
5 cholesterol in PND56 male and female rats that had been treated with Pb acetate (via s.c.
6 injection, 50 (ig/kg, continuously throughout gestation and lactation). These results
7 suggest that Pb induction of cholesterogenesis and phospholipidosis in the liver may
8 cause subtle effects at the cellular level that may lead to hepatotoxicity.
9 Kojima and Degawa (2006) examined sex-related differences in Pb-induced gene
10 expression of a rate limiting hepatic cholesterol biosynthesis enzyme, 3-hydroxy-3-
11 methylglutaryl-CoA reductase (HMGR) and its transcription factor, sterol regulatory
12 element binding protein-2 (SREBP-2). Male and female Sprague Dawley rats were
13 injected with Pb nitrate (100 (imol/kg body weight, intravenously, blood Pb not reported).
14 SREBP-2 expression was significantly increased in males and females with the increase
15 occurring earlier in male rats (6-12 hours, compared to 24-36 hours in females). In
16 contrast, expression of HMGR was significantly increased in both Pb-exposed males and
17 females at earlier time frames and greater range of onset (3-48 hours in males; 12-48
18 hours in females) compared to that of SREBP-2. Significant increases in total liver
19 cholesterol were also observed in Pb-exposed males and females at 3-48 and 24-48 hours,
20 respectively. These results suggest that the SREBP-2 and HMGR gene expressions and
21 increase in total cholesterol levels in the liver in response to Pb exposure occur earlier in
22 males compared to females and also suggest that the HMGR gene expression and
23 increase in total cholesterol levels in the liver occur before an increase in the SREBP-2
24 gene expression in each sex.
Hepatic Oxidative Stress
25 A number of studies demonstrated increased hepatic oxidative stress as a result of
26 exposure to various Pb compounds, demonstrated by increases in reactive oxygen species
27 (ROS) or decreases in antioxidant levels or enzyme activity. ROS can potentially result in
28 damage to hepatic function and structure. Several of these observations were made in
29 animals exposed to Pb in drinking water that produced blood Pb levels relevant to
30 humans. In a study examining the effects of Pb exposure to fetuses, Masso et al. (2007)
31 exposed pregnant Wistar rats to 300 ppm Pb in drinking water from GD1 to parturition,
32 or to weaning. Blood Pb levels were higher at parturition (mean [SD]: 31.5 [0.80] (ig/dL)
33 than at weaning (mean [SD]: 22.8 [0.50] (ig/dL). Pups exhibited liver damage that was
34 accompanied by an increased production of thiobarbituric acid-reactive species (TEARS,
35 an indicator of lipid peroxidation) and increased CAT activity compared to controls. In
36 addition, increased ALP and acid phosphatase activity was observed. Uzbekov et al.
November 2012 5-642 Draft - Do Not Cite or Quote
-------
1 (2007) showed differential effects by duration of maternal exposure before mating.
2 Female Wistar rats were exposed to 0.3 and 3.0 ppm Pb nitrate in drinking water, for
3 1 month or 5 months prior to pregnancy, and also continuing during pregnancy; then the
4 livers from both groups of GD20 fetuses were examined for hepatic SOD activity. The
5 pregnant control rats had a mean (SD) blood Pb level of 16.1 (0.63) (ig/dL, whereas the
6 dams exposed to 0.3 and 3.0 ppm Pb had mean blood Pb levels of 20.4 (ig/dL and
7 24.4 (ig/dL, respectively. In the GD20 fetuses from dams exposed for 1 month prior to
8 pregnancy, a concentration-dependent increase in liver SOD activity was observed,
9 whereas SOD activity was decreased in the GD20 fetuses from dams exposed for
10 5 months prior to pregnancy. The increase in SOD activity in the livers of fetuses from
11 dams exposed to 0.3 or 3.0 ppm Pb nitrate for one month suggests an initial activation of
12 SOD in response to increased free radical production, while the decrease in SOD
13 production in fetal livers from dams exposed to the same concentrations for 5 months
14 suggests that longer durations of Pb exposure impairs the antioxidant defense mechanism.
15 Increased oxidative stress also was found in animals with postnatal Pb exposure in
16 drinking water. Jurczuk et al. (2007) reported that male Wistar rats treated with 500 ppm
17 Pb in drinking water (blood Pb not reported) exhibited decreases in liver vitamin E and
18 GSH levels along with an increase in lipid peroxidation. The correlation between vitamin
19 E and lipid peroxidation suggested that vitamin E is involved in the mechanism of
20 peroxidative action of Pb in the liver. In a study examining the role of low molecular
21 weight thiols on peroxidative mechanisms, Jurczuk et al. (2006) found that male Wistar
22 rats treated with 500 ppm Pb acetate in drinking water exhibited a decrease in blood
23 ALAD as well as decreases in GSH and nonprotein sulfhydryl levels in the liver.
24 Metallothionein levels were also reported to be higher in the liver following exposure to
25 Pb. Yu et al. (2008) reported concentration-dependent increases in lipid peroxide levels
26 and decreases in GSH levels and CAT, SOD and GPx activities in livers from castrated
27 male pigs that received a diet mixed with 0, 5, 10, or 20 mg/kg Pb nitrate exposure,
28 during ages 55-100 days. The level of hepatic CuZnSOD mRNA was also reduced in
29 Pb-treated animals. The study authors suggested that this decrease in SOD mRNA
30 expression and activity of antioxidant enzymes may lead to a reduction in free radical
31 scavenging capability and increased lipid peroxidation.
32 Studies administering Pb by bolus doses had similar findings. Adegbesan and Adenuga
33 (2007) reported that lipid peroxidation was increased and SOD activity was decreased in
34 protein undernourished male Wistar rats compared to well-fed rats, and that these effects
35 were further exacerbated in protein undernourished rats injected with Pb nitrate
36 (100 (imol/kg, blood Pb not reported). Protein undernourishment also decreased GSH
37 levels and CAT activity compared to normal diet; however, co-treatment with Pb
38 mitigated the severity of these effects. GSH levels and CAT activity were still lower in
November 2012 5-643 Draft - Do Not Cite or Quote
-------
1 undernourished rats with Pb exposure compared to well-fed rats, but greater than
2 undernourished rats with no Pb exposure. The results indicated Pb treatment exacerbated
3 the effects of malnutrition on liver lipid peroxidation and altered the involvement of free
4 radicals. Male Charles-Foster rats treated with Pb acetate (0.025 mg/kg via i.p. injection,
5 blood Pb not reported) also exhibited statistically significant increases in lipid
6 peroxidation levels and decreases in SOD, CAT, and glucose-6-phosphatase
7 dehydrogenase levels in liver mitochondrial and postmitochondrial fractions (Pandya et
8 al.. 2010). Statistically nonsignificant decreases were observed in GSH levels, and in GPx
9 and GR activities in Pb-treated animals. In mice gavaged with Pb nitrate (50 mg/kg for
10 40 days), lipid peroxidation was increased, and SOD, CAT, and GSH were decreased
11 compared to controls (Sharma et al.. 2010a). Additionally, Pb nitrate treatment resulted in
12 histopathological changes in the structure of the liver: hepatocytes were damaged and
13 were marked by cytoplasmic vacuolization and pycnotic nuclei.
14 Khotimchenko and Kolinchenko (2007) also reported an increase in lipid peroxidation
15 and development of hepatitis in male albino rat liver parenchyma following intragastric
16 treatment with Pb acetate (100 mg/kg for 14 days). Lipid peroxidation was demonstrated
17 by increases in malondialdehyde (MDA) levels along with decreases in GSH and thiol
18 groups; indicating injury in the liver antioxidant system. Levels of hepatic lipid
19 peroxidation were observed to be significantly increased in rats treated with Pb acetate
20 (35 mg/kg via i.p. injection daily for 3 days, blood Pb not reported), whereas hepatic
21 GSH was significantly decreased (Upadhyay et al.. 2009). A study examining male and
22 female rat pups that were continuously exposed to Pb during gestation and lactation
23 (pregnant dams were injected [s.c.] with Pb acetate, 50 (ig/kg per day [GDO to PND21],
24 blood Pb not reported), did not find effects on GSH or MDA levels at PND56 (Filial et
25 al., 2009). In vitro exposure of cells from a hepatic human embryonic epithelial cell line
26 (WRL-68) to 5 (iM Pb acetate for 30 days resulted in increased production of ROS
27 throughout the incubation period (Hernandez-Franco et al., 2011). Concurrent with this
28 increase in ROS generation, the activities of SOD and the levels of membrane lipid
29 peroxidative damage increased throughout the first 24 days of exposure but returned to
30 normal levels by day 30.
Hepatic Apoptosis
31 Fan et al. (2009b) reported that a single i.v. injection (tail vein) of Pb nitrate
32 (200 (imol/kg in 0.5 mL) in rats resulted in an increase in the percentage of apoptotic
33 hepatocytes (mean: 2.5 [SD: 1.4]% of total hepatocytes) compared with controls (mean:
34 0.31 [SD: 0.31]%). Expression of ferritin light-chain (FTL) protein also increased (mean
35 [SD]: 3.5 [1.0]-fold increase) over that in controls. Immunohistochemical analysis
36 revealed that hepatocytes around the central vein were heavily stained by anti-FTL
November 2012 5-644 Draft - Do Not Cite or Quote
-------
1 antibodies, as were nonparenchymal cells identified as Kupffer cells. The authors
2 hypothesized that the expression of FTL in Kupffer cells may have resulted from the
3 phagocytosis of apoptotic cells. Treatment of rats with clofibrate, a lipid lowering agent,
4 did not increase FTL expression in Kupffer cells and induced hepatocellular proliferation
5 butnotapoptosis.
5.9.1.4 Summary of Effects on the Hepatic System
6 While explicit associations between hepatic injury (i.e., histopathological effects) and Pb
7 exposure have not established, evidence from epidemiologic and toxicological studies
8 have indicated that exposure to Pb can result in altered liver function and hepatic
9 oxidative stress. A few studies have reported associations of higher blood Pb levels with
10 decreased cytochrome P450 enzymes (Phase I xenobiotic metabolism) in children and
11 nonoccupationally-exposed adults. However, most evidence indicates decreases in serum
12 protein and albumin levels and increased AST, ALT, ALP, and GGT activities (indicators
13 of decreased liver function), increased oxidative stress, and decreased antioxidant status
14 in Pb-exposed workers with blood Pb levels >29 (ig/dL (Can et al., 2008; Khan et al.,
15 2008; Patil et al., 2007; Conterato et al.. In Press). The implications of the epidemiologic
16 evidence is limited because of its cross-sectional study design nature, the high blood Pb
17 levels examined, and lack of consideration for potential confounding by factors such as
18 age, diet, BMI, smoking, or other occupational exposures.
19 Similar changes in liver function enzymes have been found in mature animals exposed to
20 high levels of Pb during adulthood (Sharma et al., 2010a; Wang et al., 2010; Herman et
21 al.. 2009; Cheng et al.. 2006). and animals exposed during gestation and lactation (Pillai
22 et al.. 2009; Teijon et al., 2006). Pb exposure has been shown to impair lipid metabolism
23 in animals, as evidenced by increased hepatic cholesterogenesis, and in altered
24 triglyceride and phospholipid levels (Ademuyiwa et al., 2009; Khotimchenko and
25 Kolenchenko. 2007). Multiple studies in humans and animals have observed
26 Pb-associated hepatic oxidative stress, generally indicated by an increase in lipid
27 peroxidation along with a decrease in GSH levels and CAT, SOD, and GPx activities
28 (Pandvaetal. 2010; Sharma et al.. 2010a; Khan et al.. 2008; Yu et al.. 2008; Adegbesan
29 and Adenuga. 2007; Jurczuk et al.. 2007; Khotimchenko and Kolenchenko. 2007;
30 Jurczuk et al.. 2006). Indices of increased oxidative stress were also observed in the livers
31 of fetuses exposed to Pb throughout gestation (Masso et al.. 2007). The relevance of the
32 toxicological evidence is uncertain as many studies administered Pb as bolus doses.
33 Because of the insufficient quality of studies, the evidence is inadequate to determine if
34 there is a causal relationship between Pb exposure and hepatic effects.
November 2012 5-645 Draft - Do Not Cite or Quote
-------
5.9.2 Effects on the Gastrointestinal System
1 Gastrointestinal effects examined in relation to Pb exposure include abdominal pain,
2 constipation, and internal paralysis in humans and degeneration of the intestinal epithelial
3 mucosa and a decrease in duodenal motility in animals.
5.9.2.1 Summary of Key Findings on the Effects on the
Gastrointestinal System (2006 Pb AQCD)
4 The 2006 Pb AQCD (U.S. EPA. 2006b) stated that a number of factors influence the
5 gastrointestinal absorption of Pb; including the chemical and physical form of Pb, the age
6 at Pb intake, as well as various nutritional factors. Rats exposed to Pb acetate in drinking
7 water had degeneration of the intestinal epithelial mucosa, potentially leading to
8 malabsorption of nutrients. In suckling rat pups, increased casein micelles incidences
9 were reported as a result of Pb present in bovine and rat milk and in infant milk formula.
10 Pb ingestion through water was more toxic compared to Pb ingestion via milk. Pb
11 ingested in milk was reported to be taken up by the ileal tissue, whereas Pb administered
12 intragastrically as a soluble salt was primarily accumulated in the duodenum irrespective
13 of vehicle used for administration. Decreases in duodenal motility and the amplitude of
14 contractility in the intestinal tract were observed in rats following Pb exposure.
15 Nutritional studies examining different dietary levels of Pb, Ca2+, and vitamin D in rats
16 indicated competition in absorption between Pb and calcium. Dietary supplementation
17 with vitamin D led to an increase in intestinal absorption of Pb and calcium. In instances
18 where severe calcium deficiency was noted, ingestion of Pb caused a clear decrease in
19 1,25-dihydroxy vitamin D (1,25-(OH)2D3) levels. Overall, the 2006 Pb AQCD stated
20 that studies in rat intestine have shown that the largest amount of Pb absorption occurs in
21 the duodenum with the mechanisms of absorption involving active transport and
22 diffusion via the intestinal epithelial cells. Absorption has been reported to occur, through
23 both saturable and nonsaturable pathways, based on results from various animal studies.
24 The 2006 Pb AQCD reported evidence that symptoms associated with gastrointestinal
25 colic (abdominal pain, constipation, intestinal paralysis) were more prevalent in
26 occupationally-exposed adults with blood Pb levels > 50 (ig/dL.
5.9.2.2 Recent Epidemiologic Studies
27 Consistent with previous findings, Kuruvilla et al. (2006) reported gastrointestinal effects
28 including stomach pain and gastritis along with other non-gastrointestinal effects in 53
29 Pb-exposed painters (mean [SD] blood Pb: 8.04 [5.04] (ig/dL) in India compared with 50
November 2012 5-646 Draft - Do Not Cite or Quote
-------
1 controls (mean [SD] blood Pb level: 5.76 [4.45] (ig/dL) matched by sex, age, education,
2 income, smoking, and alcohol consumption. Prevalence of symptoms in painters did not
3 differ from that in battery workers with higher blood Pb levels (mean [SD] blood Pb level
4 of 42.40 ([25.53] (ig/dL). Despite the consistent evidence among occupational studies,
5 the implications of gastrointestinal findings in Pb-exposed workers are limited by the
6 cross-sectional study designs, high blood Pb levels associated with effects (mostly
7 > 50 (ig/dL), and limited consideration for potential confounding by factors such as age,
8 smoking, alcohol use, nutrition, or other occupational exposures.
5.9.2.3 Recent lexicological Studies
9 A few recent studies pertaining to gastrointestinal effects of Pb exposure were identified
10 that provide evidence for additional mechanisms underlying gastrointestinal damage and
11 impaired function. Santos et al. (2006) examined the impact of Pb exposure on
12 nonadrenergic noncholinergic (NANC) relaxations in rat gastric fundus. Male Wistar rats
13 treated with 80 ppm Pb acetate via drinking water for 15, 30, and 120 days (blood Pb not
14 reported) exhibited a significant difference in NANC relaxations in the gastric fundus
15 following electrical field stimulus. While frequency-dependent relaxations were observed
16 in all groups, including the control group, the relaxations were significantly inhibited in
17 rats treated with Pb acetate for all three durations. When gastric fundus strips from rats
18 were incubated with L-nitroarginine, a nitric oxide (NO) synthase inhibiter, no additional
19 inhibition in relaxations was observed. In contrast, incubation with sodium nitroprusside
20 and 8-Br-GMPc (a cyclic guanosine monophosphate [cGMP] analog), resulted in a
21 concentration-dependent relaxation in strips in the control group and in the group
22 exposed to Pb acetate for 120 days. The results suggested that long-term exposure to Pb
23 causes inhibition in NANC relaxation probably due to the modulated release of NO from
24 the NANC nerves or due to interaction with the intracellular transducer mechanism in the
25 rat gastric fundus.
26 In another study examining Pb-induced oxidative stress in the gastric mucosa, Olaleye et
27 al. (2007) treated Albino Wistar rats with 100 or 5,000 ppm Pb acetate in drinking water
28 for 15 weeks (blood Pb not reported). Exposure to Pb acetate caused a significant
29 increase in gastric mucosal damage caused by pretreatment with acidified ethanol. While
30 the basal gastric acid secretory rate was not altered, stomach response to histamine was
31 significantly higher in animals treated with Pb acetate compared to that in the controls.
32 Additionally, there was a significant increase in gastric lipid peroxidation at both the 100
33 and 5,000 ppm dose levels. In contrast, CAT, and SOD activities and nitrite levels were
34 significantly decreased in the gastric mucosa. The results indicated that Pb-induced
35 gastric damage may be mediated via increases in oxidative stress.
November 2012 5-647 Draft - Do Not Cite or Quote
-------
5.9.2.4 Summary of Gastrointestinal Effects
1 Relatively few human studies have been conducted on the gastrointestinal toxicity of Pb.
2 The evidence points to more prevalent symptoms, such as stomach pain, gastritis,
3 constipation, and intestinal paralysis, in Pb-exposed workers (Kuruvilla et al.. 2006).
4 However, the implications of gastrointestinal findings in Pb-exposed workers are limited
5 by the cross-sectional study designs, high blood Pb levels associated with effects (mostly
6 > 40 (ig/dL), and limited consideration of potential confounding by factors such as age,
7 smoking, alcohol use, nutrition, or other occupational exposures. Toxicological evidence
8 indicates that Pb is absorbed primarily in the duodenum by active transport and diffusion,
9 although variability is observed by Pb compound, age of intake, and nutritional factors.
10 There is some coherence between the evidence in Pb-exposed workers and observations
11 in animals that Pb induces damage to the intestinal mucosal epithelium, decreases
12 duodenum contractility and motility, reduces absorption of Ca2+, inhibits NANC
13 relaxations in the gastric fundus, and induces oxidative stress (lipid peroxidation,
14 decreased SOD and CAT) in the gastric mucosa (Olaleve et al., 2007; Santos et al.,
15 2006). The observation of oxidative stress was accompanied by gastric mucosal damage.
16 Because of the insufficient quantity and quality of studies, the evidence is inadequate to
17 determine if there is a causal relationship between Pb exposure and gastrointestinal
18 effects.
5.9.3 Effects on the Endocrine System
19 A summary of key findings pertaining to reproductive hormones in males and females is
20 presented in the section on Reproductive and Developmental Effects (Sections 5.8.1 and
21 5.8.2). Collective epidemiologic and toxicological evidence is inconsistent in
22 demonstrating the effects of Pb exposure on male and female sex hormone levels. Other
23 endocrine processes that are most commonly found to be impacted by Pb exposure
24 include changes in thyroid hormones, including thyroid stimulating hormone (TSH),
25 triiodothyronine (T3), and thyroxine (T4). A few studies have examined calcium and
26 cortisol.
5.9.3.1 Summary of Key Findings of the Effects on the Endocrine
System (2006 Pb AQCD)
27 The 2006 Pb AQCD (U.S. EPA. 2006b) reported that endocrine processes impacted by
28 occupational Pb exposure include thyroid hormone levels, changes in male sex hormone
29 levels, as well as changes in the production of vitamin D (1,25-(OH)2D3). However,
November 2012 5-648 Draft - Do Not Cite or Quote
-------
1 these effects were observed only with blood Pb levels exceeding 30-40 (ig/dL and in
2 studies with little or no consideration for potential confounding by factors such as sex,
3 SES, nutritional status, BMI, smoking, comorbid conditions, and other occupational
4 exposures. In addition, alterations in calcitropic hormones were found in children with
5 blood Pb levels ranging from 10-120 (ig/dL and in an opposite direction than that in
6 Pb-exposed workers.
5.9.3.2 Recent Epidemiologic Studies
7 Recent epidemiologic studies have reported associations between indicators of exposure
8 to Pb and thyroid hormone levels, in populations of children and in adults with and
9 without occupational Pb exposure; although results have not been consistent for a
10 particular hormone. Further, the implications of these findings are limited because of the
11 use of cross-sectional study design, lack of rigorous statistical analysis, and limited
12 consideration for potential confounding factors. Inconsistent associations were found in a
13 study that considered potential confounding factors. Abdelouahab et al. (2008) examined
14 a Canadian population characterized by high consumption of freshwater fish
15 contaminated with Pb and other environmental pollutants. The median concurrent blood
16 Pb level was 3.1 (ig/dL for men and 1.7 (ig/dL for women. The median blood Pb level for
17 women was lower than the limit of detection (2.1 (ig/dL), resulting in measurement error
18 of blood Pb level and greater uncertainty in the results. In an analysis stratified by sex,
19 TSH levels were negatively correlated with blood Pb in women with adjustment for age,
20 smoking status, estro-progestative intake, total plasma lipids, and Se. No associations
21 with T3 and T4 levels were found in women. TSH, T3 and T4 levels were not correlated
22 with blood Pb level in males, after adjustment for the same covariates (excluding
23 hormone intake) plus pesticide exposure, corticoid medication, concurrent alcohol
24 consumption, and occupational exposure to metals. Overall, the inconsistent associations
25 and potential influence of other exposures did not strongly demonstrate an effect of Pb
26 exposure.
27 Studies with less rigorous methods also did not clearly indicate an association between
28 blood Pb level and a particular thyroid hormone. In a Kosovo, Yugoslavia population,
29 higher pregnancy blood Pb levels were associated with lower pregnancy free T4 level
30 among the 156 women living in a highly exposed town with a smelter and battery plant,
31 but not among the 153 women living in a relatively unexposed nearby town (Lamb et al..
32 2008). The mid-pregnancy blood Pb levels were highly elevated in the industrial town
33 compared to the unexposed town (mean [SD]: 20.56 [7.38] versus 5.60 [1.99] (ig/dL). In
34 24 newborns delivered in Tokyo, Japan, neither TSH nor free T4 (sampled 4-6 days
35 postpartum) was correlated with cord blood Pb level (mean: 0.67 (ig/dL) (lijima et al..
November 2012 5-649 Draft - Do Not Cite or Quote
-------
1 2007). Neither of these studies considered potential confounding. Croes et al. (2009)
2 examined the hormone levels in 1,679 adolescents residing in nine study areas in
3 Belgium with varying exposures to multiple industrial pollutants including Pb. The
4 median concurrent blood Pb level of the participants from the nine different regions
5 ranged from 1.6 to 2.8 (ig/dL. Analyses only indicated differences in free T3, at the
6 region or neighborhood level with adjustment for age, sex, recent disease, and BMI. No
7 direct associations with blood Pb level were analyzed, thus the results could be attributed
8 to other factors that varied by location.
9 Contrasting results were found for free T4 in Pb-exposed workers. Dundar et al. (2006)
10 examined associations between blood Pb levels and thyroid function in 42 male
11 adolescent auto repair apprentice workers, with no history of prior disease, exposed long
12 term to Pb (in the auto repair apprenticeship at least 1 year). Mean blood Pb level was
13 higher in the auto repair workers compared to the 55 healthy unexposed control subjects
14 (mean [SD]: 7.3 [2.92] versus 2.08 [1.24] (ig/dL). Free T4 levels were significantly lower
15 in the auto workers compared to the control group, which had no abnormal free T4 levels
16 reported. In contrast, free T3 and TSH levels were comparable between auto workers and
17 controls. Blood Pb level was negatively correlated with free T4 levels. In contrast,
18 another study (Pekcici et al.. 2010) found higher free T4 and TSH in adult auto mechanic
19 or battery factory workers who were highly exposed to Pb (mean blood Pb: 71.1 (ig/dL)
20 compared to controls (mean blood Pb level: 0.2 (ig/dL). Free T3 levels were similar
21 between the two groups. The results from this study are likely not generalizable to the
22 general public due to the high blood Pb levels of the exposed workers.
23 Previous findings for blood Pb-associated changes in serum vitamin D (1,25-(OH)2D3)
24 in children were mixed. A recent study in New Jersey examined winter (December to
25 March) and summer (July to September) seasonal changes in the associations between
26 blood Pb level and serum 1,25-(OH)2D3 status, in 142 young, U.S. urban African-
27 American or Hispanic children (ages 1-8 years, grouped by age [1-3 year-olds and 4-8
28 year-olds] and race/ethnicity) using a repeated measures design (Kemp et al.. 2007). The
29 percentage of 1 -3 year-old African-American children (n = 49) with blood Pb levels
30 > 10 (ig/dL increased from 12.2% in winter to 22.5% in summer. This large seasonal
31 increase in blood Pb levels in these 1-3 year-old children was not accompanied by a
32 significant increase in serum 1,25-(OH)2D3 concentrations. There was also a larger
33 seasonal increase in blood Pb levels in 1-3 year-old children from both races combined
34 (n = 78) (mean [SE]: 4.94 [0.45] (ig/dL winter, 6.54 [0.82] (ig/dL summer) than in 4-8
35 year-old children from both races combined (n = 64) (mean [SE]: 3.68 [0.31] (ig/dL
36 winter, 4.16 [0.36] (ig/dL summer). However, no difference in seasonal 1,25-(OH)2D3
37 was observed in the 1-3 year-old children from both races combined. A larger winter to
38 summer increase in blood Pb level was correlated with a larger seasonal increase in
November 2012 5-650 Draft - Do Not Cite or Quote
-------
1 serum 1,25-(OH)2D3 in the 4-8 year-old children from both races combined and in the
2 4-8 year-old African American children (n = 42). In the 4-8 year-old children from both
3 races combined, there was a winter to summer increase in 1,25-(OH)2D3 (mean [SE]:
4 25.3 [1.2] (ig/L in winter versus 33.8 [1.1] (ig/L in summer.), which may account forthe
5 results in this older age group. Based on these results, the study authors concluded that
6 higher summertime increase in serum 1,25-(OH)2D3 levels in children between 4 and 8
7 years is most likely due to increased sunlight-induced vitamin D synthesis and may be a
8 contributing factor to seasonal changes in blood Pb levels via changes in gastrointestinal
9 absorption or release of Pb from bone.
10 HPA function was examined in a prospective analysis of associations between prenatal
11 maternal blood Pb levels (cord blood collected at delivery) or postnatal blood Pb levels
12 (at mean [SD] age: 2.62 [1.2] years, data obtained from family physicians or state
13 records), and saliva cortisol levels during a stress protocol in the children at age 9.5 years
14 (Gump et al.. 2008). For prenatal blood Pb, the children were divided into the following
15 quartiles: < 1, 1.1-1.4, 1.5-1.9, and 2.0-6.3 (ig/dL. For postnatal blood Pb, the quartiles
16 were: 1.5-2.8, 2.9-4.1, 4.2-5.4, and 5.5-13.1 (ig/dL. With adjustment for potential
17 confounding (by SES-related factors, HOME score, pregnancy health, maternal substance
18 abuse), blood Pb level was not associated with initial salivary cortisol levels. However,
19 following an acute stressor, which comprised submerging the dominant arm for a minute
20 in a gallon of one part ice to one part water, increasing prenatal and postnatal blood Pb
21 levels were associated with statistically significant increases in salivary cortisol
22 responses. Children in the 2nd, 3rd, and 4th prenatal blood Pb quartiles and in the 4th
23 postnatal quartile had increased salivary cortisol responses compared to children in the
24 1st quartile. When blood Pb was treated as a continuous variable, regression analysis
25 showed that both prenatal and postnatal blood Pb levels were associated salivary cortisol
26 reactivity. While associations were found in children with blood Pb levels below
27 10 (ig/dL, they could have been attributed to higher earlier childhood blood Pb levels of
28 these children who were born in 1980-1990s.
5.9.3.3 Recent lexicological Studies
29 Pb-associated changes in thyroid hormones also were found in animal studies. In a study
30 examining the effects of Pb and Cd in adult cows reared in a polluted environment in
31 India, Swarup et al. (2007) found significantly higher mean plasma T3 and T4 levels in
32 cows living near Pb/Zn smelters (mean [SD] blood Pb: 86 [6] (ig/dL) and near closed
33 Pb/operational Zn smelters (mean [SD] blood Pb: 51 [9] (ig/dL) when compared to cows
34 in unpolluted areas (mean [SD] blood Pb: 7 [1] (ig/dL). Regression analyses of the 269
35 cows showed a significant positive correlation between blood Pb levels and plasma T3
November 2012 5-651 Draft - Do Not Cite or Quote
-------
1 and T4 levels, whereas the correlation between blood Pb levels and plasma cortisol was
2 not statistically significant. Mean plasma estradiol level was significantly higher in cows
3 near closed Pb/operational Zn smelters compared to the control group of cows. Because
4 of the Pb-Zn co-exposure, the effects cannot be attributed specifically to Pb.
5 Biswas and Ghosh (2006) investigated the effect of Pb treatment on adrenal and male
6 gonadal functions in Wistar rats treated with Pb acetate (8.0 mg/kg via i.p. injection for
7 21 days, blood Pb not reported). Pb treatment significantly increased adrenal
8 steroidogenic enzyme activity and serum corticosterone levels. Accessory sex organ
9 (prostate and seminal vesicle) weights were decreased in Pb-treated animals, whereas
10 adrenal weights were increased. These effects were accompanied by a decrease in
11 spermatogenesis and serum concentrations of testosterone, FSH, and LH and by an
12 increase in the percent of spermatid degeneration. Supplementation with testosterone
13 during the last 14 days of Pb treatment was observed to ameliorate these effects.
5.9.3.4 Summary of Endocrine Effects
14 Collective epidemiologic and toxicological evidence is inconsistent in demonstrating the
15 effects of Pb exposure on male and female sex hormone levels (Sections 5.8.1 and 5.8.2)
16 and vitamin D levels. Several epidemiologic studies have reported associations between
17 indicators of Pb exposure and thyroid hormone levels in populations of children and
18 adults without (Lamb et al.. 2008) and with occupational Pb exposure (Dundar et al.
19 2006). although results have not been consistent for a particular hormone. Further, the
20 implications of these findings are limited because of the cross-sectional study design,
21 high blood Pb levels associated with effects (>30 (ig/dL), lack of rigorous statistical
22 analysis, and limited consideration for potential confounding factors. Blood Pb level was
23 positively correlated with plasma T3 and T4 levels in adult cows living near Pb-Zn
24 smelters; however, the effects could not be attributed specifically to Pb exposure (Swarup
25 et al.. 2007).
26 In a prospective study of children in New York, who were challenged with an acute
27 stressor, higher cord blood levels (as a reflection of prenatal maternal Pb blood level), or
28 2-year-old blood Pb levels, were associated with significant higher salivary cortisol in
29 response to a stress challenge at age 9 years (Gump et al.. 2008). While these associations
30 were found with blood Pb levels <10 (ig/dL, they could have been attributed to higher
31 earlier childhood blood Pb levels of these children who were born in the 1980s and
32 1990s. Biswas and Ghosh (2006) found a Pb-induced increase in corticosterone in rats,
33 albeit by i.p. Pb treatment. Cortisol and corticosterone are the major glucocorticoids in
34 humans and rodent, respectively.
November 2012 5-652 Draft - Do Not Cite or Quote
-------
1 In conclusion, epidemiologic and toxicological evidence indicates Pb-associated
2 endocrine effects such as thyroid hormones, cortisol, and vitamin D, although results are
3 not consistent. Because of the lack of insufficient quantity and quality of studies, the
4 evidence is inadequate to determine if there is a causal relationship between Pb exposure
5 and endocrine effects related to thyroid hormones, cortisol, and vitamin D.
5.9.4 Effects on Bone and Teeth
6 Primary effects on bone associated with Pb exposure or biomarker levels have included
7 an increase in osteoporosis, increased frequencies of falls and fractures, changes in bone
8 cell function as a result of replacement of bone calcium with Pb, and depression in early
9 bone growth. Other effects include tooth loss and periodontitis. Mechanistic evidence
10 from toxicological studies includes effects on cell proliferation, procollagen type I
11 production, intracellular protein, and osteocalcin in human dental pulp cell cultures.
5.9.4.1 Summary of Key Findings of the Effects on Bone and
Teeth (2006 Pb AQCD)
12 The 2006 Pb AQCD reported many effects on bone and some in teeth in animals
13 following Pb exposure. Exposure of animals to Pb during gestation and the immediate
14 postnatal period was reported to significantly depress early bone growth with the effects
15 showing concentration-dependent trends. In mature animals, long-term Pb exposure (up
16 to one year), along with poor nutrition (low calcium) reduced bone growth as well as
17 bone density. Systemic effects of Pb exposure included disruption in bone mineralization
18 during growth, alteration in bone cell differentiation and function due to alterations in
19 plasma levels of growth hormones and calcitropic hormones such as 1,25-[OH]2D3 and
20 impact on Ca2+- binding proteins and increases in Ca2+ and phosphorus concentrations in
21 the bloodstream. Bone cell cultures exposed to Pb had altered vitamin D-stimulated
22 production of osteocalcin accompanied by inhibited secretion of bone-related proteins
23 such as osteonectin and collagen. In addition, Pb exposure caused suppression in bone
24 cell proliferation most likely due to interference from factors such as growth hormone
25 (GH), epidermal growth factor (EGF), transforming growth factor-beta 1 (TGF-|31), and
26 parathyroid hormone-related protein (PTHrP).
27 As in bone, Pb exposure was found to easily substitute for Ca2+ in the teeth and was taken
28 up and incorporated into developing teeth in experimental animals. Since teeth do not
29 undergo remodeling like bone does during growth, most of the Pb in the teeth remains in
30 a state of permanent storage. High dose Pb exposure to animals (30 mg/kg body weight)
November 2012 5-653 Draft - Do Not Cite or Quote
-------
1 was found to induce the formation of a "Pb line" that is visible in both the enamel and
2 dentin and is localized in areas of recently formed tooth structure. Areas of mineralization
3 were easily evident in the enamel and the dentin within these "Pb lines." Pb has also been
4 shown to decrease cell proliferation, procollagen type I production, intracellular protein,
5 and osteocalcin in human dental pulp cell cultures. Adult rats exposed to Pb have
6 exhibited an inhibition of the posteruptive enamel proteinases, delayed teeth eruption
7 times, as well as a decrease in microhardness of surface enamel. Pb was reported to be
8 widely dispersed and incorporated into developing apatite crystal during enamel
9 formation process; however, post formation, Pb was reported to be capable of entering
10 and concentrating in specific enamel areas which were Ca2+-deficient. The
11 2006 Pb AQCD (U.S. EPA. 2006b) also reported that a number of animal studies and a
12 few epidemiologic studies each suggested that Pb is a caries-promoting element. The
13 strongest epidemiologic evidence comprised associations between concurrent blood Pb
14 level and dental caries in an NHANES analysis of thousands of children that adjusted for
15 age, sex, race/ethnicity, poverty to income ratio, exposure to cigarette smoke, geographic
16 region, head of household education, carbohydrate and calcium intake, and frequency of
17 dental visits. Other effects found in humans included bone disease (e.g., Paget's disease);
18 however, the evidence was provided by occupational or case-control studies.
5.9.4.2 Recent lexicological and Epidemiologic Studies
19 Consistent with evidence reported in the 2006 Pb AQCD, recent studies have found
20 associations between Pb exposure or biomarker levels and effects in bones of humans and
21 animals. The association between blood Pb levels and lower bone mineral density was
22 examined in several epidemiologic studies. Prospective evidence was provided by Khalil
23 et al. (2008) in 533 older women aged 65-87 years with a mean (SD) blood Pb of 5.3
24 (2.3) (ig/dL. Bone mineral density was measured in 1986-1988 (calcaneus), again in
25 1988-1990 (total hip and femoral neck), and again in 1993-1994 (calcaneus, total hip and
26 femoral neck), while blood Pb levels were measured in between the 2nd and last bone
27 analyses (during 1990-1991; and categorized as low [n = 122], medium [n = 332], and
28 high [n = 79] Pb blood levels [range: 1-21 ng/dL]). Information on falls and fractures was
29 collected every 4 months, starting after the initial enrollment (1986-1988) and continuing
30 for more than 10 years. The bone mineral density at the last measurement (1993-1994)
31 was 7% lower in the total hip (p <0.02) and 5% lower in the femoral neck (p <0.03) in the
32 high blood Pb group (> 8 (ig/dL) compared to the low blood Pb group (< 3 (ig/dL). A
33 concentration-dependent relationship was found for total hip and femoral neck bone
34 mineral density across the three blood Pb level groups. In addition, total hip, femoral
35 neck, and calcaneus bone loss was observed to be greater in the medium (blood Pb:
November 2012 5-654 Draft - Do Not Cite or Quote
-------
1 4-7 (ig/dL) and high Pb groups compared to the low Pb group, with a statistically
2 significant trend found for calcaneus bone loss. Compared to the low blood Pb level
3 group, women in the high blood Pb level group had an increased risk of non-spine
4 fracture (10.5 year interview follow-up), and women with medium or high blood Pb
5 levels had a higher risk of falls (4 year follow-up) with adjustment for age, clinic, BMI,
6 weight change between visits, smoking, chair stands (were able to stand up five times
7 from a chair, without using the arms of the chair), fracture history, estrogen use, and
8 baseline (1986-1988) bone mineral density. Nutritional factors were not considered. The
9 increased risk of lower bone density and falls leading to osteoporosis-related fractures
10 associated with blood Pb levels >4 (ig/dL are likely influenced by higher past Pb
11 exposures of these women.
12 Supporting evidence was provided by cross-sectional epidemiologic studies, although the
13 direction of the association and the magnitude, timing, frequency, and duration of Pb
14 exposure that contributed to the observed associations are uncertain. Further, most studies
15 did not consider potential confounding by nutritional factors. A large NHANES II
16 analysis of 8,654 adults > 50 years of age (Campbell and Auinger. 2007). which was
17 stratified by non-Hispanic white men (mean blood Pb: 4.9 [range: 0.7 to 48.1] (ig/dL),
18 non-Hispanic white women (mean blood Pb: 3.6 [range: 0.7 to 28.7] (ig/dL), African-
19 American men (mean blood Pb: 7.7 [range: 0.7 to 52.9] (ig/dL), and African-American
20 women (mean blood Pb: 4.5 [range: 0.7 to 23.3] (ig/dL). In analyses of covariance that
21 considered potential confounding (by age, race, sex, BMI, menopausal status, tobacco
22 use, alcohol use, physical activity, Ca2+ intake, chronic medical conditions, certain
23 medication use, and SES), non-Hispanic white men (n = 1,693, p <0.05) and women
24 (n = 1,754, p <0.10) in the highest tertile of concurrent blood Pb level had lower mean
25 total hip bone mineral density than non-Hispanic white men and women in the lowest
26 tertile of blood Pb levels (actual concentration not reported). Smaller differences were
27 observed in African-American men and women (possibly due to the smaller sample sizes
28 (n = 613, and 629, respectively). No association was observed between blood Pb levels
29 and osteoporotic fractures in any sex or race/ethnicity group.
30 Similar observations were made by Sun et al. (2008a) in 155 males and 37 females in
31 China who were occupationally-exposed to Pb (mean blood Pb: 20.22 and 15.5 (ig/dL,
32 respectively). In analyses (including all workers, plus 36 male and 21 female unexposed
33 controls stratified into groups according to blood Pb and urinary Pb levels), groups with
34 urinary Pb levels > 5 (ig/g creatinine had lower (p <0.01) bone mineral density compared
35 to groups with lower urinary Pb in each sex. Prevalence of osteoporosis increased with
36 increasing blood Pb in a linear manner. In contrast, a significant difference was observed
37 between blood Pb level and bone mineral density, but only in men with blood Pb levels
38 >30 (ig/dL. Prevalence of osteoporosis increased significantly with increasing blood Pb
November 2012 5-655 Draft - Do Not Cite or Quote
-------
1 in a linear manner. Results were not adjusted for potential confounding factors, including
2 other occupational exposures.
3 Cross-sectional epidemiologic studies also found associations between concurrent blood
4 Pb level and biological markers of bone turnover. Among 329 male (mean age: 65 years,
5 median blood Pb level: 2.2 (ig/dL) and 342 female (mean age: 62 years, median blood Pb
6 level: 1.9 (ig/dL) adults in North Carolina, Nelson et al. (2011) found in women that
7 higher blood Pb level was associated with higher uN-telopeptide cross-linked collagen
8 type I (uNTX-I, a marker of bone resorption/turnover) and uCTX-II (a marker associated
9 with the progression of radiographic knee and hip osteoarthritis) after adjusting for age,
10 BMI, race, and smoking status. In adjusted analyses of men, higher blood Pb level was
11 associated with higher uCTX-II, COMP, and C2C:CPII ratio (an indication of the balance
12 between cartilage collagen degradation and synthesis). In women, a weaker association
13 was found for COMP, a cartilage biomarker related to osteroarthritis. The results
14 indicated that blood Pb level is associated with bone turnover and mineralized cartilage
15 turnover in women, and with non-mineralized cartilage turnover in men.
16 Similarly, Machida et al. (2009) investigated bone matrix turnover in Japanese women
17 farmers, and how it is related to age-related menopause status and blood Pb level.
18 Perimenopausal women (n = 319 [age range: 49 to 55 years]) had higher geometric mean
19 blood Pb level (2.0 (ig/dL) than the other 3 groups did: premenopausal women (n = 261
20 [age range: 35 to 48 years], blood Pb level: 1.6 (ig/dL), younger postmenopausal women
21 (n = 397 [age range: 56 to 65 years], blood Pb level: 1.8 (ig/dL), or older postmenopausal
22 women (n = 248 [age range: 66 to 75 years], blood Pb level: 1.7 (ig/dL). In a model that
23 simultaneously included bone-mineral density, NTx, osteocalcin, and age, higher blood
24 Pb levels were positively associated with bone mineral density, NTx, and osteocalcin
25 (all p <0.01). In perimenopausal women, higher blood Pb level was predicted most
26 strongly by higher osteocalcin levels. Age was positively associated with higher blood Pb
27 levels in perimenopausal women only. Associations also were reported for bone-specific
28 ALP in unadjusted analyses.
29 To characterize mechanisms underlying the effects of Pb on bone, Jang et al. (2008)
30 studied the effect of Pb exposure on Ca2+-release activated Ca2+-influx (CRACI) using
31 cultures of human fetal osteoblast-like hFOB 1.19 cells (OLCs) in vitro. When cells were
32 incubated with 1,000 or 3,000 (iM Pb in the culture medium, a concentration-dependent
33 decrease on CRACI was observed, as was a concentration-dependent increase in the
34 influx of Pb into human OLC. These results suggest that Pb inhibits the measurable
35 influx of Ca2+ upon re-addition of Ca2+, which in turn, results in an influx of Pb into the
36 OLCs.
November 2012 5-656 Draft - Do Not Cite or Quote
-------
1 Studies have found inconsistent associations between higher blood Pb level and reduced
2 growth in children; however, Zuscik et al. (2007) hypothesized that Pb may alter growth
3 by altering chondrogenic commitment of mesenchymal cells and by affecting various
4 signaling pathways. Exposure of stage El 1.5 murine limb bud mesenchymal cells
5 (MSCs) to 1 (iM Pb in vitro caused increased basal and TGF-(3/BMP induction of
6 chondrogenesis, which was accompanied by nodule formation and upregulation of Sox-9,
7 type 2 collagen, and aggrecan, which are all key markers of chondrogenesis. Enhanced
8 chondrogenesis during induced ectopic bone formation also was found in mice that had
9 been pre-exposed to Pb acetate for six weeks via drinking water (55 or 233 ppm,
10 [previously shown to correspond to 14 or 40 (ig/dL blood Pb level, respectively]). MSCs
11 exposed to Pb in vitro exhibited an increase in TGF-(3, but BMP-2 signaling was
12 inhibited. Pb also induced NF-KB and inhibited AP-1 signaling. These results suggested
13 that the chondrogenesis induced by Pb exposure most likely involved modulation and
14 integration of multiple signaling pathways including TGF-(3, BMP, AP-1, and NF-KB.
15 Effects of Pb exposure on teeth were examined in a few recent cross-sectional
16 epidemiologic studies. A subset of the U.S. NHANES III (1988-1994) population was
17 selected for a large periodontitis versus Pb blood level study of both men (n = 2,500) and
18 women (n = 2,399), 30-55 years-old, that considered potential confounding by a large set
19 of factors, including nutritional status (Saraiva et al., 2007). Compared to individuals
20 with a concurrent blood Pb level of <3 (ig/dL, the prevalence ratios of periodontitis were
21 1.70 (95% CI: 1.02, 2.85) formen with concurrent blood Pb level of >7 (ig/dL and 3.80
22 (95% CI: 1.66, 8.73) for women with concurrent blood Pb level >7 (ig/dL. These results
23 were adjusted for age, NHANES III phase, cotinine levels, poverty to income ratio,
24 race/ethnicity, education, bone mineral density, diabetes, calcium intake, dental visits,
25 and menopause status in women.
26 Arora et al. (2009) examined the association between blood and bone Pb level and the
27 loss of natural teeth, in 333 men (age range: 50 to 94 years) from a subset of the Veterans
28 Affairs Normative Aging Study (NAS). Tooth loss was ascertained as the number of teeth
29 present during a dental assessment, and was categorized into three groups: 0 missing
30 teeth (n = 44), 1-8 missing teeth (n = 164), or > 9 missing teeth (n = 125). Men with > 9
31 teeth missing had significantly higher tibia and patella Pb concentrations (measured
32 within 3 years of dental assessment) compared to those with no tooth loss. Men with the
33 highest tibia Pb concentrations (>23 (ig/g) had higher odds of tooth loss (OR: 3.03 [95%
34 CI: 1.60, 5.75]) compared to men with tibia Pb levels < 15 (ig/g. Men with the highest
35 patella Pb levels (>36 (ig/g) also had higher odds of tooth loss (> 9 missing teeth versus
36 0-8 missing teeth; or > 1 missing teeth versus 0 missing teeth: OR: 2.41 [95% CI: 1.30,
37 4.49]) compared to men with patella Pb levels < 22.0 (ig/g. Men with tibia Pb levels
38 16-23 (ig/g, and men with patella Pb levels 23-36 (ig/g also had elevated odds of tooth
November 2012 5-657 Draft - Do Not Cite or Quote
-------
1 loss. Results were adjusted for age, education, smoking status, pack-years of smoking,
2 and diabetes, but nutritional factors were not considered. Tooth loss was not associated
3 with higher blood Pb levels [also measured within 3 years of dental assessment (Hu et al.,
4 1996b)1. indicating that long-term cumulative exposure to Pb is associated with increased
5 odds of tooth loss. However, because the timing of tooth loss was not ascertained and
6 bone Pb levels may represent exposures after tooth loss occurred, the directionality of
7 effects is uncertain.
5.9.4.3 Summary of Effects on Bone and Teeth
8 A few studies have indicated associations of Pb exposure or Pb biomarker levels with
9 bone disease (e.g., Paget's disease); however, the implications are limited by examination
10 of Pb-exposed workers or individuals with bone disease (i.e., case-control). Numerous
11 epidemiologic studies indicated an association between higher Pb biomarker levels and
12 lower bone density in adults. Prospective evidence was provided by a study of elderly
13 women (65-87 years-old), in which higher blood Pb levels were associated with lower
14 bone density measured after 2-4 years and greater risk of falls and osteoporosis-related
15 fractures (Khalil et al., 2008). Cross-sectional epidemiologic associations between higher
16 blood Pb levels and lower bone mineral density were found in adults without (Campbell
17 and Auinger. 2007) and with occupational Pb exposure (Sun et al., 2008a). Cross-
18 sectional studies also indicated associations between higher blood Pb levels and higher
19 markers of bone turnover in elderly populations (Nelson et al., 2011; Machidaet al.,
20 2009). In the cross-sectional epidemiologic evidence, it is difficult to determine whether
21 an increase in blood Pb level results from lower bone density or from higher bone
22 turnover, and whether these effects lead to a greater release of Pb from bone into the
23 bloodstream. Except for Sun et al. (2008a), studies adjusted for several potentially
24 important confounding factors, including age, BMI, and smoking. However, studies did
25 not consider nutritional status, which could affect the release of Pb from bone to blood.
26 To support the direction and independent effects of Pb on bone, toxicological studies
27 have found Pb-induced (gestational and postnatal) decreases in bone growth in juvenile
28 animals. Further, these toxicological studies have characterized potential modes of action,
29 by showing Pb-induced decreases in bone mineralization and bone cell differentiation,
30 inhibition of CRACI, and alterations in signaling pathways involved in skeletal
31 development (Jang et al., 2008; Zuscik et al., 2007).
32 Epidemiologic studies have found associations between blood Pb levels and effects on
33 teeth. Large NHANES analyses adjusted for several potentially important confounding
34 factors (including age, SES-related factors, and nutritional factors), and found
35 associations between concurrent blood Pb level and dental caries in children (Moss et al..
November 2012 5-658 Draft - Do Not Cite or Quote
-------
1 1999) and periodontitis in adults (Saraiva et al., 2007). Higher patella and tibia Pb levels
2 were associated with tooth loss in NAS men (Arora et al.. 2009). The results for blood Pb
3 and bone Pb levels in adults indicate that long-term, cumulative exposure to Pb exposure
4 is associated with negative effect on teeth. This epidemiologic evidence was based on
5 cross-sectional study design analyses, which precludes conclusions about the
6 directionality of effects. However, these findings are supported by toxicological evidence
7 in animals for Pb-induced increases in Pb uptake into teeth; and decreases in cell
8 proliferation, procollagen type I production, intracellular protein, and osteocalcin in cells
9 exposed to Pb in vitro.
10 The small body of epidemiologic evidence showing associations between Pb biomarker
11 levels and various bone and teeth effects (after adjusting for potential confounding by
12 age, SES-related factors, and nutritional factors), plus the supporting toxicological
13 evidence, is sufficient to conclude that there is a likely causal relationship between Pb
14 exposure and effects on bone and teeth.
5.9.5 Effects on Ocular Health
15 Ocular effects most commonly associated with exposure to Pb include formation of
16 cataracts, impaired vision, edema and retinal stippling.
5.9.5.1 Summary of Key Findings of the Effects on Ocular Health
(2006 Pb AQCD)
17 The 2006 Pb AQCD stated that various changes in the visual system were observed with
18 Pb poisoning including retinal stippling and edema, cataracts, ocular muscle paralysis and
19 impaired vision. Maternal prenatal blood Pb levels in the range of 10.5 to 32.5 ug/dL
20 were associated with supernormal retinal ERGs in children at age 5-7 years. Cataracts
21 were noted in middle-aged men with tibia bone Pb levels of 31-126 ug/g.
5.9.5.2 Recent Toxicological and Epidemiologic Studies
22 The recent cross-sectional epidemiologic studies of ocular effects in adults did not
23 produce clear evidence, and each was limited by the lack of rigorous statistical analysis
24 and lack of consideration for potential confounding. Erie et al. (2009) measured Pb and
25 Cd in retinal tissue from 36 eye donors with age-related macular degeneration (cases) and
26 25 normal control donors. Pb, but not Cd, concentration was significantly elevated in the
27 neural retina tissue of the 36 donors with macular degeneration (72 eyes; median [IQR]:
November 2012 5-659 Draft - Do Not Cite or Quote
-------
1 12.0 [8-18] ng/g Pb) versus normal control donors (50 eyes; median [IQR]: 8.0 [0-11]
2 ng/g Pb). Neither of these heavy metals were significantly elevated in the retinal pigment
3 epithelium (RPE)/choroid complex in donors with macular degeneration over normal
4 controls. Mosad et al. (2010) compared Pb, Cd, vitamin C, vitamin E, and beta carotene
5 blood levels between 45 middle-aged male smokers and nonsmokers with cataracts.
6 Blood Pb levels were elevated (p <0.0001) in 15 light (mean [SD]: 14.5 [0.41] (ig/dL), 15
7 moderate (14.5 [0.41] (ig/dL), and 15 heavy smokers (18.7 [1.24] (ig/dL) compared to 15
8 nonsmokers (12.2 [0.21] (ig/dL). Similar associations were observed for Cd blood levels
9 and lens concentrations. There was no direct analysis of the association between Pb blood
10 level or lens Pb concentration and the severity of cataracts.
11 Recent animal studies have observed Pb-induced retinal progenitor cell proliferation and
12 neurogenesis (Section 5.3.7.3). An in vitro study found increased opacity of rat (age
13 4-6 weeks) lens exposed to 1 (iM Pb nitrate with or without secondary oxidative
14 challenge after 5-8 days but not after 3 days (Neal et al.. 2010b). Thus, short-term Pb
15 exposure did not induce osmotic swelling or lens shrinkage. With a 5-day exposure, 30%
16 of the Pb-exposed lenses displayed "definite cataracts" compared to only 2.5% of control
17 lenses. By culture day 8, 100% of the exposed lenses were described either as clearly
18 opaque or definite cataracts, while only 7% of control lenses displayed these
19 characteristics, indicating that prolonged exposure of lenses to Pb induced an accelerated
20 formation of opacity/cataract compared to unexposed lenses. Pb-exposed lenses cleared
21 the media of hydrogen peroxide more rapidly than did control lenses, potentially due to
22 increased CAT activity. Exposure to hydrogen peroxide resulted in total (100%) opacity
23 in Pb-exposed lenses at culture day 7, compared to less than 20% in control cells.
24 Exposure to Pb additionally altered epithelial nutrient transport and lens histology,
25 relative to that in controls.
26 In summary, prospective epidemiologic evidence indicates associations between prenatal
27 maternal blood Pb levels of 10.5-32.5 (ig/dL and supernormal retinal ERGs in children
28 ages 5-7 years after adjusting for age, sex, and head circumference. However, the
29 relevance of supernormal ERGs is uncertain. Evidence in adults for associations between
30 eye tissue Pb levels and macular degeneration (Mosad et al.. 2010) and cataracts in adults
31 (Erie et al.. 2009) is limited by weak statistical methods and lack of consideration for
32 potential confounding to warrant conclusions. Toxicology studies have reported
33 Pb-induced retinal progenitor cell proliferation, retinal ERGs, and lens opacity
34 (Section 5.3.7.3). Because of the insufficient quantity and quality of studies in the
35 cumulative body of evidence, the evidence is inadequate to determine a causal
36 relationship between Pb exposure and ocular effects.
November 2012 5-660 Draft - Do Not Cite or Quote
-------
5.9.6 Effects on the Respiratory System
1 Blood Pb level has been associated with asthma and allergy in children in prospective and
2 cross-sectional epidemiologic studies (Section 5.6.5.2). As described in Section 5.2.4.
3 Pb exposure has been shown to induce the generation of ROS. ROS are implicated in
4 mediating increases in bronchial responsiveness and activating neural reflexes, leading to
5 decrements in lung function. Collectively, studies investigating these airway responses in
6 asthma-free populations are limited in number, lack rigorous statistical analysis, and
7 collectively do not provide strong evidence of an association with blood Pb level
8 (Section 5.6.5.3). Collectively, panel and time-series epidemiologic studies demonstrate
9 associations between short-term increases in ambient air Pb (measured in PM2 5 or PMi0
10 air samples), and decreases in lung function and increases in respiratory symptoms, and
11 asthma hospitalizations in children but not adults (Section 5.6.5.3). Toxicological studies
12 have found pulmonary inflammation induced by concentrated ambient air particles
13 (CAPs) in which Pb was one of the numerous components (Wei et al.. 2011; Duvall et al..
14 2008; Godleski et al.. 2002; Saldiva et al.. 2002). Despite this evidence for respiratory
15 effects related to air-Pb concentrations, the limitations of air-Pb studies, including the
16 limited data on the size distribution of Pb-PM (Section 3.5.3). the uncertain relationships
17 of Pb-PMio and Pb-PM2 5 with blood Pb levels, and the lack of adjustment for other
18 correlated PM chemical components preclude firm conclusions about air Pb-associated
19 respiratory effects. Because of the insufficient quantity and quality of studies in the
20 cumulative body of evidence, the evidence is inadequate to determine a causal
21 relationship between Pb exposure and respiratory effects in populations without asthma.
5.10 Cancer
22 Previous AQCDs have demonstrated that Pb is a well-established animal carcinogen.
23 Oral Pb acetate exposure to male and female rodents has consistently been shown to be a
24 kidney carcinogen in multiple separate studies, inducing adenocarcinomas and adenomas
25 after chronic exposure. Developmental Pb acetate exposure also induced kidney tumors
26 in offspring whose dams received Pb acetate in drinking water during pregnancy and
27 lactation. Gliomas of the brain have also been reported after oral Pb exposure. These
28 rodent toxicological studies have been conducted at high doses of Pb and have shown that
29 Pb is an animal carcinogen. Because of this strong body of historical data, the
30 2006 Pb AQCD states, "limited tumorigenesis studies have been conducted in animal
31 models and the focus has been more on the mechanism of neoplasia... and possible
32 immunomodulatory effects of Pb in the promotion of cancer." More recent studies have
November 2012 5-661 Draft - Do Not Cite or Quote
-------
1 focused on administration of Pb with known carcinogens or modifiers such that lifestage,
2 diet, and mechanism of action can be belter understood.
3 The previous epidemiologic studies included in the 2006 Pb AQCD (U.S. EPA. 2006b)
4 "provide [d] only very limited evidence suggestive of Pb exposure associations with
5 carcinogenic or genotoxic effects in humans," and the studies were summarized as
6 follows:
"The epidemiologic data ... suggest a relationship between Pb exposure and cancers of the
lung and the stomach... Studies of genotoxicity consistently link Pb-exposed populations
with DNA damage and micronuclei formation, although less consistently with
chromosomal aberrations."
7 The International Agency for Research on Cancer (IARC) classified inorganic Pb
8 compounds as probable human carcinogens (Group 2A of IARC classification) based on
9 sufficient evidence in animal studies (evidence in human studies was limited), and
10 organic Pb compounds as not classifiable (Group 3 of IARC classifications) (IARC.
11 2006a; Rousseau et al.. 2005). Additionally, the National Toxicology Program (NTP)
12 listed Pb and Pb compounds as "reasonably anticipated to be human carcinogens" (NTP.
13 2011). The typical cancer bioassays employed by IARC or NTP as evidence of
14 Pb-induced carcinogenicity used rodents that were continuously exposed to Pb acetate in
15 chow or drinking water for 18 months to two years in duration. These two year cancer
16 bioassays and the doses administered are typical of cancer bioassays used with other
17 chemicals.
18 In the following sections, recent epidemiologic and toxicological studies published since
19 the 2006 Pb AQCD, regarding Pb and cancer mortality and incidence are examined. In
20 addition, recent studies of Pb exposure associated with DNA and cellular damage, as well
21 as epigenetic effects, are summarized. When the information is available, the form of the
22 Pb compound under study (e.g., inorganic, organic) is indicated. In epidemiologic
23 studies, various biological indicators of Pb exposure are used including Pb measured in
24 blood and bone. The biological indicators of Pb associated with cancer-related endpoints
25 are considered in drawing conclusions about potentially important levels and timing of Pb
26 exposure. Bone Pb is indicative of cumulative Pb exposure. Blood Pb can represent more
27 recent exposure, but because it can also represent remobilized Pb occurring during times
28 of bone remodeling, blood Pb level may also be an indicator of long-term Pb exposure in
29 adults. Toxicological studies only report exposure by blood Pb or exposure dose. More
30 detailed discussion of these measures is given in Section 4.3.5. Details of the recent
31 epidemiologic and toxicological studies follow.
November 2012 5-662 Draft - Do Not Cite or Quote
-------
5.10.1 Cancer Incidence and Mortality
1 Recent studies have included epidemiologic evaluations of the associations between Pb
2 exposure and both specific cancers (such as lung cancer and brain cancer), and overall
3 cancer (cancer of any type). Table 5-49 provides an overview of the study characteristics
4 and results for the epidemiologic studies that reported effect estimates. This section also
5 evaluates toxicological evidence on the potential carcinogenicity of Pb.
November 2012 5-663 Draft - Do Not Cite or Quote
-------
Table 5-49 Summary of recent epidemiologic studies of cancer incidence and overall cancer mortality.
Reference
(In order of Study
appearance in text.) Location
Cancer Study
Outcome Population
Methodological Measure of
Details Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Overall Cancer Mortality:
Menke et al. (2006) Multiple U.S.
locations
Schoberet al. Multiple U.S.
(2006) locations
Overall NHANES III
cancer cohort with
mortality Blood Pb
measures in
1 988-1 994
At least 1 2 years
of follow-up
Blood Pb
<10 ug/dL
N=1 3,946
N for cancer
mortality = 411
Overall NHANES III
cancer cohort
mortality At |east 40 years
of age
N= 9,757
N for cancer
mortality = 543
Cohort study Blood Pb at
using Cox baseline
regression and
other techniques
Cohort study Blood Pb at
using Cox baseline
proportional
hazard regression
analysis and other
techniques
2.58 ug/dL
(geometric
mean)
Tertile 1 :
<1.93 ug/dL
Tertile 2:
1. 94-3.62 ug/dL
Tertile 3:
> 3.63 ug/dL
Blood
Pb<5 ug/dL:
67.7%
Blood Pb
5-9 ug/dL:
26.0%
Blood
Pb>=10 ug/dL:
6.3%
HR (95% Cl):
Tertile 1 :
1.00
Tertile 2:
0 72
(95% Cl: 0.46, 1.12)
Tertile 3:
1.10
(95% Cl: 0.82, 1.47)
RR (95% Cl):
Blood Pb<5 ug/dL:
1.00
Blood Pb 5-9 ug/dL:
1.44 (95% Cl: 1.12,
1.86)
Blood Pb> 10 ug/dL:
1.69 (95% Cl: 1.14,
2.52)
Note: Modification by
age assessed and
associations varied
Age, race-
ethnicity, sex,
diabetes mellitus,
body mass index,
current or former
smoking, alcohol
consumption,
physical activity,
low income, CRP,
total cholesterol,
high school
education, urban
residence,
postmenopausal
status,
hypertension, and
level of kidney
function
Sex, race /
ethnicity,
education, and
smoking status
Age used as time-
scale in models
Additional
covariates
considered but not
included: Census
region and urban
status of
residence, alcohol
intake
slightly
November 2012
5-664
Draft - Do Not Cite or Quote
-------
Reference
(In order of Study Cancer
appearance in text.) Location Outcome
Weisskopf et al. Boston, MA Overall
(2009) area cancer
mortality
Khalil et al. (2009b) Baltimore, MD, Overall
and cancer
Monongahela mortality
Valley, PA
Study
Population
Normative Aging
Study (MAS)
Included men
only (mostly
white)
Mean follow-up
period for this
study: 8.9 yr
Blood Pb
measures
available
N=1038
N Tor cancer
mortality=85
Bone Pb
measures
available
N=727
N for cancer
mortality=57
Subgroup of the
Study of
Osteoporotic
Fractures cohort
Included white
women aged
65-87;
1 2 yr (+/- 3 yr)
follow-up
N=533
N for cancer
mortality=38
Methodological Measure of
Details Pb Exposure
Cohort study Blood Pb at
using baseline,
Cox proportional Patella Pb at
hazards baseline
Cohort study Blood Pb at
using baseline
Cox proportional
hazards
regression
analysis and other
techniques
Mean Pb
(SD)
Blood Pb:
5.6 ug/dL (3.4)
Tertile 1 of
Blood Pb:
<4 ug/dL
Tertile 2 of
Blood Pb:
4-6 ug/dL
Tertile 3 of
Blood Pb:
>6 ug/dL
Tertile 1 of
patella Pb:
<22ug/g
Tertile 2 of
patella Pb:
22-35ug/g
Tertile 3 of
patella Pb:
>35 ug/g
Blood Pb Level
5.3 (2.3)ug/dL
Adjusted Effect
Estimates
HR (95% Cl):
Blood Pb Tertile 1:
1.00
Blood Pb Tertile 2:
1.03(95% Cl: 0.42,
2.55)
Blood Pb Tertile 3:
0.53 (95% Cl: 0.20,
1.39)
Patella Pb Tertile 1:
1.00
Patella Pb Tertile 2:
0.82 (95% Cl: 0.26,
2.59)
Patella Pb Tertile 3:
0.32 (95% Cl: 0.08,
1 .35)
HR (95% Cl):
Blood Pb<8 ug/dL:
1.00
Blood Pb> 8 ug/dL:
1.64(95% Cl: 0.73,
3.71)
Potential
confounders
adjusted for in
analysis
Age, smoking, and
education
Additional
covariates
considered but not
included: alcohol
intake, physical
activity, body
mass index, total
cholesterol, serum
high-density
lipoprotein,
diabetes mellitus,
race, and
hypertension
Age, clinic, BMI,
education,
smoking, alcohol
intake, estrogen
use, hypertension,
walking for
exercise,
diabetes, and total
hip BMD
November 2012
5-665
Draft - Do Not Cite or Quote
-------
Reference
(In order of
appearance in text.)
Study
Location
Cancer
Outcome
Study
Population
Methodological
Details
Measure of
Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Overall Cancer Incidence:
Absalon and Slesak
(2010)
Obhodaset al.
(2007)
Mendy et al.
(2012)
Silesia
province,
Poland
Island of Krk,
Croatia
Multiple U.S.
locations
Overall
cancer
incidence
Incidence
rates for
neoplasms
Incidence of
cancer or
"malignancy
of any kind"
Children living in
this province at
least five years
N = not specified
Individuals living
in the Island of
Krk from
1997-2001
N= 1,940
2007-2008
NHANES cohort
- at least 20
years of age
N= 1,857
Ecologic analysis
using correlations.
Cross-sectional
study using
correlations and
linear regression
Cross-sectional
study using
logistic regression
Pb-related air
pollution
measures
Soil and
vegetation
samples,
household
potable water
samples,
children's hair
samples
Concurrently
measured
creatinine-
adjusted urinary
Pb
NA
NA
Geometric
mean for
creatinine-
adjusted urinary
Pb marker:
0.59 ug/g (95%
Cl: 0.57, 0.61)
Reported
correlations between
changes in Pband
cancer incidence -
no/low correlations
observed (correlation
coefficients between
-0.3 and 0.2)
No association
observed between
Pb in the samples
and incidence of
neoplasm (numerical
results not provided)
OR (95% Cl):
Greater than log-
transformed mean
creatinine-adjusted
urinary Pb level
compared to less
than log-transformed
mean creatinine-
adjusted urinary Pb
level: 0.76 (0.44,
1.33)
None specified
Examined
correlation by sex,
no difference
reported
None specified
Age, sex,
race/ethnicity,
education level,
ratio family
income to poverty,
alcohol
consumption,
cigarette smoking,
and other heavy
metals
November 2012
5-666
Draft - Do Not Cite or Quote
-------
Reference
(In order of Study
appearance in text.) Location
Cancer Study
Outcome Population
Methodological
Details
Measure of
Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Lung Cancer:
Lundstrom et al. Sweden
(2006)
Jones et al. (2007) Humberside,
U.K.
Lung cancer Male Pb smelter
(incidence workers first
and mortality) employed for
a 3 months
between 1 928
and 1979
Followed up for
mortality from
1 955 -1 987
N=187
N lung
cancer=46
Lung cancer Male tin smelter
mortality employees
N=1,462
Nested case-
referent study
using conditional
logistic regression
Cohort study
using Poisson
regression
Median peak
blood Pb level
Median number
of years with at
least one blood
sample
obtained
Median
cumulative
blood Pb index
(sum of annual
blood Pb Level)
Personnel
record cards
and air
sampling
conducted from
1972-1991
Three exposure
scenarios
determined for
working lifetime
cumulative
exposure - all
have similar
medians of
approximately
2 mg/m3-yr
Median peak
blood Pb Level:
cases
49.7 ug/dL,
controls
55.9 ug/dL
Median number
of years with at
least one blood
sample
obtained:
cases 4.5 yr,
controls 6.0 yr
Median
cumulative
blood Pb index:
cases
186 ug/dL,
controls
246 ug/dL
~2.0 mg/m3-yr
OR (95% Cl):
Median peak blood
Pb Level:
1 .00 (0.71 , 1 .42)
Median number of
years with at least
one blood sample
obtained:
0.98(0.96, 1.01) per
10ug/dL
Median cumulative
Blood Pb index:
1.00(0.98, 1.01) per
10 ug/dL
Note: similar results
were observed when
restricted to smokers
only
RR for Pb exposure
weighted age and
time since exposure
(90% Cl): 1.54(1.14,
2.08)
Note: Similar results
for other exposure
determination
scenarios.
Matched by age
Adjusted for
smoking and As
exposure
Not specified
November 2012
5-667
Draft - Do Not Cite or Quote
-------
Reference
(In order of
appearance in text.)
Study
Location
Cancer
Outcome
Study
Population
Methodological
Details
Measure of
Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Rousseau et al.
(2007)
Montreal,
Canada
Lung cancer
incidence
Men aged 35-79
N population
based controls=
ranged from 271
to 471
depending on
exposure of
interest
N controls with
other cancers=
ranged from 737
to 1203
depending on
exposure of
interest
N lung
cancer= ranged
from 433 to 751
depending on
exposure of
interest
Population-based Interview of job
case-control study history and
using exposure matrix
unconditional
logistic regression
Ever exposed
to:
Organic Pb
3.0%
Inorganic Pb
17.0%
Pb in gasoline
emissions
38.6%
OR (95% Cl):
Organic Pb exposure
compared to no
exposure:
Lung
1.3 (95% 01:0.5,3.1)
Inorganic Pb
exposure compared
to no exposure:
Lung
1.1 (95%CI:0.7, 1.7)
Pb in gasoline
emissions exposure
compared to no
exposure:
Lung
0.8 (95% Cl: 0.6, 1.1)
Note: results are for
comparisons using
population-based
controls; results for
controls with other
types of cancers
were similar
Age, family
income, cultural
origin, proxy
status, ever
exposure to
asbestos, silica,
As, Cd, and
chromium (VI)
November 2012
5-668
Draft - Do Not Cite or Quote
-------
Reference
(In order of
appearance in text.)
Study
Location
Cancer
Outcome
Study
Population
Methodological
Details
Measure of
Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Brain Cancer:
van Wijngaarden
and Dosemeci
(2006)
Multiple U.S
locations
Brain cancer
mortality
National
Longitudinal
Mortality Study -
included
individuals with
occupational
information
-included follow-
up from
1970-1989
N= 317,968
Cohort study
using proportional
hazards, Poisson
regression
techniques, and
standardized
mortality ratios
(SMR)
Interview about
current or most
recent job within
the past 5 years
and a job
exposure matrix
NA
HR (95% Cl):
Any Pb exposure
compared to no
exposure 1.56
(95% Cl: 1.00, 2.43)
RR (95% Cl) from
Poisson regression:
1.42
(0.91, 2.20)
SMR (95% Cl):
Not exposed: 0.87
(0.70, 1.06)
Any exposure:
1.11
(0.74, 1.59)
Gender, age, race,
living in an urban
area, marital
status and
educational level
Additional
covariate
considered but not
included: Family
income (not used
due to large %
missing; additional
analysis including
it gave similar
results)
Note: Effect
estimates were
greatest among
those with high
probabilities of
exposure and
medium/high
exposure intensity
November 2012
5-669
Draft - Do Not Cite or Quote
-------
Reference
(In order of
appearance in text.)
Study
Location
Cancer
Outcome
Study
Population
Methodological
Details
Measure of
Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Rajaraman et al.
(2006)
Phoenix, AZ,
Boston, MA,
and Pittsburgh,
PA
Brain cancer
incidence
NCI Brain Tumor
Study
- included
individuals >=18
yr diagnosed
with brain cancer
less than 8 week
before
hospitalization;
frequency-
matched controls
were individuals
admitted to the
same hospitals
for non-
neoplastic
conditions
N controls
=799
N glioma
= 489
N meningioma
=197
Case-control
study using
unconditional
logistic regression
Interviews of
lifetime work
history and
exposure
databases
NA
OR (95% Cl):
Meningioma:
Ever exposure to Pb
0.8(0.5, 1.3)
Glioma:
Ever exposure to Pb
0.8(0.6, 1.1)
Note: positive
associations
between Pb
exposure and
meningioma
incidence was
observed among
individuals with
ALAD2 genotypes,
but not individuals
with ALAD1
genotypes; these
associations were
not observed for
glioma incidence
Age, sex, race /
ethnicity, hospital,
and residential
proximity to
hospital
November 2012
5-670
Draft - Do Not Cite or Quote
-------
Reference
(In order of Study Cancer Study Methodological Measure of
appearance in text.) Location Outcome Population Details Pb Exposure
Bhatti et al. (2009) Phoenix, AZ, Brain cancer NCI Brain Tumor Case-control Interviews of
Boston, MA, incidence Study study using lifetime work
and Pittsburgh, included non unconditional history and
PA Hispanic whites logistic regression exposure
> .{jj yr databases
diagnosed with
brain cancer less
than 8 week
before
hospitalization;
frequency-
matched controls
were individuals
admitted to the
same hospitals
for non-
neoplastic
conditions
N controls
=494
N glioma
= 362
N meningioma
=134
Mean Pb
(SD)
Glioma:
70.5 ug/m -yr
(193.8 ug/m i -y)
Glioblastoma
multiform:
97.5 ug/m3-yr
(233.9 ug/m -y)
Meningioma:
101.1 ug/m3-yr
(408.7 ug/m3-y)
Controls'
69.7 ug/m3-yr
(248.8 ug/rrvSO
Potential
confounders
Adjusted Effect adjusted for in
Estimates analysis
OR (95% Cl) per Age, sex, hospital,
100 ug/m3y increase and residential
in cumulative Pb proximity to the
exposure hospital
Glioma'
1.0(0.9, 1.1)
Glioblastoma
multiform:
1.0(0.9, 1.1)
Meningioma: 1.1
On *i o\
.0, 1.2)
Note: modification by
SNPs was conducted
and associations
varied by SNP
November 2012
5-671
Draft - Do Not Cite or Quote
-------
Reference
(In order of Study
appearance in text.) Location
Cancer Study
Outcome Population
Methodological
Details
Measure of Mean Pb
Pb Exposure (SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
Breast Cancer:
Pan et al Canada
.(2011)
Breast cancer National
incidence Enhanced
Cancer
Surveillance
System
(NECSS)-
population-
based sample of
cancer cases
and controls with
information
collected from
1 994-1 997
N controls
=2467
N cases=
2343
Population-based
case-control study
using
unconditional
logistic regression
Self-reported NA
previous
addresses and
their proximity
to Pb smelters
(determined
using
Environmental
Quality
Database
[EQDB])
OR (95% Cl):
Residing
>3.2 km from Pb
smelter or no nearby
smelter
1.00
Residing
0.8-3.2 km from Pb
smelter:
0.41
(0.11, 1.51)
Residing
<0.8 km from Pb
smelter:
0.61
(0.11 , 3.42)
Age, province of
residence,
education,
smoking pack
years, alcohol
consumption,
body mass index,
recreational
physical activity,
number of live
births, age at
menarche,
menopausal
status, total
energy intake, and
employment in the
industry under
consideration
Additional
covariate
considered but not
included: Family
income
Other Cancers:
Rousseau et al. Montreal,
(2007) Canada
Various Men aged 35-79
cancer N p0pu|ati0n
incidences baseS contro|s=
ranged from 271
to 471
depending on
the cancer and
exposure of
interest
N controls with
other cancers=
ranged from 697
to 2,250
depending on
exposure of
interest
N
cancer= ranged
from 60 to 442
depending on
the cancer and
exposure of
interest
Population-based
case-control study
using
unconditional
logistic regression
Interview of job Ever exposed
history and to:
exposure matrix organic Pb
3.0%
Inorganic Pb
1 7.0%
Pb in gasoline
emissions
38.6%
OR (95% Cl):
Never exposed is
referent group
Organic Pb:
Esophageal
1.7(0.5,6.4)
Stomach
3.0(1.2,7.3)
Colon
1.5(0.7,3.6)
Rectum
3.0(1.2,7.5)
Pancreas
0.9(0.1,5.2)
Prostate
1.9(0.8,4.6)
Bladder
1.7(0.7,4.2)
Kidney
Age, family
income, cultural
origin or
birthplace, and
proxy status; all
models except
those for
melanoma and
non-Hodgkin's
lymphoma were
adjusted for
smoking
November 2012
5-672
Draft - Do Not Cite or Quote
-------
Reference
(In order of
appearance in text.)
Study
Location
Cancer
Outcome
Study
Population
Methodological
Details
Measure of
Pb Exposure
Mean Pb
(SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
2.3(0.8,6.7)
Non-Hodgkin's
lymphoma
0.4(0.1,2.2)
Inorganic Pb:
Esophageal
0.6(0.3,1.2)
Stomach
0.9(0.6, 1.5)
Colon
0.8(0.5,1.1)
Rectum
0.8(0.5, 1.3)
Pancreas
0.9(0.4,1.8)
Prostate
1.1 (0.7, 1.6)
Bladder
1.1 (0.7,1.5)
Kidney
1.0(0.6, 1.7)
Melanoma
0.4(0.2,1.0)
Non-Hodgkin's
lymphoma
0.7(0.4, 1.2)
Pb in gasoline
emissions:
Esophageal
0.6(0.4,1.1)
Stomach
1.0(0.7, 1.4)
Colon
0.8(0.6,1.1)
Rectum
1.0(0.7, 1.4)
Pancreas
0.9(0.5,1.4)
Prostate
0.9(0.7, 1.2)
Bladder
0.8(0.6, 1.1)
Kidney
1.0(0.7,1.5)
Melanoma
November 2012
5-673
Draft - Do Not Cite or Quote
-------
Reference
(In order of
appearance in text.)
Study
Location
Cancer Study
Outcome Population
Methodological Measure of Mean Pb
Details Pb Exposure (SD)
Adjusted Effect
Estimates
Potential
confounders
adjusted for in
analysis
0.8(0.5,1.4)
Non-Hodgkin's
lymphoma
0.7(0.5,1.0)
Note: results are for
comparisons using
population-based
controls; results for
controls with other
types of cancers
were similar except
no association was
present between
organic Pb and rectal
cancer
November 2012
5-674
Draft - Do Not Cite or Quote
-------
Reference
(In order of Study Cancer
appearance in text.) Location Outcome
Santibanez et al. Valencia and Esophageal
(2008) Alicante, Spain cancer
incidence
Study Methodological Measure of Mean Pb
Population Details Pb Exposure (SD)
PANESOES Case-control Interviews to NA
study study using determine
included 30-80 unconditional occupational
vr old men logistic regression history and a
hospitalized in job exposure
any of the matrlx
participating
study hospitals
N controls=285
N cancer=185
(147 squamous
cell, 38 adeno.
carcinoma)
Potential
confounders
Adjusted Effect adjusted for in
Estimates analysis
OR (95% Cl): Age, hospital
All esophageal location,
cancers: educational level,
Unexposed: 1.00 smoking and
Low workplace alcohol use
Pb exposure
(< 4.9 ug/dL):
0.79 (0.43, 1 .46)
High workplace
Pb exposure
(>4.9 ug/dL):
1.69(0.57,5.03)
Esophageal
squamous cell
carcinoma:
Unexposed: 1.00
Low workplace
Pb exposure
(< 4.9 ug/dL):
0.70 (0.34, 1 .43)
High workplace Pb
exposure (>4.9
ug/dL):
0.91 (0.22, 3.75)
Adenocarcinoma:
Unexposed: 1.00
Low workplace
Pb exposure
(< 4.9 ug/dL):
0.95 (0.32, 2.82)
High workplace
Pb exposure
(>4.9 ug/dL):
5.30(1.39,20.22)
*Note: associations
not changed or
slightly increased
when restricted to
occupational
exposures > 15yr
November 2012
5-675
Draft - Do Not Cite or Quote
-------
5.10.1.1 Overall Cancer Mortality
1 Several recent cohort studies examined the association between Pb levels and cancer
2 mortality, including multiple analyses of the NHANES III population. In one NHANES
3 III analysis, the cohort of 13,946 (N for cancer mortality=411) was followed for 12 years
4 and individuals with blood Pb levels greater than 10 (ig/dL were excluded from the study
5 (mean baseline blood Pb level was 2.58 (ig/dL). No association was observed between
6 blood Pb and cancer mortality (HR of highest tertile [> 3.63 (ig/dL] compared to lowest
7 tertile [<1.93 (ig/dL]: 1.10 [95% CI: 0.82, 1.47]) (Menke et al.. 2006). Another analysis
8 of the NHANES III population, which was restricted to individuals 40 years and older at
9 the time of blood Pb collection and included 9,757 (N for cancer mortality=543)
10 individuals with all blood Pb levels (including those greater than 10 (ig/dL), reported
11 associations between blood Pb and cancer mortality (Schober et al.. 2006). In this study,
12 median follow-up time was 8.6 years. The RRs were 1.69 (95% CI: 1.14, 2.52) for
13 individuals with blood Pb levels of at least 10 (ig/dL and 1.44 (95% CI: 1.12, 1.86) for
14 blood Pb levels of 5-9 (ig/dL compared to individuals with blood Pb levels less than
15 5 (ig/dL. When stratified by age, point estimates comparing blood Pb levels of 5-9 versus
16 less than 5 (ig/dL were similar across all age groups but only statistically significant
17 among 75-84 year olds. The risks of mortality associated with blood Pb levels > 10 (ig/dL
18 in the groups aged 40-74 years and 85 years and older were elevated.
19 A study of men (primarily white) from the greater Boston, MA area enrolled in the
20 Normative Aging Study (NAS) found no association between blood or bone Pb and
21 cancer mortality in adjusted analyses (N=l,038, N for cancer mortality=85 when using
22 blood Pb measures; N=727, N for cancer mortality=57 when using bone Pb measures). At
23 baseline, the mean (SD) blood Pb level for this population was 5.6 (3.4) (ig/dL and blood
24 Pb was poorly correlated with measured bone Pb (Weisskopf et al.. 2009). As part of the
25 Study of Osteoporotic Fractures , 533 white women aged 65-87 (N for cancer
26 mortality=38) were included in a sub-study of blood Pb level and cancer mortality and
27 were followed for approximately 12 years (Khalil et al.. 2009b). The mean (SD) blood Pb
28 level at baseline was 5.3 (2.3) (ig/dL and no association was detected between blood Pb
29 and cancer mortality in the study population.
30 Overall, epidemiologic studies of blood Pb levels and cancer mortality reported
31 inconsistent results. An epidemiologic study using NHANES III data demonstrated the
32 strongest association between blood Pb and increased cancer mortality; however, other
33 studies reported weak or no associations. These cohort studies were well-conducted
34 longitudinal studies with control for potential confounders, such as age, smoking, and
November 2012 5-676 Draft - Do Not Cite or Quote
-------
1 education (see list of the potential confounders addressed in each study in Table 5-49).
2 One limitation is that the studies in populations other than NHANES cohorts each had a
3 small number of cancer mortality cases.
5.10.1.2 Overall Cancer Incidence
4 Studies of overall cancer incidence have also been performed (Table 5-49). An ecologic
5 analysis compared Pb-related air pollution over 5 year increments from 1990 to 2005
6 with incidence rates of cancer during this time period (cancer sites not specified) among
7 children (N not specified) (Absalon and Slesak. 2010). The highest Pb air pollution levels
8 were measured in 1990 when over 50% of the study area exceeded the limit of
9 1 (ig/m2-year. No correlation was observed both overall and in sex-specific analyses.
10 Another study (N= 1,940) examined correlations between Pb concentrations in soil, water,
11 vegetation, and hair samples with incidence of neoplasms (Obhodas et al.. 2007). The Pb
12 concentrations were not correlated with incidence of neoplasms. A recent study using the
13 2007-2008 NHANES cohort reported no association between higher creatinine-adjusted
14 urine Pb levels and elevated odds of having ever had cancer or a malignancy (N=l,857)
15 (Mendy et al.. 2012). The timing of cancer diagnosis in relation to the urine sample
16 collection was not identified.
17 Overall, epidemiologic studies reported no positive associations between various
18 measures of Pb exposure and overall cancer incidence. These studies are limited by their
19 ecologic and cross-sectional designs. Absalon and Slesak (2010) and Obhodas (2007) did
20 not collect biological measurements, and no control for potential confounding was
21 mentioned.
5.10.1.3 Lung Cancer
22 Most of the recent evidence regarding lung cancer incidence is provided by a few studies
23 of occupationally-exposed adults. These are described in Table 5-49. Some studies in the
24 2006 Pb AQCD (U.S. EPA. 2006b) reported associations between Pb exposure and lung
25 cancer in occupational cohorts, although the studies were limited due to possible
26 confounding by smoking or other workplace exposures. In a more recently published
27 study of smelter workers (N= 187, N for lung cancer=46), no association was observed
28 between several metrics of Pb exposure (peak blood Pb values, number of years Pb
29 samples were obtained, and cumulative blood Pb index) and lung cancer incidence and
30 mortality combined (Lundstrom et al.. 2006). The median follow-up in the study was
31 about 30 years, and the median peak blood Pb values during employment were
November 2012 5-677 Draft - Do Not Cite or Quote
-------
1 49.7 (ig/dL for lung cancer cases and 55.9 (ig/dL for controls. In a study of 1,462 tin
2 smelter workers, no association was observed between Pb exposure and lung cancer
3 mortality in unweighted analyses, but when the analyses were weighted by age and time
4 since exposure, positive associations were apparent (Jones et al.. 2007). In this study,
5 cumulative Pb exposure was calculated by combining historical air sampling data and
6 personnel record cards, which specified work histories. The median cumulative Pb
7 exposure was estimated to be approximately y 2 mg/m3«yr. It is important to note that the
8 tin smelter workers were exposed to other metals as well, such as As and antimony and
9 the study did not specify if additional potential confounders were evaluated (beyond the
10 weighting for age and time since exposure). A population-based case-control study
11 performed among men in Montreal, Canada in the 1980s assessed occupational Pb
12 exposure via interviews regarding job histories and determined the likely Pb exposures
13 associated with the job activities (Rousseau et al.. 2007). No association was apparent
14 between organic Pb, inorganic Pb, or Pb from gasoline emissions and lung cancer (N
15 ranged from 271 to 1,203 depending on the exposure of interest).
16 Studies were also conducted that compared lung tissue Pb measurements for individuals
17 with lung cancer to those without lung cancer. The controls for these studies were
18 individuals with metastases in the lung from other primary cancers (De Palmaet al..
19 2008) and individuals with non-cancerous lung diseases (De Palma et al., 2008; Kuo et
20 al.. 2006). Limitations in these studies include their cross-sectional; design, the
21 measurement of Pb in cancerous tissue, which may have altered Pb distribution, and the
22 use of controls with other cancers and lung diseases. Findings are mixed among the
23 studies. De Palma et al. (2008) reported higher Pb concentrations in the cancerous and
24 non-cancerous lung tissue of individuals with non-small cell lung cancer compared to
25 control groups, although the authors report these results may be confounded by smoking.
26 Kuo et al. (2006) found no statistical difference in Pb levels for lung tissue of individuals
27 with lung cancer compared to controls.
28 Some studies in the 2006 Pb AQCD reported associations between Pb exposure and lung
29 cancer among occupational cohorts; however, recent epidemiologic studies of lung
30 cancer reported no associations. Overall, these recent epidemiologic studies included only
31 men, limiting the generalizability. The studies by Jones et al. (2007) and Rousseau et al.
32 (2007) also have the disadvantage of not obtaining actual measures of Pb exposure or
33 biomarker levels. In addition, these studies, as well as those in the 2006 Pb AQCD, are of
34 occupational cohorts, and the relationships with Pb exposures may be confounded by
35 other workplace exposures and covariates that were not considered, such as smoking.
November 2012 5-678 Draft - Do Not Cite or Quote
-------
5.10.1.4 Brain Cancer
1 A few studies of brain cancer examined the association between cancer and occupational
2 Pb exposure using exposures determined via exposure databases and patient interviews
3 about past jobs and known exposures (Table 5-49). The National Longitudinal Mortality
4 Study, a study that included a national sample of the U.S. population (N=317,968),
5 estimated occupational Pb exposure based on current/most recent employment among
6 individuals (van Wijngaarden and Dosemeci. 2006). Although not all estimates using
7 various statistical techniques and measures of Pb exposure/intensity are statistically
8 significant, a pattern of increased associations between Pb exposure and brain cancer
9 mortality was observed in the study population. In a case-control study of brain tumors
10 (N for controls=799, N for glioma=489, n for meningioma =197), glioma was reported to
11 have no association with any Pb exposure metric. However, positive associations were
12 observed between high cumulative occupational Pb exposure and meningioma among
13 individuals with ALAD2 genotypes (OR 2.4 [95% CI: 0.7, 8.8] comparing individuals
14 ever exposed to Pb with those not exposed to Pb; OR 12.8 [95% CI: 1.4, 120.8]
15 comparing individuals with cumulative Pb exposure > 100 (ig-year/m3 to those not
16 exposed to Pb) (Rajaraman et al.. 2006). This association was not present among
17 individuals with the ALAD1 genotypes (OR 0.5 [95% CI: 0.3, 1.0] comparing individuals
18 ever exposed to Pb with those not exposed to Pb; OR 0.7 [95% CI: 0.2, 1.8] comparing
19 individuals with cumulative Pb exposure > 100 (ig-year/m3 to those not exposed to Pb).
20 Another study of the association between occupational Pb exposure (measured using self-
21 reported occupational exposure history) and brain tumors reported none or slight overall
22 associations with types of brain tumors; however, positive associations were observed
23 among individuals with certain genetic single nucleotide polymorphisms (SNPs) (N for
24 controls=494, N for glioma=362, n for meningioma =134) (Bhatti et al.. 2009). After
25 control for multiple comparisons, individuals with GPX1 variants (rs 105 0450) had
26 positive associations between cumulative Pb exposure and glioblastoma multiforme and
27 meningioma. Individuals without RAC2 variants (rs2239774) showed a positive
28 association between Pb and glioblastoma multiforme. Also, individuals withoutXDH
29 variants (rs75 74920) displayed a positive association between Pb and meningioma.
30 Overall, associations between occupational Pb exposure and brain cancer incidence and
31 mortality were found to vary according to several genetic variants. Studies of the
32 association between Pb exposure and brain cancer were not reported in the
33 2006 Pb AQCD. These studies were limited in their methods because they do not have
34 individual level biological or exposure Pb measurements and the potential for
35 confounding by other workplace exposures exist. Additional research is needed to
36 characterize these associations and the modification by various genetic variants.
November 2012 5-679 Draft - Do Not Cite or Quote
-------
5.10.1.5 Breast Cancer
1 The association between proximity to a Pb smelter and breast cancer was evaluated in a
2 non-occupational cohort. A population-based case-control study in Canada (N for
3 controls=2,467, N for cases=2,343) examined the proximity to a Pb smelter based on
4 residential addresses (Pan etal. 2011) (Table 5-49). No association was reported
5 between proximity of a Pb smelter and breast cancer incidence, but the study was limited
6 by the small number of women who resided near a Pb smelter (n=13 lived < 3.2 km from
7 Pb smelter). No biological samples to determine Pb levels in the body were used in the
8 study, nor were Pb biomarker or exposure data were available.
9 A few case-control studies examined Pb levels in biological samples among individuals
10 with and without breast tumor and/or cancer. A study of newly diagnosed breast cancer
11 patients and controls examined Pb levels in blood and hair samples and reported higher
12 Pb levels in both for cancer cases, although the difference in the Pb content in hair
13 samples was not statistically significant (Alatise and Schrauzer. 2010). Siddiqui et al.
14 (2006) observed higher blood Pb levels in women with benign and malignant tumors
15 compared to controls. Additionally, although blood Pb levels were higher among those
16 with malignant breast tumors compared to those with benign tumors, both had similar
17 levels of Pb detected in breast tissues. Another study of Pb levels present in breast tissue
18 reported no statistical difference in Pb levels (Pasha et al., 2008b). However, a study of
19 breast tissue did observe a statistically significant difference between Pb levels in the
20 breast tissue of cancer cases and controls (lonescu et al.. 2007). Finally, a study of Pb
21 levels in urine reported a positive association between urine Pb and breast cancer, but this
22 association became null when women taking nonsteroidal aromatase inhibitors but not
23 taking bisphostphonates (a combination responsible for bone loss) were excluded from
24 the analysis (McElroy et al.. 2008).
25 The 2006 Pb AQCD (U.S. EPA. 2006b) did not report any studies examining Pb levels
26 and breast cancer. Overall, recent studies suggest that women with breast cancer may
27 have higher blood Pb levels than those without breast cancer. However, results are mixed
28 in studies that compared breast tissue Pb concentrations between breast tumor and control
29 samples. These studies are limited by their study design. The samples are taken after
30 cancer is already present in the cases, thus, the directionality between tissue or blood Pb
31 levels and cancer development cannot be established. Additionally, the sample sizes are
32 often small and the studies may be underpowered (most of the studies had less than 25
33 cases (Alatise and Schrauzer. 2010; lonescu et al.. 2007; Siddiqui et al., 2006). A case-
34 control study, also limited by its method of exposure measurement, reported no
35 association between living near a Pb smelter and breast cancer (Pan et al., 2011).
November 2012 5-680 Draft - Do Not Cite or Quote
-------
5.10.1.6 Other Cancers
1 There have been a few studies of cancer types other than those listed above. The
2 2006 Pb AQCD reported evidence of an association between Pb exposure and stomach
3 cancer in several occupational cohorts. A study performed among men in Montreal,
4 Canada in the 1980s evaluated multiple cancer outcomes and estimated occupational
5 exposures to organic Pb, inorganic Pb, and Pb from gasoline emissions via interviews
6 regarding job histories and subsequent exposure approximations by chemists and
7 hygienists (N for cases and controls varied from 60 to 2,250 based on the cancer type and
8 exposure) (Rousseau et al.. 2007). Adults with occupational exposure to organic Pb
9 exposure had greater odds of having stomach cancer compared to adults without
10 occupational exposure to organic Pb. A positive association was also observed for rectal
11 cancer when population-based controls were used but was null when the control
12 population was limited to individuals with other types of cancers. No association was
13 detected for cancers of the esophagus, colon, pancreas, prostate, bladder, kidney,
14 melanoma, or non-Hodgkin's lymphoma. None of the cancers were associated with
15 occupational exposure to inorganic Pb. When occupational exposure to Pb in gasoline
16 emissions was categorized as "unexposed," "nonsubstantial level," or "substantial level,"
17 a positive association with stomach cancer was observed when cancer controls were used
18 as the comparison group; however, the association was not present when population
19 controls were utilized as the control group). Another case-control study using participant
20 interviews and a job exposure matrix, including only men, (N for controls=285, N for
21 cancer=185) reported no association between occupational Pb exposure and esophageal
22 squamous cell carcinomas, but an association was present between high occupational Pb
23 exposure and adenocarcinoma of the esophagus (Santibanez et al.. 2008). However,
24 neither of these studies quantified Pb levels in biological or exposure samples.
25 Several studies compared Pb levels in blood, tissue, and urine of individuals who have
26 cancer with Pb-levels in individuals who are cancer-free. Compared to control groups,
27 higher Pb levels were observed in the blood and bladder tissue of individuals with
28 bladder cancer (Golabek et al.. 2009). the kidney tissue of individuals with renal cell
29 carcinoma (with highest levels among those with the highest stage tumors) (Calvo et al..
30 2009). the tissue (but not serum) of individuals with laryngeal cancer (Olszewski et al..
31 2006). the blood of individuals with gastric cancer (Khorasani et al.. 2008). the plasma
32 and hair of individuals with gastrointestinal cancer (Pasha etal.. 2010). the blood and hair
33 of individuals with non-specified types of cancer (Pasha et al.. 2008c: Pasha et al.. 2007).
34 and the hair of individuals with benign tumors (Pasha et al.. 2008a). No statistical
35 difference in Pb levels was reported for colon tissue of individuals with colorectal polyps
36 (Alimonti et al.. 2008) or urine of individuals with bladder cancer (Lin et al.. 2009)
37 compared to control groups. A study examining Pb levels in kidney tissue reported the
November 2012 5-681 Draft - Do Not Cite or Quote
-------
1 highest levels of Pb in normal kidney tissue samples that were adjacent to neoplastic
2 tumors. The Pb levels reported in the kidney tissue of neoplastic tumors were elevated
3 compared to those detected in corpses without neoplastic tumors of the kidney (Cerulli et
4 al.. 2006). All of these comparison studies are limited by the inability to determine
5 temporality as Pb biomarkers were measured after the cancer diagnosis; the level of Pb
6 may be due to changes that result from having cancer, not changes that result in cancer.
7 Many of these studies attempted to control for this by including only cases who have not
8 undergone certain treatments. Additionally, studies are limited by their small sample
9 sizes and the selection of the control populations. Control populations are supposed to
10 represent the general population from which the cases are drawn; some of the control
11 subjects in these studies are individuals with diseases/conditions warranting tissue
12 resections, which are not prevalent in the general population.
13 In summary, epidemiologic studies examining the potential for associations of Pb
14 exposure with the incidence of specific cancers reported varying associations with
15 occupational Pb exposure. Associations were null for occupational Pb exposure and most
16 cancer sites examined. However, a positive association was observed between
17 occupational Pb exposure and adenocarcinoma of the esophagus as well as exposure to
18 occupational organic Pb and stomach cancer, which is supported by evidence of a
19 relationship between Pb exposure and stomach cancer in occupational cohorts reported in
20 the 2006 Pb AQCD. Associations between occupational organic Pb exposure and rectal
21 cancer and occupational exposure to Pb in gasoline emissions and stomach cancer were
22 inconsistent. These studies of various cancer sites have limited generalizability due to the
23 study populations comprising only men. In addition, there are no personal biological or
24 exposure samples used in the epidemiologic analyses and confounding by other
25 occupational exposures is possible. In other studies, biological samples were used in
26 biomarker comparisons of cancer and cancer-free individuals but as stated above, these
27 studies have multiple other limitations.
5.10.1.7 Animal Models of Carcinogenicity
28 Previous AQCDs have established that Pb has been shown to act as a carcinogen in
29 animal toxicology models, albeit at relatively high concentrations. Chronic oral
30 Pb acetate exposure to male and female rodents has consistently been shown to be a
31 kidney carcinogen in multiple separate studies, inducing adenocarcinomas and adenomas
32 after chronic exposure. Gliomas of the brain have also been reported after oral Pb
33 exposure. The kidneys are the most common target of Pb-dependent carcinogenicity
34 (Kasprzak et al.. 1985; Roller etal.. 1985; Azaretal.. 1973; Van Esch and Kroes. 1969)
35 but the testes, brain, adrenals, prostate, pituitary, and mammary gland have also been
November 2012 5-682 Draft - Do Not Cite or Quote
-------
1 affected (IARC. 2006a). The typical cancer bioassays used by IARC or NTP as evidence
2 of Pb-induced carcinogenicity were designed using rodents, typically males but
3 sometimes animals of both sexes, that were continuously exposed to Pb acetate in chow
4 (i.e., 1,000 or 10,000 ppm Pb acetate) or drinking water (i.e., 26 or 2,600 ppm Pb acetate)
5 for 18 months to two years in duration, the typical life span of a rodent (Kasprzak et al.
6 1985: Roller etal.. 1985: Azaretal. 1973: Van Esch and Kroes. 1969). These two-year
7 cancer bioassays and the doses employed are typical of cancer bioassays employed for
8 other chemicals, albeit at doses that are higher than Pb doses cited in other toxicological
9 sections of the ISA. In cancer bioassays, to obtain statistically valid data from small
10 groups of animals, doses are selected such that any dose-related effects will occur
11 frequently enough to be detected. The 2006 Pb AQCD (U.S. EPA. 2006b) pointed out
12 that because Pb is a "well-established animal carcinogen...., focus has been more on the
13 mechanism of neoplasia and possible immunomodulatory effects of Pb in the promotion
14 of cancer." This focus continues to date. More recent studies have focused on
15 administration of Pb with known carcinogens or antioxidants such that lifestage, diet, and
16 mode of action can be better understood. Developmental Pb acetate exposure also
17 induced kidney tumors in offspring whose dams received Pb acetate in drinking water
18 during pregnancy and lactation.
19 Recognition of the importance of windows of exposure in Pb-induced cancer bioassays is
20 a focus of more recent studies. In one study, gestational and lactational exposure of
21 laboratory rodents to inorganic Pb (500, 750 or 1,000 ppm Pb acetate in drinking water)
22 induced carcinogenicity in adult offspring (Waalkes etal.. 1995). Another recent study
23 considered Pb-induced carcinogenesis in laboratory animals with early life Pb exposure
24 (gestation and lactation) in which Tokar et al. (2010) examined tumorigenesis in
25 homozygous metallothionein I/II knockout mice and their corresponding wild type
26 controls (groups often mice each). The dams/mothers were exposed by drinking water to
27 2,000 or 4,000 ppm Pb acetate during gestation and lactation and compared to untreated
28 controls. Study animals were exposed in utero, through birth and lactation, and then
29 postnatally to drinking water until 8 weeks old. The Pb-exposed metallothionein I/II
30 knockout mice had increased testicular teratomas and renal and urinary bladder
31 preneoplasia. The tumor burden of Pb-exposed wild-type mice were not statistically
32 significantly different than controls. The data suggest that metallothionein can protect
33 against Pb-induced tumorigenesis. Concerns with the study are that the doses are at levels
34 of Pb to which humans would not likely be exposed and there is no metallothionein null
35 condition in humans, though there is variability in the expression of metallothionein. The
36 data do not address whether this variability would have any impact on Pb-induced
37 carcinogenesis in humans. Thus, the animal toxicology data demonstrate that Pb is a
38 well-established animal carcinogen in studies employing high-dose Pb exposure over a
39 continuous extended duration of exposure (i.e., 2 years), which is typical of cancer
November 2012 5-683 Draft - Do Not Cite or Quote
-------
1 bioassays. Newer studies are showing early-life maternal Pb exposure can contribute to
2 carcinogenicity in offspring and have shown that metallothionein is protective against
3 cancer in this pathway.
5.10.2 Cancer Biomarkers
4 A cross-sectional study of men aged 21-40 years without occupational history of
5 exposure to metals examined prostate specific antigen (PSA), a biomarker for prostate
6 cancer (N=57). Studies of Pb exposure and PSA were not reported in the 2006 Pb AQCD
7 (U.S. EPA. 2006b). This recent study reported a positive association between Pb levels
8 and PSA levels (measured in the same blood samples) in regression models adjusted for
9 the following potential confounders: age, smoking, alcohol consumption, and other
10 metals (Cd, Zn, Se, and Cu) (Pizent et al., 2009). The median concurrent blood Pb level
11 was 2.6 (ig/dL (range 1.0-10.8 (ig/dL). The authors note that the study population was
12 young and at lower risk of prostate cancer than are older men.
5.10.3 Modes of Action for Pb-induced Carcinogenicity
13 The carcinogenic mode of action of Pb is poorly understood. It is unclear whether the
14 mode of action of Pb is best understood within the framework of multistage
15 carcinogenesis, genomic instability or epigenetic modification. For example, multistage
16 carcinogenesis involves a series of cellular and molecular changes that result from the
17 progressive accumulation of mutations that induce alterations in cancer-related genes. Pb
18 does not appear to follow this paradigm, and the literature suggests it is weakly
19 mutagenic. Pb does appear to have some ability to induce DNA damage
20 (Section 5.10.3.2). However, the ability of Pb to alter gene expression through epigenetic
21 mechanism (Section 5.10.3.3) and to interact with proteins may be a means by which Pb
22 induces carcinogenicity. It is known that Pb can replace Zn in Zn-binding (Zn-finger)
23 proteins (Section 5.2). which include hormone receptors, cell-cycle regulatory proteins,
24 the Ah receptor, estrogen receptor, p53, DNA repair proteins, protamines, and histones.
25 These Zn-finger proteins all bind to specific recognition elements in DNA. Thus, Pb may
26 act at a post-translational stage to alter protein structure of Zn-finger proteins, which can
27 in turn alter gene expression, DNA repair and other cellular functions. To recapitulate,
28 cancer develops from one or a combination of multiple mechanisms including
29 modification of DNA via epigenetics or enzyme dysfunction and genetic instability or
30 mutation. These modifications then provide the cancer cells with a selective growth
31 advantage. In this schematic, Pb may contribute to epigenetic changes and chromosomal
32 aberrations.
November 2012 5-684 Draft - Do Not Cite or Quote
-------
1 The genomic instability paradigm requires a cascade of genome-wide changes caused by
2 impaired DNA repair, kinetochore assembly, cellular checkpoints, centrosome
3 duplication, microtubule dynamics or a number of cell maintenance processes. These
4 processes have been rarely studied for Pb, thus there are few data that suggest Pb may
5 interfere with some of these processes. Furthermore, the bulk of the literature in this area
6 involves Pb chromate and it is unclear if the effects are due to Pb or chromate. Epigenetic
7 modifications may lead to cancer by altering cellular functions without altering the DNA
8 sequence. The most commonly studied epigenetic change is methylation alterations. A
9 small number of studies show that Pb can induce epigenetic changes (Section 5.10.3.3).
10 but studies are still missing to clearly tie these effects to Pb-induced carcinogenesis and
11 genotoxicity. Thus, either genomic instability or epigenetic modification paradigms or
12 some combination of the two may underlie Pb-induced carcinogenicity.
13 Exposure to mixtures can also contribute to understanding of modes of action. No recent
14 studies of the protective role of Ca2+ or Zn in Pb-induced carcinogenesis or genotoxicity
15 were found. Pb can displace these and other divalent cations, affecting physiological
16 processes. There were some data suggesting that metallothionein (Section 5.10.4). which
17 sequesters Pb and makes it less bioavailable, protects rodents from Pb-induced cancers.
18 Boron, melatonin, N-acetylcysteine, turmeric and myrrh protected cells against
19 Pb-induced genotoxicity (Section 5.10.3.2) and affected antioxidant status, especially the
20 glutathione pathway. There were some data suggesting that Pb mimics or antagonizes the
21 essential micronutrient Se in rodents. These data are discussed in more detail elsewhere
22 (Section 5.10.4) and point to the relevance of mixtures in assessing toxicity.
5.10.3.1 Neoplastic Transformation Studies, Human Cell Cultures
23 Carcinogenesis can be measured in cell culture systems through neoplastic transformation
24 models that monitor change by following morphological transformation of cells,
25 i.e., formation of a focus (or foci) of cell growth. Xie et al. (2007) treated BEP2D cells
26 (human papilloma virus- immortalized human bronchial cells) with 0, 1, 5, or 10 (ig/cm2
27 PbCrO4 for 120 hours. PbCrO4 induced foci formation in a concentration-dependent
28 manner. Xie et al. (2008) treated BJhTERT cells (hTERT-immortalized human skin
29 fibroblasts) and ATLD-2 cells (hTERT-immortalized human skin fibroblasts deficient in
30 Mrel 1) with 0, 0.1, 0.5, and 1 (ig/cm2 PbCrO4 for 120 hours. PbCrO4 induced foci
31 formation in a concentration-dependent manner in the Mre 11 deficient cells. Mre 11 was
32 required to prevent PbCrO4-induced neoplastic transformation.
November 2012 5-685 Draft - Do Not Cite or Quote
-------
Immune Modulation of Tumorigenesis by Pb
1 As described in the 2006 Pb AQCD (U.S. EPA. 2006b). Pb-induced immunotoxicity can
2 contribute to increased risk of cancer, primarily as the result of suppressed Thl responses
3 and misregulated inflammation. First, Pb-induced misregulation of inflammation
4 involving innate immune cells has been shown to result in chronic insult to tissues. These
5 insults, excessive lipid and DNA oxidation production by overproduction of ROS and
6 weakened antioxidant defenses, can increase the likelihood of mutagenesis, cellular
7 instability, and tumor cell formation. For example, results from Xu et al. (2008) support
8 the association with Pb exposure and DNA damage, and investigators concluded that it is
9 a possible route to increased Pb-induced tumorigenesis. The second component of
10 increased risk of cancer involves Pb-induced suppression of Thl-dependent anti-tumor
11 immunity as acquired immunity shifts statistically significantly toward Th2 responses.
12 With cytotoxic T lymphocytes and other cell-mediated defenses dramatically lessened,
13 the capacity to resist cancer may be compromised.
5.10.3.2 DNA and Cellular Damage
14 Multiple studies have been performed examining the relationship between Pb and DNA
15 and cellular damage. Details of the recent epidemiologic and toxicological studies follow.
Epidemiologic Evidence for DNA and Cellular Damage
16 Multiple studies examined the relationship between Pb and sister chromatid exchange
17 (SCE). SCEs are exchanges of homologous DNA material between chromatids on a
18 chromosome and are a test for mutagenicity or DNA damage. A study of male policemen
19 reported mean blood Pb levels for the study population of 43.5 (ig/dL (Wiwanitkit et al.,
20 2008). In analyses dichotomized as high or low blood Pb levels (cut-off at 49.7 (ig/dL),
21 the higher blood Pb group was observed to have higher mean SCE. Another study of
22 adult males compared the SCE of storage battery manufacturing workers (mean blood Pb
23 levels of 40.14 (ig/dL) and office workers (mean blood Pb levels of 9.11 (ig/dL) (Duydu
24 et al.. 2005). The exposed workers had higher SCE levels and also a greater number of
25 cells in which the SCEs per cell were higher than the 95th percentile of the population.
26 Finally, a study of children aged 5-14 years old (mean [SD] blood Pb levels of 7.69
27 [4.29] (ig/dL) reported no correlation between blood Pb levels and SCE (Mielzvnska et
28 al.. 2006). However, the study did report a positive association between blood Pb and
29 micronuclei (MN) levels.
November 2012 5-686 Draft - Do Not Cite or Quote
-------
1 Other studies of DNA damage have reported mixed results. A study of children ages 6-11
2 years old and environmentally-exposed to Pb reported no association between blood Pb
3 and baseline DNA damage or repair ability after a peroxide challenge (children attending
4 a school far from a Pb smelter: median blood Pb level 4.6 (ig/dL; children attending a
5 school near a Pb smelter: median blood Pb level 28.6 (ig/dL) (Mendez-Gomez et al.
6 2008). Another study included adult participants aged 50-65 years and reported an
7 association between blood Pb and carcinoembryonic antigen (CEA) but not with DNA-
8 strand breaks, MN frequency, or oxidative DNA damage (median blood Pb level of the
9 study population: 3.92 (ig/dL) (De Coster et al., 2008). A study conducted among
10 workers exposed to Pb (mean blood Pb level: 30.3 (ig/dL) and unexposed controls (mean
11 blood Pb level: 3.2 (ig/dL) reported greater cytogenetic damage (measured by MN
12 frequency), chromosomal aberrations, and DNA damage in the Pb-exposed group
13 (although this was not statistically significant in linear regression models controlling for
14 age) (Grover et al., 2010). A study of painters in India, where Pb concentrations in paint
15 are high, reported a mean (SD) blood Pb level of 21.56 (6.43) (ig/dL among painters who
16 reported painting houses for 8-9 hours/day for 5-10 years (Khan et al., 2010b); the mean
17 (SD) blood Pb level was 2.84 (0.96) (ig/dL for healthy workers who had not been
18 occupationally exposed to Pb. Cytogenetic damage was higher among the painters
19 compared to the healthy controls. Another study compared the blood Pb of metal workers
20 and office workers and reported higher blood Pb levels (both current and 2 year average)
21 among the metal workers (blood Pb level > 20 (ig/dL) compared to the office workers
22 (blood Pb level <10 (ig/dL) (Olewinska et al., 2010). Overall, the workers had increased
23 DNA strand breaks versus the office workers (this held true at various blood Pb levels).
24 Finally, a study of Pb battery workers with symptoms of Pb toxicity and a group of
25 controls were examined (Shaik and Jamil. 2009). Higher chromosomal aberrations, MN
26 frequency, and DNA damage were reported for the battery workers as compared to the
27 controls. These workplace studies are limited by the lack of consideration for potential
28 confounding factors, including other occupational exposures.
lexicological Evidence for DNA and Cellular Damage
Sister Chromatic/ Exchanges
29 Pb has been shown to induce SCEs both in vivo and in vitro. Tapisso et al. (2009).
30 considered SCEs in adult Algerian mice (groups of six mice each) that were treated by
31 i.p. injection with 5 or 10 doses of 0.46 mg/kg Pb acetate. The SCE in bone marrow were
32 elevated after Pb exposure alone and increased with time. Co-exposure with Cd or Zn
33 further increased SCE levels.
November 2012 5-687 Draft - Do Not Cite or Quote
-------
1 SCE was also followed in cultured human cells. Ustundag and Duydu (2007) considered
2 the ability of N-acetylcysteine and melatonin to reduce Pb nitrate-induced SCE in a
3 single human donor. Cells were treated with 0, 1, 5, 10, or 50 (iM Pb nitrate. SCE
4 statistically significantly increased at every Pb concentration in a concentration
5 dependent manner. Both 1 and 2 mM N-acetylcysteine and melatonin were able to
6 statistically significantly reduce SCE levels in Pb-exposed cells. In another study, Turkez
7 et al. (2011) considered the ability of boron compounds, essential micronutrients, to
8 prevent Pb chloride-induced SCE in human lymphocytes. Cells were obtained from 4
9 non-smoking donors. Both 3 and 5 ppm Pb chloride induced a statistically significant
10 increase in SCE levels over controls. Boron was able to statistically significantly
11 diminish these levels. For both studies, exposure times were not provided, and the full
12 interpretation of these data is limited by the limited number of donors and the absence of
13 an exposure time for the SCE assay.
Micronuclei Formation
14 The 2006 Pb AQCD stated "studies of genotoxicity consistently find associations of Pb
15 exposure with DNA damage and MN formation" and recent studies continue to report
16 these associations. Alghazal et al. (2008b) considered the ability of Pb acetate trihydrate
17 to induce MN in bone marrow of adult Wistar rats. Animals were given a daily dose of
18 100 mg/L in their drinking water for 125 days. The mean number of MN in male and
19 female rats was statistically significantly higher in Pb-exposed animals than in unexposed
20 controls. Tapisso et al. (2009) considered Pb-induced MN in rodents. Algerian mice were
21 treated by i.p. injection with 5 or 10 doses of 0.46 mg/kg Pb acetate and compared to
22 untreated controls. The MN in bone marrow were elevated after Pb exposure and
23 increased with time
24 MN formation has also been followed in cultured human cells. Ustundag and Duydu
25 (2007) considered the ability of N-acetylcysteine and melatonin to reduce Pb nitrate-
26 induced MN in a single human donor. Cells were treated with 0, 1,5, 10, or 50 (iM
27 Pb nitrate. MN formation statistically significantly increased at the two highest Pb
28 concentrations in a concentration-dependent manner. Both 1 and 2 mM N-acetylcysteine
29 and melatonin were not able to statistically significantly reduce MN levels. In another
30 study, Turkez et al. considered the ability of boron compounds to prevent Pb chloride-
31 induced MN in human lymphocytes. Cells were obtained from 4 non-smoking donors.
32 Both 3 and 5 ppm Pb chloride induced a statistically significant increase in MN levels
33 over controls. Boron induced a statistically significant attenuation of these Pb-induced
34 levels. For both studies, exposure times were not provided, and the full interpretation of
35 these data is limited by the limited number of donors and the absence of an exposure time
36 for the MN assay. Gastaldo et al. (2007) evaluated the ability of Pb to induce MN.
November 2012 5-688 Draft - Do Not Cite or Quote
-------
1 Human endothelial HMEC cell line was treated with 1-1,000 (iM Pb nitrate for 24 hours.
2 MN increased in a statistically significant, concentration-dependent manner.
Hypoxanthine-guanine phosphoribosyltransferase Mutations
3 The potential mutagenicity of Pb in human or animal cells has been evaluated by
4 monitoring mutations at the hypoxanthine-guanine phosphoribosytransferase (HPRT)
5 locus. Li et al. (2008a) evaluated Pb acetate-induced HPRT in the non-small-cell lung
6 carcinoma tumor cell line, CL3, and in normal human diploid fibroblasts (specific tissue
7 source not reported). All cells were exposed to 0, 100, 300 or 500 (iM Pb acetate for 24
8 hours in serum-free medium ± a 1-hour pretreatment with a MKK1/2 inhibitor or a
9 PKC-alpha inhibitor. Pb alone did not induce HPRT mutations. Inhibiting the ERK
10 pathway via either inhibitor statistically significantly increased Pb-induced mutagenesis.
11 Wang et al. (2008c_), investigated Pb acetate -induced HPRT mutations in CL3 cells. All
12 cells were exposed to 0, 100, 300 or 500 (iM Pb acetate for 24 hours in serum-free
13 medium ± a 1-hour pretreatment with a PKC-alpha inhibitor or siRNA for PKC-alpha. Pb
14 alone did not induce HPRT mutations. Inhibiting PKC-alpha via either inhibitor
15 statistically significantly increased Pb-induced mutagenesis. McNeill et al. (2007)
16 examined Pb acetate induced HPRT mutations in Chinese hamster ovary AA8 cells and
17 AA8 cells overexpressing human Apel. Cells were treated with 5 (iM Pb acetate for 6
18 hours. No increases in HPRT mutations were observed after Pb exposure in either cell
19 line but with specific pathway perturbations (PKC-alpha or ERK), Pb was able to induce
20 HPRT mutations.
Chromosomal Aberrations
21 Chromosomal aberrations, an indicator of cancer risk, were followed in Pb-exposed
22 rodents (El-Ashmawv et al.. 2006). Dietary exposure to Pb acetate administered as a
23 single dose of 5,000 ppm w/w to adult male Swiss albino mice caused statistically
24 significant increased levels of chromosomal aberrations in the Pb treatment alone group,
25 particularly with respect to fragments, deletions, ring chromosomes, gaps, and end-to-end
26 associations. In addition, the authors found turmeric and myrrh powders were protective.
27 Concerns with the study include the use of only a single dose of Pb acetate along with the
28 high levels of unusual aberrations such as ring chromosomes and end-to-end associations.
29 Typically, these aberrations are rare after metal exposure, but were the most commonly
30 observed aberration in this study raising questions about the quality of the metaphase
31 preparations. An additional concern was that only 50 metaphases per dose were analyzed
32 instead of the more common 100 metaphases per dose. The authors did not explain why
33 their spectrum of aberrations was so different, why they only used one dose, or analyzed
34 fewer metaphases per dose.
November 2012 5-689 Draft - Do Not Cite or Quote
-------
1 Multiple studies considered the ability of Pb to induce chromosomal aberrations in
2 cultured human cells. The ability of Pb nitrate to induce chromosomal aberrations was
3 examined in primary human peripheral blood lymphocytes obtained from healthy,
4 nonsmoking donors (Pasha Shaik et al.. 2006). Cells were treated with 0, 1.2 or 2 mM
5 Pb nitrate for 2 hours. No increase in chromosomal aberrations was reported. Some
6 aneuploidy was observed. Concerns with the study are that only a 2-hour exposure was
7 used, which may not be long enough for DNA damage to be expressed as a chromosomal
8 aberration. It also appears from the data presentation that only three subjects were used;
9 one for a control, one for the low dose and one for the high dose. Experiments were not
10 repeated, thus given the small number of subjects, this study may not have had sufficient
11 power to detect any effects. Holmes et al. (2QQ6a), treated WHTBF-6 cells (hTERT-
12 immortalized human lung cells) with 0, 0.1, 0.5, or 1 (ig/cm2 Pb chromate for 24-120
13 hours or with 0, 0.1, 0.5, 1, 5 or 10 (ig/cm2 Pb oxide for 24 or 120 hours. Pb chromate
14 induced statistically significant, concentration-dependent increases in centrosome
15 abnormalities and aneuploidy. Wise et al. (2006a) treated BEP2D cells with 0, 0.5, 1, 5,
16 or 10 (ig/cm2 Pb chromate for 24 hours. Pb chromate induced statistically significant
17 concentration-dependent increases in chromosomal aberrations. Holmes et al. (2006b),
18 treated WHTBF-6 cells with 0, 0.1, 0.5, or 1 (ig/cm2 Pb chromate for 24-72 hours.
19 Pb chromate induced statistically significant, concentration-dependent increases in
20 chromosomal aberrations. The effects of the chromate anion cannot be ruled out as
21 causative in inducing these chromosomal aberrations. Wise et al. (2006b), treated
22 WHTBF-6 cells with 0, 0.1, 0.5, or 1 (ig/cm2 Pb chromate for 24-120 hours. Pb chromate
23 induced statistically significant, concentration-dependent increases in spindle assembly
24 and checkpoint disruption, effects of mitosis and aneuploidy. By contrast, chromate-free
25 Pb oxide did not induce centrosome amplification. The effects were likely attributable to
26 the chromate anion. Xie et al. (2007) treated BEP2D cells with 0, 1, 5, or 10 (ig/cm2
27 Pb chromate for 24 hours. Pb chromate induced statistically significant, concentration-
28 dependent increases in chromosomal aberrations and aneuploidy. Wise et al. (2010)
29 treated WHTBF-6 cells with 0, 0.1, 0.5, or 1 (ig/cm2 Pb chromate for 24 hours in a study
30 comparing 4 chromate compounds. Pb chromate induced statistically significant,
31 concentration-dependent increases in chromosomal aberrations.
32 Multiple investigators considered the ability of Pb chromate to induce chromosome
33 aberrations in rodent cell cultures. Grlickova-Duzevik et al. (2006) treated Chinese
34 hamster ovary (CHO) cells with 0, 0.1, 0.5, or 1 (ig/cm2 Pb chromate for 24 hours.
35 Specific CHO lines used included AA8 (wildtype) EM9 (XRCC1-deficient), and H9T3
36 (EM9 complemented with human XRCC1 gene). Pb chromate induced statistically
37 significant, concentration-dependent increases in chromosomal aberrations that were
38 statistically significantly increased by XRCC1 deficiency. Nestmann and Zhang (2007)
39 treated Chinese hamster ovary cells (clone WB(L)) with 0, 0.1, 0.5, 1, 5, or 10 (ig/cm2
November 2012 5-690 Draft - Do Not Cite or Quote
-------
1 Pb chromate (as pigment yellow) for 18 hours. No increases in chromosomal aberrations
2 were observed. Savery et al. (2007) treated CHO cells with 0, 0.1, 0.5, 1, or 5 (ig/cm2
3 Pb chromate for 24 hours. Specific CHO lines used included AA8 (wildtype), KO40
4 (,Fa«cg-deficient), and 40BP6 (.Fawcg-complemented). The Fancg gene plays an
5 important role in cellular resistance to DNA interstrand crosslinks, protecting against
6 genetic instability. Pb chromate induced statistically significant, concentration-dependent
7 increases in chromosomal aberrations that were increased by ^a«cg-deficiency. Camyre
8 et al. (2007) treated CHO cells with 0, 0.1, 0.5, 1, 5, or 10 (ig/cm2 Pb chromate for 24
9 hours. Specific CHO lines used included CHO-K1 (parental), xrs-6 (Ku80 deficient), and
10 2E (xrs-6 complemented with Ku80 gene). Pb chromate induced statistically significant,
11 concentration-dependent increases in chromosomal aberrations that were not affected by
12 Ku80 deficiency. Ku80 is a gene involved in nonhomologous end-joining repair and its
13 absence can contribute to genetic instability. Stackpole et al. (2007) treated CHO and
14 Chinese hamster lung (CHL) cells with 0, 0.1, 0.5, or 1 (ig/cm2 Pb chromate for 24 hours.
15 Specific CHO lines used included AA8 (wildtype), irs 1SF (XRCC3-deficient), and
16 ISFwtS (XRCC3 complemented). XRCC3 is DNA repair enzyme involved in
17 homologous recombination. CHL lines used included V79 (wildtype), irs3 (Rad51C
18 deficient) and irs3#6 (Rad51C complemented). Rad51C is a gene that encodes strand-
19 transfer proteins that are thought to be involved in recombinational repair of damaged
20 DNA and in meiotic recombination. Pb chromate induced statistically significant,
21 concentration-dependent increases in chromosomal aberrations that were statistically
22 significantly increased by both XRCC3 and Rad51C deficiency.
23 Multiple studies considered the ability of Pb chromate to induce chromosome aberrations
24 in marine mammal cell cultures. Li Chen et al. (2009) treated primary North Atlantic
25 right whale lung and skin fibroblasts with 0, 0.5, 1.0, 2.0, and 4.0 (ig/cm2 Pb chromate for
26 24 hours. Wise et al. (2009) treated primary Steller sea lion lung fibroblasts with 0, 0.1,
27 0.5, 1 and 5 (ig/cm2 Pb chromate for 24 hours. Wise et al. (2011) treated primary sperm
28 whale skin fibroblasts with 0, 0.5, 1,3,5, and 10 (ig/cm2 Pb chromate for 24 hours. In all
29 three studies, Pb chromate induced statistically significant, concentration-dependent
30 increases in chromosomal aberrations.
31 In summary, exposure of various cell models and an in vivo model to Pb (acetate,
32 chromate, or nitrate) induced significant increases in chromosomal aberration that often
33 responded in a concentration dependent manner. The use of various cell lines deficient in
34 specific DNA repair enzymes helped to elucidate which pathways may be most sensitive
35 to Pb-dependent chromosomal aberration. However, a number of studies used
36 Pb chromate exposures and the effects of the chromate anion cannot be ruled out as
37 causative in inducing these chromosomal aberrations.
November 2012 5-691 Draft - Do Not Cite or Quote
-------
COMETAssay
I Multiple studies considered the ability of Pb to induce DNA single strand breaks in
2 laboratory animals and human and animal cells using the comet assays. The COMET
3 assay measures DNA damage assessed by single cell electrophoresis of a lysed cell and
4 measurement of the fragmented DNA or tail length. Xu et al. (2008) examined DNA
5 damage in male ICR mice treated with Pb acetate. Animals (5 per group) were given
6 Pb acetate by gavage at doses of 0, 10, 50, or 100 mg/kg body weight every other day for
7 4 weeks. Pb exposure statistically significantly increased both tail length and tail moment
8 in a dose-dependent manner. Nava-Hernandez et al. (2009) considered the ability of
9 Pb acetate to induce DNA damage in primary spermatocyte DNA of male Wistar rats.
10 Animals (3 per group) were treated for 13 weeks with 0, 250, or 500 mg/L Pb in their
11 drinking water. There was statistically significantly less DNA damage in the controls
12 compared to the two treatment groups. Narayana and Al-Bader (2011) examined DNA
13 damage in liver tissue of adult male Wistar rats exposed to Pb nitrate. Animals (8 per
14 group) were treated for 60 days with doses of 0, 5,000, or 10,000 ppm Pb nitrate in their
15 drinking water. There were no statistical differences between treated groups and controls.
16 Drosophila melanogaster larvae (72 hours old) exposed to Pb nitrate (2,000, 4,000, and
17 8,000 (iM in culture media for 24 hours) yielded haemocytes that tested positive in the
18 comet assay; Pb chloride (8,000 (iM) did not cause DNA damage with the comet assay
19 (Carmona et al.. 2011).
20 Other studies used the COMET assay in cultured human cells. Pasha Shaik et al. (2006)
21 treated primary human peripheral blood lymphocytes obtained from healthy, nonsmoking
22 donors with 0, 2.1, 2.4, 2.7, 3.0, 3.3* 103 (JVI Pb nitrate for 2 hours and found dose-
23 dependent increases in Comet tail length. Concerns with the study are that apparently no
24 negative control was used. It also appears from the data presentation that only five
25 subjects were used; one for each dose. Experiments were not repeated. Thus, given the
26 small number of subjects and the absence of a negative control, this study may only be
27 detecting background levels of DNA damage. Xie et al. (2008) treated BJhTERT cells
28 (hTERT-immortalized human skin fibroblasts) and ATLD-2 cells (hTERT-immortalized
29 human skin fibroblasts deficient in Mrel 1) with 0, 0.1, 0.5, and 1 (ig/cm2 Pb chromate for
30 24 hours. Mrel 1 is a component of the MRN complex and plays a role in telomere
31 maintenance and double-strand break repair. Pb chromate induced a concentration-
32 dependent increase in DNA double strand breaks measured by the comet assay.
33 Pb chromate exposure and the effects of the chromate anion cannot be ruled out as
34 causative in inducing these aberrations. In another study, Pb nitrate exposure (30 (ig/mL)
35 induced statistically significant increased DNA damage in human liver HepG2 cells that
36 was attenuated with co-exposure with the antioxidant NAC (500 \\M) (Yedjou et al..
37 2010).
November 2012 5-692 Draft - Do Not Cite or Quote
-------
1 Other studies used the comet assay to examine Pb-induced DNA single strand breaks in
2 rodent cell cultures. Xu et al. (2006). treated PC12 cells with 0, 0.1, 1 or 10 (iM
3 Pb acetate. Both tail length and tail moment statistically significantly increased in a
4 concentration-dependent manner. Kermani et al. (2008) exposed mouse bone marrow-
5 mesenchymal stem cells to 60 (iM Pb acetate for 48 hours. There was an increase in
6 several comet assay measurements including tail length.
7 The COMET assay showed multiple positive findings after Pb exposure in rodents, flies,
8 primary human cells, and cell lines. In vivo studies with rodents exposed to Pb acetate
9 yielded significant increases in tail length and moment via COMET assays in separate
10 studies that used lymphocytes and sperm. In drosophila, Pb nitrate but not Pb chloride
11 produced significant increases with the COMET assay. Human cell culture from primary
12 cells (lymphocytes) and from cell lines (fibroblasts and liver) produced positive COMET
13 assays with separate Pb nitrate and Pb chromate exposures. Thus, the COMET assay
14 showed multiple positive findings of DNA damage after in vitro and in vivo Pb exposure.
Other Indicators of DNA Damage
15 Other studies considered the ability of Pb to induce DNA double strand breaks by
16 measuring gamma-H2A.X foci formation in cultured human cells. Xie et al. (2008)
17 treated BJhTERT cells (hTERT-immortalized human skin fibroblasts) and ATLD-2 cells
18 (hTERT-immortalized human skin fibroblasts deficient in Mrel 1) with 0, 0.1, 0.5, and
19 1 (ig/cm2 Pb chromate for 24 hours. Pb chromate induced a concentration-dependent
20 increase in DNA double strand breaks measured by gamma-H2A.X foci formation.
21 Pb chromate exposure and the effects of the chromate anion cannot be ruled out as
22 contributory. Gastaldo et al. (2007) evaluated the ability of Pb to induce DNA double
23 strand breaks with both gamma-H2A.X foci formation and pulse-field gel electrophoresis
24 in cultured human cells. The human endothelial HMEC cell line was treated with 1 to
25 1,000 (iM Pb nitrate for 24 hours. DNA double strand breaks increased in a
26 concentration-dependent manner. Wise et al. (2010) treated WHTBF-6 cells with 0, 0.1,
27 0.5, or 1 (ig/cm2 Pb chromate for 24 hours in a study comparing four chromate
28 compounds. Pb chromate induced statistically significant, concentration-dependent
29 increases in DNA double strand breaks measured by gamma-H2A.X foci formation, at a
30 similar level to the three other compounds. A few studies demonstrated the ability of Pb
31 to destabilize DNA by forming DNA-histone cross links, which can lead to histone
32 aggregation (Rabbani-Chadegani et al.. 2011; Rabbani-Chadegani et al.. 2009). In
33 extracts of rat liver, Pb nitrate (<300 (iM) was shown to react with chromatin components
34 and induce chromatin aggregation via histone-DNA cross links.
35 Genotoxicity testing of Drosophila melanogaster larvae (72 hours old) using the Wing
36 Spot test showed that neither Pb chloride nor Pb nitrate (at concentrations of 2,000, 4,000
November 2012 5-693 Draft - Do Not Cite or Quote
-------
1 and 8,000 (iM in culture media with exposure until pupation) was able to induce
2 significant increases in the frequency of wing spots (Carmona et al.. 2011). The wing
3 spot test can detect mitotic recombination and multiple mutational events such as point
4 mutations, deletions, and certain types of chromosome aberrations (Graf and Wurgler.
5 1986). Further, wing spot assays employing Pb co-exposure with gamma radiation
6 showed no effect of Pb on gamma radiation induced spotting frequency.
7 Multiple studies examined the effects of Pb on DNA repair. Most were conducted in
8 cultured cells, and one was done in an animal model. El-Ghor et al. (2011) followed
9 microsatellite instability (MSI) in Pb acetate trihydrate exposed adult male rats. MSI
10 reflects impaired DNA mismatch repair and contributes to an increased risk of cancer.
11 DNA from leukocytes of adult male albino rats exposed to Pb acetate (acute: single oral
12 dose of 467 mg/kg BW or sub-chronic: 47 mg/kg BW six days/week for 4 week) showed
13 increased MSI at three microsatellite loci (D6mit3, D9mit2, and DISMghl). This study is
14 limited by its small sample size (n=2 to 3 rodents per treatment group). Li et al. (2008a)
15 evaluated Pb acetate-induced effects on nucleotide excision repair efficiency in CL3
16 cells. All cells were exposed to 0, 100, 300 or 500 (iM Pb acetate for 24 hours in serum-
17 free medium. Pb increased nucleotide excision repair efficiency. Gastaldo et al. (2007)
18 evaluated the ability of Pb to affect DNA repair in cultured human cells. The human
19 endothelial HMEC cell line was treated with 100 (iM Pb nitrate for 24 hours. Pb inhibited
20 non-homologous end joining repair, over activated MRE11-dependent repair, and
21 increased Rad51 -related repair. Xie et al. (2008) treated BJhTERT cells (hTERT-
22 immortalized human skin fibroblasts) and ATLD-2 cells (hTERT-immortalized human
23 skin fibroblasts deficient in Mrel 1) with 0, 0.1, 0.5, and 1 (ig/cm2 Pb chromate for 24 or
24 120 hours. Mrel 1 was required to prevent Pb chromate-induced DNA double strand
25 breaks. In this finding, Pb chromate exposure and the effects of the chromate anion
26 cannot be ruled out as causative. McNeill et al, (2007) considered Pb acetate effects on
27 Apel. Chinese hamster ovary cells (AA8) were treated with 0, 0.5, 5, 50, or 500 (iM
28 Pb acetate and then whole cell extracts were used to determine AP site incision activity.
29 The data show that Pb reduced AP endonuclease function. Finally, studies considered
30 Pb-induced cellular proliferation in laboratory animals. Kermani et al. (2008) exposed
31 mouse bone marrow-mesenchymal stem cells to 0-100 (iM Pb acetate for 48 hours. As
32 measured by the MTT assay, Pb decreased cell proliferation at all concentrations tested.
33 An earlier study in rats showed Pb nitrate-induced increased proliferation of liver cells
34 after a partial hepatectomy, with more prominent effects found in males than females
35 (sexual dimorphism) (Tessitore et al., 1995). Recent studies showed similar trends in
36 males. Fortoul et al. (2005) exposed adult male CD1 mice (24 animals per group) to
37 1* 104 (iM Pb acetate, 0.006 M Cd chloride or a mixture of the two chemicals for 1 h
38 twice a week for 4 weeks by inhalation. Electron microscopy indicated Pb-induced
39 cellular proliferation in the lungs.
November 2012 5-694 Draft - Do Not Cite or Quote
-------
5.10.3.3 Epigenetics
1 Air pollution exposure is being linked increasingly with epigenetic changes in humans
2 and toxicological models (Pavanello et al., 2010; Baccarelli and Bollati. 2009; Tarantini
3 et al.. 2009; Bollati et al.. 2007). Epigenetic changes are changes in DNA expression that
4 occur without actual changes in the DNA sequence, and these changes may be heritable.
5 Epigenetic changes are mediated by histone modification, DNA methylation, miRNA
6 changes, or pathways that affect these three mediators. Differential epigenetic
7 modification has the possibility to contribute to disease. Epigenetic studies have been
8 conducted to examine the associations between Pb biomarker levels and global DNA
9 methylation markers [Alu and long interspersed nuclear element-1 (LINE-1)] in humans
10 (Wright etal.. 2010; Pilsner et al.. 2009). Wright et al. (2010) examined men from the
11 Normative Aging Study (N=517) with mean (SD) Pb levels of 20.5(14.8)g/g in tibia, 27.4
12 (19.7)g/g in patella, and 4.1 (2.4) (ig/dL in blood. In both crude and adjusted analyses,
13 patella Pb levels were inversely associated with LINE-1 methylation but not with Alu.
14 The adjusted models all included age, BMI, percent lymphocytes, with some adjusted
15 models also controlling for education, smoking, and blood Pb levels. In examination of
16 the relationship between patella Pb and LINE-1 more closely, a non-linear trend was
17 observed with a smaller magnitude of effect estimated for higher patella Pb (> 40 (ig/g).
18 No associations were observed for tibia or blood Pb and either LINE-1 or Alu. Another
19 study included maternal-infant pairs from the Early Life Exposures in Mexico to
20 Environmental Toxicants study (N=103) and measured LINE-1 and Alu methylation in
21 umbilical cord blood samples (Pilsner et al., 2009). In unadjusted models, maternal tibia
22 Pb levels one month postpartum (mean [SD]: 10.5 [8.4] (ig/g) were inversely associated
23 with Alu methylation in the cord blood. Maternal patella Pb levels one month postpartum
24 (mean [SD]: 12.9 [14.3] (ig/g) were inversely associated with LINE-1 methylation. The
25 associations persisted in models adjusted for maternal age, maternal education, infant sex,
26 smoking during pregnancy, and umbilical cord blood Pb levels (the results were no
27 longer statistically significant when umbilical cord blood was removed from the model).
28 No association was detected between umbilical cord Pb levels and the DNA methylation
29 markers. Overall, the studies consistently demonstrate an association between higher
30 patella Pb levels and lower LINE-1 methylation. Lower DNA methylation is associated
31 with increased gene expression; however, the link between global DNA methylation and
32 risk of disease, has not been established.
33 Toxicological studies have examined Pb-induced epigenetic changes and gene
34 expression, DNA repair, and mitogenesis. Glahn et al., (2008) performed a gene array
35 study in primary normal human bronchial epithelial cells from four donors after in vitro
36 treatment of the cells with 55 (ig/dL Pb chloride, 15 (ig/L Cd sulfate, 25 (ig/L Co chloride
37 or all three combined for 72 hours. The authors describe a pattern of RNA expression
November 2012 5-695 Draft - Do Not Cite or Quote
-------
1 changes indicating "... coordinated stress-response and cell-survival signaling,
2 deregulation of cell proliferation, increased steroid metabolism, and increased expression
3 of xenobiotic metabolizing enzymes." These are all known targets of possible epigenetic
4 changes, but attributing the results to epigenetic changes is complicated. In a recent
5 publication (Li etal.. 2011). exposure of HepG2 cells to a high dose of Pb (100 (iM
6 Pb acetate) resulted in ALAD gene promoter hypermethylation and decreased ALAD
7 transcription. This was in agreement with findings in battery plant workers who showed
8 ALAD hypermethylation (versus non-occupationally exposed controls) and an
9 association of this hypermethylation with elevated risk of Pb poisoning (Li et al.. 2011).
10 These latter results have implications for Pb toxicokinetics or disposition of Pb as
11 modified by ALAD.
5.10.4 Effects of Pb within Mixtures
12 Several studies considered the impact of Pb as part of a mixture on mixtures genotoxicity
13 and mutagenesis. Mendez-Gomez et al. (2008) evaluated 65 children in Mexico with high
14 environmental exposures to both As and Pb. DNA damage and decreased DNA repair
15 were seen using the comet assay and other assays but did not correlate with urinary As or
16 blood Pb levels. Tapisso et al. (2009) examined Pb alone, Pb plus Zn and Pb plus Cd-
17 induced MN in rodents. Algerian mice (groups of six mice each) were treated i.p. with 5
18 or 10 doses of 0.46 mg/kg Pb acetate and compared to untreated controls. The MN in
19 bone marrow were elevated after Pb treatment alone and increased with time.
20 Co-exposure with Cd or Zn did not further increase MN levels but did increase SCE
21 levels. Glahn et al. (2008) performed a gene array study in primary normal human
22 bronchial epithelial cells from four donors treated with 55 (ig/dL Pb chloride, 15 (ig/L Cd
23 sulfate, 25 (ig/L Co chloride or all three combined for 72 hours. There was a clear
24 interaction of all three metals impacting RNA expression.
25 Studies in the 2006 Pb AQCD (U.S. EPA. 2006b) found a protective role for calcium in
26 genotoxic and mutagenic assays with Pb co-exposure. No recent studies of the protective
27 role of calcium in Pb-induced carcinogenesis or genotoxicity were found. There were
28 some data suggesting that boron, melatonin, N-acetylcysteine, turmeric and myrrh protect
29 cells against Pb-induced genotoxicity (Section 5.10.3.2).
30 A recent study details Pb and Se interactions in virus-dependent carcinogenesis in
31 laboratory animals. Schrauzer (2008) considered the impact of Se on carcinogenesis by
32 studying four groups of weanling virgin female C3H/St mice infected with murine
33 mammary tumor virus (groups of 20-30 mice), which induces mammary tumor
34 formation. One set of two groups were fed a diet containing 0.15 ppm Se and then were
November 2012 5-696 Draft - Do Not Cite or Quote
-------
1 exposed via drinking water to acetic acid (control group) or 0.5 ppm Pb acetate. The
2 second set of two groups were fed a diet containing 0.65 ppm Se and then similarly
3 exposed to acetic acid or 0.5 ppm Pb acetate. The study was primarily focused on the
4 general effects of a low Se diet. The data suggest that Se is anticarcinogenic as in the
5 groups without Pb exposure, the animals exposed to the higher Se levels had fewer
6 mammary tumors and these tumors had a delayed onset of appearance. Pb exposure with
7 low Se caused the same delayed onset as did the higher dose of Se and also caused some
8 reduction in the tumor frequency. Pb exposure with higher Se increased the tumor
9 frequency and the onset of the tumors. Pb also induced weight loss at 14 months in both
10 exposed groups. The data suggest that there may be interactions of Pb and Se, but they
11 suggest that Pb mimics or antagonizes Se. They do not suggest that Se is protective of
12 Pb-induced toxicity or carcinogenesis.
13 In summary, the new data on Pb exposure as part of a mixture is derived from studies
14 designed with co-exposure to metals or antioxidants. Children in Mexico with
15 co-exposure to high levels of Pb and As showed elevated DNA damage and impaired
16 DNA repair. Pb and Cd co-exposure in mice elevated SCE levels but did not further
17 exacerbate MN levels above Pb exposure alone. Primary lung cells exposed to a metals
18 mixture showed an interaction at the mRNA level among the three metals tested. In other
19 genotoxicity assays, various antioxidants (melatonin, NAC, turmeric and myrrh) and
20 metals (boron) were protective against Pb-induced genotoxicity. In an animal model of
21 breast cancer, Se modified the onset and multiplicity of murine mammary tumor virus-
22 induced tumorogenicity in Pb-exposed animals. These data show that co-exposure of Pb
23 with antioxidants or metals, modifies the effect of Pb on DNA damage, DNA repair,
24 mutagenicity, genotoxicity, or tumorogenicity.
5.10.5 Summary and Causal Determination
25 Toxicological and epidemiologic studies of the association between Pb exposure and
26 cancer and cancer-related outcomes have been reviewed in the preceding sections.
27 Evaluation of the relationship between Pb exposure and cancer with respect to causality
28 was based on evidence for tumor incidence in experimental animals, associations of Pb
29 exposure with cancer incidence and mortality in humans, and evidence describing
30 potential modes of action including mutagenesis, clastogenesis, and epigenetic changes.
31 The application of the key supporting evidence from these studies to the causal
32 framework is summarized in Table 5-50 and the following text.
November 2012 5-697 Draft - Do Not Cite or Quote
-------
1 The toxicological literature provides consistent evidence of the carcinogenic potential of
2 Pb and possible contributing modes of action, including genotoxic, mutagenic and
3 epigenetic effects. In laboratory studies, chronic Pb exposure for 18 months or two years
4 to high concentrations such as 10,000 ppm Pb acetate in diet or 2,600 ppm Pb acetate in
5 drinking water has been demonstrated to be an animal carcinogen. Chronic Pb exposure
6 to male and female rodents has consistently induced kidney and brain carcinogenesis in
7 multiple separate studies, inducing various tumors, (i.e., adenocarcinomas, adenomas,
8 and gliomas. Pb has also been shown to cause mammary gland, prostate, adrenal, and
9 testicular tumors in animals. Developmental Pb acetate exposure also induced tumors in
10 offspring whose dams received Pb acetate in drinking water during pregnancy and
11 lactation. Multiple toxicological studies showed neoplastic transformation in cultured
12 cells providing an additional potential mode of action, but most used Pb chromate, and it
13 is possible that the chromate ion contributed to these findings. The toxicological and
14 epidemiologic literature provides evidence for potential carcinogenic modes of action
15 from genotoxic, mutagenic and epigenetic assays. Multiple longitudinal epidemiologic
16 studies have been performed examining the association between cancer incidence and
17 mortality and Pb exposures, estimated with biological measures and exposure databases.
18 Mixed results have been reported for cancer mortality studies; a large NHANES
19 epidemiologic study demonstrated a positive association between blood Pb and cancer
20 mortality with median 8.6 years of follow up on subjects (Schober et al., 2006). but the
21 other studies reported null results (Khalil et al.. 2009b: Weisskopf et al.. 2009; Menke et
22 al., 2006). These were well-conducted epidemiologic studies with control for important
23 potential confounders such as age, smoking, and education. Although the 2006 Pb AQCD
24 (U.S. EPA. 2006b) reported some studies that found an association between Pb exposure
25 indicators and lung cancer, recent studies mostly included occupationally-exposed adults
26 and observed no associations. Most studies of Pb and brain cancer were null among the
27 overall study population, but positive associations were observed among individuals with
28 certain genetic variants. However, the studies of Pb and brain cancer were all limited by
29 the use of occupational cohorts and interviews instead of biological measurements to
30 represent Pb exposure, and by possible confounding by several factors, including other
31 workplace exposures. A limited amount of research has been performed on other types of
32 cancer. The 2006 Pb AQCD reported evidence that suggested an association between Pb
33 exposure and stomach cancer, but in a recent study of stomach cancer the results were
34 inconsistent, reporting a positive association between organic Pb exposure and stomach
35 cancer but null findings for exposure to inorganic Pb or Pb from gasoline emissions and
36 stomach cancer.
37 Among epidemiologic studies, high Pb levels (over 40 (ig/dL in adults) were associated
38 with SCEs among adults. This association was not observed among children (mean blood
39 Pb 7.69 (ig/dL). Other epidemiologic studies of DNA damage reported inconsistent
November 2012 5-698 Draft - Do Not Cite or Quote
-------
1 results. Consistent with previous toxicological findings, Pb does appear to have genotoxic
2 activity in animal and in vitro models, inducing SCE, MN and DNA strand breaks. The
3 majority of the chromosomal aberration studies with Pb-induced significant finding used
4 Pb chromate exposure and the aberrations are likely due to the chromate. Pb does not
5 appear to be very mutagenic as the HPRT assays were typically negative unless a cell
6 signaling pathway was disturbed.
7 Mechanistic understanding of the carcinogenicity of Pb in toxicological models is
8 expanding with work on the antioxidant Se and metallothionein, a protein that binds Pb
9 and reduces its bioavailability. Low Se diet affects tumorigenesis and tumor multiplicity
10 with Pb exposure. Metallothionein has been shown to be protective against the effect of
11 Pb on carcinogenicity. Pb is clastogenic and mutagenic in some but not all models.
12 Clastogenicity and mutagenicity may be possible mechanisms contributing to cancer but
13 are not absolutely associated with the induction of cancer. Because Pb has a higher
14 atomic weight than does Zn, Pb replaces Zn at many Zn binding sites or Zn finger
15 proteins. This substitution has the potential to induce effects that can indirectly contribute
16 to carcinogenicity via interactions with hormone receptors, cell-cycle regulatory proteins,
17 tumor suppressor genes like p53, DNA repair enzymes, histones, etc. These indirect
18 effects may act at a post-translational level to negatively alter protein structure and DNA
19 repair.
20 Epigenetic changes associated with Pb exposure or biological markers, particularly,
21 methylation and effects on DNA repair, are beginning to appear in the literature.
22 Epigenetic modifications may contribute to carcinogenicity by altering DNA repair or
23 changing the expression of a tumor suppressor gene or oncogene. A small number of
24 epidemiologic studies examining Pb and global epigenetic changes demonstrated an
25 inverse association between bone Pb and LINE-1 or Alu methylation. Lower DNA
26 methylation is associated with increased gene expression, but epigenetic contributions to
27 cancer are not yet fully characterized in this emerging area of research. Toxicological
28 studies show that Pb can activate or interfere with a number of signaling and repair
29 pathways, though it is unclear whether these are due to epigenetic responses or
30 genotoxicity. Thus, an underlying mechanism is still uncertain, but likely involves
31 genomic instability, epigenetic modifications, or both.
32 In conclusion, the toxicological literature provides the strong evidence for long-term
33 exposure (i.e., 18 months or 2 years) to high concentrations of Pb (> 2,600 ppm) and
34 cancer. The consistent evidence indicating Pb-induced carcinogenicity in animal models
35 is substantiated by the mode of action findings from multiple high-quality toxicological
36 studies in animal and in vitro models from different laboratories. This is substantiated by
37 the findings of other agencies including IARC, which has classified inorganic Pb
November 2012 5-699 Draft - Do Not Cite or Quote
-------
1 compounds as a probable human carcinogen and the National Toxicology Program,
2 which has listed Pb and Pb compounds as "reasonably anticipated to be human
3 carcinogens." Strong evidence from animal toxicological studies demonstrates an
4 association between Pb and cancer, genotoxicity, mutagenicity or epigenetic
5 modification. Carcinogenicity in animal toxicology studies with relevant routes of Pb
6 exposure has been reported in the kidneys, testes, brain, adrenals, prostate, pituitary, and
7 mammary gland, albeit at high doses of Pb. Epidemiologic studies of cancer incidence
8 and mortality reported inconsistent results; one strong epidemiologic study demonstrated
9 an association between blood Pb and increased cancer mortality, but the other studies
10 reported weak or no associations. In the 2006 Pb AQCD, indicators of Pb exposure were
11 found to be associated with stomach cancer, and a recent study on stomach cancer and
12 occupational Pb exposure, reported mixed findings depending on the type of Pb exposure
13 (organic Pb, inorganic Pb, or Pb from gasoline emissions). Similarly, some studies in the
14 2006 Pb AQCD reported associations between Pb exposure indicators and lung cancer.
15 Recent epidemiologic studies of lung cancer focused on occupational exposures and
16 reported inconsistent associations. The majority of epidemiologic studies of brain cancer
17 had null results overall, but positive associations between Pb exposure indicators and
18 brain cancer were observed among individuals with certain genotypes. Overall, the
19 consistent and strong body of evidence from toxicological studies but inconsistent
20 epidemiologic evidence is sufficient to conclude that a causal relationship is likely to
21 exist between Pb exposure and cancer.
November 2012 5-700 Draft - Do Not Cite or Quote
-------
Table 5-50 Summary of evidence supporting cancer and genotoxicity
causal determinations.
Attribute in
Causal
Framework3
Key Supporting Evidence13
Pb Exposure or
Blood Levels
Associated
References13 with Effects
Cancer - Likely Causal
Consistent Consistent findings across multiple
toxicological toxicology studies using 18 month or
results of tumors two year cancer bioassays in rats
in laboratory wherein rodents are fed chow or
animals with received drinking water enriched with
chronic Pb Pb acetate, showing tumor
exposure development.
Azaretal. (1973).
Kasprzak et al. (1985).
Kolleretal. (1985),
Van Esch and Kroes (1969)
Chronic 10,000 ppm
Pb acetate diet or
2,600 ppm drinking water
Pb acetate, no blood Pb
measurement available.
Toxicological
studies of early
life Pb exposure
induced tumor
formation in
adulthood
Gestational and lactational Pb exposure
induced carcinogenicity in adult
offspring
Waalkesetal., (1995)
500, 750 and 1,000 ppm
Pb in drinking water, no
blood Pb measurement
available.
Limited and Epidemiologic studies of overall cancer
inconsistent mortality have inconsistent findings.
epidemiologic These are high-quality, longitudinal
studies studies with control for confounders,
such as age, smoking, and education.
Epidemiologic studies of specific cancer
sites were limited. Recent studies were
not consistent with previous findings of
possible associations for lung and
stomach cancers reported in the
2006 PbAQCD. Many of the
epidemiologic studies examining
specific cancer sites were case-control
studies and not all included potentially
important confounders, such as
smoking.
Menke et al. (2006)
Schoberetal. (2006)
Weisskopfetal. (2009),
Khalil (2009b)
Overall Cancer Mortality:
See Section 5.10.1.1
Specific Cancer: See Sections:
5.10.1.3 (Lung), 5.10.1.4 (Brain),
5.10.1.5 (Breast): and
Section 5.10.1.6 (Other cancers).
Evidence clearly
supports mode of
action
Mutagenic,
carcinogenic
and genotoxic
assays provide
consistent
support to the
MOA
Clastogenic
assays provide
inconsistent
support to the
MOA
Epigenetic
evidence
provides
support to the
MOA
Consistent evidence of toxicological
findings of mutagenicity,
carcinogenicity, and genotoxicity has
been reported by multiple laboratories
in animals and in vitro models using
multiple assays (MN, SCE, COMET).
Toxicological studies of the clastogenic
effects of Pb often employ
Pb chromate. The effect of the
chromate ion in contributing to the
clastogenic effects cannot be ruled out.
Bone Pb levels were inversely
associated with LINE-1 methylation in
adult men.
Maternal pregnancy bone Pb levels
were inversely associated with Alu and
LINE-1 methylation in child cord blood.
Occupational battery workers had
ALAD hypermethylation compared with
controls; cell culture study of high dose
Pb exposure caused ALAD
hypermethylation.
Epidemiology evidence of DMA and
cellular damage:
See Section 5.10.3.2
Toxicology evidence of DMA and
cellular damage:
See Section 5.10.3.2
See Section 5.10.3.2 (Toxicological
Evidence for DMA and Cellular
Damage)
See Section 5.10.3.3
Described in detail in Table II of the Preamble.
bDescribes the key evidence and references contributing the most heavily to causal determination. References to earlier sections
indicate where full body of evidence is described.
November 2012
5-701
Draft - Do Not Cite or Quote
-------
References for Chapter 5
Abam. E; Okediran. BS: Odukoya. OO: Adamson. I; Ademuyiwa. O. (2008). Reversal of ionoregulatory
disruptions in occupational lead exposure by vitamin C. Environ Toxicol Pharmacol 26: 297-304.
http://dx.doi.0rg/10.1016/i.etap.2008.05.008
Abdel Moneim. AE; Dkhil. MA; Al-Quraishy. S. (2011 a). Effects of flaxseed oil on lead acetate-induced
neurotoxicity in rats. Biol Trace Elem Res 144: 904-913. http://dx.doi.org/10.1007/sl2011-011-9055-4
Abdel Moneim. AE; Dkhil. MA; Al-Quraishy. S. (20 lib). The protective effect of flaxseed oil on lead
acetate-induced renal toxicity in rats. J Hazard Mater 194: 250-255.
http://dx.doi.0rg/10.1016/i.jhazmat.2011.07.097
Abdelouahab. N; Mergler. D; Takser. L; Vanier. C: St-Jean. M; Baldwin. M; Spear. PA; Chan. HM.
(2008). Gender differences in the effects of organochlorines, mercury, and lead on thyroid hormone
levels in lakeside communities of Quebec (Canada). Environ Res 107: 380-392.
http://dx.doi.0rg/10.1016/i.envres.2008.01.006
Aboul-Ela. EL (2002). The protective effect of calcium against genotoxicity of lead acetate administration
on bone marrow and spermatocyte cells of mice in vivo. Mutat Res Genet Toxicol Environ Mutagen
516: 1-9. http://dx.doi.org/10.1016/81383-5718(01)00332-1
Absalon. D; Slesak. B. (2010). The effects of changes in cadmium and lead air pollution on cancer
incidence in children. Sci Total Environ 408: 4420-4428.
http://dx.doi.0rg/10.1016/i.scitotenv.2010.06.030
Adegbesan. BO; Adenuga. GA. (2007). Effect of lead exposure on liver lipid peroxidative and antioxidant
defense systems of protein-undernourished rats. Biol Trace Elem Res 116: 219-225.
http://dx.doi.org/10.1007/BF02685932
Ademuyiwa. O: Agarwal. R; Chandra. R; Behari. JR. (2009). Lead-induced phospholipidosis and
cholesterogenesis in rat tissues. Chem Biol Interact 179: 314-320.
http://dx.doi.org/10.1016/i.cbi.20Q8.10.057
Ademuyiwa. O: Ugbaja. RN; Idumebor. F; Adebawo. O. (2005a). Plasma lipid profiles and risk of
cardiovascular disease in occupational lead exposure in Abeokuta, Nigeria. Lipids Health Dis 4:19.
http://dx.doi.org/10.1186/1476-511X-4-19
Ademuyiwa. O: Ugbaja. RN; Ojo. DA; Owoigbe. AO; Adeokun. SE. (2005b). Reversal of aminolevulinic
acid dehydratase (ALAD) inhibition and reduction of erythrocyte protoporphyrin levels by Vitamin C
in occupational lead exposure in Abeokuta, Nigeria. Environ Toxicol Pharmacol 20: 404-411.
http://dx.doi.0rg/10.1016/i.etap.2005.04.002
Adonaylo. VN; Oteiza. PI. (1999). Pb2+ promotes lipid oxidation and alterations in membrane physical
properties. Toxicology 132: 19-32. http://dx.doi.org/10.1016/S0300-483X(98)00134-6
Afeiche. M; Peterson. KE; Sanchez. BN; Cantonwine. D: Lamadrid-Figueroa. H; Schnaas. L; Ettinger.
AS: Hernandez-Avila. M; Hu. H; Tellez-Rojo. MM. (2011). Prenatal lead exposure and weight of 0 to
5 year-old children in Mexico City. Environ Health Perspect 119: 1436-1441.
http://dx.doi.org/10.1289/ehp.1003184
Agarwal. S: Roy. S: Ray. A; Mazumder. S: Bhattacharya. S. (2009). Arsenic trioxide and lead acetate
induce apoptosis in adult rat hepatic stem cells. Cell Biol Toxicol 25: 403-413.
http://dx.doi.org/10.1007/sl0565-008-9094-6
Aguilar. RP; Genta. S: Sanchez. S. (2008). Renal gangliosides are involved in lead intoxication. J Appl
Toxicol 28: 122-131. http://dx.doi.org/10.1002/iat.1256
November 2012 5-702 Draft - Do Not Cite or Quote
-------
Ahamed. M; Fareed. M; Kumar. A; Siddiqui. WA; Siddiqui. MKJ. (2008). Oxidative stress and
neurological disorders in relation to blood lead levels in children. Redox Rep 13: 117-122.
http://dx.doi.org/10.1179/135100008X259213
Ahamed. M; Mehrotra. PK; Kumar. P; Siddiqui. MKJ. (2009). Placenta! lead-induced oxidative stress and
preterm delivery. Environ Toxicol Pharmacol 27: 70-74. http://dx.doi.Org/10.1016/i.etap.2008.08.013
Ahamed. M; Siddiqui. MK. (2007). Low level lead exposure and oxidative stress: Current opinions
[Review]. Clin Chim Acta 383: 57-64. http://dx.doi.Org/10.1016/i.cca.2007.04.024
Ahamed. M; Singh. S: Behari. JR; Kumar. A; Siddiqui. MK. (2007). Interaction of lead with some
essential trace metals in the blood of anemic children from Lucknow, India. Clin Chim Acta 377:
92-97. http://dx.doi.0rg/10.1016/j.cca.2006.08.032
Ahamed. M; Verma. S: Kumar. A; Siddiqui. MKJ. (2005). Environmental exposure to lead and its
correlation with biochemical indices in children. Sci Total Environ 346: 48-55.
http://dx.doi.0rg/10.1016/i.scitotenv.2004.12.019
Ahamed. M; Verma. S: Kumar. A; Siddiqui. MKJ. (2006). Delta-aminolevulinic acid dehydratase
inhibition and oxidative stress in relation to blood lead among urban adolescents. Hum Exp Toxicol
25: 547-553. http://dx.doi.org/10.1191/0960327106het657oa
Aisemberg. J; Nahabedian. DE; Wider. EA; Verrengia Guerrero. NR. (2005). Comparative study on two
freshwater invertebrates for monitoring environmental lead exposure. Toxicology 210: 45-53.
http://dx.doi.0rg/10.1016/i.tox.2005.01.005
Akel. A; Wagner. CA; Kovacikova. J: Kasinathan. RS: Kiedaisch. V; Koka. S: Alper. SL; Bernhardt. I;
Wieder. T; Huber. SM; Lang. F. (2007). Enhanced suicidal death of erythrocytes from gene-targeted
mice lacking the C1-/HCO3- exchanger AE1. Am J Physiol Cell Physiol 292: C1759-C1767.
http://dx.doi.org/10.1152/aipcell.00158.2006
Akesson. A; Lundh. T; Vahter. M; Bjellerup. P; Lidfeldt. J: Nerbrand. C: Goran. S: Stromberg. U;
Skerfving. S. (2005). Tubular and glomerular kidney effects in Swedish women with low
environmental cadmium exposure. Environ Health Perspect 113: 1627-1631.
httrj://dx.doi.org/10.1289/ehp.8Q33
Al-Saleh. I; Coskun. S: Mashhour. A; Shinwari. N; El-Doush. I; Billedo. G: Jaroudi. K; Al-Shahrani. A;
Al-Kabra. M; El Din Mohamed. G. (2008a). Exposure to heavy metals (lead, cadmium and mercury)
and its effect on the outcome of in-vitro fertilization treatment. Int J Hyg Environ Health 211:
560-579. http://dx.doi.0rg/10.1016/i.iiheh.2007.09.005
Al-Saleh. I; Shinwari. N; Nester. M; Mashhour. A; Moncari. L; Mohamed. GED; Rabah. A. (2008b).
Longitudinal study of prenatal and postnatal lead exposure and early cognitive development in Al-
Kharj, Saudi Arabia: A preliminary results of cord blood lead levels. JTrop Pediatr 54: 300-307.
http://dx.doi.org/10.1093/tropej/fmn019
Alasia. DP; Emem-Chioma. PC: Wokoma. FS. (2010). Occupational and environmental lead exposure in
Port Harcourt, Nigeria: Analysis of its association with renal function indices. Niger J Med 19:
407-414.
Alatise. OI; Schrauzer. GN. (2010). Lead exposure: A contributing cause of the current breast cancer
epidemic in Nigerian women. Biol Trace Elem Res 136: 127-139.
http://dx.doi.org/10.1007/sl2011-010-8608-2
Alber. SA; Strupp. BJ. (1996). An in-depth analysis of lead effects in a delayed spatial alternation task:
assessment of mnemonic effects, side bias, and proactive interference. Neurotoxicol Teratol 18: 3-15.
http://dx.doi.org/10.1016/0892-0362(95)02026-8
Alexander. KR; Fishman. GA. (1984). Supernormal scotopic ERG in cone dystrophy. Br J Ophthalmol
68: 69-78.
Alfano. DP; Petit. TL. (1981). Behavioral effects of postnatal lead exposure: possible relationship to
hippocampal dysfunction. Behav Neural Biol 32: 319-333.
November 2012 5-703 Draft - Do Not Cite or Quote
-------
Alghazal. MA; Lenartova. V: Holovska. K; Sobekova. A; Falls. M; Legatfa. J. (2008a). Activities of
antioxidant and detoxifying enzymes in rats after lead exposure. Acta Vet (Beogr) 77: 347-354.
http://dx.doi.org/10.2754/avb200877030347
Alghazal. MA; Sutiakova. I; Kovalkovicova. N; Legath. J; Palis. M; Pistl. J: Sabo. R; Benova. K; Sabova.
L; Vaczi. P. (2008b). Induction of micronuclei in rat bone marrow after chronic exposure to lead
acetate trihydrate. Toxicol Ind Health 24: 587-593. http://dx.doi.org/10.1177/0748233708100089
Aligne. CA; Auinger. P; Byrd. RS: Weitzman. M. (2000). Risk factors for pediatric asthma: Contributions
of poverty, race, and urban residence. Am J Respir Crit Care Med 162: 873-877.
Alimonti. A; Bocca. B; Lamazza. A; Forte. G: Rahimi. S: Mattei. D; Fiori. E; lacomino. M; Schillaci. A;
De Masi. E; Pino. A. (2008). A study on metals content in patients with colorectal polyps. J Toxicol
Environ Health A 71: 342-347. http://dx.doi.org/10.1080/15287390701839133
Alinovi. R; Scotti. E; Andreoli. R; De Palma. G: Goldoni. M; Apostoli. P; Mutti. A. (2005).
[Neuroendocrine and renal effects of inorganic lead]. G Ital Med Lav Ergon 27: 33-38.
Allouche. L; Hamadouche. M; Touabti. A. (2009). Chronic effects of low lead levels on sperm quality,
gonadotropins and testosterone in albino rats. Exp Toxicol Pathol 61: 503-510.
http://dx.doi.0rg/10.1016/i.etp.2008.12.003
Alomran. AH; Shleamoon. MN. (1988). The influence of chronic lead exposure on lymphocyte
proliferative response and immunoglobulin levels in storage battery workers. J Biol Sci 19: 575-585.
Altmann. L; Weinsberg. F; Sveinsson. K; Lilienthal. H; Wiegand. H; Winneke. G. (1993). Impairment of
long-term potentiation and learning following chronic lead exposure. Toxicol Lett 66: 105-112.
http://dx.doi.org/10.1016/0378-4274(93)90085-C
Altmann. P; Maruna. RF; Maruna. H; Michalica. W: Wagner. G. (1981). [Lead detoxication effect of a
combined calcium phosphate and ascorbic acid therapy in pregnant women with increased lead burden
(author's transl)]. Wiener medizinische Wochenschrift 131: 311-314.
Alvarez. J: Garcia-Sancho. J: Herreros. B. (1986). Inhibition of Ca2+-dependent K+ channels by lead in
one-step inside-out vesicles from human red cell membranes. Biochim Biophys Acta 857: 291-294.
http://dx.doi.org/10.1016/0005-2736(86)90359-7
Amici. A; Emanuelli. M; Magni. G: Raffaelli. N; Ruggieri. S. (1997). Pyrimidine nucleotidases from
human erythrocyte possess phosphotransferase activities specific for pyrimidine nucleotides. FEES
Lett 419: 263-267. http://dx.doi.org/10.1016/S0014-5793(97)01464-6
Andersen. ZJ; Wahlin. P; Raaschou-Nielsen. O; Scheike. T; Loft. S. (2007). Ambient particle source
apportionment and daily hospital admissions among children and elderly in Copenhagen. J Expo Sci
Environ Epidemiol 17: 625-636. http://dx.doi.org/10.1038/sj.jes.7500546
Andreae. MO. (1983). Biotransformation of arsenic in the marine environment. In WH Lederer; RJ
Fensterheim (Eds.), Arsenic: Industrial, biomedical and environmental perspectives: Proceedings of
the Arsenic Symposium (pp. 378-391). New York, NY: Van Nostrand Reinhold.
Anetor. JI; Adeniyi. FAA. (1998). Decreased immune status in Nigerian workers occupationally exposed
to lead. Afr J Med Med Sci 28: 169-172.
Anjum. RM; Sainath. SB; Suneetha. Y; Reddv. SP (2010). Lead acetate induced reproductive and
paternal mediated developmental toxicity in rats. Ecotoxicol Environ Saf 74: 793-799.
http://dx.doi.0rg/10.1016/i.ecoenv.2010.10.044
Annesi-Maesano. I; Pollitt R: King. G: Bousquet. J: Hellier. G: Sahuquillo. J: Huel. G. (2003). In utero
exposure to lead and cord blood total IgE: Is there a connection? Allergy 58: 589-594.
http://dx.doi.0rg/10.1034/i.1398-9995.2003.00111.x
Antonowicz. J: Andrzejak. R; Smolik. R. (1990). Influence of heavy metal mixtures on erythrocyte
metabolism. Int Arch Occup Environ Health 62: 195-198. http://dx.doi.org/10.1007/BF00379431
November 2012 5-704 Draft - Do Not Cite or Quote
-------
Antonyuk. SV: Strange. RW: Marklund. SL; Hasnain. SS. (2009). The structure of human extracellular
copper-zinc superoxide dismutase at 1.7 A resolution: Insights into heparin and collagen binding. J
Mol Biol 338: 310-326. http://dx.doi.Org/10.1016/j.jmb.2009.03.026
Anwar. WA; Kamal. AAM. (1988). Cytogenetic effects in a group of traffic policemen in Cairo. Mutat
Res Lett 208: 225-231. http://dx.doi.org/10.1016/0165-7992(88)90065-6
Apostoli. P: Huard. C: Chaumontet C: Mattel. P; Alleskx L; Mazzoleni. G. (2000). Effects of four
inorganic lead compounds on the proliferation and junctional coupling of cultured REL liver cells. Am
JIndMed 38: 340-348.
Apostoli. P: Romeo. L; De Matteis. MC: Menegazzi. M; Faggionato. G: Vettore. L. (1988). Effects of
lead on red blood cell membrane proteins. Int Arch Occup Environ Health 61: 71-75.
http://dx.doi.org/10.1007/BF00381610
Aquilonius. SM; Hartvig. P. (1986). A Swedish county with unexpectedly high utilization of anti-
parkinsonian drugs. ActaNeurol Scand 74: 379-382.
http://dx.doi.org/10.1111/i. 1600-0404.1986.tb03529.x
Ari. E; Kava. Y; Demir. H; Asicioglu. E; Keskin. S. (2011). The Correlation of Serum Trace Elements and
Heavy Metals with Carotid Artery Atherosclerosis in Maintenance Hemodialysis Patients. Biol Trace
Elem Res 144: 351-359. http://dx.doi.org/10.1007/sl2011-011-9103-0
Ariza. ME; Bijur. GN; Williams. MV. (1998). Lead and mercury mutagenesis: Role of H2O2, superoxide
dismutase, and xanthine oxidase. Environ Mol Mutagen 31: 352-361.
http://dx.doi.org/10.1002/(SICI)1098-2280(1998)31:4<352::AID-EM8>3.0.CO:2-K
Ariza. ME; Williams. MV. (1996). Mutagenesis of AS52 cells by low concentrations of lead(II) and
mercury(II). Environ Mol Mutagen 27: 30-33.
httrj://dx.doi.org/10.1002/(SICI)1098-2280(1996)27:l<:30::AID-EM4>:3.0.CO:2-I
Ariza. ME; Williams. MV. (1999). Lead and mercury mutagenesis: Type of mutation dependent upon
metal concentration. J Biochem Mol Toxicol 13: 107-112.
http://dx.doi.org/10.1002/(SICI)1099-0461(1999)13:2<107::AID-JBT6>3.0.CO:2-0
Arnold. WJ: Kellev. WN. (1978). Adenine phosphoribosyltransferase. Methods Enzymol 51: 568-574.
http://dx.doi.org/10.1016/80076-6879(78)51079-3
Arora. M; Chan. SW: Ryan. CG: Kennedy. BJ; Walker. DM. (2005). Spatial distribution of lead in enamel
and coronal dentine of wistar rats. Biol Trace Elem Res 105: 159-170.
http://dx.doi.Org/10.1385/BTER:105:l-3:159
Arora. M; Weuve. J: Weisskopf. MG: Sparrow. D; Nie. H; Garcia. RI; Hu. H. (2009). Cumulative lead
exposure and tooth loss in men: The normative aging study. Environ Health Perspect 117: 1531-1534.
http://dx.doi.org/10.1289/ehp.0900739
Arrieta. O: Palencia. G: Garcia-Arenas. G: Morales-Espinosa. D; Hernandez-Pedro. N; Sotelo. J. (2005).
Prolonged exposure to lead lowers the threshold of pentylenetetrazole-induced seizures in rats.
Epilepsia46: 1599-1602. http://dx.doi.org/10.1111/i. 1528-1167.2005.00267.x
Astrin. KH; Bishop. DF; Wetmur. JG: Kaul. B; Davidow. B; Desnick. RJ. (1987). 5-aminolevulinic acid
dehydratase isozymes and lead toxicity. Ann N Y Acad Sci 514: 23-29.
http://dx.doi.org/10.1111/i. 1749-6632.1987.tb48757.x
Atabek. ME; Kurtoglu. S: Pirgon. O; Uzum. K; Saraymen. R. (2007). Relation of in utero lead exposure
with insulin-like growth factor-I levels and neonatal anthropometric parameters. Int J Hyg Environ
Health 210: 91-95. httrj://dx.doi.org/10.1016/i.iiheh.2006.06.007
Atchison. WD. (2003). Effects of toxic environmental contaminants on voltage-gated calcium channel
function: From past to present. J Bioenerg Biomembr 35: 507-532.
http://dx.doi.Org/10.1023/B:JOBB.0000008023.11211.13
November 2012 5-705 Draft - Do Not Cite or Quote
-------
ATS (American Thoracic Society). (2000). What constitutes an adverse health effect of air pollution? This
official statement of the American Thoracic Society was adopted by the ATS Board of Directors, July
1999. Am J Respir Crit Care Med 161: 665-673.
Attri. J; Dhawan. V; Mahmood. S: Pandhi. P; Parwana. HK; Nath. R. (2003). Effect of vitamin C
supplementation on oxidative DNA damage in an experimental model of lead-induced hypertension.
Ann Nutr Metab 47: 294-301. http://dx.doi.org/10.1159/000072402
Audesirk. G: Audesirk. T. (1991). Effects of inorganic lead on voltage-sensitive calcium channels in
N1E-115 neuroblastoma cells. Neurotoxicology 12: 519-528.
Audesirk. G: Audesirk. T. (1993). The effects of inorganic lead on voltage-sensitive calcium channels
differ among cell types and among channel subtypes. Neurotoxicology 142: 259-266.
Azar. A; Trochimowicz. HJ; Maxfield. ME. (1973). Review of lead studies in animals carried out at
Haskell Laboratory: Two year feeding study and response to hemorrhage study. In D Earth; A Berlin;
R Engel; P Recht; J Smeets (Eds.), Environmental health aspects of lead: Proceedings of an
international symposium (pp. 199-210). Luxembourg: Commission of the European Communities.
Baccarelli. A; Bollati. V. (2009). Epigenetics and environmental chemicals [Review]. Curr Opin Pediatr
21: 243-251. http://dx.doi.org/10.1097/MOP.Ob013e32832925cc
Badavi. M; Mehrgerdi. FZ; Sarkaki. A; Naseri. MK; Dianat. M. (2008). Effect of grape seed extract on
lead induced hypertension and heart rate in rat. Pak J Biol Sci 11: 882-887.
Bagchi. D; Preuss. HG. (2005). Effects of acute and chronic oral exposure of lead on blood pressure and
bone mineral density in rats. J Inorg Biochem 99: 1155-1164.
http://dx.doi.0rg/10.1016/i.jinorgbio.2005.02.011
Baghurst. PA; McMichael. AJ; Wigg. NR; Vimpani. GV; Robertson. EF; Roberts. RJ; Tong. SL. (1992).
Environmental exposure to lead and children's intelligence at the age of seven years: The Port Pirie
cohort study. N Engl J Med 327: 1279-1284. http://dx.doi.org/10.1056/NEJM199210293271805
Baker. EL: Feldman. RG: White. RA: Harlev. JP: Niles. CA: Dinse. GE: Berkev. CS. (1984).
Occupational lead neurotoxicity: A behavioural and electrophysiological evaluation: Study design and
year one results. Occup Environ Med 41: 352-361. http://dx.doi.Org/10.1136/oem.41.3.352
Baker. EL; Landrigan. PJ; Barbour. AG; Cox. DH; Folland. PS; Ligo. RN; Throckmorton. J. (1979).
Occupational lead poisoning in the United States: Clinical and biochemical findings related to blood
lead levels. Occup Environ Med 36: 314-322. http://dx.doi.Org/10.1136/oem.36.4.314
Baker. EL; White. RF; Pothier. LJ; Berkev. CS; Dinse. GE; Travers. PH; Harlev. JP; Feldman. RG.
(1985). Occupational lead neurotoxicity: Improvement in behavioural effects after reduction of
exposure. Br J Ind Med 42: 507-516. http://dx.doi.Org/10.1136/oem.42.8.507
Balbuena. P; Li. W; Ehrich. M. (2011). Assessments of tight junction proteins occludin, claudin 5 and
scaffold proteins ZO1 and ZO2 in endothelial cells of the rat blood-brain barrier: cellular responses to
neurotoxicants malathion and lead acetate. Neurotoxicology 32: 58-67.
http://dx.doi.0rg/10.1016/i.neuro.2010.10.004
Balbuena. P; Li. W; Magnin-Bissel. G; Meldrum. B; Ehrich. M. (2010). Comparison of Two Blood-Brain
Barrier in Vitro Systems: Cytotoxicity and Transfer Assessments of Malathion/oxon and Lead Acetate.
Toxicol Sci 114: 260-271. http://dx.doi.org/10.1093/toxsci/kfqOO 1
Ball. GV; Sorensen. LB. (1969). Pathogenesis of hyperuricemia in saturnine gout. N Engl J Med 280:
1199-1202. http://dx.doi.org/10.1056/neiml96905292802203
Bandeen-Roche. K; Glass. TA; Bolla. KI; Todd. AC; Schwartz. BS. (2009). Cumulative lead dose and
cognitive function in older adults. Epidemiology 20: 831-839.
http://dx.doi.org/10.1097/EDE.Ob013e3181b5flOO
November 2012 5-706 Draft - Do Not Cite or Quote
-------
Bao. OS: Lu. CY; Song. H; Wang. M; Ling. WH; Chen. WQ: Deng. XQ; Hao. YT; Rao. SO. (2009).
Behavioural development of school-aged children who live around a multi-metal sulphide mine in
Guangdong province, China: Across-sectional study. BMC Public Health 9: 217.
http://dx.doi.org/10.1186/1471-2458-9-217
Baranowska-Bosiacka. I; D^browska-Bouta. B; Struzynska. L. (2011 a). Regional changes in purines and
selected purinergic receptors in immature rat brain exposed to lead. Toxicology 279: 100-107.
htrp://dx.doi.org/10.1016/i.tox.2010.09.016
Baranowska-Bosiacka. I; Dziedziejko. V; Safranow. K; Gutowska. I; Marchlewicz. M; Dolegowska. B;
Rac. ME; Wiszniewska. B; Chlubek. D. (2009). Inhibition of erythrocyte phosphoribosyltransferases
(APRT and HPRT) by Pb2+: A potential mechanism of lead toxicity. Toxicology 259: 77-83.
htrp://dx.doi.org/10.1016/i.tox.2009.02.005
Baranowska-Bosiacka. I; Gutowska. I; Marchetti. C: Rutkowska. M; Marchlewicz. M; Kolasa. A;
Prokopowicz. A; Wiernicki. I; Piotrowska. K; Baskiewicz. M; Safranow. K; Wiszniewska. B;
Chlubek. D. (20 lib). Altered energy status of primary cerebellar granule neuronal cultures from rats
exposed to lead in the pre- and neonatal period. Toxicology 280: 24-32.
http://dx.doi.0rg/10.1016/i.tox.2010.ll.004
Baranowska-Bosiacka. I; Hlynczak. AJ. (2003). The effect of lead ions on the energy metabolism of
human erythrocytes in vitro. Comp Biochem Physiol C Toxicol Pharmacol 134: 403-416.
http://dx.doi.org/10.1016/S1532-0456(03)00008-5
Baranowska-Bosiacka. I; Hlynczak. AJ. (2004). Effect of lead ions on rat erythrocyte purine content. Biol
Trace Elem Res 100: 259-273. http://dx.doi.Org/10.1385/BTER:100:3:259
Barbeito. AG: Martinez-Palma. L; Vargas. MR; Pehar. M; Mafiav. N; Beckman. JS: Barbeito. L; Cassina.
P (2010). Lead exposure stimulates VEGF expression in the spinal cord and extends survival in a
mouse model of ALS. Neurobiol Dis 37: 574-580. http://dx.doi.Org/10.1016/i.nbd.2009.ll.007
Barbosa. F. Jr; Sandrim. VC: Uzuelli. JA; Gerlach. RF; Tanus-Santos. JE. (2006c). eNOS genotype-
dependent correlation between whole blood lead and plasma nitric oxide products concentrations.
Nitric Oxide 14: 58-64. http://dx.doi.Org/10.1016/i.niox.2005.09.007
Barker. DJ; Osmond. C: Golding. J; Kuh. D; Wadsworth. ME. (1989). Growth in utero, blood pressure in
childhood and adult life, and mortality from cardiovascular disease. BMJ 298: 564-567.
Barker. DJP; Bagbv. SP (2005). Developmental antecedents of cardiovascular disease: a historical
perspective. J Am Soc Nephrol 16: 2537-2544. http://dx.doi.org/10.1681/ASN.2005020160
Baronas. ET; Lee. JW: Alden. C: Hsieh. FY. (2007). Biomarkers to monitor drug-induced
phospholipidosis. Toxicol Appl Pharmacol 218: 72-78. http://dx.doi.Org/10.1016/i.taap.2006.10.015
Basha. MR; Murali. M; Siddiqi. HK; Ghosal. K; Siddiqi. OK; Lashuel. HA; Ge. YW: Lahiri. DK; Zawia.
NH. (2005). Lead (Pb) exposure and its effect on APP proteolysis and A beta aggregation. FASEB J
19: 2083-2084. http://dx.doi.org/10.1096/fi.05-4375fje
Basha. MR; Wei. W: Brydie. M; Razmiafshari. M; Zawia. NH. (2003). Lead-induced developmental
perturbations in hippocampal Spl DNA-binding are prevented by zinc supplementation: In vivo
evidence for Pb and Zn competition. Int J Dev Neurosci 21: 1-12.
htrp://dx.doi.org/10.1016/S0736-5748(02)00137-5
Batalha. J. R.: Saldiva P. H; Clarke. RW: Coull. BA; Stearns. RC: Lawrence. J: Murthy. GG: Koutrakis.
P; Godleski. JJ. (2002). Concentrated ambient air particles induce vasoconstriction of small
pulmonary arteries in rats. Environ Health Perspect 110: 1191-1197.
Batuman. V. (1993). Lead nephropathy, gout, and hypertension [Review]. Am J Med Sci 305: 241-247.
Batuman. V; Landv. E; Maesaka. JK; Wedeen. RP (1983). Contribution of lead to hypertension with renal
impairment. N Engl J Med 309: 17-21. http://dx.doi.org/10.1056/NEJM198307073090104
Bayer. SA. (1989). Cellular aspects of brain development [Review]. Neurotoxicology 10: 307-320.
November 2012 5-707 Draft - Do Not Cite or Quote
-------
Baykov. B; Gunova. M; Stovanov. M; Nevchev. H; Stefanova. T; Nicolova. N. (1996). Designing an
artificial ecological mesocosm for the study of Cd and Pb impact on the immune system of
experimental animals. Toxicol Lett 89: 5-10.
Bear. MF; Malenka. RC. (1994). Synaptic plasticity: LTP and LTD. Curr Opin Neurobiol 4: 389-399.
http://dx.doi.org/10.1016/0959-4388(94)90101-5
Beaudin. SA; Stangle. DE; Smith. PR; Levitsky. DA; Strupp. BJ. (2007). Succimer chelation normalizes
reactivity to reward omission and errors in lead-exposed rats. Neurotoxicol Teratol 29: 188-202.
http://dx.doi.0rg/10.1016/i.ntt.2006.ll.004
Bechara. EJH; Medeiros. MHG: Monteiro. HP; Hermes-Lima. M; Pereira. B; Demasi. M; Costa. CA;
Abdalla. DSP: Onuki. J; Wendel. CMA; Mascio. PP. (1993). A free radical hypothesis of lead
poisoning and inborn porphyrias associated with 5-aminolevulinic acid overload. Quimica Nova 16:
385-392.
Behl. M; Zhang. Y; Zheng. W. (2009). Involvement of insulin-degrading enzyme in the clearance of beta-
amyloid at the blood-CSF barrier: Consequences of lead exposure. Cerebrospinal Fluid Research 6:
11. http://dx.doi.org/10.1186/1743-8454-6-ll
Bell. ML; Ebisu. K; Peng. RD; Samet. JM; Dominici. F. (2009). Hospital admissions and chemical
composition of fine particle air pollution. Am J Respir Crit Care Med 179: 1115-1120.
http://dx.doi.org/10.1164/rccm.200808-1240OC
Belles-Isles. M; Ayotte. P; Dewailly. E; Weber. JP; Roy. R. (2002). Cord blood lymphocyte functions in
newborns from a remote maritime population exposed to organochlorines and methylmercury. J
Toxicol Environ Health A 65: 165-182. http://dx.doi.org/10.1080/152873902753396794
Bellinger. D; Hu. H; Titlebaum. L; Needleman. HL. (1994a). Attentional correlates of dentin and bone
lead levels in adolescents. Arch Environ Health 49: 98-105.
http://dx.doi.org/10.1080/00039896.1994.9937461
Bellinger. D; Leviton. A; Allred. E; Rabinowitz. M. (1994b). Pre- and postnatal lead exposure and
behavior problems in school-aged children. Environ Res 66: 12-30.
http://dx.doi.org/10.1006/enrs.1994.1041
Bellinger. D; Leviton. A; Sloman. J. (1990). Antecedents and correlates of improved cognitive
performance in children exposed in utero to low levels of lead. Environ Health Perspect 89: 5-11.
Bellinger. D; Leviton. A; Waternaux. C: Needleman. H; Rabinowitz. M. (1987). Longitudinal analyses of
prenatal and postnatal lead exposure and early cognitive development. N Engl J Med 316: 1037-1043.
http://dx.doi.org/10.1056/NEJM198704233161701
Bellinger. D; Needleman. HL. (2003). Intellectual impairment and blood lead levels [Letter]. N Engl J
Med 349: 500. http://dx.doi.org/10.1056/NEJM200307313490515
Bellinger. D; Sloman. J: Leviton. A; Rabinowitz. M; Needleman. HL; Waternaux. C. (1991). Low-level
lead exposure and children's cognitive function in the preschool years. Pediatrics 87: 219-227.
Bellinger. D; Stiles. KM; Needleman. HL. (1992). Low-level lead exposure, intelligence and academic
achievement: A long-term follow-up study. Pediatrics 90: 855-861.
Bellinger. DC. (2004a). Assessing environmental neurotoxicant exposures and child neurobehavior:
Confounded by confounding? Epidemiology 15: 383-384.
http://dx.doi.org/10.1097/01.ede.0000129525.15064.a4
Bellinger. DC. (2004b). What is an adverse effect? A possible resolution of clinical and epidemiological
perspectives on neurobehavioral toxicity. Environ Res 95: 394-405.
http://dx.doi.0rg/10.1016/i.envres.2003.07.013
Bellinger. DC. (2007). Interpretation of small effect sizes in occupational and environmental
neurotoxicology: Individual versus population risk. Neurotoxicology 28: 245-251.
http://dx.doi.0rg/10.1016/i.neuro.2006.05.009
November 2012 5-708 Draft - Do Not Cite or Quote
-------
Bellinger. DC. (2008). Email from David Bellinger, Harvard Medical School, to Jee-Young Kim, EPA.
SUBJECT: Request for additional information on Bellinger and Needleman (2003). (EPA-HQ-
OAR-2006-0735-5156). U.S. Environmental Protection Agency.
http://www.regulations.gov/#!documentDetail:D=EPA-HQ-OAR-2006-0735-5156
Belloni-Olivi. L; Annadata. M; Goldstein. GW: Bressler. JP (1996). Phosphorylation of membrane
proteins in erythrocytes treated with lead. Biochem J 315: 401-406.
Bener. A; Almehdi. AM; Alwash. R; Al-Neamy. FRM. (200 la). A pilot survey of blood lead levels in
various types of workers in the United Arab Emirates. Environ Int 27: 311-314.
http://dx.doi.org/10.1016/S0160-4120(01)00061-7
Bener. A; Obineche. E; Gillett. M; Pasha. MAH; Bishawi. B. (200Ib). Association between blood levels
of lead, blood pressure and risk of diabetes and heart disease in workers. Int Arch Occup Environ
Health 74: 375-378. http://dx.doi.org/10.1007/s004200100231
Bennet. C: Bettaiya. R; Rajanna. S: Baker. L; Yallapragada. PR; Brice. JJ: White. SL; Bokara. KK.
(2007). Region specific increase in the antioxidant enzymes and lipid peroxidation products in the
brain of rats exposed to lead. Free Radic Res 41: 267-273.
http://dx.doi.org/10.1080/10715760600889855
Berenson. GS. (2002). Childhood risk factors predict adult risk associated with subclinical cardiovascular
disease: The Bogalusa Heart Study [Review]. Am J Cardiol 90: 3L-7L.
http://dx.doi.org/10.1016/S0002-9149(02)02953-3
Bergdahl. IA; Grubb. A; Schiitz. A; Desnick. RJ; Wetmur. JG: Sassa. S: Skerfving. S. (1997a). Lead
binding to delta-aminolevulinic acid dehydratase (ALAD) in human erythrocytes. Basic Clin
Pharmacol Toxicol 81: 153-158. http://dx.doi.Org/10.llll/i.1600-0773.1997.tb02061.x
Bergdahl. IA; Schutz. A; Gerhardsson. L; Jensen. A; Skerfving. S. (1997b). Lead concentrations in human
plasma, urine and whole blood. Scand J Work Environ Health 23: 359-363.
Bergdahl. IA; Sheveleva. M; Schutz. A; Artamonova. VG: Skerfving. S. (1998). Plasma and blood lead in
humans: Capacity-limited binding to delta-aminolevulinic acid dehydratase and other lead-binding
components. Toxicol Sci 46: 247-253. http://dx.doi.Org/10.1093/toxsci/46.2.247
Bergeret. A; Pouget. E; Tedone. R: Meygert. T; Cadot. R: Descotes. J. (1990). Neutrophil functions in
lead-exposed workers. Hum Exp Toxicol 9: 231-233. http://dx.doi.org/10.1177/096032719000900405
Berglund. AMM; Ingvarsson. PK; Danielsson. H; Nyholm. NEI. (2010). Lead exposure and biological
effects in pied flycatchers (Ficedula hypoleuca) before and after the closure of a lead mine in northern
Sweden. Environ Pollut 158: 1368-1375. http://dx.doi.Org/10.1016/j.envpol.2010.01.005
Berkowitz. Z; Price-Green. P; Bove. FJ; Kave. WE. (2006). Lead exposure and birth outcomes in five
communities in Shoshone County, Idaho. Int J Hyg Environ Health 209: 123-132.
http://dx.doi.0rg/10.1016/i.ijheh.2005.ll.001
Berrahal. AA; Lasram. M; El Elj. N; Kerkeni. A; Gharbi. N; El-Fazaa. S. (2011). Effect of age-dependent
exposure to lead on hepatotoxicity and nephrotoxicity in male rats. Environ Toxicol 26: 68-78.
http://dx.doi.org/10.1002/tox.20530
Berrahal. AA; Nehdi. A; Haijaii. N; Gharbi. N; El-Fazaa. S. (2007). Antioxidant enzymes activities and
bilirubin level in adult rat treated with lead. C R Biol 330: 581-588.
http://dx.doi.0rg/10.1016/i.crvi.2007.05.007
Berry. WD. Jr; Moriartv. CM; Lau. YS. (2002). Lead attenuation of episodic growth hormone secretion in
male rats. Int J Toxicol 21: 93-98.
Bhattacharya. A; Shukla. R: Dietrich. K; Bornschein. R; Berger. O. (1995). Effect of early lead exposure
on children's postural balance. Dev Med Child Neurol 37: 861-878.
Bhattacharva. A; Shukla. R: Dietrich. KN; Bornschein. RL. (2006). Effect of early lead exposure on the
maturation of children's postural balance: A longitudinal study. Neurotoxicol Teratol 3: 376-385.
http://dx.doi.0rg/10.1016/i.ntt.2006.02.003
November 2012 5-709 Draft - Do Not Cite or Quote
-------
Bhatti. P: Stewart. PA; Hutchinson. A; Rothman. N; Linet. MS: Inskip. PD; Rajaraman. P. (2009). Lead
exposure, polymorphisms in genes related to oxidative stress, and risk of adult brain tumors. Cancer
Epidemiol Biomarkers Prev 18: 1841-1848. http://dx.doi.org/10.1158/1055-9965.EPI-09-0197
Bielarczyk. H; Tian. X; Suszkiw. JB. (1996). Cholinergic denervation-like changes in rat hippocampus
following developmental lead exposure. Brain Res 708: 108-115.
http://dx.doi.org/10.1016/0006-8993(95)01315-6
Bihaqi. SW: Huang. H; Wu. J: Zawia. NH. (2011). Infant exposure to lead (Pb) and epigenetic
modifications in the aging primate brain: Implications for Alzheimer's disease. J Alzheimers Dis 27:
819-833. http://dx.doi.org/10.3233/JAD-2011-111013
Bilban. M. (1998). Influence of the work environment in a Pb-Zn mine on the incidence of cytogenetic
damage in miners. Am J Ind Med 34: 455-463.
http://dx.doi.org/10.1002/(SICI)1097-0274(199811)34:5<455::AID-AJIM6>3.0.CO:2-P
Billick. IH; Curran. AS: Shier. DR. (1979). Analysis of pediatric blood lead levels in New York City for
1970-1976. Environ Health Perspect 31: 183-190.
Bishayi. B; Sengupta. M. (2006). Synergism in immunotoxicological effects due to repeated combined
administration of arsenic and lead in mice. Int Immunopharmacol 6: 454-464.
http://dx.doi.0rg/10.1016/i.intimp.2005.09.011
Biswas. NM; Ghosh. PK. (2006). Protection of adrenal and male gonadal functions by androgen in lead-
treated rats. Kathmandu Univ Med J 4: 218-221.
Bitto. E; Bingman. CA; Wesenberg. GE; McCoy. JG: Phillips. GN. Jr. (2006). Structure of pyrimidine 5'-
nucleotidase type 1: Insight into mechanism of action and inhibition during lead poisoning. J Biol
Chem 281: 20521-20529. http://dx.doi.org/10.1074/ibc.M602000200
Blackman. SS. Jr. (1936). Intranuclear inclusion bodies in the kidney and liver caused by lead poisoning.
Bull Johns Hopkins Hosp 58: 384-402.
Blaklev. BR; Sisodia. CS: Mukkur. TK. (1980). The effect of methylmercury, tetraethyl lead, and sodium
arsenite on the humoral immune response in mice. Toxicol Appl Pharmacol 52: 245-254.
http://dx.doi.org/10.1016/0041-008X(80)90111-8
Bleecker. ML; Ford. DP; Celio. MA; Vaughan. CG: Lindgren. KN. (2007a). Impact of cognitive reserve
on the relationship of lead exposure and neurobehavioral performance. Neurology 69: 470-476.
http://dx.doi.org/10.1212/01.wnl.0000266628.43760.8c
Bleecker. ML; Ford. DP; Vaughan. CG: Walsh. KS: Lindgren. KN. (2007b). The association of lead
exposure and motor performance mediated by cerebral white matter change. Neurotoxicology 28:
318-323. http://dx.doi.0rg/10.1016/i.neuro.2006.04.008
Bleecker. ML; Lindgren. KN: Ford. DP. (1997). Differential contribution of current and cumulative
indices of lead dose to neuropsychological performance by age. Neurology 48: 639-645.
Bloom. MS: Louis. GM; Sundaram. R; Kostyniak. PJ; Jain. J. (2011 a). Associations between blood
metals and fecundity among women residing in New York State. Reprod Toxicol 31: 158-163.
http://dx.doi.0rg/10.1016/i.reprotox.2010.09.013
Bloom. MS: Parsons. PJ: Kim. D; Steuerwald. AJ; Vaccari. S: Cheng. G: Fujimoto. VY (20lib). Toxic
trace metals and embryo quality indicators during in vitro fertilization (IVF). Reprod Toxicol 31:
164-170. http://dx.doi.0rg/10.1016/i.reprotox.2010.ll.011
Bloom. MS: Parsons. PJ: Steuerwald. AJ: Schisterman. EF; Browne. RW: Kim. K; Coccaro. GA; Conti.
GC: Naravan. N; Fujimoto. VY. (2010). Toxic trace metals and human oocytes during in vitro
fertilization (IVF). Reprod Toxicol 29: 298-305. http://dx.doi.Org/10.1016/j.reprotox.2010.01.003
Boerrigter. G: Burnett. JC. Jr. (2009). Soluble guanylate cyclase: Not a dull enzyme [Editorial].
Circulation 119: 2752-2754. http://dx.doi.org/10.1161/CIRCULATIONAHA.109.860288
November 2012 5-710 Draft - Do Not Cite or Quote
-------
Boillee. S: Yamanaka. K; Lobsiger. CS: Copeland. NG: Jenkins. NA; Kassiotis. G: Kollias. G: Cleveland.
DW. (2006). Onset and progression in inherited ALS determined by motor neurons and microglia.
Science 312: 1389-1392. http://dx.doi.org/10.1126/science.1123511
Bokara. KK; Blaylock. I; Denise. SB; Bettaiya. R; Raj anna. S: Yallapragada. PR. (2009). Influence of
lead acetate on glutathione and its related enzymes in different regions of rat brain. J Appl Toxicol 29:
452-458. http://dx.doi.org/10.1002/iat.1423
Bolin. CM; Basha. R; Cox. D; Zawia. NH; Malonev. B; Lahiri. DK; Cardozo-Pelaez. F. (2006). Exposure
to lead (Pb) and the developmental origin of oxidative DNA damage in the aging brain. FASEB J 20:
788-790. http://dx.doi.org/10.1096/fi.05-5091fie
Bollati. V: Baccarelli. A; Hou. L; Bonzini. M; Fustinoni. S: Cavallo. D; Byun. HM; Jiang. J: Marinelli. B;
Pesatori. AC: Bertazzi. PA; Yang. AS. (2007). Changes in DNA methylation patterns in subjects
exposed to low-dose benzene. Cancer Res 67: 876-880.
http://dx.doi.org/10.1158/0008-5472.CAN-06-2995
Bonacker. D; Stoiber. T; Bohm. KJ; Prots. I; Wang. M; Unger. E; Thier. R; Bolt. HM; Degen. GH. (2005).
Genotoxicity of inorganic lead salts and disturbance of microtubule function. Environ Mol Mutagen
45: 346-353. http://dx.doi.org/10.1002/em.20100
Borgstahl. GEO; Parge. HE; Hickev. MJ; Beyer. WF. Jr; Hallewell. RA; Tainer. JA. (1992). The structure
of human mitochondria! manganese superoxide dismutase reveals a novel tetrameric interface of two
4-helix bundles. Cell 71: 107-118. http://dx.doi.org/10.1016/0092-8674(92)90270-M
Boscolo. P: Di Gioacchino. M; Sabbioni. E; Benvenuti. F; Conti. P; Reale. M; Bavazzano. P; Giuliano. G.
(1999). Expression of lymphocyte subpopulations, cytokine serum levels, and blood and urinary trace
elements in asymptomatic atopic men exposed to an urban environment. Int Arch Occup Environ
Health 72: 26-32. http://dx.doi.org/10.1007/s004200050330
Boscolo. P: Di Gioacchino. M; Sabbioni. E; Di Giacomo. F; Reale. M; Volpe. AR; Di Sciascio. MB;
Conti. P: Giuliano. G. (2000). Lymphocyte subpopulations, cytokines and trace elements in
asymptomatic atopic women exposed to an urban environment. Life Sci 67: 1119-1126.
http://dx.doi.org/10.1016/S0024-3205(00)00712-8
Bouchard. MF; Bellinger. DC: Weuve. J: Matthews-Bellinger. J: Gilman. SE; Wright. RO: Schwartz. J:
Weisskopf. MG. (2009). Blood lead levels and major depressive disorder, panic disorder, and
generalized anxiety disorder in US young adults. Arch Gen Psychiatry 66: 1313-1319.
http://dx.doi.org/10. lOOl/archgenpsvchiatry.2009.164
Boudene. C: Despaux-Pages. N; Comoy. E; Bohuon. C. (1984). Immunological and enzymatic studies of
erythrocytic delta-aminolevulinate dehydratase: Comparison of results obtained in normal and lead-
exposed subjects. Int Arch Occup Environ Health 55: 87-96. http://dx.doi.org/10.1007/BF00378071
Bound. JP; Harvey. PW: Francis. BJ; Awwad. F; Gatrell. AC. (1997). Involvement of deprivation and
environmental lead in neural tube defects: A matched case-control study. Arch Dis Child 76: 107-112.
http://dx.doi.0rg/10.1136/adc.76.2.107
Bouton. CM; Frelin. LP; Forde. CE; Arnold Godwin. H; Pevsner. J. (2001). Synaptotagmin I is a
molecular target for lead. JNeurochem 76: 1724-1735.
http://dx.doi.0rg/10.1046/i.1471-4159.2001.00168.x
Bowers. TS: Beck. BD. (2006). What is the meaning of non-linear dose-response relations between blood
lead concentrations and IQ? Neurotoxicology 27: 520-524.
http://dx.doi.0rg/10.1016/i.neuro.2006.02.001
Bragadin. M; Marton. D; Manente. S. (2007). Trialkyllead compounds induce the opening of the MTP
pore in rat liver mitochondria. J Inorg Biochem 101: 876-878.
http://dx.doi.0rg/10.1016/i.iinorgbio.2007.01.016
Bragadin. M; Marton. D; Murgia. M; Rizzoli. V; Scutari. G: Deana. R. (1998). Interactions of tri alky lie ad
compounds with rat liver mitochondria. J Inorg Biochem 69: 259-262.
http://dx.doi.org/10.1016/S0162-0134(97)10036-8
November 2012 5-711 Draft - Do Not Cite or Quote
-------
Bratton. GR; Hinev. JK; Dees. WL. (1994). Lead (Pb) alters the norepinephrine-induced secretion of
luteinizing hormone releasing hormone from the medium eminence of adult male rats in vitro. Life Sci
55: 563-571. http://dx.doi.org/10.1016/0024-3205(94)00482-X
Braude. S: Rovston. D; Coe. C: Barnes. PJ. (1984). Histamine increases lung permeability by an H2-
receptor mechanism. Lancet 2: 372-374. http://dx.doi.org/10.1016/S0140-6736(84)90542-7
Braun. JM; Froehlich. TE; Daniels. JL; Dietrich. KN; Hornung. R; Auinger. P; Lanphear. BP (2008).
Association of environmental toxicants and conduct disorder in U.S. children: NHANES 2001-2004.
Environ Health Perspect 116: 956-962. http://dx.doi.org/10.1289/ehp. 11177
Braun. JM; Kahn. RS: Froehlich. T; Auinger. P; Lanphear. BP. (2006). Exposures to environmental
toxicants and attention deficit hyperactivity disorder in U.S. children. Environ Health Perspect 114:
1904-1909. http://dx.doi.org/10.1289/ehp.9478
Bravo. Y; Quiroz. Y; Ferrebuz. A; Vaziri. ND; Rodriguez-Iturbe. B. (2007). Mycophenolate mofetil
administration reduces renal inflammation, oxidative stress, and arterial pressure in rats with lead-
induced hypertension. Am J Physiol Renal Physiol 293: F616-F623.
http://dx.doi.org/10.1152/ajprenal.00507.2006
Brender. JD; Suarez. L; Felkner. M; Gilani. Z; Stinchcomb. D; Moody. K; Henry. J; Hendricks. K. (2006).
Maternal exposure to arsenic, cadmium, lead, and mercury and neural tube defects in offspring.
Environ Res 101: 132-139. http://dx.doi.Org/10.1016/j.envres.2005.08.003
Brockel. BJ; Cory-Slechta. DA. (1998). Lead, attention, and impulsive behavior: Changes in a fixed-ratio
waiting-for-reward paradigm. Pharmacol Biochem Behav 60: 545-552.
http://dx.doi.org/10.1016/S0091-3057(98)00023-9
Brockel. BJ: Cory-Slechta. DA. (1999a). The effects of postweaning low-level Pb exposure on sustained
attention: a study of target densities, stimulus presentation rate, and stimulus predictability.
Neurotoxicology 20: 921-933.
Brockel. BJ: Cory-Slechta. DA. (1999b). Lead-induced decrements in waiting behavior: involvement of
D2-like dopamine receptors. Pharmacol Biochem Behav 63: 423-434.
Brondum. J. (2007). Environmental exposures and ADHD [Letter]. Environ Health Perspect 115: A398;
author reply A399. http://dx.doi.org/10.1289/ehp.10274
Brondum. J. (2011). ADHD, lead, and PCBs: Appropriate comparison studies [Letter]. Environ Health
Perspect 119: a282-a282. http://dx.doi.org/10.1289/ehp.1103513
Brubaker. CJ: Dietrich. KN; Lanphear. BP: Cecil. KM. (2010). The influence of age of lead exposure on
adult gray matter volume. Neurotoxicology 31: 259-266.
http://dx.doi.0rg/10.1016/i.neuro.2010.03.004
Brubaker. CJ: Schmithorst. VJ: Haynes. EN; Dietrich. KN; Egelhoff. JC: Lindquist. DM; Lanphear. BP:
Cecil. KM. (2009). Altered myelination and axonal integrity in adults with childhood lead exposure: A
diffusion tensor imaging study. Neurotoxicology 30: 867-875.
http://dx.doi.0rg/10.1016/i.neuro.2009.07.007
Bryant-Stephens. T. (2009). Asthma disparities in urban environments. J Allergy Clin Immunol 123:
1199-1206. http://dx.doi.0rg/10.1016/j.jaci.2009.04.030
Buccelletti. E; Gilardi. E; Scaini. E; Galiuto. L; Persiani. R; Biondi. A; Basile. F; Silveri. NG. (2009).
Heart rate variability and myocardial infarction: systematic literature review and metanalysis
[Review]. Eur Rev Med Pharmacol Sci 13: 299-307.
Buchanan. LH; Counter. SA; Ortega. F. (2011). Environmental lead exposure and otoacoustic emissions
in Andean children. J Toxicol Environ Health A 74: 1280-1293.
http://dx.doi.org/10.1080/15287394.2011.587106
Budtz-Jorgensen. E; Keiding. N; Grandjean. P. (2001). Benchmark dose calculation from epidemiological
data. Biometrics 57: 698-706. http://dx.doi.Org/10.llll/i.0006-341X.2001.00698.x
November 2012 5-712 Draft - Do Not Cite or Quote
-------
Bunn. TL; Dietert. RR; Ladies. GS: Holsapple. MR (200la). Developmental immunotoxicology
assessment in the rat: Age, gender, and strain comparisons after exposure to Pb. Toxicol Methods 11:
41-58. http://dx.doi.org/10.1080/105172301300055151
Bunn. TL; Marsh. JA; Dietert. RR. (2000). Gender differences in developmental immunotoxicity to lead
in a chicken: Analysis following a single early low-level exposure in ovo. J Toxicol Environ Health A
61: 677-693. http://dx.doi.org/10.1080/00984100050195152
Bunn. TL; Parsons. PJ: Kao. E; Dietert. RR. (200Ib). Exposure to lead during critical windows of
embryonic development: Differential immunotoxic outcome based on stage of exposure and gender.
Toxicol Sci 64: 57-66. http://dx.doi.Org/10.1093/toxsci/64.l.57
Bunn. TL; Parsons. PJ: Kao. E; Dietert. RR. (2001c). Gender-based profiles of developmental
immunotoxicity to lead in the rat: Assessment in juveniles and adults. J Toxicol Environ Health A 64:
223-240. http://dx.doi.org/10.1080/15287390152543708
Burger. J: Gochfeld. M. (2005). Effects of lead on learning in herring gulls: An avian wildlife model for
neurobehavioral deficits. Neurotoxicology 26: 615-624. http://dx.doi.Org/10.1016/i.neuro.2005.01.005
Burns. JM; Baghurst PA; Sawyer. MG: McMichael. AJ; Tong. SL. (1999). Lifetime low-level exposure
to environmental lead and children's emotional and behavioral development at ages 11-13 years: The
Port Pirie cohort study. Am J Epidemiol 149: 740-749.
Bussolaro. D; Neto. FF; Gargioni. R; Fernandes. LC: Randi. MAP; Pelletier. E; Oliveira Ribeiro. CA.
(2008). The immune response of peritoneal macrophages due to exposure to inorganic lead in the
house mouse Mus musculus. Toxicol In Vitro 22: 254-260. http://dx.doi.Org/10.1016/i.tiv.2007.09.003
Cabaravdic. M; Mijanovic. M; Kusturica. J: Cabaravdic. A. (2010). Occupational exposure of workers at
gas station to inorganic lead. Med Arh 64: 107-109.
Caffo. B; Chen. S: Stewart. W: Bolla. K; Yousem. D; Davatzikos. C: Schwartz. BS. (2008). Are brain
volumes based on magnetic resonance imaging mediators of the associations of cumulative lead dose
with cognitive function? Am J Epidemiol 167: 429-437. http://dx.doi.org/10.1093/aje/kwm326
Cai. MY; Arenaz. P. (1998). Antimutagenic effect of crown ethers on heavy metal-induced sister
chromatid exchanges. Mutagenesis 13: 27-32. http://dx.doi.Org/10.1093/mutage/13.l.27
Calderon-Salinas. JV: Quintanar-Escorcia. MA; Gonzalez-Martinez. MT; Hernandez-Luna. CE. (1999a).
Lead and calcium transport in human erythrocyte. Hum Exp Toxicol 18: 327-332.
http://dx.doi.org/10.1191/096032799678840138
Calderon-Salinas. JV: Quintanar-Escorza. MA; Hernandez-Luna. CE; Gonzalez-Martinez. MT. (1999b).
Effect of lead on the calcium transport in human erythrocyte. Hum Exp Toxicol 18: 146-153.
http://dx.doi.org/10.1177/096032719901800303
Calvo. FB; Santos. D: Rodrigues. CJ: Krug. FJ; Marumo. JT; Schor. N; Bellini. MH. (2009). Variation in
the distribution of trace elements in renal cell carcinoma. Biol Trace Elem Res 130: 107-113.
htrp://dx.doi.org/10.1007/sl2011-009-8325-x
Camoratto. AM; White. LM; Lau. YS: Ware. GO: Berry. WD; Moriartv. CM. (1993). Effect of exposure
to low level lead on growth and growth hormone release in rats. Toxicology 83: 101-114.
http://dx.doi.org/10.1016/0300-483X(93)90095-A
Campagna. D: Huel. G: Hellier. G: Girard. F; Sahuquillo. J: Fagot-Campagna. A; Godin. J: Blot. P.
(2000). Negative relationships between erythrocyte Ca-pump activity and lead levels in mothers and
newborns. Life Sci 68: 203-215. http://dx.doi.org/10.1016/S0024-3205(00)00928-0
Campbell. JR; Auinger. P. (2007). The association between blood lead levels and osteoporosis among
adults-results from the Third National Health and Nutrition Examination Survey (NHANES III).
Environ Health Perspect 115: 1018-1022. http://dx.doi.org/10.1289/ehp.9716
November 2012 5-713 Draft - Do Not Cite or Quote
-------
Camyre. E; Wise. SS: Milligan. P; Gordon. N; Goodale. B; Stackpole. M; Patzlaff N; Aboueissa. AM;
Wise. JP. Sr. (2007). Ku80 deficiency does not affect paniculate chromate-induced chromosome
damage and cytotoxicity in Chinese hamster ovary cells. Toxicol Sci 97: 348-354.
http://dx.doi.org/10.1093/toxsci/kfm045
Can. S: Bagci. C: Ozaslan. M; Bozkurt. AI; Cengiz. B; Cakmak. EA; Kocabas. R; Karadag. E;
Tarakcioglu. M. (2008). Occupational lead exposure effect on liver functions and biochemical
parameters. Acta Physiol Hung 95: 395-403. http://dx.doi.Org/10.1556/APhysiol.95.2008.4.6
Canfield. R. (2008). Email from Dr. Richard Canfield, Cornell University, to Jee-Young Kim, USEPA
Subject: Request for additional information on Canfield et al. (2003). (EPA-HQ-
OAR-2006-0735-5811). U.S. Environmental Protection Agency.
http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR-2006-0735-5811
Canfield. RL; Gendle. MH; Cory-Slechta. DA. (2004). Impaired neuropsychological functioning in lead-
exposed children. Dev Neuropsychol 26: 513-540. http://dx.doi.org/10.1207/sl5326942dn2601 8
Canfield. RL; Henderson. CR. Jr; Cory-Slechta. DA; Cox. C: Jusko. TA; Lanphear. BP (2003a).
Intellectual impairment in children with blood lead concentrations below 10 micrograms per deciliter.
NEnglJMed348: 1517-1526. http://dx.doi.org/10.1056/NEJMoa022848
Canfield. RL: Kreher. DA; Cornwell. C: Henderson. CR. Jr. (2003b). Low-level lead exposure, executive
functioning, and learning in early childhood. Child Neuropsychol 9: 35-53.
http://dx.doi.org/10.1076/chin.9.1.35.14496
Cantarow. A; Trumper. M. (1944). Lead poisoning. Hagerstown, MD: Williams & Wilkins Co.
Cantonwine. D; Hu. H; Sanchez. BN; Lamadrid-Figueroa. H; Smith. D; Ettinger. AS: Mercado-Garcia. A;
Hernandez-Avila. M; Wright. RO: Tellez-Rojo. MM. (2010a). Critical windows of fetal lead exposure:
Adverse impacts on length of gestation and risk of premature delivery. J Occup Environ Med 52:
1106-1111. http://dx.doi.org/10.1097/JOM.Ob013e3181f86fee
Cantonwine. D: Hu. H; Tellez-Rojo. MM; Sanchez. BN; Lamadrid-Figueroa. H; Ettinger. AS: Mercado-
Garcia. A; Hernandez-Avila. M; Wright. RO. (2010b). HFE gene variants modify the association
between maternal lead burden and infant birthweight: A prospective birth cohort study in Mexico City,
Mexico. Environ Health Global Access Sci Source 9: 43. http://dx.doi.org/10.1186/1476-069X-9-43
Cao. XJ; Huang. SH; Wang. M; Chen. JT; Ruan. DY. (2008). S-adenosyl-L-methionine improves
impaired hippocampal long-term potentiation and water maze performance induced by developmental
lead exposure in rats. Eur J Pharmacol 595: 30-34. http://dx.doi.Org/10.1016/i.ejphar.2008.07.061
Cao. XJ; Wang. M; Chen. WH; Zhu. DM; She. JO: Ruan. DY. (2009). Effects of chronic administration of
melatonin on spatial learning ability and long-term potentiation in lead-exposed and control rats.
Biomed Environ Sci 22: 70-75. http://dx.doi.org/10.1016/S0895-3988(09)60025-8
Carafoli. E. (2005). The Theodor Bucher Lecture - Calcium - a universal carrier of biological signals.
Delivered on 3 July 2003 at the Special FEES Meeting in Brussels. FEES J 272: 1073-1089.
http://dx.doi.0rg/10.llll/i.1742-4658.2005.04546.x
Cardenas. A; Roels. H; Bernard. AM; Barbon. R; Buchet. JP: Lauwerys. RR; Rosello. J: Ramis. I; Mutti.
A; Franchini. I; Pels. LM; Stolte. H; De Broe. ME; Nuyts. GD; Taylor. SA; Price. RG. (1993).
Markers of early renal changes induced by industrial pollutants: II Application to workers exposed to
lead. Br J Ind Med 50: 28-36. http://dx.doi.Org/10.1136/oem.50.l.28
Carey. JB; Allshire. A; Van Pelt. FN. (2006). Immune modulation by cadmium and lead in the acute
reporter antigen-popliteal lymph node assay. Toxicol Sci 91: 113-122.
http://dx.doi.org/10.1093/toxsci/kfi 142
Carmignani. M; Boscolo. P; Poma. A; Volpe. AR. (1999). Kininergic system and arterial hypertension
following chronic exposure to inorganic lead. Int Immunopharmacol 44: 105-110.
http://dx.doi.org/10.1016/S0162-3109(99)00115-0
November 2012 5-714 Draft - Do Not Cite or Quote
-------
Carmignani. M; Volpe. AR; Boscolo. P; Qiao. N; MDi. G: Grilli. A; Felaco. M. (2000). Catcholamine and
nitric oxide systems as targets of chronic lead exposure in inducing selective functional impairment.
Life Sci 68: 401-415. http://dx.doi.org/10.1016/S0024-3205(00)00954-l
Carmona. ER; Creus. A; Marcos. R. (2011). Genotoxicity testing of two lead-compounds in somatic cells
of Drosophila melanogaster. Mutat Res 724: 35-40. http://dx.doi.Org/10.1016/i.mrgentox.2011.05.008
Carreras. HA; Wannaz. ED; Pignata. ML. (2009). Assessment of human health risk related to metals by
the use of biomonitors in the province of Cordoba, Argentina. Environ Pollut 157: 117-122.
http://dx.doi.0rg/10.1016/i.envpol.2008.07.018
Casado. MF; Cecchini. AL; Simao. ANC: Oliveira. RD; Cecchini. R. (2007). Free radical-mediated pre-
hemolytic injury in human red blood cells subjected to lead acetate as evaluated by
chemiluminescence. Food Chem Toxicol 45: 945-952. http://dx.doi.Org/10.1016/j.fct.2006.12.001
Castranova. V: Bowman. L; Reasor. MJ; Miles. PR. (1980). Effects of heavy metal ions on selected
oxidative metabolic processes in rat alveolar macrophages. Toxicol Appl Pharmacol 53: 14-23.
http://dx.doi.org/10.1016/0041-008X(80)90375-0
Caylak. E; Aytekin. M; Halifeoglu. I. (2008). Antioxidant effects of methionine, alpha-lipoic acid, N-
acetylcysteine and homocysteine on lead-induced oxidative stress to erythrocytes in rats. Exp Toxicol
Pathol 60: 289-294. http://dx.doi.Org/10.1016/i.etp.2007.ll.004
CDC (Centers for Disease Control and Prevention). (2004). The health consequences of smoking: A
report of the Surgeon General. Washington, DC: U.S. Department of Health and Human Services.
http ://www. surgeongeneral.gov/librarv/smokingconsequences/
CDC (Centers for Disease Control and Prevention). (20 lib). State-based prevalence data of ADHD
diagnosis. Atlanta, GA. Retrieved from http://www.cdc.gov/ncbddd/adhd/prevalence.html
CDC (Centers for Disease Control and Prevention). (2012). Tested and confirmed elevated blood lead
levels by state, year and blood lead level group for children <72 months. Atlanta, GA. Retrieved from
http://www.cdc.gov/nceh/lead/data/national.htm
Cecil. K; Yuan. W: Holland. S: Wessel. S: Dietrich. K; Ris. D; Lanphear. B. (2005). The influence of
childhood lead exposure on language function in young adults: An FMRI study [Abstract]. Proc Int
Soc Magn Reson Med 13: 1441.
Cecil. KM; Brubaker. CJ: Adler. CM; Dietrich. KN; Altave. M; Egelhoff. JC: Wessel. S: Elangovan. I;
Hornung. R; Jarvis. K; Lanphear. BP (2008). Decreased brain volume in adults with childhood lead
exposure. PLoS Med 5: el 12. http://dx.doi.org/10.1371/iournal.pmed.0050112
Cecil. KM; Dietrich. KN; Altave. M; Egelhoff. JC: Lindquist. DM; Brubaker. CJ: Lanphear. BP. (2011).
Proton magnetic resonance spectroscopy in adults with childhood lead exposure. Environ Health
Perspect 119: 403-408. http://dx.doi.org/10.1289/ehp.1002176
Celik. A; Ogenler. O; Comelekoglu. U. (2005). The evaluation of micronucleus frequency by acridine
orange fluorescent staining in peripheral blood of rats treated with lead acetate. Mutagenesis 20:
411-415. http://dx.doi.org/10.1093/mutage/gei055
Cerulli. N; Campanella. L; Grossi. R: Politi. L; Scandurra. R: Soda. G: Gallo. F; Damiani. S: Alimonti. A;
Petrucci. F; Caroli. S. (2006). Determination of Cd, Cu, Pb and Zn in neoplastic kidneys and in renal
tissue of fetuses, newborns and corpses. J Trace Elem Med Biol 20: 171-179.
http://dx.doi.0rg/10.1016/i.itemb.2006.03.002
Chai. S: Webb. RC. (1988). Effects of lead on vascular reactivity [Review]. Environ Health Perspect 78:
85-89.
Chakrabortv. I; Sharma. A; Talukder. G. (1987). Antagonistic and synergistic effects of lead and selenium
in Rattus norvegicus. Toxicol Lett 37: 21-26. http://dx.doi.org/10.1016/0378-4274(87)90162-7
Chancellor. AM; Slatterv. JM; Fraser. H; Warlow. CP (1993). Risk factors for motor neuron disease: A
case-control study based on patients from the Scottish Motor Neuron Disease Register. J Neurol
Neurosurg Psychiatry 56: 1200-1206. http://dx.doi.org/10.1136/innp.56.ll.1200
November 2012 5-715 Draft - Do Not Cite or Quote
-------
Chandramouli. K; Steer. CD; Ellis. M; Emond. AM. (2009). Effects of early childhood lead exposure on
academic performance and behaviour of school age children. Arch Dis Child 94: 844-848.
http://dx.doi.org/10.1136/adc.2008.149955
Chang. HR: Chen. SS: Tsao. DA: Cheng. JT: Ho. CK: Yu. HS. (1997). Change of cardiac beta-
adrenoceptors in lead-exposed rats. Toxicology 123: 27-32.
httrj://dx.doi.org/10.1016/S0300-483X(97)00100-5
Chang. HR; Tsao. DA; Yu. HS: Ho. CK. (2005). The change of "beta"-adrenergic system after cessation
of lead exposure. Toxicology 207: 73-80. http://dx.doi.Org/10.1016/i.tox.2004.08.018
Chang. MK; Raggatt. LJ; Alexander. KA; Kuliwaba. JS: Fazzalari. NL; Schroder. K; Maylin. ER; Ripoll.
VM; Hume. DA; Pettit AR. (2008a). Osteal tissue macrophages are intercalated throughout human
and mouse bone lining tissues and regulate osteoblast function in vitro and in vivo. J Immunol 181:
1232-1244.
Chang. SH: Cheng. BH: Lee. SL: Chuang. HY: Yang. CY: Sung. FC: Wu. TN. (2006). Low blood lead
concentration in association with infertility in women. Environ Res 101: 380-386.
http://dx.doi.0rg/10.1016/i.envres.2005.10.004
Chang. WC: Chang. CC: Wang. YS: Wang. YS: Weng. WT; Yoshioka. T; Juo. SH. (2011). Involvement of
the epidermal growth factor receptor in Pb(2+)-induced activation of cPLA(2)/COX-2 genes and
PGE(2) production in vascular smooth muscle cells. Toxicology 279: 45-53.
http://dx.doi.0rg/10.1016/i.tox.2010.09.004
Chang. YF: Teng. HC: Cheng. SY: Wang. CT: Chiou. SH: Kao. LS: Kao. FJ: Chiou. A: Yang. DM.
(2008b). Orail-STIMl formed store-operated Ca[sub]2+[/sub] channels (SOCs) as the molecular
components needed for Pb[sub]2+[/sub] entry in living cells. Toxicol Appl Pharmacol 227: 430-439.
http://dx.doi.0rg/10.1016/i.taap.2007.ll.025
Chao. SL: Moss. JM; Harry. GJ. (2007). Lead-induced alterations of apoptosis and neurotrophic factor
mRNA in the developing rat cortex, hippocampus, and cerebellum. J Biochem Mol Toxicol 21:
265-272. http://dx.doi.org/10.1002/ibt.20191
Chaurasia. SS: Gupta. P: Maiti. PK; Kar. A. (1998). Possible involvement of lipid peroxidation in the
inhibition of type I iodothyronine 5'-monodeiodinase activity by lead in chicken liver. J Appl Toxicol
18: 299-300. http://dx.doi.org/10.1002/(SICI)1099-1263(199807/08)18:4<299::AID-
JAT493>3.0.CO:2-J
Chaurasia. SS: Kar. A. (1997). Protective effects of vitamin E against lead-induced deterioration of
membrane associated type-I iodothyronine 5'-monodeiodinase (5'D-I) activity in male mice.
Toxicology 124: 203-209. http://dx.doi.org/10.1016/S0300-483X(97)00155-8
Chen. A; Cai. B; Dietrich. KN; Radcliffe. J: Rogan. WJ. (2007). Lead exposure, IQ, and behavior in urban
5- to 7-year-olds: Does lead affect behavior only by lowering IQ? Pediatrics 119: e650-e658.
http://dx.doi.org/10.1542/peds.2006-1973
Chen. A; Dietrich. KN; Ware. JH; Radcliffe. J: Rogan. WJ. (2005). IQ and blood lead from 2 to 7 years of
age: Are the effects in older children the residual of high blood lead concentrations in 2-year-olds?
Environ Health Perspect 113: 597-601. http://dx.doi.org/10.1289/ehp.7625
Chen. B: Lamberts. LV: Behets. GJ: Zhao. TT: Zhou. MX: Liu. G: Hou. XH: Guan. GJ: D'Haese. PC.
(2009a). Selenium, lead, and cadmium levels in renal failure patients in China. Biol Trace Elem Res
131: 1-12. http://dx.doi.org/10.1007/sl2011-009-8340-v
Chen. HH; Chan. MH. (2002). Developmental lead exposure differentially alters the susceptibility to
chemoconvulsants in rats. Toxicology 173: 249-257. http://dx.doi.org/10.1016/S0300-483X(02)00032-
X
Chen. HI; Chiu. YW: Hsu. YK; Li. WF; Chen. YC: Chuang. HY. (2010a). The association of
metallothionein-4 gene polymorphism and renal function in long-term lead-exposed workers. Biol
Trace Elem Res 137: 55-62. http://dx.doi.org/10.1007/sl2011-009-8564-x
November 2012 5-716 Draft - Do Not Cite or Quote
-------
Chen. J; Mercer. G: Rotfa. SR; Abraham. L; Lutz. P; Ercal. N; Neal. RE. (20lib). Sub-chronic lead
exposure alters kidney proteome profiles. Hum Exp Toxicol 30: 1616-1625.
http://dx.doi.org/10.1177/0960327110396521
Chen. PC: Pan. IJ; Wang. JD. (2006a). Parental exposure to lead and small for gestational age births. Am
J Ind Med 49: 417-422. http://dx.doi.org/10.1002/aiim.20313
Chen. PS: Tan. AY. (2007). Autonomic nerve activity and atrial fibrillation [Review]. Heart Rhythm 4:
S61-S64. http://dx.doi.0rg/10.1016/i.hrthm.2006.12.006
Chen. S: Golemboski. KA; Piepenbrink. M; Dietert. RR. (2004). Developmental immunotoxicity of lead
in the rat: Influence of maternal diet. J Toxicol Environ Health A 67: 495-511.
http://dx.doi.org/10.1080/15287390490276520
Chen. S: Golemboski. KA; Sanders. FS: Dietert. RR. (1999). Persistent effect of in utero meso-2,3-
dimercaptosuccinic acid (DMSA) on immune function and lead-induced immunotoxicity. Toxicology
132: 67-79. http://dx.doi.org/10.1016/S0300-483X(98)00139-5
Chen. S: Miller. TE; Golemboski. KA: Dietert. RR. (1997). Suppression of macrophage metabolite
production by lead glutamate in vitro is reversed by meso-2,3-dimercaptosuccinic acid (DMSA). In
Vitro Toxicol 10: 351-358.
Chen. XK; Yang. Q; Smith. G: Krewski. D; Walker. M; Wen. SW. (2006c). Environmental lead level and
pregnancy-induced hypertension. Environ Res 100: 424-430.
http://dx.doi.0rg/10.1016/i.envres.2005.07.006
Chen. Y; Zhao. JX; Liu. JW: Cui. J: Li. L; Tian. W. (2008c). Lack of association of delta-aminolevulinic
acid dehydratase genotype with blood lead levels in environmentally exposed children of Uygur and
Han populations. Acta Paediatr 97: 1717-1720. http://dx.doi.Org/10.llll/j.1651-2227.2008.01003.x
Cheng. Y; Schwartz. J: Sparrow. D; Aro. A; Weiss. ST; Hu. H. (2001). Bone lead and blood lead levels in
relation to baseline blood pressure and the prospective development of hypertension: The Normative
Aging Study. Am J Epidemiol 153: 164-171. http://dx.doi.Org/10.1093/aje/153.2.164
Cheng. Y; Schwartz. J: Vokonas. PS: Weiss. ST; Aro. A; Hu. H. (1998). Electrocardiographic conduction
disturbances in association with low-level lead exposure (the Normative Aging Study). Am J Cardiol
82: 594-599. http://dx.doi.org/10.1016/S0002-9149(98)00402-0
Cheng. YJ; Yang. BC: Hsieh. WC: Huang. BM; Liu. MY. (2002). Enhancement of TNF-alpha expression
does not trigger apoptosis upon exposure of glial cells to lead and lipopolysaccharide. Toxicology 178:
183-191. http://dx.doi.org/10.1016/S0300-483X(02)00225-l
Cheng. YJ; Yang. BC: Liu. MY. (2006). Lead increases lipopolysaccharide-induced liver injury through
tumor necrosis factor-alpha overexpression by monocytes/macrophages: Role of protein kinase C and
p42/44 mitogen-activated protein kinase. Environ Health Perspect 114: 507-513.
http://dx.doi.org/10.1289/ehp.8550
Chettv. CS: Reddv. GR; Murthy. KS: Johnson. J: Saiwan. K; Desaiah. D. (2001). Perinatal lead exposure
alters the expression of neuronal nitric oxide synthase in rat brain. Int J Toxicol 20: 113-120.
http://dx.doi.org/10.1080/109158101317097692
Chettv. CS: Vemuri. MC: Campbell. K; Suresh. C. (2005). Lead-induced cell death of human
neuroblastoma cells involves GSH deprivation. Cell Mol Biol Lett 10: 413-423.
Chia. SE: Zhou. H: Tham. MT: Yap. E: Dong. NV: Tu. NH: Chia. KS. (2005). Possible influence of
"delta"-aminolevulinic acid dehydratase polymorphism and susceptibility to renal toxicity of lead: A
study of a Vietnamese population. Environ Health Perspect 113: 1313-1317.
http://dx.doi.org/10.1289/ehp.7904
Chia. SE; Zhou. HJ; Theng. TM; Yap. E. (2007). Possibilities of newer ALAD polymorphism influencing
human susceptibility to effects of inorganic lead on the neurobehavioral functions. Neurotoxicology
28: 312-317. http://dx.doi.Org/10.1016/i.neuro.2006.04.003
November 2012 5-717 Draft - Do Not Cite or Quote
-------
Chia. SE; Zhou. HJ; Yap. E; Tham. MT; Dong. NV; Hong Tu. NT; Chia. KS. (2006). Association of renal
function and delta-aminolevulinic acid dehydratase polymorphism among Vietnamese and Singapore
workers exposed to inorganic lead. Occup Environ Med 63: 180-186.
http://dx.doi.org/10.1136/oem.2005.021154
Chiodo. LM; Covington. C: Sokol. RJ; Hannigan. JH; Jannise. J: Ager. J; Greenwald. M; Delanev-Black.
V (2007). Blood lead levels and specific attention effects in young children. Neurotoxicol Teratol 29:
538-546. http://dx.doi.0rg/10.1016/i.ntt.2007.04.001
Chiodo. LM; Jacobson. SW: Jacobson. JL. (2004). Neurodevelopmental effects of postnatal lead exposure
at very low levels. Neurotoxicol Teratol 26: 359-371. http://dx.doi.Org/10.1016/i.ntt.2004.01.010
Chiu. TY; Teng. HC: Huang. PC: Kao. FJ; Yang. DM. (2009). Dominant role of Orail with STIM1 on the
cytosolic entry and cytotoxicity of lead ions. Toxicol Sci 110: 353-362.
http://dx.doi.org/10.1093/toxsci/kfp099
Cho. SC: Kim. BN: Hong. YC: Shin. MS: Yoo. HJ: Kim. JW: Bhang. SY: Cho. IH: Kim. HW. (2010).
Effect of environmental exposure to lead and tobacco smoke on inattentive and hyperactive symptoms
and neurocognitive performance in children. J Child Psychol Psychiatry 51: 1050-1057.
http://dx.doi.0rg/10.llll/i.1469-7610.2010.02250.x
Chobanian. AV; Bakris. GL; Black. HR; Cushman. WC: Green. LA; Izzo. JL. Jr; Jones. DW: Materson.
BJ; Oparil. S: Wright. JT. Jr; Roccella. EJ. (2003). Seventh report of the Joint National Committee on
Prevention, Detection, Evaluation, and Treatment of High Blood Pressure. Hypertension 42:
1206-1252. http://dx.doi.org/10.1161/01.HYP.0000107251.49515.c2
Choie. DP; Richter. GW. (1972). Lead poisoning: Rapid formation of intranuclear inclusions. Science
177: 1194-1195. http://dx.doi.org/10.1126/science.177.4055.1194
Choie. DP; Puchter. GW: Young. LB. (1975). Biogenesis of intranuclear lead-protein inclusions in mouse
kidney. Pathol Res Pract 155: 197-203.
Chou. YH; Woon. PY; Huang. WC: Shiurba. R; Tsai. YT; Wang. YS: Hsieh. TJ; Chang. WC: Chuang.
HY; Chang. WC. (2011). Divalent lead cations induce cyclooxygenase-2 gene expression by
epidermal growth factor receptor/nuclear factor-kappa B signaling in A43 Icarcinoma cells. Toxicol
Lett 203: 147-153. http://dx.doi.Org/10.1016/i.toxlet.2011.03.017
Chuang. HY: Kuo. CH: Chiu. YW: Ho. CK: Chen. CJ: Wu. TN. (2007). A case-control study on the
relationship of hearing function and blood concentrations of lead, manganese, arsenic, and selenium.
Sci Total Environ 387: 79-85. http://dx.doi.Org/10.1016/i.scitotenv.2007.07.032
Chuang. HY; Schwartz. J: Gonzales-Cossio. T; Lugo. MC: Palazuelos. E; Aro. A; Hu. H; Hernandez-
Avila. M. (2001). Interrelations of lead levels in bone, venous blood, and umbilical cord blood with
exogenous lead exposure through maternal plasma lead in peripartum women. Environ Health
Perspect 109: 527-532. http://dx.doi.org/10.1289/ehp.01109527
Church. HJ: Day. JP; Braithwaite. RA; Brown. SS. (1993a). Binding of lead to a metallothionein-like
protein in human erythrocytes. J Inorg Biochem 49: 55-68.
http://dx.doi.org/10.1016/0162-0134(93)80048-E
Church. HJ: Day. JP: Braithwaite. RA; Brown. SS. (1993b). The speciation of lead in erythrocytes in
relation to lead toxicity: Case studies of two lead-exposed workers. Neurotoxicology 14: 359-364.
Ciubar. R; Ciofrangeamu C: Mitran. V; Cimpean. A; Stanescu. A; lordachescu. D. (2007). The effect of
lead ions on human erythrocytes in vitro. Rev Chim 58: 895-897.
Clark. CS: Bornschein. RL; Succop. P; Que Hee. SS: Hammond. PB; Peace. B. (1985). Condition and
type of housing as an indicator of potential environmental lead exposure and pediatric blood lead
levels. Environ Res 38: 46-53. http://dx.doi.org/10.1016/0013-9351(85)90071-4
November 2012 5-718 Draft - Do Not Cite or Quote
-------
Glaus Henn. B; Schnaas. L; Ettinger. AS: Schwartz. J; Lamadrid-Figueroa. H; Hernandez-Avila. M;
Amarasiriwardena. C: Hu. H; Bellinger. DC: Wright. RO: Tellez-Rojo. MM. (2012). Associations of
early childhood manganese and lead co-exposure with neurodevelopment. Environ Health Perspect
120: 126-131. http://dx.doi.org/10.1289/ehp.1003300
Cline. HT; Witte. S: Jones. KW. (1996). Low lead levels stunt neuronal growth in a reversible manner.
PNAS 93: 9915-9920.
Coban. TA; Senturk. M; Ciftci. M; Kufrevioglu. OI. (2007). Effects of some metal ions on human
erythrocyte glutathione reductase: an in vitro study. Protein Pept Lett 14: 1027-1030.
http://dx.doi.org/10.2174/092986607782541060
Cocco. P: Fadda. D; Atzeri. S: Avataneo. G: Meloni. M; Flore. C. (2007). Causes of death among lead
smelters in relation to the glucose-6-phosphate dehydrogenase polymorphism. Occup Environ Med
64: 414-416. http://dx.doi.org/10.1136/oem.2006.028779
Cockcroft. DW: Gault. MH. (1976). Prediction of creatinine clearance from serum creatinine. Nephron
16:31-41.
Cohen. BS: Li. W: Xiong. JQ; Lippmann. M. (2000). Detecting H+ in ultrafine ambient aerosol using iron
nano-film detectors and scanning probe microscopy. Appl Occup Environ Hyg 15: 80-89.
http://dx.doi.org/10.1080/104732200301881
Cohen. N; Modai. D; Golik. A; Weissgarten. J; Peller. S: Katz. A; Averbukh. Z; Shaked. U. (1989).
Increased concanavalin A-induced suppressor cell activity in humans with occupational lead exposure.
Environ Res 48: 1-6. http://dx.doi.org/10.1016/80013-9351(89)80079-9
Columbano. A; Endoh. T; Denda. A; Noguchi. O; Nakae. D: Hasegawa. K; Ledda-Columbano. GM;
Zedda. AI; Konishi. Y. (1996). Effects of cell proliferation and cell death (apoptosis and necrosis) on
the early stages of rat hepatocarcinogenesis. Carcinogenesis 17: 395-400.
http://dx.doi.0rg/10.1093/carcin/17.3.395
Columbano. A; Ledda-Columbano. GM; Coni. P; Pani. P. (1987). Failure of mitogen-induced cell
proliferation to achieve initiation of rat liver carcinogenesis. Carcinogenesis 8: 345-347.
http://dx.doi.Org/10.1093/carcin/8.2.345
Columbano. A; Ledda-Columbano. GM: Ennas. MG: Curto. M; Chelo. A; Pani. P. (1990). Cell
proliferation and promotion of rat liver carcinogenesis: Different effect of hepatic regeneration and
mitogen induced hyperplasia on the development of enzyme-altered foci. Carcinogenesis 11: 771-776.
http://dx.doi.0rg/10.1093/carcin/ll.5.771
Company. R; Serafim. A; Lopes. B; Cravo. A; Kalman. J: Riba. I; DelValls. TA; Blasco. J: Delgado. J:
Sarmiento. AM; Nieto. JM; Shepherd. TJ; Nowell. G: Bebianno. MJ. (2011). Source and impact of
lead contamination on delta-aminolevulinic acid dehydratase activity in several marine bivalve species
along the Gulf of Cadiz. AquatToxicol 101: 146-154. http://dx.doi.Org/10.1016/i.aquatox.2010.09.012
Comper. WD; Russo. LM. (2009). The glomerular filter: An imperfect barrier is required for perfect renal
function [Review]. Curr Opin Nephrol Hypertens 18: 336-342.
http://dx.doi.org/10.1097/Mnh.Ob013e32832cb96a
Coni. P; Pichiri-Coni. G: Ledda-Columbano. GM: Semple. E; Raialakshmi. S: Rao. PM; Sarma. DSR;
Columbano. A. (1992). Stimulation of DNA synthesis by rat plasma following in vivo treatment with
three liver mitogens. Cancer Lett 61: 233-238. http://dx.doi.org/10.1016/0304-3835(92)90293-5
Conroy. MC: Adkinson. NF. Jr; Lichtenstein. LM. (1977). Measurement of IgE on human basophils:
Relation to serum IgE and anti-IgE-induced histamine release. J Immunol 118: 1317-1321.
Conterato. GMM; Augusti. PR; Somacal. S: Einsfeld. L; Sobieski. R; Torres. JRV: Emanuelli. T. (2007).
Effect of lead acetate on cytosolic thioredoxin reductase activity and oxidative stress parameters in rat
kidneys. Basic Clin Pharmacol Toxicol 101: 96-100.
http://dx.doi.0rg/10.llll/i.1742-7843.2007.00084.x
November 2012 5-719 Draft - Do Not Cite or Quote
-------
Conterato. GMM; Bulcao. RP; Sobieski. R; Moro. AM; Charao. MF; de Freitas. FA; de Almeida. FL;
Moreira. APL; Roehrs. M; Tonello. R; Batista. BL; Grotto. D; Barbosa. F. Jr; Garcia. SC; Emanuelli.
T\ (In Press) Blood thioredoxin reductase activity, oxidative stress and hematological parameters in
painters and battery workers: Relationship with lead and cadmium levels in blood. J Appl Toxicol.
http://dx.doi.org/10.1002/iat. 1731
Cook. JA; Hoffmann. EO; Di Luzio. NR. (1975). Influence of lead and cadmium on the susceptibility of
rats to bacterial challenge. Proc Soc Exp Biol Med 150: 741-747.
Coon. S: Stark. A; Peterson. E; Gloi. A; Kortsha. G: Pounds. J: Chettle. D; Gorell. J. (2006). Whole-body
lifetime occupational lead exposure and risk of Parkinson's disease. Environ Health Perspect 114:
1872-1876. http://dx.doi.org/10.1289/ehp.9102
Coonev. G: Bell. A; Stavrou. C. (1991). Low level exposures to lead and neurobehavioural development:
The Sydney study at seven years. In JG Farmer (Ed.), International conference: Heavy metals in the
environment, Vol 1; Edinburgh - September 1991 (pp. 16-19). Edinburgh, United Kingdom: CEP
Consultants, Ltd.
Coonev. GH; Bell. A; McBride. W: Carter. C. (1989a). Low-level exposures to lead: the Sydney lead
study. Dev Med Child Neural 31: 640-649.
Coonev. GH; Bell. A; McBride. W: Carter. C. (1989b). Neurobehavioural consequences of prenatal low
level exposures to lead. Neurotoxicol Teratol 11: 95-104.
Cooper. GP; Manalis. RS. (1984). Interactions of lead and cadmium on acetylcholine release at the frog
neuromuscular junction. Toxicol Appl Pharmacol 74: 411-416.
http://dx.doi.org/10.1016/0041-008X(84)90294-l
Coratelli. P; Giannattasio. M; Lomonte. C: Marzolla. R: Rana. F; L'Abbate. N. (1988). Enzymuria to
detect tubular injury in workers exposed to lead: A 12-month follow-up. In C Bianchi; V Bocci; FA
Carone; R Rabkin (Eds.), Kidney and proteins in health and disease: Fifth international symposium in
health and disease; July 1987; Montecatini Terme, Italy (pp. 207-211). Basel, Switzerland: S. Karger.
Coria. C; Cabello. A; Tassara. E; Lopez. E; Rosales. H; Perez. M; Zavala. C; Munoz. P; Orellana. G;
Inostroza. MI; Contreras. L; Kirsten. L. (2009). [Long term consequences among children exposed to
lead poisoning]. Rev Med Chil 137: 1037-1044. http://dx.doi.org/10.4067/S0034-98872009000800006
Coria. F; Berciano. MT; Berciano. J; Lafarga. M. (1984). Axon membrane remodeling in the lead-induced
demyelinating neuropathy of the rat. Brain Res 291: 369-372.
http://dx.doi.org/10.1016/0006-8993(84)91271-x
Corongiu. FP; Milia. A. (1982). Rise of hepatic glutathione concentration induced in rats by chronic lead
nitrate treatment: Its role in aflatoxin Bl intoxication. Res Comm Chem Pathol Pharmacol 38: 97-112.
Cory-Slechta. DA. (1990). Exposure duration modifies the effects of low level lead on fixed-interval
performance. Neurotoxicology 11: 427-441.
Cory-Slechta. DA. (1994). Neurotoxicant-induced changes in schedule-controlled behavior. In LW Chang
(Ed.), Principles of neurotoxicology (pp. 313-344). New York, NY: Marcel Dekker, Inc.
Cory-Slechta. DA; Pokora. MJ; Widzowski. DV. (1991). Behavioral manifestations of prolonged lead
exposure initiated at different stages of the life cycle: II. Delayed spatial alternation. Neurotoxicology
12: 761-776.
Cory-Slechta. DA; Pokora. MJ; Widzowski. DV. (1992). Postnatal lead exposure induces supersensitivity
to the stimulus properties of D2D3 agonist. Brain Res 598: 162-172.
http://dx.doi.org/10.1016/0006-8993(92)90180-H
Cory-Slechta. DA; Stern. S; Weston. D; Allen. JL; Liu. S. (2010). Enhanced learning deficits in female
rats following lifetime pb exposure combined with prenatal stress. Toxicol Sci 117: 427-438.
http://dx.doi.org/10.1093/toxsci/kfq221
November 2012 5-720 Draft - Do Not Cite or Quote
-------
Costa. LG: Fox. DA. (1983). A selective decrease of cholinergic muscarinic receptors in the visual cortex
of adult rats following developmental lead exposure. Brain Res 276: 259-266.
http://dx.doi.org/10.1016/0006-8993(83)90733-3
Costa. M; Zhitkovich. A; Gargas. M; Paustenbach. D; Finlev. B; Kuykendall. J: Billings. R; Carlson. TJ;
Wetterhahn. K; Xu. J; Patierno. S: Bogdanffy. M. (1996). Interlaboratory validation of a new assay for
DNA-protein crosslinks. Mutat Res Genet Toxicol 369: 13-21.
http://dx.doi.org/10.1016/S0165-1218(96)90043-9
Counter. SA; Buchanan. LH; Ortega. F. (2008). Zinc protoporphyrin levels, blood lead levels and
neurocognitive deficits in Andean children with chronic lead exposure. Clin Biochem 41: 41-47.
http://dx.doi.0rg/10.1016/i.clinbiochem.2007.10.002
Counter. SA; Buchanan. LH; Ortega. F. (2009). Neurocognitive screening of lead-exposed andean
adolescents and young adults. J Toxicol Environ Health A 72: 625-632.
http://dx.doi.org/10.1080/15287390902769410
Counter. SA; Buchanan. LH: Ortega. F; Rifai. N; Shannon. MW. (2007). Comparative analysis of zinc
protoporphyrin and blood lead levels in lead-exposed Andean children. Clin Biochem 40: 787-792.
http://dx.doi.0rg/10.1016/i.clinbiochem.2007.03.003
Counter. SA; Buchanan. LH: Ortega. F; van der Velde. J: Borg. E. (2011). Assessment of auditory
brainstem function in lead-exposed children using stapedius muscle reflexes. J Neurol Sci 306: 29-37.
http://dx.doi.0rg/10.1016/i.jns.2011.04.003
Courtois. E; Marques. M; Barrientos. A; Casado. S: Lopez-Farre. A. (2003). Lead-induced
downregulation of soluble guanylage cyclase in isolated rat aortic segments mediated by reactive
oxygen species and cyclooxygenase-2. JAm SocNephrol 14: 1464-1470.
http://dx.doi.org/10.1097/01.ASN.0000064947.14997.69
Coyle. JT; Tsai. G. (2004). The NMDA receptor glycine modulatory site: A therapeutic target for
improving cognition and reducing negative symptoms in schizophrenia [Review].
Psychopharmacology 174: 32-38. htrp://dx.doi.org/10.1007/s00213-003-1709-2
Croes. K; Baevens. W: Bruckers. L; Den Hond. E; Koppen. G: Nelen. V; Van de Mieroop. E; Keune. H;
Dhooge. W: Schoeters. G: Van Larebeke. N. (2009). Hormone levels and sexual development in
Flemish adolescents residing in areas differing in pollution pressure. Int J Hyg Environ Health 212:
612-625. http://dx.doi.0rg/10.1016/i.ijheh.2009.05.002
Crooks. PR; Ghosh. MC: Haller. RG: Tong. WH; Rouault. TA. (2010). Posttranslational stability of the
heme biosynthetic enzyme ferrochelatase is dependent on iron availability and intact iron-sulfur
cluster assembly machinery. Blood 115: 860-869. http://dx.doi.org/10.1182/blood-2009-09-243105
Cullen. C: Singh. A; Dykeman. A; Rice. D; Foster. W. (1993). Chronic lead exposure induces
ultrastructural alterations in the monkey seminal vesicle. J Submicrosc Cytol Pathol 25: 127-135.
Daggett DA; Oberlev. TD; Nelson. SA; Wright. LS: Kornguth. SE; Siegel. FL. (1998). Effects of lead on
rat kidney and liver: GST expression and oxidative stress. Toxicology 128: 191-206.
http://dx.doi.org/10.1016/S0300-483X(98)00080-8
Danadevi. K; Rozati. R; Saleha Banu. B; Hanumanth Rao. P; Grover. P. (2003). DNA damage in workers
exposed to lead using comet assay. Toxicology 187: 183-193.
htrp://dx.doi.org/10.1016/S0300-483X(03)00054-4
Daniels. SR. (2011). International differences in secular trends in childhood blood pressure: A puzzle to
be solved [Editorial]. Circulation 124: 378-380.
http://dx.doi.org/10.1161/CIRCULATIONAHA. 111.042747
Davenport. A; Murcutt. G: Whiting. S. (2009). Cross-sectional audit of blood lead levels in regular
outpatient haemodialysis patients dialysing in north London. Nephrology 14: 476-481.
http://dx.doi.0rg/10.llll/i.1440-1797.2009.01087.x
November 2012 5-721 Draft - Do Not Cite or Quote
-------
David. O: Clark. J; Voeller. K. (1972). Lead and hyperactivity. Lancet 300: 900-903.
http://dx.doi.org/10.1016/S0140-6736(72)92534-2
David. OJ: Hoffman. SP; Sverd. J: Clark. J; Voeller. K. (1976). Lead and hyperactivity: Behavorial
response to chelation: Apilot study. Am J Psychiatry 133: 1155-1158.
Davidson. R; MacKinnon. JG. (1981). Several tests for model specification in the presence of alternative
hypotheses. Econometrica49: 781-793.
de Burbure. C: Buchet. JP; Leroyer. A; Nisse. C: Haguenoer. JM; Mutti. A; Smerhovsky. Z; Cikrt. M;
Trczinka-Ochocka. M; Razniewska. G: Jakubowski. M; Bernard. A. (2006). Renal and neurologic
effects of cadmium, lead, mercury, and arsenic in children: Evidence of early effects and multiple
interactions at environmental exposure levels. Environ Health Perspect 114: 584-590.
http:/7dx.doi.org/10.1289/ehp.82Q2
De Coster. S: Koppen. G: Bracke. M; Schroijen. C: Den Hond. E; Nelen. V; Van De Mieroop. E;
Bruckers. L; Bilau. M; Baevens. W: Schoeters. G: van Larebeke. N. (2008). Pollutant effects on
genotoxic parameters and tumor-associated protein levels in adults: Across sectional study. Environ
Health Global Access Sci Source 7: 26. http://dx.doi.org/10.1186/1476-069X-7-26
De Guise. S: Bernier. J: Lapierre. P; Dufresne. MM; Dubreuil. P; Fornier. M. (2000). Immune function of
bovine leukocytes after in vitro exposure to selected heavy metals. Am J Vet Res 61: 339-344.
http://dx.doi.org/10.2460/aivr.2000.61.339
De. M; Ghosh. S: Palit. S: Ghosh. A; Talukder. G: Sharma. A. (1995). Clastogenic effects in human
samples following prolonged exposure in metal industry. Bull Environ Contam Toxicol 54: 357-362.
http://dx.doi.org/10.1007/BF00195105
De Palma. G: Goldoni. M; Catalani. S: Carbognani. P; Poli. D; Mozzoni. P; Acampa. O: Internullo. E;
Rusca. M; Apostoli. P. (2008). Metallic elements in pulmonary biopsies from lung cancer and control
subjects. Acta Biomed 79: 43-51.
de Restrepo. HG: Sicard. D; Torres. MM. (2000). DNA damage and repair in cells of lead exposed
people. Am J Ind Med 38: 330-334. http://dx.doi.org/10.1002/1097-0274(200009)38:3<330::AID-
AJIM13>3.0.CO:2-Z
de Souza Lisboa. SF; Goncalves. G: Komatsu. F; Queiroz. CAS: Almeida. AA; Moreira. EG. (2005).
Developmental lead exposure induces depressive-like behavior in female rats. Drug Chem Toxicol 28:
67-77. http://dx.doi.org/10.1081/dct-200039696
Dearth. RK; Hinev. JK; Srivastava. V: Burdick. SB; Bratton. GR; Dees. WL. (2002). Effects of lead (Pb)
exposure during gestation and lactation on female pubertal development in the rat. Reprod Toxicol 16:
343-352. http://dx.doi.org/10.1016/S0890-6238(02)00037-0
Degawa. M; Arai. H; Miura. S: Hashimoto. Y. (1993). Preferential inhibitions of hepatic P450IA2
expression and induction by lead nitrate in the rat. Carcinogenesis 14: 1091-1094.
http://dx.doi.0rg/10.1093/carcin/14.6.1091
Dehlink. E; Baker. AH; Yen. E; Nurko. S: Fiebiger. E. (2010). Relationships between levels of serum IgE,
cell-bound IgE, and IgE-receptors on peripheral blood cells in a pediatric population. PLoS ONE 5:
e!2204. http://dx.doi.org/10.1371/iournal.pone.0012204
Del Donno. M; Chetta. A; Foresi. A; Gavaruzzi. G: Ugolotti. G: Olivieri. D. (1997). Lung epithelial
permeability and bronchial responsiveness in subjects with stable asthma. Chest 111: 1255-1260.
http://dx.doi.0rg/10.1378/chest.lll.5.1255
Delville. Y. (1999). Exposure to lead during development alters aggressive behavior in golden hamsters.
Neurotoxicol Teratol 21: 445-449.
Dempster. EL; Pidslev. R; Schalkwyk. LC: Owens. S: Georgiades. A; Kane. F; Kalidindi. S: Picchioni. M;
Kravariti. E; Toulopoulou. T; Murray. RM; Mill. J. (2011). Disease-associated epigenetic changes in
monozygotic twins discordant for schizophrenia and bipolar disorder. Hum Mol Genet 20: 4786-4796.
http://dx.doi.org/10.1093/hmg/ddr416
November 2012 5-722 Draft - Do Not Cite or Quote
-------
Den Hond E. D. hooge W. B..nickers L. S..choeters G. N..elen V. van de Mieroop E. K..oppen G. B.jlau
M. S..chroijen C. K..eune H. B..aevens W. van Larebeke N. (2011). Internal exposure to pollutants and
sexual maturation in Flemish adolescents. J Expo Sci Environ Epidemiol 21: 224-233.
http://dx.doi.0rg/10.1038/ies.2010.2
Deng. H; Callender. R; Schramm. VL; Grubmever. C. (2010). Pyrophosphate activation in hypoxanthine—
guanine phosphoribosyltransferase with transition state analogue. Biochemistry 49: 2705-2714.
http://dx.doi.org/10.1021M100012u
Deng. W: Poretz. RD. (2002). Protein kinase C activation is required for the lead-induced inhibition of
proliferation and differentiation of cultured oligodendroglial progenitor cells. Brain Res 929: 87-95.
http://dx.doi.org/10.1016/S0006-8993(01)03385-6
Denham. M; Schell. LM; Deane. G: Gallo. MV; Ravenscroft. J: DeCaprio. AP. (2005). Relationship of
lead, mercury, mirex, dichlorodiphenyldichloroethylene, hexachlorobenzene, and polychlorinated
biphenyls to timing of menarche among Akwesasne Mohawk girls. Pediatrics 115: e!27-e!34.
http://dx.doi.org/10.1542/peds.2004-1161
Dentener. MA; Greve. JW: Maessen. JG: Buurman. WA. (1989). Role of tumour necrosis factor in the
enhanced sensitivity of mice to endotoxin after exposure to lead. Immunopharmacol Immunotoxicol
11: 321-334. http://dx.doi.org/10.3109/08923978909005373
Despres. C: Beuter. A; Richer. F; Poitras. K; Veilleux. A; Ayotte. P; Dewailly. E; Saint-Amour. D;
Muckle. G. (2005). Neuromotor functions in Inuit preschool children exposed to Pb, PCBs, and Hg.
Neurotoxicol Teratol 27: 245-257. http://dx.doi.Org/10.1016/i.ntt.2004.12.001
Devaraian. P. (2007). Emerging biomarkers of acute kidney injury. In C Ronco; R Bellomo; JA Kellum
(Eds.), Acute kidney injury (pp. 203-212). Basel, Switzerland: Karger.
http://dx.doi.org/10.1159/000102085
Deveci. E. (2006). Ultrastructural effects of lead acetate on brain of rats. Toxicol Ind Health 22: 419-422.
http://dx.doi.org/10.1177/07482337060220100101
Devi. CB; Reddv. GH; Prasanthi. RPJ; Chettv. CS: Reddv. GR. (2005). Developmental lead exposure
alters mitochondrial monoamine oxidase and synaptosomal catecholamine levels in rat brain. Int J
DevNeurosci 23: 375-381. http://dx.doi.org/10.1016/Ujdevneu.2004.ll.003
Devi. KD; Banu. BS: Grover. P; Jamil. K. (2000). Genotoxic effect of lead nitrate on mice using SCGE
(comet assay). Toxicology 145: 195-201. http://dx.doi.org/10.1016/S0300-483X(00)00154-2
Dhir. H; Ghosh. S: Sharma. A; Talukder. G. (1992a). Interaction between two group IV metals-lead and
zirconium—in bone marrow cells of Mus musculus in vivo. Biometals 5: 81-86.
http://dx.doi.org/10.1007/BF01062218
Dhir. H; Roy. AK; Sharma. A. (1993). Relative efficiency of Phyllanthus emblica fruit extract and
ascorbic acid in modifying lead and aluminium-induced sister-chromatid exchanges in mouse bone
marrow. Environ Mol Mutagen 21: 229-236. http://dx.doi.org/10.1002/em.2850210305
Dhir. H; Roy. AK; Sharma. A; Talukder. G. (1990). Modification of clastogenicity of lead and aluminium
in mouse bone marrow cells by dietary ingestion of Phyllanthus emblica fruit extract. Mutat Res
Genet Toxicol 241: 305-312. http://dx.doi.org/10.1016/0165-1218(90)90029-2
Dhir. H; Sharma. A; Talukder. G. (1992b). Modifying effect of iron on lead-induced clastogenicity in
mouse bone marrow cells. Biol Trace Elem Res 34: 279-286. http://dx.doi.org/10.1007/BF02783683
Di Lorenzo. L; Silvestroni. A; Martino. MG: Gagliardi. T; Corfiati. M; Soleo. L. (2006). Evaluation of
peripheral blood neutrophil leucocytes in lead-exposed workers. Int Arch Occup Environ Health 79:
491-498. http://dx.doi.org/10.1007/s00420-005-0073-4
Di Lorenzo. L; Vacca. A; Corfiati. M; Lovreglio. P: Soleo. L. (2007). Evaluation of tumor necrosis factor-
alpha and granulocyte colony-stimulating factor serum levels in lead-exposed smoker workers. Int J
Immunopathol Pharmacol 20: 239-247.
November 2012 5-723 Draft - Do Not Cite or Quote
-------
Dietert. RR. (2011). Role of developmental immunotoxicity and immune dysfunction in chronic disease
and cancer [Review]. Reprod Toxicol 31: 319-326. http://dx.doi.Org/10.1016/j.reprotox.2010.09.006
Dietert. RR: McCabe. MJ. Jr. (2007). Lead immunotoxicity. In RW Luebke; RV House; I Kimber (Eds.),
Immunotoxicology and immunopharmacology (3rd ed., pp. 207-223). Boca Raton, FL: CRC Press.
Dietert. RR: Piepenbrink. MS. (2006). Perinatal immunotoxicity: Why adult exposure assessment fails to
predict risk [Review]. Environ Health Perspect 114: 477-483. http://dx.doi.org/10.1289/ehp.8566
Dietrich. KN; Berger. OG: Succop. PA. (1993 a). Lead exposure and the motor developmental status of
urban six-year-old children in the Cincinnati Prospective Study. Pediatrics 91: 301-307.
Dietrich. KN; Berger. OG: Succop. PA; Hammond. PB; Bornschein. RL. (1993b). The developmental
consequences of low to moderate prenatal and postnatal lead exposure: Intellectual attainment in the
Cincinnati Lead Study Cohort following school entry. Neurotoxicol Teratol 15: 37-44.
http://dx.doi.org/10.1016/0892-0362(93)90043-N
Dietrich. KN; Krafft KM; Bier. M; Succop. PA; Berger. O; Bornschein. RL. (1986). Early effects of fetal
lead exposure: Neurobehavioral findings at 6 months. International Journal for Biosocial Research 8:
151-168.
Dietrich. KN; Krafft. KM; Bornschein. RL; Hammond. PB; Berger. O; Succop. PA; Bier. M. (1987a).
Low-level fetal lead exposure effect on neurobehavioral development in early infancy. Pediatrics 80:
721-730.
Dietrich. KN; Krafft. KM; Shukla. R; Bornschein. RL; Succop. PA. (1987b). The neurobehavioral effects
of early lead exposure. In SR Schroeder (Ed.), Toxic substances and mental retardation:
Neurobehavioral toxicology and teratology (pp. 71-95). Washington, DC: American Association on
Mental Deficiency.
Dietrich. KN: Ris. MD; Succop. PA; Berger. OG: Bornschein. RL. (2001). Early exposure to lead and
juvenile delinquency. Neurotoxicol Teratol 23: 511-518.
http://dx.doi.org/10.1016/S0892-0362(01)00184-2
Dietrich. KN: Succop. PA; Berger. OG: Hammond. PB: Bornschein. RL. (1991). Lead exposure and the
cognitive development of urban preschool children: The Cincinnati Lead Study cohort at age 4 years.
Neurotoxicol Teratol 13: 203-211. http://dx.doi.org/10.1016/0892-0362(91)90012-L
Dietrich. KN: Succop. PA; Berger. OG: Keith. RW. (1992). Lead exposure and the central auditory
processing abilities and cognitive development of urban children: The Cincinnati Lead Study cohort at
age 5 years. Neurotoxicol Teratol 14: 51-56. http://dx.doi.org/10.1016/0892-0362(92)90028-9
Ding. Y; Gonick. HC: Vaziri. ND. (2000). Lead promotes hydroxyl radical generation and lipid
peroxidation in cultured aortic endothelial cells. Am J Hypertens 13: 552-555.
http://dx.doi.org/10.1016/S0895-7061(99)00226-5
Ding. Y; Gonick. HC: Vaziri. ND; Liang. K; Wei. L. (2001). Lead-induced hypertension. III. Increased
hydroxyl radical production. Am J Hypertens 14: 169-173.
http://dx.doi.org/10.1016/S0895-7061(00)01248-6
Diouf. A; Garcon. G: Diop. Y; Ndiave. B; Thiaw. C: Fall. M; Kane-Barry. O: Ba. D; Haguenoer. JM;
Shirali. P. (2006). Environmental lead exposure and its relationship to traffic density among
Senegalese children: A cross-sectional study. Hum Exp Toxicol 25: 637-644.
http://dx.doi.org/10.1177/0960327106074591
Dogru. MI; Dogru. AK; Gul. M; Esrefoglu. M; Yurekli. M; Erdogan. S: Ates. B. (2008). The effect of
adrenomedullin on rats exposed to lead. J Appl Toxicol 28: 140-146.
http://dx.doi.org/10.1002/iat. 1259
Dogu. O: Louis. ED; Tamer. L; Unal. O: Yilmaz. A; Kaleagasi. H. (2007). Elevated blood lead
concentrations in essential tremor: A case-control study in Mersin, Turkey. Environ Health Perspect
115: 1564-1568. http://dx.doi.org/10.1289/ehp.10352
November 2012 5-724 Draft - Do Not Cite or Quote
-------
Donald. JM; Cutler. MG: Moore. MR. (1987). Effects of lead in the laboratory mouse. Development and
social behaviour after lifelong exposure to 12 microM lead in drinking fluid. Neuropharmacology 26:
391-399. http://dx.doi.org/10.1016/0028-3908(87)90194-8
Donald. JM; Cutler. MG: Moore. MR; Bardlev. M. (1986). Effects of lead in the laboratory mouse-2:
Development and social behaviour after lifelong administration of a small dose of lead acetate in
drinking fluid. Neuropharmacology 25: 151-160.
Donaldson. WE; Knowles. SO. (1993). Is lead toxicosis a reflection of altered fatty acid composition of
membranes? [Review]. Comp Biochem Physiol C Toxicol Pharmacol 104: 377-379.
httrj://dx.doi.org/10.1016/0742-8413(93)90003-4
Dong. S: Liang. D; An. N; Jia. L; Shan. Y; Chen. C: Sun. K; Niu. F; Li. H; Fu. S. (2009). The role of
MAPK and FAS death receptor pathways in testicular germ cell apoptosis induced by lead. Acta
Biochim Biophys Sin 41: 800-807. http://dx.doi.org/10.1093/abbs/gmp069
Dorman. RV; Freeman. EJ. (2002). Lead-dependent effects on arachidonic acid accumulation and the
proliferation of vascular smooth muscle. J Biochem Mol Toxicol 16: 245-253.
http://dx.doi.org/10.1002/ibt.10045
Dorsev. CD; Lee. BK; Bolla. KI; Weaver. VM; Lee. SS: Lee. GS: Todd. AC: Shi. W: Schwartz. BS.
(2006). Comparison of patella lead with blood lead and tibia lead and their associations with
neurobehavioral test scores. J Occup Environ Med 48: 489-496.
http://dx.doi.org/10.1097/01.jom.0000199678.86629.3b
Dou. C: Zhang. J. (2011). Effects of lead on neurogenesis during zebrafish embryonic brain development.
J Hazard Mater 194: 277-282. http://dx.doi.org/10.1016/i.ihazmat.2011.07.106
Dowd. JB; Aiello. AE. (2009). Socioeconomic differentials in immune response. Epidemiology 20:
902-908. http://dx.doi.org/10.1097/EDE.Ob013e3181bb5302
Dowd. TL; Gupta. RK. (1991). 19F-NMR study of the effect of lead on intracellular free calcium in
human platelets. Biochim Biophys Acta 1092: 341-346.
http://dx.doi.org/10.1016/S0167-4889(97)90010-0
Dowd. TL; Rosen. JF; Gundberg. CM; Gupta. RK. (1994). The displacement of calcium from osteocalcin
at submicromolar concentrations of free lead. Biochim Biophys Acta 1226: 131-137.
http://dx.doi.org/10.1016/0925-4439(94)90020-5
Dowd. TL: Rosen. JF; Gupta. RK. (1990). 31PNMR and saturation transfer studies of the effect of PB2+
on cultured osteoblastic bone cells. J Biol Chem 265: 20833-20838.
Dowd. TL; Rosen. JF: Mints. L; Gundberg. CM. (2001). The effect of Pb2+ on the structure and
hydroxyapatite binding properties of osteocalcin. Biochim Biophys Acta 1535: 153-163.
http://dx.doi.org/10.1016/S0925-4439(00)00094-6
Dumitrescu. E; Trif. A; Argherie. D. (2007). Consequences of chronic exposure of female rats to lead
acetate on ovarian and uterine histoarhitectonic. Lucrari Stiintifice, Seria C: Medicina Veterinara LII:
198-205.
Dumitrescu. E; Trif. A; Florin. M. (2008a). The consequences of female rats chronic exposure to lead
acetate in the biomarkers emphasizing the hormonal disrupting potential of the reproductive function
for in vivo evaluation. Bull Univ Agric Sci Vet Med Cluj-Napoca Vet Med 65: 382-387.
Dumitrescu. E; Trif. A; Muselin. F. (2008b). The impact of lead acetate on some markers of
developmental toxicity in female rats (body weight, litter size, litter weight until weaning). Lucrari
Stiintifice, Seria C: Medicina Veterinara 53: 192-197.
Dumitrescu. E; Trif. A; Petrovici. S. (2008c). Lead acetate impact on some markers of female
reproductive system performances (litter size, sex ratio) and physical development (vaginal opening)
in rats. Bull Univ Agric Sci Vet Med Cluj-Napoca Vet Med 65: 283-287.
November 2012 5-725 Draft - Do Not Cite or Quote
-------
Dundar. B; Oktem. F; Arslan. MK; Delibas. N; Baykal. B; Arslan. C: Gultepe. M; Ilhan. IE. (2006). The
effect of long-term low-dose lead exposure on thyroid function in adolescents. Environ Res 101:
140-145. http://dx.doi.0rg/10.1016/i.envres.2005.10.002
Dursun. N; Arifoglu. C: Suer. C: Keskinol. L. (2005). Blood pressure relationship to nitric oxide, lipid
peroxidation, renal function, and renal blood flow in rats exposed to low lead levels. Biol Trace Elem
Res 104: 141-149. http://dx.doi.Org/10.1385/BTER:104:2:141
DuVal. G: Fowler. BA. (1989). Preliminary purification and characterization studies of a low molecular
weight, high affinity cytosolic lead-binding protein in rat brain. Biochem Biophys Res Commun 159:
177-184. http://dx.doi.org/10.1016/0006-291X(89)92420-0
Duvall. RM; Norris. GA; Dailev. LA; Burke. JM; McGee. JK; Gilmour. MI; Gordon. T; Devlin. RB.
(2008). Source apportionment of paniculate matter in the U.S. and associations with lung
inflammatory markers. Inhal Toxicol 20: 671-683. http://dx.doi.org/10.1080/08958370801935117
Duvdu. Y; Dur. A; Siizen. HS. (2005). Evaluation of increased proportion of cells with unusually high
sister chromatid exchange counts as a cytogenetic biomarker for lead exposure. Biol Trace Elem Res
104: 121-129. http://dx.doi.Org/10.1385/BTER:104:2:121
Duvdu. Y; Suzen. HS: Aydin. A; Gander. O; Uysal. H; Isimer. A; Vural. N. (2001). Correlation between
lead exposure indicators and sister chromatid exchange (SCE) frequencies in lymphocytes from
inorganic lead exposed workers. Arch Environ Contam Toxicol 41: 241-246.
http://dx.doi.org/10.1007/s002440010244
Dyatlov. VA; Lawrence. DA. (2002). Neonatal lead exposure potentiates sickness behavior induced by
Listeria monocytogenes infection of mice. Brain Behav Immun 16: 477-492.
http://dx.doi.org/10.1006/brbi.2001.0641
Dyatlov. VA; Platoshin. AV; Lawrence. DA; Carpenter. DO. (1998). Lead potentiates cytokine- and
glutamate-mediated increases in permeability of the blood-brain barrier. Neurotoxicology 19:
283-291.
Eder. K; Reichlmayr-Lais. AM; Kirchgessner. M. (1990). Activity of Na-KATPase and Ca-MgATPase in
red blood cell membranes of lead-depleted rats. J Trace Elem Electrolytes Health Dis 4: 21-24.
Egle. PM; Shelton. KR. (1986). Chronic lead intoxication causes a brain-specific nuclear protein to
accumulate in the nuclei of cells lining kidney tubules. J Biol Chem 261: 2294-2298.
Ehrlich. R; Robins. T; Jordaan. E; Miller. S: Mbuli. S: Selbv. P; Wynchank. S: Cantrell. A; De Broe. M;
D'Haese. P; Todd. A; Landrigan. P. (1998). Lead absorption and renal dysfunction in a South African
battery factory. Occup Environ Med 55: 453-460. http://dx.doi.Org/10.1136/oem.55.7.453
Ekinci. D; Bevdemir. S: Kufrevioglu. OI. (2007). In vitro inhibitory effects of some heavy metals on
human erythrocyte carbonic anhydrases. J Enzyme Inhib Med Chem 22: 745-750.
http://dx.doi.org/10.1080/14756360601176048
El-Ashmawv. IM; Ashry. KM; El-Nahas. AF; Salama. OM. (2006). Protection by turmeric and myrrh
against liver oxidative damage and genotoxicity induced by lead acetate in mice. Basic Clin
Pharmacol Toxicol 98: 32-37. http://dx.doi.Org/10.llll/i.1742-7843.2006.pto 228.x
El-Fawal. HAN; Waterman. SJ: De Feo. A; Shamy. MY. (1999). Neuroimmunotoxicology: Humoral
assessment of neurotoxicity and autoimmune mechanisms [Review]. Environ Health Perspect 107:
767-775.
El-Ghor, AA; Noshy, MM; Bid, JI. (2011). Lead acetate and arsenic trioxide induce instability of
microsatellites at three different fragile sites (6q21, 9q32-9q33 and 15pl4) within the genome of the
rat. Mutat Res 726: 195-199. http://dx.doi.Org/10.1016/i.mrgentox.2011.10.003
El-Nekeetv. AA; El-Kadv. AA; Soliman. MS: Hassan. NS: Abdel-Wahhab. MA. (2009). Protective effect
of Aquilegia vulgaris (L.) against lead acetate-induced oxidative stress in rats. Food Chem Toxicol 47:
2209-2215. http://dx.doi.0rg/10.1016/i.fct.2009.06.019
November 2012 5-726 Draft - Do Not Cite or Quote
-------
El-Neweshy. MS: El-Saved. YS. (2011). Influence of vitamin C supplementation on lead-induced
histopathological alterations in male rats. Exp Toxicol Pathol 63: 221-227.
http://dx.doi.0rg/10.1016/i.etp.2009.12.003
El-Sokkary. GH; Abdel-Rahman. GH; Kamel. ES. (2005). Melatonin protects against lead-induced
hepatic and renal toxicity in male rats. Toxicology 213: 25-33.
http://dx.doi.0rg/10.1016/i.tox.2005.05.003
El Shafai. A; Zohdv. N; El Mulla. K; Hassan. M; Morad. N. (2011). Light and electron microscopic study
of the toxic effect of prolonged lead exposure on the seminiferous tubules of albino rats and the
possible protective effect of ascorbic acid. Food Chem Toxicol 49: 734-743.
http://dx.doi.0rg/10.1016/i.fct.2010.ll.033
Blinder. CG: Friberg. L; Lind. B; Nilsson. B; Svartengren. M; Overmark. I. (1986). Decreased blood lead
levels in residents of Stockholm for the period 1980-1984. Scand J Work Environ Health 12: 114-120.
Elmarsafawv. SF; Jain. NB; Schwartz. J; Sparrow. D; Nie. HL; Hu. H. (2006). Dietary calcium as a
potential modifier of the relationship of lead burden to blood pressure. Epidemiology 17: 531-537.
http://dx.doi.org/10.1097/01.ede.0000231285.86968.2b
Elwood. RK; Kennedy. S: Belzberg. A; Hogg. JC: Pare. PP. (1983). Respiratory mucosal permeability in
asthma. Am Rev Respir Dis 128: 523-527.
Emmerson. BT. (1965). The renal excretion of urate in chronic lead nephropathy. Intern Med J 14:
295-303.
Emmerson. BT; Ravenscroft. PJ. (1975). Abnormal renal urate homeostasis in systemic disorders.
Nephron 14: 62-80. http://dx.doi.org/10.1159/000180436
Ercal. N; Treeratphan. P; Hammond. TC: Matthews. RH; Grannemann. NH; Spitz. DR. (1996). In vivo
indices of oxidative stress in lead-exposed C57BL/6 mice are reduced by treatment with meso-2,3-
dimercaptosuccinic acid or N-acetylcysteine. Free Radic Biol Med 21: 157-161.
http://dx.doi.org/10.1016/0891-5849(96)00020-2
Ergurhan-Ilhan. I; Cadir. B; Koyuncu-Arslan. M; C. A; Gultepe. FM; Ozkan. G. (2008). Level of
oxidative stress and damage in erythrocytes in apprentices indirectly exposed to lead. Pediatrics
International 50: 45-50. http://dx.doi.Org/10.llll/i.1442-200X.2007.02442.x
Erie. JC: Good. JA; Butz. JA. (2009). Excess lead in the neural retina in age-related macular
degeneration. Am J Ophthalmol 148: 890-894. http://dx.doi.Org/10.1016/i.ajo.2009.07.001
Erjefalt. I; Persson. CGA. (1991). Allergen, bradykinin, and capsaicin increase outward but not inward
macromolecular permeability of guinea-pig tracheobronchial mucosa. Clin Exp Allergy 21: 217-224.
http://dx.doi.0rg/10.llll/i.1365-2222.1991.tb00833.x
Ernhart. CB; Morrow-Tlucak. M; Marler. MR; Wolf. AW. (1987). Low level lead exposure in the prenatal
and early preschool periods: Early preschool development. Neurotoxicol Teratol 9: 259-270.
http://dx.doi.org/10.1016/0892-0362(87)90011-0
Ernhart. CB; Morrow-Tlucak. M; Wolf. AW. (1988). Low level lead exposure and intelligence in the
preschool years. Sci Total Environ 71: 453-459.
Ernhart. CB: Morrow-Tlucak. M; Wolf. AW: Super. D; Drotar. D. (1989). Low level lead exposure in the
prenatal and early preschool periods: intelligence prior to school entry. Neurotoxicol Teratol 11:
161-170.
Ettinger. AS: Lamadrid-Figueroa. H; Tellez-Rojo. MM; Mercado-Garcia. A; Peterson. KE; Schwartz. J:
Hu. H; Hernandez-Avila. M. (2009). Effect of calcium supplementation on blood lead levels in
pregnancy: A randomized placebo-controlled trial. Environ Health Perspect 117: 26-31.
http://dx.doi.org/10.1289/ehp.11868
Ettinger. AS: Tellez-Rojo. MM; Amarasiriwardena. C: Bellinger. D; Peterson. K; Schwartz. J: Hu. H;
Hernandez-Avila. M. (2004a). Effect of breast milk lead on infant blood lead levels at 1 month of age.
Environ Health Perspect 112: 1381-1385.
November 2012 5-727 Draft - Do Not Cite or Quote
-------
Bum. KD; Nie. LH; Schwartz. J; Vokonas. PS: Sparrow. D; Hu. H; Weisskopf. MG. (2011). Prospective
cohort study of lead exposure and electrocardiographic conduction disturbances in the Department of
Veterans Affairs Normative Aging Study. Environ Health Perspect 119: 490-494.
http://dx.doi.org/10.1289/ehp.1003279
Evans. M; Fored. CM; Nise. G: Bellocco. R; Nyren. O; Blinder. CG. (2010). Occupational lead exposure
and severe CKD: A population-based case-control and prospective observational cohort study in
Sweden. Am J Kidney Dis 55: 497-506. http://dx.doi.Org/10.1053/i.aikd.2009.ll.012
Ewers. U: Stiller-Winkler. R; Idel. H. (1982). Serum immunoglobulin, complement C3, and salivary IgA
levels in lead workers. Environ Res 29: 351-357. http://dx.doi.org/10.1016/0013-9351(82)90036-6
Exon. JH; Koller. LD; Kerkvliet. NI. (1979). Lead-cadmium interaction: Effects on viral-induced
mortality and tissue residues in mice. Arch Environ Health 34: 469-475.
Factor-Litvak. P; Graziano. JH; Kline. JK; Popovac. D; Mehmeti. A; Ahmedi. G: Shrout. P; Murphy. MJ;
Gashi. E; Haxhiu. R; Raiovic. L; Nenezic. DU: Stein. ZA. (1991). Aprospective study of birthweight
and length of gestation in a population surrounding a lead smelter in Kosovo, Yugoslavia. Int J
Epidemiol 20: 722-728. http://dx.doi.Org/10.1093/iie/20.3.722
Factor-Litvak. P; Kline. JK; Popovac. D: Hadzialjevic. S: Lekic. V: Preteni-Rexhepi. E; Capuni-Paracka.
S: Slavkovich. V; Graziano. J. (1996). Blood lead and blood pressure in young children.
Epidemiology 7: 633-637.
Factor-Litvak. P; Slavkovich. V; Liu. X; Popovac. D: Preteni. E; Capuni-Paracka. S: Hadzialjevic. S:
Lekic. V: Lolacono. N; Kline. J: Graziano. J. (1998). Hyperproduction of erythropoietin in nonanemic
lead-exposed children. Environ Health Perspect 106: 361-364.
Factor-Litvak. P; Wasserman. G: Kline. JK: Graziano. J. (1999). The Yugoslavia prospective study of
environmental lead exposure. Environ Health Perspect 107: 9-15.
http://dx.doi.org/10.1289/ehp.991079
Fadrowski. JJ; Navas-Acien. A; Tellez-Plaza. M; Guallar. E; Weaver. VM; Furth. SL. (2010). Blood lead
level and kidney function in US adolescents: The Third National Health and Nutrition Examination
Survey. Arch Intern Med 170: 75-82. http://dx.doi.org/10.1001/archinternmed.2009.417
Fahmy. MA. (1999). Lead acetate genotoxicity in mice. Cytologia (Tokyo) 64: 357-365.
http://dx.doi.org/10.1508/cvtologia.64.357
Faith. RE: Luster. MI; Kimmel. CA. (1979). Effect of chronic developmental lead exposure on cell-
mediated immune functions. Clin Exp Immunol 35: 413-420.
Fan. G: Feng. C: Li. Y; Wang. CH; Yan. J: Li. W: Feng. JG: Shi. XL; Bi. YY (2009a). Selection of
nutrients for prevention or amelioration of lead-induced learning and memory impairment in rats. Ann
Occup Hyg 53: 341-351. http://dx.doi.org/10.1093/annhyg/mep019
Fan. G: Feng. C: Wu. F: Ye. W: Lin. F: Wang. C: Yan. J: Zhu. G: Xiao. Y: Bi. Y. (2010). Methionine
choline reverses lead-induced cognitive and N-methyl-d-aspartate receptor subunit 1 deficits.
Toxicology 272: 23-31. http://dx.doi.Org/10.1016/i.tox.2010.03.018
Fan. Y; Yamada. T; Shimizu. T; Nanashima. N; Akita. M; Suto. K; Tsuchida. S. (2009b). Ferritin
expression in rat hepatocytes and Kupffer cells after lead nitrate treatment. Toxicol Pathol 37:
209-217. http://dx.doi.org/10.1177/0192623308328544
Fang. F: Kwee. LC: Allen. KD: Umbach. DM: Ye. W: Watson. M: Keller. J: Oddone. EZ: Sandier. DP:
Schmidt. S: Kamel. F. (2010). Association between blood lead and the risk of amyotrophic lateral
sclerosis. Am J Epidemiol 171: 1126-1133. http://dx.doi.org/10.1093/aje/kwq06
Farant. JP; Wigfield. DC. (1987). Interaction of divalent metal ions with normal and lead-inhibited human
erythrocytic porphobilinogen synthase in vitro. Toxicol Appl Pharmacol 89: 9-18.
http://dx.doi.org/10.1016/0041-008X(87)90171-2
Farant. JP: Wigfield. DC. (1990). The effects of copper, zinc, mercury, and cadmium on rabbit
erythrocytic porphobilinogen synthase in vivo. J Anal Toxicol 14: 222-226.
November 2012 5-728 Draft - Do Not Cite or Quote
-------
Faraone. SV: Doyle. AE. (2001). The nature and heritability of attention-deficit/hyperactivity disorder
[Review]. Child Adolesc Psychiatr ClinNAm 10: 299-316, viii-ix.
Farmand. F; Ehdaie. A; Roberts. CK; Sindhu. RK. (2005). Lead-induced dysregulation of superoxide
dismutases, catalase, glutathione peroxidase, and guanylate cyclase. Environ Res 98: 33-39.
http://dx.doi.0rg/10.1016/i.envres.2004.05.016
Farrer. DG: Hueber. S: Laiosa. MD; Eckles. KG: McCabe. MJ. Jr. (2008). Reduction of myeloid
suppressor cell derived nitric oxide provides a mechanistic basis of lead enhancement of alloreactive
CD4+ T cell proliferation. Toxicol Appl Pharmacol 229: 135-145.
http://dx.doi.0rg/10.1016/i.taap.2007.12.011
Farrer. DG: Hueber. SM; McCabe. MJ. Jr. (2005). Lead enhances CD4+ T cell proliferation indirectly by
targeting antigen presenting cells and modulating antigen-specific interactions. Toxicol Appl
Pharmacol 207: 125-137. http://dx.doi.Org/10.1016/i.taap.2004.12.017
Favalli. L; Chiari. MC: Piccinini. F; Rozza. A. (1977). Experimental investigations on the contraction
induced by lead in arterial smooth muscle. Acta Pharmacol Toxicol 41: 412-420.
http://dx.doi.org/10.1111/i. 1600-0773.1977.tb03362.x
Fazli-Tabaei. S: Fahim. M; Khoshbaten. A. (2006). Acute lead exposure and contraction of rat isolated
aorta induced by Dl-dopaminergic and alpha-adrenergic drugs. Arch Iran Med 9: 119-123.
Fehlau. R; Grygorczyk. R; Fuhrmann. GF; Schwarz. W. (1989). Modulation of the Ca2+- or Pb2+-
activated K+-selective channels in human red cells. II. Parallelisms to modulation of the activity of a
membrane-bound oxidoreductase. Biochim Biophys Acta 978: 37-42.
http://dx.doi.org/10.1016/0005-2736(89)90495-l
Feinberg. AP (2007). Phenotypic plasticity and the epigenetics of human disease [Review]. Nature 447:
433-440. http://dx.doi.org/10.1038/nature05919
Pels. LM; Wunsch. M; Baranowski. J; Norska-Borowka. I; Price. RG: Taylor. SA; Patel. S: De Broe. M;
Elsevier. MM; Lauwerys. R: Roels. H; Bernard. A; Mutti. A; Gelpi. E; Rosello. J: Stolte. H. (1998).
Adverse effects of chronic low level lead exposure on kidney function: A risk group study in children.
Nephrol Dial Transplant 13: 2248-2256. http://dx.doi.Org/10.1093/ndt/13.9.2248
Ferguson. C: Kern. M; Audesirk. G. (2000). Nanomolar concentrations of inorganic lead increase Ca2+
efflux and decrease intracellular free Ca2+ ion concentrations in cultured rat hippocampal neurons by
a calmodulin-dependent mechanism. Neurotoxicology 21: 365-378.
Ferguson. MA; Vaidva. VS: Bonventre. JV. (2008). Biomarkers of nephrotoxic acute kidney injury.
Toxicology 245: 182-193. http://dx.doi.Org/10.1016/i.tox.2007.12.024
Fergusson. DM; Boden. JM; Horwood. LJ. (2008). Dentine lead levels in childhood and criminal
behaviour in late adolescence and early adulthood. J Epidemiol Community Health 62: 1045-1050.
http://dx.doi.org/10.1136/iech.2007.072827
Fergusson. DM; Fergusson. JE; Horwood. LJ: Kinzett. NG. (1988a). A longitudinal study of dentine lead
levels, intelligence, school performance and behaviour Part II Dentine lead and cognitive ability. J
Child Psychol Psychiatry 29: 793-809.
Fergusson. DM; Fergusson. JE; Horwood. LJ: Kinzett. NG. (1988b). A longitudinal study of dentine lead
levels, intelligence, school performance and behaviour: Part I. Dentine lead levels and exposure to
environmental risk factors. J Child Psychol Psychiatry 29: 781-792.
Fergusson. DM; Horwood. LJ: Lynskev. MT. (1993). Early dentine lead levels and subsequent cognitive
and behavioural development. J Child Psychol Psychiatry 34: 215-227.
http://dx.doi.0rg/10.llll/i.1469-7610.1993.tb00980.x
Fergusson. DM; Horwood. LJ: Lynskev. MT. (1997). Early dentine lead levels and educational outcomes
at 18 years. J Child Psychol Psychiatry 38: 471-478.
November 2012 5-729 Draft - Do Not Cite or Quote
-------
Fernandez-Cabezudo. MJ; All. SAE; Ullah. A; Hasan. MY; Kosanovic. M; Fahim. MA; Adem. A; al-
Ramadi. BK. (2007). Pronounced susceptibility to infection by Salmonella enterica serovar
Typhimurium in mice chronically exposed to lead correlates with a shift to Th2-type immune
responses. Toxicol Appl Pharmacol 218: 215-226. http://dx.doi.Org/10.1016/i.taap.2006.ll.018
Fine. PR; Thomas. CW: Suhs. RH; Cohnberg. RE; Flashner. BA. (1972). Pediatric blood lead levels. A
study in 14 Illinois cities of intermediate population. JAMA 221: 1475-1479.
Fiorim. J; Ribeiro Junior. RF; Silveira. EA; Padilha. AS: Vescovi. MV; de Jesus. HC: Stefanon. I;
Salaices. M; Vassallo. DV. (2011). Low-level lead exposure increases systolic arterial pressure and
endothelium-derived vasodilator factors in rat aortas. PLoS ONE 6: e!7117.
http://dx.doi.org/10.1371/iournal.pone.0017117
Fischbein. A; Tsang. P; Luo. JCJ: Roboz. JP; Jiang. JD; Bekesi. JG. (1993). Phenotypic aberrations of the
CD3+ and CD4+ cells and functional impairments of lymphocytes at low-level occupational exposure
to lead. Clin Immunol Immunopathol 66: 163-168. http://dx.doi.org/10.1006/clin.1993.1020
Flegal. AR; Smith. DR. (1992). Lead levels in preindustrial humans [Letter]. N Engl J Med 326:
1293-1294.
Flohe. SB; Briiggemann. J: Herder. C: Goebel. C: Kolb. H. (2002). Enhanced proinflammatory response
to endotoxin after priming of macrophages with lead ions. J Leukoc Biol 71: 417-424.
Flora. SJS: Saxena. G: Gautam. P; Kaur. P; Gill. KD. (2007). Response of lead-induced oxidative stress
and alterations in biogenic amines in different rat brain regions to combined administration of DMSA
and MiADMSA. Chem Biol Interact 170: 209-220. http://dx.doi.Org/10.1016/i.cbi.2007.08.003
Flora. SJS: Singh. S: Tandon. SK. (1989). Thiamine and zinc in prevention or therapy of lead
intoxication. J Int Med Res 17: 68-75.
Foglieni. C: Fulgenzi. A; Ticozzi. P; Pellegatta. F; Sciorati. C: Belloni. D; Ferrero. E; Ferrero. ME.
(2006). Protective effect of EDTA preadministration on renal ischemia. BMC Nephrol 7: 5.
http://dx.doi.org/10.1186/1471-2369-7-5
Foller. M; Huber. SM; Lang. F. (2008). Erythrocyte programmed cell death [Review]. IUBMB Life 60:
661-668. http://dx.doi.org/10.1002/iub. 106
Forst. LS: Freels. S: Persky. V. (1997). Occupational lead exposure and hearing loss. J Occup Environ
Med 39: 658-660.
Fortier. M; Omara. F; Bernier. J: Brousseau. P: Fournier. M. (2008). Effects of physiological
concentrations of heavy metals both individually and in mixtures on the viability and function of
peripheral blood human leukocytes in vitro. J Toxicol Environ Health A 71: 1327-1337.
http://dx.doi.org/10.1080/15287390802240918
Fortoul. TI; Saldivar O. L; Espejel-Mavab. G: Bazarro N. P; Mussali-Galante. P: Avila-Casado. M. del C:
Colin-Barenque. L; Avila-Costa. MR. (2005). Inhalation of cadmium, lead or its mixture effects on the
bronchiolar structure and its relation with metal tissue concentrations. Environ Toxicol Pharmacol 19:
329-334. http://dx.doi.0rg/10.1016/i.etap.2004.08.007
Fortune. T; Lurie. DI. (2009). Chronic low-level lead exposure affects the monoaminergic system in the
mouse superior olivary complex. J Comp Neurol 513: 542-558. http://dx.doi.org/10.1002/cne.21978
Foster. WG. (1992). Reproductive toxicity of chronic lead exposure in the female cynomolgus monkey.
Reprod Toxicol 6: 123-131.
Foster. WG: McMahon. A; YoungLai. EV; Hughes. EG: Rice. DC. (1993). Reproductive endocrine
effects of chronic lead exposure in the male cynomolgus monkey. Reprod Toxicol 7: 203-209.
Fowler. BA; DuVal. G. (1991). Effects of lead on the kidney: Roles of high-affinity lead-binding proteins.
Environ Health Perspect 91: 77-80. http://dx.doi.org/10.1289/ehp.919177
Fowler. BA: Kahng. MW: Smith. PR; Conner. EA; Laughlin. NK. (1993). Implications of lead binding
proteins for risk assessment of lead exposure. J Expo Sci Environ Epidemiol 3: 441-448.
November 2012 5-730 Draft - Do Not Cite or Quote
-------
Fowler. BA; Kimmel. CA; Woods. JS: McConnell. EE; Grant. LD. (1980). Chronic low-level lead
toxicity in the rat: III An integrated assessment of long-term toxicity with special reference to the
kidney. Toxicol Appl Pharmacol 56: 59-77. http://dx.doi.org/10.1016/0041-008X(80)90131-3
Fox. DA; Campbell. ML; Blocker. YS. (1997). Functional alterations and apoptotic cell death in the retina
following developmental or adult lead exposure. Neurotoxicology 18: 645-664.
Fox. DA; Chu. LWF. (1988). Rods are selectively altered by lead: II. Ultrastructure and quantitative
histology. Exp Eye Res 46: 613-625. http://dx.doi.org/10.1016/S0014-4835(88)80017-4
Fox. DA; Farber. DB. (1988). Rods are selectively altered by lead: I. Electrophysiology and biochemistry.
Exp Eye Res 46: 597-611. http://dx.doi.org/10.1016/S0014-4835(88)80016-2
Fox. DA; Kala. SV: Hamilton. WR; Johnson. JE; O'Callaghan. JA. (2008). Low-level human equivalent
gestational lead exposure produces supernormal scotopic electroretinograms, increased retinal
neurogenesis, and decreased retinal dopamine utilization in rats. Environ Health Perspect 116:
618-625. http://dx.doi.org/10.1289/ehp.11268
Fox. DA; Katz. LM. (1992). Developmental lead exposure selectively alters the scotopic ERG component
of dark and light adaptation and increases rod calcium content. Vision Res 32: 249-255.
http://dx.doi.org/10.1016/0042-6989(92)90134-5
Fox. DA; Katz. LM; Farber. DB. (1991). Low level developmental lead exposure decreases the
sensitivity, amplitude and temporal resolution of rods. Neurotoxicology 12: 641-654.
Fox. DA; Opanashuk. L; Zharkovsky. A; Weiss. B. (2010). Gene-chemical interactions in the developing
mammalian nervous system: Effects on proliferation, neurogenesis and differentiation.
Neurotoxicology 31: 589-597. http://dx.doi.Org/10.1016/i.neuro.2010.03.007
Fox. DA; Sillman. AJ. (1979). Heavy metals affect rod, but not cone, photoreceptors. Science 206: 78-80.
http://dx.doi.org/10.1126/science.314667
Fracasso. ME; Perbellini. L; Solda. S: Talamini. G: Franceschetti. P. (2002). Lead induced DNA strand
breaks in lymphocytes of exposed workers: Role of reactive oxygen species and protein kinase
C. Mutat Res Genet Toxicol Environ Mutagen 515: 159-169.
http://dx.doi.org/10.1016/S1383-5718(02)00012-8
Fraser. S: Muckle. G: Despres. C. (2006). The relationship between lead exposure, motor function and
behaviour in Inuit preschool children. Neurotoxicol Teratol 28: 18-27.
http://dx.doi.0rg/10.1016/i.ntt.2005.10.008
Fried. LF. (2009). Creatinine and cystatin C: What are the values? [Comment]. Kidney Int 75: 578-580.
http://dx.doi.org/10.1038/Ki.2008.688
Froehlich. TE; Lanphear. BP; Auinger. P; Hornung. R; Epstein. JN; Braun. J; Kahn. RS. (2009).
Association of tobacco and lead exposures with attention-deficit/hyperactivity disorder. Pediatrics
124: E1054-E1063. http://dx.doi.org/10.1542/peds.2009-0738
Froehlich. TE; Lanphear. BP; Dietrich. KN; Cory-Slechta. DA; Wang. N; Kahn. RS. (2007). Interactive
effects of a DRD4 polymorphism, lead and sex on executive functions in children. Biol Psychiatry 62:
243-249. http://dx.doi.0rg/10.1016/i.biopsvch.2006.09.039
Fujita. H; Orii. Y; Sano. S. (1981). Evidence of increased synthesis of delta-aminolevulinic acid
dehydratase in experimental lead-poisoned rats. Biochim Biophys Acta 678: 39-50.
http://dx.doi.org/10.1016/0304-4165(81)90045-3
Fujita. H; Sato. K; Sano. S. (1982). Increase in the amount of erythrocyte delta-aminolevulinic acid
dehydratase in workers with moderate lead exposure. Int Arch Occup Environ Health 50: 287-297.
http://dx.doi.org/10.1007/BF00378090
Fujiwara. Y; Kaii. T; Sakurai. S: Sakamoto. M; Kozuka. H. (1997). Inhibitory effect of lead on the repair
of wounded monolayers of cultured vascular endothelial cells. Toxicology 117: 193-198.
http://dx.doi.org/10.1016/S0300-483X(96)03575-5
November 2012 5-731 Draft - Do Not Cite or Quote
-------
Fukumoto. K; Karai. I; Horiguchi. S. (1983). Effect of lead on erythrocyte membranes. Occup Environ
Med 40: 220-223. http://dx.doi.Org/10.1136/oem.40.2.220
Fuller. GM; Zhang. Z. (2001). Transcriptional control mechanism of fibrinogen gene expression. Ann N Y
Acad Sci 936: 469-479. http://dx.doi.Org/10.llll/i.1749-6632.2001.tb03534.x
Fulton. M; Raab. G: Thomson. G: Laxen. D; Hunter. R; Hepburn. W. (1987). Influence of blood lead on
the ability and attainment of children in Edinburgh. Lancet 329: 1221-1226.
http://dx.doi.org/10.1016/S0140-6736(87)92683-3
Gainer. JH. (1977). Effects of heavy metals and of deficiency of zinc on mortality rates in mice infected
with encephalomyocarditis virus. Am J Vet Res 38: 869-872.
Gao. A; Lu. XT; Li. QY; Tian. L. (2010a). Effect of the delta-aminolevulinic acid dehydratase gene
polymorphism on renal and neurobehavioral function in workers exposed to lead in China. Sci Total
Environ 408: 4052-4055. http://dx.doi.Org/10.1016/i.scitotenv.2010.04.024
Gao. D; Kasten-Jolly. J; Lawrence. DA. (2006). The paradoxical effects of lead in interferon-gamma
knockout BALB/c mice. Toxicol Sci 89: 444-453. http://dx.doi.org/10.1093/toxsci/kfj043
Gao. D: Mondal. TK; Lawrence. DA. (2007). Lead effects on development and function of bone marrow-
derived dendritic cells promote Th2 immune responses. Toxicol Appl Pharmacol 222: 69-79.
http://dx.doi.0rg/10.1016/i.taap.2007.04.001
Gao. SJ: Jin. YL; Unverzagt. FW: Ma. F; Hall. KS: Murrell. JR; Cheng. YB; Shen. JZ; Ying. B; Ji. RD;
Matesan. J: Liang. C: Hendrie. HC. (2008). Trace element levels and cognitive function in rural
elderly Chinese. J Gerontol ABiol Sci Med Sci 63: 635-641.
Garavan. H; Morgan. RE; Levitsky. DA; Hermer-Vazquez. L; Strupp. BJ. (2000). Enduring effects of
early lead exposure: Evidence for a specific deficit in associative ability. Neurotoxicol Teratol 22:
151-164.
Garcia-Leston. J: Roma-Torres. J: Vilares. M; Pinto. R; Cunha. LM; Prista. J: Teixeira. JP; Mayan. O;
Pasaro. E; Mendez. J: Laffon. B. (2011). Biomonitoring of a population of Portuguese workers
exposed to lead. Mutat Res Genet Toxicol Environ Mutagen 721: 81-88.
http://dx.doi.0rg/10.1016/i.mrgentox.2011.01.001
Garcon. G: Leleu. B; Marez. T; Zerimech. F; Jean-Marie. HP; Daniel. FB; Shirali. P. (2007).
Biomonitoring of the adverse effects induced by the chronic exposure to lead and cadmium on kidney
function: Usefulness of alpha-glutathione S-transferase. Sci Total Environ 377: 165-172.
http://dx.doi.0rg/10.1016/i.scitotenv.2007.02.002
Gargioni. R; Neto. FF; Buchi. DF; Randi. MAP; Franco. CRC: Paludo. KS: Pelletier. E; Ferraro. MVM;
Cestari. MM; Bussolaro. D; Ribeiro. CAP. (2006). Cell death and DNA damage in peritoneal
macrophages of mice (Mus musculus) exposed to inorganic lead. Cell Biol Int 30: 615-623.
http://dx.doi.0rg/10.1016/i.cellbi.2006.03.010
Garza. A; Vega. R: Soto. E. (2006). Cellular mechanisms of lead neurotoxicity [Review]. Med Sci Monit
12: RA57-RA65.
Gastaldo. J: Viau. M; Bencokova. Z; Joubert. A; Charvet AM; Balosso. J: Foray. N. (2007). Lead
contamination results in late and slowly repairable DNA double-strand breaks and impacts upon the
ATM-dependent signaling pathways. Toxicol Lett 173: 201-214.
http://dx.doi.0rg/10.1016/i.toxlet.2007.08.003
Gautam. P; Flora. SJ. (2010). Oral supplementation of gossypin during lead exposure protects alteration
in heme synthesis pathway and brain oxidative stress in rats. Nutrition 26: 563-570.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.12.070
Gedeon. Y; Ramesh. GT; Wellman. PJ; Jadhav. AL. (2001). Changes in mesocorticolimbic dopamine and
D1/D2 receptor levels after low level lead exposure: Atime course study. Toxicol Lett 123: 217-226.
http://dx.doi.org/10.1016/S0378-4274(01)00408-8
November 2012 5-732 Draft - Do Not Cite or Quote
-------
Gent. JF; Koutrakis. P; Belanger. K; Triche. E; Holford. TR; Bracken. MB; Leaderer. BP. (2009).
Symptoms and medication use in children with asthma and traffic-related sources of fine particle
pollution. Environ Health Perspect 117: 1168-1174. http://dx.doi.org/10.1289/ehp.0800335
Gerr. F; Letz. R; Stokes. L; Chettle. D; McNeill. F; Kave. W. (2002). Association between bone lead
concentration and blood pressure among young adults. Am J Ind Med 42: 98-106.
Ghering. AB; Jenkins. LMM; Schenck. BL; Deo. S: Mayer. RA; Pikaart. MJ; Omichinski. JG: Godwin.
HA. (2005). Spectroscopic and functional determination of the interaction of Pb2+ with GATA
proteins. JAm Chem Soc 127: 3751-3759. http://dx.doi.org/10.1021/ia0464544
Ghosh-Narang. J; Jones. TM; Menke. A; Todd. AC: Muntner. P; Batuman. V. (2007). Parathyroid
hormone status does not influence blood and bone lead levels in dialysis patients. Am J Med Sci 334:
415-420. http://dx.doi.org/10.1097/MAJ.Ob013e318068b237
Giddabasappa. A; Hamilton. WR; Chanev. S: Xiao. W: Johnson. JE; Mukherjee. S: Fox. DA. (2011).
Low-level gestational lead exposure increases retinal progenitor cell proliferation and rod
photoreceptor and bipolar cell neurogenesis in mice. Environ Health Perspect 119: 71-77.
http://dx.doi.org/10.1289/ehp.1002524
Giedd. JN; Lalonde. FM; Celano. MJ: White. SL; Wallace. GL; Lee. NR; Lenroot. RK. (2009).
Anatomical brain magnetic resonance imaging of typically developing children and adolescents
[Review]. J Am Acad Child Adolesc Psychiatry 48: 465-470.
http://dx.doi.org/10.1097/CHI.Ob013e31819f2715
Gilbert. ME; Kelly. ME; Samsam. TE; Goodman. JH. (2005). Chronic developmental lead exposure
reduces neurogenesis in adult rat hippocampus but does not impair spatial learning. Toxicol Sci 86:
365-374. http://dx.doi.org/10.1093/toxsci/kFil56
Gilbert. ME; Mack. CM; Laslev. SM. (1999). Chronic developmental lead exposure and hippocampal
long-term potentiation: biphasic dose-response relationship. Neurotoxicology 20: 71-82.
Gilbert. SG: Rice. DC. (1987). Low-level lifetime lead exposure produces behavioral toxicity (spatial
discrimination reversal) in adult monkeys. Toxicol Appl Pharmacol 91: 484-490.
Gilsanz. V; Chalfant. J: Kalkwarf. H; Zemel. B; Lappe. J: Oberfield. S: Shepherd. J: Wren. T; Winer. K.
(2011). Age at onset of puberty predicts bone mass in young adulthood. J Pediatr 158: 100-105,
105.el01-102. http://dx.doi.0rg/10.1016/i.ipeds.2010.06.054
Giridhar. J: Isom. GE. (1990). Interaction of lead acetate with atrial natriuretic factor in rats. Life Sci 46:
569-576. http://dx.doi.org/10.1016/0024-3205(90)90124-A
Gittleman. R; Eskenazi. B. (1983). Lead and hyperactivity revisited: An investigation of non-
disadvantaged children. Arch Gen Psychiatry 40: 827-833.
http://dx.doi.org/10.1001/archpsvc.1983.01790070017002
Glahn. F; Schmidt-Heck. W: Zellmer. S: Guthke. R: Wiese. J: Golka. K; Hergenroder. R; Degen. GH;
Lehmann. T; Hermes. M; Schormann. W: Brulport. M; Bauer. A; Bedawv. E; Gebhardt. R: Hengstler.
JG: Foth. H. (2008). Cadmium, cobalt and lead cause stress response, cell cycle deregulation and
increased steroid as well as xenobiotic metabolism in primary normal human bronchial epithelial cells
which is coordinated by at least nine transcription factors. Arch Toxicol 82: 513-524.
http://dx.doi.org/10.1007/s00204-008-0331-9
Glass. TA; Bandeen-Roche. K; McAtee. M; Bolla. K; Todd. AC: Schwartz. BS. (2009). Neighborhood
psychosocial hazards and the association of cumulative lead dose with cognitive function in older
adults. Am J Epidemiol 169: 683-692. http://dx.doi.org/10.1093/aie/kwn390
Glenn. BS: Bandeen-Roche. K; Lee. BK; Weaver. VM; Todd. AC: Schwartz. BS. (2006). Changes in
systolic blood pressure associated with lead in blood and bone. Epidemiology 17: 538-544.
http://dx.doi.org/10.1097/01.ede.0000231284.19078.4b
Glymour. M. (2012). Data clarification for Glymour et al 2005. Available online at
http ://www.regulations. gov/#! documentDetail;D=EPA-HQ-ORD-2011 -0051 -0027
November 2012 5-733 Draft - Do Not Cite or Quote
-------
Glymour. MM; Weuve. J: Berkman. LF; Kawachi. I; Robins. JM. (2005). When is baseline adjustment
useful in analyses of change? An example with education and cognitive change. Am J Epidemiol 162:
267-278. http://dx.doi.org/10.1093/aje/kwil87
Godleski. JJ: Clarke. RW: Coull. BA: Saldiva. PHN: Jiang. NF: Lawrence. J: Koutrakis. P. (2002).
Composition of inhaled urban air particles determines acute pulmonary responses. Ann Occup Hyg
46: 419-424. http://dx.doi.org/10.1093/annhyg/46.suppl 1.419
Goering. PL; Fowler. BA. (1984). Regulation of lead inhibition of delta-aminolevulinic acid dehydratase
by a low molecular weight, high affinity renal lead-binding protein. J Pharmacol Exp Ther 231: 66-71.
Goering. PL; Fowler. BA. (1985). Mechanism of renal lead-binding protein reversal of delta-
aminolevulinic acid dehydratase inhibition by lead. J Pharmacol Exp Ther 234: 365-371.
Goering. PL; Fowler. BA. (1987a). Kidney zinc-thionein regulation of delta-aminolevulinic acid
dehydratase inhibition by lead. Arch Biochem Biophys 253: 48-55.
http://dx.doi.org/10.1016/0003-9861(87)90635-7
Goering. PL; Fowler. BA. (1987b). Regulatory roles of high-affinity metal-binding proteins in mediating
lead effects on delta-aminolevulinic acid dehydratase. Ann N Y Acad Sci 514: 235-247.
http://dx.doi.org/10.1111/i. 1749-6632.1987.tb48778.x
Goering. PL; Mistry. P; Fowler. BA. (1986). A low molecular weight lead-binding protein in brain
attenuates lead inhibition of delta-aminolevulinic acid dehydratase: Comparison with a renal lead-
binding protein. J Pharmacol Exp Ther 237: 220-225.
Golabek. T; Darewicz. B; Borawska. M; Markiewicz. R; Socha. K; Kudelski. J. (2009). Lead
concentration in the bladder tissue and blood of patients with bladder cancer. Scand J Urol Nephrol
43: 467-470. http://dx.doi.org/10.3109/00365590903198991
Goldstein. GW. (1993). Evidence that lead acts as a calcium substitute in second messenger metabolism
[Review]. Neurotoxicology 14: 97-101.
Gollenberg. AL; Hediger. ML; Lee. PA; Himes. JH; Buck Louis. GM. (2010). Association between lead
and cadmium and reproductive hormones in peripubertal U.S. girls. Environ Health Perspect 118:
1782-1787. http://dx.doi.org/10.1289/ehp.1001943
Gomaa. A; Hu. H; Bellinger. D; Schwartz. J: Tsaih. SW: Gonzalez-Cossio. T; Schnaas. L; Peterson. K;
Aro. A; Hernandez-Avila. M. (2002). Maternal bone lead as an independent risk factor for fetal
neurotoxicity: A prospective study. Pediatrics 110: 110-118.
Gomez-Ramirez. P..; Martinez-Lopez. E..; Maria-Mojica. P..: Leon-Ortega. M..; Garcia-Fernandez. A..
1 (2011). Blood lead levels and 5-ALAD inhibition in nestlings of Eurasian Eagle Owl (Bubo bubo)
to assess lead exposure associated to an abandoned mining area. Ecotoxicology 20: 131-138.
http://dx.doi.org/10.1007/sl0646-010-0563-3
Gong. Z; Evans. HL. (1997). Effect of chelation with meso-dimercaptosuccinic acid (DMSA) before and
after the appearance of lead-induced neurotoxicity in the rat. Toxicol Appl Pharmacol 144: 205-214.
http://dx.doi.org/10.1006/taap.1997.8118
Gonick. HC: Cohen. AH; Ren. Q; Saldanha. LF; Khalil-Manesh. F; Anzalone. J: Sun. YY. (1996). Effect
of 2,3-dimercaptosuccinic acid on nephrosclerosis in the Dahl rat I role of reactive oxygen species.
Kidney Int 50: 1572-1581. http://dx.doi.org/10.1038/ki.1996.473
Gonick. HC: Ding. Y; Bondv. SC: Ni. Z; Vaziri. ND. (1997). Lead-induced hypertension: Interplay of
nitric oxide and reactive oxygen species. Hypertension 30: 1487-1492.
Gonick. HC: Ding. Y; Vaziri. ND. (1998). Effect of low lead exposure on eicosanoid excretion in rats.
Prostaglandins Other Lipid Mediat 55: 77-82. http://dx.doi.org/10.1016/S0090-6980(98)00010-0
Gonick. HC: Khalil-Manesh. F; Raghavan. SRV. (1985). Characterization of human erythrocyte lead-
binding protein. In TD Lekkas (Ed.), Heavy metals in the environment: Athens ~ September 1985 (pp.
313-316). Edinburgh, U.K.: CEP Consultants.
November 2012 5-734 Draft - Do Not Cite or Quote
-------
Gorell. JM; Johnson. CC: Rybicki. BA; Peterson. EL; Kortsha. GX; Brown. GG: Richardson. RJ. (1997).
Occupational exposures to metals as risk factors for Parkinson's disease. Neurology 48: 650-658.
Goswami. K; Gachhui. R; Bandopadhyav. A. (2005). Hepatorenal dysfunctions in lead pollution. J
Environ Eng Sci 47: 75-80.
Governa. M; Valentino. M; Visona. I. (1987). In vitro impairment of human granulocyte functions by
lead. Arch Toxicol 59: 421-425. http://dx.doi.org/10.1007/BF00316208
Gover. RA. (1968). The renal tubule in lead poisoning: I. mitochondrial swelling and aminoaciduria. Lab
Invest 19: 71-77.
Gover. RA; Cherian. MG: Delaquerriere-Richardson. L. (1978). Renal effects of repeated administration
of calcium disodium ethylenediaminetetraacetate during excessive exposure to lead in rats. J Environ
Pathol Toxicol Oncol 1: 403-410.
Gover. RA; Krall. A; Kimball. JP. (1968). The renal tubule in lead poisoning: II. In vitro studies of
mitochondrial structure and function. Lab Invest 19: 78-83.
Gover. RA: Leonard. PL; Moore. JF; Rhyne. B; Krigman. MR. (1970a). Lead dosage and the role of the
intranuclear inclusion body: An experimental study. Arch Environ Health 20: 705-711.
Gover. RA: May. P; Gates. MM; Krigman. MR. (1970b). Lead and protein content of isolated intranuclear
inclusion bodies from kidneys of lead-poisoned rats. Lab Invest 22: 245-251.
Gover. RA; Rhyne. BC. (1973). Pathological effects of lead [Review]. Int Rev Exp Pathol 12: 1-77.
Gozdzik-Zolnierkiewicz. T; Moszvnski. B. (1969). VIII nerve in experimental lead poisoning. Acta
Otolaryngol 68: 85-89. http://dx.doi.org/10.3109/00016486909121546
Grabowska. M; Guminska. M. (1996). The effect of lead on lactate formation, ATP level and membrane
ATPase activities in human erythrocytes in vitro. Int J Occup Med Environ Health 9: 265-274.
Gracia. CR; Sammel. MD; Coutifaris. C: Guzick. DS: Barnhart. KT. (2005). Occupational exposures and
male infertility. Am J Epidemiol 162: 729-733. http://dx.doi.org/10.1093/aie/kwi269
Graf. U: Wiirgler. FE. (1986). The present status of validation of the wing spot test in Drosophila. Prog
Clin Biol Res 209B: 391-398.
Gramigni. E; Tadini-Buoninsegni. F; Bartolommei. G: Santini. G: Chelazzi. G: Moncelli. MR. (2009).
Inhibitory effect of Pb2+ on the transport cycle of the Na+,K+ATPase. Chem Res Toxicol 22:
1699-1704. http://dx.doi.org/10.1021/tx9001786
Grant. LD; Kimmel. CA; West. GL; Martinez-Vargas. CM; Howard. JL. (1980). Chronic low-level lead
toxicity in the rat: II. Effects on postnatal physical and behavioral development. Toxicol Appl
Pharmacol 56: 42-58. http://dx.doi.org/10.1016/0041-008X(80)90130-l
Graves. AB; Van Duijn. CM; Chandra. V: Fratiglioni. L; Heyman. A; Jorm. AF; Kokmen. E; Kondo. K;
Mortimer. JA; Rocca. WA; Shalat. SL; Soininen. H; Hofman. A. (1991). Occupational exposures to
solvents and lead as risk factors for Alzheimer's disease: a collaborative re-analysis of case-control
studies. Int J Epidemiol 2: S58-S61.
Graziano. J: Slavkovich. V; Liu X Factor-Litvak. P; Todd. A. (2004). A prospective study of prenatal and
childhood lead exposure and erythropoietin production. J Occup Environ Med 46: 924-929.
Graziano. JH; Slavkovic. V: Factorlitvak. P; Popovac. D; Ahmedi. X; Mehmeti. A. (1991). Depressed
serum erythropoietin in pregnant women with elevated blood lead. Arch Environ Occup Health 46:
347-350.
Greene. T; Ernhart. CB. (1993). Dentine lead and intelligence prior to school entry: A statistical
sensitivity analysis. J Clin Epidemiol 46: 323-339. http://dx.doi.org/10.1016/0895-4356(93)90147-S
Greene. T; Ernhart. CB; Boyd. TA. (1992). Contributions of risk factors to elevated blood and dentine
lead levels in preschool children. Sci Total Environ 115: 239-260.
November 2012 5-735 Draft - Do Not Cite or Quote
-------
Grizzo. LT; Cordellini. S. (2008). Perinatal lead exposure affects nitric oxide and cyclooxygenase
pathways in aorta of weaned rats. Toxicol Sci 103: 207-214. http://dx.doi.org/10.1093/toxsci/kfnO 18
Grlickova-Duzevik. E; Wise. SS: Munroe. RC: Thompson. WD; Wise. JP. Sr. (2006). XRCC1 protects
against paniculate chromate-induced chromosome damage and cytotoxicity in Chinese hamster ovary
cells. Toxicol Sci 92: 409-415. http://dx.doi.org/!0.1093/toxsci/kfl021
Grover. P; Rekhadevi. PV: Danadevi. K; Vuyyuri. SB; Mahboob. M; Rahman. MF. (2010). Genotoxicity
evaluation in workers occupationally exposed to lead. Int J Hyg Environ Health 213: 99-106.
http://dx.doi.0rg/10.1016/i.ijheh.2010.01.005
Guilarte. TR; McGlothan. JL. (1998). Hippocampal NMDA receptor mRNA undergoes subunit specific
changes during developmental lead exposure. Brain Res 790: 98-107.
http://dx.doi.org/10.1016/S0006-8993(98)00054-7
Guilarte. TR; Toscano. CD; Mcglothan. JL; Weaver. SA. (2003). Environmental enrichment reverses
cognitive and molecular deficits induced by developmental lead exposure. Ann Neurol 53: 50-56.
http://dx.doi.org/10.1002/ana.10399
Gulson. B; Mahaffev. KR; Jameson. CW: Patison. N; Law. AJ; Mizon. KJ; Korsch. MJ; Pederson. D.
(1999). Impact of diet on lead in blood and urine in female adults and relevance to mobilization of
lead from bone stores. Environ Health Perspect 107: 257-263. http://dx.doi.org/10.1289/ehp.99107257
Gulson. B; Mizon. KJ: Korsch. MJ: Palmer. JM; Donnelly. JB. (2003). Mobilization of lead from human
bone tissue during pregnancy and lactation: A summary of long-term research. Sci Total Environ 303:
79-104. http://dx.doi.org/10.1016/S0048-9697(02)00355-8
Gulson. B; Mizon. KJ: Palmer. JM; Korsch. MJ: Taylor. AJ: Mahaffev. KR. (2004a). Blood lead changes
during pregnancy and postpartum with calcium supplementation. Environ Health Perspect 112:
1499-1507. http://dx.doi.org/10.1289/ehp.6548
Gulson. BL; Cameron. MA; Smith. AJ: Mizon. KJ: Korsch. MJ: Vimpani. G: McMichael. AJ: Pisaniello.
D; Jameson. CW: Mahaffev. KR. (1998c). Blood lead urine lead relationships in adults and children.
Environ Res 2: 152-160. http://dx.doi.org/10.1006/enrs.1997.3810
Gump. BB; Mackenzie. JA; Bendinskas. K; Morgan. R; Dumas. AK; Palmer. CD; Parsons. PJ. (2011).
Low-level Pb and cardiovascular responses to acute stress in children: the role of cardiac autonomic
regulation. Neurotoxicol Teratol 33: 212-219. http://dx.doi.Org/10.1016/i.ntt.2010.10.001
Gump. BB; Reihman. J; Stewart. P; Lonky. E; Darvill. T; Matthews. KA. (2007). Blood lead (Pb) levels:
A potential environmental mechanism explaining the relation between socioeconomic status and
cardiovascular reactivity in children. Health Psychol 26: 296-304.
http://dx.doi.0rg/10.1037/0278-6133.26.3.296
Gump. BB; Reihman. J: Stewart. P: Lonky. E; Granger. DA; Matthews. KA. (2009). Blood lead (Pb)
levels: Further evidence for an environmental mechanism explaining the association between
socioeconomic status and psychophysiological dysregulation in children. Health Psychol 28: 614-620.
http://dx.doi.org/10.1037/a0015611
Gump. BB; Stewart. P: Reihman. J: Lonky. E; Darvill. T; Matthews. KA; Parsons. PJ. (2005). Prenatal
and early childhood blood lead levels and cardiovascular functioning in 9 1/2 year old children.
Neurotoxicol Teratol 27: 655-665. http://dx.doi.Org/10.1016/i.ntt.2005.04.002
Gump. BB; Stewart. P: Reihman. J: Lonky. E; Darvill. T; Parsons. PJ: Granger. DA. (2008). Low-level
prenatal and postnatal blood lead exposure and adrenocortical responses to acute stress in children.
Environ Health Perspect 116: 249-255. http://dx.doi.org/10.1289/ehp.10391
Gundacker. C: Frohlich. S: Graf-Rohrmeister. K; Eibenberger. B; Jessenig. V; Gicic. D: Prinz. S:
Wittmann. KJ: Zeisler. H; Valiant. B; Pollak. A; Husslein. P. (2010). Perinatal lead and mercury
exposure in Austria. Sci Total Environ 408: 5744-5749.
http://dx.doi.0rg/10.1016/i.scitotenv.2010.07.079
November 2012 5-736 Draft - Do Not Cite or Quote
-------
Guo. TL; Mudzinski. SP; Lawrence. DA. (1996a). The heavy metal lead modulates the expression of both
TNF-alpha and TNF-alpha receptors in lipopolysaccharide-activated human peripheral blood
mononuclear cells. J Leukoc Biol 59: 932-939.
Guo. TL; Mudzinski. SP: Lawrence. DA. (1996b). Regulation of HLA-DR and invariant chain expression
by human peripheral blood mononuclear cells with lead, interferon-gamma, or interleukin-4. Cell
Immunol 171: 1-9. http://dx.doi.org/10.1006/cimm.1996.0166
Gupta. P: Husain. MM; Shankar. R; Seth. PK; Maheshwari. RK. (2002). Lead exposure enhances virus
multiplication and pathogenesis in mice. Vet Hum Toxicol 44: 205-210.
Gustafson. A; Hedner. P; Schiitz. A; Skerfving. S. (1989). Occupational lead exposure and pituitary
function. Int Arch Occup Environ Health 61: 277-281. http://dx.doi.org/10.1007/BF00381426
Gustavsson. P; Plato. N; Hallqvist. J: Hogstedt. C: Lewne. M; Reuterwall. C: Scheele. P. (2001). A
population-based case-referent study of myocardial infarction and occupational exposure to motor
exhaust, other combustion products, organic solvents, lead, and dynamite Stockholm heart
epidemiology program (SHEEP) study group. Epidemiology 12: 222-228.
Habermann. E; Crowell. K; Janicki. P. (1983). Lead and other metals can substitute for Ca2+ in
calmodulin. Arch Toxicol 54: 61-70. http://dx.doi.org/10.1007/BF00277816
Hage. FG: Szalai. AJ. (2007). C-reactive protein gene polymorphisms, C-reactive protein blood levels,
and cardiovascular disease risk [Review]. JAm Coll Cardiol 50: 1115-1122.
http://dx.doi.0rg/10.1016/i.iacc.2007.06.012
HaMai. D; Bondv. SC. (2004). Oxidative basis of manganese neurotoxicity [Review]. Ann N Y Acad Sci
1012: 129-141. http://dx.doi.org/10.1196/annals.1306.010
Hamed. EA; Meki. AR; Abd El-Mottaleb. NA. (2010). Protective effect of green tea on lead-induced
oxidative damage in rat's blood and brain tissue homogenates. J Physiol Biochem 66: 143-151.
http://dx.doi.org/10.1007/sl3105-010-0019-5
Hanas. JS: Rodgers. JS: Bantle. JA; Cheng. YG. (1999). Lead inhibition of DNA-binding mechanism of
Cys(2)His(2) zinc finger proteins. Mol Pharmacol 56: 982-988.
Hansen. JA; Audet. D; Spears. BL; Healy. KA; Brazzle. RE; Hoffman. DJ; Dailev. A; Beyer. WN.
(201 la). Lead exposure and poisoning of songbirds using the Coeur d'Alene River Basin, Idaho, USA.
Integr Environ Assess Manag 7: 587-595. http://dx.doi.org/10.1002/ieam.201
Hara. N; Yamada. K; Terashima. M; Osago. H; Shimoyama. M; Tsuchiya. M. (2003). Molecular
identification of human glutamine- and ammonia-dependent NAD synthetases: Carbon-nitrogen
hydrolase domain confers glutamine dependency. J Biol Chem 278: 10914-10921.
http://dx.doi.org/10.1074/ibc.M209203200
Haraguchi. T; Ishizu. H; Takehisa. Y; Kawai. K; Yokota. O: Terada. S: Tsuchiya. K; Ikeda. K; Morita. K;
Horike. T; Kira. S: Kuroda. S. (2001). Lead content of brain tissue in diffuse neurofibrillary tangles
with calcification (DNTC): the possibility of lead neurotoxicity. Neuroreport 12: 3887-3890.
Harry. GJ: Schmitt TJ; Gong. Z; Brown. H; Zawia. N; Evans. HL. (1996). Lead-induced alterations of
glial fibrillary acidic protein (GFAP) in the developing rat brain. Toxicol Appl Pharmacol 139: 84-93.
http://dx.doi.org/10.1006/taap. 1996.0145
Hartwig. A; Schlepegrell. R; Beversmann. D. (1990). Indirect mechanism of lead-induced genotoxicity in
cultured mammalian cells. Mutat Res Genet Toxicol 241: 75-82.
http://dx.doi.org/10.1016/0165-1218(90)90110-N
Hashemzadeh-Gargari. H; Guilarte. TR. (1999). Divalent cations modulate N-methyl-D-aspartate receptor
function at the glycine site. J Pharmacol Exp Ther 290: 1356-1362.
Hashmi. NS: Kachru. DN; Khandelwal. S: Tandon. SK. (1989). Interrelationship between iron deficiency
and lead intoxication (part 2). Biol Trace Elem Res 22: 299-307.
http://dx.doi.org/10.1007/BF02916618
November 2012 5-737 Draft - Do Not Cite or Quote
-------
Hauser. R; Sergevev. O: Korrick. S: Lee. MM; Revich. B; Gitin. E; Burns. JS: Williams. PL. (2008).
Association of blood lead levels with onset of puberty in Russian boys. Environ Health Perspect 116:
976-980. http://dx.doi.org/10.1289/ehp.10516
Hausmann. OV; Gentinetta. T; Fux. M; Ducrest. S: Pichler. WJ: Dahinden. CA. (2011). Robust
expression of CCR3 as a single basophil selection marker in flow cytometry. Allergy 66: 85-91.
http://dx.doi.0rg/10.llll/i.1398-9995.2010.02431.x
He. LH; Poblenz. AT; Medrano. CJ: Fox. DA. (2000). Lead and calcium produce rod photoreceptor cell
apoptosis by opening the mitochondria! permeability transition pore. J Biol Chem 275: 12175-12184.
http://dx.doi.org/10.1074/ibc.275.16.12175
He. S: Dong. W: Deng. Q; Weng. S: Sun. W. (2003). Seeing more clearly: Recent advances in
understanding retinal circuitry [Review]. Science 302: 408-411.
http://dx.doi.org/10.1126/science.1085457
He. T; Hirsch. HVB; Ruden. DM; Lnenicka. GA. (2009). Chronic lead exposure alters presynaptic
calcium regulation and synaptic facilitation in Drosophila larvae. Neurotoxicology 30: 777-784.
http://dx.doi.0rg/10.1016/i.neuro.2009.08.007
Hechtenberg. S: Beversmann. D. (1991). Inhibition of sarcoplasmic reticulum Ca2+ATPase activity by
cadmium, lead and mercury. Enzyme 45: 109-115.
Hegazy. RM; Hamdv. R; Kamel. HF. (2011). Modulation of IgE levels in lead exposed children by
parental cigarette smoking, Qualyobia Governate, Egypt. Int J Pharma Bio Sci 2: 372-385.
Heier. LS: Lien. IB; Stromseng. AE; Ljones. M; Rosseland. BO; Tollefsen. KE; Salbu. B. (2009).
Speciation of lead, copper, zinc and antimony in water draining a shooting range: Time dependant
metal accumulation and biomarker responses in brown trout (Salmo trutta L.). Sci Total Environ 407:
4047-4055. http://dx.doi.Org/10.1016/i.scitotenv.2009.03.002
Heiman. AS: Tonner. LE. (1995). The acute effect of lead acetate on glucocorticoid regulation of tyrosine
aminotransferase in hepatoma cells. Toxicology 100: 57-68.
http://dx.doi.org/10.1016/0300-483X(95)03061-J
Hellstrom. L; Blinder. CG: Dahlberg. B; Lundberg. M; Jarup. L; Persson. B; Axelson. O. (2001).
Cadmium exposure and end-stage renal disease. Am J Kidney Dis 38: 1001-1008.
http://dx.doi.org/10.1053/aikd.2001.28589
Hemdan. NYA; Emmrich. F; Adham. K; Wichmann. G: Lehmann. I; El-Massrv. A; Ghoneim. H;
Lehmann. J: Sack. U. (2005). Dose-dependent modulation of the in vitro cytokine production of
human immune competent cells by lead salts. Toxicol Sci 86: 75-83.
http://dx.doi.org/10.1093/toxsci/kfil77
Hemphill. FE; Kaeberle. ML; Buck. WB. (1971). Lead suppression of mouse resistance to Salmonella
typhimurium. Science 172: 1031-1032. http://dx.doi.org/10.1126/science. 172.3987.1031
Hengstler. JG: Bolm-Audorff. U; Faldum. A; Janssen. K; Reifenrath. M; Gotte. W: Jung. D; Maver-
Popken. O; Fuchs. J: Gebhard. S: Bienfait. HG: Schlink. K; Dietrich. C: Faust. D; Epe. B; Oesch. F.
(2003). Occupational exposure to heavy metals: DNA damage induction and DNA repair inhibition
prove co-exposures to cadmium, cobalt and lead as more dangerous than hitherto expected.
Carcinogenesis 24: 63-73. http://dx.doi.Org/10.1093/carcin/24.l.63
Heo. Y; Lee. BK; Ahn. KD; Lawrence. DA. (2004). Serum IgE elevation correlates with blood lead levels
in battery manufacturing workers. Hum Exp Toxicol 23: 209-213.
http://dx.doi.org/10.1191/0960327104ht442oa
Heo. Y; Lee. WT; Lawrence. DA. (1997). In vivo the environmental pollutants lead and mercury induce
oligoclonal T cell responses skewed toward type-2 reactivities. Cell Immunol 179: 185-195.
http://dx.doi.org/10.1006/cimm.1997.1160
November 2012 5-738 Draft - Do Not Cite or Quote
-------
Heo. Y; Lee. WT; Lawrence. DA. (1998). Differential effects of lead and cAMP on development and
activities of Thl- and Th2-lymphocytes. Toxicol Sci 43: 172-185.
http://dx.doi.0rg/10.1093/toxsci/43.2.172
Heo. Y; Mondal. TK; Gao. D; Kasten-Jolly. J; Kishikawa. H; Lawrence. DA. (2007). Posttranscriptional
inhibition of interferon-gamma production by lead. Toxicol Sci 96: 92-100.
http://dx.doi.org/10.1093/toxsci/kfll82
Heo. Y; Parsons. PJ: Lawrence. DA. (1996). Lead differentially modifies cytokine production in vitro and
in vivo. Toxicol Appl Pharmacol 138: 149-157. http://dx.doi.org/10.1006/taap.1996.0108
Herman. DS: Geraldine. M; T. V. (2009). Influence of minerals on lead-induced alterations in liver
function in rats exposed to long-term lead exposure. J Hazard Mater 166: 1410-1414.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.12.070
Hermes-Lima. M; Pereira. B; Bechara. EJH. (1991). Are free radicals involved in lead poisoning?
[Review]. Xenobiotica 21: 1085-1090. http://dx.doi.org/10.3109/00498259109039548
Hernandez-Avila. M; Smith. D; Meneses. F; Sanin. LH; Hu. H. (1998). The influence of bone and blood
lead on plasma lead levels in environmentally exposed adults. Environ Health Perspect 106: 473-477.
http://dx.doi.org/10.1289/ehp.98106473
Hernandez-Franco. P; Silva. M; Valverde. M; Rojas. E. (2011). Induction of oxidative stress by low doses
of lead in human hepatic cell line WRL-68. Biometals 24: 951-958.
http://dx.doi.org/10.1007/sl0534-011-9453-2
Hernandez-Ochoa. I; Sanchez-Gutierrez. M; Solis-Heredia. MJ; Quintanilla-Vega. B. (2006).
Spermatozoa nucleus takes up lead during the epididymal maturation altering chromatin condensation.
Reprod Toxicol 21: 171-178. http://dx.doi.Org/10.1016/i.reprotox.2005.07.015
Hernandez-Serrato. MI; Fortoul. TI; Rojas-Martinez. R; Mendoza-Alvarado. LR; Canales-Trevino. L;
Bochichio-Riccardelli. T; Avila-Costa. MR; Olaiz-Fernandez. G. (2006). Lead blood concentrations
and renal function evaluation: Study in an exposed Mexican population. Environ Res 100: 227-231.
http://dx.doi.0rg/10.1016/i.envres.2005.03.004
Herold. C. (2010). NMDA and D2-like receptors modulate cognitive flexibility in a color discrimination
reversal task in pigeons. Behav Neurosci 124: 381-390. http://dx.doi.org/10.1037/a0019504
Hevdari. A; Norouzzadeh. A; Khoshbaten. A; Asgari. A; Ghasemi. A; Naiafi. S: Badalzadeh. R. (2006).
Effects of short-term and subchronic lead poisoning on nitric oxide metabolites and vascular
responsiveness in rat. Toxicol Lett 166: 88-94. http://dx.doi.Org/10.1016/i.toxlet.2006.05.014
Hilbertz. U; Kramer. U; De Ruiter. N; Baginski. B. (1986). Effects of cadmium and lead on oxidative
metabolism and phagocytosis by mouse peritoneal macrophages. Toxicology 39: 47-57.
Hilson. JA; Strupp. BJ. (1997). Analyses of response patterns clarify lead effects in olfactory reversal and
extradimensional shift tasks: Assessment of inhibitory control, associative ability, and memory. Behav
Neurosci 111: 532-542.
Holdstein. Y; Pratt. H; Goldsher. M; Rosen. G: Shenhav. R: Linn. S: Mor. A; Barkai. A. (1986). Auditory
brainstem evoked potentials in asymptomatic lead-exposed subjects. J Laryngol Otol 100: 1031-1036.
Hollowav. WR. Jr; Thor. DH. (1987). Low level lead exposure during lactation increases rough and
tumble play fighting of juvenile rats. Neurotoxicol Teratol 9: 51-57.
Holmes. AL; Wise. SS: Sandwick. SJ: Lingle. WL; Negron. VC: Thompson. WD; Wise. JP. Sr. (2006a).
Chronic exposure to lead chromate causes centrosome abnormalities and aneuploidy in human lung
cells. Cancer Res 66: 4041-4048. http://dx.doi.org/10.1158/0008-5472.can-05-3312
Holmes. AL; Wise. SS: Sandwick. SJ: Wise. JP. (2006b). The clastogenic effects of chronic exposure to
paniculate and soluble Cr(VI) in human lung cells. Mutat Res Genet Toxicol Environ Mutagen 610:
8-13. http://dx.doi.0rg/10.1016/i.mrgentox.2006.06.006
November 2012 5-739 Draft - Do Not Cite or Quote
-------
Holtzman. D; Olson. JE; DeVries. C: Bensch. K. (1987). Lead toxicity in primary cultured cerebral
astrocytes and cerebellar granular neurons. Toxicol Appl Pharmacol 89: 211-225.
http://dx.doi.org/10.1016/0041-008X(87)90042-l
Hon. KL; Ching. GK; Hung. EC: Leung. TF. (2009). Serum lead levels in childhood eczema [Letter]. Clin
Exp Dermatol 34: e508-e509. http://dx.doi.Org/10.llll/i.1365-2230.2009.03596.x
Hon. KLE; Wang. SS: Hung. ECW: Lam. HS: Lui. HHK; Chow. CM; Ching. GKW: Fok. TF; Ng. PC:
Leung. TF. (2010). Serum levels of heavy metals in childhood eczema and skin diseases: Friends or
foes. Pediatric Allergy and Immunology 21: 831-836.
http://dx.doi.0rg/10.llll/i.1399-3038.2010.01022.x
Hong. YC: Hwang. SS: Kim. JH; Lee. KH; Lee. HJ; Lee. KH; Yu. SD; Kim. PS. (2007b). Metals in
particulate pollutants affect peak expiratory flow of schoolchildren. Environ Health Perspect 115:
430-434. http://dx.doi.org/10.1289/ehp.9531
Horiguchi. S: Matsumura. S: Fukumoto. K; Karai. I; Endo. G: Teramoto. K; Shinagawa. K; Kiyota. I;
Wakitani. F; Takise. S: Kawarava. T. (1991). Erythrocyte deformability in workers exposed to lead.
Osaka City Med J 37: 149-155.
Hornung. R. (2008). Email from Richard Hornung, Cincinnati Children's Hospital Medical Center
(CCHMC) to Jee-Young Kim, USEPA Subject: Request for additional information on Lanphear et al.
(2005). (EPA-HQ-OAR-2006-0735-5814). U.S. Environmental Protection Agency.
http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR-2006-0735-5814
Hornung. R; Lanphear. B; Kietrich. K. (2006). Response to: "What is the meaning of non-linear dose-
response relationships between blood lead concentration and IQ?" [letter] [Letter]. Neurotoxicology
27: 635.
Hornung. RW: Lanphear. BP; Dietrich. KN. (2009). Age of greatest susceptibility to childhood lead
exposure: Anew statistical approach. Environ Health Perspect 117: 1309-1312.
http://dx.doi.org/10.1289/ehp.0800426
Hotz. P; Buchet. JP; Bernard. A; Lison. D; Lauwerys. R. (1999). Renal effects of low-level environmental
cadmium exposure: 5-year follow-up of a subcohort from the Cadmibel study. 354: 1508-1513.
http://dx.doi.org/10.1016/S0140-6736(99)91145-5
Howard. JK. (1974). Human erythrocyte glutathione reductase and glucose 6-phosphate dehydrogenase
activities in normal subjects and in persons exposed to lead. Clin Sci Mol Med 47: 515-520.
Hsiao. CL; Wu. KH: Wan. KS. (2011). Effects of environmental lead exposure on T-helper cell-specific
cytokines in children. J Immunotoxicol 8: 284-287. http://dx.doi.org/10.3109/1547691X.2011.592162
Hsiao. CY; Wu. HDI; Lai. JS: Kuo. HW. (2001). A longitudinal study of the effects of long-term exposure
to lead among lead battery factory workers in Taiwan (1989-1999). Sci Total Environ 279: 151-158.
http://dx.doi.org/10.1016/S0048-9697(01)00762-8
Hsieh. SJ: Chiu. YW: Li. WF; Wu. CH; Chen. HI; Chuang. HY. (2009a). Increased concentrations of
serum inhibin B among male workers with long-term moderate lead exposure. Sci Total Environ 407:
2603-2607. http://dx.doi.Org/10.1016/i.scitotenv.2008.12.056
Hsieh. TJ; Chen. YC: Li. CW: Liu. GC: Chiu. YW: Chuang. HY. (2009b). A proton magnetic resonance
spectroscopy study of the chronic lead effect on the basal ganglion and frontal and occipital lobes in
middle-age adults. Environ Health Perspect 117: 941-945. http://dx.doi.org/10.1289/ehp.0800187
Hsu. CW: Lin. JL: Lin-Tan. DT: Yen. TH: Huang. WH: Ho. TC: Huang. YL: Yeh. LM: Huang. LM.
(2009a). Association of environmental cadmium exposure with inflammation and malnutrition in
maintenance haemodialysis patients. Nephrol Dial Transplant 24: 1282-1288.
http://dx.doi.org/10.1093/ndt/gfn602
Hsu. JM. (1981). Lead toxicity as related to glutathione metabolism. JNutr 111: 26-33.
November 2012 5-740 Draft - Do Not Cite or Quote
-------
Hsu. PC: Chang. HY; Quo. YL; Liu. YC: Shih. TS. (2009b). Effect of smoking on blood lead levels in
workers and role of reactive oxygen species in lead-induced sperm chromatin DNA damage. Fertil
Steril91: 1096-1103. http://dx.doi.Org/10.1016/i.fertnstert.2008.01.005
Hsu. PC: Hsu. CC: Liu. MY; Chen. LY; Quo. YL. (1998a). Lead-induced changes in spermatozoa
function and metabolism. J Toxicol Environ Health A 55: 45-64.
http://dx.doi.org/10.1080/009841098158610
Hsu. PC: Liu. MY; Hsu. CC: Chen. LY; Quo. YL. (1998b). Effects of vitamin E and/or C on reactive
oxygen species-related lead toxicity in the rat sperm. Toxicology 128: 169-179.
http://dx.doi.org/10.1016/S0300-483X(98)00068-7
Hu. H. (1991). A 50-year follow-up of childhood plumbism: Hypertension, renal function, and
hemoglobin levels among survivors. Am J Dis Child 145: 681-687.
Hu. H; Aro. A; Pavton. M; Korrick. S: Sparrow. (1996a). The relationship of bone and blood lead to
hypertension: The Normative Aging Study. JAMA 275: 1171-1176.
http://dx.doi.org/10.1001/iama.1996.03530390037031
Hu. H; Hernandez-Avila. M. (2002). Lead, bones, women, and pregnancy: The poison within? Am J
Epidemiol 156: 1088-1091. http://dx.doi.org/10.1093/aje/kwfl64
Hu. H; Pavton. M; Korrick. S: Aro. A; Sparrow. D; Weiss. ST; Rotnitzky. A. (1996b). Determinants of
bone and blood lead levels among community-exposed middle-aged to elderly men: The Normative
Aging Study. Am J Epidemiol 144: 749-759.
Hu. H; Tellez-Rojo. MM; Bellinger. D; Smith. D; Ettinger. AS: Lamadrid-Figueroa. H; Schwartz. J:
Schnaas. L; Mercado-Garca. A; Hernandez-Avila. M. (2006). Fetal lead exposure at each stage of
pregnancy as a predictor of infant mental development. Environ Health Perspect 114: 1730-1735.
http://dx.doi.org/10.1289/ehp.9067
Hu. P; Wang. M; Chen. WH; Liu. J: Chen. L; Yin. ST; Yong. W: Chen. JT; Wang. HL; Ruan. DY
(2008a). Quercetin relieves chronic lead exposure-induced impairment of synaptic plasticity in rat
dentate gyrus in vivo. Naunyn Schmiedebergs Arch Pharmacol 378: 43-51.
http://dx.doi.org/10.1007/s00210-008-0301-z
Hu. OS: Fu. HJ: Ren. TL: Wang. SY: Zhou. W: Song. H: Han. YF: Dong. SZ. (2008b). Maternal
low-level lead exposure reduces the expression of PSA-NCAM and the activity of sialyltransferase in
the hippocampi of neonatal rat pups. Neurotoxicology 29: 675-681.
http://dx.doi.0rg/10.1016/i.neuro.2008.04.002
Huang. BM; Lai. HY; Liu. MY. (2002). Concentration dependency in lead-inhibited steroidogenesis in
MA-10 mouse Leydig tumor cells. J Toxicol Environ Health A 65: 557-567.
http://dx.doi.org/10.1080/15287390252808000
Huang. BM; Liu. MY. (2004). Inhibitory actions of lead on steroidogenesis in MA-10 mouse Leydig
tumor cells. Arch Androl 50: 5-9. http://dx.doi.org/10.1080/01485010490250434
Huang. H; Bihaqi. SW: Cui. L; Zawia. NH. (2011 a). In vitro Pb exposure disturbs the balance between
Ap production and elimination: the role of ApPP and neprilysin. Neurotoxicology 32: 300-306.
http://dx.doi.0rg/10.1016/i.neuro.2011.02.001
Huang. J; Wu. J: Li. T; Song. X; Zhang. B; Zhang. P; Zheng. X. (20lib). Effect of exposure to trace
elements in the soil on the prevalence of neural tube defects in a high-risk area of China. Biomed
Environ Sci 24: 94-101. http://dx.doi.org/10.3967/0895-3988.2011.02.002
Huang. M; Krepkiy. D; Hu. W: Petering. PH. (2004). Zn-, Cd-, and Pb-transcription factor IIIA:
Properties, DNA binding, and comparison with TFIIIA-fmger 3 metal complexes. J Inorg Biochem 98:
775-785. http://dx.doi.0rg/10.1016/i.iinorgbio.2004.01.014
Huang. XP; Feng. ZY; Zhai. WL; Xu. JH. (1988). Chromosomal aberrations and sister chromatid
exchanges in workers exposed to lead. Biomed Environ Sci 1: 382-387.
November 2012 5-741 Draft - Do Not Cite or Quote
-------
Hudson. CA; Cao. L; Kasten-Jolly. J; Kirkwood. JN; Lawrence. DA. (2003). Susceptibility of lupus-
prone NZM mouse strains to lead exacerbation of systemic lupus erythematosus symptoms. J Toxicol
Environ Health A 66: 895-918. http://dx.doi.org/10.1080/15287390306456
Huel. G: Sahuquillo. J: Debotte. G: Oury. JF; Takser. L. (2008). Hair mercury negatively correlates with
calcium pump activity in human term newborns and their mothers at delivery. Environ Health Perspect
116: 263-267. http://dx.doi.org/10.1289/ehp.10381
Huo. X; Peng. L; Xu. XJ; Zheng. LK; Qiu. B; Qi. ZL; Zhang. B; Han. D; Piao. ZX. (2007). Elevated
blood lead levels of children in Guiyu, an electronic waste recycling town in China. Environ Health
Perspect 115: 1113-1117. http://dx.doi.org/10.1289/ehp.9697
Huseman. CA; Moriartv. CM; Angle. CR. (1987). Childhood lead toxicity and impaired release of
thyrotropin-stimulating hormone. Environ Res 42: 524-533.
http://dx.doi.org/10.1016/80013-9351(87)80219-0
Huseman. CA; Varma. MM; Angle. CR. (1992). Neuroendocrine effects of toxic and low blood lead
levels in children [Letter]. Pediatrics 90: 186-189.
Hwang. KY; Lee. BK; Bressler. JP; Bolla. KI; Stewart. WF; Schwartz. BS. (2002). Protein kinase C
activity and the relations between blood lead and neurobehavioral function in lead workers. Environ
Health Perspect 110: 133-138. http://dx.doi.org/10.1289/ehp.02110133
Hwang. YH; Chiang. HY; Yen-Jean. MC: Wang. JD. (2009). The association between low levels of lead
in blood and occupational noise-induced hearing loss in steel workers. Sci Total Environ 408: 43-49.
http://dx.doi.0rg/10.1016/i.scitotenv.2009.09.016
Hwua. YS: Yang. JL. (1998). Effect of 3-aminotriazole on anchorage independence and mutagenicity in
cadmium- and lead-treated diploid human fibroblasts. Carcinogenesis 19: 881-888.
http://dx.doi.0rg/10.1093/carcin/19.5.881
IARC (International Agency for Research on Cancer). (2006a). Inorganic and organic lead compounds.
Lyon, France. http://monographs.iarc.fr/ENG/Monographs/vol87/index.php
lavicoli. I; Carelli. G: III. SE; Castellino. N; Calabrese. EJ. (2004). Effects of low doses of dietary lead on
puberty onset in female mice. Reprod Toxicol 19: 35-41.
http://dx.doi.0rg/10.1016/i.reprotox.2004.06.013
lavicoli. I; Carelli. G: Stanek. EJ: Castellino. N; Li. Z; Calabrese. EJ. (2006a). Low doses of dietary lead
are associated with a profound reduction in the time to the onset of puberty in female mice. Reprod
Toxicol 22: 586-590. http://dx.doi.Org/10.1016/i.reprotox.2006.03.016
lavicoli. I: Carelli. G: Stanek. EJ. Ill; Castellino. N; Calabrese. EJ. (2006b). Below background levels of
blood lead impact cytokine levels in male and female mice. Toxicol Appl Pharmacol 210: 94-99.
http://dx.doi.0rg/10.1016/i.taap.2005.09.016
lavicoli. I: Sgambato. A; Carelli. G: Ardito. R; Cittadini. A; Castellino. N. (2001). Lead-related effects on
rat fibroblasts. Mol Cell Biochem 222: 35-40. http://dx.doi.Org/10.1023/A:1017930401906
Ignasiak. Z; Slawinska. T; Rozek. K; Little. BB; Malina. RM. (2006). Lead and growth status of
schoolchildren living in the copper basin of south-western Poland: Differential effects on bone growth.
Ann Hum Biol 33: 401-414. http://dx.doi.org/10.1080/03014460600730752
lijima. K; Otake. T; Yoshinaga. J: Ikegami. M; Suzuki. E; Naruse. H; Yamanaka. T; Shibuva. N;
Yasumizu. T; Kato. N. (2007). Cadmium, lead, and selenium in cord blood and thyroid hormone status
of newborns. Biol Trace Elem Res 119: 10-18. http://dx.doi.org/10.1007/sl2011-007-0057-l
Ikebuchi. H; Teshima. R; Suzuki. K; Terao. T; Yamane. Y. (1986). Simultaneous induction of
Pb-metallothionein-like protein and Zn-thionein in the liver of rats given lead acetate. Biochem J 233:
541-546.
Ingelsson. E; Gona. P; Larson. MG: Lloyd-Jones. DM; Kannel. WB; Vasan. RS: Lew. D. (2008). Altered
blood pressure progression in the community and its relation to clinical events. Arch Intern Med 168:
1450-1457. http://dx.doi.org/10.1001/archinte.168.13.1450
November 2012 5-742 Draft - Do Not Cite or Quote
-------
Inglis. JA; Henderson. DA; Emmerson. BT. (1978). The pathology and pathogenesis of chronic lead
nephropathy occurring in Queensland. J Pathol 124: 65-76. http://dx.doi.org/10.1002/path.1711240202
Institoris. L; Kovacs. D; Kecskemeti-Kovacs. I; Lukacs. A; Szabo. A; Lengyel. Z; Papp. A;
Nagymaitenyi. L; Desi. I. (2006). Immunotoxicological investigation of subacute combined exposure
with low doses of PB, HG and CD in rats. Acta Biol Hung 57: 433-439.
http://dx.doi.Org/10.1556/ABiol.57.2006.4.5
lonescu. JG: Novotny. J; Stejskal. V: Latsch. A; Blaurock-Busch. E; Eisenmann-Klein. M. (2007). Breast
tumours strongly accumulate transition metals. Maedica 2: 5-9.
Iranpour. R; Besharati. AA; Nasseri. F; Hashemipour. M; Balali-Mood. M; Kelishadi. R. (2007).
Comparison of blood lead levels of mothers and cord blood in intrauterine growth retarded neonates
and normal term neonates. Saudi Med J 28: 877-880.
Irgens. A; Kruger. K; Skorve. AH; Irgens. LM. (1998). Reproductive outcome in offspring of parents
occupationally exposed to lead in Norway. Am J Ind Med 34: 431-437.
Iwata. T; Yano. E; Karita. K; Dakeishi. M; Murata. K. (2005). Critical dose of lead affecting postural
balance in workers. Am J Ind Med 48: 319-325. http://dx.doi.org/10.1002/ajim.20220
Jabeen. R; Tahir. M; Waqas. S. (2010). Teratogenic effects of lead acetate on kidney. J Ayub Med Coll
Abbottabad 22: 76-79.
Jackson. LW: Howards. PP; Wactawski-Wende. J: Schisterman. EF. (2011). The association between
cadmium, lead and mercury blood levels and reproductive hormones among healthy, premenopausal
women. Hum Reprod 26: 2887-2895. http://dx.doi.org/10.1093/humrep/der250
Jadhav. SH; Sarkar. SN; Ram. GC: Tripathi. HC. (2007). Immunosuppressive effect of subchronic
exposure to a mixture of eight heavy metals, found as groundwater contaminants in different areas of
India, through drinking water in male rats. Arch Environ Contam Toxicol 53: 450-458.
http://dx.doi.org/10.1007/s00244-006-0177-l
Jagetia. GC: Aruna. R. (1998). Effect of various concentrations of lead nitrate on the induction of
micronuclei in mouse bone marrow. Mutat Res Genet Toxicol Environ Mutagen 415: 131-137.
http://dx.doi.org/10.1016/S1383-5718(98)00052-7
Jain. NB; Potula. V: Schwartz. J: Vokonas. PS: Sparrow. D; Wright. RO: Nie. H; Hu. H. (2007). Lead
levels and ischemic heart disease in a prospective study of middle-aged and elderly men: The VA
Normative Aging Study. Environ Health Perspect 115: 871-875. http://dx.doi.org/10.1289/ehp.9629
Jamieson. JA; Shuhyta. JN; Taylor. CG. (2007). Lead does not affect transcription of intestinal zinc-
binding proteins in growing rats. Exp Biol Med 232: 744-753.
Jamieson. JA: Stringer. DM; Zahradka. P; Taylor. CG. (2008). Dietary zinc attenuates renal lead
deposition but metallothionein is not directly involved. Biometals 21: 29-40.
http://dx.doi.org/10.1007/sl0534-007-9090-v
Jang. HO; Kim. JS: Kwon. WC: Kim. JK; Ko. MS: Kim. DH; Kim. WI; Jeon. YC: Chung. IK; Shin. SH;
Chung. J; Bae. MK; Yun. I. (2008). The effect of lead on calcium release activated calcium influx in
primary cultures of human osteoblast-like cells. Arch Pharm Res 31: 188-194.
http://dx.doi.org/10.1007/sl2272-001-1140-3
Jang. WH: Lim. KM: Kim. K: Noh. JY: Kang. S: Chang. YK: Chung. JH. (2011). Low level of lead can
induce phosphatidylserine exposure and erythrophagocytosis: Anew mechanism underlying lead-
associated anemia. Toxicol Sci 122: 177-184. http://dx.doi.org/10.1093/toxsci/kfr079
Janjua. NZ; Delzell. E; Larson. RR; Meleth. S: Kristensen. S: Kabagambe. E; Sathiakumar. N. (2009).
Determinants of low birth weight in urban Pakistan. Public Health Nutr 12: 789-798.
http://dx.doi.org/10.1017/S1368980008002942
Javakumar. T; Sridhar. MP; Bharathprasad. TR; Ilavaraia. M; Govindasamy. S: Balasubramanian. MP
(2009). Experimental studies of Achyranthes aspera (L) preventing nephrotoxicity induced by lead in
albino rats. J Health Sci 55: 701-708. http://dx.doi.org/10.1248/ihs.55.701
November 2012 5-743 Draft - Do Not Cite or Quote
-------
Jedrychowski. W: Perera. F; Maugeri. U; Miller. RL; Rembiasz. M; Flak. E; Mroz. E; Majewska. R;
Zembala. M. (2011). Intrauterine exposure to lead may enhance sensitization to common inhalant
allergens in early childhood: Aprospective prebirth cohort study. Environ Res 111: 119-124.
http://dx.doi.0rg/10.1016/i.envres.2010.ll.002
Jedrychowski. W: Perera. FP; Jankowski. J; Mrozek-Budzyn. D; Mroz. E; Flak. E; Edwards. S: Skarupa.
A; Lisowska-Miszczyk. I. (2009b). Very low prenatal exposure to lead and mental development of
children in infancy and early childhood: Krakow prospective cohort study. Neuroepidemiology 32:
270-278. http://dx.doi.org/10.1159/000203075
Jehan. ZS: Motlag. DB. (1995). Metal induced changes in the erythrocyte membrane of rats. Toxicol Lett
78: 127-133. http://dx.doi.org/10.1016/0378-4274(94)03245-3
Jelliffe-Pawlowski. LL; Miles. SO; Courtney. JG: Materna. B; Charlton. V. (2006). Effect of magnitude
and timing of maternal pregnancy blood lead (Pb) levels on birth outcomes. J Perinatol 26: 154-162.
http://dx.doi.org/10.1038/si.irj.7211453
Jett. DA; Kuhlmann. AC: Farmer. SJ: Guilarte. TR. (1997). Age-dependent effects of developmental lead
exposure on performance in the Morris water maze. Pharmacol Biochem Behav 57: 271-279.
http://dx.doi.org/10.1016/S0091-3057(96)00350-4
Jiang. YM; Long. LL; Zhu. XY; Zheng. H; Fu. X; Ou. SY; Wei. PL; Zhou. HL; Zheng. W. (2008).
Evidence for altered hippocampal volume and brain metabolites in workers occupationally exposed to
lead: A study by magnetic resonance imaging and 1H magnetic resonance spectroscopy. Toxicol Lett
181: 118-125. http://dx.doi.0rg/10.1016/j.toxlet.2008.07.009
Jin. YP; Liao. YJ; Lu. CW: Li. GX; Yu. F; Zhi. XP; Xu. JH; Liu. SX; Liu. MM; Yang. J. (2006). Health
effects in children aged 3-6 years induced by environmental lead exposure. Ecotoxicol Environ Saf
63: 313-317. http://dx.doi.Org/10.1016/i.ecoenv.2005.05.011
Johnson. BL; Elbatawi. M; Xintaras. C: Baker. EL. Jr; Hanninen. H; Seppalainen. AM. (1987).
Prevention of neurotoxic illness in working populations. In BL Johnson; M Elbatawi; C Xintaras; EL
Baker, Jr.; H Hanninen; AM Seppalainen (Eds.). New York: John Willey & Sons.
Johnson. RJ; Kang. DH; Feig. D; Kivlighn. S; Kanellis. J; Watanabe. S; Turtle. KR; Rodriguez-Iturbe. B;
Herrera-Acosta. J; Mazzali. M. (2003). Is there a pathogenetic role for uric acid in hypertension and
cardiovascular and renal disease? Hypertension 41: 1183-1190.
http://dx.doi.org/10.1161/01.HYP.0000069700.62727.C5
Johnston. C; Mash. EJ; Miller. N; Ninowski. JE. (2012). Parenting in adults with attention-
deficit/hyperactivity disorder (ADHD) [Review]. Clin Psychol Rev 32: 215-228.
http://dx.doi.0rg/10.1016/i.cpr.2012.01.007
Jomova. K; Jenisova. Z; Feszterova. M; Baros. S; Liska. J; Hudecova. D; Rhodes. CJ; Valko. M. (2011).
Arsenic: Toxicity, oxidative stress and human disease [Review]. J Appl Toxicol 31: 95-107.
http://dx.doi.org/10.1002/iat.1649
Jomova. K; Valko. M. (2011). Advances in metal-induced oxidative stress and human disease. Toxicology
283: 65-87. http://dx.doi.Org/10.1016/j.tox.2011.03.001
Jones. AYM; Lam. PKW; Dean. E. (2006). Respiratory health of bus drivers in Hong Kong. Int Arch
Occup Environ Health 79: 414-418. http://dx.doi.org/10.1007/s00420-005-0061-8
Jones. AYM; Lam. PKW; Gohel. MDI. (2008). Respiratory health of road-side vendors in a large
industrialized city. Environ Sci Pollut Res Int 15: 150-154. http://dx.doi.org/10.1065/espr2006.12.368
Jones. EA; Wright. JM; Rice. G; Buckley. BT; Magsumbol. MS; Barr. DB; Williams. BL. (2010). Metal
exposures in an inner-city neonatal population. Environ Int 36: 649-654.
http://dx.doi.0rg/10.1016/i.envint.2010.04.007
Jones. SR; Atkin. P; Holroyd. C; Lutman. E; Batlle. JVI; Wakeford. R; Walker. P. (2007). Lung cancer
mortality at a U.K. tin smelter. Occup Med (Lond) 57: 238-245.
http://dx.doi.org/10.1093/occmed/kqll53
November 2012 5-744 Draft - Do Not Cite or Quote
-------
Joseph. CLM; Havstad. S: Ownbv. PR; Peterson. EL; Maliarik. M; McCabe. MJ. Jr; Barone. C: Johnson.
CC. (2005). Blood lead level and risk of asthma. Environ Health Perspect 113: 900-904.
http://dx.doi.org/10.1289/ehp.7453
Jurczuk. M; Brzoska. MM; Moniuszko-Jakoniuk. J. (2007). Hepatic and renal concentrations of vitamins
E and C in lead- and ethanol-exposed rats. An assessment of their involvement in the mechanisms of
peroxidative damage. Food Chem Toxicol 45: 1478-1486. http://dx.doi.Org/10.1016/i.fct.2007.02.007
Jurczuk. M; Moniuszko-Jakoniuk. J; Brzoska. MM. (2006). Involvement of some low-molecular thiols in
the peroxidative mechanisms of lead and ethanol action on rat liver and kidney. Toxicology 219:
11-21. http://dx.doi.0rg/10.1016/i.tox.2005.10.022
Jusko. TA; Henderson. CR; Lanphear. BP; Cory-Slechta. DA; Parsons. PJ: Canfield. RL. (2008). Blood
lead concentrations < 10 microg/dL and child intelligence at 6 years of age. Environ Health Perspect
116: 243-248. http://dx.doi.org/10.1289/ehp.10424
Kaczynska. K; Walski. M; Szereda-Przestaszewska. M. (2011). Ultrastructural changes in lung tissue after
acute lead intoxication in the rat. J Electron Microsc (Tokyo) 60: 289-294.
http://dx.doi.org/10.1093/imicro/dfr035
Kaji. T; Suzuki. M; Yamamoto. C: Mishima. A; Sakamoto. M; Kozuka. H. (1995). Severe damage of
cultured vascular endothelial cell monolayer after simultaneous exposure to cadmium and lead. Arch
Environ Contam Toxicol 28: 168-172. http://dx.doi.org/10.1007/BF00217612
Kalman. J: Riba. I; Blasco. J: Delvalls. TA. (2008). Is delta-aminolevulinic acid dehydratase activity in
bivalves from south-west Iberian Peninsula a good biomarker of lead exposure? Mar Environ Res 66:
38-40. http://dx.doi.0rg/10.1016/j.marenvres.2008.02.016
Kamal. M; Fathy. MM; Taher. E; Hasan. M; Tolba. M. (2011). Assessment of the role of paraoxonase
gene polymorphism (Q192R) and paraoxonase activity in the susceptibility to atherosclerosis among
lead-exposed workers. Ann Saudi Med 31: 481-487. http://dx.doi.org/10.4103/0256-4947.84625
Kamel. F; Umbach. DM; Hu. H; Munsat. TL; Shefner. JM; Taylor. JA; Sandier. DP. (2005). Lead
exposure as a risk factor for amyotrophic lateral sclerosis. Neurodegener Dis 2: 195-201.
http://dx.doi.org/10.1159/000089625
Kamel. F; Umbach. DM; Munsat. TL; Shefner. JM; Hu. H; Sandier. DP. (2002). Lead exposure and
amyotrophic lateral sclerosis. Epidemiology 13: 311-319.
Kamel. F; Umbach. DM; Stallone. L; Richards. M; Hu. H; Sandier. DP. (2008). Association of lead
exposure with survival in amyotrophic lateral sclerosis. Environ Health Perspect 116: 943-947.
http://dx.doi.org/10.1289/ehp.11193
Kanduc. D; Rossiello. MR; Aresta. A; Cavazza. C: Quagliariello. E; Farber. E. (1991). Transitory DNA
hypomethylation during liver cell proliferation induced by a single dose of lead nitrate. Arch Biochem
Biophys 286: 212-216. http://dx.doi.org/10.1016/0003-9861(91)90030-M
Kannel. WB. (2000a). Elevated systolic blood pressure as a cardiovascular risk factor [Review]. Am J
Cardiol 85: 251-255. http://dx.doi.org/10.1016/S0002-9149(99)00635-9
Kannel. WB. (2000b). Risk stratification in hypertension: New insights from the Framingham Study. Am
JHypertens 13: 3S-10S. http://dx.doi.org/10.1016/S0895-7061(99)00252-6
Kapuku. GK; Harshfield. GA; Davis. HC: Treiber. FA. (2006). Early markers of cardiovascular disease
[Review]. Vascul Pharmacol 45: 277-280. http://dx.doi.Org/10.1016/i.vph.2006.08.009
Karaca. T; Simsek. N. (2007). Effects of spirulina on the number of ovary mast cells in lead-induced
toxicity in rats. Phytother Res 21: 44-46. http://dx.doi.org/10.1002/ptr.2015
Karimooy. HN; Mood. MB; Hosseini. M; Shadmanfar. S. (2010). Effects of occupational lead exposure
on renal and nervous system of workers of traditional tile factories in Mashhad (northeast of Iran).
Toxicol Ind Health 26: 633-638. http://dx.doi.org/10.1177/0748233710377774
November 2012 5-745 Draft - Do Not Cite or Quote
-------
Karita. K; Yano. E; Dakeishi. M; Iwata. T; Murata. K. (2005). Benchmark dose of lead inducing anemia
at the workplace. Risk Anal 25: 957-962. http://dx.doi.Org/10.llll/i.1539-6924.2005.00652.x
Karmaus. W: Brooks. KR; Nebe. T; Witten. J; Obi-Osius. N; Kruse. H. (2005). Immune function
biomarkers in children exposed to lead and organochlorine compounds: A cross-sectional study.
Environ Health Global Access Sci Source 4: 5. http://dx.doi.org/10.1186/1476-069X-4-5
Kasperczyk. A; Kasperczyk. S: Horak. S: Ostalowska. A; Grucka-Mamczar. E; Romuk. E; Olejek. A;
Birkner. E. (2008). Assessment of semen function and lipid peroxidation among lead exposed men.
Toxicol Appl Pharmacol 228: 378-384. http://dx.doi.Org/10.1016/i.taap.2007.12.024
Kasperczyk. S: Kasperczyk. J; Ostalowska. A; Zalejska-Fiolka. J: Wielkoszynski. T; Swietochowska. E;
Birkner. E. (2009). The role of the antioxidant enzymes in erythrocytes in the development of arterial
hypertension among humans exposed to lead. Biol Trace Elem Res 130: 95-106.
http://dx.doi.org/10.1007/sl2011-009-8323-z
Kasprzak. KS: Hoover. KL; Poirier. LA. (1985). Effects of dietary calcium acetate on lead subacetate
carcinogenicity in kidneys of male Sprague- Dawley rats. Carcinogenesis 6: 279-282.
http://dx.doi.Org/10.1093/carcin/6.2.279
Kasten-Jolly. J: Heo. Y; Lawrence. DA. (2010). Impact of developmental lead exposure on splenic
factors. Toxicol Appl Pharmacol 247: 105-115. http://dx.doi.Org/10.1016/i.taap.2010.06.003
Kaufman. AS. (2001). Do low levels of lead produce IQ loss in children? A careful examination of the
literature. Arch Clin Neuropsychol 16: 303-341.
Kazi. TG: Jalbani. N; Kazi. N; Jamali. MK; Arain. MB; Afridi. HI; Kandhro. A; Pirzado. Z. (2008).
Evaluation of toxic metals in blood and urine samples of chronic renal failure patients, before and
after dialysis. Ren Fail 30: 737-745. http://dx.doi.org/10.1080/08860220802212999
Ke. H. (2004). Implications of PDE4 structure on inhibitor selectivity across PDE families. Int J Impot
Res 16: S24-S27. http://dx.doi.org/10.1038/sj.ijir.3901211
Kemp. FW: Neti. PVS. V: Howell. RW: Wenger. P; Louria. DB; Bogden. JD. (2007). Elevated blood lead
concentrations and vitamin D deficiency in winter and summer in young urban children. Environ
Health Perspect 115: 630-635. http://dx.doi.org/10.1289/ehp.9389
Kempe. DA; Lang. PA; Eisele. K; Klarl. BA; Wieder. T; Huber. SM; Duranton. C: Lang. F. (2005).
Stimulation of erythrocyte phosphatidylserine exposure by lead ions. Am J Physiol 288: C396-C402.
http://dx.doi.org/10.1152/ajpcell.00115.2004
Kempermann. G: Krebs. J: Fabel. K. (2008). The contribution of failing adult hippocampal neurogenesis
to psychiatric disorders [Review]. Curr Opin Psychiatry 21: 290-295.
http://dx.doi.org/10.1097/YCO.Ob013e3282fad375
Kerkvliet. NI; Baecher-Steppan. L. (1982). Immunotoxicology studies on lead: Effects of exposure on
tumor growth and cell-mediated immunity after syngeneic or allogeneic stimulation.
Immunopharmacology 4: 213-224. http://dx.doi.org/10.1016/0162-3109(82)90003-0
Kermani. S: Karbalaie. K; Madani. SH; Jahangirnejad. AA; Eslaminejad. MB; Nasr-Esfahani. MH;
Baharvand. H. (2008). Effect of lead on proliferation and neural differentiation of mouse bone
marrow-mesenchymal stem cells. Toxicol In Vitro 22: 995-1001.
http://dx.doi.0rg/10.1016/i.tiv.2008.02.009
Kern. M; Audesirk. G. (2000). Stimulatory and inhibitory effects of inorganic lead on calcineurin.
Toxicology 150: 171-178. httrj://dx.doi.org/10.1016/S0300-483X(00)00258-4
Kern. M; Wisniewski. M; Cabell. L; Audesirk. G. (2000). Inorganic lead and calcium interact positively
in activation of calmodulin. Neurotoxicology 21: 353-363.
Khairullina. AY; Olshanskava. TV; Filimonenko. DS: Yasinski. VM; Slobozhanina. El; Kozlova. NM.
(2008). Study of optical and nanostructural metal-induced changes in erythrocyte membranes by
scattering and atomic force microscopy. Optics and Spectroscopy 105: 154-160.
http://dx.doi.org/10.1134/S0030400X08070242
November 2012 5-746 Draft - Do Not Cite or Quote
-------
Khalil-Manesh. F; Gonick. HC: Cohen. A; Bergamaschi. E; Mutti. A. (1992a). Experimental model of
lead nephropathy: II. Effect of removal from lead exposure and chelation treatment with
dimercaptosuccinic acid (DMSA). Environ Res 58: 35-54.
http://dx.doi.org/10.1016/80013-9351(05)80203-8
Khalil-Manesh. F; Gonick. HC: Cohen. AH. (1993a). Experimental model of lead nephropathy III
Continuous low-level lead administration. Arch Environ Occup Health 48: 271-278.
httrj://dx.doi.org/10.1080/00039896.1993.9940372
Khalil-Manesh. F; Gonick. HC: Cohen. AH; Alinovi. R; Bergamaschi. E; Mutti. A; Rosen. VJ. (1992b).
Experimental model of lead nephropathy. I. Continuous high-dose lead administration. Kidney Int 41:
1192-1203. http://dx.doi.org/10.1038/ki.1992.181
Khalil-Manesh. F; Gonick. HC: Weiler. EWJ: Prins. B; Weber. MA; Purdv. R; Ren. Q. (1994). Effect of
chelation treatment with dimercaptosuccinic acid (DMSA) on lead-related blood pressure changes.
Environ Res 65: 86-99. http://dx.doi.org/10.1006/enrs.1994.1023
Khalil-Manesh. F: Gonick. HC: Weiler. EWJ: Prins. B: Weber. MA: Purdv. RE. (1993b). Lead-induced
hypertension: Possible role of endothelial factors. Am J Hypertens 6: 723-729.
Khalil. N. (2010). Re: Blood lead mortality study: Khalil et al., 2009. Khalil,
N. http://www.regulations.gov/#!documentDetail:D=EPA-HQ-ORD-2011-0051-0006
Khalil. N: Caulev. JA: Wilson. JW: Talbott. EO: Morrow. L: Hochberg. MC: Hillier. TA: Muldoon. SB:
Cummings. SR. (2008). Relationship of blood lead levels to incident nonspine fractures and falls in
older women: The Study of Osteoporotic Fractures. J Bone Miner Res 23: 1417-1425.
http://dx.doi.org/10.1359/ibmr.080404
Khalil. N; Morrow. LA; Needleman. H; Talbott. EO: Wilson. JW: Caulev. JA. (2009a). Association of
cumulative lead and neurocognitive function in an occupational cohort. Neuropsychology 23: 10-19.
http://dx.doi.org/10.1037/a0013757
Khalil. N; Wilson. JW: Talbott. EO: Morrow. LA; Hochberg. MC: Hillier. TA; Muldoon. SB; Cummings.
SR: Caulev. JA. (2009b). Association of blood lead concentrations with mortality in older women: a
prospective cohort study. Environ Health Global Access Sci Source 8: 15.
http://dx.doi.org/10.1186/1476-069x-8-15
Khan. D; Qawum. S: Saleem. S: Ansari. W: Khan. F. (2010a). Lead exposure and its adverse health
effects among occupational workers children. Toxicol Ind Health 26: 497.
http://dx.doi.org/10.1177/0748233710373085
Khan. DA; Qawum. S: Saleem. S: Khan. FA. (2008). Lead-induced oxidative stress adversely affects
health of the occupational workers. Toxicol Ind Health 24: 611-618.
http://dx.doi.org/10.1177/0748233708098127
Khan. MI; Ahmad. I; Mahdi. AA; Akhtar. MJ; Islam. N; Ashquin. M; Venkatesh. T. (201 Ob). Elevated
blood lead levels and cytogenetic markers in buccal epithelial cells of painters in India: Genotoxicity
in painters exposed to lead containing paints. Environ Sci Pollut Res Int 17: 1347-1354.
http://dx.doi.org/10.1007/sll356-010-0319-x
Khan. MI; Islam. N; Sahasrabuddhe. AA; Mahdi. AA; Siddiqui. H; Ashquin. M; Ahmad. I. (2011).
Ubiquitous hazardous metal lead induces TNF-a in human phagocytic THP-1 cells: Primary role of
ERK 1/2. J Hazard Mater 189: 255-264. http://dx.doi.Org/10.1016/i.ihazmat.2011.02.027
Kharoubi. O: Slimani. M; Aoues. A; Seddik. L. (2008a). Prophylactic effects of Wormwood on lipid
peroxidation in an animal model of lead intoxication. Indian J Nephrol 18: 51-57.
http://dx.doi.org/10.4103/0971-4065.42333
Kharoubi. O: Slimani. M; Krouf. D; Seddik. L; Aoues. A. (2008b). Role of wormwood (Artemisia
absinthium) extract on oxidative stress in ameliorating lead induced haematotoxicity. African Journal
of Traditional, Complementary and Alternative Medicines 5: 263-270.
November 2012 5-747 Draft - Do Not Cite or Quote
-------
Khorasani. G: Shokrzade. M; Salehifar. E; Asadi. M; Shabankhani. B; Rezaeinejad. S. (2008). The
comparison of lead and zinc plasma levels in gastric cancer patients with healthy volunteers. Res J
Biol Sci 3: 631-634. http://dx.doi.org/10.3923/rjbsci.2008.631.634
Khotimchenko. MY; Kolenchenko. EA. (2007). Efficiency of low-esterified pectin in toxic damage to the
liver inflicted by lead treatment. Bull Exp Biol Med 144: 60-62.
httrj://dx.doi.org/10.1007/sl0517-007-0254-0
Kim. D; Lawrence. DA. (2000). Immunotoxic effects of inorganic lead on host resistance of mice with
different circling behavior preferences. Brain Behav Immun 14: 305-317.
http://dx.doi.org/10.1006/brbi.2000.0609
Kim. HS: Lee. SS: Lee. GS: Hwangbo. Y; Ahn. KD; Lee. BK. (2004). The protective effect of delta-
aminolevulinic acid dehydratase 1-2 and 2-2 isozymes against blood lead with higher hematologic
parameters. Environ Health Perspect 112: 538-541. http://dx.doi.org/10.1289/ehp.6464
Kim. JH: Lee. KH: Yoo. DH: Kang. D: Cho. SH: Hong. YC. (2007). GSTM1 and TNF-alpha gene
polymorphisms and relations between blood lead and inflammatory markers in a non-occupational
population. Mutat Res Genet Toxicol Environ Mutagen 629: 32-39.
http://dx.doi.0rg/10.1016/i.mrgentox.2007.01.004
Kim. R; Landrigan. C: Mossmann. P; Sparrow. D: Hu. H. (1997). Age and secular trends in bone lead
levels in middle-aged and elderly men: Three-year longitudinal follow-up in the Normative Aging
Study. Am J Epidemiol 146: 586-591.
Kim. R; Rotnitsky. A; Sparrow. D: Weiss. ST; Wager. C: Hu. H. (1996). A longitudinal study of low-level
lead exposure and impairment of renal function The Normative Aging Study. JAMA 275: 1177-1181.
http://dx.doi.org/10.1001/iama.1996.03530390043032
Kim. Y: Kim. BN: Hong. YC: Shin. MS: Yoo. HJ: Kim. JW: Bhang. SY: Cho. SC. (2009b). Co-exposure
to environmental lead and manganese affects the intelligence of school-aged children.
Neurotoxicology 30: 564-571. http://dx.doi.Org/10.1016/i.neuro.2009.03.012
Kimber. I; Stonard. MD; Gidlow. DA; Niewola. Z. (1986). Influence of chronic low-level exposure to
lead on plasma immunoglobulin concentration and cellular immune function in man. Int Arch Occup
Environ Health 57: 117-125. http://dx.doi.org/10.1007/BF00381379
Kiraly. E; Jones. DG. (1982). Dendritic spine changes in rat hippocampal pyramidal cells after postnatal
lead treatment: Agolgi study. Exp Neurol 77: 236-239.
http://dx.doi.org/10.1016/0014-4886(82)90158-3
Kiran Kumar. B; Prabhakara Rao. Y; Noble. T; Weddington. K; McDowell. VP; Rajanna. S: Bettaiya. R.
(2009). Lead-induced alteration of apoptotic proteins in different regions of adult rat brain. Toxicol
Lett 184: 56-60. http://dx.doi.Org/10.1016/i.toxlet.2008.10.023
Kirberger. M; Yang. JJ. (2008). Structural differences between Pb2+- and Ca2+-binding sites in proteins:
Implications with respect to toxicity. J Inorg Biochem 102: 1901-1909.
http://dx.doi.0rg/10.1016/i.jinorgbio.2008.06.014
Kishi. R; Ikeda. T; Miyake. H; Uchino. E; Tsuzuki. T; Inoue. K. (1983). Effects of low lead exposure on
neuro-behavioral function in the rat. Arch Environ Occup Health 38: 25-33.
Kishikawa. H; Song. R; Lawrence. DA. (1997). Interleukin-12 promotes enhanced resistance to Listeria
monocytogenes infection of lead-exposed mice. Toxicol Appl Pharmacol 147: 180-189.
http://dx.doi.org/10.1006/taap.1997.8308
Kishimoto. T; Oguri. T; Ueda. D: Tada. M. (1995). Effect of lead on tube formation by cultured human
vascular endothelial cells. Arch Toxicol 69: 718-721. http://dx.doi.org/10.1007/s002040050238
Kiziler. AR; Aydemir. B; Onaran. I; Alici. B; Ozkara. H; Gulyasar. T; Akyolcu. MC. (2007). High levels
of cadmium and lead in seminal fluid and blood of smoking men are associated with high oxidative
stress and damage in infertile subjects. Biol Trace Elem Res 120: 82-91.
http://dx.doi.org/10.1007/sl2011-007-8020-8
November 2012 5-748 Draft - Do Not Cite or Quote
-------
Klann. E; Shelton. KR. (1989). The effect of lead on the metabolism of a nuclear matrix protein which
becomes prominent in lead-induced intranuclear inclusion bodies. J Biol Chem 264: 16969-16972.
Klein. D; Y-JY. W: Kamyab. S: Okuda. H; Sokol. RZ. (1994). Effects of toxic levels of lead on gene
regulation in the male axis: increase in messenger ribonucleic acids and intracellular stores of
gonadotrophs within the central nervous system. Biol Reprod 50: 802-811. http://dx.doi.org/10.1095/
biolreprod50.4.802
Knowles. SO: Donaldson. WE. (1990). Dietary modification of lead toxicity: Effects on fatty acid and
eicosanoid metabolism in chicks. Comp Biochem Physiol C Toxicol Pharmacol 95: 99-104.
httrj://dx.doi.org/10.1016/0742-8413(90)90088-O
Knowles. SO: Donaldson. WE. (1997). Lead disrupts eicosanoid metabolism, macrophage function, and
disease resistance in birds. Biol Trace Elem Res 60: 13-26. http://dx.doi.org/10.1007/BF02783306
Kojima. M; Degawa. M. (2006). Gender-related difference in altered gene expression of a sterol
regulatory element binding protein, SREBP-2, by lead nitrate in rats: Correlation with development of
hypercholesterolemia. J Appl Toxicol 26: 381-384. http://dx.doi.org/10.1002/jat.1138
Kojima. M; Sekikawa. K; Nemoto. K; Degawa. M. (2005). Tumor necrosis factor-alpha-independent
downregulation of hepatic cholesterol 7 alpha-hydroxylase gene in mice treated with lead nitrate.
Toxicol Sci 87: 537-542. http://dx.doi.org/10.1093/toxsci/kfi267
Koka. S: Huber. SM; Boini. KM; Lang. C: Foller. M; Lang. F. (2007). Lead decreases parasitemia and
enhances survival of Plasmodium berghei-infected mice. Biochem Biophys Res Commun 363:
484-489. http://dx.doi.0rg/10.1016/i.bbrc.2007.08.173
Kolesarova. A; Roychoudhury. S: Slivkova. J: Sirotkin. A; Capcarova. M; Massanyi. P. (2010). In vitro
study on the effects of lead and mercury on porcine ovarian granulosa cells. J Environ Sci Health A
Tox Hazard Subst Environ Eng 45: 320-331. http://dx.doi.org/10.1080/10934520903467907
Koller. LD. (1973). Immunosuppression produced by lead, cadmium, and mercury. Am J Vet Res 34:
1457-1458.
Koller. LD; Kerkvliet. NI; Exon. JH. (1985). Neoplasia induced in male rats fed lead acetate, ethyl urea,
and sodium nitrite. Toxicol Pathol 13: 50-57. http://dx.doi.org/10.1177/019262338501300107
Koller. LD; Kovacic. S. (1974). Decreased antibody formation in mice exposed to lead. Nature 250:
148-150. http://dx.doi.org/10.1038/250148aO
Korashy. HM; Ei-Kadi. AOS. (2008). The role of redox-sensitive transcription factors NF-kappa B and
AP-1 in the modulation of the Cyplal gene by mercury, lead, and copper. Free Radic Biol Med 44:
795-806. http://dx.doi.0rg/10.1016/i.freeradbiomed.2007.ll.003
Korashy. HM; El-Kadi. AOS. (2004). Differential effects of mercury, lead and copper on the constitutive
and inducible expression of aryl hydrocarbon receptor (AHR)-regulated genes in cultured hepatoma
Hepa Iclc7 cells. Toxicology 201: 153-172. http://dx.doi.Org/10.1016/i.tox.2004.04.011
Korashy. HM; El-Kadi. AOS. (2008). NF-kappa B and AP-1 are key signaling pathways in the
modulation of NAD(P)H: Quinone oxidoreductase 1 gene by mercury, lead, and copper. J Biochem
Mol Toxicol 22: 274-283. http://dx.doi.org/10.1002/ibt.20238
Kordas. K; Canfield. RL; Lopez. P; Rosado. JL; Vargas. GG: Cebrian. ME; Rico. JA; Ronquillo. D;
Stoltzfus. RJ. (2006). Deficits in cognitive function and achievement in Mexican first-graders with
low blood lead concentrations. Environ Res 100: 371-386.
http://dx.doi.0rg/10.1016/i.envres.2005.07.007
Kordas. K; Casavantes. KA; Mendoza. C: Lopez. P; Ronquillo. D; Rosado. JL; Vargas. GG: Stoltzfus. RJ.
(2007). The association between lead and micronutrient status, and children's sleep, classroom
behavior, and activity. Arch Environ Occup Health 62: 105-112.
http://dx.doi.Org/10.3200/AEOH.62.2.105-112
November 2012 5-749 Draft - Do Not Cite or Quote
-------
Kordas. K; Ettinger. AS: Bellinger. DC: Schnaas. L; Tellez Rojo. MM; Hernandez-Avila. M; Hu. H;
Wright. RO. (2011). A dopamine receptor (DRD2) but not dopamine transporter (DAT1) gene
polymorphism is associated with neurocognitive development of Mexican preschool children with
lead exposure. J Pediatr 159: 638-643. http://dx.doi.Org/10.1016/j.jpeds.2011.03.043
Kordas. K; Ettinger. AS: Lamadrid-Figueroa. H; Tellez-Rojo. MM; Hernandez-Avila. M; Hu. H; Wright.
RO. (2009). Methylenetetrahydrofolate reductase (MTHFR) C677T, A1298C and Gl 793 A genotypes,
and the relationship between maternal folate intake, tibia lead and infant size at birth. Br J Nutr 102:
907-914. http://dx.doi.org/10.1017/s0007114509318280
Kottgen. A; Selvin. E; Stevens. LA; Levey. AS: Van Lente. F; Coresh. J. (2008). Serum cystatin C in the
United States: The Third National Health and Nutrition Examination Survey (NHANES III). Am J
Kidney Dis 51: 385-394. http://dx.doi.Org/10.1053/i.ajkd.2007.ll.019
Kowolenko. M; Tracy. L; Lawrence. D. (1991). Early effects of lead on bone marrow cell responsiveness
in mice challenged with Listeria monocytogenes. Fundam Appl Toxicol 17: 75-82.
http://dx.doi.org/10.1093/toxsci/17.L75
Kowolenko. M; Tracy. L; Mudzinski. S: Lawrence. DA. (1988). Effect of lead on macrophage function. J
LeukocBiol 43: 357-364.
Krieg. EF. Jr. (2007). The relationships between blood lead levels and serum follicle stimulating hormone
and luteinizing hormone in the third national health and nutrition examination survey. Environ Res
104: 374-382. http://dx.doi.Org/10.1016/i.envres.2006.09.009
Krieg. EF. Jr; Butler. MA. (2009). Blood lead, serum homocysteine, and neurobehavioral test
performance in the third National Health and Nutrition Examination Survey. Neurotoxicology 30:
281-289. http://dx.doi.0rg/10.1016/i.neuro.2008.12.014
Krieg. EF. Jr; Butler. MA; Chang. MH; Liu. TB; Yesupriya. A; Lindegren. ML; Dowling. N. (2009). Lead
and cognitive function in ALAD genotypes in the Third National Health and Nutrition Examination
Survey. Neurotoxicol Teratol 31: 364-371. http://dx.doi.Org/10.1016/i.ntt.2009.08.003
Krieg. EF. Jr; Butler. MA; M-h. C: Liu. T; Yesupriya. A; Dowling. N; Lindegren. ML. (2010). Lead and
cognitive function in VDR genotypes in the Third National Health and Nutrition Examination Survey.
Neurotoxicol Teratol 32: 262-272. http://dx.doi.Org/10.1016/i.ntt.2009.12.004
Krieg. EF. Jr: Chrislip. DW: Crespo. CJ: Brightwell. WS: Ehrenberg. RL: Otto. DA. (2005). The
relationship between blood lead levels and neurobehavioral test performance in NHANES III and
related occupational studies. Public Health Rep 120: 240-251.
Krishnamoorthv. MS: Parthiban. N; Muthu. P; Paul. V: Balagopal. G: Kumaravel. TS. (1993). Effect of
acute pretreatment of lead on picrotoxin-induced convulsions in rats. J Appl Toxicol 13: 155-159.
Kristal-Boneh. E; Coller. D; Froom. P; Harari. G: Ribak. J. (1999). The association between occupational
lead exposure and serum cholesterol and lipoprotein levels. Am J Public Health 89: 1083-1087.
Krocova. Z; Macela. A; Kroca. M; Hernychova. L. (2000). The immunomodulatory effect(s) of lead and
cadmium on the cells of immune system in vitro. Toxicol In Vitro 14: 33-40.
Kuhlmann. AC: McGlothan. JL; Guilarte. TR. (1997). Developmental lead exposure causes spatial
learning deficits in adult rats. Neurosci Lett 233: 101-104.
http://dx.doi.org/10.1016/S0304-3940(97)00633-2
Kuo. CY; Wong. RH; Lin. JY; Lai. JC: Lee. H. (2006). Accumulation of chromium and nickel metals in
lung tumors from lung cancer patients in Taiwan. J Toxicol Environ Health A 69: 1337-1344.
http://dx.doi.org/10.1080/15287390500360398
Kuruvilla. A; Pillav. VV: Adhikari. P; Venkatesh. T; Chakrapani. M; Rao. HTJ; Bastia. BK; Raieev. A;
Saralava. KM; Rai. M. (2006). Clinical manifestations of lead workers of Mangalore, India. Toxicol
Ind Health 22: 405-413. http://dx.doi.org/10.1177/0748233706074174
November 2012 5-750 Draft - Do Not Cite or Quote
-------
Laden. F; Neas. LM; Dockery. DW: Schwartz. J. (2000). Association of fine paniculate matter from
different sources with daily mortality in six US cities. Environ Health Perspect 108: 941-947.
http://dx.doi.org/10.1289/ehp.00108941
Lai. CC: Lin. HH; Chen. CW: Chen. SH; Chiu. TH. (2002). Excitatory action of lead on rat sympathetic
preganglionic neurons in vitro and in vivo. Life Sci 71: 1035-1045.
http://dx.doi.org/10.1016/S0024-3205(02)01789-7
Lai. LH; Chou. SY; Wu. FY; Chen. JJH; Kuo. HW. (2008a). Renal dysfunction and hyperuricemia with
low blood lead levels and ethnicity in community-based study. Sci Total Environ 401: 39-43.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.04.004
Lai. B; Murtfay. RC: Anand. M; Chandra. SV: Kumar. R; Tripathi. O: Srimal. RC. (1991). Cardiotoxicity
and hypertension in rats after oral lead exposure. Drug Chem Toxicol 14: 305-318.
http://dx.doi.org/10.3109/01480549109002192
Lamadrid-Figueroa. H; Tellez-Rojo. MM; Hernandez-Avila. M; Trejo-Valdivia. B; Solano-Gonzalez. M;
Mercado-Garcia. A; Smith. D; Hu. H; Wright. RO. (2007). Association between the plasma/whole
blood lead ratio and history of spontaneous abortion: A nested cross-sectional study. BMC Pregnancy
Childbirth 7: 22. http://dx.doi.org/10.1186/1471-2393-7-22
Lamb. MR; Janevic. T; Liu. X; Cooper. T; Kline. J: Factor-Litvak. P. (2008). Environmental lead
exposure, maternal thyroid function, and childhood growth. Environ Res 106: 195-202.
http://dx.doi.0rg/10.1016/i.envres.2007.09.012
Landrigan. PJ; Suk. WA; Amler. RW. (1999). Chemical wastes, children's health, and the Superfund Basic
Research Program [Review]. Environ Health Perspect 107: 423-427.
Lang. F; Gulbins. E; Lerche. H; Huber. SM; Kempe. PS: Foller. M. (2008). Eryptosis, a window to
systemic disease [Review]. Cell Physiol Biochem 22: 373-380. http://dx.doi.org/10.1159/000185448
Lanphear. BP; Dietrich. K; Auinger. P; Cox. C. (2000). Cognitive deficits associated with blood lead
concentrations <10 microg/dL in US children and adolescents. Public Health Rep 115: 521-529.
Lanphear. BP; Hornung. R; Khoury. J: Yolton. K; Baghurst. P; Bellinger. DC: Canfield. RL; Dietrich.
KN; Bornschein. R; Greene. T; Rothenberg. SJ: Needleman. HL; Schnaas. L; Wasserman. G:
Graziano. J: Roberts. R. (2005). Low-level environmental lead exposure and children's intellectual
function: An international pooled analysis. Environ Health Perspect 113: 894-899.
Lara-Tejero. M; Pamer. EG. (2004). T cell responses to Listeria monocytogenes [Review]. Curr Opin
Microbiol 7: 45-50. http://dx.doi.Org/10.1016/j.mib.2003.12.002
Larsson. A; Malm. J: Grubb. A; Hansson. LO. (2004). Calculation of glomerular filtration rate expressed
in mL/min from plasma cystatin C values in mg/L. Scand J Clin Lab Invest 64: 25-30.
Laschi-Loquerie. A; Decotes. J: Tachon. P; Evreux. JC. (1984). Influence of lead acetate on
hypersensitivity: Experimental study. J Immunopharm 6: 87-93.
http://dx.doi.org/10.3109/08923978409026461
Laslev. SM; Gilbert. ME. (2002). Rat hippocampal glutamate and GABA release exhibit biphasic effects
as a function of chronic lead exposure level. Toxicol Sci 66: 139-147.
http://dx.doi.org/10.1093/toxsci/66.1.139
Laties. AM; Zrenner. E. (2002). Viagra (sildenafil citrate) and ophthalmology [Review]. Prog Retin Eye
Res 21: 485-506. http://dx.doi.org/10.1016/sl350-9462(02)00013-7
Lau. YS: Camoratto. AM; White. LM; Moriartv. CM. (1991). Effect of lead on TRH and GRF binding in
rat anterior pituitary membranes. Toxicology 68: 169-179.
http://dx.doi.org/10.1016/0300-483X(91)90019-W
Laughlin. NK; Luck. ML; Lasky. RE. (2008). Postnatal lead effects on the development of visual spatial
acuity in rhesus monkeys (Macaca mulatta). Dev Psychobiol 50: 608-614.
http://dx.doi.org/10.1002/dev.20315
November 2012 5-751 Draft - Do Not Cite or Quote
-------
Laughlin. NK; Luck. ML; Lasky. RE. (2009). Early lead exposure effects on an auditory threshold task in
the rhesus monkey (Macaca mulatta). Dev Psychobiol 51: 289-300.
http://dx.doi.org/10.1002/dev.20364
Lawrence. DA. (1981a). Heavy metal modulation of lymphocyte activities - II. Lead, an in vitro mediator
of B-cell activation. International Journal of Immunopharmacology 3: 153-161.
http://dx.doi.org/10.1016/0192-0561(81)90006-0
Lawrence. DA. (198 Ib). In vivo and in vitro effects of lead on humoral and cell- mediated immunity.
Infect Immun 31: 136-143.
Lawton. LJ; Donaldson. WE. (1991). Lead-induced tissue fatty acid alterations and lipid peroxidation.
Biol Trace Elem Res 28: 83-97. http://dx.doi.org/10.1007/BF02863075
Lazarus. JH. (2005). Thyroid disorders associated with pregnancy: Etiology, diagnosis, and management
[Review]. Treat Endocrinol 4: 31-41.
Leasure. JL; Giddabasappa. A; Chanev. S: Johnson. JE; Pothakos. K; Lau. YS: Fox. DA. (2008). Low-
level human equivalent gestational lead exposure produces sex-specific motor and coordination
abnormalities and late-onset obesity in year-old mice. Environ Health Perspect 116: 355-361.
http://dx.doi.org/10.1289/ehp.10862
Ledda-Columbano. GM; Coni. O; Curto. M; Giacomini. O; Faa. G: Sarma. DSR; Columbano. A. (1992).
Mitogen-induced liver hyperplasia does not substitute for compensatory regeneration during
promotion of chemical hepatocarcinogenesis. Carcinogenesis 13: 379-383.
http://dx.doi.0rg/10.1093/carcin/13.3.379
Lee. BK; Lee. GS: Stewart. WF; Ahn. KD; Simon. D; Kelsev. KT; Todd. AC: Schwartz. BS. (200la).
Associations of blood pressure and hypertension with lead dose measures and polymorphisms in the
vitamin D receptor and delta-aminolevulinic acid dehydratase genes. Environ Health Perspect 109:
383-389.
Lee. DH; Blomhoff. R; Jacobs. DR. Jr. (2004). Is serum gamma glutamyltransferase a marker of oxidative
stress? [Review]. Free Radic Res 38: 535-539. http://dx.doi.org/10.1080/10715760410001694026
Lee. DH; Lim. JS: Song. K; Boo. Y; Jacobs. DR. (2006a). Graded associations of blood lead and urinary
cadmium concentrations with oxidative-stress-related markers in the US population: Results from the
Third National Health and Nutrition Examination Survey. Environ Health Perspect 114: 350-354.
http://dx.doi.org/10.1289/ehp.8518
Lee. JE; Chen. S: Golemboski. KA; Parsons. PJ: Dietert. RR. (200Ib). Developmental windows of
differential lead-induced immunotoxicity in chickens. Toxicology 156: 161-170.
http://dx.doi.org/10.1016/S0300-483X(00)00350-4
Lee. JE; Naqi. SA; Kao. E; Dietert. RR. (2002). Embryonic exposure to lead: Comparison of immune and
cellular responses in unchallenged and virally stressed chickens. Arch Toxicol 75: 717-724.
http://dx.doi.org/10.1007/s00204-001-0283-9
Lee. JJ: Battles. AH. (1994). Lead toxicity via arachidonate signal transduction to growth responses in the
splenic macrophage. Environ Res 67: 209-219. http://dx.doi.org/10.1006/enrs.1994.1075
Lee. MK; Cho. SY; Kim. DJ; Jang. JY; Shin. KH; Park. SA; Park. EM; Lee. JS: Choi. MS: Kim. MJ.
(2005). Du-zhong (Eucommia ulmoides Oliv.) cortex water extract alters heme biosynthesis and
erythrocyte antioxidant defense system in lead-administered rats. J Med Food 8: 86-92.
http://dx.doi.0rg/10.1089/imf.2005.8.86
Lee. MY; Shin. JH; Han. HS: Chung. JH. (2006b). In vivo effects of lead on erythrocytes following
chronic exposure through drinking water. Arch Pharm Res 29: 1158-1163.
Lee. TH; Tseng. MC: Chen. CJ: Lin. JL. (2009). Association of high body lead store with severe
intracranial carotid atherosclerosis. Neurotoxicology 30: 876-880.
http://dx.doi.0rg/10.1016/i.neuro.2009.07.004
November 2012 5-752 Draft - Do Not Cite or Quote
-------
Legate. ME; Barhoumi. R; Burghardt. RC: Tiffany-Castiglioni. E. (1993). Low-level lead exposure in
cultured astroglia: Identification of cellular targets with vital fluorescent probes. Neurotoxicology 14:
267-272.
Lenroot. RK; Giedd. JN. (2006). Brain development in children and adolescents: insights from anatomical
magnetic resonance imaging [Review]. Neurosci Biobehav Rev 30: 718-729.
http://dx.doi.0rg/10.1016/i.neubiorev.2006.06.001
Leret. ML; Garcia-Uceda. F; Antonio. MT. (2002). Effects of maternal lead administration on
monoaminergic, GABAergic and glutamatergic systems. Brain Res Bull 58: 469-473.
http://dx.doi.org/10.1016/S0361-9230(02)00819-5
Lesenev. AM; Deme. D; Legue. O: Ohavon. R; Chanson. P; Sales. JP; Carvalho. DP; Dupuy. C: Virion.
A. (1999). Biochemical characterization of a Ca2+/NAD(P)H-dependent H2O2 generator in human
thyroid tissue. Biochimie 81: 373-380. http://dx.doi.org/10.1016/S0300-9084(99)80084-4
Levey. AS: Bosc. JP: Lewis. JB; Greene. T; Rogers. N; Roth. D. (1999). A more accurate method to
estimate glomerular filtration rate from serum creatinine: A new prediction equation. Ann Intern Med
130: 461-470.
Levey. AS: Greene. T; Kusek. JW: Beck. G. (2000). A simplified equation to predict glomerular filtration
rate from serum creatinine [Abstract]. J Am Soc Nephrol 11: A0828.
Levey. AS: Stevens. LA; Schmid. CH; Zhang. YP; Castro. AF; Feldman. HI; Kusek. JW: Eggers. P; Van
Lente. F; Greene. T; Coresh. J. (2009). A new equation to estimate glomerular filtration rate. Ann
Intern Med 150: 604-612.
Levin. ED; Bowman. RE; Wegert. S: Vuchetich. J. (1987). Psychopharmacological investigations of a
lead-induced long-term cognitive deficit in monkeys. Psychopharmacology 91: 334-341.
Leviton. A; Bellinger. D; Allred. EN; Rabinowitz. M; Needleman. H; Schoenbaum. S. (1993). Pre- and
postnatal low-level lead exposure and children's dysfunction in school. Environ Res 60: 30-43.
http://dx.doi.org/10.1006/enrs.1993.1003
Lewis. M; Pitts. D. (2004). Inorganic lead exposure in the rat activates striatal cFOS expression at lower
blood levels and inhibits amphetamine-induced cFOS expression at higher blood levels. J Pharmacol
ExpTher310: 815-820. http://dx.doi.org/10.1124/ipet.103.063941
Li Chen. T; Wise. SS: Kraus. S: Shaffiev. F; Levine. KM; Thompson. WD; Romano. T; O'Hara. T; Wise.
JP. Sr. (2009). Particulate hexavalent chromium is cytotoxic and genotoxic to the North Atlantic right
whale (Eubalaena glacialis) lung and skin fibroblasts. Environ Mol Mutagen 50: 387-393.
http://dx.doi.org/10.1002/em.20471
Li. JP; Wang. CY; Tang. YA; Lin. YW: Yang. JL. (2008a). Role of extracellular signal-regulated kinase
(ERK) signaling in nucleotide excision repair and genotoxicity in response to As(III) and Pb(II). Pure
Appl Chem 80: 2735-2750. http://dx.doi.org/10.1351/pac200880122735
Li. N: Yu. ZL: Wang. L: Zheng. YT: Jia. JX: Wang. O: Zhu. MJ: Liu. XH: Xia. X: Li. WJ. (2009c). Early-
life lead exposure affects the activity of TNF-alpha and expression of SNARE complex in
hippocampus of mouse pups. Biol Trace Elem Res 132: 227-238.
http://dx.doi.org/10.1007/sl2011-009-8551-2
Li. N; Yu. ZL: Wang. L; Zheng. YT; Jia. JX; Wang. Q; Zhu. MJ: Liu. XL; Xia. X; Li. WJ. (201 Ob).
Increased tau phosphorylation and beta amyloid in the hipocampus of mouse pups by early life lead
exposure. Acta Biol Hung 61: 123-134. http://dx.doi.Org/10.1556/ABiol.61.2010.2.l
Li. S: Chen. W: Srinivasan. SR: Bond. MG: Tang. R; Urbina. EM; Berenson. GS. (2003). Childhood
cardiovascular risk factors and carotid vascular changes in adulthood: The Bogalusa Heart Study.
JAMA 290: 2271-2276. http://dx.doi.org/10.1001/iama.290.17.2271
Li. S: Zhao. Z; Zhou. X; Liu. S. (2008c). The effect of lead on intracellular Ca2+ in mouse
lymphocytes. Toxicol In Vitro 22: 1815-1819. http://dx.doi.Org/10.1016/i.tiv.2008.08.005
November 2012 5-753 Draft - Do Not Cite or Quote
-------
Li. TY; Zhang. X; Wei. XP; Liu. YF; Qu. P; Liu. YX; Chen. J. (2011). [Impact of antioxidant vitamins
and heavy metal levels at birth on neurodevelopment of children assessed at two years of age].
Zhonghua Er Ke Za Zhi 49: 439-444.
Lidsky. TI; Schneider. JS. (2003). Lead neurotoxicity in children: basic mechanisms and clinical
correlates [Review]. Brain 126: 5-19. http://dx.doi.org/10.1093/brain/awg014
Lidsky. TI; Schneider. JS. (2004). Lead and public health: Review of recent findings, re-evaluation of
clinical risks. J Environ Monit 6: 36N-42N. http://dx.doi.org/10.1039/B403259b
Lilienthal. H; Kohler. K; Turfeld. M; Winneke. G. (1994). Persistent increases in scotopic b-wave
amplitudes after lead exposure in monkeys. Exp Eye Res 59: 203-209.
http://dx.doi.org/10.1006/exer. 1994.1098
Lilienthal. H; Lenaerts. C: Winneke. G: Hennekes. R. (1988). Alteration of the visual evoked potential
and the electroretinogram in lead-treated monkeys. Neurotoxicol Teratol 10: 417-422.
http://dx.doi.org/10.1016/0892-0362(88)90002-5
Lilienthal. H; Winneke. G. (1996). Lead effects on the brain stem auditory evoked potential in monkeys
during and after the treatment phase. Neurotoxicol Teratol 18: 18:17-32.
Lilis. R; Fischbein. A; Eisinger. J: Blumberg. WE; Diamond. S: Anderson. HA; Rom. W: Rice. C:
Sarkozi. L; Kon. S: Selikoff. IJ. (1977). Prevalence of lead disease among secondary lead smelter
workers and biological indicators of lead exposure. Environ Res 14: 255-285.
http://dx.doi.org/10.1016/0013-9351(77)90037-8
Lim. SY: Dohertv. JD: McBride. K: Miller-Ihli. NJ: Carmona. GN: Stark. KD: Salem. N. (2005). Lead
exposure and (n-3) fatty acid deficiency during rat neonatal development affect subsequent spatial task
performance and olfactory discrimination. JNutr 135: 1019-1026.
Lin-Tan. DT; Lin. JL; Wang. LH; Wang. LM; Huang. LM; Liu. L; Huang. JY; Huang. YL. (2007a).
Fasting glucose levels in predicting 1-year all-cause mortality in patients who do not have diabetes
and are on maintenance hemodialysis. J Am Soc Nephrol 18: 2385-2391.
http://dx.doi.org/10.1681/Asn.2006121409
Lin-Tan. DT; Lin. JL; Yen. TH; Chen. KH; Huang. YL. (2007b). Long-term outcome of repeated lead
chelation therapy in progressive non-diabetic chronic kidney diseases. Nephrol Dial Transplant 22:
2924-2931. http://dx.doi.org/10.1093/ndt/gfm342
Lin. CN; Wang. LH; Shen. KH. (2009). Determining urinary trace elements (Cu, Zn, Pb, As, and Se) in
patients with bladder cancer. J Clin Lab Anal 23: 192-195. http://dx.doi.org/10.1002/jcla.20318
Lin. JL: Lin-Tan. DT: Chen. KH; Hsu. CW: Yen. TH; Huang. WH; Huang. YL. (2010). Blood lead levels
association with 18-month all-cause mortality in patients with chronic peritoneal dialysis. Nephrol
Dial Transplant 25: 1627-1633. http://dx.doi.org/10.1093/ndt/gfp663
Lin. JL: Lin-Tan. DT: Hsu. CW: Yen. TH: Chen. KH: Hsu. HH: Ho. TC: Hsu. KH. (2011). Association of
blood lead levels with mortality in patients on maintenance hemodialysis. Am J Med 124: 350-358.
http://dx.doi.0rg/10.1016/i.amjmed.2010.10.022
Lin. JL; Lin-Tan. DT; Hsu. KH; Yu. CC. (2003). Environmental lead exposure and progression of chronic
renal diseases in patients without diabetes. N Engl J Med 348: 277-286.
http://dx.doi.org/10.1097/01.ASN.0000118529.01681
Lin. JL; Lin-Tan. DT; Li. YJ; Chen. KH; Huang. YL. (2006a). Low-level environmental exposure to lead
and progressive chronic kidney diseases. Am J Med 119: 1-9.
http://dx.doi.0rg/10.1016/i.amjmed.2006.01.005
Lin. JL: Lin-Tan. DT: Yen. TH: Hsu. CW: Jena. CC: Chen. KH: Hsu. KH: Huang. YL. (2008). Blood lead
levels, malnutrition, inflammation, and mortality in patients with diabetes treated by long-term
hemodialysis. Am J Kidney Dis 51: 107-115. http://dx.doi.Org/10.1053/i.aikd.2007.10.002
November 2012 5-754 Draft - Do Not Cite or Quote
-------
Lin. JL; Lin-Tan. DT; Yu. CC: Li. YJ; Huang. YY; Li. KL. (2006b). Environmental exposure to lead and
progressive diabetic nephropathy in patients with type II diabetes. Kidney Int 69: 2049-2056.
http://dx.doi.org/10.1038/sj.ki.5001505
Lin. RH; Lee. CH; Chen. WK; Lin-Shiau. SY. (1994). Studies on cytotoxic and genotoxic effects of
cadmium nitrate and lead nitrate in Chinese hamster ovary cells. Environ Mol Mutagen 23: 143-149.
http://dx.doi.org/10.1002/em.2850230212
Lin. TA; Tai-yi. J. (2007). Benchmark dose approach for renal dysfunction in workers exposed to lead.
Environ Toxicol 22: 229-233. http://dx.doi.org/10.1002/tox.20260
Lind. SE; Park. JS: Drexler. JW. (2009). Pyrithione and 8-hydroxyquinolines transport lead across
erythrocyte membranes. Transl Res 154: 153-159. http://dx.doi.Org/10.1016/i.trsl.2009.06.002
Lison. D; Raguzzi. F; Lauwerys. R. (1990). Comparison of the effects of auranofin, heavy metals and
retinoids on protein kinase C in vitro and on a protein kinase C mediated response in macrophages.
Pharmacol Toxicol 67: 239-242. http://dx.doi.Org/10.llll/i.1600-0773.1990.tb00820.x
Little. BB; Spalding. S: Walsh. B; Keves. DC: Wainer. J; Pickens. S: Rovster. M; Villanacci. J: Gratton. T.
(2009). Blood lead levels and growth status among African-American and Hispanic children in Dallas,
Texas-1980 and 2002: Dallas Lead Project II. Ann Hum Biol 36: 331-341.
http://dx.doi.org/10.1080/03014460902806615
Liu. HT; Niu. RY; Wang. JM; He. Y; Wang. JD. (2008). Changes caused by fluoride and lead in energy
metabolic enzyme activities in the reproductive system of male offspring rats. Fluoride 41: 184-191.
Liu. J; Han. D; Li. Y; Zheng. L; Gu. C: Piao. Z; Au. WW: Xu. X; Huo. X. (201 Ob). Lead affects apoptosis
and related gene XIAP and Smac expression in the hippocampus of developing rats. Neurochem Res
35: 473-479. http://dx.doi.org/10.1007/sll064-009-0083-9
Liu. J; Kershaw. WC: Klaassen. CD. (1991). The protective effect of metallothionein on the toxicity of
various metals in rat primary hepatocyte culture. Toxicol Appl Pharmacol 107: 27-34.
http://dx.doi.org/10.1016/0041-008X(91)90327-B
Liu. J: Xu. X: Wu. K: Piao. Z: Huang. J: Quo. Y: Li. W: Zhang. Y: Chen. A: Huo. X. (20lib). Association
between lead exposure from electronic waste recycling and child temperament alterations.
Neurotoxicology 32: 458-464. http://dx.doi.Org/10.1016/i.neuro.2011.03.012
Liu. L; Wong. T; Pozza. M; Lingenhoehl. K; Wang. Y; Sheng. M; Auberson. Y; Wang. Y. (2004). Role of
NMDA receptor subtypes in governing the direction of hippocampal synaptic plasticity. Science 304:
1021-1024. http://dx.doi.org/10.1126/science.1096615
Liu. MY; Hsieh. WC: Yang. BC. (2000). In vitro aberrant gene expression as the indicator of lead-
induced neurotoxicity in U-373MG cells. Toxicology 147: 59-64.
http://dx.doi.org/10.1016/S0300-483X(00)00186-4
Llanos. MN; Ronco. AM. (2009). Fetal growth restriction is related to placenta! levels of cadmium, lead
and arsenic but not with antioxidant activities. Reprod Toxicol 27: 88-92.
http://dx.doi.0rg/10.1016/i.reprotox.2008.ll.057
Lodi. S: Sharma. V: Kansal. L. (2011). The protective effect of Rubia cordifolia against lead nitrate-
induced immune response impairment and kidney oxidative damage. Indian J Pharmacol 43: 441-444.
http://dx.doi.org/10.4103/0253-7613.83118
Lohmann. C: Bonhoeffer. T. (2008). A role for local calcium signaling in rapid synaptic partner selection
by dendritic filopodia. Neuron 59: 253-260. http://dx.doi.Org/10.1016/j.neuron.2008.05.025
Lolin. Y; O'Gorman. P. (1988). An intra-erythrocyctic low molecular weight lead-binding protein in acute
and chronic lead exposure and its possible protective role in lead toxicity. Ann Clin Biochem 25:
688-697.
Long. GJ: Pounds. JG: Rosen. JF (1992). Lead intoxication alters basal and parathyroid hormone-
regulated cellular calcium homeostasis in rat osteosarcoma (ROS 17/28) cells. Calcif Tissue Int 50:
451-458. http://dx.doi.org/10.1007/BF00296777
November 2012 5-755 Draft - Do Not Cite or Quote
-------
Long. GJ: Rosen. JF. (1992). Lead perturbs epidermal growth factor (EOF) modulation of intracellular
calcium metabolism and collagen synthesis in clonal rat osteoblastic (ROS 17/28) cells. Toxicol Appl
Pharmacol 114: 63-70. http://dx.doi.org/10.1016/0041-008X(92)90097-C
Long. GJ: Rosen. JF; Pounds. JG. (1990). Cellular lead toxicity and metabolism in primary and clonal
osteoblastic bone cells. Toxicol Appl Pharmacol 102: 346-361.
Long. GJ: Rosen. JF: Schanne. FAX. (1994). Lead activation of protein kinase C from rat brain:
Determination of free calcium, lead, and zinc by 19F NMR. J Biol Chem 269: 834-837.
Long. Y; Li. Q; Li. J; Cui. Z. (2011). Molecular analysis, developmental function and heavy metal-
induced expression of ABCC5 in zebrafish. Comp Biochem Physiol B Biochem Mol Biol 158: 46-55.
http://dx.doi.0rg/10.1016/i.cbpb.2010.09.005
Lopez-Larrubia. P; Cauli. O. (2011). Alterations of apparent diffusion coefficient (ADC) in the brain of
rats chronically exposed to lead acetate. Toxicology 281: 1-6.
http://dx.doi.0rg/10.1016/i.tox.2010.12.009
Lopez. CM; Pineiro. AE; Nunez. N; Avagnina. AM; Villaamil. EC: Roses. OE. (2000). Thyroid hormone
changes in males exposed to lead in the Buenos Aires area (Argentina). Pharmacol Res 42: 599-602.
http://dx.doi.org/10.1006/phrs.2000.0734
Louis. ED; Applegate. L; Graziano. JH; Parides. M; Slavkovich. V; Bhat. HK. (2005). Interaction
between blood lead concentration and delta-amino-levulinic acid dehydratase gene polymorphisms
increases the odds of essential tremor. Mov Disord 20: 1170-1177.
http://dx.doi.org/10.1002/mds.20565
Louis. ED; Jurewicz. EC: Applegate. L; Factor-Litvak. P; Parides. M; Andrews. L; Slavkovich. V;
Graziano. JH; Carroll. S: Todd. A. (2003). Association between essential tremor and blood lead
concentration. Environ Health Perspect 111: 1707-1711.
Lu. H; Guizzetti. M; Costa. LG. (2002). Inorganic lead activates the mitogen-activated protein kinase
kinase-mitogen-activated protein kinase-p90RSK signaling pathway in human astrocytoma cells via a
protein kinase C-dependent mechanism. J Pharmacol Exp Ther 300: 818-823.
Lundstrom. NG: Englyst. V: Gerhardsson. L; Jin. T; Nordberg. G. (2006). Lung cancer development in
primary smelter workers: A nested case-referent study. J Occup Environ Med 48: 376-380.
http://dx.doi.org/10.1097/01.iom.0000201556.95982.95
Lustberg. M; Silbergeld. E. (2002). Blood lead levels and mortality. Arch Intern Med 162: 2443-2449.
Lutz. PM; Wilson. TJ; Ireland. J: Jones. AL; Gorman. JS: Gale. NL; Johnson. JC: Hewett. JE. (1999).
Elevated immunoglobulin E (IgE) levels in children with exposure to environmental lead. Toxicology
134: 63-78. http://dx.doi.org/10.1016/S0300-483X(99)00036-0
Lynes. MA; Fontenot. AP; Lawrence. DA; Rosenspire. AJ; Pollard. KM. (2006). Gene expression
influences on metal immunomodulation [Review]. Toxicol Appl Pharmacol 210: 9-16.
http://dx.doi.0rg/10.1016/i.taap.2005.04.021
MacDonald. JF: Jackson. MF; Beazely. MA. (2006). Hippocampal long-term synaptic plasticity and
signal amplification of NMDAreceptors [Review]. Crit Rev Neurobiol 18: 71-84.
Machida. M; Sun. SJ: Oguma. E; Kavama. F (2009). High bone matrix turnover predicts blood levels of
lead among perimenopausal women. Environ Res 109: 880-886.
http://dx.doi.0rg/10.1016/i.envres.2009.06.005
Maciejczyk. P; Chen. LC. (2005). Effects of subchronic exposures to concentrated ambient particles
(CAPs) in mice: VIII. Source-related daily variations in in vitro responses to CAPs. Inhal Toxicol 17:
243-253. http://dx.doi.org/10.1080/08958370590912914
Madhavi. D; Devi. KR; Rao. KK; Reddv. PP. (2007). Modulating effect of Phyllanthus fruit extract
against lead genotoxicity in germ cells of mice. J Environ Biol 28: 115-117.
November 2012 5-756 Draft - Do Not Cite or Quote
-------
Maeda. K; Sugino. H; Hirose. T; Kitagawa. H; Nagai. T; Mizoguchi. H; Takuma. K; Yamada. K. (2007).
Clozapine prevents a decrease in neurogenesis in mice repeatedly treated with phencyclidine. J
Pharmacol Sci 103: 299-308. http://dx.doi.org/10.1254/iphs.FP0061424
Magalhaes. C: Socodato. R; Paes-de-Carvalho. R. (2006). Nitric oxide regulates the proliferation of chick
embryo retina cells by a cyclic GMP-independent mechanism. Int J Dev Neurosci 24: 53-60.
http://dx.doi.0rg/10.1016/i.ijdevneu.2005.10.004
Magyar. JS: Weng. TC: Stern. CM; Dye. DF; Rous. BW: Payne. JC: Bridgewater. BM; Mijovilovich. A;
Parkin. G: Zaleski. JM; Penner-Hahn. JE; Godwin. HA. (2005). Reexamination of lead(II)
coordination preferences in sulfur-rich sites: Implications for a critical mechanism of lead poisoning. J
Am Chem Soc 127: 9495-9505. http://dx.doi.org/10.1021/ia0424530
Mahaffev. KR; Capar. SG: Gladen. BC: Fowler. BA. (1981). Concurrent exposure to lead, cadmium, and
arsenic. Effects on toxicity and tissue metal concentrations in the rat. Transl Res 98: 463-481.
Mahaffev. KR; Fowler. BA. (1977). Effects of concurrent administration of lead, cadmium, and arsenic in
the rat. Environ Health Perspect 19: 165-171. http://dx.doi.org/10.1289/ehp.7719165
Mahram. M; Mousavinasab. N; Dinmohammadi. H; Soroush. S: Sarkhosh. F. (2007). Effect of living in
lead mining area on growth. Indian J Pediatr 74: 555-559. http://dx.doi.org/10.1007/sl2098-007-0107-
x
Maitani. T; Watahiki. A; Suzuki. KT. (1986). Induction of metallothionein after lead administration by
three injection routes in mice. Toxicol Appl Pharmacol 83: 211-217.
http://dx.doi.org/10.1016/0041-008X(86)90298-X
Maizlish. NA; Parra. G: Feo. O. (1995). Neurobehavioural evaluation of Venezuelan workers exposed to
inorganic lead. Occup Environ Med 52: 408-414. http://dx.doi.Org/10.1136/oem.52.6.408
Malveaux. FJ; Conroy. MC: Adkinson. NF. Jr; Lichtenstein. LM. (1978). IgE receptors on human
basophils: Relationship to serum IgE concentration. J Clin Invest 62: 176-181.
http://dx.doi.org/10.1172/JCI109103
Malvezzi. CK; Moreira. EG: Vassilieff. I; Vassilieff. VS: Cordellini. S. (2001). Effect of L-arginine,
dimercaptosuccinic acid (DMSA) and the association of L-arginine and DMSA on tissue lead
mobilization and blood pressure level in plumbism. Braz J Med Biol Res 34: 1341-1346.
http://dx.doi.org/10.1590/S0100-879X2001001000016
Marchlewicz. M; Baranowska-Bosiacka. I; Kolasa. A; Kondarewicz. A; Chlubek. D; Wiszniewska. B.
(2009). Disturbances of energetic metabolism in rat epididymal epithelial cells as a consequence of
chronic lead intoxication. Biometals 22: 877-887. http://dx.doi.org/10.1007/sl0534-009-9238-z
Marcus. DK; Fulton. JJ; Clarke. EJ. (2010). Lead and conduct problems: Ameta-analysis. J Clin Child
Adolesc Psychol 39: 234-241. http://dx.doi.org/10.1080/15374411003591455
Marie. C: Hall. JE. (2011). Obesity, metabolic syndrome and diabetic nephropathy [Review]. Contrib
Nephrol 170: 28-35. http://dx.doi.org/10.1159/000324941
Markovac. J: Goldstein. GW. (1988a). Lead activates protein kinase C in immature rat brain microvessels.
Toxicol Appl Pharmacol 96: 14-23. http://dx.doi.org/10.1016/0041-008X(88)90242-6
Markovac. J: Goldstein. GW. (1988b). Picomolar concentrations of lead stimulate brain protein kinase
C. Nature 334: 71-73. http://dx.doi.org/10.1038/334071aO
Marques. CC: Nunes. AC: Pinheiro. T; Lopes. PA; Santos. MC: Viegas-Crespo. AM; Ramalhinho. MG:
Mathias. ML. (2006). An assessment of time-dependent effects of lead exposure in algerian mice (Mus
spretus) using different methodological approaches. Biol Trace Elem Res 109: 75-90.
http://dx.doi.Org/10.1385/BTER:109:l:075
Marques. M; Millas. I; Jimenez. A; Garcia-Colis. E; Rodriguez-Feo. JA; Velasco. S: Barrientos. A;
Casado. S: Lopez-Farre. A. (2001). Alteration of the soluble guanylate cyclase system in the vascular
wall of lead-induced hypertension in rats. J Am Soc Nephrol 12: 2594-2600.
November 2012 5-757 Draft - Do Not Cite or Quote
-------
Martin. D; Glass. TA; Bandeen-Roche. K; Todd. AC: Shi. WP; Schwartz. BS. (2006). Association of
blood lead and tibia lead with blood pressure and hypertension in a community sample of older adults.
Am J Epidemiol 163: 467-478. http://dx.doi.org/10.1093/aje/kwj060
Martinez-Haro. M; Green. AJ; Mateo. R. (2011). Effects of lead exposure on oxidative stress biomarkers
and plasma biochemistry in waterbirds in the field. Environ Res 111: 530-538.
http://dx.doi.0rg/10.1016/i.envres.2011.02.012
Martinez. T; Lartigue. J: Juarez. F; Avila-perez. P; Marquez. C: Zarazua. G: Tejeda. S. (2007).
206Pb/207Pb ratios in dry deposit samples from the Metropolitan Zone of Mexico Valley. Journal of
Radioanal Chem 273: 577-582. http://dx.doi.org/10.1007/sl0967-007-0913-7
Mas-Oliva. J. (1989). Effect of lead on the erythrocyte (Ca2+,Mg2+)ATPase activity: Calmodulin
involvement. Mol Cell Biochem 89: 87-93. http://dx.doi.org/10.1007/BF00228283
Massadeh. AM; Al-Safi. SA; Momani. IF; Al-Mahmoud. M; Alkofahi. AS. (2007). 115. Biol Trace Elem
Res 115: 157-167. http://dx.doi.org/10.1007/BF02686027
Massanyi. P; Lukac. N; Makarevich. AV; Chrenek. P; Forgacs. Z; Zakrzewski. M; Stawarz. R; Toman. R;
Lazor. P; Flesarova. S. (2007). Lead-induced alterations in rat kidneys and testes in vivo. J Environ
Sci Health A Tox Hazard Subst Environ Eng 42: 671-676.
http://dx.doi.org/10.1080/10934520701244474
Masso-Gonzalez. EL; Antonio-Garcia. MT. (2009). Natural antioxidants protect against lead-induced
damage during pregnancy and lactation in rat's pups. Ecotoxicol Environ Saf 72: 2137-2142.
http://dx.doi.0rg/10.1016/i.ecoenv.2009.03.013
Masso. EL; Corredor. L; Antonio. MT. (2007). Oxidative damage in liver after perinatal intoxication with
lead and/or cadmium. J Trace Elem Med Biol 21: 210-216.
http://dx.doi.0rg/10.1016/i.itemb.2007.03.002
Matovic. V: Buha. A; Bulat. Z; Dukic-Cosic. D. (2011). Cadmium toxicity revisited: Focus on oxidative
stress induction and interactions with zinc and magnesium [Review]. Arh Hig Rada Toksikol 62:
65-76. http://dx.doi.org/10.2478/10004-1254-62-2011-2075
Maue'l. J: Ransijn. A; Buchmuller-Rouiller. Y. (1989). Lead inhibits intracellular killing of Leishmania
parasites and extracellular cytolysis of target cells by macrophages exposed to macrophage activating
factor. J Leukoc Biol 45: 401-409.
Mazumdar. M; Bellinger. DC: Gregas. M; Abanilla. K; Bacic. J: Needleman. HL. (2011). Low-level
environmental lead exposure in childhood and adult intellectual function: A follow-up study. Environ
Health Global Access Sci Source 10: 24. http://dx.doi.org/10.1186/1476-069X-10-24
McCabe. MJ. Jr; Lawrence. DA. (1990). The heavy metal lead exhibits B cell-stimulatory factor activity
by enhancing B cell la expression and differentiation. J Immunol 145: 671-677.
McCabe. MJ. Jr; Lawrence. DA. (1991). Lead, a major environmental pollutant, is immunomodulatory by
its differential effects on CD4+T cell subsets. Toxicol Appl Pharmacol 111: 13-23.
http://dx.doi.org/10.1016/0041-008X(91)90129-3
McCabe. MJ. Jr; Singh. KP; Reiners. JJ. Jr. (1999). Lead intoxication impairs the generation of a delayed
type hypersensitivity response. Toxicology 139: 255-264.
http://dx.doi.org/10.1016/S0300-483X(99)00147-X
McElroy. JA; Shafer. MM; Gangnon. RE; Crouch. LA; Newcomb. PA. (2008). Urinary lead exposure and
breast cancer risk in a population-based case-control study. Cancer Epidemiol Biomarkers Prev 17:
2311-2317. http://dx.doi.org/10.1158/1055-9965.epi-08-0263
McLachlin. JR; Gover. RA; Cherian. MG. (1980). Formation of lead-induced inclusion bodies in primary
rat kidney epithelial cell cultures: Effect of actinomycin D and cycloheximide. Toxicol Appl
Pharmacol 56: 418-431. http://dx.doi.org/10.1016/0041-008X(80)90076-9
November 2012 5-758 Draft - Do Not Cite or Quote
-------
McMichael. AJ; Baghurst PA; Vimpani. GV: Robertson. EF; Wigg. NR; Tong. SL. (1992).
Sociodemographic factors modifying the effect of environmental lead on neuropsychological
development in early childhood. Neurotoxicol Teratol 14: 321-327.
http://dx.doi.org/10.1016/0892-0362(92)90038-C
McMichael. AJ: Baghurst. PA; Wigg. NR; Vimpani. GV: Robertson. EF; Roberts. RJ. (1988). Port Pirie
cohort study: Environmental exposure to lead and children's abilities at the age of four years. N Engl J
Med 319: 468-475. http://dx.doi.org/10.1056/NEJM19880825319080
McNeill. PR; Wong. HK; Naravana. A; Wilson. DM. (2007). Lead promotes abasic site accumulation and
co-mutagenesis in mammalian cells by inhibiting the major abasic endonuclease Apel. Mol Carcinog
46: 91-99. http://dx.doi.org/10.1002/mc.20196
Meek. WH; Williams. CL; Cermak. JM; Blusztajn. JK. (2007). Developmental periods of choline
sensitivity provide an ontogenetic mechanism for regulating memory capacity and age-related
dementia. Frontiers in Integrative Neuroscience 1: 7. http://dx.doi.org/10.3389/neuro.07.007.2007
Meeker. JD; Rossano. MG: Protas. B; Diamond. MP; Puscheck. E; Daly. D; Paneth. N; Wirth. JJ. (2008).
Cadmium, lead, and other metals in relation to semen quality: Human evidence for molybdenum as a
male reproductive toxicant. Environ Health Perspect 116: 1473-1479.
http://dx.doi.org/10.1289/ehp.11490
Meeker. JD; Rossano. MG: Protas. B; Padmanahban. V; Diamond. MP; Puscheck. E; Daly. D; Paneth. N;
Wirth. JJ. (2010). Environmental exposure to metals and male reproductive hormones: Circulating
testosterone is inversely associated with blood molybdenum. Fertil Steril 93: 130-140.
http://dx.doi.0rg/10.1016/i.fertnstert.2008.09.044
Mendez-Armenta. M; Nava-Ruiz. C: Fernandez-Valverde. F; Sanchez-Garcia. A; Rios. C. (2011).
Histochemical changes in muscle of rats exposed subchronically to low doses of heavy metals.
Environ Toxicol Pharmacol 32: 107-112. http://dx.doi.Org/10.1016/i.etap.2011.03.001
Mendez-Gomez. J; Garcia-Vargas. GG: Lopez-Carrillo. L; Calderon-Aranda. ES: Gomez. A; Vera. E;
Valverde. M; Cebrian. ME; Rojas. E. (2008). Genotoxic effects of environmental exposure to arsenic
and lead on children in region Lagunera, Mexico. Ann N Y Acad Sci 1140: 358-367.
http://dx.doi.org/10.1196/annals.1454.027
Mendiola. J: Moreno. JM; Roca. M; Vergara-Juarez. N; Martinez-Garcia. MJ; Garcia-Sanchez. A; Elvira-
Rendueles. B; Moreno-Grau. S: Lopez-Espin. JJ: Ten. J: Bernabeu. R; Torres-Cantero. AM. (2011).
Relationships between heavy metal concentrations in three different body fluids and male reproductive
parameters: A pilot study. Environ Health Global Access Sci Source 10: 6.
http://dx.doi.org/10.1186/1476-069X-10-6
Mendv. A; Gasana. J: Vieira. ER. (2012). Urinary heavy metals and associated medical conditions in the
US adult population. Int J Environ Health Res 22: 105-118.
http://dx.doi.org/10.1080/09603123.2011.605877
Menegazzi. M; Carcereri-De Prati. A; Suzuki. H; Shinozuka. H; Pibiri. M; Piga. R; Columbano. A;
Ledda-Columbano. GM. (1997). Liver cell proliferation induced by nafenopin and cyproterone acetate
is not associated with increases in activation of transcription factors NF-kappaB and AP-1 or with
expression of tumor necrosis factor alpha. Hepatology 25: 585-592.
http://dx.doi.org/10.1002/hep.510250316
Meng. XM; Zhu. DM; Ruan. DY; She. JQ: Luo. L. (2005). Effects of chronic lead exposure on H MRS of
hippocampus and frontal lobes in children. Neurology 64: 1644-1647.
http://dx.doi.org/10.1212/01.WNL.0000160391.58004.D4
Menke. A; Muntner. P; Batuman. V; Silbergeld. EK; Guallar. E. (2006). Blood lead below 0.48
micromol/L (10 microg/dL) and mortality among US adults. Circulation 114: 1388-1394.
http://dx.doi.org/10.1161/circulationaha.106.628321
November 2012 5-759 Draft - Do Not Cite or Quote
-------
Mesdaghinia. A; Yazdanpanah. H; Seddighi. M; Banafshe. HR; Hevdari. A. (2010). Effect of short-term
lead exposure on PTZ-induced seizure threshold in mice. Toxicol Lett 199: 6-9.
http://dx.doi.0rg/10.1016/i.toxlet.2010.07.012
Messerschmidt. A; Huber. R; Wieghardt. K; Poulos. T. (2001). Handbook of metalloproteins. In A
Messerschmidt; R Huber; K Wieghardt; T Poulos (Eds.). Chichester, U.K.: Wiley.
http://dx.doi.org/10.1002/0470028637
Mielzvnska. D; Siwinska. E; Kapka. L; Szyfter. K; Knudsen. LE; Merlo. DF. (2006). The influence of
environmental exposure to complex mixtures including PAHs and lead on genotoxic effects in
children living in Upper Silesia, Poland. Mutagenesis 21: 295-304.
http://dx.doi.org/10.1093/mutage/gel037
Miller. TE; Golemboski. KA; Ha. RS; Bunn. T; Sanders. FS; Dietert. RR. (1998). Developmental
exposure to lead causes persistent immunotoxicity in Fischer 344 rats. Toxicol Sci 42: 129-135.
http://dx.doi.org/10.1006/toxs.1998.2424
Min. JY; Min. KB; Cho. SI; Kim. R; Sakong. J; Paek. D. (2007). Neurobehavioral function in children
with low blood lead concentrations. Neurotoxicology 28: 421-425.
http://dx.doi.0rg/10.1016/i.neuro.2006.3.007
Min. JY; Min. KB; Kim. R; Cho. SI; Paek. D. (2008a). Blood lead levels and increased bronchial
responsiveness. Biol Trace Elem Res 123: 41-46. http://dx.doi.org/10.1007/sl2011-008-8099-6
Min. KB; Min. JY; Cho. SI; Kim. R; Kim. H; Paek. D. (2008b). Relationship between low blood lead
levels and growth in children of white-collar civil servants in Korea. Int J Hyg Environ Health 211:
82-87. http://dx.doi.0rg/10.1016/j.ijheh.2007.03.003
Min. MYO; Singer. LT; Kirchner. HL; Minnes. S; Short. E; Hussain. Z; Nelson. S. (2009). Cognitive
development and low-level lead exposure in poly-drug exposed children. Neurotoxicol Teratol 31:
225-231. http://dx.doi.0rg/10.1016/i.ntt.2009.03.002
Minozzo. R; Deimling. LI; Gigante. LP; Santos-Mello. R. (2004). Micronuclei in peripheral blood
lymphocytes of workers exposed to lead. Mutat Res Genet Toxicol Environ Mutagen 565: 53-60.
http://dx.doi.0rg/10.1016/i.mrgentox.2004.09.003
Minozzo. R; Deimling. LI; Santos-Mello. R. (2010). Cytokinesis-blocked micronucleus cytome and
comet assays in peripheral blood lymphocytes of workers exposed to lead considering folate and
vitamin B12 status. Mutat Res Genet Toxicol Environ Mutagen 697: 24-32.
http://dx.doi.0rg/10.1016/i.mrgentox.2010.01.009
Miranda. ML; Edwards. SE; Swamy. GK; Paul. CJ; Neelon. B. (2010). Blood lead levels among pregnant
women: Historical versus contemporaneous exposures. Int J Environ Res Public Health 7: 1508-1519.
http://dx.doi.org/10.3390/iierph7041508
Miranda. ML; Kim. D; Galeano. M; Paul. CJ; Hull. AP; Morgan. SP (2007a). The relationship between
early childhood blood lead levels and performance on end of grade tests. Environ Health Perspect 115:
1242-1247. http://dx.doi.org/10.1289/ehp.9994
Miranda. ML; Kim. D; Reiter. J; Overstreet Galeano. MA; Maxson. P. (2009). Environmental contributors
to the achievement gap. Neurotoxicology 30: 1019-1024.
http://dx.doi.0rg/10.1016/i.neuro.2009.07.012
Mishra. KP. (2009). Lead exposure and its impact on immune system: A review [Review]. Toxicol In
Vitro 23: 969-972. http://dx.doi.Org/10.1016/i.tiv.2009.06.014
Mishra. KP; Chauhan. U.K.; Naik. S. (2006a). Effect of lead exposure on serum immunoglobulins and
reactive nitrogen and oxygen intermediate. Hum Exp Toxicol 25: 661-665.
http://dx.doi.org/10.1177/0960327106070453
Mishra. KP; Rani. R; Yadav. VS; Naik. S. (2010). Effect of lead exposure on lymphocyte subsets and
activation markers. Immunopharmacol Immunotoxicol 32: 446-449.
http://dx.doi.org/10.3109/08923970903503668
November 2012 5-760 Draft - Do Not Cite or Quote
-------
Mishra. KP; Singh. VK; Rani. R; Yadav. VS: Chandran. V; Srivastava. SP; Seth. PK. (2003). Effect of
lead exposure on the immune response of some occupationally exposed individuals. Toxicology 188:
251-259. http://dx.doi.org/10.1016/S0300-483X(03)00091-X
Misonou. H; Morishima-Kawashima. M; Ihara. Y. (2000). Oxidative stress induces intracellular
accumulation of amyloid beta-protein (Abeta) in human neuroblastoma cells. Biochemistry 39:
6951-6959. http://dx.doi.org/10.1021M000169p
Mistry. P; Lucier. GW: Fowler. BA. (1985). High-affinity lead binding proteins in rat kidney cytosol
mediate cell-free nuclear translocation of lead. J Pharmacol Exp Ther 232: 462-469.
Mistry. P; Mastri. C: Fowler. BA. (1986). Influence of metal ions on renal cytosolic lead-binding proteins
and nuclear uptake of lead in the kidney. Biochem Pharmacol 35: 711-713.
http://dx.doi.org/10.1016/0006-2952(86)90371-0
Moffitt. TE; Caspi. A; Rutter. M; Silva. PA. (2001). Sex differences in antisocial behavior: Conduct
disorder, delinquency, and violence in the Dunedin Longitudinal Study. Cambridge, U.K.: University
of Cambridge Press.
Mohammad. A; Ali. N; Reza. B; Ali. K. (2010). Effect of ascorbic acid supplementation on nitric oxide
metabolites and systolic blood pressure in rats exposed to lead. Indian J Pharmacol 42: 78-81.
http://dx.doi.org/10.4103/0253-7613.64501
Mohammad. IK; Mahdi. AA; Raviraja. A; Naimul. I; Iqbal. A; Thuppil. V. (2008). Oxidative stress in
painters exposed to low lead levels. Arh Hig Rada Toksikol 59: 161-169.
http://dx.doi.org/10.2478/10004-1254-59-2008-1883
Molero. L; Carrasco. C: Marques. M; Vaziri. ND; Mateos-Caceres. PJ: Casado. S: Macava. C: Barrientos.
A; Lopez-Farre. AJ. (2006). Involvement of endothelium and endothelin-1 in lead-induced smooth
muscle cell dysfunction in rats. Kidney Int 69: 685-690. http://dx.doi.org/10.1038/sj.ki.5000103
Molina. RM; Phattanarudee. S: Kim. J: Thompson. K; Wessling-Resnick. M; Maher. TJ; Brain. JD.
(2011). Ingestion of Mn and Pb by rats during and after pregnancy alters iron metabolism and
behavior in offspring. Neurotoxicology 32: 413-422. http://dx.doi.Org/10.1016/i.neuro.2011.03.010
Monteiro. HP; Abdalla. DSP: Augusto. O: Bechara. EJH. (1989). Free radical generation during delta-
aminolevulinic acid autoxidation: Induction by hemoglobin and connections with porphyrinpathies.
Arch Biochem Biophys 271: 206-216. http://dx.doi.org/10.1016/0003-9861(89)90271-3
Monteiro. HP; Abdalla. DSP: Faljoni-Alario. A; Bechara. EJH. (1986). Generation of active oxygen
species during coupled autoxidation of oxyhemoglobin and delta-aminolevulinic acid. Biochim
Biophys Acta 881: 100-106. http://dx.doi.org/10.1016/0304-4165(86)90102-9
Monteiro. HP; Bechara. EJH: Abdalla. DSP. (1991). Free radicals involvement in neurological porphyrias
and lead poisoning. Mol Cell Biochem 103: 73-83. http://dx.doi.org/10.1007/BF00229595
Montenegro. MF; Barbosa. F. Jr; Sandrim. VC: Gerlach. RF; Tanus-Santos. JE. (2006). A polymorphism
in the delta-aminolevulinic acid dehydratase gene modifies plasma/whole blood lead ratio. Arch
Toxicol 80: 394-398. http://dx.doi.org/10.1007/s00204-005-0056-v
Moore. CF; Gaiewski. LL; Laughlin. NK; Luck. ML; Larson. JA; Schneider. ML. (2008). Developmental
lead exposure induces tactile defensiveness in rhesus monkeys (Macaca mulatta). Environ Health
Perspect 116: 1322-1326. http://dx.doi.org/10.1289/ehp.11203
Moore. JF; Gover. RA. (1974). Lead-induced inclusion bodies: Composition and probable role in lead
metabolism. Environ Health Perspect 7: 121-127. http://dx.doi.org/10.1289/ehp.747121
Moore. JF; Gover. RA; Wilson. M. (1973). Lead-induced inclusion bodies: Solubility, amino acid content,
and relationship to residual acidic nuclear proteins. Lab Invest 29: 488-494.
Moorhouse. SR; Garden. S: Drewitt PN; Elev. BP; Hargreaves. PJ: Felling. D. (1988). The effect of
chronic low level lead exposure on blood-brain barrier function in the developing rat. Biochem
Pharmacol 37: 4539-4547. http://dx.doi.org/10.1016/0006-2952(88)90670-3
November 2012 5-761 Draft - Do Not Cite or Quote
-------
Moorman. WJ: Skaggs. SR; Clark. JC: Turner. TW: Sharpnack. DP; Murrell. JA; Simon. SD; Chapin.
RE; Schrader. SM. (1998). Male reproductive effects of lead, including species extrapolation for the
rabbit model. Reprod Toxicol 12: 333-346. http://dx.doi.org/10.1016/S0890-6238(98)00010-0
Morita. Y; Sakai. T; Araki. S: Araki. T; Masuvama. Y. (1997). Nicotinamide adenine dinucleotide
synthetase activity in erythrocytes as a tool for the biological monitoring of lead exposure. Int Arch
Occup Environ Health 70: 195-198. http://dx.doi.org/10.1007/s004200050206
Morris. S: van Aardt. WJ: Ahern. MD. (2005). The effect of lead on the metabolic and energetic status of
the Yabby, Cherax destructor, during environmental hypoxia. Aquat Toxicol 75: 16-31.
http://dx.doi.0rg/10.1016/i.aquatox.2005.07.001
Mosad. SM; Ghanem. AA; El-Fallal. HM; El-Kannishy. AM; El Baiomy. AA; Al-Diastv. AM; Arafa. LF.
(2010). Lens cadmium, lead, and serum vitamins C, E, and beta carotene in cataractous smoking
patients. Curr Eye Res 35: 23-30. http://dx.doi.org/10.3109/02713680903362880
Moser. R; Oberlev. TD; Daggett. DA; Friedman. AL; Johnson. JA; Siegel. FL. (1995). Effects of lead
administration on developing rat kidney: I Glutathione S-transferase isoenzymes. Toxicol Appl
Pharmacol 131: 85-93. http://dx.doi.org/10.1006/taap.1995.1050
Moss. ME; Lanphear. BP; Auinger. R (1999). Association of dental caries and blood lead levels. JAMA
281: 2294-2298. http://dx.doi.org/10.1001/iama.281.24.2294
Mudipalli. A. (2007). Lead hepatotoxicity & potential health effects [Review]. Indian J Med Res 126:
518-527.
Mudzinski. SP; Rudofsky. UH; Mitchell. DG: Lawrence. DA. (1986). Analysis of lead effects on in vivo
antibody-mediated immunity in several mouse strains. Toxicol Appl Pharmacol 83: 321-330.
http://dx.doi.org/10.1016/0041-008X(86)90309-l
Muldoon. SB; Caulev. JA: Kuller. LH; Morrow. L; Needleman. HL; Scott. J: Hooper. FJ. (1996). Effects
of blood lead levels on cognitive function of older women. Neuroepidemiology 15: 62-72.
http://dx.doi.org/10.1159/000109891
Miiller. S: Gillert. KE; Krause. C: Gross. U: L'Age-Stehr. J: Diamantstein. T. (1977). Suppression of
delayed type hypersensitivity of mice by lead. Experientia 33: 667-668.
http://dx.doi.org/10.1007/BF01946564
Miiller. YM; Kobus. K; Schatz. JC: Ammar. D; Nazari. EM. (2012). Prenatal lead acetate exposure
induces apoptosis and changes GFAP expression during spinal cord development. Ecotoxicol Environ
Saf 75: 223229. http://dx.doi.Org/10.1016/i.ecoenv.2011.08.004
Muntner. P; He. J: Vupputuri. S: Coresh. J: Batuman. V. (2003). Blood lead and chronic kidney disease in
the general United States population: Results from NHANES III. Kidney Int 63: 1044-1050.
http://dx.doi.0rg/10.1046/i.1523-1755.2003.00812.x
Muntner. P; Menke. A; Batuman. V; Rabito. FA; He. J: Todd. AC. (2007). Association of tibia lead and
blood lead with end-stage renal disease: A pilot study of African-Americans. Environ Res 104:
396-401. http://dx.doi.0rg/10.1016/i.envres.2007.04.001
Muntner. P; Menke. A; DeSalvo. KB; Rabito. FA; Batuman. V. (2005). Continued decline in blood lead
levels among adults in the United States - The National Health and Nutrition Examination Surveys.
Arch Intern Med 165: 2155-2161. http://dx.doi.org/10.1001/archinte.165.18.2155
Murakami. K; Feng. G: Chen. SG. (1993). Inhibition of brain protein kinase C subtypes by lead. J
Pharmacol Exp Ther 264: 757-761.
Murata. K; Iwata. T; Dakeishi. M; Karita. K. (2009). Lead toxicity: Does the critical level of lead
resulting in adverse effects differ between adults and children? [Review]. J Occup Health 51: 1-12.
http://dx.doi.org/10.1539/ioh.K8003
Murthy. RC: Gupta. SK; Saxena. DK. (1995). Nuclear alterations during acrosomal cap formation in
spermatids of lead-treated rats. Reprod Toxicol 9: 483-489.
November 2012 5-762 Draft - Do Not Cite or Quote
-------
Muwakkit. S: Nuwayhid. I; Nabulsi. M; al Haij. R; Khoury. R; Mikati. M; Abboud. MR. (2008). Iron
deficiency in young Lebanese children: Association with elevated blood lead levels. J Pediatr Hematol
Oncol 30: 382-386. http://dx.doi.org/10.1097/MPH.Ob013e318165b283
Naha. N; Chowdhury. AR. (2006). Inorganic lead exposure in battery and paint factory: Effect on human
sperm structure and functional activity. J UOEH 28: 157-171.
Naha. N; Manna. B. (2007). Mechanism of lead induced effects on human spermatozoa after occupational
exposure. Kathmandu Univ Med J 5: 85-94.
Naicker. N; Norris. SA; Mathee. A; Becker. P; Richter. L. (2010). Lead exposure is associated with a
delay in the onset of puberty in South African adolescent females: Findings from the birth to twenty
cohort. Sci Total Environ 408: 4949-4954. http://dx.doi.Org/10.1016/i.scitotenv.2010.07.037
Nakagawa. K. (1989). Hepatic glutathione metabolism in mice acutely treated with lead acetate. Jpn J
Pharmacol 51: 173-179. http://dx.doi.org/10.1254/iip.51.173
Nakagawa. K. (1991). Decreased glutathione S-transferase activity in mice livers by acute treatment with
lead, independent of alteration in glutathione content. Toxicol Lett 56: 13-17.
http://dx.doi.org/10.1016/0378-4274(91)90085-K
Nakaiima. T; Deguchi. T; Kagawa. K; Hikita. H; Ueda. K; Katagishi. T; Ohkawara. T; Kakusui. M;
Kimura. H; Okanoue. T; Kashima. K; Ashihara. T. (1995). Identification of apoptotic hepatocytes in
situ in rat liver after lead nitrate administration. J Gastroenterol 30: 725-730.
http://dx.doi.org/10.1007/BF02349638
Nakao. K; Kibavashi. K; Taki. T; Koyama. H. (2010). Changes in the brain after intracerebral
implantation of a lead pellet in the rat. J Neurotrauma 27: 1925-1934.
http://dx.doi.org/10.1089/neu.2010.1379
Nakhoul. F; Kayne. LH; Brautbar. N; Hu. MS: Mcdonough. A; Eggena. P; Golub. MS: Berger. M; Chang.
CT; Jamgotchian. N; Lee. DBN. (1992). Rapid hypertensinogenic effect of lead: Studies in the
spontaneously hypertensive rat. Toxicol Ind Health 8: 89-102.
Nampoothiri. LP; Agarwal. A; Gupta. S. (2007). Effect of co-exposure to lead and cadmium on
antioxidant status in rat ovarian granulose cells. Arch Toxicol 81: 145-150.
http://dx.doi.org/10.1007/s00204-006-0133-x
Nampoothiri. LP; Gupta. S. (2006). Simultaneous effect of lead and cadmium on granulosa cells: A
cellular model for ovarian toxicity. Reprod Toxicol 21: 179-185.
http://dx.doi.0rg/10.1016/i.reprotox.2005.07.010
Nampoothiri. LP; Gupta. S. (2008). Biochemical effects of gestational coexposure to lead and cadmium
on reproductive performance, placenta, and ovary. J Biochem Mol Toxicol 22: 337-344.
http://dx.doi.org/10.1002/ibt.20246
Nandi. S: Gupta. PS: Selvaraju. S: Roy. SC: Ravindra. JP (2010). Effects of exposure to heavy metals on
viability, maturation, fertilization, and embryonic development of buffalo (Bubalus bubalis) oocytes in
vitro. Arch Environ Contam Toxicol 58: 194-204. http://dx.doi.org/10.1007/s00244-009-9342-7
Naravana. K; Al-Bader. M. (2011). Ultrastructural and DNA damaging effects of lead nitrate in the liver
[Review]. Exp Toxicol Pathol 63: 43-51. http://dx.doi.Org/10.1016/i.etp.2009.09.007
Nation. JR; Gleaves. DH. (2001). Low-level lead exposure and intelligence in children [Comment]. Arch
ClinNeuropsychol 16: 375-388.
Nava-Hernandez. MP; Hauad-Marroquin. LA; Bassol-Mavagoitia. S: Garcia-Arenas. G: Mercado-
Hernandez. R; Echavarri-Guzman. MA; Cerda-Flores. RM. (2009). Lead-, cadmium-, and arsenic-
induced DNA damage in rat germinal cells. DNA Cell Biol 28: 241-248.
http://dx.doi.org/10.1089/dna.2009.0860
November 2012 5-763 Draft - Do Not Cite or Quote
-------
Navarro-Moreno. LG: Quintanar-Escorza. MA; Gonzalez. S: Mondragon. R; Cerbon-Solorzano. J;
Valdes. J: Calderon-Salinas. JV. (2009). Effects of lead intoxication on intercellular junctions and
biochemical alterations of the renal proximal tubule cells. Toxicol In Vitro 23: 1298-1304.
http://dx.doi.0rg/10.1016/i.tiv.2009.07.020
Navas-Acien. A; Schwartz. BS: Rothenberg. SJ: Hu. H; Silbergeld. EK; Guallar. E. (2008). Bone lead
levels and blood pressure endpoints: Ameta-analysis. Epidemiology 19: 496-504.
http://dx.doi.org/10.1097/EDE.Ob013e31816a2400
Navas-Acien. A; Selvin. E; Sharrett. AR; Calderon-Aranda. E; Silbergeld. E; Guallar. E. (2004). Lead,
cadmium, smoking, and increased risk of peripheral arterial disease. Circulation 109: 3196-3201.
http://dx.doi.org/10.1161/01.CIR.0000130848.18636.B2
Navas-Acien. A; Silbergeld. EK; Sharrett. AR; Calderon-Aranda. E; Selvin. E; Guallar. E. (2005). Metals
in urine and peripheral arterial disease. Environ Health Perspect 113: 164-169.
http://dx.doi.org/10.1289/ehp.7329
Navas-Acien. A; Tellez-Plaza. M; Guallar. E; Muntner. P; Silbergeld. E; Jaar. B; Weaver. V. (2009). Blood
cadmium and lead and chronic kidney disease in US adults: Ajoint analysis. Am J Epidemiol 170:
1156-1164. http://dx.doi.org/10.1093/aie/kwp248
Naves. M; Diaz-Lopez. JB; Gomez. C: Rodriguez-Rebollar. A; Cannata-Andia. JB. (2005). Determinants
of incidence of osteoporotic fractures in the female Spanish population older than 50. Osteoporos Int
16: 2013-2017. http://dx.doi.org/10.1007/s00198-005-1983-4
Nawrot TS: Thijs. L; Den Hond. EM; Roels. HA; Staessen. JA. (2002). An epidemiological re-appraisal
of the association between blood pressure and blood lead: Ameta-analysis. J Hum Hypertens 16:
123-131. http://dx.doi.org/10.1038/si/ihh/1001300
Navak. BN; Ray. M; Persaud. TVN. (1989a). Maternal and fetal chromosomal aberrations in mice
following prenatal exposure to subembryotoxic doses of lead nitrate. Acta Anat 135: 185-188.
http://dx.doi.org/10.1159/000146751
Navak. BN; Ray. M; Persaud. TVN; Nigli. M. (1989b). Relationship of embryotoxicity to genotoxicity of
lead nitrate in mice. Exp Pathol 36: 65-73.
Neal. A; Stansfield. K; Worlev. P; Thompson. R; Guilarte. T. (2010a). Lead exposure during
synaptogenesis alters vesicular proteins and impairs vesicular release: Potential role of NMD A
receptor-dependent BDNF signaling. Toxicol Sci 116: 249-263.
http://dx.doi.org/10.1093/toxsci/kfqlll
Neal. RE; Lin. C: Isom. R; Vaishnav. K; Zigler. JS. Jr. (2010b). Opacification of lenses cultured in the
presence of Pb. Mol Vis 16: 2137-2145.
Neaton. JD; Kuller. L; Stamler. J: Wentworth. DN. (1995). Impact of systolic and diastolic blood pressure
on cardiovascular mortality. In JH Laragh; BM Brenner (Eds.), Hypertension: Pathophysiology,
diagnosis, and management (2nd ed., pp. 127-144). New York, NY: Raven Press Ltd.
Needleman. HL; Bellinger. D. (2001). Studies of lead exposure and the developing central nervous
system: A reply to Kaufman [Comment]. Arch Clin Neuropsychol 16: 359-374.
http://dx.doi.org/10.1016/S0887-6177(00)00087-l
Needleman. HL; Gatsonis. CA. (1990). Low-level lead exposure and the IQ of children: Ameta-analysis
of modern studies. JAMA 263: 673-678. http://dx.doi.org/10.1001/iama.1990.03440050067035
Needleman. HL; Gunnoe. C: Leviton. A; Reed. R: Peresie. H; Maher. C: Barrett. P. (1979). Deficits in
psychologic and classroom performance of children with elevated dentine lead levels. N Engl J Med
300: 689-695. http://dx.doi.org/10.1056/NEJM197903293001301
Needleman. HL; Leviton. A; Bellinger. D. (1982). Lead-associated intellectual deficit [letter] [Letter]. N
Engl J Med 306: 367. http://dx.doi.org/10.1056/NEJM198202113060619
November 2012 5-764 Draft - Do Not Cite or Quote
-------
Needleman. HL; McFarland. C: Ness. RB; Fienberg. SE; Tobin. MJ. (2002). Bone lead levels in
adjudicated delinquents: A case control study. Neurotoxicol Teratol 24: 711-717.
http://dx.doi.org/10.1016/S0892-0362(02)00269-6
Needleman. HL; Riess. JA; Tobin. MJ: Biesecker. GE; Greenhouse. JB. (1996). Bone lead levels and
delinquent behavior. JAMA 275: 363-369. http://dx.doi.org/10.1001/iama.1996.03530290033034
Needleman. HL: Schell. A; Bellinger. D; Leviton. A; Allred. EN. (1990). The long-term effects of
exposure to low doses of lead in childhood: An 11-year follow-up report. N Engl J Med 322: 83-88.
http://dx.doi.org/10.1056/NEJM199001113220203
Nehez. M; Lorencz. R; Desi. I. (2000). Simultaneous action of cypermethrin and two environmental
pollutant metals, cadmium and lead, on bone marrow cell chromosomes of rats in subchronic
administration. Ecotoxicol Environ Saf 45: 55-60. http://dx.doi.org/10.1006/eesa.1999.1831
Nelson. AE; Chaudhary. S: Kraus. VB; Fang. F; Chen. JC: Schwartz. TA; Shi. XA; Renner. JB; Stabler.
TV: Helmick. CG: Caldwell. K: Poole. AR: Jordan. JM. (2011). Whole blood lead levels are
associated with biomarkers of joint tissue metabolism in African American and white men and
women: The Johnston County Osteoarthritis Project. Environ Res 111: 1208-1214.
http://dx.doi.0rg/10.1016/i.envres.2011.08.002
Nelson. MM; Espy. KA. (2009). Low-Level Lead Exposure and Contingency-Based Responding in
Preschoolers: An Exploratory Study. Dev Neuropsychol 34: 494-506.
http://dx.doi.org/10.1080/87565640902964565
Nemoto. K; Ito. S: Yoshida. C: Miyata. M; Kojima. M; Degawa. M. (2011). Hepatic expression of
spermatogenic genes and their transiently remarkable downregulations in Wistar-Kyoto rats in
response to lead-nitrate administration: strain-difference in the gene expression patterns. J Toxicol Sci
36: 357-364.
Nemsadze. K; Sanikidze. T; Ratiani. L; Gabunia. L; Sharashenidze. T. (2009). Mechanisms of lead-
induced poisoning [Review]. Georgian Med News92-96.
Nenov. VD; Taal. MW: Sakharova. OV: Brenner. BM. (2000). Multi-hit nature of chronic renal disease
[Review]. Curr Opin Nephrol Hypertens 9: 85-97.
Nestmann. ER; Zhang. BW. (2007). Chromosome aberration test of Pigment Yellow 34 (lead chromate) in
Chinese hamster ovary cells. Mutat Res Genet Toxicol Environ Mutagen 633: 126-132.
http://dx.doi.0rg/10.1016/i.mrgentox.2007.05.012
Neuberger. JS: Hu. SC: Drake. KD; Jim. R. (2009). Potential health impacts of heavy-metal exposure at
the Tar Creek Superfund site, Ottawa County, Oklahoma. Environ Geochem Health 31: 47-59.
http://dx.doi.org/10.1007/sl0653-008-9154-0
Newland. MC: Yezhou. S: Logdberg. B; Berlin. M. (1994). Prolonged behavioral effects of in utero
exposure to lead or methyl mercury: Reduced sensitivity to changes in reinforcement contingencies
during behavioral transitions and in steady state. Toxicol Appl Pharmacol 126: 6-15.
http://dx.doi.org/10.1006/taap.1994.1084
Ni. Z; Hou. S: Barton. CH; Vaziri. ND. (2004). Lead exposure raises superoxide and hydrogen peroxide
in human endothelial and vascular smooth muscle cells. Kidney Int 66: 2329-2336.
http://dx.doi.0rg/10.llll/i.1523-1755.2004.66032.x
Nicolescu. R: Petcu. C: Cordeanu. A; Fabritius. K; Schlumpf. M; Krebs. R: Kramer. U; Winneke. G.
(2010). Environmental exposure to lead, but not other neurotoxic metals, relates to core elements of
ADHD in Romanian children: Performance and questionnaire data. Environ Res 110: 476-483.
http://dx.doi.0rg/10.1016/i.envres.2010.04.002
Nigg. J; Knottnerus. G: Martel. M; Nikolas. M; Cavanagh. K; Karmaus. W: Rapplev. M. (2008). Low
blood lead levels associated with clinically diagnosed attention-deficit/hyperactivity disorder and
mediated by weak cognitive control. Biol Psychiatry 63: 325-331.
http://dx.doi.0rg/10.1016/i.biopsvch.2007.07.013
November 2012 5-765 Draft - Do Not Cite or Quote
-------
Nigg. JT; Nikolas. M; Mark Knottnerus. G: Cavanagh. K; Friderici. K. (2010). Confirmation and
extension of association of blood lead with attention-deficit/hyperactivity disorder (ADHD) and
ADHD symptom domains at population-typical exposure levels. J Child Psychol Psychiatry 51: 58-65.
http://dx.doi.0rg/10.llll/i.1469-7610.2009.02135.x
Nikolova. P; Kavaldzhieva. B. (1991). The effect of certain heavy metals (Mn and Pb) on parameters of
erythrocyte energy metabolism. J Hyg Epidemiol Microbiol Immunol 35: 361-365.
Niu. RY; Sun. ZL; Cheng. ZT; Li. ZG: Wang. JD. (2009). Decreased learning ability and low
hippocampus glutamate in offspring rats exposed to fluoride and lead. Environ Toxicol Pharmacol 28:
254-258. http://dx.doi.0rg/10.1016/i.etap.2009.04.012
NKF (National Kidney Foundation). (2002). K/DOQI clinical practice guidelines for chronic kidney
disease: Evaluation, classification, and stratification, Part 4: Definition and stages of chronic kidney
disease. Am J Kidney Dis 39: S46-S75. http://dx.doi.org/10.1053/ajkd.2002.30943
Nordberg. M; Winblad. B; Fratiglioni. L; Basun. H. (2000). Lead concentrations in elderly urban people
related to blood pressure and mental performance: Results from a population-based study. Am J Ind
Med 38: 290-294. httrj://dx.doi.org/10.1002/1097-0274(200009)38:3<290::AID-AJIM7>3.0.CO:2-T
Novak. J; Banks. RO. (1995). Lead and nickel alter the cardiorenal actions of endothelin in the rat. Proc
Soc Exp Biol Med 208: 191-198.
Nowak. P; Szczerbak. G: Nitka. D; Kostrzewa. RM; Sitkiewicz. T; Brus. R. (2008). Effect of prenatal
lead exposure on nigrostriatal neurotransmission and hydroxyl radical formation in rat neostriatum:
Dopaminergic-nitrergic interaction. Toxicology 246: 83-89.
http://dx.doi.0rg/10.1016/i.tox.2007.12.026
NRC (National Research Council). (2000). Toxicological effects of methylmercury. Washington, DC:
National Academy Press, http://books.nap.edu/catalog.php7record id=9899
Nriagu. J: Afeiche. M; Linder. A; Arowolo. T; Ana. G: Sridhar. MKC: Oloruntoba. EO; Obi. E; Ebenebe.
JC: Orisakwe. OE; Adesina. A. (2008). Lead poisoning associated with malaria in children of urban
areas of Nigeria. Int J Hyg Environ Health 211: 591-605. http://dx.doi.org/10.1016/Ujheh.2008.05.001
NTP (National Toxicology Program). (2011). Report on carcinogens: Twelfth edition. Washington, DC:
U.S. Department of Health and Human Services, http://ntp.niehs.nih.gov/ntp/roc/twelfth/rocl2.pdf
Nuvts. GD: Van Vlem. E; Thys. J: De Leersnijder. D; D'Haese. PC: Elseviers. MM; De Broe. ME. (1995).
New occupational risk factors for chronic renal failure. Lancet 346: 7-11.
http://dx.doi.org/10.1016/S0140-6736(95)92648-8
O'Byrne. PM; Dolovich. M; Dirks. R; Roberts. RS: Newhouse. MT. (1984). Lung epithelial permeability:
Relation to nonspecific airway responsiveness. Journal of Applied Physiology: Respiratory,
environmental and exercise physiology 57: 77-84.
O'Byrne. PM; Dolovich. M; Duvall. A; Newhouse. MT. (1982). Lung epithelial permeability after
histamine challenge [Abstract]. Am Rev Respir Dis 125: 280.
Oberlev. TD; Friedman. AL; Moser. R; Siegel. FL. (1995). Effects of lead administration on developing
rat kidney: II Functional, morphologic, and immunohistochemical studies. Toxicol Appl Pharmacol
131: 94-107. http://dx.doi.org/10.1006/taap.1995.1051
Oberto. A; Marks. N; Evans. HL; Guidotti. A. (1996). Lead (Pb+2) promotes apoptosis in newborn rat
cerebellar neurons: Pathological implications. J Pharmacol Exp Ther 279: 435-442.
Obhodas. J: Tucak-Zoric. S: Kutle. A; Valkovic. V. (2007). Indications for synergetic and antagonistic
effects between trace elements in the environment to human health. Coll Antropol 31: 209-219.
Oishi. H; Nakashima. M; Totoki. T; Tomokuni. K. (1996). Chronic lead exposure may inhibit
endothelium-dependent hyperpolarizing factor in rats. J Cardiovasc Pharmacol 28: 558-563.
http://dx.doi.org/10.1097/00005344-199610000-00013
November 2012 5-766 Draft - Do Not Cite or Quote
-------
Olafiranye. O: Zizi. F; Brimah. P; Jean-Louis. G: Makarvus. AN; Mcfarlane. S: Ogedegbe. G. (2011).
Management of hypertension among patients with coronary heart disease. Int J Hypertens 2011:
653903. http://dx.doi.org/10.4061/2011/653903
Olaleve. SB; Adaramoye. OA; Erigbali. PP; Adeniyi. OS. (2007). Lead exposure increases oxidative
stress in the gastric mucosa of HCl/ethanol-exposed rats. World J Gastroenterol 13: 5121-5126.
Olewinska. E; Kasperczvk. A; Kapka. L; Kozlowska. A; Pawlas. N; Dobrakowski. M; Birkner. E;
Kasperczyk. S. (2010). Level of DNA damage in lead-exposed workers. Ann Agric Environ Med 17:
231-236.
Oliveira. H; Spano. M; Santos. C: Pereira. MDL. (2009). Lead chloride affects sperm motility and
acrosome reaction in mice: Lead affects mice sperm motility and acrosome reaction. Cell Biol Toxicol
25: 341-353. http://dx.doi.org/10.1007/sl0565-008-9088-4
Olivero-Verbel. J: Duarte. D; Echenique. M; Guette. J: Johnson-Restrepo. B; Parsons. PJ. (2007). Blood
lead levels in children aged 5-9 years living in Cartagena, Colombia. Sci Total Environ 372: 707-716.
http://dx.doi.0rg/10.1016/i.scitotenv.2006.10.025
Olszewski. J: Latusinski. J: Kita. A; Pietkiewicz. P; Starostecka. B; Majak. J. (2006). Comparative
assessment of aluminum and lead concentrations in serum and tissue bioptates in patients with
laryngeal papilloma or cancer. B-ENT 2: 47-49.
Ong. CN; Lee. WR. (1980a). Distribution of lead-203 in human peripheral blood in vitro. Br J Ind Med
37: 78-84. http://dx.doi.Org/10.1136/oem.37.l.78
Ong. CN; Lee. WR. (1980b). High affinity of lead for fetal haemoglobin. Br J Ind Med 37: 292-298.
http://dx.doi.0rg/10.1136/oem.37.3.292
Onuegbu. AJ; Olisekodiaka. MJ; Nwaba. El; Adeveve. AD; Akinola. FFS. (2011). Assessment of some
renal indices in people occupationally exposed to lead. Toxicol Ind Health 27: 475-479.
http://dx.doi.org/10.1177/0748233710390020
Opler. MGA; Brown. AS: Graziano. J: Desai. M; Zheng. W: Schaefer. C: Factor-Litvak. P; Susser. ES.
(2004). Prenatal lead exposure, delta-aminolevulinic acid, and schizophrenia. Environ Health Perspect
112: 548-552. http://dx.doi.org/10.1289/ehp.10464
Opler. MGA: Buka. SL; Groeger. J: McKeague. I; Wei. C: Factor-Litvak. P; Bresnahan. M; Graziano. J:
Goldstein. JM; Seidman. LJ; Brown. AS: Susser. ES. (2008). Prenatal exposure to lead, delta-
aminolevulinic acid, and schizophrenia: Further evidence. Environ Health Perspect 116: 1586-1590.
http://dx.doi.org/10.1289/ehp.10464
Orisakwe. OE; Nwachukwu. E; Osadolor. HB; Afonne. OJ: Okocha. CE. (2007). Liver and kidney
function tests amongst paint factory workers in Nkpor, Nigeria. Toxicol Ind Health 23: 161-165.
http://dx.doi.org/10.1177/0748233707081908
Oriin. E; Yalcm. SS: Aykut. O: Orhan. G: Morgil. GK; Yurdakok. K; Uzun. R. (2011). Breast milk lead
and cadmium levels from suburban areas of Ankara. Sci Total Environ 409: 2467-2472.
http://dx.doi.0rg/10.1016/i.scitotenv.2011.02.035
Oskarsson. A; Fowler. BA. (1985). Effects of lead inclusion bodies on subcellular distribution of lead in
rat kidney: The relationship to mitochondria! function. Exp Mol Pathol 43: 397-408.
http://dx.doi.org/10.1016/0014-4800(85)90076-0
Oskarsson. A; Squibb. KS: Fowler. BA. (1982). Intracellular binding of lead in the kidney: The partial
isolation and characterization of postmitochondrial lead binding components. Biochem Biophys Res
Commun 104: 290-298. http://dx.doi.org/10.1016/0006-291X(82)91973-8
Osman. K; Pawlas. K; Schutz. A; Gazdzik. M; Sokal. JA; Vahter. M. (1999). Lead exposure and hearing
effects in children in Katowice, Poland. Environ Res 80: 1-8. http://dx.doi.org/10.1006/enrs.1998.3886
Osterode. W: Barnas. D; Geissler. K. (1999). Dose dependent reduction of erythroid progenitor cells and
inappropriate erythropoietin response in exposure to lead: new aspects of anaemia induced by lead.
Occup Environ Med 56: 106-109.
November 2012 5-767 Draft - Do Not Cite or Quote
-------
Osterode. W: Ulberth. F. (2000). Increased concentration of arachidonic acid in erythrocyte membranes in
chronically lead-exposed men. J Toxicol Environ Health A 59: 87-95.
http://dx.doi.org/10.1080/009841000156998
Ostro. B; Feng. WY; Broadwin. R; Green. S: Lipsett. M. (2007). The effects of components of fine
paniculate air pollution on mortality in California: Results from CALFINE. Environ Health Perspect
115: 13-19. http://dx.doi.org/10.1289/ehp.9281
Oteiza. PI; Kleinman. CG: Demasi. M; Bechara. EJH. (1995). 5-Aminolevulinic acid induces iron release
from ferritin. Arch Biochem Biophys 316: 607-611. http://dx.doi.org/10.1006/abbi.1995.1080
Otto. DA; Fox. DA. (1993). Auditory and visual dysfunction following lead exposure [Review].
Neurotoxicology 14: 191-207.
Overmann. SR. (1977). Behavioral effects of asymptomatic lead exposure during neonatal development
in rats. Toxicol Appl Pharmacol 41: 459-471. http://dx.doi.org/10.1016/S0041-008X(77)80002-l
Ozsov. SY; Ozsov. B; Ozyildiz. Z; Aytekin. I. (2010). Protective effect of L-c ami tine on experimental
lead toxicity in rats: A clinical, histopathological and immunohistochemical study. Biotech Histochem
86: 436-443. http://dx.doi.org/10.3109/10520295.2010.529825
Pace. BM; Lawrence. DA; Behr. MJ; Parsons. PJ: Dias. JA. (2005). Neonatal lead exposure changes
quality of sperm and number of macrophages in testes of BALB/c mice. Toxicology 210: 247-256.
http://dx.doi.0rg/10.1016/i.tox.2005.02.004
Paglia. DE; Valentine. WN. (1975). Characteristics of a pyrimidine-specific 5'-nucleotidase in human
erythrocytes. J Biol Chern 250: 7973-7979.
Palaniappan. K; Roy. A; Balakrishnan. K; Gopalakrishnan. L; Mukherjee. B; Hu. H; Bellinger. DC.
(2011). Lead exposure and visual-motor abilities in children from Chennai, India. Neurotoxicology 32:
465-470. http://dx.doi.0rg/10.1016/i.neuro.2011.03.011
Palus. J: Rydzynski. K; Dziubaltowska. E; Wvszynska. K; Nataraian. AT; Nilsson. R. (2003). Genotoxic
effects of occupational exposure to lead and cadmium. Mutat Res Genet Toxicol Environ Mutagen
540: 19-28. http://dx.doi.org/10.1016/S1383-5718(03)00167-0
Pan. SY; Morrison. H; Gibbons. L; Zhou. J: Wen. SW: DesMeules. M; Mao. Y. (2011). Breast cancer risk
associated with residential proximity to industrial plants in Canada. J Occup Environ Med 53:
522-529. http://dx.doi.org/10.1097/JOM.Ob013e318216dOb3
Pandva. C: Pillai. P: Nampoothiri. LP; Bhatt. N; Gupta. S: Gupta. S. (2012). Effect of lead and cadmium
co-exposure on testicular steroid metabolism and antioxidant system of adult male rats. Andrologia
44: 813-822. http://dx.doi.Org/10.llll/i.1439-0272.2010.01137.x
Pandva. CD; Pillai. PP; Gupta. SS. (2010). Lead and cadmium co-exposure mediated toxic insults on
hepatic steroid metabolism and antioxidant system of adult male rats. Biol Trace Elem Res 134:
307-317. http://dx.doi.org/10.1007/sl2011-009-8479-6
Park. SK; Elmarsafawv. S: Mukherjee. B; Spiro. A. Ill; Vokonas. PS: Nie. H; Weisskopf. MG: Schwartz.
J; Hu. H. (2010). Cumulative lead exposure and age-related hearing loss: The VA Normative Aging
Study. Hear Res 269: 48-55. http://dx.doi.Org/10.1016/i.heares.2010.07.004
Park. SK; Hu. H; Wright. RO: Schwartz. J: Cheng. Y; Sparrow. D; Vokonas. PS: Weisskopf. MG.
(2009b). Iron metabolism genes, low-level lead exposure, and QT interval. Environ Health Perspect
117: 80-85. http://dx.doi.org/10.1289/ehp.11559
Park. SK; Mukherjee. B; Xia. X; Sparrow. D; Weisskopf. MG: Nie. H; Hu. H. (2009c). Bone lead level
prediction models and their application to examine the relationship of lead exposure and hypertension
in the third National Health and Nutrition Examination Survey. J Occup Environ Med 51: 1422-1436.
http://dx.doi.org/10.1097/JOM.Ob013e3181bf6c8d
Park. SK; Schwartz. J: Weisskopf. M; Sparrow. D: Vokonas. PS: Wright. RO: Coull. B; Nie. H; Hu. H.
(2006). Low-level lead exposure, metabolic syndrome, and heart rate variability: The VA Normative
Aging Study. Environ Health Perspect 114: 1718-1724.
November 2012 5-768 Draft - Do Not Cite or Quote
-------
Parkinson. DK; Ryan. C: Bromet EJ; Connell. MM. (1986). A psychiatric epidemiologic study of
occupational lead exposure. Am J Epidemiol 123: 261-269.
Parr. PR; Harris. EJ. (1976). The effect of lead on the calcium-handling capacity of rat heart
mitochondria. Biochemistry 158: 289-294.
Pasha. Q: Malik. SA; Iqbal. J: Shah. MH. (2007). Characterization and distribution of the selected metals
in the scalp hair of cancer patients in comparison with normal donors. Biol Trace Elem Res 118:
207-216. http://dx.doi.org/10.1007/sl2011-007-0035-7
Pasha. Q: Malik. SA; Iqbal. J: Shaheen. N; Shah. MH. (2008a). Comparative distribution of the scalp hair
trace metal contents in the benign tumour patients and normal donors. Environ Monit Assess 147:
377-388. http://dx.doi.org/10.1007/sl0661-007-0127-z
Pasha. Q: Malik. SA; Iqbal. J: Shaheen. N; Shah. MH. (2008b). Comparative evaluation of trace metal
distribution and correlation in human malignant and benign breast tissues. Biol Trace Elem Res 125:
30-40. http://dx.doi.org/10.1007/sl2011-008-8158-z
Pasha. Q: Malik. SA; Shah. MH. (2008c). Statistical analysis of trace metals in the plasma of cancer
patients versus controls. J Hazard Mater 153: 1215-1221.
http://dx.doi.0rg/10.1016/i.jhazmat.2007.09.115
Pasha. Q: Malik. SA; Shaheen. N; Shah. MH. (2010). Investigation of trace metals in the blood plasma
and scalp hair of gastrointestinal cancer patients in comparison with controls. Clin Chim Acta 411:
531-539. http://dx.doi.0rg/10.1016/i.cca.2010.01.010
Pasha Shaik. A; Sankar. S: Reddv. SC: Das. PG: Jamil. K. (2006). Lead-induced genotoxicity in
lymphocytes from peripheral blood samples of humans: In vitro studies. Drug Chem Toxicol 29:
111-124. http://dx.doi.org/10.1080/01480540500408739
Pastor-Barriuso. R; Banegas. JR; Damian. J: Appel. LJ; Guallar. E. (2003). Systolic blood pressure,
diastolic blood pressure, and pulse pressure: An evaluation of their joint effect on mortality. Ann
Intern Med 139: 731-739.
Patel. AB; Prabhu. AS. (2009). Determinants of lead level in umbilical cord blood. Indian Pediatr 46:
791-793.
Patil. AJ; Bhagwat VR; Patil. JA; Dongre. NN; Ambekar. JG: Das. KK. (2006a). Biochemical aspects of
lead exposure in silver jewelry workers in western Maharashtra (India). J Basic Clin Physiol
Pharmacol 17: 213-229.
Patil. AJ; Bhagwat. VR: Patil. JA; Dongre. NN; Ambekar. JG: Das. KK. (2007). Occupational lead
exposure in battery manufacturing workers, silver jewelry workers, and spray painters in western
Maharashtra (India): Effect on liver and kidney function. J Basic Clin Physiol Pharmacol 18: 87-100.
http://dx.doi.Org/10.1515/JBCPP.2007.18.2.87
Patil. AJ: Bhagwat. VR: Patil. JA; Dongre. NN; Ambekar. JG: Jailkhani. R; Das. KK. (2006b). Effect of
lead (Pb) exposure on the activity of superoxide dismutase and catalase in battery manufacturing
workers (BMW) of Western Maharashtra (India) with reference to heme biosynthesis. Int J Environ
Res Public Health 3: 329-337.
Pavanello. S: Bollati. V: Pesatori. AC: Kapka. L; Bolognesi. C: Bertazzi. PA; Baccarelli. A. (2009).
Global and gene-specific promoter methylation changes are related to anti-B[a]PDE-DNA adduct
levels and influence micronuclei levels in polycyclic aromatic hydrocarbon-exposed individuals. Int J
Cancer 125: 1692-1697. http://dx.doi.org/10.1002/iic.24492
Pavanello. S: Pesatori. AC: Dioni. L; Hoxha. M; Bollati. V: Siwinska. E; Mielzynska. D; Bolognesi. C:
Bertazzi. PA; Baccarelli. A. (2010). Shorter telomere length in peripheral blood lymphocytes of
workers exposed to polycyclic aromatic hydrocarbons. Carcinogenesis 31: 216-221.
http://dx.doi.org/10.1093/carcin/bgp278
Pavton. M; Hu. H; Sparrow. D; Weiss. ST. (1994). Low-level lead exposure and renal function in the
Normative Aging Study. Am J Epidemiol 140: 821-829.
November 2012 5-769 Draft - Do Not Cite or Quote
-------
Pavton. M; Riggs. KM; Spiro. A. Ill: Weiss. ST; Hu. H. (1998). Relations of bone and blood lead to
cognitive function: The VA Normative Aging Study. Neurotoxicol Teratol 20: 19-27.
Pekcici. R; Kavlakoglu. B; Yilmaz. S: Sahin. M; Delibasi. T. (2010). Effects of lead on thyroid functions
in lead-exposed workers. Central European Journal of Medicine 5: 215-218.
http://dx.doi.org/10.2478/sll536-009-0092-8
Peng. S: Hajela. RK: Atchison. WD. (2002). Characteristics of block by Pb2+ of function of human
neuronal L-, N-, and R-type Ca2+ channels transiently expressed in human embryonic kidney 293
cells. Mol Pharmacol 62: 1418-1430. http://dx.doi.org/10.1124/mol.62.6.1418
Pentvala. S: Ruggeri. J; Veerraiu. A; Yu. Z; Bhatia. A; Desaiah. D; Vig. P. (2010). Microsomal Ca2+ flux
modulation as an indicator of heavy metal toxicity. Indian J Exp Biol 48: 737-743.
Perez-Bravo. F; Ruz. M; Moran-Jimenez. MJ; Olivares. M; Rebolledo. A; Codoceo. J: Sepulveda. V:
Jenkin. A; Santos. JL; Fontanellas. A. (2004). Association between aminolevulinate dehydrase
genotypes and blood lead levels in children from a lead-contaminated area in Antofagasta, Chile. Arch
Environ Contam Toxicol 47: 276-280. http://dx.doi.org/10.1007/s00244-004-2215-l
Perlstein. T; Weuve. J: Schwartz. J: Sparrow. D; Wright. R; Litonjua. A; Nie. H; Hu. H. (2007).
Cumulative community-level lead exposure and pulse pressure: The Normative Aging Study. Environ
Health Perspect 115: 1696-1700. http://dx.doi.org/10.1289/ehp.10350
Peters. JL; Kubzansky. L; McNeely. E; Schwartz. J: Spiro. A. Ill; Sparrow. D: Wright. RO: Nie. H; Hu. H.
(2007). Stress as a potential modifier of the impact of lead levels on blood pressure: The Normative
Aging Study. Environ Health Perspect 115: 1154-1159. http://dx.doi.org/10.1289/ehp.10002
Petit. TL; Leboutillier. JC. (1979). Effects of lead exposure during development on neocortical dendritic
and synaptic structure. Exp Neural 64: 482-492. http://dx.doi.org/10.1016/0014-4886(79)90226-7
Piao. F; Cheng. F; Chen. H; Li. G: Sun. X; Liu. S: Yamauchi. T; Yokoyama. K. (2007). Effects of zinc
coadministration on lead toxicities in rats. Ind Health 45: 546-551.
Piccinini. F; Favalli. L; Chiari. MC. (1977). Experimental investigations on the conctraction induced by
lead in arterial smooth muscle. Toxicology 8: 43-51. http://dx.doi.org/10.1016/0300-483X(77)90022-l
Pillai. P: Pandva. C: Bhatt N; Gupta. SS. (2012). Biochemical and reproductive effects of
gestational/lactational exposure to lead and cadmium with respect to testicular steroidogenesis,
antioxidant system, endogenous sex steroid and cauda-epididymal functions. Andrologia 44: 92-101.
http://dx.doi.0rg/10.llll/i.1439-0272.2010.01109.x
Pillai. P; Pandva. C: Gupta. S: Gupta. S. (2010). Biochemical and molecular effects of gestational and
lactational coexposure to lead and cadmium on ovarian steroidogenesis are associated with oxidative
stress in Fl generation rats. J Biochem Mol Toxicol 24: 384-394. http://dx.doi.org/10.1002/ibt.20351
Pillai. P: Patel. R: Pandva. C: Gupta. S. (2009). Sex-specific effects of gestational and lactational
coexposure to lead and cadmium on hepatic phase I and phase II xenobiotic/steroid-metabolizing
enzymes and antioxidant status. J Biochem Mol Toxicol 23: 419-431.
http://dx.doi.org/10.1002/ibt.20305
Pilsner. JR; Hu. H; Ettinger. A; Sanchez. BN; Wright. RO: Cantonwine. D: Lazarus. A; Lamadrid-
Figueroa. H; Mercado-Garcia. A; Tellez-Rojo. MM; Hernandez-Avila. M. (2009). Influence of
prenatal lead exposure on genomic methylation of cord blood DNA. Environ Health Perspect 117:
1466-1471. http://dx.doi.org/10.1289/ehp.0800497
Pilsner. JR; Hu. H; Wright. RO: Kordas. K; Ettinger. AS: Sanchez. BN; Cantonwine. D: Lazarus. AL;
Cantoral. A; Schnaas. L; Tellez-Rojo. MM; Hernandez-Avila. M. (2010). Maternal MTHFR genotype
and haplotype predict deficits in early cognitive development in a lead-exposed birth cohort in Mexico
City. Am J Clin Nutr 92: 226-234. http://dx.doi.org/10.3945/aicn.2009.28839
Pine. MD; Hinev. JK; Dearth. RK; Bratton. GR; Dees. WL. (2006). IGF-1 administration to prepubertal
female rats can overcome delayed puberty caused by maternal Pb exposure. Reprod Toxicol 21:
104-109. http://dx.doi.0rg/10.1016/i.reprotox.2005.07.003
November 2012 5-770 Draft - Do Not Cite or Quote
-------
Pineda-Zavaleta. AP; Garcia-Vargas. G: Borja-Aburto. VH; Acosta-Saavedra. LC: Aguilar. EV; Gomez-
Mufioz. A; Cebrian. ME; Calderon-Aranda. ES. (2004). Nitric oxide and superoxide anion production
in monocytes from children exposed to arsenic and lead in region Lagunera, Mexico. Toxicol Appl
Pharmacol 198: 283-290. http://dx.doi.Org/10.1016/i.taap.2003.10.034
Pinkerton. LE; Biagini. RE; Ward. EM; Hull. RD; Deddens. JA; Boeniger. MF; Schnorr. TM; MacKenzie.
BA; Luster. MI. (1998). Immunologic findings among lead-exposed workers. Am J Ind Med 33:
400-408. httrj://dx.doi.org/10.1002/(SICI)1097-0274(199804)33:4<400::AID-AJIMll>3.0.CO:2-2
Pinto. D; Ceballos. JM; Garcia. G: Guzman. P; Del Razo. LM; Vera. E; Gomez. H; Garcia. A; Gonsebatt.
ME. (2000). Increased cytogenetic damage in outdoor painters. Mutat Res Genet Toxicol Environ
Mutagen467: 105-111. http://dx.doi.org/10.1016/S1383-5718(00)00024-3
Pirkle. JL; Brodv. DJ: Gunter. EW: Kramer. RA; Paschal. DC: Flegal. KM; Matte. TD. (1994). The
decline in blood lead levels in the United States: The National Health and Nutrition Examination
Surveys (NHANES). JAMA 272: 284-291. http://dx.doi.org/10.1001/jama.1994.03520040046039
Pizent. A; Colak. B; Kljakovic Gaspic. Z; Telisman. S. (2009). Prostate-specific antigen (PSA) in serum
in relation to blood lead concentration and alcohol consumption in men. Arh Hig Rada Toksikol 60:
69-78. http://dx.doi.org/10.2478/10004-1254-60-2009-1901
Pizent. A; Macan. J; Jurasovic. J: Varnai. VM; Milkovic-Kraus. S: Kanceljak-Macan. B. (2008).
Association of toxic and essential metals with atopy markers and ventilatory lung function in women
and men. Sci Total Environ 390: 369-376. http://dx.doi.Org/10.1016/j.scitotenv.2007.10.049
Plusquellec. P; Muckle. G: Dewailly. E; Ayotte. P; Begin. G: Desrosiers. C: Despres. C: Saint-Amour. D;
Poitras. K. (2010). The relation of environmental contaminants exposure to behavioral indicators in
Inuit preschoolers in Arctic Quebec. Neurotoxicology 31: 17-25.
http://dx.doi.0rg/10.1016/i.neuro.2009.10.008
Plusquellec. P: Muckle. G: Dewailly. E; Ayotte. P: Jacobson. SW: Jacobson. JL. (2007). The relation of
low-level prenatal lead exposure to behavioral indicators of attention in Inuit infants in Arctic Quebec.
Neurotoxicol Teratol 29: 527-537. http://dx.doi.Org/10.1016/i.ntt.2007.07.002
Pocock. SJ: Smith. M; Baghurst P (1994). Environmental lead and children's intelligence: A systematic
review of the epidemiological evidence [Review]. Br Med J 309: 1189-1197.
http://dx.doi.org/10.1136/bmj.309.6963.1189
Pollack. AZ; Schisterman. EF; Goldman. LR; Mumford. SL; Albert. PS: Jones. RL; Wactawski-Wende. J.
(2011). Cadmium, lead, and mercury in relation to reproductive hormones and anovulation in
premenopausal women. Environ Health Perspect 119: 1156-1161.
http://dx.doi.org/10.1289/ehp.1003284
Pollitt. E. (2005). Are the psychological tests valid? [Letter]. Am J Clin Nutr 82: 201; author reply
201-201; author reply 202.
Porgba. R; Gac. P; Poreba. M; Antonowicz-Juchniewicz. J; Andrzejak. R. (2011 a). Relationship between
occupational exposure to lead and local arterial stiffness and left ventricular diastolic function in
individuals with arterial hypertension. Toxicol Appl Pharmacol 254: 342-348.
http://dx.doi.0rg/10.1016/i.taarj.2011.05.009
Poreba. R; Gac. P; Poreba. M; Derkacz. A; Pilecki. W; Antonowicz-Juchniewicz. J; Andrzejak. R. (2010).
[Relationship between chronic exposure to lead, cadmium and manganese, blood pressure values and
incidence of arterial hypertension]. Med Pr 61: 5-14.
Poreba. R; Poreba. M; Gac. P; Andrzejak. R. (2011). Ambulatory blood pressure monitoring and
structural changes in carotid arteries in normotensive workers occupationally exposed to lead. Hum
Exp Toxicol 30: 1174-1180. http://dx.doi.org/10.1177/0960327110391383
Poreba. R; Poreba. M; Gac. P; Pilecki. W; Andrzejak. R. (2010). Impaired endothelial function assessed
by flow-mediated vasodilatation in workers occupationally exposed to lead. Environ Toxicol
Pharmacol 29: 266-270. http://dx.doi.Org/10.1016/i.etap.2010.03.001
November 2012 5-771 Draft - Do Not Cite or Quote
-------
Porgba. R; Porgba. M; Gac. P; Steinmetz-Beck. A; Beck. B; Pilecki. W: Andrzejak. R; Sobieszczanska.
M. (20lib). Electrocardiographic changes in workers occupationally exposed to lead. Ann
Noninvasive Electrocardiol 16: 33-40. http://dx.doi.Org/10.llll/i.1542-474X.2010.00406.x
Poulos. L; Qammaz. S: Athanaselis. S: Maravelias. C: Koutselinis. A. (1986). Statistically significant
hematopoietic effects of low blood lead levels. Arch Environ Occup Health 41: 384-386.
Prentice. RC: Kopp. SJ. (1985). Cardiotoxicity of lead at various perfusate calcium concentrations:
Functional and metabolic responses of the perfused rat heart. Toxicol Appl Pharmacol 81: 491-501.
http://dx.doi.org/10.1016/0041-008X(85)90420-X
Prins. JM; Park. S: Lurie. PL (2010). Decreased expression of the voltage-dependent anion channel in
differentiated PC-12 and SH-SY5Ycells following low-level Pb exposure. Toxicol Sci 113: 169-176.
http://dx.doi.org/10.1093/toxsci/kfp249
Prospective Studies Collaboration. (2002). Age-specific relevance of usual blood pressure to vascular
mortality: A meta-analysis of individual data for one million adults in 61 prospective studies. Lancet
360: 1903-1913. http://dx.doi.org/10.1016/80140-6736(02)11911-8
Prozialeck. WC: Grunwald. GB; Dev. PM; Reuhl. KR; Parrish. AR. (2002). Cadherins and NCAM as
potential targets in metal toxicity [Review]. Toxicol Appl Pharmacol 182: 255-265.
http://dx.doi.org/10.1006/taap.2002.9422
Pugh Smith. P; Nriagu. JO. (2011). Lead poisoning and asthma among low-income and African American
children in Saginaw, Michigan. Environ Res 111: 81-86.
http://dx.doi.0rg/10.1016/i.envres.2010.ll.007
Purdv. RE; Smith. JR; Ding. Y; Oveisi. F; Vaziri. ND; Gonick. HC. (1997). Lead-induced hypertension is
not associated with altered vascular reactivity in vitro. Am J Hypertens 10: 997-1003.
http://dx.doi.org/10.1016/S0895-7061(97)00108-8
Putnam. CD; Arvai. AS: Bourne. Y; Tainer. JA. (2000). Active and inhibited human catalase structures:
Ligand and NADPH binding and catalytic mechanism. J Mol Biol 296: 295-309.
http://dx.doi.org/10.1006/imbi.1999.3458
Pvatt. DW: Zheng. JH; Stillman. WS: Irons. RD. (1996). Inorganic lead activates NF-kappa B in primary
human CD4+ T lymphocytes. Biochem Biophys Res Commun 227: 380-385.
http://dx.doi.org/10.1006/bbrc. 1996.1516
Qian. Y; Harris. ED; Zheng. Y; Tiffany-Castiglioni. E. (2000). Lead targets GRP78, a molecular
chaperone, in C6 rat glioma cells. Toxicol Appl Pharmacol 163: 260-266.
http://dx.doi.org/10.1006/taap.1999.8878
Qian. Y; Zheng. Y; Ramos. KS: Tiffany-Castiglioni. E. (2005). GRP78 compartmentalized redistribution
in Pb-treated glia: Role of GRP78 in lead-induced oxidative stress. Neurotoxicology 26: 267-275.
http://dx.doi.0rg/10.1016/i.neuro.2004.09.002
Qian. ZM; Morgan. EH. (1990). Effect of lead on the transport of transferrin-free and transferrin-bound
iron into rabbit reticulocytes. Biochem Pharmacol 40: 1049-1054.
http://dx.doi.org/10.1016/0006-2952(90)90492-4
Qiao. YF; Jiang. YS: Pang. DZ. (2006). [Expression of renal nuclear factor-kappaB, transforming growth
factor-beta and fibronectin of rats exposed to lead]. Zhonghua Lao Dong Wei Sheng Zhi Ye Bing Za
Zhi 24: 139-142.
Qu. W: Diwan. BA; Liu. J: Gover. RA; Dawson. T; Horton. JL; Cherian. MG: Waalkes. MP (2002). The
metallothionein-null phenotype is associated with heightened sensitivity to lead toxicity and an
inability to form inclusion bodies. Am J Pathol 160: 1047-1056.
http://dx.doi.org/10.1016/S0002-9440(10)64925-5
Queirolo. El; Ettinger. AS: Stoltzfus. RJ: Kordas. K. (2010). Association of anemia, child and family
characteristics with elevated blood lead concentrations in preschool children from Montevideo,
Uruguay. Arch Environ Occup Health 65: 94-100. http://dx.doi.org/10.1080/19338240903390313
November 2012 5-772 Draft - Do Not Cite or Quote
-------
Queiroz. MLS: Almeida. M; Gallao. MI; Hoehr. NF. (1993). Defective neutrophil function in workers
occupationally exposed to lead. Pharmacol Toxicol 72: 73-77.
http://dx.doi.0rg/10.llll/i.1600-0773.1993.tb00293.x
Queiroz. MLS: Costa. FF; Bincoletto. C: Perlingeiro. RCR: Dantas. PCM; Cardoso. MP; Almeida. M.
(1994a). Engulfment and killing capabilities of neutrophils and phagocytic splenic function in persons
occupationally exposed to lead. International Journal of Immunopharmacology 16: 239-244.
http://dx.doi.org/10.1016/0192-0561(94)90018-3
Queiroz. MLS: da Rocha. MC: Torello. CO: de Souza Queiroz. J; Bincoletto. C: Morgano. MA; Romano.
MR; Paredes-Gamero. EJ; Barbosa. CMV; Calgarotto. AK. (2011). Chlorella vulgaris restores bone
marrow cellularity and cytokine production in lead-exposed mice. Food Chem Toxicol 49: 2934-2941.
http://dx.doi.0rg/10.1016/i.fct.2011.06.056
Queiroz. MLS: Perlingeiro. RCR: Bincoletto. C: Almeida. M; Cardoso. MP; Dantas. PCM. (1994b).
Immunoglobulin levels and cellular immune function in lead exposed workers. Immunopharmacol
Immunotoxicol 16: 115-128. http://dx.doi.org/10.3109/08923979409029904
Quinlan. GJ: Halliwell. B; Moorhouse. CP; Gutteridge. JMC. (1988). Action of lead(II) and
aluminium(III) ions on iron-stimulated lipid peroxidation in liposomes, erythrocytes and rat liver
microsomal fractions. Biochim Biophys Acta 962: 196-200.
http://dx.doi.org/10.1016/0005-2760(88)90159-2
Quintanar-Escorza. MA; Gonzalez-Martinez. MT; del Pilar. IQM; Calderon-Salinas. JV. (2010).
Oxidative damage increases intracellular free calcium [Ca2+]i concentration in human erythrocytes
incubated with lead. Toxicol In Vitro 24: 1338-1346. http://dx.doi.Org/10.1016/i.tiv.2010.05.002
Quintanar-Escorza. MA; Gonzalez-Martinez. MT; Navarro. L; Maldonado. M; Arevalo. B; Calderon-
Salinas. JV. (2007). Intracellular free calcium concentration and calcium transport in human
erythrocytes of lead-exposed workers. Toxicol Appl Pharmacol 220: 1-8.
http://dx.doi.0rg/10.1016/i.taap.2006.10.016
Quintanilla-Vega. B; Smith. PR; Kahng. MW: Hernandez. JM; Albores. A; Fowler. BA. (1995). Lead-
binding proteins in brain tissue of environmentally lead-exposed humans. Chem Biol Interact 98:
193-209. http://dx.doi.org/10.1016/0009-2797(95)03646-6
Rabbani-Chadegani. A; Abdosamadi. S: Fani. N; Mohammadian. S. (2009). A comparison of the effect of
lead nitrate on rat liver chromatin, DNA and histone proteins in solution. Arch Toxicol 83: 565-570.
http://dx.doi.org/10.1007/s00204-008-0362-2
Rabbani-Chadegani. A. Ill: Fani. N; Abdossamadi. S: Shahmir. N. (2011). Toxic effects of lead and nickel
nitrate on rat liver chromatin components. J Biochem Mol Toxicol 25: 127-134.
http://dx.doi.org/10.1002/ibt.20368
Rabinowitz. MB. (1988). Lead and pregnancy. Birth 15: 236-241.
http://dx.doi.Org/10.llll/i.1523-536X.1988.tb01117.x
Rabinowitz. MB; Alfred. EN; Bellinger. DC: Leviton. A; Needleman. HL. (1990). Lead and childhood
propensity to infectious and allergic disorders: Is there an association? Bull Environ Contam Toxicol
44: 657-660. http://dx.doi.org/10.1007/BF01701784
Rabinowitz. MB; Wang. JD; Soong. WT. (1992). Children's classroom behavior and lead in Taiwan. Bull
Environ Contam Toxicol 48: 282-288. http://dx.doi.org/10.1007/BF00194385
Rafalowska. U: Struzynska. L; Dabrowska-Bouta. B; Lenkiewicz. A. (1996). Is lead toxicosis a reflection
of altered energy metabolism in brain synaptosomes? [Review]. Acta Neurobiol Exp (Wars) 56:
611-617.
Rafique. M; Khan. N; Perveen. K; Naqvi. A. (2009). The effects of lead and zinc on the quality of semen
of albino rats. J Coll Physicians Surg Pak 19: 510-513. http://dx.doi.org/08.2009/JCPSP.510513
November 2012 5-773 Draft - Do Not Cite or Quote
-------
Raghavan. SRV: Culver. BD; Gonick. HC. (1980). Erythrocyte lead-binding protein after occupational
exposure: I Relationship to lead toxicity. Environ Res 22: 264-270.
http://dx.doi.org/10.1016/0013-9351(80)90138-3
Raghavan. SRV: Culver. BD; Gonick. HC. (1981). Erythrocyte lead-binding protein after occupational
exposure: II. Influence on lead inhibition of membrane Na+, K+ - adenosinetriphosphatase. J Toxicol
Environ Health 7: 561-568. http://dx.doi.org/10.1080/15287398109530001
Raghavan. SRV: Gonick. HC. (1977). Isolation of low-molecular-weight lead-binding protein from
human erythrocytes. Proc Soc Exp Biol Med 155: 164-167.
Raine. A; Lencz. T; Bihrle. S: LaCasse. L; Colletti. P. (2000). Reduced prefrontal gray matter volume and
reduced autonomic activity in antisocial personality disorder. Arch Gen Psychiatry 57: 119-127.
Raitakari. OT; Juonala. M; Kahonen. M; Taittonen. L; Laitinen. T; Maki-Torkko. N; Jarvisalo. MJ; Uhari.
M; Jokinen. E; Ronnemaa. T; Akerblom. HK; Viikari. JS. (2003). Cardiovascular risk factors in
childhood and carotid artery intima-media thickness in adulthood: The Cardiovascular Risk in Young
Finns Study. JAMA290: 2277-2283. http://dx.doi.org/10.1001/iama.290.17.2277
Rajah. T; Ahuja. YR. (1995). In vivo genotoxic effects of smoking and occupational lead exposure in
printing press workers. Toxicol Lett 76: 71-75. http://dx.doi.org/10.1016/0378-4274(94)03200-9
Rajah. TT; Ahuja. YR. (1996). In vivo genotoxicity of alcohol consumption and lead exposure in printing
press workers. Alcohol 13: 65-68. http://dx.doi.org/10.1016/0741-8329(95)02014-4
Rajan. P; Kelsev. KT; Schwartz. JD; Bellinger. DC: Weuve. J: Sparrow. D; Spiro. A. Ill; Smith. TJ; Nie.
H; Hu. H; Wright. RO. (2007). Lead burden and psychiatric symptoms and the modifying influence of
the delta-aminolevulinic acid dehydratase (ALAD) polymorphism: The VA Normative Aging Study.
Am J Epidemiol 166: 1400-1408. http://dx.doi.org/10.1093/aie/kwm220
Rajan. P; Kelsev. KT; Schwartz. JD; Bellinger. DC: Weuve. J: Spiro. A. Ill; Sparrow. D; Smith. TJ: Nie.
H; Weisskopf. MG: Hu. H; Wright. RO. (2008). Interaction of the delta-aminolevulinic acid
dehydratase polymorphism and lead burden on cognitive function: The VA Normative Aging Study. J
Occup Environ Med 50: 1053-1061. http://dx.doi.org/10.1097/JOM.Ob013e3181792463
Rajaraman. P; Stewart. PA; Samet. JM; Schwartz. BS: Linet. MS: Zahm. SH; Rothman. N; Yeager. M;
Fine. HA; Black. PM; Loeffler. J: Shapiro. WR; Selker. RG: Inskip. PP. (2006). Lead, genetic
susceptibility, and risk of adult brain tumors. Cancer Epidemiol Biomarkers Prev 15: 2514-2520.
http://dx.doi.org/10.1158/1055-9965.EPI-06-0482
Ramesh. GT; Manna. SK; Aggarwal. BB; Jadhav. AL. (1999). Lead activates nuclear transcription factor -
kB, activator protein-1, and amino-terminal c-Jun kinase in pheochromocytoma cells. Toxicol Appl
Pharmacol 155: 280-286. http://dx.doi.org/10.1006/taap.1999.8624
Ramesh. GT; Manna. SK; Aggarwal. BB; Jadhav. AL. (2001). Lead exposure activates nuclear factor
kappa B, activator protein-1, c-Jun N-terminal kinase and caspases in the rat brain. Toxicol Lett 123:
195-207. http://dx.doi.org/10.1016/S0378-4274(01)00395-2
Rana. SVS. (2008). Metals and apoptosis: Recent developments. J Trace Elem Med Biol 22: 262-284.
http://dx.doi.0rg/10.1016/i.itemb.2008.08.002
Razmiafshari. M; Kao. J: dAvignon. A; Zawia. NH. (2001). NMR identification of heavy metal-binding
sites in a synthetic zinc finger peptide: Toxicological implications for the interactions of xenobiotic
metals with zinc finger proteins. Toxicol Appl Pharmacol 172: 1-10.
http://dx.doi.org/10.1006/taap.2001.9132
Razmiafshari. M; Zawia. NH. (2000). Utilization of a synthetic peptide as a tool to study the interaction
of heavy metals with the zinc finger domain of proteins critical for gene expression in the developing
brain. Toxicol Appl Pharmacol 166: 1-12. http://dx.doi.org/10.1006/taap.2000.8950
Reddv. GR; Zawia. NH. (2000). Lead exposure alters Egr-1 DNA-binding in the neonatal rat brain. Int J
DevNeurosci 18: 791-795. http://dx.doi.org/10.1016/S0736-5748(00)00048-4
November 2012 5-774 Draft - Do Not Cite or Quote
-------
Rees. PJ: Shelton. D; Chan. TB; Eiser. N; Clark. TJH; Maisev. MN. (1985). Effects of histamine on lung
permeability in normal and asthmatic subjects. Thorax 40: 603-606.
Regunathan. S: Sundaresan. R. (1984). Pyruvate metabolism in the brain of young rats intoxicated with
organic and inorganic lead. JNeurochem 43: 1346-1351.
http://dx.doi.org/10.1111/i.l471-4159.1984.tb05393.x
Rehman. S: Rehman. S: Chandra. O: Abdulla. M. (1995). Evaluation of malondialdehyde as an index of
lead damage in rat brain homogenates. Biometals 8: 275-279. http://dx.doi.org/10.1007/BF00141599
Reif. A; Fritzen. S: Finger. M; Strobel. A; Lauer. M; Schmitt. A; Lesch. K. (2006). Neural stem cell
proliferation is decreased in schizophrenia, but not in depression. Mol Psychiatry 11: 514-522.
http://dx.doi.org/10.1038/sj.mp.4001791
Rendon-Ramirez. A; Cerbon-Solorzano. J; Maldonado-Vega. M; Quintanar-Escorza. MA; Calderon-
Salinas. JV. (2007). Vitamin-E reduces the oxidative damage on 5-aminolevulinic dehydratase induced
by lead intoxication in rat erythrocytes. Toxicol In Vitro 21: 1121-1126.
http://dx.doi.0rg/10.1016/i.tiv.2007.04.019
Reuhl. KR; Puce. DC: Gilbert. SG: Mallett J. (1989). Effects of chronic developmental lead exposure on
monkey neuroanatomy: Visual system. Toxicol Appl Pharmacol 99: 501-509.
http://dx.doi.org/10.1016/0041-008X(89)90157-9
Revis. NW: Zinsmeister. AR; Bull. R. (1981). Atherosclerosis and hypertension induction by lead and
cadmium ions: An effect prevented by calcium ion. PNAS 78: 6494-6498.
Reza. B; Ali. N; Azhdar. H; Alireza. A; Ali. K. (2008). Effects of low-level lead exposure on blood
pressure and function of the rat isolated heart. Indian J Pharmacol 40: 69-72.
http://dx.doi.org/10.4103/0253-7613.41041
Rhodes. D; Spiro. A. Ill; Aro. A; Hu. H. (2003). Relationship of bone and blood lead levels to psychiatric
symptoms: The Normative Aging Study. J Occup Environ Med 45: 1144-1151.
http://dx.doi.org/10.1097/01.jom.0000094995.23808.7b
Rice. C: Ghorai. JK; Zalewski. K; Weber. DN. (2011). Developmental lead exposure causes startle
response deficits in zebrafish. Aquat Toxicol 105: 600-608.
http://dx.doi.0rg/10.1016/i.aquatox.2011.08.014
Rice. D: Barone. S. (2000). Critical periods of vulnerability for the developing nervous system: Evidence
from humans and animal models [Review]. Environ Health Perspect 108: 511-533.
http://dx.doi.org/10.1289/ehp.00108s3511
Rice. DC. (1990). Lead-induced behavioral impairment on a spatial discrimination reversal task in
monkeys exposed during different periods of development. Toxicol Appl Pharmacol 106: 327-333.
http://dx.doi.org/10.1016/0041-008X(90)90251-O
Rice. DC. (1992a). Effect of lead during different developmental periods in the monkey on concurrent
discrimination performance. Neurotoxicology 13: 583-592.
Rice. DC. (1992b). Lead exposure during different developmental periods produces different effects on FI
performance in monkeys tested as juveniles and adults. Neurotoxicology 13: 757-770.
Rice. DC. (1997). Effects of lifetime lead exposure in monkeys on detection of pure tones. Fundam Appl
Toxicol 36: 112-118. http://dx.doi.org/10.1006/faat.1996.2268
Rice. DC. (1998). Effects of lifetime lead exposure on spatial and temporal visual function in monkeys.
Neurotoxicology 19: 893-902.
Rice. DC: Gilbert. SG. (1985). Low lead exposure from birth produces behavioral toxicity (DRL) in
monkeys. Toxicol Appl Pharmacol 80: 421-426. http://dx.doi.org/10.1016/0041-008X(85)90386-2
Rice. DC: Gilbert. SG. (1990a). Lack of sensitive period for lead-induced behavioral impairment on a
spatial delayed alternation task in monkeys. Toxicol Appl Pharmacol 103: 364-373.
http://dx.doi.org/10.1016/0041-008X(90)90236-N
November 2012 5-775 Draft - Do Not Cite or Quote
-------
Rice. DC: Gilbert. SG. (1990b). Sensitive periods for lead-induced behavioral impairment (nonspatial
discrimination reversal) in monkeys. Toxicol Appl Pharmacol 102: 101-109.
http://dx.doi.org/10.1016/0041-008X(90)90087-B
Rice. DC: Karpinski. KF. (1988). Lifetime low-level lead exposure produces deficits in delayed
alternation in adult monkeys. Neurotoxicol Teratol 10: 207-214.
http:/7dx.doi.org/10.1016/0892-0362(88)90019-0
Richardt. G: Federolf. G: Habermann. E. (1986). Affinity of heavy metal ions to intracellular Ca2+-
binding proteins. Biochem Pharmacol 35: 1331-1335.
httrj://dx.doi.org/10.1016/0006-2952(86)90278-9
Richetti. S: Rosemberg. D; Ventura-Lima. J; Monserrat. J; Bogo. M; Bonan. C. (2010).
Acetylcholinesterase activity and antioxidant capacity of zebrafish brain is altered by heavy metal
exposure. Neurotoxicology 32: 116-122. http://dx.doi.Org/10.1016/i.neuro.2010.ll.001
Riddell. TJ; Solon. O; Quimbo. SA; Tan. CM; Butrick. E; Peabodv. JW. (2007). Elevated blood-lead
levels among children living in the rural Philippines. Bull World Health Organ 85: 674-680.
Pus. MD; Dietrich. KN; Succop. PA; Berger. OG: Bornschein. RL. (2004). Early exposure to lead and
neuropsychological outcome in adolescence. J Int Neuropsychol Soc 10: 261-270.
http://dx.doi.org/10.1017/S1355617704102154
Rizzi. E; Castro. MM; Fernandes. K; Barbosa. F; Arisi. GM; Garcia-Cairasco. N; Bendhack. LM; Tanus-
Santos. JE; Gerlach. RF. (2009). Evidence of early involvement of matrix metalloproteinase-2 in lead-
induced hypertension. Arch Toxicol 83: 439-449. http://dx.doi.org/10.1007/s00204-008-0363-l
Robins. JM; Cullen. MR; Connors. BB; Kayne. RD. (1983). Depressed thyroid indexes associated with
occupational exposure to inorganic lead. Arch Intern Med 143: 220-224.
http://dx.doi.0rg/10.1001/archinte.143.2.220
Robinson. G: Baumann. S: Kleinbaum. D; Barton. C: Schroeder. S: Mushak. P; Otto. D. (1985). Effects
of low to moderate lead exposure on brainstem auditory evoked potentials in children. In
Neurobehavioral methods in occupational and environmental health: extended abstracts from the
second international symposium; August; Copenhagen, Denmark. Copenhagen, Denmark: World
Health Organization.
Rodriguez-Iturbe. B; Sindhu. RK; Quiroz. Y; Vaziri. ND. (2005). Chronic exposure to low doses of lead
results in renal infiltration of immune cells, NF-kappaB activation, and overexpression of
tubulointerstitial angiotensin II. Antioxid Redox Signal 7: 1269-1274.
http://dx.doi.0rg/10.1089/ars.2005.7.1269
Rodriguez-Iturbe. B; Vaziri. ND; Herrera-Acosta. J; Johnson. RJ. (2004). Oxidative stress, renal
infiltration of immune cells, and salt-sensitive hypertension: All for one and one for all [Review]. Am
J Physiol 286: F606-F616. http://dx.doi.org/10.1152/aiprenal.00269.2003
Roels. H; Lauwerys. R; Konings. J; Buchet. JP; Bernard. A; Green. S; Bradley. D; Morgan. W; Chettle.
EX (1994). Renal function and hyperfiltration capacity in lead smelter workers with high bone lead.
Occup Environ Med 51: 505-512. http://dx.doi.Org/10.1136/oem.51.8.505
Romeo. R; Aprea. C; Boccalon. P; OrsiD; Porcelli. B; Sartorelli. P. (1996). Serum erthropoietin and blood
lead concentrations. Int Arch Occup Environ Health 69: 73-75.
Roncal. C; Mu. W; Reungjui. S; Kim. KM; Henderson. GN; Ouyang. X; Nakagawa. T; Johnson. RJ.
(2007). Lead, at low levels, accelerates arteriolopathy and tubulointerstitial injury in chronic kidney
disease. Am J Physiol Renal Physiol 293: F1391-F1396.
http://dx.doi.org/10.1152/ajprenal.00216.2007
Rondo. PH; Carvalho. MDF; Souza. MC; Moraes. F. (2006). Lead, hemoglobin, zinc protoporphyrin and
ferritin concentrations in children. Rev Saude Publica 40: 71-76.
http://dx.doi.org/10.1590/S0034-89102006000100012
November 2012 5-776 Draft - Do Not Cite or Quote
-------
Ronis. MJ; Badger. TM; Shema. SJ: Roberson. PK; Shaikh. F. (1998a). Effects on pubertal growth and
reproduction in rats exposed to lead perinatally or continuously throughout development. J Toxicol
Environ Health A 53: 327-341. http://dx.doi.org/10.1080/009841098159312
Ronis. MJJ; Aronson. J; Gao. GG: Hogue. W: Skinner. RA; Badger. TM; Lumpkin. CK. Jr. (2001).
Skeletal effects of developmental lead exposure in rats. Toxicol Sci 62: 321-329.
http://dx.doi.0rg/10.1093/toxsci/62.2.321
Ronis. MJJ: Badger. TM: Shema. SJ: Roberson. PK: Shaikh. F. (1996). Reproductive toxicity and growth
effects in rats exposed to lead at different periods during development. Toxicol Appl Pharmacol 136:
361-371. http://dx.doi.org/10.1006/taap.1996.0044
Ronis. MJJ: Gandv. J: Badger. T. (1998b). Endocrine mechanisms underlying reproductive toxicity in the
developing rat chronically exposed to dietary lead. J Toxicol Environ Health A 54: 77-99.
http://dx.doi.org/10.1080/009841098158935
Roshan. VD; Assali. M; Moghaddam. AH; Hosseinzadeh. M; Myers. J. (2011). Exercise training and
antioxidants: effects on rat heart tissue exposed to lead acetate. Int J Toxicol 30: 190-196.
http://dx.doi.org/10.1177/1091581810392809
Rossi-George. A; Virgolini. M; Weston. D; Thiruchelvam. M; Corv-Slechta. D. (2011). Interactions of
lifetime lead exposure and stress: Behavioral, neurochemical and HPA axis effects. Neurotoxicology
32: 83-99. http://dx.doi.Org/10.1016/i.neuro.2010.09.004
Rossi-George. A; Virgolini. MB; Weston. D; Corv-Slechta. DA. (2009). Alterations in glucocorticoid
negative feedback following maternal Pb, prenatal stress and the combination: A potential biological
unifying mechanism for their corresponding disease profiles. Toxicol Appl Pharmacol 234: 117-127.
http://dx.doi.0rg/10.1016/i.taap.2008.10.003
Rothenberg. SJ: Poblano. A; Garza-Morales. S. (1994b). Prenatal and perinatal low level lead exposure
alters brainstem auditory evoked responses in infants. Neurotoxicology 15: 695-699.
Rothenberg. SJ: Poblano. A; Schnaas. L. (2000). Brainstem auditory evoked response at five years and
prenatal and postnatal blood lead. Neurotoxicol Teratol 22: 503-510.
http://dx.doi.org/10.1016/S0892-0362(00)00079-9
Rothenberg. SJ: Rothenberg. JC. (2005). Testing the dose-response specification in epidemiology: Public
health and policy consequences for lead. Environ Health Perspect 113: 1190-1195.
http://dx.doi.org/10.1289/ehp.7691
Rothenberg. SJ: Schnaas. L; Salgado-Valladares. M; Casanueva. E; Geller. AM; Hudnell. HK; Fox. DA.
(2002b). Increased ERG a- and b-wave amplitudes in 7- to 10-year-old children resulting from
prenatal lead exposure. Invest Ophthalmol Vis Sci 43: 2036-2044.
Rotruck. JT; Pope. AL; Ganther. HE; Swanson. AB; Hafeman. DG: Hoekstra. WG. (1973). Selenium:
Biochemical role as a component of glutathione peroxidase. Science 179: 588-590.
http://dx.doi.org/10.1126/science.179.4073.588
Rousseau. MC: Parent. ME; Nadon. L; Latrellie. B; Siemiatvcki. J. (2007). Occupational exposure to lead
compounds and risk of cancer among men: A population-based case-control study. Am J Epidemiol
166: 1005-1014. http://dx.doi.org/10.1093/aie/kwml83
Rousseau. MC: Straif. K; Siemiatvcki. J. (2005). IARC carcinogen update [Letter]. Environ Health
Perspect 113: A580-A581. http://dx.doi.org/10.1289/ehp.113-a580
Roy. A; Bellinger. D; Hu. H; Schwartz. J: Ettinger. AS: Wright. RO: Bouchard. M; Palaniappan. K;
Balakrishnan. K. (2009a). Lead exposure and behavior among young children in Chennai, India.
Environ Health Perspect 117: 1607-1611. http://dx.doi.org/10.1289/ehp.0900625
Roy. A; Hu. H; Bellinger. DC: Mukherjee. B; Modali. R; Nasaruddin. K; Schwartz. J: Wright. RO:
Ettinger. AS: Palaniapan. K; Balakrishnan. K. (2011). Hemoglobin, lead exposure, and intelligence
quotient: Effect modification by the DRD2 Taq IA polymorphism. Environ Health Perspect 119:
144-149. http://dx.doi.org/10.1289/ehp.0901878
November 2012 5-777 Draft - Do Not Cite or Quote
-------
Roy. AA; Baxla. SP; Gupta. T; Bandvopadhyava. R; Tripathi. SN. (2009b). Particles emitted from indoor
combustion sources: Size distribution measurement and chemical analysis. Inhal Toxicol 21: 837-848.
http://dx.doi.org/10.1080/08958370802538050
Roy. AK; Dhir. H; Sharma. A. (1992). Modification of metal-induced micronuclei formation in mouse
bone marrow erythrocytes by Phyllanthus fruit extract and ascorbic acid. Toxicol Lett 62: 9-17.
http://dx.doi.org/10.1016/0378-4274(92)90072-R
Roy. NK; Rossman. TG. (1992). Mutagenesis and comutagenesis by lead compounds. Mutat Res 298:
97-103. http://dx.doi.org/10.1016/0165-1218(92)90034-W
Rubio. J: Riqueros. MI; Gasco. M; Yucra. S: Miranda. S: Gonzales. GF. (2006). Lepidium meyenii
(Maca) reversed the lead acetate induced - Damage on reproductive function in male rats. Food Chem
Toxicol 44: 1114-1122. http://dx.doi.Org/10.1016/i.fct.2006.01.007
Rybicki. BA; Johnson. CC: Uman. J; Gorell. JM. (1993). Parkinson's disease mortality and the industrial
use of heavy metals in Michigan. Mov Disord 8: 87-92. http://dx.doi.org/10.1002/mds.870080116
Sa. I; da Costa. MJP; Cunha. EM. (2012). Lead hepatotoxicology: A study in an animal model. Toxicol
Ind Health 28: 108-113. http://dx.doi.org/10.1177/0748233711407240
Sabbioni. E; Marafante. E. (1976). Identification of lead-binding components in rat liver: In vivo study.
Chem Biol Interact 15: 1-20. http://dx.doi.org/10.1016/0009-2797(76)90124-l
Saimi. Y; Kung. C. (2002). Calmodulin as an ion channel subunit [Review]. Annu Rev Physiol 64:
289-311. http://dx.doi.org/10.1146/annurev.phvsiol.64.100301.111649
Sainath. SB; Meena. R; Supriya. C. h; Reddv. KP; Reddv. PS. (2011). Protective role of Centella asiatica
on lead-induced oxidative stress and suppressed reproductive health in male rats. Environ Toxicol
Pharmacol32: 146-154. http://dx.doi.Org/10.1016/i.etap.2011.04.005
Sakai. T; Morita. Y. (1996). delta-Aminolevulinic acid in plasma or whole blood as a sensitive indicator
of lead effects, and its relation to the other heme-related parameters. Int Arch Occup Environ Health
68: 126-132. http://dx.doi.org/10.1007/BF00381245
Sakata. S: Shimizu. S: Ogoshi. K; Hirai. K; Ohno. Y; Kishi. T; Sherchand. JB; Utsumi. M; Shibata. M;
Takaki. M; Ueda. M; Mori. I. (2007). Inverse relationship between serum erythropoietin and blood
lead concentrations in Kathmandu tricycle taxi drivers. Int Arch Occup Environ Health 80: 342-345.
http://dx.doi.org/10.1007/s00420-006-0125-4
Sakuma. H; Kusama. M; Yamaguchi. K; Matsuki. T; Sugawara. S. (1984). The distribution of cigarette
smoke components between mainstream and sidestream smoke: II. Bases. Beitraege zur
Tabakforschung International 12: 199-209.
Salawu. EO; Adeevo. OA; Falokun. OP; Yusuf. UA; Oyerinde. A; Adeleke. AA. (2009). Tomato
(Lycopersicon esculentum) prevents lead-induced testicular toxicity. Journal of Human Reproductive
Sciences 2: 30-34. http://dx.doi.org/10.4103/0974-1208.51346
Saldiva. PHN; Clarke. RW: Coull. BA; Stearns. RC: Lawrence. J: Murthy. GGK; Diaz. E; Koutrakis. P;
Suh. H; Tsuda. A; Godleski. JJ. (2002). Lung inflammation induced by concentrated ambient air
particles is related to particle composition. Am J Respir Crit Care Med 165: 1610-1617.
http://dx.doi.org/10.1164/rccm.2106102
Saleh. M; Mathison. JC: Wolinski. MK; Bensinger. SJ: Fitzgerald. P; Droin. N; Ulevitch. RJ; Green. PR;
Nicholson. DW. (2006). Enhanced bacterial clearance and sepsis resistance in caspase-12-deficient
mice. Nature 440: 1064-1068. http://dx.doi.org/10.1038/nature04656
Sanchez-Fructuoso. AI; Blanco. J: Cano. M; Ortega. L; Arroyo. M; Fernandez. C: Prats. D; Barrientos. A.
(2002a). Experimental lead nephropathy: Treatment with calcium disodium
ethylenediaminetetraacetate. Am J Kidney Dis 40: 59-67. http://dx.doi.org/10.1053/ajkd.2002.33936
Sanchez-Fructuoso. AI; Cano. M; Arroyo. M; Fernandez. C: Prats. D; Barrientos. A. (2002b). Lead
mobilization during calcium disodium ethylenediaminetetraacetate chelation therapy in treatment of
chronic lead poisoning. Am J Kidney Dis 40: 51-58. http://dx.doi.org/10.1053/aikd.2002.33913
November 2012 5-778 Draft - Do Not Cite or Quote
-------
Sandercock. GRH; Brodie. DA. (2006). The role of heart rate variability in prognosis for different modes
of death in chronic heart failure [Review]. Pacing Clin Electrophysiol 29: 892-904.
http://dx.doi.0rg/10.llll/i.1540-8159.2006.00457.x
Sandhir. R; Gill. KD. (1995). Effect of lead on lipid peroxidation in liver of rats. Biol Trace Elem Res 48:
91-97. http://dx.doi.org/10.1007/BF02789081
Sandhir. R; Julka. D; Gill. KD. (1994). Lipoperoxidative damage on lead exposure in rat brain and its
implications on membrane bound enzymes. Pharmacol Toxicol 74: 66-71.
http://dx.doi.0rg/10.llll/i.1600-0773.1994.tb01077.x
Sanna. E; Vallascas. E. (2011). Hair lead levels to evaluate the subclinical impact of lead on growth in
Sardinian children (Italy). Am J Hum Biol 23: 740-746. http://dx.doi.org/10.1002/ajhb.21203
Santibanez. M; Vioque. J: Alguacil. J: Barber. X; de la Hera. G: Kauppinen. T. (2008). Occupational
exposures and risk of oesophageal cancer by histological type: a case-control study in eastern Spain.
Occup Environ Med 65: 774-781. http://dx.doi.org/10.1136/oem.2007.037929
Santos. MRV; Marchioro. M; Antoniolli. AR. (2006). Lead effects on non-adrenergic non-cholinergic
relaxations in the rat gastric fundus. Toxicol In Vitro 20: 38-42.
http://dx.doi.0rg/10.1016/i.tiv.2005.05.005
Saraiva. MCP; Taichman. RS: Braun. T; Nriagu. J: Eklund. SA; Burt. BA. (2007). Lead exposure and
periodontitis in US adults. J Periodontal Res 42: 45-52.
http://dx.doi.0rg/10.llll/i.1600-0765.2006.00913.x
Sarasua. SM; Mueller. P; Kathman. S: Campagna. D; Uddin. MS: White. MC. (2003). Confirming the
utility of four kidney biomarker tests in a longitudinal follow-up study. Ren Fail 25: 797-817.
Sarasua. SM; Vogt RF; Henderson. LO; Jones. PA; Lybarger. JA. (2000). Serum immunoglobulins and
lymphocyte subset distributions in children and adults living in communities assessed for lead and
cadmium exposure. J Toxicol Environ Health A 60: 1-15. http://dx.doi.org/10.1080/009841000156556
Sarnat. JA; Marmur. A; Klein. M; Kim. E; Russell. AG: Sarnat. SB; Mulholland. JA; Hopke. PK; Tolbert.
PE. (2008). Fine particle sources and cardiorespiratory morbidity: An application of chemical mass
balance and factor analytical source-apportionment methods. Environ Health Perspect 116: 459-466.
http://dx.doi.org/10.1289/ehp.10873
Saverv. LC: Grlickova-Duzevik. E; Wise. SS: Thompson. WD; Hinz. JA: Thompson. LH; Wise. JP, Sr.
(2007). Role of the Fancg gene in protecting cells from paniculate chromate-induced chromosome
instability. Mutat Res Genet Toxicol Environ Mutagen 626: 120-127.
http://dx.doi.0rg/10.1016/i.mrgentox.2006.09.005
Schanne. FAX; Dowd. TL; Gupta. RK; Rosen. JF. (1989). Lead increases free Ca2+ concentration in
cultured osteoblastic bone cells: simultaneous detection of intracellular free Pb2+ by 19F NMR.
PNAS 86: 5133-5135.
Schanne. FAX; Long. GJ: Rosen. JF. (1997). Lead induced rise in intracellular free calcium is mediated
through activation of protein kinase C in osteoblastic bone cells. Biochim Biophys Acta 136: 247-254.
http://dx.doi.org/10.1016/S0925-4439(97)00006-9
Schell. LM; Denham. M; Stark. AD; Gomez. M; Ravenscroft. J: Parsons. PJ: Aydermir. A; Samelson. R.
(2003). Maternal blood lead concentration, diet during pregnancy, and anthropometry predict neonatal
blood lead in a socioeconomically disadvantaged population. Environ Health Perspect 111: 195-200.
http://dx.doi.org/10.1289/ehp.5592
Schell. LM; Denham. M; Stark. AD; Parsons. PJ: Schulte. EE. (2009). Growth of infants' length, weight,
head and arm circumferences in relation to low levels of blood lead measured serially. Am J Hum Biol
21: 180-187. http://dx.doi.org/10.1002/aihb.20842
Schirrmacher. K; Wiemann. M; Bingmann. D; Busselberg. D. (1998). Effects of lead, mercury, and
methyl mercury on gap junctions and [CA2+]i in bone cells. Calcif Tissue Int 63: 134-139.
http://dx.doi.org/10.1007/s002239900503
November 2012 5-779 Draft - Do Not Cite or Quote
-------
Schmitt. CJ: Whyte. JJ: Brumbaugh. WG: Tillitt DE. (2005). Biochemical effects of lead, zinc, and
cadmium from mining on fish in the Tri-States District of northeastern Oklahoma, USA. Environ
Toxicol Chem 24: 1483-1495. http://dx.doi.Org/10.1897/04-332R.l
Schmitt. CJ: Whyte. JJ: Roberts. AP; Annis. ML; May. TW: Tilitt. DE. (2007b). Biomarkers of metals
exposure in fish from lead-zinc mining areas of southeastern Missouri, USA. Ecotoxicol Environ Saf
67: 31-47. http://dx.doi.Org/10.1016/i.ecoenv.2006.12.011
Schnaas. L; Rothenberg. SJ: Flores. MF; Martinez. S: Hernandez. C: Osorio. E; Velasco. SR; Perroni. E.
(2006). Reduced intellectual development in children with prenatal lead exposure. Environ Health
Perspect 114: 791-797. http://dx.doi.org/10.1289/ehp.8552
Schnaas. L; Rothenberg. SJ: Perroni. E; Martinez. S: Hernandez. C: Hernandez. RM. (2000). Temporal
pattern in the effect of postnatal blood lead level on intellectual development of young children.
Neurotoxicol Teratol 22: 805-810. http://dx.doi.org/10.1016/S0892-0362(00)00101-X
Schneider. JS: Anderson. DW: Wade. TV; Smith. MG: Leibrandt. P; Zuck. L; Lidsky. TI. (2005).
Inhibition of progenitor cell proliferation in the dentate gyrus of rats following post-weaning lead
exposure. Neurotoxicology 26: 141-145. http://dx.doi.Org/10.1016/j.neuro.2004.06.006
Schober. SE; Mirel. LB; Graubard. BI; Brodv. DJ: Flegal. KM. (2006). Blood lead levels and death from
all causes, cardiovascular disease, and cancer: Results from the NHANES III Mortality Study. Environ
Health Perspect 114: 1538-1541. http://dx.doi.org/10.1289/ehp.9123
Schrauzer. GN. (1987). Effects of selenium antagonists on cancer susceptibility: New aspects of chronic
heavy metal toxicity. J UOEH 9: 208-215.
Schrauzer. GN. (2008). Effects of selenium and low levels of lead on mammary tumor development and
growth in MMTV-infected female mice. Biol Trace Elem Res 125: 268-275.
http://dx.doi.org/10.1007/sl2011-008-8172-l
Schwark. WS: Haluska. M; Blackshear. P; Magana. T. (1985). Lifetime lead intoxication: influence on the
amygdaloid kindling model of epileptogenesis. Toxicology 36: 49-60.
http://dx.doi.org/10.1016/0300-483X(85)90006-X
Schwartz. BS: Chen. S: Caffo. B; Stewart. WF; Bolla. KI; Yousem. D; Davatzikos. C. (2007). Relations
of brain volumes with cognitive function in males 45 years and older with past lead exposure.
Neuroimage 37: 633-641. http://dx.doi.Org/10.1016/j.neuroimage.2007.05.035
Schwartz. BS: Lee. BK; Bandeen-Roche. K; Stewart. W: Bolla. K; Links. J: Weaver. V; Todd. A. (2005).
Occupational lead exposure and longitudinal decline in neurobehavioral test scores. Epidemiology 16:
106-113. http://dx.doi.org/10.1097/01.ede.0000147109.62324.51
Schwartz. BS: Lee. BK; Lee. GS: Stewart. WF; Lee. SS: Hwang. KY; Ahn. KD; Kim. YB; Bolla. KI;
Simon. D; Parsons. PJ; Todd. AC. (2001). Associations of blood lead, dimercaptosuccinic acid-
chelatable lead, and tibia lead with neurobehavioral test scores in South Korean lead workers. Am J
Epidemiol 153: 453-464. http://dx.doi.Org/10.1093/aie/153.5.453
Schwartz. BS; Lee. BK; Lee. GS; Stewart. WF; Simon. D; Kelsev. K; Todd. AC. (2000a). Associations of
blood lead, dimercaptosuccinic acid-chelatable lead, and tibia lead with polymorphisms in the vitamin
D receptor and delta-aminolevulinic acid dehydratase genes. Environ Health Perspect 108: 949-954.
http://dx.doi.org/10.2307/3435053
Schwartz. BS; Lee. BK; Stewart. W; Ahn. KD; Kelsev. K; Bresssler. J. (1997a). Associations of subtypes
of hemoglobin with delta-aminolevulinic acid dehydratase genotype and dimercaptosuccinic acid-
chelatable lead levels. Arch Environ Occup Health 52: 97-103.
http://dx.doi.org/10.1080/00039899709602871
Schwartz. BS; Lee. BK; Stewart. W; Sithisarankul. P; Strickland. PT; Ahn. KD; Kelsev. K. (1997b).
delta-Aminolevulinic acid dehydratase genotype modifies four hour urinary lead excretion after oral
administration of dimercaptosuccinic acid. Occup Environ Med 54: 241-246.
November 2012 5-780 Draft - Do Not Cite or Quote
-------
Schwartz. J. (1991). Lead, blood pressure, and cardiovascular disease in men and women. Environ Health
Perspect 91: 71-75. http://dx.doi.org/10.1289/ehp.919171
Schwartz. J. (1994). Low-level lead exposure and children's IQ: A meta-analysis and search for a
threshold. Environ Res 65: 42-55. http://dx.doi.org/10.1006/enrs.1994.1020
Schwartz. J: Landrigan. PJ; Baker. EL. Jr; Orenstein. WA; von Lindern. IH. (1990). Lead-induced
anemia: dose-response relationships and evidence for a threshold. Am J Public Health 80: 165-168.
Schwartz. J: Otto. D. (1987). Blood lead, hearing thresholds, and neurobehavioral development in
children and youth. Arch Environ Occup Health 42: 153-160.
http://dx.doi.org/10.1080/00039896.1987.9935814
Schwartz. J: Otto. D. (1991). Lead and minor hearing impairment. Arch Environ Occup Health 46:
300-305. http://dx.doi.org/10.1080/00039896.1991.9934391
Sciarillo. WG: Alexander. G: Farrell. KP (1992). Lead exposure and child behavior. Am J Public Health
82: 1356-1360.
Scinicariello. F; Murray. HE; Moffett. DB; Abadin. HG: Sexton. MJ; Fowler. BA. (2007). Lead and delta-
aminolevulinic acid dehydratase polymorphism: Where does it lead? A meta-analysis. Environ Health
Perspect 115: 35-41. http://dx.doi.org/10.1289/ehp.9448
Scinicariello. F; Yesupriya. A; Chang. MH; Fowler. BA. (2010). Modification by ALAD of the
association between blood lead and blood pressure in the U.S. population: Results from the Third
National Health and Nutrition Examination Survey. Environ Health Perspect 118: 259-264.
http://dx.doi.org/10.1289/ehp.0900866
Seki. T; Arai. Y. (1993). Highly polysialylated NCAM expression in the developing and adult rat spinal
cord [Letter]. Dev Brain Res 73: 141-145. httrj://dx.doi.org/10.1016/0165-3806(93)90056-G
Selevan. SG: Rice. DC: Hogan. KA; Euling. SY; Pfahles-Hutchens. A; Bethel. J. (2003). Blood lead
concentration and delayed puberty in girls. N Engl J Med 348: 1527-1536.
http://dx.doi.org/10.1056/NEJMoa020880
Selvin-Testa. A; Lopez-Costa. JJ: Nessi-de Avinon. AC: Saavedra. JP (1991). Astroglial alterations in rat
hippocampus during chronic lead exposure. Glia 4: 384-392. http://dx.doi.org/10.1002/glia.440040406
Selye. H; Tuchweber. B; Bertok. L. (1966). Effect of lead acetate on the susceptibility of rats to bacterial
endotoxins. J Bacteriol 91: 884-890.
Senger. MR; Rico. EP; Arizi. MDB; Frazzon. APG: Dias. RD; Bogo. MR; Bonan. CD. (2006). Exposure
to Hg2+ and Pb2+ changes NTPDase and ecto-5 '-nucleotidase activities in central nervous system of
zebrafish (Danio rerio). Toxicology 226: 229-237. http://dx.doi.Org/10.1016/j.tox.2006.07.012
Shabani. A; Rabbani. A. (2000). Lead nitrate induced apoptosis in alveolar macrophages from rat lung.
Toxicology 149: 109-114. http://dx.doi.org/10.1016/S0300-483X(00)00232-8
Shadick. NA; Kim. R; Weiss. S: Liang. MH; Sparrow. D: Hu. H. (2000). Effect of low level lead exposure
on hyperuricemia and gout among middle aged and elderly men: the normative aging study. J
Rheumatol27: 1708-1712.
Shah. F; Kazi. TG: Afridi. HI; Baig. JA; Khan. S: Kolachi. NF; Wadhwa. SK; Shah. AQ. (2010).
Environmental exposure of lead and iron deficit anemia in children age ranged 1-5 years: A cross
sectional study. Sci Total Environ 408: 5325-5330. http://dx.doi.Org/10.1016/i.scitotenv.2010.07.091
Shaik. AP; Jamil. K. (2009). Individual susceptibility and genotoxicity in workers exposed to hazardous
materials like lead. J Hazard Mater 168: 918-924. http://dx.doi.Org/10.1016/i.jhazmat.2009.02.129
Shakoor. A; Gupta. PK; Singh. YP; Kataria. M. (2000). Beneficial effects of aluminum on the progression
of lead-induced nephropathy in rats. Basic Clin Pharmacol Toxicol 87: 258-260.
http://dx.doi.0rg/10.1034/i.1600-0773.2000.pto870603.x
November 2012 5-781 Draft - Do Not Cite or Quote
-------
Shan. G: Tang. T; Zhang. X. (2009). The protective effect of ascorbic acid and thiamine supplementation
against damage caused by lead in the testes of mice. J Huazhong Univ Sci Technolog Med Sci 29:
68-72. http://dx.doi.org/10.1007/sll596-009-0114-4
Sharifi. AM; Darabi. R; Akbarloo. N; Larijani. B; Khoshbaten. A. (2004). Investigation of circulatory and
tissue ACE activity during development of lead-induced hypertension. Toxicol Lett 153: 233-238.
http://dx.doi.0rg/10.1016/i.toxlet.2004.04.013
Sharifi. AM; Mousavi. SH. (2008). Studying the effects of lead on DNA fragmentation and proapoptotic
bax and antiapoptotic Bcl-2 protein expression in PC12 cells. Toxicol Mech Meth 18: 75-79.
http://dx.doi.org/10.1080/15376510701665814
Sharifi. AM; Mousavi. SH; Jorjani. M. (2010). Effect of chronic lead exposure on pro-apoptotic Bax and
anti-apoptotic Bcl-2 protein expression in rat hippocampus in vivo. Cell Mol Neurobiol 30: 769-774.
http://dx.doi.org/10.1007/sl0571-010-9504-l
Sharma. A; Sharma. V: Kansal. L. (2010a). Amelioration of lead-induced hepatotoxicity by Allium
sativum extracts in Swiss albino mice. Libyan Journal of Medicine 5: 4621.
http://dx.doi.org/10.4176/091107
Sharma. V: Sharma. A; Kansal. L. (201 Ob). The effect of oral administration of Allium sativum extracts
on lead nitrate induced toxicity in male mice. Food Chem Toxicol 48: 928-936.
http://dx.doi.0rg/10.1016/i.fct.2010.01.002
Shelkovnikov. SA; Gonick. HC. (2001). Influence of lead on rat thoracic aorta contraction and relaxation.
Am J Hypertens 14: 873-878. http://dx.doi.org/10.1016/S0895-7061(01)02149-5
Shelton. KR: Cunningham. JG: Klann. E; Merchant. RE; Egle. PM; Bigbee. JW. (1990). Low-abundance
32-kilodalton nuclear protein specifically enriched in the central nervous system. J Neurosci Res 25:
287-294. http://dx.doi.org/10.1002/inr.490250304
Shelton. KR: Egle. PM. (1982). The proteins of lead-induced intranuclear inclusion bodies. J Biol Chem
257: 11802-11807.
Shih. RA; Glass. TA; Bandeen-Roche. K; Carlson. MC: Bolla. KI; Todd. AC: Schwartz. BS. (2006).
Environmental lead exposure and cognitive function in community-dwelling older adults. Neurology
67: 1556-1562. http://dx.doi.org/10.1212/01.wnl.0000239836.26142.c5
Shin. JH; Lim. KM; Noh. JY; Bae. ON: Chung. SM; Lee. MY; Chung. JH. (2007). Lead-induced
procoagulant activation of erythrocytes through phosphatidylserine exposure may lead to thrombotic
diseases. Chem Res Toxicol 20: 38-43. http://dx.doi.org/10.1021/tx060114+
Shinkai. Y; Yamamoto. C: Kaii. T. (2010). Lead induces the expression of ER chaperones GRP78 and
GRP94 in vascular endothelial cells via the JNK-AP-1 pathway. Toxicol Sci 114: 378-386.
http://dx.doi.org/10.1093/toxsci/kfq008
Shinozuka. H; Ohmura. T; Katval. SL; Zedda. AI; Ledda-Columbano. GM; Columbano. A. (1996).
Possible roles of nonparenchymal cells in hepatocyte proliferation induced by lead nitrate and by
tumor necrosis factor alpha. Hepatology 23: 1572-1577. http://dx.doi.org/10.1002/hep.510230638
Siddiqui. MKJ; Jyoti; Singh. S: Mehrotra. PK; Singh. K; Sarangi. R. (2006). Comparison of some trace
elements concentration in blood, tumor free breast and tumor tissues of women with benign and
malignant breast lesions: An Indian study. Environ Int 32: 630-637.
http://dx.doi.0rg/10.1016/i.envint.2006.02.002
Silberstein. T; Saphier. O; Paz-Tal. O: Trimarchi. JR; Gonzalez. L; Keefe. PL. (2006). Lead concentrates
in ovarian follicle compromises pregnancy. J Trace Elem Med Biol 20: 205-207.
http://dx.doi.0rg/10.1016/i.jtemb.2006.05.001
Silkin. YA; Silkina. EN; Sherstobitov. AO; Gusev. GP (2001). Activation of potassium channels in
erythrocytes of marine teleost Scorpaena porcus. Membr Cell Biol 14: 773-782.
November 2012 5-782 Draft - Do Not Cite or Quote
-------
Silva. PA; Hughes. P; Williams. S: Faed. JM. (1988). Blood lead, intelligence, reading attainment, and
behaviour in eleven year old children in Dunedin, New Zealand. J Child Psychol Psychiatry 29: 43-52.
http://dx.doi.0rg/10.llll/i.1469-7610.1988.tb00687.x
Silveira. EA; Lizardo. JH; Souza. LP; Stefanon. I; Vassallo. DV. (2010). Acute lead-induced
vasoconstriction in the vascular beds of isolated perfused rat tails is endothelium-dependent. Braz J
Med Biol Res 43: 492-499. http://dx.doi.org/10.1590/S0100-879X2010007500027
Simoes. MR; Ribeiro Junior. RF; Vescovi. MV; de Jesus. HC: Padilha. AS: Stefanon. I; Vassallo. DV:
Salaices. M; Fioresi. M. (2011). Acute lead exposure increases arterial pressure: role of the Renin-
Angiotensin system. PLoS ONE 6: el8730. http://dx.doi.org/10.1371/iournal.pone.0018730
Simons. TJB. (1986). Passive transport and binding of lead by human red blood cells. J Physiol 378:
267-286.
Simons. TJB. (1993a). Lead-calcium interactions in cellular lead toxicity [Review]. Neurotoxicology 14:
77-85.
Simons. TJB. (1993b). Lead transport and binding by human erythrocytes in vitro. Pflugers Arch 423:
307-313. http://dx.doi.org/10.1007/BF00374410
Simons. TJB. (1995). The affinity of human erythrocyte porphobilinogen synthase for Zn2+ and Pb2+.
FEES J 234: 178-183. http://dx.doi.Org/10.llll/i.1432-1033.1995.178 c.x
Simsek. N; Karadeniz. A; Kalkan. Y; Keles. ON; Unal. B. (2009). Spirulina platensis feeding inhibited
the anemia- and leucopenia-induced lead and cadmium in rats. J Hazard Mater 164: 1304-1309.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.09.041
Singh. A; Cullen. C: Dykeman. A; Rice. D; Foster. W. (1993a). Chronic lead exposure induces
ultrastructural alterations in the monkey testis. J Submicrosc Cytol Pathol 25: 479-486.
Singh. B; Chandran. V; Bandhu. HK; Mittal. BR; Bhattacharya. A; Jindal. SK; Varma. S. (2000). Impact
of lead exposure on pituitary-thyroid axis in humans. Biometals 13: 187-192.
http://dx.doi.Org/10.1023/A:1009201426184
Singh. J; Pritchard. DE; Carlisle. PL; Mclean. JA; Montaser. A; Orenstein. JM; Patierno. SR. (1999).
Internalization of carcinogenic lead chromate particles by cultured normal human lung epithelial cells:
Formation of intracellular lead-inclusion bodies and induction of apoptosis. Toxicol Appl Pharmacol
161: 240-248. http://dx.doi.org/10.1006/taap.1999.8816
Singh. US: Saxena. DK; Singh. C: Murthy. RC: Chandra. SV. (1991). Lead-induced fetal nephrotoxicity
in iron-deficient rats. Reprod Toxicol 5:211-217. http://dx.doi.org/10.1016/0890-6238(91)90053-I
Sivaprasad. R; Nagarai. M; Varalakshmi. P. (2003). Combined efficacies of lipoic acid and meso-2,3-
dimercaptosuccinic acid on lead-induced erythrocyte membrane lipid peroxidation and antioxidant
status in rats. Hum Exp Toxicol 22: 183-192. http://dx.doi.org/10.1191/0960327103ht335oa
Skoczynska. A; Stojek. E. (2005). The impact of subchronic lead poisoning on the vascular effect of nitric
oxide in rats. Environ Toxicol Pharmacol 19: 99-106. http://dx.doi.Org/10.1016/j.etap.2004.05.004
Slivkova. J: Popelkova. M; Massanyi. P; Toporcerova. S: Stawarz. R; Formicki. G: Lukac. N; Putaa. A;
Guzik. M. (2009). Concentration of trace elements in human semen and relation to spermatozoa
quality. J Environ Sci Health A Tox Hazard Subst Environ Eng 44: 370-375.
http://dx.doi.org/10.1080/10934520802659729
Smejkalova. J. (1990). The chromosomal aberrations investigation in children permanently living in the
lead polluted area. Sb Ved Pr Fak Karlovy Univ Hradci Kralovea 33: 539-564.
Smith. CM; Hu. H; Wang. X; Kelsev. KT. (1995a). ALA-D genotype is not associated with HT or HB
levels among workers exposed to low levels of lead. Med Lav 86: 229-235.
Smith. CM; Wang. X; Hu. H; Kelsev. KT. (1995b). A polymorphism in the delta-aminolevulinic acid
dehydratase gene may modify the pharmacokinetics and toxicity of lead. Environ Health Perspect 103:
248-253. http://dx.doi.org/10.1289/ehp.95103248
November 2012 5-783 Draft - Do Not Cite or Quote
-------
Smith. PR; Kahng. MW: Quintanilla-Vega. B; Fowler. BA. (1998). High-affinity renal lead-binding
proteins in environmentally-exposed humans. Chem Biol Interact 115: 39-52.
http://dx.doi.org/10.1016/S0009-2797(98)00060-X
Snyder. JE; Filipov. NM; Parsons. PJ: Lawrence. DA. (2000). The efficiency of maternal transfer of lead
and its influence on plasma IgE and splenic cellularity of mice. Toxicol Sci 57: 87-94.
http://dx.doi.0rg/10.1093/toxsci/57.l.87
Snyder. RD; Lachmann. PJ. (1989). Thiol involvement in the inhibition of DNA repair by metals in
mammalian cells. Mol Toxicol 2: 117-128.
Sobekova. A; Holovska. K; Lenartova. V: Legatfa. J: Javorsky. P. (2009). The alteration of glutathione
peroxidase activity in rat organs after lead exposure. Acta Physiol Hung 96: 37-44.
http://dx.doi.Org/10.1556/APhysiol.96.2009.l.4
Sokol. RZ. (1987). Hormonal effects of lead acetate in the male rat: mechanism of action. Biol Reprod
37: 1135-1138. http://dx.doi.0rg/10.1095/biolreprod37.5.1135
Sokol. RZ. (1989). Reversibility of the toxic effect of lead on the male reproductive axis. Reprod Toxicol
3: 175-180.
Sokol. RZ; Berman. N. (1991). The effect of age of exposure on lead-induced testicular toxicity.
Toxicology 69: 269-278. http://dx.doi.org/10.1016/0300-483X(91)90186-5
Sokol. RZ; Madding. CE; Swerdloff. RS. (1985). Lead toxicity and the hypothalamic-pituitary-testicular
axis. Biol Reprod 33: 722-728. http://dx.doi.org/10.1095/biolreprod33.3.722
Sokol. RZ; Okuda. H; Nagler. HM; Berman. N. (1994). Lead exposure in vivo alters the fertility potential
of sperm in vitro. Toxicol Appl Pharmacol 124: 310-316.
Solliwav. BM; Schaffer. A; Pratt. H; Yannai. S. (1996). Effects of exposure to lead on selected
biochemical and haematological variables. Basic Clin Pharmacol Toxicol 78: 18-22.
Solon. O; Riddell. TJ; Quimbo. SA; Butrick. E; Aylward. GP; Lou Bacate. M; Peabodv. JW. (2008).
Associations between cognitive function, blood lead concentration, and nutrition among children in
the central Philippines. J Pediatr 152: 237-243. http://dx.doi.Org/10.1016/i.jpeds.2007.09.008
Songdej. N; Winters. PC: McCabe. MJ. Jr; van Wijngaarden. E. (2010). A population-based assessment of
blood lead levels in relation to inflammation. Environ Res 110: 272-277.
http://dx.doi.0rg/10.1016/i.envres.2009.12.008
Spector. JT; Navas-Acien. A; Fadrowski. J: Guallar. E; Jaar. B; Weaver. VM. (2011). Associations of
blood lead with estimated glomerular filtration rate using MDRD, CKD-EPI and serum cystatin C-
based equations. Nephrol Dial Transplant 26: 2786-2792. http://dx.doi.org/10.1093/ndt/gfq773
SRC (Syracuse Research Corporation). (2002). Baseline human health risk assessment: Eureka Mills -
Eureka, Utah. Denver, CO: U.S. Environmental Protection Agency.
http://www.epa.gov/region8/r8risk/pdf/hhra_eureka.pdf
Srivastava. D; Hurwitz. RL; Fox. DA. (1995). Lead- and calcium-mediated inhibition of bovine rod
cGMPphosphodiesterase: Interactions with magnesium. Toxicol Appl Pharmacol 134: 43-52.
http://dx.doi.org/10.1006/taap.1995.1167
Srivastava. V: Dearth. RK; Hinev. JK; Ramirez. LM; Bratton. GR; Dees. W. (2004). The effects of
low-level Pb on steroidogenic acute regulatory protein (StAR) in the prepubertal rat ovary. Toxicol Sci
77: 35-40. http://dx.doi.org/10.1093/toxsci/kfg249
Stacchiotti. A; Morandini. F; Bettoni. F; Schena. I; Lavazza. A; Grigolato. PG: Apostoli. P; Rezzani. R;
Aleo. MF. (2009). Stress proteins and oxidative, damage in a renal derived cell line exposed to
inorganic mercury and lead. Toxicology 264: 215-224. http://dx.doi.Org/10.1016/j.tox.2009.08.014
November 2012 5-784 Draft - Do Not Cite or Quote
-------
Stackpole. MM; Wise. SS: Goodale. BC: Duzevik. EG: Munroe. RC: Thompson. WD; Thacker. J;
Thompson. LH; Hinz. JM; Wise. JP. (2007). Homologous recombination repair protects against
paniculate chromate-induced chromosome instability in Chinese hamster cells. Mutat Res-Fundam
Mol Mech Mutagen 625: 145-154. http://dx.doi.Org/10.1016/i.mrfmmm.2007.06.003
Staessen. JA; Lauwerys. RR; Buchet. JP: Bulpitt. CJ: Rondia. D; Van Renterghem. Y; Amery. A. (1992).
Impairment of renal function with increasing blood lead concentrations in the general population. N
EnglJ Med 327: 151-156. http://dx.doi.org/10.1056/NEJM199207163270303
Staessen. JA; Nawrot. T; Den Hond. E; Thijs. L; Fagard. R; Hoppenbrouwers. K; Koppen. G: Nelen. V;
Schoeters. G: Vanderschueren. D; Van Hecke. E; Verschaeve. L; Vlietinck. R; Roels. HA. (2001).
Renal function, cytogenetic measurements, and sexual developments in adolescents in relation to
environmental pollutants: Afeasibility study of biomarkers. Lancet 357: 1660-1669.
http://dx.doi.org/10.1016/S0140-6736(00)04822-4
Stangle. DE; Smith. PR; Beaudin. SA; Strawderman. MS: Levitsky. DA; Strupp. BJ. (2007). Succimer
chelation improves learning, attention, and arousal regulation in lead-exposed rats but produces lasting
cognitive impairment in the absence of lead exposure. Environ Health Perspect 115: 201-209.
http://dx.doi.org/10.1289/ehp.9263
Stayner. L; Steenland. K; Dosemeci. M; Hertz-Picciotto. I. (2003). Attenuation of exposure-response
curves in occupational cohort studies at high exposure levels. Scand J Work Environ Health 29:
317-324.
Steenland. NK; Thun. MJ; Ferguson. CW: Port. FK. (1990). Occupational and other exposures associated
with male end-stage renal disease: A case/control study. Am J Public Health 80: 153-157.
Steinthorsdottir. SD; Eliasdottir. SB; Indridason. OS: Agustsdottir. IM; Palsson. R; Edvardsson. VO.
(2011). Prevalence of hypertension in 9- to 10-year-old Icelandic school children. J Clin Hypertens
(Greenwich) 13: 774-779. http://dx.doi.Org/10.llll/j.1751-7176.2011.00496.x
Sterling. G: O'Neill. K; McCaffertv. M; O'Neill. J. (1982). Effect of chronic lead ingestion by rats on
glucose metabolism and acetylcholine synthesis in cerebral cortex slices. JNeurochem 39: 592-596.
http://dx.doi.org/10.1111/i.l471-4159.1982.tb03989.x
Stevens. LA; Coresh. J: Schmid. CH; Feldman. HI; Froissart. M; Kusek. J: Rossert. J: Van Lente. F;
Bruce. RD; Zhang. YP; Greene. T; Levey. AS. (2008). Estimating GFR using serum cystatin C alone
and in combination with serum creatinine: A pooled analysis of 3,418 individuals with CKD. Am J
Kidney Dis 51: 395-406. http://dx.doi.Org/10.1053/i.ajkd.2007.ll.018
Stewart. PW: Elaine. C: Cohen. M; Burright. RG: Donovick. PJ. (1996). Acute and longer term effects of
meso-2,3 dimercaptosuccinic acid (DMSA) on the behavior of lead-exposed and control mice. Physiol
Behav 59: 849-855. http://dx.doi.org/10.1016/0031-9384(95)02185-X
Stewart. WF; Schwartz. BS: Davatzikos. C: Shen. D; Liu. D; Wu. X; Todd. AC: Shi. W: Bassett. S:
Youssem. D. (2006). Past adult lead exposure is linked to neurodegeneration measured by brain MRI.
Neurology 66: 1476-1484. http://dx.doi.org/10.1212/01.wnl.0000216138.69777.15
Stewart. WF; Schwartz. BS: Simon. D; Kelsev. K; Todd. AC. (2002). ApoE genotype, past adult lead
exposure, and neurobehavioral function. Environ Health Perspect 110: 501-505.
http://dx.doi.org/10.1289/ehp.02110501
Stiles. KM; Bellinger. DC. (1993). Neuropsychological correlates of low-level lead exposure in school-
age children: a prospective study. Neurotoxicol Teratol 15: 27-35.
Stoleski. S: Karadzinska-Bislimovska. J: Stikova. E; Risteska-Kuc. S: Mijakoski. D; Minov. J. (2008).
Adverse effects in workers exposed to inorganic lead. Arh Hig Rada Toksikol 59: 19-29.
http://dx.doi.org/10.2478/10004-1254-59-2008-1866
Stromberg. U; Schiitz. A; Skerfving. S. (1995). Substantial decrease of blood lead in Swedish children,
1978-94, associated with petrol lead. Occup Environ Med 52: 764-769.
http://dx.doi.org/10.1136/oem.52.ll.764
November 2012 5-785 Draft - Do Not Cite or Quote
-------
Struzvnska. L; Dabrowska-Bouta. B; Koza. K; Sulkowski. G. (2007). Inflammation-like glial response in
lead-exposed immature rat brain. Toxicol Sci 95: 156-162. http://dx.doi.org/10.1093/toxsci/kfl 134
Struzynska. L; Dabrowska-Bouta. B; Rafalowska. U. (1997a). Acute lead toxicity and energy metabolism
in rat brain synaptosomes. Acta Neurobiol Exp (Wars) 57: 275-281.
Struzynska. L; Walski. M; Gadamski. R; Dabrowska-Bouta. B; Rafalowska. U. (1997b). Lead-induced
abnormalities in blood-brain barrier permeability in experimental chronic toxicity. Mol Chem
Neuropathol 31: 207-224. http://dx.doi.org/10.1007/BF02815125
Su. Y; Ding. Y; Jiang. M; Hu. X; Zhang. Z. (2007). Protein 4.2 Komatsu (D175Y) associated with the
lack of interaction with ankyrin in human red blood cells. Blood Cells Mol Dis 38: 221-228.
http://dx.doi.0rg/10.1016/i.bcmd.2006.ll.004
Sun. H; Wang. HL; Wang. S. (2007). D-serine relieves chronic lead exposure-impaired long-term
potentiation in the CA1 region of the rat hippocampus in vitro. Neurosci Lett 417: 118-122.
http://dx.doi.0rg/10.1016/i.neulet.2007.01.085
Sun. L; Hu. J; Zhao. Z; Li. L; Cheng. H. (2003). Influence of exposure to environmental lead on serum
immunoglobulin in preschool children. Environ Res 92: 124-128.
http://dx.doi.org/10.1016/80013-9351(02)00090-7
Sun. LR; Suszkiw. JB. (1995). Extracellular inhibition and intracellular enhancement of Ca2+ currents by
Pb2+ in bovine adrenal chromaffin cells. J Neurophysiol 74: 574-581.
Sun. Y; Sun. D; Zhou. Z; Zhu. G: Zhang. H; Chang. X; Lei. L; Jin. T. (2008a). Osteoporosis in a Chinese
population due to occupational exposure to lead. Am J Ind Med 51: 436-442.
http://dx.doi.org/10.1002/aiim.20567
Sun. Y; Sun. DH; Zhou. ZJ; Zhu. GY; Lei. LJ; Zhang. HY; Chang. XL; Jin. TY. (2008b). Estimation of
benchmark dose for bone damage and renal dysfunction in a Chinese male population occupationally
exposed to lead. Ann Occup Hyg 52: 527-533. http://dx.doi.org/10.1093/annhyg/men031
Sundstrom. R; Muntzing. K; Kalimo. H; Sourander. P. (1985). Changes in the integrity of the blood-brain
barrier in suckling rats with low dose lead encephalopathy. Acta Neuropathol 68: 1-9.
http://dx.doi.org/10.1007/BF00688948
Surkan. PJ: Schnaas. L; Wright. RJ: Tellez-Rojo. MM; Lamadrid-Figueroa. H; Hu. H; Hernandez-Avila.
M; Bellinger. DC: Schwartz. J: Perroni. E; Wright. RO. (2008). Maternal self-esteem, exposure to
lead, and child neurodevelopment. Neurotoxicology 29: 278-285.
http://dx.doi.0rg/10.1016/i.neuro.2007.ll.006
Surkan. PJ: Zhang. A; Trachtenberg. F; Daniel. DB; McKinlav. S: Bellinger. DC. (2007).
Neuropsychological function in children with blood lead levels <10 ^g/dL. Neurotoxicology 28:
1170-1177. http://dx.doi.0rg/10.1016/i.neuro.2007.07.007
Suszkiw. J: Toth. G: Murawsky. M; Cooper. GP (1984). Effects of Pb2+ and Cd2+ on acetylcholine
release and Ca2+ movements in synaptosomes and subcellular fractions from rat brain and Torpedo
electric organ. Brain Res 323: 31-46. http://dx.doi.org/10.1016/0006-8993(84)90262-2
Suszkiw. JB. (2004). Presynaptic disruption of transmitter release by lead. Neurotoxicology 25: 599-604.
http://dx.doi.0rg/10.1016/i.neuro.2003.09.009
Suzuki. T; Morimura. S: Diccianni. MB; Yamada. R: Hochi. SI; Hirabavashi. M; Yuki. A; NomuraK;
Kitagawa. T; Imagawa. M; Muramatsu. M. (1996). Activation of glutathione transferase P gene by
lead requires glutathione transferase P enhancer I. J Biol Chem 271: 1626-1632.
http://dx.doi.0rg/10.1074/ibc.271.3.1626
Swanson. J: Baler. RD; Volkow. ND. (2011). Understanding the effects of stimulant medications on
cognition in individuals with attention-deficit hyperactivity disorder: a decade of progress [Review].
Neuropsychopharmacology 36: 207-226. http://dx.doi.org/10.1038/npp.2010.160
November 2012 5-786 Draft - Do Not Cite or Quote
-------
Swarup. D: Naresh. R; Varshnev. VP; Balagangatharathilagar. M; Kumar. P; Nandi. D; Patra. RC. (2007).
Changes in plasma hormones profile and liver function in cows naturally exposed to lead and
cadmium around different industrial areas. Res Vet Sci 82: 16-21.
http://dx.doi.0rg/10.1016/i.rvsc.2006.05.002
Taketani. S: Tanaka. A; Tokunaga. R. (1985). Reconstitution of heme-synthesizing activity from ferric ion
and porphyrins, and the effect of lead on the activity. Arch Biochem Biophys 242: 291-296.
http://dx.doi.org/10.1016/0003-9861(85)90504-l
Tanner. CM; Chen. B; Wang. W: Peng. M; Liu. Z; Liang. X; Kao. LC: Gillev. DW: Goetz. CG:
Schoenberg. BS. (1989). Environmental factors and Parkinson's disease: A case-control study in
China. Neurology 39: 660-664.
Tapisso. JT; Marques. CC: Mathias. ML; Ramalhinho. MG. (2009). Induction of micronuclei and sister
chromatid exchange in bone-marrow cells and abnormalities in sperm of Algerian mice (Mus spretus)
exposed to cadmium, lead and zinc. Mutat Res Genet Toxicol Environ Mutagen 678: 59-64.
http://dx.doi.0rg/10.1016/i.mrgentox.2009.07.001
Tarantini. L; Bonzini. M; Apostoli. P; Pegoraro. V; Bollati. V: Marinelli. B; Cantone. L; Rizzo. G: Hou.
L; Schwartz. J; Bertazzi. PA; Baccarelli. A. (2009). Effects of paniculate matter on genomic DNA
methylation content and iNOS promoter methylation. Environ Health Perspect 117: 217-222.
http://dx.doi.org/10.1289/ehp.11898
Task Force of the European Society of Cardiology and the North American Society of Pacing and
Electrophysiologv. (1996). Heart rate variability: Standards of measurement, physiological
interpretation and clinical use. Circulation 93: 1043-1065.
Tavakoli-Nezhad. M; Barron. AJ; Pitts. DK. (2001). Postnatal inorganic lead exposure decreases the
number of spontaneously active midbrain dopamine neurons in the rat. Neurotoxicology 22: 259-269.
http://dx.doi.org/10.1016/S0161-813X(01)00010-9
te Velde. SJ: Ferreira. I; Twisk. JW: Stehouwer. CD; van Mechelen. W: Kemper. HC: Study. AGaHL.
(2004). Birthweight and arterial stiffness and blood pressure in adulthood—results from the
Amsterdam Growth and Health Longitudinal Study. Int J Epidemiol 33: 154-161.
http://dx.doi.org/10.1093/iie/dvh011
Technische Universitat Braunschweig. (2011). BRENDA: Comprehensive enzyme information system.
Available online at http://www.brenda-enzymes.org (accessed January 10, 2011).
Teijon. C: Blanco. MD; Romero. CS: Beneit. JV: Villarino. AL; Guerrero. S: Olmo. R. (2010). Study of
response of thymic and submaxillary lymph node lymphocytes to administration of lead by different
routes. Biol Trace Elem Res 135: 74-85. http://dx.doi.org/10.1007/sl2011-009-8495-6
Teijon. C: Olmo. R; Blanco. D: Romero. A; Teijon. JM. (2006). Low doses of lead: Effects on
reproduction and development in rats. Biol Trace Elem Res 111: 151-165.
http://dx.doi.Org/10.1385/BTER:lll:l:151
Telisman. S: Colak. B; Pizent. A; Jurasovic. J: Cvitkovic. P. (2007). Reproductive toxicity of low-level
lead exposure in men. Environ Res 105: 256-266. http://dx.doi.Org/10.1016/i.envres.2007.05.011
Tellez-Rojo. M. (2008). Email from Martha Tellez Rojo, Direction of Evaluation of Programs and
Quantitative Methods, Center of Investigation in Evaluation and Polls, National institute of Public
Health to Jee-Young Kim, USEPA, Subject - Questions on the Tellez-Rojo et al. (2006) paper. (EPA-
HQ-OAR-2006-0735-5123). U.S. Environmental Protection Agency.
http://www.regulations.gov/#!documentDetail;D=EPA-HQ-OAR-2006-0735-5123
Tellez-Rojo. MM; Bellinger. DC: Arroyo-Quiroz. C: Lamadrid-Figueroa. H; Mercado-Garcia. A;
Schnaas-Arrieta. L; Wright. RO: Hernandez-Avila. M; Hu. H. (2006). Longitudinal associations
between blood lead concentrations lower than 10 microg/dL and neurobehavioral development in
environmentally exposed children in Mexico City. Pediatrics 118: e323-e330.
http://dx.doi.org/10.1542/peds.2005-3123
November 2012 5-787 Draft - Do Not Cite or Quote
-------
Tepper. LB; Levin. LS. (1975). A survey of air and population lead levels in selected American
communities. In TB Griffin; JH Knelson (Eds.), Lead (pp. 152-196). Stuttgart, Germany: Georg
Thieme Publishers.
Terziivanov. D; Filipova. M; Janku. I; Balik. J; Filip. V; Stika. L. (1982). Changes in electroretinogram
and serum potassium during L-DOPA treatment in parkinsonism. Arch Psychiatr Z Neurol 232:
507-513. http://dx.doi.org/10.1007/BF00344065
Teschke. K; Olshan. AF; Daniels. JL; De Roos. AJ; Parks. CG: Schulz. M; Vaughan. TL. (2002).
Occupational exposure assessment in case-control studies: Opportunities for improvement [Review].
Occup Environ Med 59: 575-594. http://dx.doi.Org/10.1136/oem.59.9.575
Tessitore. L; Sesca. E; Pani. P; Dianzani. MU. (1995). Sexual dimorphism in the regulation of cell
turnover during liver hyperplasia. Chem Biol Interact 97: 1-10.
http://dx.doi.org/10.1016/0009-2797(94)03602-l
Thind. IS: Khan. MY. (1978). Potentiation of the neurovirulence of langat virus infection by lead
intoxication in mice. Exp Mol Pathol 29: 342-347. http://dx.doi.org/10.1016/0014-4800(78)90076-X
Tian. L; Lawrence. DA. (1995). Lead inhibits nitric oxide production in vitro by murine splenic
macrophages. Toxicol Appl Pharmacol 132: 156-163. http://dx.doi.org/10.1006/taap.1995.1096
Tian. L; Lawrence. DA. (1996). Metal-induced modulation of nitric oxide production in vitro by murine
macrophages: Lead, nickel, and cobalt utilize different mechanisms. Toxicol Appl Pharmacol 141:
540-547. http://dx.doi.org/10.1006/taap.1996.0320
Tian. Y; Green. PG: Stamova. B; Hertz-Picciotto. I; Pessah. IN; Hansen. R; Yang. X; Gregg. JP;
Ashwood. P; Jickling. G: Van de Water. J; Sharp. FR. (2011). Correlations of gene expression with
blood lead levels in children with autism compared to typically developing controls. Neurotox Res 19:
1-13. http://dx.doi.org/10.1007/sl2640-009-9126-x
Tiffany-Castiglioni. E; Sierra. EM; Wu. JN; Rowles. TK. (1989). Lead toxicity in neuroglia [Review].
Neurotoxicology 10: 417-443.
Tokar. EJ; Diwan. BA; Waalkes. MP (2010). Early life inorganic lead exposure induces testicular
teratoma and renal and urinary bladder preneoplasia in adult metallothionein-knockout mice but not in
wild type mice. Toxicology 276: 5-10. http://dx.doi.Org/10.1016/i.tox.2010.06.006
Tomokuni. K; Ichiba. M. (1990). Effect of lead on the activity of erythrocyte porphobilinogen deaminase
in vivo and in vitro. Toxicol Lett 50: 137-142. http://dx.doi.org/10.1016/0378-4274(90)90003-5
Tomoum. HY; Mostafa. GA; Ismail. NA; Ahmed. SM. (2010). Lead exposure and its association with
pubertal development in school-age Egyptian children: Pilot study. Pediatrics International 52: 89-93.
http://dx.doi.Org/10.llll/i.1442-200X.2009.02893.x
Tomsig. JL; Suszkiw. JB. (1993). Intracellular mechanism of Pb(2+)-induced norepinephrine release from
bovine chromaffin cells. Am J Physiol Cell Physiol 265: C1630-C1636.
Tomsig. JL; Suszkiw. JB. (1995). Multisite interactions between Pb2+ and protein kinase C and its role in
norepinephrine release from bovine adrenal chromaffin cells. JNeurochem 64: 2667-2673.
http://dx.doi.0rg/10.1046/i.1471-4159.1995.64062667.x
Tonelli. M; Wiebe. N; Hemmelgarn. B; Klarenbach. S: Field. C: Manns. B; Thadhani. R; Gill. J. (2009).
Trace elements in hemodialysis patients: A systematic review and meta-analysis [Review]. BMC Med
7: 25. http://dx.doi.org/10.1186/1741-7015-7-25
Tong. IS: Lu. Y. (2001). Identification of confounders in the assessment of the relationship between lead
exposure and child development. Ann Epidemiol 11: 38-45.
http://dx.doi.org/10.1016/S1047-2797(00)00176-9
Tong. S: Baghurst. P: McMichael. A; Sawyer. M; Mudge. J. (1996). Lifetime exposure to environmental
lead and children's intelligence at 11-13 years: The Port Pirie cohort study. Br Med J 312: 1569-1575.
http://dx.doi.org/10.1136/bmi.312.7046.1569
November 2012 5-788 Draft - Do Not Cite or Quote
-------
Tong. S: Baghurst. PA; Sawyer. MG: Burns. J: McMichael. AJ. (1998). Declining blood lead levels and
changes in cognitive function during childhood: the Port Pirie cohort study. JAMA 280: 1915-1919.
http://dx.doi.org/10.1001/iama.280.22.1915
Tong. S: McMichael. AJ: Baghurst. PA. (2000). Interactions between environmental lead exposure and
sociodemographic factors on cognitive development. Arch Environ Health 55: 330-335.
http://dx.doi.org/10.1080/00039890009604025
Trope. I; Lopez-Villegas. D; Cecil. KM; Lenkinski. RE. (2001). Exposure to lead appears to selectively
alter metabolism of cortical gray matter. Pediatrics 107: 1437-1443.
http://dx.doi.org/10.1542/peds. 107.6.1437
Tsaih. SW: Korrick. S: Schwartz. J; Amarasiriwardena. C: Aro. A; Sparrow. D; Hu. H. (2004). Lead,
diabetes, hypertension, and renal function: The Normative Aging Study. Environ Health Perspect 112:
1178-1182. http://dx.doi.org/10.1289/ehp.7024
Tsao. DA; Yu. HS: Cheng. JT; Ho. CK; Chang. HR. (2000). The change of beta-adrenergic system in
lead-induced hypertension. ToxicolAppl Pharmacol 164: 127-133.
http://dx.doi.org/10.1006/taap.1999.8871
Tsuang. M. (2000). Schizophrenia: Genes and environment [Review]. Biol Psychiatry 47: 210-220.
http://dx.doi.org/10.1016/S0006-3223(99)00289-9
Tuppurainen. M; Wagar. G: Kurppa. K; Sakari. W: Wambugu. A; Froseth. B; Alho. J: Nykyri. E. (1988).
Thyroid function as assessed by routine laboratory tests of workers with long-term lead exposure.
Scand J Work Environ Health 14: 175-180.
Turgut. S: Polat. A; Inan. M; Turgut. G: Emmungil. G: Bican. M; Karakus. TY; Gene. O. (2007).
Interaction between anemia and blood levels of iron, zinc, copper, cadmium and lead in children.
Indian J Pediatr 74: 827-830. http://dx.doi.org/10.1007/sl2098-007-0147-2
Turi. GJ: Ellis. R; Wattie. JN; Labiris. NR; Inman. MD. (2011). The effects of inhaled house dust mite on
airway barrier function and sensitivity to inhaled methacholine in mice. Am J Physiol Lung Cell Mol
Physiol 300: L185-L190. http://dx.doi.org/10.1152/ajplung.00271.2010
Turkez. H; Geyikoglu. F; Tatar. A; Keles. MS: Kaplan. I. (2011). The effects of some boron compounds
against heavy metal toxicity in human blood. Exp Toxicol Pathol 64: 93-101.
http://dx.doi.0rg/10.1016/i.etp.2010.06.011
U.S. EPA (U.S. Environmental Protection Agency). (1978). National ambient air quality standard for
lead: final rules and proposed rulemaking. Fed Reg 43: 46246-46263.
U.S. EPA (U.S. Environmental Protection Agency). (1986a). Air quality criteria for lead [EPA Report].
(EPA/600/8-83/028aF-dF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
U.S. EPA (U.S. Environmental Protection Agency). (1986b). Air quality criteria for lead: Volume I of IV
[EPA Report]. (EPA-600/8-83/028aF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
U.S. EPA (U.S. Environmental Protection Agency). (1986c). Lead effects on cardiovascular function,
early development, and stature: An addendum to U.S. EPAAir Quality Criteria for Lead (1986) [EPA
Report]. (EPA-600/8-83/028aF). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (1990a). Air quality criteria for lead: Supplement to
the 1986 addendum [EPA Report]. (EPA/600/8-89/049F). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2006b). Air quality criteria for lead: Volume I of II
[EPA Report]. (EPA/600/R-05/144aF). Research Triangle Park, NC.
http ://cfpub. epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
U.S. EPA (U.S. Environmental Protection Agency). (2006c). Air quality criteria for lead: Volume II of II
[EPA Report]. (EPA/600/R-05/144bF). Research Triangle Park, NC.
http ://cfpub .epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
November 2012 5-789 Draft - Do Not Cite or Quote
-------
U.S. EPA (U.S. Environmental Protection Agency). (2006g). Table 5-7, studies reporting lead-induced
shifts in Thl versus Th2 cytokines [EPA Report]. In Air quality criteria for lead: Volume I of II (pp.
5-225 - 225-226). (EPA/600/R-5/144aF). Research Triangle Park, NC.
U.S. EPA (U.S. Environmental Protection Agency). (2006h). Table AX5.9.6., pattern of lead-induced
macrophage immunotoxicity [EPA Report]. In Air quality criteria for lead: Volume II of II (pp.
AX5-155). (EPA/600/R-05/144bF). Research Triangle Park, NC.
U.S. Renal Data System. (2009). USRDS 2009 annual data report: Atlas of chronic kidney disease and
end-stage renal disease in the United States. Bethesda, MD: National Institutes of Health, National
Institute of Diabetes and Digestive and Kidney Diseases, http://www.usrds.org/adr 2009.htm
Ueda. D; Kishimoto. T; Dekio. S: Tada. M. (1997). Inhibitory effect of lead on tube formation by cultured
human vascular endothelial cells. Hum Cell 10: 283-291.
Ujihara. H; Sasa. M; Ban. T. (1995). Selective blockade of P-type calcium channels by lead in cultured
hippocampal neurons. Jpn J Pharmacol 67: 267-269. http://dx.doi.org/10.1254/ijp.67.267
Ukaejiofo. EO; Thomas. N; Ike. SO. (2009). Haematological assessment of occupational exposure to lead
handlers in Enugu urban, Enugu State, Nigeria. Niger J Clin Pract 12: 58-64.
Undeger. U; Basaran. N; Canpinar. H; Kansu. E. (1996). Immune alterations in lead-exposed workers.
Toxicology 109: 167-172. http://dx.doi.org/10.1016/0300-483X(96)03333-l
Upadhyav. AK; Mathur. R; Bhadauria. M; Nirala. SK. (2009). Therapeutic influence of zinc and ascorbic
acid against lead induced biochemical alterations. Therapie 64: 383-388.
http://dx.doi.org/10.2515/therapie/2009055
Urch. B; Brook. JR; Wasserstein. D; Brook. RD; Raiagopalan. S: Corey. P; Silverman. F. (2004). Relative
contributions of PM2.5 chemical constituents to acute arterial vasoconstriction in humans. Inhal
Toxicol 16: 345-352. http://dx.doi.org/10.1080/08958370490439489
Ustundag. A; Duydu. Y. (2007). The influence of melatonin and N-acetylcysteine in delta-aminolevulinic
acid and lead induced genotoxicity in lymphocytes in vitro. Biol Trace Elem Res 117: 53-64.
http://dx.doi.org/10.1007/BF02698083
Uzbekov. MG: Bubnova. NI; Kulikova. GV (2007). Effect of prenatal lead exposure on superoxide
dismutase activity in the brain and liver of rat fetuses. Bull Exp Biol Med 144: 783-785.
http://dx.doi.org/10.1007/sl0517-007-0431-l
Uzu. G: Sauvain. JJ; Baeza-Squiban. A; Riediker. M; Hohl. MSS: Val. S: Tack. K; Denys. S: Pradere. P;
Dumat. C. (2011). In vitro assessment of the pulmonary toxicity and gastric availability of lead-rich
particles from a lead recycling plant. Environ Sci Technol 45: 7888-7895.
http://dx.doi.org/10.1021/es200374c
Vaglenov. A; Carbonell. E; Marcos. R. (1998). Biomonitoring of workers exposed to lead genotoxic
effects, its modulation by polyvitamin treatment and evaluation of the induced radioresistance. Mutat
Res Genet Toxicol Environ Mutagen 418: 79-92. http://dx.doi.org/10.1016/S1383-5718(98)00111-9
Vaglenov. A; Creus. A; Laltchev. S: Petkova. V: Pavlova. S: Marcos. R. (2001). Occupational exposure to
lead and induction of genetic damage. Environ Health Perspect 109: 295-298.
Valencia. I; Castillo. EE; Chamorro. G: Bobadilla. RA; Castillo. C. (2001). Lead induces endothelium-
and Ca2+-independent contraction in rat aortic rings. Pharmacol Toxicol 89: 177-182.
http://dx.doi.0rg/10.llll/i.0901-9928.2001.890406.x
Valentino. M; Governa. M; Marchiseppe. I; Visona. I. (1991). Effects of lead on polymorphonuclear
leukocyte (PMN) functions in occupationally exposed workers. Arch Toxicol 65: 685-688.
http://dx.doi.org/10.1007/BF02098038
Valentino. M; Rapisarda. V: Santarelli. L; Bracci. M; Scorcelletti. M; Di Lorenzo. L; Cassano. F; Soleo.
L_. (2007). Effect of lead on the levels of some immunoregulatory cytokines in occupationally exposed
workers. Hum Exp Toxicol 26: 551-556. http://dx.doi.org/10.1177/0960327107073817
November 2012 5-790 Draft - Do Not Cite or Quote
-------
Valverde. M; Fortoul. TI; Diaz-Barriga. F; Maiia. J: Del Castillo. ER. (2002). Genotoxicity induced in
CD-I mice by inhaled lead: Differential organ response. Mutagenesis 17: 55-61.
http://dx.doi.0rg/10.1093/mutage/17.l.55
Van de Vvver. FL; D'Haese. PC: Visser. WJ: Elseviers. MM; Knippenberg. LJ; Lamberts. LV; Wedeen.
RP; de Broe. ME. (1988). Bone lead in dialysis patients. Kidney Int 33: 601-607.
Van Esch. GJ: Kroes. R. (1969). The induction of renal tumours by feeding basic lead acetate to mice and
hamsters. Br J Cancer 23: 765-771. http://dx.doi.org/10.1038/bjc. 1969.95
Van Larebeke. N; Koppen. G: Nelen. V; Schoeters. G: Van Loon. H; Albering. H; Riga. L; Vlietinck. R;
Kleinjans. J. (2004). Differences in HPRT mutant frequency among middle-aged Flemish women in
association with area of residence and blood lead levels. Biomarkers 9: 71-84.
http://dx.doi.org/10.1080/13547500310001652160
van Wijngaarden. E; Campbell. JR; Cory-Slechta. DA. (2009). Bone lead levels are associated with
measures of memory impairment in older adults. Neurotoxicology 30: 572-580.
http://dx.doi.0rg/10.1016/i.neuro.2009.05.007
van Wijngaarden. E; Dosemeci. M. (2006). Brain cancer mortality and potential occupational exposure to
lead: Findings from the National Longitudinal Mortality Study, 1979-1989. Int J Cancer 119:
1136-1144. http://dx.doi.org/10.1002/ijc.21947
Vander. AJ. (1988). Chronic effects of lead on the renin-angiotensin system [Review]. Environ Health
Perspect 78: 77-83. http://dx.doi.org/10.1289/ehp.887877
Vargas-Robles. H; Romo. E; Sanchez-Mendoza. A; Rios. AP; Soto. V: Avila-Casado. C: Medina. A;
Escalante. B. (2007). Lead exposure effect on angiotensin II renal vasoconstriction. Hum Exp Toxicol
26: 499-507. http://dx.doi.org/10.1177/0960327106077597
Vassallo. DV: Lebarch. EC: Moreira. CM; Wiggers. GA; Stefanon. I. (2008). Lead reduces tension
development and the myosin ATPase activity of the rat right ventricular myocardium. Braz J Med Biol
Res 41: 789-795. http://dx.doi.org/10.1590/S0100-879X2008000900008
Vaziri. ND. (2008a). Causal link between oxidative stress, inflammation, and hypertension. Iran J Kidney
Dis2: 1-10.
Vaziri. ND. (2008b). Mechanisms of lead-induced hypertension and cardiovascular disease [Review]. Am
J Physiol Heart Circ Physiol 295: H454-H465. http://dx.doi.org/10.1152/ajpheart.00158.2008
Vaziri. ND; Ding. Y. (2001). Effect of lead on nitric oxide synthase expression in coronary endothelial
cells: Role of superoxide. Hypertension 37: 223-226. http://dx.doi.Org/10.1161/01.HYP.37.2.223
Vaziri. ND; Ding. Y; Ni. Z. (1999a). Nitric oxide synthase expression in the course of lead-induced
hypertension. Hypertension 34: 558-562.
Vaziri. ND; Ding. Y; Ni. Z; Barton. CH. (2005). Bradykinin down-regulates, whereas arginine analogs
up-regulates, endothelial nitric-oxide synthase expression in coronary endothelial cells. J Pharmacol
ExpTher313: 121-126. http://dx.doi.org/10.1124/ipet.104.076497
Vaziri. ND; Ding. Y; Ni. Z; Gonick. HC. (1997). Altered nitric oxide metabolism and increased oxygen
free radical activity in lead-induced hypertension: effect of lazaroid therapy. Kidney Int 52:
1042-1046. http://dx.doi.org/10.1038/ki.1997.426
Vaziri. ND; Khan. M. (2007). Interplay of reactive oxygen species and nitric oxide in the pathogenesis of
experimental lead-induced hypertension [Review]. Clin Exp Pharmacol Physiol 34: 920-925.
http://dx.doi.0rg/10.llll/i.1440-1681.2007.04644.x
Vaziri. ND; Liang. K; Ding. Y. (1999b). Increased nitric oxide inactivation by reactive oxygen species in
lead-induced hypertension. Kidney Int 56: 1492-1498.
http://dx.doi.0rg/10.1046/i.1523-1755.1999.00670.x
November 2012 5-791 Draft - Do Not Cite or Quote
-------
Vaziri. ND; Lin. CY; Farmand. F; Sindhu. RK. (2003). Superoxide dismutase, catalase, glutathione
peroxidase and NADPH oxidase in lead-induced hypertension. Kidney Int 63: 186-194.
http://dx.doi.0rg/10.1046/i.1523-1755.2003.00711.x
Vaziri. ND; Wang. XQ. (1999). cGMP-mediated negative-feedback regulation of endothelial nitric oxide
synthase expression by nitric oxide. Hypertension 34: 1237-1241.
Vaziri. ND; Wang. XQ: Oveisi. F; Rad. B. (2000). Induction of oxidative stress by glutathione depletion
causes severe hypertension in normal rats. Hypertension 36: 142-146. http://dx.doi.org/10.1161/
Ol.HYP.36.1.142
Vega-Dienstmaier. JM; Salinas-Pielago. JE; Gutierrez-Campos. MDR; Mandamiento-Ayquipa. RD; Yara-
Hokama. MDC: Ponce-Canchihuaman. J; Castro-Morales. J. (2006). Lead levels and cognitive
abilities in Peruvian children. Rev Bras Psiquiatr 28: 33-39.
http://dx.doi.org/10.1590/S1516-44462006000100008
Veranth. JM: Moss. TA: Chow. JC: Labban. R: Nichols. WK: Walton. JC: Walton. JG: Yost. GS. (2006).
Correlation of in vitro cytokine responses with the chemical composition of soil-derived paniculate
matter. Environ Health Perspect 114: 341-349. http://dx.doi.org/10.1289/ehp.8360
Verina. T; Rohde. CA; Guilarte. TR. (2007). Environmental lead exposure during early life alters granule
cell neurogenesis and morphology in the hippocampus of young adult rats. Neuroscience 145:
1037-1047. http://dx.doi.0rg/10.1016/i.neuroscience.2006.12.040
Verma. SK; Dua. R; Gill. KD. (2005). Impaired energy metabolism after co-exposure to lead and ethanol.
Basic Clin Pharmacol Toxicol 96: 475-479. http://dx.doi.Org/10.llll/i.1742-7843.2005.pto 96611.x
Vetter. SW: Leclerc. E. (2003). Novel aspects of calmodulin target recognition and activation. Eur J
Biochem 270: 404-414. http://dx.doi.Org/10.1046/i.1432-1033.2003.03414.x
Vigeh. M; Yokoyama. K; Kitamura. F; Afshinrokh. M; Bevgi. A; Niroomanesh. S. (2010). Early
pregnancy blood lead and spontaneous abortion. Women Health 50: 756-766.
http://dx.doi.org/10.1080/03630242.2010.532760
Vigeh. M; Yokoyama. K; Sevedaghamiri. Z; Shinohara. A; Matsukawa. T; Chiba. M; Yunesian. M.
(2011). Blood lead at currently acceptable levels may cause preterm labour. Occup Environ Med 68:
231-234. http://dx.doi.org/10.1136/oem.2009.050419
Vimpani. GV: Wigg. NR; Robertson. EF; McMichael. AJ; Baghurst. PA; Roberts. RJ. (1985). The Port
Pirie cohort study: blood lead concentration and childhood developmental assessment. In LJ
Goldwater; LM Wysocki; RA Volpe (Eds.), Edited proceedings: Lead environmental health - the
current issues (pp. 139-146). Durham, NC: Duke University.
Vinceti. M; Guidetti. D; Bergomi. M; Caselgrandi. E; Vivoli. R; Olmi. M; Rinaldi. L; Rovesti. S: Solime.
K (1997). Lead, cadmium, and selenium in the blood of patients with sporadic amyotrophic lateral
sclerosis. Ital JNeurol Sci 18: 87-92. http://dx.doi.org/10.1007/BF01999568
Virgolini. MB; Chen. K; Weston. DP; Bauter. MR; Cory-Slechta. DA. (2005). Interactions of chronic
lead exposure and intermittent stress: Consequences for brain catecholamine systems and associated
behaviors and HPA axis function. Toxicol Sci 87: 469-482. http://dx.doi.org/10.1093/toxsci/kfi269
Virgolini. MB; Rossi-George. A; Lisek. R; Weston. DP; Thiruchelvam. M; Cory-Slechta. DA. (2008a).
CNS effects of developmental Pb exposure are enhanced by combined maternal and offspring stress.
Neurotoxicology 29: 812-827. http://dx.doi.Org/10.1016/i.neuro.2008.03.003
Virgolini. MB; Rossi-George. A; Weston. D; Cory-Slechta. DA. (2008b). Influence of low level maternal
Pb exposure and prenatal stress on offspring stress challenge responsivity. Neurotoxicology 29:
928-939. http://dx.doi.0rg/10.1016/i.neuro.2008.09.010
Vogetseder. A; Picard. N; Gaspert. A; Walch. M; Kaissling. B; Le Hir. M. (2008). Proliferation capacity of
the renal proximal tubule involves the bulk of differentiated epithelial cells. Am J Physiol Cell Physiol
294: C22-C28. http://dx.doi.org/10.1152/aipcell.00227.2007
November 2012 5-792 Draft - Do Not Cite or Quote
-------
Vvskocil. A; Semecky. V; Fiala. Z; Cizkova. M; Viau. C. (1995). Renal alterations in female rats
following subchronic lead exposure. J Appl Toxicol 15: 257-262.
http://dx.doi.org/10.1002/iat.2550150405
Waalkes. MP: Diwan. BA; Ward. JM; Devor. DE; Gover. RA. (1995). Renal tubular tumors and atypical
hyperplasias in B6C3F1 mice exposed to lead acetate during gestation and lactation occur with
minimal chronic nephropathy. Cancer Res 55: 5265-5271.
Waalkes. MP: Liu. J; Gover. RA; Diwan. BA. (2004). Metallothionein-I/II double knockout mice are
hypersensitive to lead-induced kidney carcinogenesis: Role of inclusion body formation. Cancer Res
64: 7766-7772. http://dx.doi.org/10.1158/0008-5472.CAN-04-2220
Wananukul. W: Sura. T; Salaitanawatwong. P. (2006). Polymorphism of delta-aminolevulinic acid
dehydratase and its effect on blood lead levels in Thai workers. Arch Environ Occup Health 61: 67-72.
http://dx.doi.Org/10.3200/AEOH.61.2.67-72
Wang. A; Wang. Q; Song. Q; Xu. J. (2009a). [Study of ALAD and VDR gene polymorphisms associated
with lead nephrotoxicity susceptibility]. Wei Sheng Yan Jiu 38: 326-329.
Wang. CH; Zhang. Y; Liang. JC: Shan. G: Wang. Y; Shi. Q. (2006a). Impacts of ascorbic acid and
thiamine supplementation at different concentrations on lead toxicity in testis. Clin Chim Acta 370:
82-88. http://dx.doi.0rg/10.1016/j.cca.2006.01.023
Wang. CY; Lin. YW: Yang. JL. (2008c). Activation of protein kinase C alpha signaling prevents
cytotoxicity and mutagenicity following lead acetate in CL3 human lung cancer cells. Toxicology 250:
55-61. http://dx.doi.0rg/10.1016/j.tox.2008.06.001
Wang. FT; Hu. H; Schwartz. J: Weuve. J: Spiro. AS. Ill; Sparrow. D; Nie. HL; Silverman. EK; Weiss. ST;
Wright. RO. (2007a). Modifying effects of the HFE polymorphisms on the association between lead
burden and cognitive decline. Environ Health Perspect 115: 1210-1215.
http://dx.doi.org/10.1289/ehp.9855
Wang. G: Chen. XQ; Lipsky. MM; Whittaker. MH; Fowler. BA. (2005a). Effect of combined lead,
cadmium and arsenic exposure on the expression of cellular protective factors in rat kidneys at LOEL
dose levels [Abstract]. Toxicologist 85: 237.
Wang. GS: Fowler. BA. (2008). Roles of biomarkers in evaluating interactions among mixtures of lead,
cadmium and arsenic [Review]. Toxicol Appl Pharmacol 233: 92-99.
http://dx.doi.0rg/10.1016/i.taap.2008.01.017
Wang. HL; Chen. XT; Yang. B; Ma. FL; Wang. S: Tang. ML; Hao. MG: Ruan. DY. (2008d). Case-control
study of blood lead levels and attention deficit hyperactivity disorder in Chinese children. Environ
Health Perspect 116: 1401-1406. http://dx.doi.org/10.1289/ehp.11400
Wang. L; Chen. DW: Wang. H; Liu. ZP (2009b). Effects of lead and/or cadmium on the expression of
metallothionein in the kidney of rats. Biol Trace Elem Res 129: 190-199.
http://dx.doi.org/10.1007/sl2011-008-8288-3
Wang. L; Wang. H; Hu. MZ; Cao. J: Chen. DW: Liu. Z. (2009c). Oxidative stress and apoptotic changes
in primary cultures of rat proximal tubular cells exposed to lead. Arch Toxicol 83: 417-427.
http://dx.doi.org/10.1007/s00204-009-0425-z
Wang. L; Wang. H; Li. J: Chen. D; Liu. Z. (20 lib). Simultaneous effects of lead and cadmium on primary
cultures of rat proximal tubular cells: interaction of apoptosis and oxidative stress. Arch Environ
Contam Toxicol 61: 500-511. http://dx.doi.org/10.1007/s00244-011-9644-4
Wang. L; Wang. Z; Liu. J. (2010d). Protective effect of N-acetylcysteine on experimental chronic lead
nephrotoxicity in immature female rats. Hum Exp Toxicol 29: 581-591.
http://dx.doi.org/10.1177/0960327109357270
Wang. Q; Luo. WJ: Zhang. WB; Dai. ZM; Chen. YM; Chen. JY (2007b). Iron supplementation protects
against lead-induced apoptosis through MAPK pathway in weanling rat cortex. Neurotoxicology 28:
850-859. http://dx.doi.0rg/10.1016/i.neuro.2007.04.004
November 2012 5-793 Draft - Do Not Cite or Quote
-------
Wang. Q; Zhao. HH; Chen. JW: Hao. QL; Gu. KD; Zhu. YX; Zhou. YK; Ye. LX. (201 Of), delta-
Aminolevulinic acid dehydratase activity, urinary delta-aminolevulinic acid concentration and zinc
protoporphyrin level among people with low level of lead exposure. Int J Hyg Environ Health 213:
52-58. http://dx.doi.0rg/10.1016/j.ijheh.2009.08.003
Wang. TL; Kao. TH; Inbarai. BS: Su. YT; Chen. BH. (2010g). Inhibition effect of poly(y-glutamic acid)
on lead-induced toxicity in mice. J Agric Food Chem 58: 12562-12567.
http://dx.doi.org/10.1021/ifl034509
Wang. YY; Sui. KX; Hong. LI; Ma. HY. (2009e). The effects of lead exposure on placenta! NF-kappaB
expression and the consequences for gestation. Reprod Toxicol 27: 190-195.
http://dx.doi.0rg/10.1016/i.reprotox.2008.12.006
Wasserman. G: Graziano. JH; Factor-Litvak. R; Popovac. D; Morina. N; Musabegovic. A; Vrenezi. N;
Capuni-Paracka. S: Lekic. V: Preteni-Redjepi. E; Hadzialjevic. S: Slavkovich. V; Kline. J: Shrout. P;
Stein. Z. (1992). Independent effects of lead exposure and iron deficiency anemia on developmental
outcome at age 2 years. J Pediatr 121: 695-703. http://dx.doi.org/10.1016/80022-3476(05)81895-5
Wasserman. GA; Factor-Litvak. P. (2001). Methodology, inference and causation: Environmental lead
exposure and childhood intelligence [Comment]. Arch Clin Neuropsychol 16: 343-352.
http://dx.doi.org/10.1016/S0887-6177(00)00085-8
Wasserman. GA; Factor-Litvak. P; Liu. X; Todd. AC: Kline. JK; Slavkovich. V; Popovac. D; Graziano.
JH. (2003). The relationship between blood lead, bone lead and child intelligence. Child Neuropsychol
9: 22-34. http://dx.doi.Org/10.1076/chin.9.l.22.14497
Wasserman. GA: Graziano. JH: Factor-Litvak. P; Popovac. D: Morina. N; Musabegovic. A; Vrenezi. N;
Capuni-Paracka. S: Lekic. V: Preteni-Redjepi. E; Hadzialjevic. S: Slavkovich. V; Kline. J: Shrout. P:
Stein. Z. (1994). Consequences of lead exposure and iron supplementation on childhood development
at age 4 years. Neurotoxicol Teratol 16: 233-240. http://dx.doi.org/10.1016/0892-0362(94)90044-2
Wasserman. GA: Liu. X; Lolacono. NJ; Factor-Litvak. P; Kline. JK; Popovac. D: Morina. N;
Musabegovic. A; Vrenezi. N; Capuni-Paracka. S: Lekic. V: Preteni-Redjepi. E; Hadzialjevic. S:
Slavkovich. V; Graziano. JH. (1997). Lead exposure and intelligence in 7-year-old children: The
Yugoslavia prospective study. Environ Health Perspect 105: 956-962.
Wasserman. GA: Liu. X; Pine. DS: Graziano. JH. (2001). Contribution of maternal smoking during
pregnancy and lead exposure to early child behavior problems. Neurotoxicol Teratol 23: 13-21.
http://dx.doi.org/10.1016/S0892-0362(00)00116-l
Wasserman. GA: Musabegovic. A; Liu. X; Kline. J: Factor-Litvak. P; Graziano. JH. (2000). Lead
exposure and motor functioning in 4 1/2-year-old children: The Yugoslavia prospective study. J
Pediatr 137: 555-561. http://dx.doi.org/10.1067/mpd.2000.109111
Wasserman. GA: Staghezza-Jaramillo. B; Shrout. P: Popovac. D: Graziano. J. (1998). The effect of lead
exposure on behavior problems in preschool children. Am J Public Health 88: 481-486.
http://dx.doi.Org/10.2105/AJPH.88.3.481
Waterman. SJ: El-Fawal. HAN; Snyder. CA. (1994). Lead alters the immunogenicity of two neural
proteins: A potential mechanism for the progression of lead-induced neurotoxicity. Environ Health
Perspect 102: 1052-1056.
Watts. SW: Chai. S: Webb. RC. (1995). Lead acetate-induced contraction in rabbit mesenteric artery:
Interaction with calcium and protein kinase C. Toxicology 99: 55-65.
http://dx.doi.org/10.1016/0300-483X(94)03003-K
Weaver. V; Jaar. B. (2010). UpToDate: Lead nephropathy and lead-related nephrotoxicity. Available
online at http://www.uptodate.com/contents/lead-nephropathv-and-lead-related-nephrotoxicitv
(accessed October 25, 2010).
Weaver. VM. (2010). Further analysis of Korean lead worker data set with a focus on determining the
functional form of the exposure-response relationship. Available online
November 2012 5-794 Draft - Do Not Cite or Quote
-------
Weaver. VM; Ellis. LR; Lee. BK; Todd. AC: Shi. W: Ahn. KD; Schwartz. BS. (2008). Associations
between patella lead and blood pressure in lead workers. Am J Ind Med 51: 336-343.
http://dx.doi.org/10.1002/aiim.20573
Weaver. VM: Griswold. M: Todd. AC: Jaar. BG: Ahn. KD: Thompson. CB: Lee. BK. (2009).
Longitudinal associations between lead dose and renal function in lead workers. Environ Res 109:
101-107. http://dx.doi.org/10.1016/i.envres.20Q8.09.005
Weaver. VM: Jarr. BG: Schwartz. BS: Todd. AC: Ahn. KD: Lee. SS: Wen. J; Parsons. PJ: Lee. BK.
(2005a). Associations among lead dose biomarkers, uric acid, and renal function in Korean lead
workers. Environ Health Perspect 113: 36-42. http://dx.doi.org/10.1289/ehp.7317
Weaver. VM: Kim. NS: Jaar. BG: Schwartz. BS: Parsons. PJ: Steuerwald. AJ; Todd. AC: Simon. D; Lee.
BK. (2011). Associations of low-level urine cadmium with kidney function in lead workers. Occup
Environ Med 68: 250-256. http://dx.doi.org/10.1136/oem.2010.056077
Weaver. VM: Lee. BK: Ahn. KD: Lee. GS: Todd. AC: Stewart. WF: Wen. J: Simon. DJ: Parsons. PJ:
Schwartz. BS. (2003a). Associations of lead biomarkers with renal function in Korean lead workers.
Occup Environ Med 60: 551-562. http://dx.doi.Org/10.1136/oem.60.8.551
Weaver. VM; Lee. BK: Todd. AC: Ahn. KD: Shi. W: Jaar. BG: Kelsev. KT; Lustberg. ME; Silbergeld.
EK; Parsons. PJ: Wen. J: Schwartz. BS. (2006). Effect modification by delta-aminolevulinic acid
dehydratase, vitamin D receptor, and nitric oxide synthase gene polymorphisms on associations
between patella lead and renal function in lead workers. Environ Res 102: 61-69.
http://dx.doi.0rg/10.1016/i.envres.2006.01.001
Weaver. VM; Schwartz. BS: Ahn. KD: Stewart. WF; Kelsev. KT; Todd. AC: Wen. J: Simon. DJ: Lustberg.
ME; Parsons. PJ: Silbergeld. EK; Lee. BK. (2003b). Associations of renal function with
polymorphisms in the "delta"-aminolevulinic acid dehydratase, vitamin D receptor, and nitric oxide
synthase genes in Korean lead workers. Environ Health Perspect 111: 1613-1619.
Weaver. VM: Schwartz. BS: Jaar. BG: Ahn. KD: Todd. AC: Lee. SS: Kelsev. KT: Silbergeld. EK:
Lustberg. ME: Parsons. PJ: Wen. J: Lee. BK. (2005b). Associations of uric acid with polymorphisms
in the "delta"-aminolevulinic acid dehydratase, vitamin D receptor, and nitric oxide synthase genes in
Korean lead workers. Environ Health Perspect 113: 1509-1515. http://dx.doi.org/10.1289/ehp.7927
Wedrychowski. A: Schmidt. WN: Hnilica. LS. (1986). The in vivo cross-linking of proteins and DNAby
heavy metals. J Biol Chem 261: 3370-3376.
Wei. H: Wei. D: Yi. S: Zhang. F: Ding. W. (2011). Oxidative stress induced by urban fine particles in
cultured EA.hy926 cells. Hum Exp Toxicol 30: 579-590. httrj://dx.doi.org/10.1177/0960327110374207
Weiss. B. (1988). Neurobehavioral toxicity as a basis for risk assessment [Review]. Trends Pharmacol Sci
9: 59-62.
Weiss. B. (1990). Risk assessment: The insidious nature of neurotoxicity and the aging brain [Review].
Neurotoxicology 11: 305-314.
Weisskopf. MG: Jain. N; Nie. HL: Sparrow. D: Vokonas. P: Schwartz. J: Hu. H. (2009). A prospective
study of bone lead concentration and death from all causes, cardiovascular diseases, and cancer in the
department of veterans affairs normative aging study. Circulation 120: 1056-1064.
http://dx.doi.org/10.1161/circulationaha.108.827121
Weisskopf. MG: Proctor. SP: Wright. RO: Schwartz. J: Spiro. A. Ill: Sparrow. D: Nie. HL: Hu. H.
(2007b). Cumulative lead exposure and cognitive performance among elderly men. Epidemiology 18:
59-66. htrp://dx.doi.org/10.1097/01.ede.0000248237.35363.29
Weisskopf. MG: Weuve. J: Nie. H: Saint-Hilaire. MH: Sudarsky. L: Simon. DK: Hersh. B: Schwartz. J:
Wright. RO: Hu. H. (2010). Association of cumulative lead exposure with Parkinson's Disease.
Environ Health Perspect 118: 1609-1613. http://dx.doi.org/10.1289/ehp.1002339
November 2012 5-795 Draft - Do Not Cite or Quote
-------
Weisskopf. MG: Wright. RO: Schwartz. J; Spiro. A. Ill: Sparrow. D; Aro. A; Hu. H. (2004). Cumulative
lead exposure and prospective change in cognition among elderly men: The VA Normative Aging
Study. Am J Epidemiol 160: 1184-1193. http://dx.doi.org/10.1093/aje/kwh333
Wellenius. GA; Coull. BA; Godleski. JJ; Koutrakis. P; Okabe. K; Savage. ST. (2003). Inhalation of
concentrated ambient air particles exacerbates myocardial ischemia in conscious dogs. Environ Health
Perspect 111: 402-408. http://dx.doi.org/10.1289/ehp.5775
Wells. E; Jarrett. J: Hong Lin. Y; Caldwell. K; Hibbeln. J: Apelberg. B; Herbstman. J: Halden. R; Witter.
F; Goldman. L. (2011 a). Body burdens of mercury, lead, selenium and copper among Baltimore
newborns. Environ Res 111: 411-417. http://dx.doi.Org/10.1016/j.envres.2010.12.009
Wells. EM; Navas-Acien. A; Herbstman. JB; Apelberg. BJ; Silbergeld. EK; Caldwell. KL; Jones. RL;
Halden. RU; Witter. FR; Goldman. LR. (20lib). Low level lead exposure and elevations in blood
pressure during pregnancy. Environ Health Perspect 119: 664-669.
http://dx.doi.org/10.1289/ehp.1002666
Westerink. RH; Vijverberg. HP. (2002). Ca2+-independent vesicular catecholamine release in PC 12 cells
by nanomolar concentrations of Pb2+. JNeurochem 80: 861-867.
http://dx.doi.0rg/10.1046/i.0022-3042.2001.00751.x
Wetmur. JG. (1994). Influence of the common human delta-aminolevulinate dehydratase polymorphism
on lead body burden [Review]. Environ Health Perspect 102: 215-219.
Wetmur. JG: Lehnert. G: Desnick. RJ. (1991b). The delta-aminolevulinate dehydratase polymorphism:
Higher blood lead levels in lead workers and environmentally exposed children with the 1-2 and 2-2
isozymes. Environ Res 56: 109-119. http://dx.doi.org/10.1016/80013-9351(05)80001-5
Weuve. J: Kelsev. KT; Schwartz. J: Bellinger. D; Wright. RO: Raian. P; Spiro. A. Ill; Sparrow. D; Aro. A;
Hu. H. (2006). Delta-aminolevulinic acid dehydratase polymorphism and the relation between low
level lead exposure and the Mini-Mental Status Examination in older men: The Normative Aging
Study. Occup Environ Med 63: 746-753. http://dx.doi.org/10.1136/oem.2006.027417
Weuve. J: Korrick. SA; Weisskopf. MA; Ryan. LM; Schwartz. J: Nie. HL; Grodstein. F; Hu. H. (2009).
Cumulative exposure to lead in relation to cognitive function in older women. Environ Health
Perspect 117: 574-580. http://dx.doi.org/10.1289/ehp.11846
Whitworth. JA; Mangos. GJ: Kelly. JJ. (2000). Gushing, cortisol, and cardiovascular disease [Review].
Hypertension 36: 912-916. http://dx.doi.Org/10.1161/01.HYP.36.5.912
WHO (World Health Organization). (1948). Preamble to the Constitution of the World Health
Organization as adopted by the International Health Conference, New York, 19-22 June, 1946. In
Constitution of the World Health Organization (pp. 2). Geneva, Switzerland.
http://whqlibdoc.who.int/hist/official records/constitution.pdf
WHO (World Health Organization). (1992). International statistical classification of diseases and related
health problem, tenth revision. Geneva, Switzerland, http ://www.who, int/classifications/icd/en/
Wiebe. JP; Barr. KJ. (1988). Effect of prenatal and neonatal exposure to lead on the affinity and number
of estradiol receptors in the uterus. J Toxicol Environ Health 24: 451-460.
http://dx.doi.org/10.1080/15287398809531176
Wiebe. JP: Barr. KJ: Buckingham. KD. (1988). Effect of prenatal and neonatal exposure to lead on
gonadotropin receptors and steroidogenesis in rat ovaries. J Toxicol Environ Health 24: 461-476.
Wiemann. M; Schirrmacher. K; Busselberg. D. (1999). Interference of lead with the calcium release
activated calcium flux of osteoblast-like cells. Calcif Tissue Int 65: 479-485.
http://dx.doi.org/10.1007/s002239900736
Wilker. E; Korrick. S: Nie. LH; Sparrow. D: Vokonas. P: Coull. B; Wright. RO: Schwartz. J: Hu. H.
(2011). Longitudinal changes in bone lead levels: The VA Normative Aging Study. J Occup Environ
Med 53: 850-855. http://dx.doi.org/10.1097/JOM.Ob013e31822589a9
November 2012 5-796 Draft - Do Not Cite or Quote
-------
Williams. BL; Pennock-Roman. M; Suen. HK; Magsumbol. MS: Ozdenerol. E. (2007). Assessing the
impact of the local environment on birth outcomes: A case for HLM. J Expo Sci Environ Epidemiol
17: 445-457. http://dx.doi.org/10.1038/sj.jes.7500537
Williams. PL; Sergevev. O: Lee. MM; Korrick. SA; Burns. JS: Humblet. O: DelPrato. J: Revich. B;
Hauser. R. (2010). Blood lead levels and delayed onset of puberty in a longitudinal study of Russian
boys. Pediatrics 125: 1088-1096. http://dx.doi.org/10.1542/peds.2009-2575
Wise. JR Sr; Stearns. DM; Wetterhahn. KE; Patierno. SR. (1994). Cell-enhanced dissolution of
carcinogenic lead chromate particles: the role of individual dissolution products in clastogenesis.
Carcinogenesis 15: 2249-2254. http://dx.doi.org/10.1093/carcin/15.10.2249
Wise. JP. Sr: Wise. SS: Lacerte. C: Wise. JP; Aboueissa. AM. (2011). The genotoxicity of paniculate and
soluble chromate in sperm whale (physeter macrocephalus) skin fibroblasts. Environ Mol Mutagen 52:
43-49. http://dx.doi.org/10.1002/em.20579
Wise. SS: Holmes. AL; Ketterer. ME; Hartsock. WJ: Fomchenko. E; Katsifis. S: Thompson. WD; Wise.
JP. Sr. (2004). Chromium is the proximate clastogenic species for lead chromate-induced
clastogenicity in human bronchial cells. Mutat Res Genet Toxicol Environ Mutagen 560: 79-89.
http://dx.doi.0rg/10.1016/i.mrgentox.2004.02.009
Wise. SS: Holmes. AL; Moreland. JA; Xie. H; Sandwick. SJ: Stackpole. MM; Fomchenko. E; Teufack. S:
May. AJ. Jr; Katsfis. SP; Wise. JP. Sr. (2005). Human lung cell growth is not stimulated by lead ions
after lead chromate-induced genotoxicity. Mol Cell Biochem 279: 75-84.
http://dx.doi.org/10.1007/sll010-005-8217-0
Wise. SS: Holmes. AL: Qin. Q; Xie. H; Katsifis. SP: Thompson. WD: Wise. JR Sr. (2010). Comparative
genotoxicity and cytotoxicity of four hexavalent chromium compounds in human bronchial cells.
Chem Res Toxicol 23: 365-372. http://dx.doi.org/10.1021/tx900363j
Wise. SS: Holmes. AL; Wise. JP. Sr. (2006a). Paniculate and soluble hexavalent chromium ate cytotoxic
and genotoxic to human lung epithelial cells. Mutat Res Genet Toxicol Environ Mutagen 610: 2-7.
http://dx.doi.0rg/10.1016/i.mrgentox.2006.06.005
Wise. SS: Holmes. AL: Xie. H; Thompson. WD: Wise. JP. Sr. (2006b). Chronic exposure to paniculate
chromate induces spindle assembly checkpoint bypass in human lung cells. Chem Res Toxicol 19:
1492-1498. http://dx.doi.org/10.1021/tx0601410
Wise. SS: Shafflev. F; Lacerte. C: Goertz. CEC: Dunn. JL; Gulland. FMD; Aboueissa. AE; Zheng. TZ;
Wise. JP. (2009). Paniculate and soluble hexavalent chromium are cytotoxic and genotoxic to Steller
sea lion lung cells. Aquat Toxicol 91: 329-335. http://dx.doi.Org/10.1016/i.aquatox.2008.12.004
Wiwanitkit. V: Suwansaksri. J: Soogarun. S. (2008). White blood cell sister chromatid exchange among a
sample of Thai subjects exposed to lead: Lead-induced genotoxicity. Toxicol Environ Chem 90:
765-768. http://dx.doi.org/10.1080/02772240701712758
Wolf. J: Daley. AJ. (2007). Microbiological aspects of bacterial lower respiratory tract illness in children:
Atypical pathogens. Paediatr Respir Rev 8: 212-220. http://dx.doi.Org/10.1016/i.prrv.2007.07.004
Wolff. MS: Britton. JA; Boguski. L; Hochman. S: Malonev. N; Serra. N; Liu. ZS: Berkowitz. G: Larson.
S: Forman. J. (2008). Environmental exposures and puberty in inner-city girls. Environ Res 107:
393-400. http://dx.doi.0rg/10.1016/i.envres.2008.03.006
Wozniak. K; Blasiak. J. (2003). In vitro genotoxicity of lead acetate: Induction of single and double DNA
strand breaks and DNA-protein cross-links. Mutat Res Genet Toxicol Environ Mutagen 535: 127-139.
http://dx.doi.org/10.1016/S1383-5718(02)00295-4
Wright. JP: Dietrich. KN; Ris. MD; Hornung. RW: Wessel. SD; Lanphear. BP; Ho. M; Rae. MN. (2008).
Association of prenatal and childhood blood lead concentrations with criminal arrests in early
adulthood. PLoS Med 5: 732-740. http://dx.doi.org/10.1371/iournal.pmed.0050101
November 2012 5-797 Draft - Do Not Cite or Quote
-------
Wright. RO: Schwartz. J; Wright. RJ: Bollati. V: Tarantini. L; Park. SK; Hu. H; Sparrow. D; Vokonas. P;
Baccarelli. A. (2010). Biomarkers of lead exposure and DNA methylation within retrotransposons.
Environ Health Perspect 118: 790-795. http://dx.doi.org/10.1289/ehp.0901429
Wright. RO: Tsaih. SW: Schwartz. J; Spiro. A; McDonald. K; Weiss. ST; Hu. H. (2003). Lead exposure
biomarkers and mini-mental status exam scores in older men. Epidemiology 14: 713-718.
htrp://dx.doi.org/10.1097/01.EDE.0000081988.85964.db
Wu. J: Basha. MR; Brock. B; Cox. DP; Cardozo-Pelaez. F; Mcpherson. CA; Harry. J: Rice. DC: Malonev.
B; Chen. D; Lahiri. DK; Zawia. NH. (2008a). Alzheimer's disease (AD)-like pathology in aged
monkeys after infantile exposure to environmental metal lead (pb): Evidence for a developmental
origin and environmental link for AD. JNeurosci 28: 3-9.
http://dx.doi.org/10.1523/ineurosci.4405-07.2008
Wu. J: Basha. R; Zawia. NH. (2008b). The environment, epigenetics and amyloidogenesis [Review]. J
Mol Neurosci 34: 1-7. http://dx.doi.org/10.1007/sl2031-007-0009-4
Wu. MT; Kelsev. K; Schwartz. J: Sparrow. D: Hu. H. (2003a). A delta-aminolevulinic acid dehydratase
(ALAD) polymorphism may modify the relationship of low-level lead exposure to uricemia and renal
function: The normative aging study. Environ Health Perspect 111: 335-341.
http://dx.doi.org/10.1289/ehp.5504
Wu. T; Buck. GM; Mendola. P. (2003b). Blood lead levels and sexual maturation in U.S. girls: The Third
National Health and Nutrition Examination Survey, 1988-1994. Environ Health Perspect 111:
737-741. http://dx.doi.org/10.1289/ehp.6008
Wu. TN; Shen. CY; Lai. JS: Goo. CF; Ko. KN; Chi. HY; Chang. PY; Liou. SH. (2000). Effects of lead
and noise exposures on hearing ability. Arch Environ Health 55: 109-114.
http://dx.doi.org/10.1080/00039890009603396
Xie. H; Holmes. AL; Wise. SS: Huang. S: Peng. C: Wise. JP. Sr. (2007). Neoplastic transformation of
human bronchial cells by lead chromate particles. Am J Respir Cell Mol Biol 37: 544-552.
htrp://dx.doi.org/10.1165/rcmb.2007-0058OC
Xie. H; Wise. SS: Wise. JP. Sr. (2008). Deficient repair of particulate hexavalent chromium-induced DNA
double strand breaks leads to neoplastic transformation. Mutat Res Genet Toxicol Environ Mutagen
649: 230-238. http://dx.doi.Org/10.1016/i.mrgentox.2007.09.008
Xie. Y; Chiba. M; Shinohara. A; Watanabe. H; Inaba. Y. (1998). Studies on lead-binding protein and
interaction between lead and selenium in the human erythrocytes. Ind Health 36: 234-239.
http://dx.doi.org/10.2486/indhealth.36.234
Xu. J; Ji. LD; Xu. LH. (2006). Lead-induced apoptosis in PC 12 cells: Involvement of p53, Bcl-2 family
and caspase-3. Toxicol Lett 166: 160-167. http://dx.doi.Org/10.1016/i.toxlet.2006.06.643
Xu. J; Lian. LJ; Wu. C: Wang. XF; Fu. WY; Xu. LH. (2008). Lead induces oxidative stress, DNA damage
and alteration of p53, Bax and Bcl-2 expressions in mice. Food Chem Toxicol 46: 1488-1494.
http://dx.doi.0rg/10.1016/i.fct.2007.12.016
Xu. J: Yan. CH: Yang. B: Xie. HF: Zou. XY: Zhong. L: Gao. Y: Tian. Y: Shen. XM. (2009c). The role of
metabotropic glutamate receptor 5 in developmental lead neurotoxicity. Toxicol Lett 191: 223-230.
http://dx.doi.0rg/10.1016/i.toxlet.2009.09.001
Xu. SZ; Raj anna. B. (2006). Glutamic acid reverses Pb2+-induced reductions of NMDA receptor subunits
in vitro. Neurotoxicology 27: 169-175. http://dx.doi.Org/10.1016/i.neuro.2005.08.005
Yamanaka. K; Chun. SJ: Boillee. S: Fujimori-Tonou. N; Yamashita. H; Gutmann. DH; Takahashi. R;
Misawa. H; Cleveland. DW. (2008). Astrocytes as determinants of disease progression in inherited
amyotrophic lateral sclerosis. Nat Neurosci 11: 251-253. http://dx.doi.org/10.1038/Nn2047
Yanez. L; Garcia-Nieto. E; Rojas. E; Carrizales. L; Mejia. J: Calderon. J: Razo. I; Diaz-Barriga. F. (2003).
DNA damage in blood cells from children exposed to arsenic and lead in a mining area. Environ Res
93: 231-240. http://dx.doi.Org/10.1016/i.envres.2003.07.005
November 2012 5-798 Draft - Do Not Cite or Quote
-------
Yang. Y; Ma. Y; Ni. L; Zhao. S: Li. L; Zhang. J; Fan. M; Liang. C: Cao. J: Xu. L. (2003). Lead exposure
through gestation-only caused long-term learning/memory deficits in young adult offspring. Exp
Neural 184: 489-495. http://dx.doi.org/10.1016/S0014-4886(03)00272-3
Yang. Y; Raine. A; Lencz. T; Bihrle. S: LaCasse. L; Colletti. P. (2005). Volume reduction in prefrontal
gray matter in unsuccessful criminal psychopaths. Biol Psychiatry 57: 1103-1108.
http://dx.doi.0rg/10.1016/i.biopsvch.2005.01.021
Yazbeck. C: Thiebaugeorges. O: Moreau. T; Goua. V: Debotte. G: Sahuquillo. J: Forhan. A; Foliguet B;
Magnin. G: Slama. R; Charles. MA; Huel. G. (2009). Maternal blood lead levels and the risk of
pregnancy-induced hypertension: The EDEN cohort study. Environ Health Perspect 117: 1526-1530.
htrp://dx.doi.org/10.1289/ehp.080Q488
Ye. SH. (1993). Hypoxanthine phosphoribosyl transferase assay of lead mutagenicity on keratinocytes.
Acta Pharmacol Sin 14: 145-147.
Yeates. DB; Hameister. WM. (1992). Alveolar epithelial permeability in baboons: Histamine and
capsaicin. J Physiol 450: 363-374.
Yedjou. CG: Milner. JN; Howard. CB; Tchounwou. PB. (2010). Basic apoptotic mechanisms of lead
toxicity in human leukemia (HL-60) cells. Int J Environ Res Public Health 7: 2008-2017.
http://dx.doi.org/10.3390/ijerph7052008
Yen. TH: Lin. JL: Lin-Tan. DT: Hsu. CW: Chen. KH: Hsu. HH. (2011). Blood cadmium level's
association with 18-month mortality in diabetic patients with maintenance haemodialysis. Nephrol
Dial Transplant 26: 998-1005. http://dx.doi.org/l0.1093/ndt/gfq448
Yiin. SJ: Lin. TH. (1995). Lead-catalyzed peroxidation of essential unsaturated fatty acid. Biol Trace
Elem Res 50: 167-172. http://dx.doi.org/10.1007/BF02789419
Yin. Y; Zhang. T; Dai. Y; Bao. Y; Chen. X; Lu. X. (2008). The effect of plasma lead on anembryonic
pregnancy. AnnN YAcad Sci 1140: 184-189. http://dx.doi.org/10.1196/annals.1454.042
Youssef. SAH; El-Sanousi. AA; Afifi. NA; El-Brawv. AMA. (1996). Effect of subclinical lead toxicity on
the immune response of chickens to Newcastle disease virus vaccine. Res Vet Sci 60: 13-16.
http://dx.doi.org/10.1016/S0034-5288(96)90123-X
Yu. CC: Lin. JL; Lin-Tan. DT. (2004). Environmental exposure to lead and progression of chronic renal
diseases: A four-year prospective longitudinal study. JAm Soc Nephrol 15: 1016-1022.
http://dx.doi.org/10.1097/01.ASN.0000118529.01681.4F
Yu. DY; Li. WF; Deng. B; Mao. XF. (2008). Effects of lead on hepatic antioxidant status and transcription
of superoxide dismutase gene in pigs. Biol Trace Elem Res 126: 121-128.
http://dx.doi.org/10.1007/sl2011-008-8198-4
Yu. J; Fujishiro. H; Miyataka. H; Oyama. TM; Hasegawa. T; Seko. Y; Miura. N; Himeno. S. (2009).
Dichotomous effects of lead acetate on the expression of metallothionein in the liver and kidney of
mice. Biol Pharm Bull 32: 1037-1042. http://dx.doi.org/10.1248/bpb.32.1037
Yuan. W: Holland. SK; Cecil. KM; Dietrich. KN; Wessel. SD; Altave. M; Hornung. RW: Ris. MD;
Egelhoff. JC: Lanphear. BP (2006). The impact of early childhood lead exposure on brain
organization: A functional magnetic resonance imaging study of language function. Pediatrics 118:
971-977. http://dx.doi.org/10.1542/peds.2006-0467
Yiicesov. B; Turhan. A; lire. M; Jmir. T; Karakava. A. (1997a). Effects of occupational lead and cadmium
exposure on some immunoregulatory cytokine levels in man. Toxicology 123: 143-147.
http://dx.doi.org/10.1016/S0300-483X(97)00107-8
Yiicesov. B; Turhan. A; lire. M; Jmir. T; Karakava. A. (1997b). Simultaneous effects of lead and
cadmium on NK cell activity and some phenotypic parameters. Immunopharmacol Immunotoxicol 19:
339-348. http://dx.doi.org/10.3109/08923979709046980
November 2012 5-799 Draft - Do Not Cite or Quote
-------
Yun. SW: Hover. S. (2000). Effects of low-level lead on glycolytic enzymes and pyruvate dehydrogenase
of rat brain in vitro: Relevance to sporadic Alzheimer's disease? Journal of Neural Transmission 107:
355-368. http://dx.doi.org/10.1007/s007020050030
Zailina. H; Junidah. R; Josephine. Y; Jamal. HH. (2008). The influence of low blood lead concentrations
on the cognitive and physical development of primary school children in Malaysia. Asia Pac J Public
Health 20: 317-326. http://dx.doi.org/10.1177/1010539508322697
Zawia. NH; Basha. MR. (2005). Environmental risk factors and the developmental basis for Alzheimer's
disease [Review]. RevNeurosci 16: 325-337.
Zawia. NH; Harry. GJ. (1995). Exposure to lead-acetate modulates the developmental expression of
myelin genes in the rat frontal lobe. Int J Dev Neurosci 13: 639-644.
http://dx.doi.org/10.1016/0736-5748(95)00032-C
Zawia. NH: Sharan. R; Brydie. M; Oyama. T; Crumpton. T. (1998). Spl as a target site for metal-induced
perturbations of transcriptional regulation of developmental brain gene expression. Dev Brain Res
107: 291-298. http://dx.doi.org/10.1016/S0165-3806(98)00023-6
Zeisel. S: Blusztajn. J. (1994). Choline and human nutrition [Review]. Annu Rev Nutr 14: 269-296.
http://dx.doi.org/10.1146/annurev.nu.14.070194.001413
Zelikoff. JT; Li. JH; Hartwig. A; Wang. XW: Costa. M; Rossman. TG. (1988). Genetic toxicology of lead
compounds. Carcinogenesis 9: 1727-1732. http://dx.doi.Org/10.1093/carcin/9.10.1727
Zelikoff. JT; Parsons. E; Schlesinger. RB. (1993). Inhalation of paniculate lead oxide disrupts pulmonary
macrophage-mediated functions important for host defense and tumor surveillance in the lung.
Environ Res 62: 207-222. http://dx.doi.org/10.1006/enrs.1993.1106
Zeller. I; Knoflach. M; Seubert. A; Kreutmaver. S: Stelzmiiller. M; Wallnoefer. E; Blunder. S: Frotschnig.
S: Messner. B; Willeit. J: Debbage. P; Wick. G: Kiechl. S: Laufer. G: Bernhard. D. (2010). Lead
contributes to arterial intimal hyperplasia through nuclear factor erythroid 2-related factor-mediated
endothelial interleukin 8 synthesis and subsequent invasion of smooth muscle cells. Arterioscler
Thromb Vase Biol 30: 1733-1740. http://dx.doi.org/10.1161/ATVBAHA.110.211011
Zeng. H; Chattarji. S: Barbarosie. M; Rondi-Reig. L; Philpot. B; Miyakawa. T; Bear. M; Tonegawa. S.
(2001). Forebrain-specific calcineurin knockout selectively impairs bidirectional synaptic plasticity
and working/episodic-like memory. Cell 107: 617-629.
http://dx.doi.org/10.1016/S0092-8674(01)00585-2
Zentner. LE; Rondo. PH; Mastroeni. SS. (2006). Lead contamination and anthropometry of the newborn
baby. J Trap Pediatr 52: 369-371. http://dx.doi.org/10.1093/tropej/fml009
Zentner. LEA; Rondo. PHC: Duran. MC: Oliveira. JM. (2008). Relationships of blood lead to calcium,
iron, and vitamin C intakes in Brazilian pregnant women. Clin Nutr 27: 100-104.
http://dx.doi.0rg/10.1016/i.clnu.2007.10.005
Zeyrek. D; Soran. M; Cakmak. A; Kocyigit. A; Iscan. A. (2009). Serum copper and ainc levels in mothers
and cord blood of their newborn infants with neural tube defects: A case-control study. Indian Pediatr
46: 675-680.
Zhang. A; Hu. H; Sanchez. BN; Ettinger. AS: Park. SK; Cantonwine. D: Schnaas. L; Wright. RO:
Lamadrid-Figueroa. H; Tellez-Rojo. MM. (2011 a). Association between prenatal lead exposure and
blood pressure in female offspring. Environ Health Perspect 120: 445-450.
http://dx.doi.org/10.1289/ehp.1103736
Zhang. A; Park. SK; Wright. RO: Weisskopf. MG: Mukherjee. B; Nie. H; Sparrow. D: Hu. H. (2010a).
HFE H63D polymorphism as a modifier of the effect of cumulative lead exposure on pulse pressure:
The Normative Aging Study. Environ Health Perspect 118: 1261-1266.
http://dx.doi.org/10.1289/ehp. 1002251
November 2012 5-800 Draft - Do Not Cite or Quote
-------
Zhang. LF; Peng. SQ: Sheng. W: Li. BL; Gang. H; Dong. YS. (2007a). Direct effects of lead (Pb2+) on
the relaxation of in vitro cultured rat aorta to acetylcholine. Toxicol Lett 170: 104-110.
http://dx.doi.0rg/10.1016/i.toxlet.2007.02.004
Zhang. LF; Peng. SQ: Wang. S. (2005). Influence of lead (Pb2+) on the reactions of in vitro cultured rat
aorta to 5-hydroxytryptamine. Toxicol Lett 159: 71-82. http://dx.doi.Org/10.1016/i.toxlet.2005.04.009
Zhang. LF; Peng. SQ: Wang. S. (2009a). Decreased aortic contractile reaction to 5-hydroxytryptamine in
rats with long-term hypertension induced by lead (Pb2+) exposure. Toxicol Lett 186: 78-83.
http://dx.doi.0rg/10.1016/i.toxlet.2009.01.004
Zhao. Y; Wang. L; Shen. HB; Wang. ZX; Wei. QY; Chen. F. (2007). Association between delta-
aminolevulinic acid dehydratase (ALAD) polymorphism and blood lead levels: A meta-regression
analysis. J Toxicol Environ Health 70: 1986-1994. http://dx.doi.org/10.1080/15287390701550946
Zhao. ZY: Li. R; Sun. L; Li. ZY: Yang. RL. (2004). Effect of lead exposure on the immune function of
lymphocytes and erythrocytes in preschool children. J Zhejiang Univ Sci 5: 1001-1004.
http://dx.doi.org/10.1007/BF02947614
Zheng. C: Skold. M; Li. J: Nennesmo. I; Fadeel. B; Henter. J. (2007). VEGF reduces astrogliosis and
preserves neuromuscular junctions in ALS transgenic mice. Biochem Biophys Res Commun 363:
989-993. http://dx.doi.0rg/10.1016/i.bbrc.2007.09.088
Zhou. J; Xu. YH; Chang. HF. (1985). The effects of lead ion on immune function of rabbit alveolar
macrophages: Quantitation of immune phagocytosis and rosette formation by 51Cr in vitro. Toxicol
Appl Pharmacol 78: 484-487.
Zhu. M; Fitzgerald. EF; Gelberg. KH; Lin. S: Druschel. CM. (2010). Maternal low-level lead exposure
and fetal growth. Environ Health Perspect 118: 1471-1475. http://dx.doi.org/10.1289/ehp.0901561
Zimmermann. MB; Muthawa. S: Moretti. D; Kurpad. A; Hurrell. RF. (2006). Iron fortification reduces
blood lead levels in children in Bangalore, India. Pediatrics 117: 2014-2021.
http://dx.doi.org/10.1542/peds.2005-2440
Zoeger. N; Roschger. P: Hofstaetter. JG: Jokubonis. C: Pepponi. G: Falkenberg. G: Fratzl. P:
Berzlanovich. A; Osterode. W: Streli. C: Wobrauschek. P. (2006). Lead accumulation in tidemark of
articular cartilage. Osteoarthritis Cartilage 14: 906-913. http://dx.doi.Org/10.1016/j.joca.2006.03.001
Zuo. PJ; Qu. W: Cooper. RN; Gover. RA; Diwan. BA; Waalkes. MP (2009). Potential role of alpha-
synuclein and metallothionein in lead-induced inclusion body formation. Toxicol Sci 111: 100-108.
http://dx.doi.org/10.1093/toxsci/kfpl32
Zuscik. MJ; Ma. L; Buckley. T; Puzas. JE; Drissi. H; Schwarz. EM; O'Keefe. RJ. (2007). Lead induces
chondrogenesis and alters transforming growth factor-beta and bone morphogenetic protein signaling
in mesenchymal cell populations. Environ Health Perspect 115: 1276-1282.
http://dx.doi.org/10.1289/ehp.10028
November 2012 5-801 Draft - Do Not Cite or Quote
-------
6 POTENTIALLY AT-RISK POPULATIONS
1 The NAAQS are intended to protect public health with an adequate margin of safety. In
2 so doing, protection is provided for both the population as a whole and those groups
3 potentially at increased risk for health effects from exposure to the air pollutant for which
4 each NAAQS is set (Preface to this ISA). To facilitate the identification of populations at
5 increased risk for Pb-related health effects, studies have evaluated various factors that
6 may contribute to susceptibility and/or vulnerability to Pb. The definitions of
7 susceptibility and vulnerability vary across studies, but in most instances "susceptibility"
8 refers to biological or intrinsic factors (e.g., age, sex) while "vulnerability" refers to
9 nonbiological or extrinsic factors (e.g., socioeconomic status [SES]) (U.S. EPA. 2010.
10 2009a). Additionally, in some cases, the terms "at-risk" and "sensitive" populations have
11 been used to encompass these concepts more generally. In this ISA, "at-risk" groups are
12 defined as those with characteristics that increase the risk of Pb-related health effects in a
13 population. These characteristics include various factors, such as genetic background,
14 race and ethnicity, sex, age, diet, pre-existing disease, SES, and characteristics that may
15 modify exposure or the response to Pb.
16 Individuals, and ultimately populations, could experience increased risk for air pollutant
17 induced health effects via multiple avenues. A group with intrinsically increased risk
18 would have one or more factors that increase risk for an effect through a biological
19 mechanism. In general, people in this category would have a steeper concentration-
20 risk relationship, compared to those not in the category. Potential factors that are often
21 considered intrinsic include genetic background and sex. A group of people could also
22 have extrinsically increased risk, which would be through an external, non-biological
23 factor. Examples of extrinsic factors include SES and diet. Some groups are at risk of
24 increased internal dose at a given exposure concentration. In addition, some groups
25 could have greater exposure (concentration x time), regardless of the delivered dose.
26 Finally, there are those who might be placed at increased risk for experiencing a
27 greater exposure by being exposed at a higher concentration. For example, groups of
28 people living near Pb smelters.
29 Some factors described above are multifaceted and may influence the risk of an air
30 pollutant related health effect through a combination of avenues. For example, SES may
31 affect access to medical care, which itself may contribute to the presence of preexisting
32 diseases and conditions considered as intrinsic factors. Additionally, children's outdoor
33 activities can lead to more hand-to-mouth contact with contaminated soil than adults,
34 which leads to increased intake dose and exposure. At the same time, children have
November 2012 6-1 Draft - Do Not Cite or Quote
-------
1 biological (i.e., intrinsic) differences from adults that may influence their uptake,
2 metabolism, storage, and excretion.
3 The emphasis of this chapter is to identify and understand the factors that potentially
4 increase the risk of Pb-related health effects, regardless of whether the increased risk is
5 due to intrinsic factors, extrinsic factors, increased dose/exposure, or a combination due
6 to the often interconnectedness of factors. The following sections examine factors that
7 potentially lead to increased risk of Pb-related health effects and characterize the overall
8 weight of evidence for each factor.
9 Approach to Classifying Potential At-Risk Factors
10 To identify factors that potentially lead to some populations being at greater risk of
11 Pb-related health effects, the evidence across relevant scientific disciplines (i.e., exposure
12 sciences, dosimetry, toxicology, and epidemiology) was evaluated. In this systematic
13 approach, the collective evidence is used to examine coherence of effects across
14 disciplines and determine biological plausibility. The collective results across the
15 scientific disciplines comprise the overall weight of evidence that is used to determine
16 whether a specific factor results in a population being at increased risk of an air pollutant
17 related health effect. The first section of this chapter (Section 6.1) summarizes
18 physiological factors that influence Pb levels in the body. The second section of this
19 chapter (Section 6.2) summarizes information on factors potentially related to differential
20 Pb exposure. The studies presented in this section supplement the material provided in
21 Chapter 3_ and Chapter_4_by examining how factors such as age, sex, race and ethnicity,
22 SES, proximity to Pb sources, and residential factors may affect Pb exposure or blood Pb
23 levels. The third section of this chapter (Section 6.3) discusses the epidemiologic and
24 toxicological studies evaluated in Chapter_5 that provide information on factors
25 potentially related to increased risk of Pb-induced health effects. To examine whether Pb
26 differentially affects certain populations, epidemiologic studies conduct stratified
27 analyses to identify the presence or absence of effect measure modification. A thorough
28 evaluation of potential effect measure modifiers may help identify populations that are at
29 increased risk for Pb-related health effects. Highlighted studies include only those where
30 the population was stratified into subgroups (e.g., males versus females or smokers
31 versus nonsmokers) for comparative analysis. In the case of many biomarker studies and
32 the epidemiologic studies considered, this approach allowed for a comparison between
33 populations exposed to similar Pb concentrations and within the same study design.
34 Toxicological studies also provide evidence of Pb effects and biological plausibility for
35 factors that may lead to increased risk for Pb-related health effects. Included
36 toxicological studies may have categorized the study populations by different factors,
37 such as age, sex, diet/nutrition status, and genetics, or are those that examined animal
November 2012 6-2 Draft - Do Not Cite or Quote
-------
1 models of disease. These epidemiologic and toxicological studies provide the scientific
2 basis for an overall weight of the evidence evaluation for the identification of specific
3 populations potentially at risk of Pb-related health effects. Details on the magnitude of
4 effects for studies in this third section (Section 6.3) are included in summaries of the
5 studies presented in Chapter_5.
6 Numerous studies that focused on only one potentially at-risk population were described
7 in previous chapters (Chapter_5) but are not discussed in detail in this chapter because
8 they lacked stratified analysis with adequate comparison groups. For example, pregnancy
9 is a lifestage with potentially increased risk for mothers and fetuses, but because there are
10 no comparison groups for stratified analyses, these studies were presented in Chapter_5
11 but are not included here. Additionally, it is understood that some of the stratified
12 variables/factors discussed in this third section (Section 6.3) may not be effect measure
13 modifiers but instead may be mediators of Pb-related health effects. Mediators are factors
14 that fall on the causal pathway between Pb exposure and health outcomes, whereas effect
15 measure modifiers are factors that result in changes in the measured associations between
16 Pb exposure and health effects. Because mediators are caused by Pb exposure and are
17 also intermediates in the disease pathway that is studied, mediators are not correctly
18 termed "at-risk" factors. Some of the factors discussed in this third section could be
19 mediators and/or modifiers. These are noted in Table 6-5.
20 Building on the causal framework discussed in detail in the Preamble and used
21 throughout the ISA, conclusions are made regarding the strength of evidence for each
22 factor that may contribute to increased risk of a Pb-related health effect based on the
23 evaluation and synthesis of evidence across scientific disciplines. The conclusions drawn
24 considered the "Aspects to Aid in Judging Causality" discussed in Table I of the
25 Preamble. The categories considered for evaluating the potential increased risk of an air
26 pollutant-related health effect are "adequate evidence," "suggestive evidence,"
27 "inadequate evidence," and "evidence of no effect." They are described in more detail in
28 Table 6-1.
November 2012 6-3 Draft - Do Not Cite or Quote
-------
Table 6-1 Classification of evidence for potential at-risk factors.
Health Effects
Adequate
evidence
Suggestive
evidence
Inadequate
evidence
Evidence of no
effect
There is substantial, consistent evidence within a discipline to conclude that a factor results
in a population or lifestage being at increased risk of air pollutant-related health effect(s)
relative to some reference population or lifestage. Where applicable this includes
coherence across disciplines. Evidence includes multiple high-quality studies.
The collective evidence suggests that a factor results in a population or lifestage being at
increased risk of an air pollutant-related health effect relative to some reference population
or lifestage, but the evidence is limited due to some inconsistency within a discipline or,
where applicable, a lack of coherence across disciplines.
The collective evidence is inadequate to determine if a factor results in a population or
lifestage being at increased risk of an air pollutant-related health effect relative to some
reference population or lifestage. The available studies are of insufficient quantity, quality,
consistency, and/or statistical power to permit a conclusion to be drawn.
There is substantial, consistent evidence within a discipline to conclude that a factor does
not result in a population or lifestage being at increased risk of air pollutant-related health
effect(s) relative to some reference population or lifestage. Where applicable this includes
coherence across disciplines. Evidence includes multiple high-quality studies.
6.1 Physiological Factors that Influence the Internal
Distribution of Pb
1 Blood and bone Pb measures are influenced to varying degrees by biokinetic processes
2 (e.g., absorption, distribution, metabolism, excretion), which are discussed in detail in
3 Chapter_4. These processes can be affected by multiple factors, such as age, genetics,
4 diet, and co-exposure with other metals and non-metals.
5 Age influences the biokinetic response to Pb within the body. Infants may be considered
6 an at-risk population because Pb easily crosses the placental barrier and accumulates in
7 fetal tissue during gestation (Pillai et al., 2009; Wang et al., 2009e; Uzbekov et al., 2007).
8 This transfer of Pb from mother to fetus is partly due to the remobilization of the
9 mother's bone stores (O'Flaherty. 1998; Franklin et al., 1997). This also results in
10 increased maternal blood Pb levels (Lamadrid-Figueroa et al.. 2006; Gulson et al.. 2004a;
11 Hertz-Picciotto et al.. 2000; Gulson etal.. 1997; Lagerkvist et al., 1996; Schuhmacher et
12 al.. 1996; Rothenberg et al.. 1994a). Bone growth rate is high during childhood. The
13 majority of a child's Pb body burden is not permanently incorporated in the bone, but
14 some Pb does remain in the bone until older age (McNeill et al.. 2000; O'Flaherty. 1995;
15 Leggett. 1993). Older adults are more likely to have age-related degeneration of bones
16 and organ systems and a possible redistribution of Pb stored in the bones into the blood
17 stream (Popovic et al., 2005; Garrido Latorre et al., 2003; Gulson et al., 2002).
November 2012
6-4
Draft - Do Not Cite or Quote
-------
1 Various genes can also affect Pb biomarker concentrations. Genetic variants of the
2 vitamin D receptor (VDR) in humans have been associated with varied bone and plasma
3 Pb levels (Rezende et al. 2008; Theppeang et al.. 2004; Schwartz et al. 2000a). Multiple
4 studies have also examined the association between the aminolevulinate dehydratase
5 (ALAD) polymorphism and blood Pb levels and found that the ALAD-2 polymorphism
6 may be biologically related to varying Pb levels, although some studies report no
7 difference for ALAD alleles (see also Section 5.2.3) (Mivaki et al., 2009; Shaik and
8 Jamil. 2009: Sobin et al.. 2009: Chen et al.. 2008c: Rabstein et al.. 2008: Scinicariello et
9 al.. 2007: Zhao et al.. 2007: Montenegro et al.. 2006: Wananukul et al.. 2006).
10 It is well established that diets sufficient in minerals such as calcium, iron, and zinc offer
11 some protection from Pb exposure by preventing or competing with Pb for absorption in
12 the GI tract. A study in China reported that children who regularly consumed breakfast
13 had lower blood Pb levels than those children that did not eat breakfast (Liu et al..
14 201 la). Diets designed to limit or reduce caloric intake and induce weight loss have been
15 associated with increased blood Pb levels in adult animals (Han et al.. 1999). A
16 toxicological study reported negative effects of Pb on osmotic fragility, TEARS
17 production, catalase activity, and other oxidative parameters, but most of these effects
18 were reduced to the levels observed in the control group when the rats were given
19 supplementation of zinc and vitamins (Masso-Gonzalez and Antonio-Garcia. 2009).
20 Toxicological studies by Jamieson et al. (2008: 2006) also reported that a zinc-deficient
21 diet increases bone and renal Pb content and impairs skeletal growth and mineralization.
22 A zinc-supplemented diet attenuated bone and renal Pb content. Toxicological studies
23 have shown that dietary deficiency of calcium induces increased Pb absorption and
24 retention (Fullmer. 1992: Mykkanen and Wasserman. 1981: SixandGoyer. 1970).
25 Increased calcium intake reduces accumulation of Pb in bone and mobilization of Pb
26 during pregnancy and lactation (Bogden et al.. 1995). Additionally, studies have reported
27 that iron deficiencies may result in higher Pb absorption or altered biokinetics (Schell et
28 al.. 2004: Marcus and Schwartz. 1987: Mahaffev and Annest. 1986).
29 In summary, age, genetics, and diet affect the biokinetics of Pb, which in turn affects the
30 internal distribution of Pb. These factors were discussed in greater detail in Chapter_4
31 where more information on overall biokinetic and physiological factors affecting Pb
32 distribution is provided.
November 2012 6-5 Draft - Do Not Cite or Quote
-------
6.2 Population Characteristics Potentially Related to Differential
Pb Exposure
1 Elevated or differential Pb exposure and related biomarker levels (such as blood Pb),
2 have been shown to be statistically related to several population characteristics, including
3 age, sex, race and ethnicity, SES, proximity to Pb sources, and residential factors (U.S.
4 EPA, 2006b). In most cases, exposure, absorption, and biokinetics of Pb are all
5 influenced to varying degrees by such characteristics. Additionally, the relative
6 importance of such population characteristics in affecting exposure, absorption, and
7 biokinetics varies among individuals in a population and is difficult to quantify. This
8 section presents recent studies demonstrating a relationship between each population
9 characteristic and exposure status. The studies presented in this section build upon the
10 current body of literature suggesting that population characteristics differentially
11 influence Pb exposure; the new literature does not alter previous understanding of the
12 differential influence of population characteristics on Pb exposure. Differential response
13 to given Pb exposures is discussed in Section 6.3.
6.2.1 Age
6.2.1.1 Early Childhood
14 Typically, children have increased exposure to Pb compared with adults because
15 children's behaviors and activities include increased hand-to-mouth contact, crawling,
16 and poor hand-washing that typically result in increased ingestion compared with adults
17 (U.S. EPA. 2006b). Children can also have increased Pb exposure because outdoor
18 activities can lead to hand-to-mouth contact with contaminated soil. For example, Zahran
19 et al. (2010) observed that a 1% reduction in soil Pb concentration led to a 1.55 (ig/dL
20 reduction in median blood Pb levels (p <0.05) among New Orleans children.
21 Age of the children may influence blood Pb levels through a combination of behavioral
22 and biokinetic factors. The 2009-2010 NHANES data are presented in Table 6-2 by age
23 and sex. Among children, highest blood Pb levels occurred in the 1-5 year age group
24 (children under age 1 were not included), and within this subgroup (not shown on the
25 table), 1-year old children had the highest blood Pb levels (99th percentile: 9.47 ug/dL)
26 (NCHS. 2010). It is possible that high blood Pb levels among these young children may
27 also be related to in utero exposures resulting from maternal Pb remobilization from bone
28 stores from historic exposures (Miranda et al.. 2010) or from contemporaneous Pb
29 exposures if the mothers had appreciable current Pb exposure. Jones et al. (2009a)
November 2012 6-6 Draft - Do Not Cite or Quote
-------
1 analyzed the NHANES datasets for the years 1999-2004 to study trends in blood Pb
2 among two different age groups of children over time (see Table 6-3). They observed
3 greater percentages of children aged 1-2 years having blood Pb levels of 2.5 to <5 (ig/dL,
4 5 to <7.5 (ig/dL, and > 10 (ig/dL, compared with 3-5 year-old children, but no age
5 difference was noted for the 7.5 to <10 (ig/dL bracket. At the same time, 1-2 year-old
6 children had lower percentages of blood Pb levels <1 (ig/dL and 1 to <2.5 (ig/dL
7 compared with children ages 3-5 years old. This implies that there is a shift in the
8 distribution of blood Pb levels as children age, even during early childhood. These
9 distribution differences may be attributable to differences in exposure (including
10 behavioral influences, such as hand-to-mouth contact and crawling in proximity to
11 contaminated surfaces), residual contributions from the mother's Pb burden, age-
12 dependent variability in biokinetics or diet (e.g., milk versus solid diets). Yapici et al.
13 (2006) studied the relationship between blood Pb level and age among a cohort of
14 children between 6 and 73 months of age with elevated blood Pb levels (87.6% of study
15 group with blood Pb greater than 20 (ig/dL) living near a Turkish coal mine. They
16 observed a low but statistically significant negative correlation between blood Pb and age
17 (r = -0.38,p<0.001).
November 2012 6-7 Draft - Do Not Cite or Quote
-------
Table 6-2 Blood Pb levels by age and sex, 2009-2010 NHANES.
Age
1-5 yr
6-11 yr
12-19 yr
20-59 yr
60+ yr
Overall
Sex
Total
Male
Female
Total
Male
Female
Total
Male
Female
Total
Male
Female
Total
Male
Female
Total
Male
Female
N
836
429
407
1009
521
488
1183
632
551
3856
1843
2013
1909
941
968
8793
4366
4427
Avg.
1.61
1.59
1.64
1.05
1.10
0.99
0.84
0.98
0.69
1.50
1.88
1.15
2.09
2.46
1.73
1.50
1.75
1.25
Std. Dev.
1.49
1.32
1.65
0.74
0.73
0.75
0.68
0.69
0.62
1.83
2.33
1.10
1.51
1.78
1.07
1.57
1.88
1.13
5%
0.53
0.51
0.54
0.42
0.45
0.38
0.33
0.40
0.30
0.44
0.56
0.40
0.72
0.87
0.65
0.43
0.50
0.39
25%
0.85
0.83
0.90
0.61
0.66
0.58
0.50
0.58
0.44
0.72
0.92
0.61
1.16
1.39
1.01
0.72
0.84
0.63
50%
1.21
1.22
1.20
0.83
0.88
0.79
0.69
0.80
0.57
1.08
1.37
0.89
1.69
1.99
1.43
1.10
1.29
0.96
75%
1.81
1.84
1.77
1.22
1.30
1.12
0.96
1.11
0.79
1.70
2.12
1.35
2.53
2.90
2.14
1.76
2.05
1.48
95%
4.00
4.09
3.69
2.36
2.37
2.35
1.82
2.09
1.31
3.53
4.49
2.63
4.79
5.56
3.75
3.66
4.31
2.97
99%
8.03
7.49
9.59
4.29
4.18
3.98
3.10
3.91
2.25
7.27
9.68
4.41
8.28
9.89
5.42
7.21
8.62
5.17
Source: (NCHS. 2010)
November 2012
6-8
Draft - Do Not Cite or Quote
-------
Table 6-3
Pb Units:
M9/dL
(95% Cl)
Overall 2
Sex
Female 1
Male 1
Age
1-2 yr 1
3-5 yr 1
Race/Ethnicity
Non-
Hispanic
Black
Mexican
American
Non-
Hispanic
White
Percentage of children within six categories/brackets of blood Pb
levels, 1999-2004 NHANES.
7.5 to
Geometric 1to<2.5 2.5to<5 5to<7.5 <10 >10
N mean <1 |jg/dL, % |jg/dL,% |Jg/dU% MO/dU % MO/dU % |jg/dL,%
,532 1.9(1.8-2.0) 14.0 55.0 23.6 4.5 1.5 1.4
(11.6-16.6) (52.1-57.9) (21.1-26.1) (3.3-5.9) (1.0-2.1) (1.0-2.0)
,211 1.9(1.7-2.0) 14.1 54.5 23.9 4.5 1.4 1.7
(10.8-17.7) (51.1-57.8) (20.3-27.8) (3.3-5.8) (0.8-2.3) (0.9-2.6)
,321 1.9(1.7-2.0) 14.0 55.5 23.2 4.6 1.5 1.3
(11.4-16.7) (51.4-59.5) (20.3-26.3) (3.0-6.5) (0.9-2.3) (0.7-2.6)
,231 2.1 (2.0-2.2) 10.6 51.0 27.9 6.7 1.4 2.4
(7.7-13.9) (46.7-55.3) (24.9-31.0) (5.0-8.6) (0.8-2.2) (1.4-3.5)
,301 1.7(1.6-1.9) 16.2 57.6 20.7 3.1 1.5 0.9
(12.9-19.9) (53.8-61.4) (17.9-23.7) (1.9-4.6) (0.8-2.3) (0.4-1.5)
755 2.8 (2.5-3.0) 4.0 (2.5-5.7) 42.5 36.2 9.4 4.6 3.4
(37.8-47.2) (33.1-39.3) (6.9-12.2) (3.0-6.5) (1.8-5.5)
812 1.9(1.7-2.0) 10.9 61.0 22.1 3.4 1.3 1.2
(8.6-13.4) (56.9-65.1) (18.0-26.5) (2.2-5.0) (0.6-2.2) (0.4-2.6)
731 1.7(1.6-1.8) 17.6 57.1 19.7 3.6 0.8 1.2
(14.0-21.5) (52.4-61.7) (16.1-23.5) (1.9-5.8) (0.3-1.6) (0.6-2.0)
Poverty-Income Ratio (PIR)
1
2
3
4
5
6
£1.3 1
>1.3 1
Source: Reprinted
,302 2.4(2.2-2.5) 6.7(4.6-9.2) 49.3 32.5 6.9 2.8 1.8
(44.9-53.7) (28.6-36.4) (2.2-8.8) (1.7-4.1) (1.1-2.7)
,070 1.5(1.4-1.6) 19.9 60.4 16.0 2.3 0.6 0.8
(16.3-23.8) (56.9-63.8) (12.9-19.3) (1.2-3.7) (0.1-1.4) (0.3-1.6)
with permission of the American Academy of Pediatrics; Jones et al. (2009a)
Fetal and child Pb biomarkers have been demonstrated to relate to maternal Pb
biomarkers as reported in the 2006 Pb AOCD (U.S. EPA. 2006b). Kordas et al. (2010)
observed that maternal hair Pb concentration was a statistically significant predictor of
child hair Pb concentration ((3 = 0.37 ± 0.07, p <0.01). Elevated blood Pb levels among
mothers present a potential exposure route to their children in utero or through breast
milk; see Miranda et al. (2010).
November 2012
6-9
Draft - Do Not Cite or Quote
-------
6.2.1.2 Older Adulthood
1 Blood Pb levels tend to be higher in older adults compared with the general adult
2 population, as described in the 2006 Pb AQCD (U.S. EPA. 2006b). Table 6-2 presents
3 2009-2010 NHANES data broken down by age group and shows that blood Pb levels
4 were highest in the among participants 65 years old or older, in comparison with adults
5 aged 20-64 years and with adolescents. In a study of blood Pb and saliva Pb in a mostly
6 female population in Detroit, Nriagu et al. (2006) found that age was a statistically
7 significant positive predictor of blood Pb (p <0.001). Average blood Pb levels among 14-
8 to 24-year-old subjects was 2.60 ± 0.16 ug/dL compared with 4.29 ± 0.56 ug/dL among
9 subjects aged 55 years or older. Higher average and median levels among older adults
10 could potentially be due to a shared experience of higher historical Pb exposures stored in
11 bone in conjunction with remobilization of stored Pb during bone loss (Section 4.2).
12 Theppeang et al. (2008b) studied Pb concentrations in the blood, tibia, and patella of
13 subjects age 50-70 as part of the Baltimore Memory Study. They found a statistically
14 significant relationship between age and tibia Pb ((3 = 0.37, p <0.01 in a model including
15 age, race/ethnicity, Yale energy index, and 2 diet variables; (3 = 0.57, p <0.01 in a model
16 including age, sex, and an interaction term for sex and age, which was also statistically
17 significant at p = 0.03). Theppeang et al. (2008b) also noted that patella Pb
18 concentrations were also positively associated with age, although the authors did not
19 present the data or significance levels. A statistically significant relationship was not
20 observed between the log-transform of blood Pb and age ((3 = 0.007, p = 0.11), although
21 the age range of subjects may not have been sufficient to discern a difference in blood Pb
22 level.
23 Miranda et al. (2010) observed that older pregnant women (ages 30-34 years and 35-39
24 years) had statistically significant higher odds of having greater blood Pb levels than
25 younger pregnant women (25- to 29-year-olds) in the reference age category. These
26 results could be related to a historical component to Pb exposure among mothers. These
27 findings were also consistent with observations that Pb storage in bones increased with
28 age before subsequent release with bone loss occurring during pregnancy, as described in
29 Section 4.2 and summarized in Section 6.1.
November 2012 6-10 Draft - Do Not Cite or Quote
-------
6.2.2 Sex
1 The AQCD (U.S. EPA. 2006b) described several studies showing higher Pb biomarker
2 levels in male adults compared with female adults. The 2009-2010 NHANES showed
3 that overall, males have significantly higher blood Pb levels (average: 1.75 ug/dL) than
4 females (average: 1.25 ug/dL) (p <0.0005). Among adults aged 20-59 years, average
5 blood Pb levels were 64% higher for males compared with females (p <0.0005). Among
6 adults 60 years or older, average blood Pb levels were 30% higher for males compared
7 with females (p <0.0005) (NCHS. 2010). In their study of Pb burden among Baltimore
8 adults aged 50-70 years, Theppeang et al. (2008b) observed that average blood Pb levels
9 were statistically significantly higher (p <0.01) among men (4.4 (ig/dL) than women
10 (3.1 (ig/dL). For average tibia Pb levels, Theppeang et al. (2008b) noted no difference
11 (p = 0.12) between men (18.0 (ig/g) and women (19.4 (ig/g).
12 Among U.S. children, the 2009-2010 NHANES data showed that blood Pb levels were
13 higher among girls than boys for the 1- to 5-years age group (Table 6-2). Blood Pb levels
14 became slightly higher among boys for the 6- to 11-years age group, and levels were
15 substantially higher among adolescent males than females 12- to 19-years old. The
16 2009-2010 NHANES data suggest that sex-based differences in blood Pb levels are not
17 substantial until adolescence.
6.2.3 Race and Ethnicity
18 Higher blood Pb and bone Pb levels among African Americans have been well
19 documented (U.S. EPA. 2006b). Model results presented in the 2006 Pb AQCD have
20 demonstrated not just elevations in blood Pb among African Americans but also
21 significant associations between blood Pb and race (U.S. EPA. 2006b). Recent studies are
22 consistent with those previous findings. For instance, Levin et al. (2008) and Jones et al.
23 (2009a) both analyzed NHANES survey data to examine trends in childhood blood Pb
24 levels. Data from the Jones et al. (2QQ9a) study, using NHANES data (NCHS. 2009.
25 2008) from 1988-1991 and 1999-2004 are shown in Figure 6-1. The authors found that
26 differences among children from different racial/ethnic groups with regard to the
27 percentage with blood Pb levels > 2.5 ug/dL over the period 1999-2004 have decreased
28 since the period of 1988-1991. The non-Hispanic black group still had higher percentages
29 with blood Pb levels > 2.5 ug/dL compared with non-Hispanic whites and Mexican
30 Americans, with large observable differences for blood Pb levels between 2.5 and
31 <10 ug/dL. It is notable that the distributions of blood Pb levels among Mexican
32 American and non-Hispanic white children were nearly identical in the 1999-2004
33 dataset. Theppeang et al. (2008b) also explored the effect of race and ethnicity on several
November 2012 6-11 Draft - Do Not Cite or Quote
-------
1 Pb biomarkers in a study of older adults living in Baltimore, MD. They observed a
2 statistically significant difference between African American (AA) and Caucasian (C)
3 subjects with respect to tibia Pb (AA: 21.8 (ig/g, C: 16.7 (ig/g, p <0.01) but not patella Pb
4 (AA: 7.1 ng/g, C: 7.1 ng/g, p = 0.46) or blood Pb levels (AA: 3.6 ng/dL, C: 3.6 ng/dL,
5 p = 0.69). Greater tibia (but lower patella) Pb levels may indicate greater historical
6 exposure among African Americans compared to Caucasians in the Baltimore population
7 studied by Theppeang et al. (2008b).
November 2012 6-12 Draft - Do Not Cite or Quote
-------
70
60 -
C 50
OJ
5 40
M—
O
£ 30
OJ
u
i_
o! 20
10 -
1988-1991
<1 l-<2.5 2.5-< 5 5-<7.5 7.5-<10 > 10
1999 - 2004
(U
1_
•a
u
M-
o
(U
u
70 n
60 -
50 -
40 -
30 -
20
10 -
o 4-
<1 l-<2.5 2.5-<5 5-<7.5 7.5 - <10 > 10
Blood Pb Level (ug/dL)
•••+•• Non-Hispanic black^^^Mexican American —^— Non-Hispanic white
Note: from the NHANES survey, 1988-1991 (top) and 1999-2004 (bottom).
Data used with permission of the American Academy of Pediatrics, Jones et al. (2009a)
Figure 6-1 Percent distribution of blood Pb levels by race/ethnicity among
U.S. children (1-5 years).
November 2012
6-13
Draft - Do Not Cite or Quote
-------
1 Differences in potential exposure among ethnic and racial groups have also been noted in
2 a study in the greater metropolitan New Orleans area. Campanella and Mielke (2008)
3 found that, in Census blocks where surface soil Pb levels were less than 20 mg/kg, the
4 population was 36% black, 55% white, 3.0% Asian, and 6.0% Hispanic, based on the
5 2000 Census, with the percentage based on the total number living in Census blocks with
6 the same soil Pb levels. In contrast, they found that for Census blocks in which soil Pb
7 levels were between 1,000 and 5,000 mg/kg, the population was 62% black, 34% white,
8 1% Asian, and 4% Hispanic (Figure 6-2), although the total population size generally
9 declined with soil Pb concentration, with the Census blocks with soil Pb of 1,000-5,000
10 mg/kg having less than half the population of that in the <20 mg/kg blocks. As described
11 in Section 6.2.4. the differences observed by Campanella and Mielke (2008) may also be
12 attributable to SES factors, or SES may be a confounding factor in the relationship
13 between Pb soil levels and race/ethnicity of nearby residents.
November 2012 6-14 Draft - Do Not Cite or Quote
-------
50,000 -i
45,000
40,000
35,000
30,000
o
| 25,000
Q.
O
°~ 20,000
15,000
10,000
5,000 -
0 -
i— I
Jl
—
Jl
i— i
Jl
-n
_n
• Black
• White
D Asian
D Hispanic
^ ^ In
V «,?
Soil Pb Concentration (mg/kg)
Note: By Census 2000 race/ethnicity demographic groups.
Source: Data used with permission of Springer Science; Campanella and Mielke (2008).
Figure 6-2 Soil Pb concentration exposure among the population of three
parishes within greater metropolitan New Orleans.
November 2012
6-15
Draft - Do Not Cite or Quote
-------
6.2.4 Socioeconomic Status (SES)
1 Socioeconomic factors have sometimes been associated with Pb exposure biomarkers.
2 Previous results reported in the 2006 Pb AQCD found negative associations between
3 income or other SES metrics and blood Pb, although these relationships were not always
4 statistically significant (U.S. EPA. 2006b). Nriagu et al. (2006) performed a multiple
5 regression analysis of blood Pb and saliva Pb levels on various Socioeconomic,
6 demographic, and exposure variables among an adult population in Detroit, Michigan.
7 Blood and saliva Pb were both used as indicators of Pb in unbound plasma that is
8 available to organs. Nriagu et al. (2006) found that education (p <0.001), income
9 (p <0.001), and employment status (p = 0.04) were all statistically significant predictors
10 of blood Pb levels, with blood Pb decreasing with some scatter as education and income
11 level increased. Statistically significant relationships were also reported by Nriagu et al.
12 (2006) for saliva Pb level with respect to education (p <0.001), income (p <0.001), and
13 employment (p = 0.06). However, the highest educational attainment and income
14 categories had higher saliva Pb levels compared with other groups; Nriagu et al. (2006)
15 attributed these inconsistencies to small sample sizes among the high educational
16 attainment and income categories.
17 On a national level, the difference in blood Pb levels that have historically been seen to
18 exist between different income levels has been decreasing. For example, Levin et al.
19 (2008) cited 1991 -1994 NHANES data [analyzed in Pirkle et al. (1994)1 that the
20 percentage of children aged 1-5 years with blood Pb levels > 10 ug/dL was 4.5% for the
21 lowest income group compared with 0.7% for the highest income group. Levin et al.
22 (2008) also analyzed data from the 1999-2002 NHANES and found no statistically
23 significant difference between the percent of children with blood Pb levels above
24 10 ug/dL for Medicaid-enrolled children (1.7%) compared with non-enrolled children
25 (1.3%). However, Medicaid-enrolled children did have higher median blood Pb levels
26 (2.6 ug/dL) compared to children not enrolled in Medicaid (1.7 ug/dL). Adding data for
27 2003-2004 to the analysis (i.e., for 1999-2004), widened the difference between Medicaid
28 enrolled and non-enrolled children with regard to percentage having blood Pb levels
29 > 10 ug/dL (1.9% versus 1.1%), but the difference was still not statistically significant (p
30 >0.05) and median blood Pb levels for the two groups did not change (Levin et al., 2008).
31 Likewise, Jones et al. (2009a) analyzed blood Pb levels with respect to poverty-income
32 ratio (PIR), which is the ratio of family income to the poverty threshold appropriate for a
33 given family size. They found statistically significant differences in median blood Pb for
34 PIR < 1.3 compared with PIR >1.3. The percentage of 1- to 5-year-old children having
35 blood Pb > 10 ug/dL was higher for PIR < 1.3 (1.8 versus 0.8); however, this difference
36 was not statistically significant. Additionally, in residential areas of metropolitan New
37 Orleans with soil concentrations below 20,000 mg/kg, Campanella and Mielke (2008)
November 2012 6-16 Draft - Do Not Cite or Quote
-------
1 observed a linear increase in surface soil Pb concentration with decreasing median
2 household income, suggesting a relationship of potential exposure with household
3 income. The census block-averaged median household income in areas with soil Pb
4 between 2.5 and 20 mg/kg was $40,000 per year, while the corresponding median income
5 in areas with soil Pb between 5,000 and 20,000 mg/kg was $24,000 per year. The highest
6 soil concentrations (20,000 mg/kg and above) was associated with a median income of
7 $27,000.
6.2.5 Proximity to Pb Sources
8 Air and soil Pb concentrations are higher in some industrialized and urbanized areas, as
9 described in Sections 3.2. 3.3. 3.5 and 4.1. as a result of historical and contemporary Pb
10 sources. The highest air Pb concentrations measured using the Pb-TSP monitoring
11 network have been measured at monitors located near sources emitting Pb. Elevated soil
12 Pb concentrations have also been measured in urbanized areas compared with less
13 urbanized or rural locations (Filippelli et al.. 2005). Air Pb concentrations exhibit high
14 spatial variability even at low concentrations (-0.01 ug/m3) (Martuzevicius et al.. 2004).
15 Proximity to an industrial source likely contributes to higher Pb exposures, as described
16 in the 2006 Pb AQCD (U.S. EPA. 2006^) for several studies of Superfund and other
17 industrial sites. This is consistent with the observation of higher air concentrations at
18 source oriented Pb monitoring sites compared with non-source oriented sites in the
19 2008-2010 data presented in Section 3.5.
20 Jones et al. (2010) found that neonates born near a Pb-contaminated hazardous waste site
21 had significantly higher umbilical cord blood Pb levels (median: 2.2 ug/dL [95% CI: 1.5,
22 3.3 ug/dL]) compared with a reference group of neonates not living near a potentially
23 contaminated site (median: 1.1 ug/dL [95% CI: 0.8, 1.3 ug/dL]), suggesting that
24 Pb-contaminated hazardous waste sites contribute to neonatal Pb levels. The population
25 studied in Jones et al. (2010) was 88% African American; 75% had a high school degree
26 or equivalent, while 20% had a college degree and 5% attended but did not graduate from
27 high school. However, Jones et al. (2010) did not analyze covariation between exposure
28 and maternal characteristics, so it cannot be determined if differences in characteristics
29 among the maternal groups (which did and did not report nearby hazardous waste sites)
30 confounded these results.
31 Studies have suggested that concentration of Pb in soil, a potential exposure media, is
32 related to land use type and historical sources, as described in Section 3.6.1. For instance,
33 Wu et al., (2010) observed that bioavailable Pb concentrations in Los Angeles surface
34 soil samples were significantly associated with traffic-related variables and parcel age
November 2012 6-17 Draft - Do Not Cite or Quote
-------
1 (i.e., length of time since the parcel was first developed), with parcel age being a highly
2 significant predictor of bioavailable soil Pb in most models (p <0.0001). Zahran et al.
3 (2010) observed that surface soil Pb levels in 46 Census tracts of metropolitan New
4 Orleans dropped following Hurricanes Katrina and Rita, from 330 mg/kg to 200 mg/kg
5 (averages of median measurements across all Census tracts for 2000 and 2006) and
6 attributed this observation to coverage by relatively cleaner river sediments. Blood Pb
7 levels obtained from children (ages 0-6 years) also declined subsequent to the hurricanes;
8 statistical modeling of the changes in soil and blood Pb estimated the decline to be
9 1.55 (ig/dL for each 1% reduction in soil Pb (p < 0.05).
6.2.6 Residential Factors
10 Findings from a recent study of the association between blood Pb and housing factors by
11 Dixon et al. (2009). which analyzed data from the NHANES national survey for
12 1999-2004, are consistent with those from previous studies presented in the
13 2006 Pb AQCD that observed positive associations between increased blood Pb and
14 increased house dust Pb levels (U.S. EPA. 2006b; Lanphear et al.. 1998; Laxen et al..
15 1987). Dixon et al. (2009) used NHANES data from 1999-2004 to perform a linear
16 regression of blood Pb among children 12-60 months old on several factors including
17 year of home construction, floor surface condition, floor dust Pb level, windowsill dust
18 Pb level, and renovation in homes built before 1978. They found that blood Pb (log
19 transformed) was significantly associated with homes being built after 1950 (p = 0.014),
20 windowsill Pb level (p = 0.002), dust Pb level (p <0.001), and occurrence of renovation
21 in pre-1978 homes (p = 0.045). Detailed results of this regression are shown in Table 6-4.
22 As part of the same study, Gaitens et al. (2009) performed a regression analysis of floor
23 dust Pb (PbD) and windowsill dust Pb on several factors. Floor dust Pb (log transformed)
24 was significantly associated with the following housing-related factors: floor surface
25 condition (p <0.001), windowsill dust Pb (log transformed) (p O.001), year of
26 construction (p <0.001), and renovation in a pre-1950 home (p <0.001). Windowsill dust
27 Pb (log transformed) was significantly associated with the following housing-related
28 factors: year of construction (p O.001), window surface condition (0.001), and
29 deteriorated indoor paint (p = 0.028).
November 2012 6-18 Draft - Do Not Cite or Quote
-------
Table 6-4 Regression of log-transformed blood Pb level of children 12-60
months old on various factors related to housing condition, from
1999-2004 NHANES dataset.
Variables
Overall p-value Levels3
Estimate (SE)
p-Value
Intercept
Age (in years)
Year of construction
PIR
Race/ethnicity
Country of birth
Floor surface/condition x log floor PbD
Floor surface/condition x (log floor PbD)2
Floor surface/condition x (log floor PbD)3
Log windowsill PbD
Home-apartment type
Anyone smoke inside the home
Log cotinine concentration (ng/dL) in blood
Window cabinet or wall renovation in a
pre-1 978 home
0.172
<0.001 Age
Age2
Age3
Age4
0.014 Intercept for missing
1990-present
1978-1989
1960-1977
1950-1959
1940-1949
Before 1940
<0.001 Intercept for missing
Slope
<0.001 Non-Hispanic white
Non-Hispanic black
Hispanic
Other
0.002 Missing
U.S."
Mexico
Elsewhere
<0.001 Intercept for missing
Not smooth and cleanable
Smooth and cleanable or carpeted
Not smooth and cleanable
Smooth and cleanable or carpeted
Uncarpeted not smooth and cleanable
Smooth and cleanable or carpeted
0.002 Intercept for missing
Slope
<0.001 Intercept for missing
Mobile home or trailer
One family house detached
One family house attached
Apartment (1-9 units)
Apartment (a 10 units)
0.015 Missing
Yes
No
0.004 Intercept for missing
Slope
0.045 Missing
Yes
No
-0.517 (0.373)
2.620 (0.628)
-1.353(0.354)
0.273 (0.083)
-0.019 (0.007)
-0.121 (0.052)
-0.198(0.058)
-0.196(0.060)
-0.174(0.056)
-0.207 (0.065)
-0.012 (0.072)
0.000
0.053 (0.065)
-0.053 (0.012)
0.000
0.247 (0.035)
-0.035 (0.030)
0.128(0.070)
-0.077 (0.219)
0.000
0.353 (0.097)
0.154(0.121)
0.178(0.094)
0.386 (0.089)
0.205 (0.032)
0.023(0.015)
0.027 (0.008)
-0.020 (0.014)
-0.009 (0.004)
0.053 (0.040)
0.041 (0.011)
-0.064 (0.097)
0.127(0.067)
-0.025 (0.046)
0.000
0.069 (0.060)
-0.133(0.056)
0.138(0.140)
0.100(0.040)
0.000
-0.150(0.063)
0.039(0.012)
-0.008(0.061)
0.097 (0.047)
0.000
0.172
<0.001
<0.001
0.002
0.008
0.024
0.001
0.002
0.003
0.003
0.870
—
0.420
<0.001
—
<0.001
0.251
0.073
0.728
-
<0.001
0.209
0.065
<0.001
<0.001
0.124
0.001
0.159
0.012
0.186
<0.001
0.511
0.066
0.596
—
0.256
0.022
0.331
0.015
—
0.023
0.002
0.896
0.045
-
"Children: n = 2,155 (age 10-60 months); R2 = 40%
""Includes the 50 states and the District of Columbia
Source: Dixon et al. (2009).
November 2012
6-19
Draft - Do Not Cite or Quote
-------
1 Renovation activities on older homes have been shown to produce excess Pb dust
2 concentrations. Gaitens et al. (2009) performed a regression analysis on dust Pb
3 concentrations from 1994-2004 NHANES on demographic and housing variables and
4 found that renovation of windows, cabinets, or walls in a pre-1950 home was
5 significantly associated with floor dust Pb concentration (p <0.001). Paint scraping within
6 the last twelve months was nearly significantly associated with windowsill dust Pb
7 concentration (p = 0.053). Dixon et al. (2009) performed a regression analysis on log-
8 transformed blood Pb levels from NHANES (1999-2004) on several demographic and
9 housing variables and found that renovation of windows, cabinets, or walls in pre-1978
10 homes was significantly associated with blood Pb concentration (p = 0.045). A case study
11 by Mielke et al. (2001) reports on elevated indoor and outdoor dust Pb levels at two
12 houses where exterior paint has been either power sanded (without confinement of
13 released material) or hand scraped (with collection of released material) to prepare for
14 repainting. The latter approach appeared to yield lower dust Pb levels, although given the
15 extremely limited dataset, conclusions are uncertain. In an occupational study of men
16 performing home renovations in the U.K., window renovation and wood-stripping
17 workers specializing in renovation of old houses had significantly higher median blood
18 Pb levels compared with all workers in similar occupations (wood strippers: 37 (ig/dL;
19 window renovators: 32 (ig/dL; all workers: 13.7 (ig/dL; p <0.001) (Mason et al.. 2005).
6.3 Factors Potentially Related to Increased Risk of Pb-lnduced
Health Effects
20 This section evaluates factors examined in recent studies as effect measure modifiers that
21 potentially increase the risk of various Pb-related health effects. There was limited
22 evidence from the 2006 Pb AQCD (U.S. EPA. 2006b) for many of potential at-risk
23 factors described below. Where available, information on conclusions regarding at-risk
24 populations from the 2006 Pb AQCD is included in the subsections.
6.3.1 Age
25 Below is information from epidemiologic and toxicological studies regarding studies of
26 increased risk for Pb-related health effects among children and older adults. Other age
27 groups, such as adolescents, have not been evaluated here, if they were not part of
28 stratified studies of lifestage.
November 2012 6-20 Draft - Do Not Cite or Quote
-------
6.3.1.1 Childhood
1 According to the 2000 Census, 28.6% of individuals living in the U.S. were under the age
2 of 20, with 6.8% aged 0-4 years, 7.3% aged 5-9 years, 7.3% aged 10-14 years, and 7.2%
3 aged 15-19 years (SSDAN CensusScope. 2010a). It is recognized that Pb can cross the
4 placenta and affect the developing nervous system of the fetus (Sections 4.2.2.4 and
5 5.3.2.1) and there is strong evidence of increased risk to the neurocognitive effects of Pb
6 exposure during several lifestages throughout gestation, childhood, and into adolescence
7 (for more detail, Section 5.3.2.1). However, most recent studies among children do not
8 have adequate comparison groups between children of various age groups or between
9 children and adults, and were therefore only presented in Chapter_5.
10 A study including multiple U.S. locations examined associations of blood Pb levels with
11 various immune parameters among individuals living near Pb industrial sites and
12 matched controls (Sarasua et al.. 2000). For several of these endpoints, the association in
13 the youngest group (ages 6-35 months) and the oldest group (ages 16-75 years) were in
14 opposite directions. For example, among children ages 6-35 months, the associations
15 between blood Pb levels and Immunoglobulin A (IgA), Immunoglobulin M (IgM), and
16 B-cell abundance were positive, whereas the associations among 16-75 year olds were
17 negative. The opposite associations were also present for T cell abundance. Ig antibodies,
18 which are produced by activated B cells, are important mediators of the humoral immune
19 response to antigens. T cells are important mediators of cell-mediated immune responses
20 that involve activation of other immune cells and cytokines. These findings by Sarasua et
21 al. (2000) indicate that very young children may be at increased risk for Pb-associated
22 activation of humoral immune responses and perturbations in cell-to-cell interactions that
23 underlie allergic, asthma, and inflammatory responses (for more information, see
24 Sections 5.6.2.1 and 5.6.3).
25 A study among Lebanese children examined the association between blood Pb levels and
26 transferrin saturation (TS) less than 12% and iron-deficiency anemia (IDA) (Muwakkit et
27 al.. 2008). A positive association was detected for blood Pb levels > 10 (ig/dL and both
28 TS less than 12% and IDA among children aged 11-23 months old; however, null
29 associations were observed among children 24-35 months old. Calculations were not
30 performed for children aged 36-75 months because there were no children in the highest
31 Pb group (> 10 (ig/dL) with either TS <12% or IDA. The authors noted that it is difficult
32 to know whether the Pb levels were "a cause or a result of IDA levels since previous
33 studies linked iron deficiency with Pb toxicity.
34 Overall evidence indicates early childhood as a lifestage of increased risk for Pb-related
35 health effects. Both epidemiologic studies summarized above reported associations
36 among the youngest age groups, although different age cut-points were used with one
November 2012 6-21 Draft - Do Not Cite or Quote
-------
1 study including only infants 35 months of age and younger. Toxicological studies provide
2 support for increased health effects of Pb among younger age groups. Toxicological
3 studies have reported that younger animals, whose nervous systems are developing
4 (i.e., laying down and pruning neuronal circuits) and whose junctional barrier systems in
5 the brain (i.e., the blood brain barrier) and GI system (i.e., gut closure) are immature, are
6 more at risk from the effects Pb exposure (Fullmer et al.. 1985). In sum, there are
7 consistent findings, coherent across disciplines that adequate evidence exists to conclude
8 that children are an at-risk population.
6.3.1.2 Older Adulthood
9 The number of Americans over the age of 65 will be increasing in upcoming years
10 (estimated to increase from 12.4% of the U.S. population to 19.7% between 2000 to
11 2030, which is approximately 35 million and 71.5 million individuals, respectively)
12 (SSDAN CensusScope. 2010a: U.S. Census Bureau. 2010). As of the 2000 Census, 7.2%
13 of the U.S. population were ages 60-69, 5.8% were 70-79, and 3.3% were age 80 and
14 older (SSDAN CensusScope. 2010a).
15 A study using the NHANES III cohort examined blood Pb levels and mortality among
16 individuals less than 60 years old and individuals 60 years and older (Menke et al., 2006).
17 Positive hazard ratios were observed in both age groups but the hazard ratios were greater
18 in those less than 60 years old. The interactions terms were not statistically significant. A
19 similar study using the NHANES III cohort examined the relationship between blood Pb
20 levels and mortality from all-cause, cardiovascular disease, and cancer broken down into
21 more specific age groups (Schober et al.. 2006). Point estimates were elevated for the
22 association comparing blood Pb levels > 10 (ig/dL to blood Pb levels <5 (ig/dL and all-
23 cause mortality for all age groups (40-74, 75-84, and 85+ year olds), although the
24 association for 75-84 year olds did not reach statistical significance. The association was
25 also present when comparing blood Pb levels of 5-9 (ig/dL to blood Pb levels <5 (ig/dL
26 among 40-74 year olds and 75-84 year olds, but not among those 85 years and older.
27 None of the associations between blood Pb and cardiovascular disease-related mortality
28 reached statistical significance but the point estimates for cardiovascular disease-related
29 mortality comparing blood Pb levels > 10 (ig/dL to blood Pb levels <5 (ig/dL were
30 elevated among all age groups. Finally, the association between blood Pb levels
31 > 10 (ig/dL and cancer mortality was positive among those 40-74 years old and 85 years
32 and older but the association was null for those 75-84 years old. Among 75-84 year olds
33 the association was positive comparing blood Pb levels of 5-9 (ig/dL to <5 (ig/dL. The
34 other age groups had similar point estimates but the associations were not statistically
35 significant.
November 2012 6-22 Draft - Do Not Cite or Quote
-------
1 A study using the Normative Aging Study cohort reported an interaction between Pb and
2 age (Wright et al.. 2003). The inverse association between age and cognitive function was
3 greater among those with high blood or patella Pb levels. Effect estimates were in the
4 same direction for tibia Pb but the interaction was not statistically significant.
5 Finally, a study of current and former Pb workers reported that an interaction term of Pb
6 and age (dichotomous cutpoint at 67th percentile but exact age not given) examined in
7 models of Pb (measured from blood and patella) and blood pressure was not statistically
8 significant (Weaver et al., 2008). Thus, no modification by age was observed in this study
9 of Pb and blood pressure.
10 Toxicological studies have demonstrated Pb-related health effects among older
11 populations. The kidneys of older animals appear to be more at-risk for Pb-related health
12 effects from the same dose of Pb (i.e., continuous 50 mg/L Pb acetate drinking water)
13 than younger animals (Berrahal et al.. 2011). Increased risk related to older age is also
14 observed for effects on the brain. Recent studies have demonstrated the importance of Pb
15 exposure during early development in promoting the emergence of Alzheimer's like
16 pathologies in aged animals. Development of pathologies of old age in brains of aged
17 animals that were exposed to Pb earlier in life has been documented in
18 psychopathological effects in adults (mice and monkeys), (for more details see
19 Section 5.3.10.1). These pathologies include the development of neurofibrillary tangles
20 and increased amyloid precursor protein and its product beta-amyloid (Bashaet al., 2005;
21 Zawia and Basha. 2005). Some of these findings were seen in animals that no longer had
22 elevated blood Pb levels.
23 In summary, results for age-related modification of the association between Pb and
24 mortality had mixed results. Limited evidence was available for the associations between
25 Pb and cognitive function or other health effects among older adults. Toxicological
26 studies have shown increases in Pb-related health effects by age that may be relevant in
27 humans. Future studies will be instrumental in understanding older age as a factor that
28 potentially affects the risk of Pb-related outcomes.
6.3.2 Sex
29 The distribution of males and females in the U.S. is similar. In 2000, 49.1% of the U.S.
30 population was male and 50.9% was female. The distribution of sex varied by age with a
31 greater prevalence of females > 65 years old compared to males (SSDAN CensusScope.
32 2010a). The 2006 Pb AQCD reported that boys are often found to have higher blood Pb
33 levels than girls, but findings were "less clear" regarding differences in Pb-related health
34 effects between males and females (U.S. EPA, 2006b).
November 2012 6-23 Draft - Do Not Cite or Quote
-------
1 Multiple epidemiologic studies have examined Pb-related effects on cognition stratified
2 by sex. In previous studies using the Cincinnati Lead Study cohort, Dietrich et al. Q987b)
3 and Ris et al. (2004) observed interactions between blood Pb (prenatal and postnatal) and
4 sex; associations of prenatal and postnatal blood Pb and subsequent decrements in
5 memory, attention, and visuoconstruction were observed only among male adolescents.
6 More recently, Wright et al. (2008) examined early life blood Pb levels and criminal
7 arrests in adulthood. The risks attributable to Pb exposure were greater among males than
8 females. Additionally, the association between childhood blood Pb levels and adult gray
9 matter volume loss was greater among males than females (Cecil et al.. 2008). In an
10 expanded analysis of the developmental trajectory of childhood blood Pb levels on adult
11 gray matter, researchers found that associations between yearly mean blood Pb levels and
12 volume of gray matter loss were more pronounced in the frontal lobes of males than
13 females (Brubakeretal.. 2010). Multiple studies were also conducted in Port Pirie,
14 Australia that examined blood Pb levels at various ages throughout childhood and
15 adolescence (Tong et al.. 2000; Baghurst et al.. 1992; McMichael et al.. 1992). These
16 studies observed Pb effects on cognition deficits were stronger in girls throughout
17 childhood and into early adolescence. A study in Poland also investigated the association
18 between umbilical cord blood Pb levels and cognitive deficits and reported a positive
19 association for boys at 36 months but not for girls (Jedrychowski et al.. 2009a). No
20 association was detected for boys or girls at 24 months.
21 An epidemiologic study examined the association between concurrent blood Pb levels
22 and kidney function among 12-20 year olds using the NHANES III study cohort
23 (Fadrowski et al.. 2010). The results were stratified by sex and no effect measure
24 modification was apparent.
25 Similarly, a study of current and former Pb workers examined an interaction term
26 between sex and Pb for the study of blood Pb and blood pressure (Weaver et al.. 2008).
27 No modification by sex was present.
28 Epidemiologic studies have also been performed to assess differences between males and
29 females for Pb-related effects on various biomarkers. A study comprised mostly of
30 females reported positive associations between blood Pb and total immunoglobulin E
31 (IgE) for women not taking hormone replacement therapy or oral contraceptives (Pizent
32 et al.. 2008). No association was reported in males, but other associations, such as
33 bronchial reactivity and reactive skin prick tests were observed in the opposite of the
34 expected direction, which questions the validity of the results among the male study
35 participants. Analysis of an NHANES dataset detected no association between blood Pb
36 levels and inflammatory markers (Songdei et al.. 2010). Although there was no clear
37 pattern, a few of the associations were positive between blood Pb and C-reactive protein
November 2012 6-24 Draft - Do Not Cite or Quote
-------
1 for males but not females. A study of children living at varying distances from a Pb
2 smelter in Mexico reported that blood Pb was associated with increased release of
3 superoxide anion from macrophages, which was greater among males than females
4 (Pineda-Zavaleta et al.. 2004).
5 Epidemiologic investigations of cancer have also examined the associations by sex. A
6 study of the association between occupational exposure to Pb and brain tumors reported
7 no sex-specific associations for gliomas, but a positive association for cumulative Pb
8 exposure and meningiomas for males but not females (Rajaraman et al.. 2006). An
9 ecologic analysis of Pb pollution levels and cancer incidence among children reported
10 weak correlations overall and the weak correlations were more apparent among males,
11 whereas no correlation was observed among females (Absalon and Slesak. 2010).
12 A study of all-cause and cardiovascular mortality using the NHANES III cohort reported
13 no modification of the association between blood Pb and all-cause or cardiovascular
14 mortality by sex (Menke et al., 2006). This did not differ among women when classified
15 as pre- or post-menopausal.
16 Toxicological studies have also reported sex differences in Pb-related effects to various
17 organ systems. Donald et al. (1986) reported a different time course of enhanced social
18 investigatory behavior between male and female mice exposed to Pb. In a subsequent
19 publication, Donald et al. (1987) showed that non-social behavior in mice decreased in
20 females and increased in males exposed to Pb. Males also had a shorter latency to
21 aggression with Pb treatment versus controls. Pb affected mood disorders differently for
22 males and females. Behavioral testing in rats showed males experienced emotional
23 changes and females depression-like changes with Pb exposure (de Souza Lisboa et al..
24 2005). In another study, gestational exposure to Pb impaired memory retrieval in male
25 rats at all 3 doses of Pb exposure; memory retrieval was only impaired in low-dose
26 female rats (Yang et al.. 2003). Sex-specific differences in mice were also observed for
27 gross motor skills; at the lowest Pb dose, balance and coordination were most affected
28 among males (Leasure et al.. 2008).
29 Pb and stress are co-occurring factors that act in a sex-divergent manner to affect
30 behavior, neurochemistry, and corticosterone levels. Pb and stress act synergistically to
31 affect fixed interval operant behavior and corticosterone in female rat offspring. Virgolini
32 et al. (2008a) found that effects on the offspring's central nervous system by
33 developmental Pb exposure (maternal exposure and transferred to the offspring through
34 lactation) were enhanced by combined maternal and offspring stress and females were
35 most at risk. Behavioral related outcomes after gestational and lactational Pb exposure
36 (with and without stress) exhibited sex-differences in exposed offspring (Virgolini et al..
37 2008b). Pb-induced changes in brain neurochemistry, with or without concomitant stress
November 2012 6-25 Draft - Do Not Cite or Quote
-------
1 exposure, are complex with differences varying by brain region, neurotransmitter type,
2 and sex of the animal.
3 The brain is known to have a sexually dimorphic area in the hypothalamus, termed the
4 sexually dimorphic nucleus (SDN). Lesions in this area affect sex-specific phenotypes
5 including behavior. Across species the SDN has a greater cell number and larger size in
6 males versus females. This sexually dichotomous area is especially vulnerable to
7 perturbation during fetal life and the early postnatal period. This may be one area of the
8 brain that could explain some of the sexually dichotomous effects that are seen with Pb
9 exposure. One study supporting this line of thought showed that high-dose in utero Pb
10 exposure (pup blood Pb level 64 (ig/dL at birth) induced reductions in SDN volume in
11 35% of Pb-exposed male rats (McGivern et al., 1991). Interestingly, another chemical
12 that is known to cause a hypothalamic lesion in this area, monosodium glutamate, is
13 associated with adult onset obesity (Olney. 1969); adult onset obesity is seen in the Pb
14 literature.
15 Obesity in adult offspring exposed to low-dose Pb in utero was reported for male but not
16 female mice (Leasure et al., 2008). Obesity was also found in male rat offspring exposed
17 in utero to high doses of Pb that persisted to 5 weeks of age/end of the study, but among
18 female rats, body weight remained elevated over controls only to 3 weeks of age (Yang et
19 al.. 2003). Additionally, low-dose Pb exposure induced retinal decrements in exposed
20 male mice offspring (Leasure et al., 2008).
21 A toxicological study of Pb and antioxidant enzymes in heart and kidney tissue reported
22 that male and female rats had differing enzymatic responses, although the amount of Pb
23 in the heart tissue or the disposition of Pb also varied between males and females
24 (Sobekova et al.. 2009; Alghazal et al.. 2008a). The authors reported these results could
25 be due to greater deposition of Pb in female rats or greater clearance of Pb by males
26 (Sobekova et al.. 2009).
27 Multiple associations between Pb and various health endpoints have been examined for
28 effect measure modification by sex and results have been inconsistent. Although not
29 observed in all endpoints, some studies reported differences between the associations for
30 males and females, especially in neurological studies. However, studies on cognition
31 from the Cincinnati Lead Study cohort and a study in Poland reported males to be an at-
32 risk population, whereas studies from Australia pointed to females as an at-risk
33 population. A difference in sex is supported by toxicological studies. Further research is
34 needed to confirm the presence or absence of sex-specific associations between Pb and
35 various health outcomes and to determine in which sex the associations are greater.
November 2012 6-26 Draft - Do Not Cite or Quote
-------
6.3.3 Genetics
1 The 2006 Pb AQCD stated that, "genetic polymorphisms in certain genes have been
2 implicated as influencing the absorption, retention, and toxicokinetics of Pb in humans"
3 (U.S. EPA. 2006b). The majority of discussion there focused on the aminolevulinate
4 dehydratase (ALAD) and vitamin D receptor (VDR) polymorphisms. These two genes, as
5 well as additional genes examined in recent studies, are discussed below.
6.3.3.1 Aminolevulinate Dehydratase
6 The aminolevulinate dehydratase (ALAD) gene encodes for an enzyme that catalyzes the
7 second step in the production of heme and is also the principal Pb-binding protein (U.S.
8 EPA. 2006b). Studies have examined whether ALAD variants altered associations
9 between Pb and various health effects.
10 Associations between Pb and brain tumors observed in an epidemiologic study varied by
11 ALAD genotype status (Raiaraman et al.. 2006). Positive associations between Pb
12 exposure (determined via interview about occupational exposures) and meningioma were
13 reported among ALAD2 individuals, but this association was not found among
14 individuals who had the ALAD1 allele. No associations were observed between Pb and
15 glioma regardless of ALAD genotype.
16 Studies investigating the association between Pb levels and cognitive function have also
17 examined modification by ALAD polymorphisms. The evidence is provided by an
18 NHANES analysis (Krieg et al.. 2009) as well as multiple analyses from the NAS cohort
19 examining different tests of cognitive function (Rajan et al.. 2008; Weuve et al.. 2006). In
20 the study using a cohort from NHANES III, for several indices of cognitive function,
21 associations with concurrent blood Pb levels were more pronounced in groups with CC
22 and CG ALAD genotypes (i.e., ALAD2 carriers) (Krieg et al.. 2009). In the NAS cohort
23 of men, Weuve et al. (2006) found that concurrent blood Pb level but not bone Pb level
24 was associated with a larger decrease in a test of general cognitive function among
25 ALAD2 carriers. Another NAS study examined functioning of specific cognitive
26 domains (e.g., vocabulary, memory, visuospatial skills) and found variable evidence for
27 effect modification by ALAD genotype across tests (Rajan et al.. 2008). For example,
28 among ALAD2 carriers, concurrent blood Pb level was associated with a more
29 pronounced decrease in vocabulary score but less pronounced decrease in a memory
30 index and no difference in the associations with other cognitive tests. For tibia and patella
31 Pb levels, ALAD genotype was found to modify associations with different tests, for
32 example, executive function and perceptual speed. It is not clear why the direction of
November 2012 6-27 Draft - Do Not Cite or Quote
-------
1 effect modification would vary among different cognitive domains. The limited number
2 of populations examined, and the different cognitive tests performed in each study, make
3 it difficult to conclusively summarize findings for effect modification by ALAD variants.
4 However, in the limited available body of evidence, blood and bone Pb levels were
5 generally associated with lower cognitive function in ALAD2 carriers.
6 A study of current and former workers exposed to Pb examined the association between
7 blood Pb and blood pressure and reported no modification by ALAD genotype (Weaver
8 et al., 2008). However, another study of blood Pb and blood pressure reported
9 interactions between blood Pb and ALAD, but this varied by race/ethnicity (non-Hispanic
10 white, non-Hispanic black, and Mexican American) (Scinicariello et al.. 2010).
11 Individuals with ALAD2 variants had greater associations between Pb and kidney
12 effects; among those with the variant, higher Pb was associated with higher glomerular
13 filtration measures (Weaver et al.. 2006; Weaver et al.. 2005b: Weaver et al.. 2003b). A
14 study of workers at a battery plant storage facility in China reported workers with the
15 ALAD2 allele demonstrated greater associations between blood Pb levels and renal
16 injury (Gao et al.. 2010a). Another study of renal function among Pb workers in Asia also
17 reported greater associations between blood Pb concentrations and renal function by
18 ALAD, especially at high blood Pb levels (Chia et al.. 2006).
6.3.3.2 Vitamin D Receptor
19 The vitamin D receptor (VDR) is a regulator of calcium absorption and metabolism. A
20 recent study of the NHANES III population examined the association between blood Pb
21 levels and various neurocognitive tests with assessment of effect measure modification
22 by SNPs and haplotypes of VDR (Krieg et al.. 2010). The results were varied, even
23 among specific SNPs and haplotypes, with some variants being associated with greater
24 modification of the relationship between Pb and one type of neurocognitive test
25 compared to the modification of the relationship between Pb and other neurocognitive
26 tests. In an epidemiologic study of blood Pb levels and blood pressure among a group of
27 current and former Pb-exposed workers, no modification was reported by VDR (Weaver
28 et al.. 2008).
November 2012 6-28 Draft - Do Not Cite or Quote
-------
6.3.3.3 Methylenetetrahydrofolate reductase
1 Methylenetetrahydrofolate reductase (MTHFR) catalyzes the conversion of
2 5,10-methylenetetrahydrofolate to 5-methyltetrahydrofolate, which in turn, is involved in
3 homocysteine remethylation to the amino acid methionine. A study in Mexico of the
4 association between Pb and Bayley's Mental Development Index (MDI) score at 24
5 months reported no effect measure modification by MTHFR 677T allele (Pilsner et al..
6 2010). Another study in Mexico examined the association between maternal Pb and birth
7 weight (Kordas et al.. 2009). No modification of the Pb-birth weight association by
8 MTHFR was observed.
6.3.3.4 Hemochromatosis
9 The hemochromatosis (HFE) gene encodes a protein believed to be involved in iron
10 absorption. A difference was observed between the association of tibia Pb levels and
11 cognitive function for men with and without HFE allele variants (Wang et al.. 2007a). No
12 association between tibia Pb and cognitive function was present for men with HFE
13 wildtype, but a decline in function was associated with tibia Pb levels among men with
14 any HFE allele variant. A study of bone Pb levels and HFE reported no difference in
15 effect estimates for bone Pb and pulse pressure between different HFE variants and HFE
16 wild-type (Zhang et al.. 2010a). An interaction was observed between an HFE variant in
17 mothers and maternal tibia Pb in a study of maternal Pb and birth weight (Cantonwine et
18 al.. 2010b). The inverse association between maternal tibia Pb levels and birth weight
19 was stronger for those infants whose mothers had the HFE variant. The interaction was
20 not present between the HFE variants and maternal blood Pb or cord blood Pb
21 concentrations.
6.3.3.5 Other Genetic Polymorphisms
22 Some other genetic polymorphisms were also examined as to whether they modify
23 Pb-related health effects, but only limited data were available for these polymorphisms.
24 These include dopamine receptor D4 (DRD4), dopamine receptor D2 (DRD2), dopamine
25 transporter (DAT1), glutathione S-transferase Mu 1 (GSTM1), tumor necrosis factor-
26 alpha (TNF-a), endothelial nitric oxide synthase (eNOS), and various SNPS.
27 A prospective birth cohort reported that increasing blood Pb levels were associated with
28 poorer rule learning and reversal, spatial span, and planning in their study population
29 (Froehlich et al.. 2007). These inverse associations were exacerbated among those
November 2012 6-29 Draft - Do Not Cite or Quote
-------
1 lacking DRD4-7. A study of prenatal and postnatal Pb levels in Mexico City reported no
2 modification of the associations between Pb levels and neurocognitive development by
3 DRD2 or DAT 1 (Kordas et al.. 2011).
4 A study of university students in South Korea reported blood Pb levels to be associated
5 with biomarkers of inflammation among individuals with GSTM1 null genotype and not
6 among individuals with GSTM1 present (Kim et al.. 2007). This study of blood Pb levels
7 and inflammation also examined individuals with TNF-a GG, GA, or AA alleles. An
8 association was present for those with TNF-a GG but not for those with TNF-a GA or
9 AA.
10 A study of blood Pb and plasma NOX reported no overall association but did report an
11 inverse correlation among subjects with the eNOS TC+CC genotype (Barbosa et al..
12 2006c). No correlation was observed for subjects with the eNOS TT genotype; however
13 the number of subjects in this group was small, especially for those with high blood Pb
14 levels.
15 One study examined how the association between occupational Pb exposure and brain
16 tumors varied among multiple single nucleotide polymorphisms (SNPs) (Bhatti et al..
17 2009). No effect measure modification of the association between Pb and glioma was
18 observed for any of the SNPs. GPX1 (the gene encoding for glutathione peroxidase 1)
19 modified the association for glioblastoma multiforme and meningioma. The association
20 between Pb and glioblastoma multiforme was also modified by a RAC2 (the gene
21 encoding for Rac2) variant, and the association between Pb and meningioma was also
22 modified by XDH (the gene encoding for xanthine dehydrogenase) variant.
23 Overall, studies of ALAD observed increased Pb-related health effects associated with
24 certain gene variants. Other genes, such as VDR, HFE, DRD4, GSTM1, TNF-a, and
25 eNOS, may also affect the risk of Pb-related health effects but conclusions are limited
26 due to the small number of studies.
6.3.4 Pre-existing Diseases/Conditions
27 Studies have also been performed to examine whether certain morbidities increase an
28 individual's risk of Pb-related effects on health. Recent studies have explored
29 relationships for autism, diabetes, and hypertension.
November 2012 6-30 Draft - Do Not Cite or Quote
-------
6.3.4.1 Autism
1 Rates of individuals with autism have increased in recent years. A study reported a
2 prevalence rate in 2006 of 9.0 per 1,000 individuals (95% CI: 8.6, 9.3) determined from a
3 monitoring network (Autism and Developmental Disabilities Monitoring Network) with
4 11 sites across the U.S. (CDC. 2009).
5 A cross-sectional study of children with and without autism examined the association
6 between blood Pb levels and various immune function and inflammation genes (Tian et
7 al., 2011). Blood Pb levels of children with and without autism were associated with
8 expression of the genes under study; however, the associations observed were in opposite
9 directions (for children with autism, increased blood Pb levels were associated with
10 increased expression, whereas for children without autism, increased blood Pb levels
11 were associated with decreased expression).
6.3.4.2 Diabetes
12 Approximately 8% of U.S. adults have diabetes (Pleis et al., 2009). A few studies have
13 been conducted to investigate the possibility of diabetes as a modifying factor for Pb and
14 various health outcomes.
15 Differences in the association between bone and blood Pb levels and renal function for
16 individuals with and without diabetes at baseline were examined using the Normative
17 Aging Study cohort (Tsaih et al., 2004). Tibia and blood Pb levels were positively
18 associated with measures of poor renal function among individuals with diabetes but not
19 among individuals without diabetes. However, this association was no longer statistically
20 significant after the exclusion of individuals who were hypertensive or who used diuretic
21 medications. Another study with this cohort reported no associations between bone Pb
22 and heart rate variability, which did not differ among those with and without diabetes
23 (Park et al. 2006V
24 The NHANES III data were used to evaluate whether the association between blood Pb
25 and both all-cause and cardiovascular mortality varied among individuals with and
26 without diabetes (Menke et al.. 2006). The 95% CIs among those with diabetes were
27 large and no difference was apparent among those with and without diabetes.
28 Overall, recent epidemiologic studies found that associations between Pb concentrations
29 and health outcomes did not differ for individuals with and without diabetes. However,
30 results from the 2006 Pb AQCD found that individuals with diabetes are at "increased
31 risk of Pb-associated declines in renal function" (U.S. EPA. 2006b). Future research
November 2012 6-31 Draft - Do Not Cite or Quote
-------
1 examining associations between Pb and renal function, as well as other health outcomes,
2 among individuals with and without diabetes will inform further on the potential for
3 increased risk among individuals with diabetes.
6.3.4.3 Hypertension
4 Hypertension affects approximately 24% of adults in the U.S. and the prevalence of
5 hypertension increases with age (61% of individuals > 75 years old have hypertension)
6 (Pleis et al. 2009).
7 The Normative Aging Study mentioned above evaluating modification of the association
8 between Pb levels and renal function by diabetes also examined modification by
9 hypertensive status (Tsaih et al.. 2004). The association between tibia Pb and renal
10 function, measured by change in serum creatinine, was present among individuals with
11 hypertension but not among individuals that were normotensive. Models of the follow-up
12 serum creatinine levels demonstrated an association with blood Pb for individuals with
13 hypertension but not individuals without hypertension (this association was not present
14 when using tibia or patella Pb). Another study using this population examined
15 modification of the association between bone Pb and heart rate variability, measured by
16 low frequency power, high frequency power, and their ratio (Park et al., 2006). Although
17 a statistically significant association between bone Pb and heart rate variability was not
18 observed among individuals with or without hypertension, the estimates were different,
19 with greater odds for individuals with hypertension (bone Pb levels were positively
20 related to low frequency power and the ratio of low frequency to high frequency power
21 and were inversely related to high frequency power).
22 A study using the NHANES III cohort reported a positive association between blood Pb
23 levels and both all-cause and cardiovascular mortality for individuals with and without
24 hypertension but the associations did not differ based on hypertensive status (Menke et
25 al.. 2006).
26 The 2006 Pb AQCD reported that individuals with hypertension had increased risk of
27 Pb-related effects on renal function (U.S. EPA. 2006b). This is supported by recent
28 epidemiologic studies. As described above, studies of Pb-related effects on renal function
29 and heart rate variability have observed some differences among individuals with
30 hypertension, but the difference between adults with and without hypertension was not
31 observed for Pb-related mortality.
November 2012 6-32 Draft - Do Not Cite or Quote
-------
1 Overall, studies of Pb-related health effects related to pre-existing conditions have some
2 evidence of a potential increased risk of Pb-related health effects. The evidence is
3 consistent for Pb-related renal effects and hypertension but is limited for other pre-
4 existing conditions.
6.3.5 Smoking Status
5 The rate of smoking among adults 18 years and older in the U.S. is approximately 20%
6 and about 21% of individuals identify as former smokers (Pleis et al., 2009). Studies of
7 Pb and various health effects have examined smoking as an effect measure modifier.
8 A study of blood Pb levels and all-cause and cardiovascular mortality reported no
9 modification of this association by smoking status, measured as current, former, or never
10 smokers (Menke et al.. 2006). The Normative Aging Study also examined the association
11 between blood and bone Pb levels and renal function and also reported no interaction
12 with smoking status (Tsaih et al.. 2004).
13 A study of Pb-exposed workers and controls reported similar levels of absolute neutrophil
14 counts (ANC) across Pb exposure categories among non-smokers (Di Lorenzo et al..
15 2006). However, among current smokers, higher Pb exposure was associated with higher
16 ANC. Additionally, a positive relationship was observed between higher blood Pb levels
17 and TNF-a and granulocyte colony-stimulating factor (G-CSF) among both smokers and
18 nonsmokers, but this association was greater among smokers (Di Lorenzo et al.. 2007). A
19 recent study of fertile and infertile men examined blood and seminal plasma Pb levels for
20 smokers and non-smokers (Kiziler et al.. 2007). The blood and seminal plasma Pb levels
21 were higher for smokers of both fertile and infertile groups. Additionally, the Pb levels
22 were lowest among non-smoking fertile men and highest among smoking infertile men.
23 Prenatal smoking exposure was examined in a study of children's concurrent blood Pb
24 levels and prevalence of attention-deficit/hyperactivity disorder (ADHD) among children
25 aged 8-15 years. An interaction was observed between children's current blood Pb levels
26 and prenatal tobacco smoke exposure; those children with high Pb levels and prenatal
27 tobacco smoke exposure had the highest odds of ADHD (Troehlich et al.. 2009).
28 Overall, the studies have inconsistent findings on whether smoking modifies the
29 relationship between Pb levels and health effects. Future studies of Pb-related health
30 effects and current, former, and prenatal smoking exposures among various health
31 endpoints will aid in determining changes in risk by this factor.
November 2012 6-33 Draft - Do Not Cite or Quote
-------
6.3.6 Socioeconomic Status
1 Based on the 2000 Census data, 12.4% of Americans live in poverty (poverty threshold
2 for family of 4 was $17,463) (SSDAN CensusScope. 2010c). Few studies have compared
3 blood Pb level effect estimates among groups in different sociodemographic strata.
4 Larger blood Pb-associated decreases in cognitive function were found with lower SES in
5 some studies (Ris et al.. 2004; Tong et al.. 2000; Bellinger et al.. 1990). In contrast, a
6 meta-analysis of eight studies found a smaller decrement in Full Scale Intelligence
7 Quotient (FSIQ) for studies in disadvantaged populations than for studies in advantaged
8 populations (Schwartz. 1994). While results indicate that blood Pb level is associated
9 with FSIQ deficits in both higher and lower sociodemographic groups, they do not clearly
10 indicate whether groups with different socioeconomic status differ in Pb-related changes
11 for cognitive function.
6.3.7 Race/Ethnicity
12 Based on the 2000 Census, 69.1% of the U.S. population is comprised of non-Hispanic
13 whites. Approximately 12.1% of people reported their race/ethnicity as non-Hispanic
14 black and 12.6% reported being Hispanic (SSDAN CensusScope. 2010b). Studies of
15 multiple Pb-related health outcomes examined effect measure modification by
16 race/ethnicity.
17 A study of adults from the NHANES III cohort examined the association between blood
18 Pb levels and all-cause and cardiovascular mortality (Menke et al.. 2006). Stratified
19 analyses were conducted for non-Hispanic whites, non-Hispanic blacks, and Mexican
20 Americans and no interaction for race/ethnicity was reported. Other studies have also
21 used NHANES cohorts to study blood Pb levels and hypertension (Scinicariello et al..
22 2010; Muntner et al.. 2005). While no association was observed between blood Pb and
23 hypertension for non-Hispanic whites or Hispanics, a positive association was reported
24 for non-Hispanic blacks in a study using the NHANES III cohort (Scinicariello et al..
25 2010). In another study, although none of the associations between blood Pb levels and
26 hypertension were statistically significant, increased odds were observed among
27 non-Hispanic blacks and Mexican Americans but not for non-Hispanic whites (Muntner
28 et al.. 2005).
November 2012 6-34 Draft - Do Not Cite or Quote
-------
1 A study of girls aged 8-18 years from the NHANES III cohort reported an inverse
2 association between blood Pb levels and pubertal development among blacks and
3 Mexican Americans (Selevan et al., 2003). For non-Hispanic whites, the associations
4 were in the same direction but did not reach statistical significance. Of note, less than 3%
5 of non-Hispanic whites had blood Pb levels over 5 (ig/dL, whereas 11.6% and 12.8% of
6 blacks and Mexican Americans, respectively, had blood Pb levels greater than 5 (ig/dL.
7 A study linking educational testing data for 4th grade students in North Carolina reported
8 declines in reading and mathematics scores with increasing levels of blood Pb (Miranda
9 et al.. 2007a). Although not quantitatively reported, a figure in the study depicted the
10 association stratified by race, and the slopes appeared to be similar for white and black
11 children.
12 Blood Pb and asthma incidence was examined for white and black children living in
13 Michigan (Joseph et al.. 2005). When utilizing separate referent groups for the two races,
14 the only association is an increase among whites (although not statistically significant),
15 but when restricting to the highest blood Pb levels, the association was no longer
16 apparent. Whites with low blood Pb levels were used as the referent group for both races
17 in additional analysis. Although the estimates were elevated for black children compared
18 to white children (including at the lowest blood Pb levels), the confidence intervals for
19 the associations overlapped indicating a lack of a difference by race.
20 The results of these recent epidemiologic studies provide some evidence that there may
21 be race/ethnicity-related increased risk with higher Pb levels for certain outcomes,
22 although the overall understanding of potential effect measure modification by
23 race/ethnicity is limited by the small number of studies. Additionally, these results may
24 be confounded by other factors, such as socioeconomic status or nutritional factors.
6.3.8 Body Mass Index
25 In the U.S. self-reported rates of obesity were 26.7% in 2009, up from 19.8% in 2000
26 (Sherry etal.. 2010). The NHANES III cohort was utilized in a study of blood Pb levels
27 and all-cause and cardiovascular mortality, which included assessment of the associations
28 by obesity (Menke et al.. 2006). Positive associations were observed among individuals
29 within both categories of body mass index (BMI; normal [<25 kg/m2] and
30 overweight/obese [> 25 kg/m2], determined using measured values of height and weight)
31 but there was no difference in the association between the two categories. Using the
32 Normative Aging Study data, an investigation of bone Pb levels and heart rate variability
33 was performed and reported slight changes in the association based on the presence of
34 metabolic syndrome; however, none of the changes resulted in associations that were
November 2012 6-35 Draft - Do Not Cite or Quote
-------
1 statistically significant (Park et al.. 2006). Overall, no modification by BMI or obesity
2 was observed among recent epidemiologic studies, but the available epidemiologic and
3 supporting toxicological studies are limited.
6.3.9 Alcohol Consumption
4 There are a limited number of studies examining alcohol as a factor affecting Pb-related
5 risk. A study using the Normative Aging Study cohort investigated whether the
6 association between blood and bone Pb levels and renal function would be modified by
7 an individual's alcohol consumption (Tsaih et al.. 2004). No interaction with alcohol
8 consumption was observed. However, a toxicological study reported that ethanol
9 potentiated the effect of Pb exposure by decreasing renal total protein sulfhydryls
10 (endogenous antioxidants) in rats. Pb and ethanol also decreased other endogenous renal
11 antioxidants (glutathione and non-protein sulfhydryls) (Jurczuk et al.. 2006). Overall,
12 evidence to determine if alcohol consumption is a potential at-risk factor is of limited
13 quantity and consistency.
6.3.10 Nutritional Factors
14 Different components of diet may affect the association between Pb concentrations and
15 health outcomes. Recent epidemiologic and toxicological studies of specific mineral
16 intakes/dietary components are detailed below.
6.3.10.1 Calcium
17 Using the Normative Aging Study (NAS) cohort, researchers examined the association
18 between Pb levels and hypertension, modified by calcium intake (Elmarsafawy et al..
19 2006). The associations between Pb levels (measured and modeled separately for blood,
20 patella, and tibia) and hypertension did not differ based on dichotomized calcium intake
21 (800 mg/day).
November 2012 6-36 Draft - Do Not Cite or Quote
-------
6.3.10.2 Iron
1 The 2006 Pb AQCD included studies that indicated individuals with iron-deficiency and
2 malnourishment had greater inverse associations between Pb and cognition (U.S. EPA,
3 2006b). A recent epidemiologic study of pubertal development among girls observed
4 inverse associations between blood Pb and inhibin B. This association was modified by
5 iron deficiency; girls with iron deficiency had a stronger inverse association between Pb
6 and inhibin B than those who were iron sufficient (Gollenberg et al., 2010). Toxicological
7 studies also reported that iron-deficient diets exacerbate or potentiate the effect of Pb. A
8 study of pregnant rats given an iron-deficient diet and exposed to Pb through drinking
9 water over GD6-GD14, had decreased litter size, more pups with reduced fetal weight
10 and reduced crown-rump length, increased litter resorption, and a higher dam blood Pb
11 level in the highest exposure groups (Singh et al.. 1993b: Saxenaetal.. 1991). Thus, in
12 this model, iron deficiency makes rat dams more at risk for Pb-dependent embryo and
13 fetotoxicity (Singh et al.. 1993b).
6.3.10.3 Folate
14 A study by Kordas et al. (2009) examined Pb levels and birth size among term births in
15 Mexico City. The authors reported no interaction between maternal tibia Pb and folate
16 levels.
6.3.10.4 Protein
17 No recent epidemiologic studies have evaluated protein intake as a factor affecting
18 Pb-related health effects. However, a toxicological study demonstrated that differences in
19 maternal protein intake levels could affect the extent of Pb-induced immunotoxicity
20 among offspring (Chen et al., 2004).
21 In sum, the evidence is limited for most dietary factors but evidence for iron deficiency as
22 a factor that potentially increases risk of Pb-induced effects is present and coherent in
23 epidemiologic and toxicological studies.
November 2012 6-37 Draft - Do Not Cite or Quote
-------
6.3.11 Stress
1 A study of bone (tibia and patella) Pb levels and hypertension reported modification of
2 the association by perceived stress levels (Peters et al.. 2007). Among individuals with
3 greater perceived stress levels, stronger associations between blood Pb levels and
4 hypertension were present. Among the same study population, higher perceived stress
5 was also reported to affect the association between blood Pb levels and cognitive
6 function; the higher stress group showed a greater inverse association between Pb and
7 cognitive function than those in the low stress group (Peters et al.. 2008). In another
8 study, the inverse association between tibia Pb levels and some measures of cognitive
9 function were similarly strengthened by neighborhood psychosocial hazards (Glass et al..
10 2009).
11 Toxicological studies have demonstrated that early life exposure to Pb and maternal
12 stress can result in toxicity related to multiple systems (Rossi-George et al.. 2009: Cory-
13 Slechtaetal.. 2008: Virgolini et al.. 2008a: Virgolini et al.. 2008b). including
14 dysfunctional corticosterone responses (Rossi-George et al.. 2009: Virgolini et al..
15 2008b). Additionally, toxicological studies have demonstrated that stressors to the
16 immune system can also affect associations with Pb exposure. Chickens with low Pb
17 exposure in ovo, with additional viral stressors, had increased immune cell mobilization
18 and trafficking dysfunction (Lee et al.. 2002). Similarly, mice with neonatal Pb exposure,
19 and an additional immune challenge, had a sickness behavior phenotype, likely driven by
20 IL-6 production (Dyatlov and Lawrence. 2002).
21 Although examined in a limited number of studies, recent epidemiologic studies observed
22 modification of the association between Pb and various nervous system health effects by
23 stress-level. Increased risk of Pb-related health effects by stress is further supported by
24 toxicological studies.
6.3.12 Maternal Self-Esteem
25 Maternal self-esteem has been shown to modify associations between blood Pb levels and
26 health effects in children. Surkan et al. (2008) studied the association between children's
27 blood Pb levels and Bayley's MDI and Psychomotor Development Index (PDI) among
28 mother-child pairs. High maternal self-esteem was independently associated with higher
29 MDI score and also appeared to attenuate the negative effects of the child's increased
30 blood Pb levels on MDI and PDI scores. Greater decreases in MDI and PDI were
31 associated with increased blood Pb levels among children whose mothers were in the
32 lower quartiles of self-esteem.
November 2012 6-38 Draft - Do Not Cite or Quote
-------
6.3.13 Cognitive Reserve
1 Cognitive reserve has been defined as "the maintenance of cognitive performance in spite
2 of ongoing underlying brain pathology" (Bleecker et al.. 2007a). A study of Pb smelter
3 workers reported that an inverse association between lifetime weighted blood Pb levels
4 and cognitive function was present among workers with low cognitive reserve (measured
5 using a reading achievement test) but no association was present in workers with high
6 cognitive reserve (Bleecker et al.. 2007a). Inverse associations between lifetime-weighted
7 blood Pb levels and motor functions existed among all workers regardless of cognitive
8 reserve. No other recent epidemiologic studies were performed examining cognitive
9 reserve as a factor affecting risk of Pb-related health outcomes, thus providing limited
10 evidence to conclude that cognitive reserve is a potential at-risk factor.
6.3.14 Other Metal Exposure
11 The 2006 Pb AQCD reported that the majority of studies that examined other toxicants
12 did so as confounders and not as effect measure modifiers (U.S. EPA. 2006b). Recent
13 epidemiologic studies have begun to explore the possible interaction between Pb
14 exposure and co-exposures with other metals. These studies, as well as toxicological
15 studies of these metals, are described below.
6.3.14.1 Cadmium
16 In a study of girls in the NHANES III cohort, inverse associations were observed
17 between blood Pb and inhibin B concentrations (Gollenberg et al.. 2010). These inverse
18 associations were stronger among girls with high cadmium (Cd) and high Pb compared to
19 those with high Pb and low Cd. Additionally, higher blood Pb and Cd levels together
20 were positively associated with albuminuria and reduced estimated glomerular filtration
21 rate, compared to those with the lowest levels of Pb and Cd (Navas-Acien et al.. 2009).
22 Toxicological studies reported that in rats, the addition of Cd to Pb exposure reduced the
23 histological signs of renal toxicity from each element alone; however, urinary excretion
24 of porphyrins were increased, indicating that although measured tissue burdens of Pb
25 were reduced, the biologically available fraction of Pb was actually increased (Wang and
26 Fowler. 2008). In other studies, Cd synergistically exacerbated Pb-dependent renal
27 mitochondrial dysfunction (Wang et al.. 2009c).
November 2012 6-39 Draft - Do Not Cite or Quote
-------
1 Overall, epidemiologic and toxicological studies have reported increased risk of
2 Pb-related health effects among those with high Cd levels as well; however, the number
3 of studies examining both metals is small.
6.3.14.2 Manganese
4 Among children in South Korea taking part in a study of IQ, an interaction was reported
5 between Pb and manganese (Mn) blood levels (Kim et al.. 2009b). Children with high
6 blood Mn levels were observed to have reductions in full scale IQ and verbal IQ
7 associated with increased blood Pb levels, whereas no association between blood Pb
8 levels and full scale IQ and verbal IQ were noted among those children with low blood
9 Mn levels. No effect measure modification by Mn was observed for the association
10 between blood Pb levels and performance IQ. A study performed among children in
11 Mexico City observed greater decreases in neurodevelopment with increases in blood
12 levels of Pb and Mn at 12 months, compared to decreases in neurodevelopment observed
13 for increased Pb levels with low levels of Mn (Claus Henn et al.. 2012). No interaction
14 was observed between the two metals and neurodevelopment at 24 months.
15 Overall, studies have reported increased risk of various health effects with exposure to
16 other metals in addition to Pb; however, this is limited by the small number of studies.
17 Toxicological studies, when available, have provided support for these findings.
6.4 Summary
18 Table 6-5 provides an overview of the factors examined as potentially increasing the risk
19 of Pb-related health effects based on the recent evidence integrated across disciplines.
20 They are classified according to the criteria discussed in the introduction to this chapter.
November 2012 6-40 Draft - Do Not Cite or Quote
-------
Table 6-5 Summary of evidence for factors that potentially increase the risk of
Pb-related health effects.
i
2
o
3
4
5
6
1
8
9
10
11
12
13
14
Factor Evaluated
Childhood (Sections 6.2.1 , 6.3.1)
Older Adulthood (Sections 6.2.1, 6.3.1)
Sex (Sections 6.2.2, 6.3.2)
Genetics (Section 6.3.3)
Pre-existing Disease3 (Section 6.3.4)
Smoking Status (Section 6.3.5)
Socioeconomic Status (SES) (Sections 6.2.4, 6.3.6)
Race/Ethnicitv (Sections 6.2.3, 6.3.7)
Proximity to Pb Sources (Section 6.2.5)
Residential Factors (Section 6.2.6)
Body Mass Index (BMI) (Section 6.3.8)
Alcohol Consumption (Section 6.3.9)
Nutrition (Section 6.3.10)
Stress (Section 6.3.11)
Maternal Self-Esteem (Section 6.3.12)
Cognitive Reserve3 (Section 6.3.13)
Other Metals (Section 6.3.14)
Classification
Adequate
Suggestive
Suggestive
Suggestive
Suggestive
Inadequate
Suggestive
Adequate
Adequate
Adequate
Inadequate
Inadequate
Adequate
Suggestive
Inadequate
Inadequate
Suggestive
Possible mediator
There are consistent findings, coherent across disciplines that adequate evidence exists to
conclude that childhood is an at-risk lifestage. Among children, the youngest age groups
were observed to be most at risk of elevated blood Pb levels, with levels decreasing with
increasing age of the children. Children may have increased exposure to Pb compared
with adults because children's behaviors and activities (including increased hand-to-
mouth contact, crawling, and poor hand-washing), differences in diets, and biokinetic
factors. Recent epidemiologic studies of infants/children detected increased risk of
Pb-related health effects, and this was supported by toxicological studies, providing
adequate evidence to conclude that children are an at-risk population. However, this is
based on a limited number of epidemiologic studies, and more studies are needed for
comparing various age groups and examining adolescents.
For adults, elevated Pb biomarkers were associated with increasing age. It is generally
thought that these elevated levels are related to remobilization of stored Pb during bone
loss and/or higher historical Pb exposures. Studies of older adults had inconsistent
November 2012
6-41
Draft - Do Not Cite or Quote
-------
1 findings for effect measure modification of Pb-related mortality but no difference was
2 observed for other health effects. However, toxicological studies support the possibility
3 of age-related differences in Pb-related health effects. The overall evidence is suggestive
4 that older adults are a potential at-risk population based on limited epidemiologic
5 evidence but support from toxicological studies and differential exposure studies.
6 Some studies suggest that males at some ages have higher blood Pb levels than
7 comparably aged females; this was supported by stratifying the total sample of NHANES
8 subjects. Sex-based differences appeared to be prominent among the adolescent and adult
9 age groups but were not observed among the youngest age groups (1-5 years and 6-11
10 years). Studies of effect measure modification of Pb and various health endpoints by sex
11 were inconsistent; although it appears that there are some differences in associations for
12 males and females. This is also observed in toxicological studies. Overall, there is
13 suggestive evidence to conclude that sex is a potential at-risk factor, limited due to
14 inconsistencies between whether males or females are at greater risk of certain outcomes.
15 Regarding race and ethnicity, recent data suggest that the difference in blood Pb levels
16 between black and white subjects is decreasing over time, but black subjects still tend to
17 have higher Pb body burden and Pb exposures than white subjects. Compared to whites,
18 non-white populations were observed to be more at risk of Pb-related health effects;
19 however, this could be related to confounding by factors such as SES or differential
20 exposure levels, which was noted in some of the epidemiologic studies. Studies of
21 race/ethnicity provide adequate evidence that race/ethnicity is an at-risk factor based on
22 the higher exposure observed among non-white populations and some modification
23 observed in studies of associations between Pb levels and health effects.
24 Similar to race and ethnicity, the gap between SES groups with respect to Pb body burden
25 appears to be diminishing. Studies of SES and its relationship with Pb-related health
26 effects are limited and different studies demonstrate increased risk among higher or lower
27 SES groups, providing limited evidence to determine if SES is an at-risk factor for
28 Pb-related health effects. However, biomarkers of Pb exposure have been shown to be
29 higher among lower SES groups even in recent studies in which differences among SES
30 groups have lessened. Therefore, the evidence is suggestive to conclude that low SES is a
31 potential at-risk factor for Pb-related health effects.
32 There is evidence associating proximity to areas with Pb sources, including areas with
33 large industrial sources, with increased Pb body burden and risk of Pb exposure. High
34 concentrations of ambient air Pb have been measured near sources, compared with large
35 urban areas without sources. Additionally, high Pb exposures have been documented near
36 Superfund sites.
November 2012 6-42 Draft - Do Not Cite or Quote
-------
1 Studies utilizing the NHANES dataset have reported increased Pb biomarker measures
2 related to increase house dust Pb levels, homes built after 1950, and renovation of
3 pre-1978 homes. These findings were consistent with those of several high quality
4 studies. Thus, there is adequate evidence that residing in a residence with Pb exposures
5 will increase the risk of Pb-related health effects.
6 There is suggestive evidence to conclude that various genes are potentially modifying the
7 associations between Pb and health effects. Epidemiologic and toxicological studies
8 reported that ALAD variants may increase the risk of Pb-related health effects. Other
9 genes examined that may also affect risk of Pb-related health effects were VDR, DRD4,
10 GSTM1, TNF-a, eNOS, and HFE, although the number of studies examining effect
11 measure modification by these genes was small.
12 Among nutritional factors, diets sufficient in minerals such as Ca2+, Fe, and Zn offer
13 some protection from Pb exposure by preventing or competing with Pb for absorption in
14 the GI tract. Additionally, those with iron deficiencies were observed to be an at-risk
15 population for Pb-related health effects in both epidemiologic and toxicological studies.
16 Thus, there is adequate evidence across disciplines that some nutritional factors
17 contribute to a population being at increased risk. Other nutritional factors, such as Ca2+,
18 Zn, and protein intake, demonstrated the potential to modify associations between Pb and
19 health effects in toxicological studies. Recent epidemiologic studies of these factors were
20 either not performed or observed no effect modification. Folate was also examined in an
21 epidemiologic study of birth size but no interaction was reported between Pb and folate.
22 There was suggestive evidence for several other factors as potentially increasing the risk
23 of Pb-related health effects: pre-existing diseases/conditions, stress, and co-exposure with
24 other metals. Pre-existing diseases/conditions have the potential to affect the risk of
25 Pb-related health effects. Recent epidemiologic studies did not support modification of
26 associations between Pb and health endpoints by the prevalence of diabetes; however,
27 past studies have found individuals with diabetes to be an at-risk population with regard
28 to renal function. Hypertension was observed to be a factor affecting risk in both past and
29 recent epidemiologic studies. Studies of Pb levels and both renal effects and heart rate
30 variability demonstrated greater odds of the associations among hypertensive individuals
31 compared to those that are normotensive. Epidemiologic studies also examined autism as
32 potential factors affecting Pb-related health effects; differences were observed but few
33 studies were available to examine this factor. Stress was evaluated as a factor that
34 potentially increases the risk of Pb-related health outcomes and although limited by the
35 small number of epidemiologic studies, increased stress was observed to negatively
36 impact the association between Pb and health endpoints. Toxicological studies supported
37 this finding. Finally, interactions between Pb and co-exposure with other metals were
November 2012 6-43 Draft - Do Not Cite or Quote
-------
1 evaluated in recent epidemiologic and toxicological studies of health effects. High levels
2 of other metals, such as Cd and Mn, were observed to result in greater effects for the
3 associations between Pb and various health endpoints but evidence was limited due to the
4 small number of studie s.
5 Finally, there was inadequate evidence to conclude that smoking, BMI, alcohol
6 consumption, maternal self-esteem, and cognitive reserve are potential at-risk factors due
7 to limited quantities of studies regarding their effect on Pb-related health outcomes.
8 Epidemiologic studies examining smoking as a factor potentially affecting risk reported
9 mixed findings. It is possible that smoking modifies the effects of only some Pb-related
10 health outcomes. In the limited number of studies, modification of associations between
11 Pb and various health effects (mortality and heart rate variability) was not observed for
12 BMI/obesity. Also, no modification was observed in an epidemiologic study of renal
13 function examining alcohol consumption as a modifier, but a toxicological study
14 supported the potential of alcohol to affect risk. Maternal self-esteem was examined in an
15 epidemiologic study and individuals with mothers who had lower self-esteem had greater
16 Pb-related decreases in MDI and PDI. An epidemiologic study evaluated cognitive
17 reserve as a modifier of the associations between Pb and cognitive and motor functions.
18 Cognitive reserve was an effect measure modifier for the association between Pb and
19 cognitive function but not motor function.
November 2012 6-44 Draft - Do Not Cite or Quote
-------
References for Chapter 6
Absalon. D; Slesak. B. (2010). The effects of changes in cadmium and lead air pollution on cancer
incidence in children. Sci Total Environ 408: 4420-4428.
http://dx.doi.0rg/10.1016/i.scitotenv.2010.06.030
Alghazal. MA; Lenartova. V: Holovska. K; Sobekova. A; Palis. M; Legath. J. (2008a). Activities of
antioxidant and detoxifying enzymes in rats after lead exposure. Acta Vet (Beogr) 77: 347-354.
http://dx.doi.org/10.2754/avb200877030347
Baghurst. PA; McMichael. AJ; Wigg. NR; Vimpani. GV: Robertson. EF; Roberts. RJ: Tong. SL. (1992).
Environmental exposure to lead and children's intelligence at the age of seven years: The Port Pirie
cohort study. N Engl J Med 327: 1279-1284. http://dx.doi.org/10.1056/NEJM199210293271805
Barbosa. F. Jr; Sandrim. VC: Uzuelli. JA; Gerlach. RF; Tanus-Santos. JE. (2006c). eNOS genotype-
dependent correlation between whole blood lead and plasma nitric oxide products concentrations.
Nitric Oxide 14: 58-64. http://dx.doi.Org/10.1016/i.niox.2005.09.007
Basha. MR: Murali. M: Siddiqi. HK: Ghosal. K: Siddiqi. OK: Lashuel. HA: Ge. YW: Lahiri. DK: Zawia.
NH. (2005). Lead (Pb) exposure and its effect on APP proteolysis and A beta aggregation. FASEB J
19: 2083-2084. http://dx.doi.org/10.1096/fi.05-4375fie
Bellinger. D; Leviton. A; Sloman. J. (1990). Antecedents and correlates of improved cognitive
performance in children exposed in utero to low levels of lead. Environ Health Perspect 89: 5-11.
Berrahal. AA; Lasram. M; El Elj. N; Kerkeni. A; Gharbi. N; El-Fazaa. S. (2011). Effect of age-dependent
exposure to lead on hepatotoxicity and nephrotoxicity in male rats. Environ Toxicol 26: 68-78.
http://dx.doi.org/10.1002/tox.20530
Bhatti. P: Stewart. PA; Hutchinson. A; Rothman. N; Linet. MS: Inskip. PD; Rajaraman. P. (2009). Lead
exposure, polymorphisms in genes related to oxidative stress, and risk of adult brain tumors. Cancer
Epidemiol Biomarkers Prev 18: 1841-1848. http://dx.doi.org/10.1158/1055-9965.EPI-09-0197
Bleecker. ML; Ford. DP; Celio. MA; Vaughan. CG: Lindgren. KN. (2007a). Impact of cognitive reserve
on the relationship of lead exposure and neurobehavioral performance. Neurology 69: 470-476.
http://dx.doi.org/10.1212/01.wnl.0000266628.43760.8c
Bogden. JD; Kemp. FW: Han. S: Murphy. M; Fraiman. M; Czerniach. D; Flynn. CJ: Banua. ML;
Scimone. A; Castrovilly. L; Gertner. SB. (1995). Dietary calcium and lead interact to modify maternal
blood pressure, erythropoiesis, and fetal and neonatal growth in rats during pregnancy and lactation. J
Nutr 125: 990-1002.
Brubaker. CJ: Dietrich. KN; Lanphear. BP; Cecil. KM. (2010). The influence of age of lead exposure on
adult gray matter volume. Neurotoxicology 31: 259-266.
http://dx.doi.0rg/10.1016/i.neuro.2010.03.004
Campanella. R; Mielke. HW. (2008). Human geography of New Orleans' high-lead geochemical setting.
Environ Geochem Health 30: 531-540. http://dx.doi.org/10.1007/sl0653-008-9190-9
Cantonwine. D: Hu. H; Tellez-Rojo. MM; Sanchez. BN; Lamadrid-Figueroa. H; Ettinger. AS: Mercado-
Garcia. A; Hernandez-Avila. M; Wright. RO. (2010b). HFE gene variants modify the association
between maternal lead burden and infant birthweight: A prospective birth cohort study in Mexico City,
Mexico. Environ Health Global Access Sci Source 9: 43. http://dx.doi.org/10.1186/1476-069X-9-43
CDC (Centers for Disease Control and Prevention). (2009). Prevalence of autism spectrum disorders ~
Autism and developmental disabilities monitoring network, United States, 2006. MMWR Surveill
Summ 58: 1-20.
November 2012 6-45 Draft - Do Not Cite or Quote
-------
Cecil. KM; Brubaker. CJ: Adler. CM; Dietrich. KN; Altave. M; Egelhoff. JC: Wessel S: Elangovan. I;
Hornung. R; Jarvis. K; Lanphear. BP. (2008). Decreased brain volume in adults with childhood lead
exposure. PLoS Med 5: el 12. http://dx.doi.org/10.1371/iournal.pmed.0050112
Chen. S: Golemboski. KA; Piepenbrink. M; Dietert. RR. (2004). Developmental immunotoxicity of lead
in the rat: Influence of maternal diet. J Toxicol Environ Health A 67: 495-511.
http://dx.doi.org/10.1080/15287390490276520
Chen. Y; Zhao. JX; Liu. JW: Cui. J: Li. L; Tian. W. (2008c). Lack of association of delta-aminolevulinic
acid dehydratase genotype with blood lead levels in environmentally exposed children of Uygur and
Han populations. Acta Paediatr 97: 1717-1720. http://dx.doi.Org/10.llll/j.1651-2227.2008.01003.x
Chia. SE; Zhou. HJ; Yap. E; Tham. MT; Dong. NV; Hong Tu. NT; Chia. KS. (2006). Association of renal
function and delta-aminolevulinic acid dehydratase polymorphism among Vietnamese and Singapore
workers exposed to inorganic lead. Occup Environ Med 63: 180-186.
http://dx.doi.org/10.1136/oem.2005.021154
Glaus Henn. B; Schnaas. L; Ettinger. AS: Schwartz. J: Lamadrid-Figueroa. H; Hernandez-Avila. M;
Amarasiriwardena. C: Hu. H; Bellinger. DC: Wright. RO: Tellez-Rojo. MM. (2012). Associations of
early childhood manganese and lead co-exposure with neurodevelopment. Environ Health Perspect
120: 126-131. http://dx.doi.org/10.1289/ehp.1003300
Corv-Slechta. DA; Virgolini. MB; Rossi-George. A; Thiruchelvam. M; Lisek. R; Weston. D. (2008).
Lifetime consequences of combined maternal lead and stress [Review]. Basic Clin Pharmacol Toxicol
102: 218-227. http://dx.doi.Org/10.llll/i.1742-7843.2007.00189.x
de Souza Lisboa. SF; Goncalves. G: Komatsu. F; Queiroz. CAS: Almeida. AA; Moreira. EG. (2005).
Developmental lead exposure induces depressive-like behavior in female rats. Drug Chem Toxicol 28:
67-77. http://dx.doi.org/10.1081/dct-200039696
Di Lorenzo. L; Silvestroni. A; Martino. MG: Gagliardi. T; Corfiati. M; Soleo. L. (2006). Evaluation of
peripheral blood neutrophil leucocytes in lead-exposed workers. Int Arch Occup Environ Health 79:
491-498. http://dx.doi.org/10.1007/s00420-005-0073-4
Di Lorenzo. L; Vacca. A; Corfiati. M; Lovreglio. P; Soleo. L. (2007). Evaluation of tumor necrosis factor-
alpha and granulocyte colony-stimulating factor serum levels in lead-exposed smoker workers. Int J
Immunopathol Pharmacol 20: 239-247.
Dietrich. KN; Krafft. KM; Shukla. R: Bornschein. RL; Succop. PA. (1987b). The neurobehavioral effects
of early lead exposure. In SR Schroeder (Ed.), Toxic substances and mental retardation:
Neurobehavioral toxicology and teratology (pp. 71-95). Washington, DC: American Association on
Mental Deficiency.
Dixon. SL; Gaitens. JM; Jacobs. DE; Strauss. W: Nagaraia. J: Pivetz. T; Wilson. JW: Ashley. PJ. (2009).
Exposure of US children to residential dust lead, 1999-2004: II. The contribution of lead-contaminated
dust to children's blood lead levels. Environ Health Perspect 117: 468-474.
http://dx.doi.org/10.1289/ehp.11918
Donald. JM; Cutler. MG: Moore. MR. (1987). Effects of lead in the laboratory mouse. Development and
social behaviour after lifelong exposure to 12 microM lead in drinking fluid. Neuropharmacology 26:
391-399. http://dx.doi.org/10.1016/0028-3908(87)90194-8
Donald. JM: Cutler. MG: Moore. MR; Bardlev. M. (1986). Effects of lead in the laboratory mouse~2:
Development and social behaviour after lifelong administration of a small dose of lead acetate in
drinking fluid. Neuropharmacology 25: 151-160.
Dyatlov. VA; Lawrence. DA. (2002). Neonatal lead exposure potentiates sickness behavior induced by
Listeria monocytogenes infection of mice. Brain Behav Immun 16: 477-492.
http://dx.doi.org/10.1006/brbi.2001.0641
Elmarsafawv. SF; Jain. NB; Schwartz. J: Sparrow. D; Nie. HL; Hu. H. (2006). Dietary calcium as a
potential modifier of the relationship of lead burden to blood pressure. Epidemiology 17: 531-537.
http://dx.doi.org/10.1097/01.ede.0000231285.86968.2b
November 2012 6-46 Draft - Do Not Cite or Quote
-------
Fadrowski. JJ; Navas-Acien. A; Tellez-Plaza. M; Guallar. E; Weaver. VM; Furth. SL. (2010). Blood lead
level and kidney function in US adolescents: The Third National Health and Nutrition Examination
Survey. Arch Intern Med 170: 75-82. http://dx.doi.org/10.1001/archinternmed.2009.417
Filippelli. GM; Laidlaw. MAS: Latimer. JC: Raftis. R. (2005). Urban lead poisoning and medical
geology: An unfinished story. Geological Society of America Today 15: 4-11.
httrj://dx.doi.org/10.1130/1052-5173(2005)015<4:ULPAMG>2.0.CO:2
Franklin. CA; Inskip. MJ; Baccanale. CL; Edwards. CM; Manton. WI; Edwards. E; O'Flahertv. EJ.
(1997). Use of sequentially administered stable lead isotopes to investigate changes in blood lead
during pregnancy in a nonhuman primate (Macaca fascicularis). Toxicol Sci 39: 109-119.
Froehlich. TE; Lanphear. BP; Auinger. P; Hornung. R; Epstein. JN; Braun. J; Kahn. RS. (2009).
Association of tobacco and lead exposures with attention-deficit/hyperactivity disorder. Pediatrics
124: E1054-E1063. http://dx.doi.org/10.1542/peds.2009-0738
Froehlich. TE; Lanphear. BP; Dietrich. KN; Cory-Slechta. DA; Wang. N; Kahn. RS. (2007). Interactive
effects of a DRD4 polymorphism, lead and sex on executive functions in children. Biol Psychiatry 62:
243-249. http://dx.doi.0rg/10.1016/i.biopsvch.2006.09.039
Fullmer. CS. (1992). Intestinal interactions of lead and calcium [Review]. Neurotoxicology 13: 799-807.
Fullmer. CS: Edelstein. S: Wasserman. RH. (1985). Lead-binding properties of intestinal calcium-binding
proteins. J Biol Chem 260: 6816-6819.
Gaitens. JM; Dixon. SL; Jacobs. DE; Nagaraia. J: Strauss. W: Wilson. JW: Ashley. PJ. (2009). Exposure
of US children to residential dust lead, 1999-2004:1. Housing and demographic factors. Environ
Health Perspect 117: 461-467. http://dx.doi.org/10.1289/ehp.11917
Gao. A; Lu. XT; Li. QY; Tian. L. (2010a). Effect of the delta-aminolevulinic acid dehydratase gene
polymorphism on renal and neurobehavioral function in workers exposed to lead in China. Sci Total
Environ 408: 4052-4055. http://dx.doi.Org/10.1016/i.scitotenv.2010.04.024
Garrido Latorre. F; Hernandez-Avila. M; Orozco. JT; Medina. CAA; Aro. A; Palazuelos. E; Hu. H.
(2003). Relationship of blood and bone lead to menopause and bone mineral density among middle-
age women in Mexico City. Environ Health Perspect 111: 631-636.
Glass. TA; Bandeen-Roche. K; McAtee. M; Bolla. K; Todd. AC: Schwartz. BS. (2009). Neighborhood
psychosocial hazards and the association of cumulative lead dose with cognitive function in older
adults. Am J Epidemiol 169: 683-692. http://dx.doi.org/10.1093/aie/kwn390
Gollenberg. AL; Hediger. ML; Lee. PA; Himes. JH; Buck Louis. GM. (2010). Association between lead
and cadmium and reproductive hormones in peripubertal U.S. girls. Environ Health Perspect 118:
1782-1787. http://dx.doi.org/10.1289/ehp.1001943
Gulson. B; Mizon. K; Smith. H; Eisman. J: Palmer. J: Korsch. M; Donnelly. J: Waite. K. (2002). Skeletal
lead release during bone resorption: Effect of bisphosphonate treatment in a pilot study. Environ
Health Perspect 110: 1017-1023.
Gulson. B; Mizon. KJ; Palmer. JM; Korsch. MJ: Taylor. AJ; Mahaffev. KR. (2004a). Blood lead changes
during pregnancy and postpartum with calcium supplementation. Environ Health Perspect 112:
1499-1507. http://dx.doi.org/10.1289/ehp.6548
Gulson. BL; Jameson. CW: Mahaffev. KR; Mizon. KJ: Korsch. MJ: Vimpani. G. (1997). Pregnancy
increases mobilization of lead from maternal skeleton. Transl Res 130: 51-62.
http://dx.doi.org/10.1016/S0022-2143(97)90058-5
Han. S: Li. W: Jamil. U; Dargan. K; Orefice. M; Kemp. FW: Bogden. JD. (1999). Effects of weight loss
and exercise on the distribution of lead and essential trace elements in rats with prior lead exposure.
Environ Health Perspect 107: 657-662. http://dx.doi.org/10.1289/ehp.99107657
Hertz-Picciotto. I; Schramm. M; Watt-Morse. M; Chantala. K; Anderson. J: Osterloh. J. (2000). Patterns
and determinants of blood lead during pregnancy. Am J Epidemiol 152: 829-837.
http://dx.doi.0rg/10.1093/aie/152.9.829
November 2012 6-47 Draft - Do Not Cite or Quote
-------
Jamieson. JA; Stringer. DM; Zahradka. P; Taylor. CG. (2008). Dietary zinc attenuates renal lead
deposition but metallothionein is not directly involved. Biometals 21: 29-40.
http://dx.doi.org/10.1007/sl0534-007-9090-v
Jamieson. JA; Taylor. CG: Weiler. HA. (2006). Marginal zinc deficiency exacerbates bone lead
accumulation and high dietary zinc attenuates lead accumulation at the expense of bone density in
growing rats. Toxicol Sci 92: 286-294. http://dx.doi.org/10.1093/toxsci/kfi201
Jedrychowski. W: Perera. F; Jankowski. J; Mrozek-Budzyn. D; Mroz. E; Flak. E; Edwards. S: Skarupa.
A; Lisowska-Miszczyk. I. (2009a). Gender specific differences in neurodevelopmental effects of
prenatal exposure to very low-lead levels: The prospective cohort study in three-year olds. Early Hum
Dev 85: 503-510. http://dx.doi.Org/10.1016/j.earlhumdev.2009.04.006
Jones. EA; Wright. JM; Rice. G: Buckley. BT; Magsumbol. MS: Barr. DB; Williams. BL. (2010). Metal
exposures in an inner-city neonatal population. Environ Int 36: 649-654.
http://dx.doi.0rg/10.1016/i.envint.2010.04.007
Jones. RL; Homa. DM; Meyer. PA; Brodv. DJ: Caldwell. KL; Pirkle. JL; Brown. MJ. (2009a). Trends in
blood lead levels and blood lead testing among US children aged 1 to 5 Years, 1988-2004. Pediatrics
123: e376-e385. http://dx.doi.org/10.1542/peds.2007-3608
Joseph. CLM; Havstad. S: Ownbv. PR; Peterson. EL; Maliarik. M; McCabe. MJ. Jr; Barone. C: Johnson.
CC. (2005). Blood lead level and risk of asthma. Environ Health Perspect 113: 900-904.
http://dx.doi.org/10.1289/ehp.7453
Jurczuk. M; Moniuszko-Jakoniuk. J; Brzoska. MM. (2006). Involvement of some low-molecular thiols in
the peroxidative mechanisms of lead and ethanol action on rat liver and kidney. Toxicology 219:
11-21. http://dx.doi.0rg/10.1016/i.tox.2005.10.022
Kim. JH: Lee. KH: Yoo. DH: Kang. D: Cho. SH: Hong. YC. (2007). GSTM1 and TNF-alpha gene
polymorphisms and relations between blood lead and inflammatory markers in a non-occupational
population. Mutat Res Genet Toxicol Environ Mutagen 629: 32-39.
http://dx.doi.0rg/10.1016/i.mrgentox.2007.01.004
Kim. Y; Kim. BN; Hong. YC: Shin. MS: Yoo. HJ; Kim. JW: Bhang. SY; Cho. SC. (2009b). Co-exposure
to environmental lead and manganese affects the intelligence of school-aged children.
Neurotoxicology 30: 564-571. http://dx.doi.Org/10.1016/i.neuro.2009.03.012
Kiziler. AR; Aydemir. B; Onaran. I; Alici. B; Ozkara. H; Gulyasar. T; Akyolcu. MC. (2007). High levels
of cadmium and lead in seminal fluid and blood of smoking men are associated with high oxidative
stress and damage in infertile subjects. Biol Trace Elem Res 120: 82-91.
http://dx.doi.org/10.1007/sl2011-007-8020-8
Kordas. K; Ettinger. AS: Bellinger. DC: Schnaas. L; Tellez Rojo. MM; Hernandez-Avila. M; Hu. H;
Wright. RO. (2011). A dopamine receptor (DRD2) but not dopamine transporter (DAT1) gene
polymorphism is associated with neurocognitive development of Mexican preschool children with
lead exposure. J Pediatr 159: 638-643. http://dx.doi.Org/10.1016/j.jpeds.2011.03.043
Kordas. K; Ettinger. AS: Lamadrid-Figueroa. H; Tellez-Rojo. MM; Hernandez-Avila. M; Hu. H; Wright.
RO. (2009). Methylenetetrahydrofolate reductase (MTHFR) C677T, A1298C and Gl 793 A genotypes,
and the relationship between maternal folate intake, tibia lead and infant size at birth. Br J Nutr 102:
907-914. http://dx.doi.org/10.1017/s0007114509318280
Kordas. K; Queirolo. El; Ettinger. AS: Wright. RO: Stoltzfus. RJ. (2010). Prevalence and predictors of
exposure to multiple metals in preschool children from Montevideo, Uruguay. Sci Total Environ 408:
4488-4494. http://dx.doi.Org/10.1016/i.scitotenv.2010.06.041
Krieg. EF. Jr; Butler. MA; Chang. MH; Liu. TB; Yesupriya. A; Lindegren. ML; Dowling. N. (2009). Lead
and cognitive function in ALAD genotypes in the Third National Health and Nutrition Examination
Survey. Neurotoxicol Teratol 31: 364-371. http://dx.doi.Org/10.1016/i.ntt.2009.08.003
November 2012 6-48 Draft - Do Not Cite or Quote
-------
Krieg. EF. Jr; Butler. MA; M-h. C: Liu. T; Yesupriya. A; Dowling. N; Lindegren. ML. (2010). Lead and
cognitive function in VDR genotypes in the Third National Health and Nutrition Examination Survey.
Neurotoxicol Teratol 32: 262-272. http://dx.doi.Org/10.1016/i.ntt.2009.12.004
Lagerkvist. BJ; Ekesrydh. S: Englyst. V: Nordberg. GF; Soderberg. HA; Wiklund. DE. (1996). Increased
blood lead and decreased calcium levels during pregnancy: A prospective study of Swedish women
living near a smelter. Am J Public Health 86: 1247-1252.
Laidlaw. MAS: Filippelli. GM. (2008). Resuspension of urban soils as a persistent source of lead
poisoning in children: A review and new directions. Appl Geochem 23: 2021-2039.
http://dx.doi.0rg/10.1016/i.apgeochem.2008.05.009
Lamadrid-Figueroa. H; Tellez-Rojo. MM; Hernandez-Cadena. L; Mercado-Garcia. A; Smith. D; Solano-
Gonzalez. M; Hernandez-Avila. M; Hu. H. (2006). Biological markers of fetal lead exposure at each
stage of pregnancy. J Toxicol Environ Health A 69: 1781-1796.
http://dx.doi.org/10.1080/15287390600630195
Lanphear. BP; Matte. TD; Rogers. J: Clickner. RP; Dietz. B; Bornschein. RL; Succop. P; Mahaffev. KR;
Dixon. S: Galke. W: Rabinowitz. M; Farfel. M; Rohde. C: Schwartz. J: Ashley. P; Jacobs. DE. (1998).
The contribution of lead-contaminated house dust and residential soil to children's blood lead levels: A
pooled analysis of 12 epidemiologic studies. Environ Res 79: 51-68.
http://dx.doi.org/10.1006/enrs.1998.3859
Laxen. DPH; Raab. GM; Fulton. M. (1987). Children's blood lead and exposure to lead in household dust
and water ~ a basis for an environmental standard for lead in dust. Sci Total Environ 66: 235-244.
Leasure. JL; Giddabasappa. A; Chanev. S: Johnson. JE; Pothakos. K; Lau. YS: Fox. DA. (2008). Low-
level human equivalent gestational lead exposure produces sex-specific motor and coordination
abnormalities and late-onset obesity in year-old mice. Environ Health Perspect 116: 355-361.
http://dx.doi.org/10.1289/ehp.10862
Lee. JE; Naqi. SA; Kao. E; Dietert. RR. (2002). Embryonic exposure to lead: Comparison of immune and
cellular responses in unchallenged and virally stressed chickens. Arch Toxicol 75: 717-724.
http://dx.doi.org/10.1007/s00204-001-0283-9
Leggett. RW. (1993). An age-specific kinetic model of lead metabolism in humans. Environ Health
Perspect 101: 598-616.
Levin. R; Brown. MJ; Kashtock. ME; Jacobs. DE; Whelan. EA; Rodman. J: Schock. MR; Padilla. A;
Sinks. T. (2008). Lead exposures in US children, 2008: Implications for prevention. Environ Health
Perspect 116: 1285-1293. httrj://dx.doi.org/10.1289/ehp.H241
Liu. J; McCaulev. L; Compher. C: Yan. C: Shen. X; Needleman. H; Pinto-Martin. JA. (2011 a). Regular
breakfast and blood lead levels among preschool children. Environ Health Global Access Sci Source
10: 28. http://dx.doi.org/10.1186/1476-069X-10-28
Mahaffev. KR; Annest. JL. (1986). Association of erythrocyte protoporphyrin with blood lead level and
iron status in the second national health and nutrition examination survey, 1976-1980. Environ Res 41:
327-338. http://dx.doi.org/10.1016/80013-9351(86)80194-3
Marcus. AH; Schwartz. J. (1987). Dose-response curves for erythrocyte protoporphyrin vs blood lead:
Effects of iron status. Environ Res 44: 221-227. http://dx.doi.org/10.1016/S0013-9351(87)80230-X
Martuzevicius. D; Grinshpun. SA; Reponen. T; Gorny. RL; Shukla. R; Lockev. J: Hu. S: McDonald. R;
Biswas. P: Kliucininkas. L; LeMasters. G. (2004). Spatial and temporal variations of PM2.5
concentration and composition throughout an urban area with high freeway density: The Greater
Cincinnati study. Atmos Environ 38: 1091-1105. http://dx.doi.Org/10.1016/j.atmosenv.2003.ll.015
Mason. H; Gallagher. F; Sen. D. (2005). Window renovation and exposure to lead—an observational
study. Occup Med (Lond) 55: 631-634. http://dx.doi.org/10.1093/occmed/kqil59
November 2012 6-49 Draft - Do Not Cite or Quote
-------
Masso-Gonzalez. EL; Antonio-Garcia. MT. (2009). Natural antioxidants protect against lead-induced
damage during pregnancy and lactation in rat's pups. Ecotoxicol Environ Saf 72: 2137-2142.
http://dx.doi.0rg/10.1016/i.ecoenv.2009.03.013
McGivern. RF; Sokol. RZ; Berman. NG. (1991). Prenatal lead exposure in the rat during the third week
of gestation: Long-term behavioral, physiological and anatomical effects associated with reproduction.
Toxicol Appl Pharmacol 110: 206-215. http://dx.doi.org/10.1016/S0041-008X(05)80003-l
McMichael. AJ; Baghurst. PA; Vimpani. GV: Robertson. EF; Wigg. NR; Tong. SL. (1992).
Sociodemographic factors modifying the effect of environmental lead on neuropsychological
development in early childhood. Neurotoxicol Teratol 14: 321-327.
http://dx.doi.org/10.1016/0892-0362(92)90038-C
McNeill. FE; Stokes. L; Brito. JA; Chettle. PR; Kave. WE. (2000). 109Cd K x-ray fluorescence
measurements of tibial lead content in young adults exposed to lead in early childhood. Occup
Environ Med 57: 465-471. http://dx.doi.Org/10.1136/oem.57.7.465
Menke. A; Muntner. P; Batuman. V; Silbergeld. EK; Guallar. E. (2006). Blood lead below 0.48
micromol/L (10 microg/dL) and mortality among US adults. Circulation 114: 1388-1394.
http://dx.doi.org/10.1161/circulationaha.106.628321
Mielke. HW: Laidlaw. MA; Gonzales. CR. (2011c). Estimation of leaded (Pb) gasoline's continuing
material and health impacts on 90 US urbanized areas [Review]. Environ Int 37: 248-257.
http://dx.doi.0rg/10.1016/i.envint.2010.08.006
Mielke. HW: Powell. ET; Shah. A; Gonzales. CR: Mielke. PW. (2001). Multiple metal contamination
from house paints: Consequences of power sanding and paint scraping in New Orleans. Environ
Health Perspect 109: 973-978. http://dx.doi.org/10.1289/ehp.01109973
Miranda. ML; Edwards. SE; Swamy. GK; Paul. CJ: Neelon. B. (2010). Blood lead levels among pregnant
women: Historical versus contemporaneous exposures. Int J Environ Res Public Health 7: 1508-1519.
http://dx.doi.org/10.3390/iierph7041508
Miranda. ML; Kim. D; Galeano. M; Paul. CJ: Hull. AP; Morgan. SP (2007a). The relationship between
early childhood blood lead levels and performance on end of grade tests. Environ Health Perspect 115:
1242-1247. http://dx.doi.org/10.1289/ehp.9994
Miyaki. K; Lwin. H; Masaki. K; Song. YX; Takahashi. Y; Muramatsu. M; Nakavama. T. (2009).
Association between a polymorphism of aminolevulinate dehydrogenase (ALAD) gene and blood lead
levels in Japanese subjects. Int J Environ Res Public Health 6: 999-1009.
http://dx.doi.org/10.3390/ijerph6030999
Montenegro. MF; Barbosa. F. Jr; Sandrim. VC: Gerlach. RF; Tanus-Santos. JE. (2006). A polymorphism
in the delta-aminolevulinic acid dehydratase gene modifies plasma/whole blood lead ratio. Arch
Toxicol 80: 394-398. http://dx.doi.org/10.1007/s00204-005-0056-v
Muntner. P; Menke. A; DeSalvo. KB; Rabito. FA; Batuman. V. (2005). Continued decline in blood lead
levels among adults in the United States - The National Health and Nutrition Examination Surveys.
Arch Intern Med 165: 2155-2161. http://dx.doi.org/10.1001/archinte.165.18.2155
Muwakkit. S: Nuwayhid. I; Nabulsi. M; al Haij. R; Khoury. R; Mikati. M; Abboud. MR. (2008). Iron
deficiency in young Lebanese children: Association with elevated blood lead levels. J Pediatr Hematol
Oncol 30: 382-386. http://dx.doi.org/10.1097/MPH.Ob013e318165b283
Mykkanen. HM; Wasserman. RH. (1981). Gastrointestinal absorption of lead (203Pb) in chicks: Influence
of lead, calcium, and age. JNutr 111: 1757-1765.
Navas-Acien. A; Tellez-Plaza. M; Guallar. E; Muntner. P; Silbergeld. E; Jaar. B; Weaver. V. (2009). Blood
cadmium and lead and chronic kidney disease in US adults: Ajoint analysis. Am J Epidemiol 170:
1156-1164. http://dx.doi.org/10.1093/aje/kwp248
NCHS (National Center for Health Statistics). (2008). 2007-2008 NHANES: Demographics. Retrieved
from ftp://ftp.cdc.gov/pub/Health Statistics/NCHS/nhanes/2007-2008/DEMO E.xpt
November 2012 6-50 Draft - Do Not Cite or Quote
-------
NCHS (National Center for Health Statistics). (2009). NHANES: 2007-2008 data documentation,
codebook, and frequencies: Blood cadmium and lead. Atlanta, GA: Centers for Disease Control and
Prevention, http://www.cdc.gov/nchs/nhanes/nhanes2007-2008/pbcd e.htm
Nriagu. J: Burt. B; Linder. A; Ismail. A; Sohn. W. (2006). Lead levels in blood and saliva in a low-income
population of Detroit, Michigan. Int J Hyg Environ Health 209: 109-121.
http://dx.doi.0rg/10.1016/i.iiheh.2005.ll.005
O'Flahertv. EJ. (1995). Physiologically based models for bone-seeking elements: V. Lead absorption and
disposition in childhood [Review]. Toxicol Appl Pharmacol 131: 297-308.
http://dx.doi.org/10.1006/taap. 1995.1072
O'Flahertv. EJ. (1998). A physiologically based kinetic model for lead in children and adults [Review].
Environ Health Perspect 106: 1495-1503. http://dx.doi.org/10.1289/ehp.98106s61495
Olnev. JW. (1969). Brain lesions, obesity and others disturbances in mice treated with monosodium
glutamate. Science 164: 719-721. http://dx.doi.org/10.1126/science.164.3880.719
Park. SK; Schwartz. J: Weisskopf. M; Sparrow. D; Vokonas. PS: Wright. RO: Coull. B; Nie. H; Hu. H.
(2006). Low-level lead exposure, metabolic syndrome, and heart rate variability: The VA Normative
Aging Study. Environ Health Perspect 114: 1718-1724.
Peters. JL; Kubzansky. L; McNeely. E; Schwartz. J: Spiro. A. Ill; Sparrow. D; Wright. RO: Nie. H; Hu. H.
(2007). Stress as a potential modifier of the impact of lead levels on blood pressure: The Normative
Aging Study. Environ Health Perspect 115: 1154-1159. http://dx.doi.org/10.1289/ehp.10002
Peters. JL; Wright. RJ: Weisskopf. MG: Spiro. A. Ill; Schwartz. J: Sparrow. D: Nie. H; Hu. H; Wright.
RO. (2008). Interaction of stress and lead burden on cognition in older men: The VA Normative Aging
Study [Abstract]. Epidemiology 19: S134. http://dx.doi.org/10.1097/01.ede.0000339926.08729.35
Pillai. P; Patel. R; Pandva. C: Gupta. S. (2009). Sex-specific effects of gestational and lactational
coexposure to lead and cadmium on hepatic phase I and phase II xenobiotic/steroid-metabolizing
enzymes and antioxidant status. J Biochem Mol Toxicol 23: 419-431.
http://dx.doi.org/10.1002/ibt.20305
Pilsner. JR; Hu. H; Wright. RO: Kordas. K; Ettinger. AS: Sanchez. BN; Cantonwine. D: Lazarus. AL;
Cantoral. A; Schnaas. L; Tellez-Rojo. MM; Hernandez-Avila. M. (2010). Maternal MTHFR genotype
and haplotype predict deficits in early cognitive development in a lead-exposed birth cohort in Mexico
City. Am J Clin Nutr 92: 226-234. http://dx.doi.org/10.3945/aicn.2009.28839
Pineda-Zavaleta. AP; Garcia-Vargas. G: Borja-Aburto. VH; Acosta-Saavedra. LC: Aguilar. EV; Gomez-
Mufioz. A; Cebrian. ME; Calderon-Aranda. ES. (2004). Nitric oxide and superoxide anion production
in monocytes from children exposed to arsenic and lead in region Lagunera, Mexico. Toxicol Appl
Pharmacol 198: 283-290. http://dx.doi.Org/10.1016/i.taap.2003.10.034
Pirkle. JL: Brodv. DJ: Gunter. EW: Kramer. RA; Paschal. DC: Flegal. KM; Matte. TD. (1994). The
decline in blood lead levels in the United States: The National Health and Nutrition Examination
Surveys (NHANES). JAMA 272: 284-291. http://dx.doi.org/10.1001/iama.1994.03520040046039
Pizent. A; Macan. J: Jurasovic. J: Varnai. VM; Milkovic-Kraus. S: Kanceljak-Macan. B. (2008).
Association of toxic and essential metals with atopy markers and ventilatory lung function in women
and men. Sci Total Environ 390: 369-376. http://dx.doi.Org/10.1016/i.scitotenv.2007.10.049
Pleis. JR: Lucas. JW: Ward. BW. (2009). Summary health statistics for U.S. adults: National Health
Interview Survey, 2008. (DHHS 2010-1570). Hyattsville, MD: National Center for Health Statistics.
http://www.cdc.gov/nchs/data/series/sr 10/srlO 242.pdf
Popovic. M; McNeil!. FE; Chettle. PR; Webber. CE; Lee. CV: Kave. WE. (2005). Impact of occupational
exposure on lead levels in women. Environ Health Perspect 113: 478-484.
http://dx.doi.org/10.1289/ehp.7386
November 2012 6-51 Draft - Do Not Cite or Quote
-------
Rabstein. S: Unfried. K; Ranft. U: Illig. T; Kolz. M; Mambetova. C: Vlad. M; Roman. C: Weiss. T;
Becker. D; Bruning. T; Pesch. B. (2008). Lack of association of delta-aminolevulinate dehydratase
polymorphisms with blood lead levels and hemoglobin in Romanian women from a lead-contaminated
region. J Toxicol Environ Health A 71: 716-724. http://dx.doi.org/10.1080/15287390801985190
Rajan. P; Kelsev. KT; Schwartz. JD; Bellinger. DC: Weuve. J: Spiro. A. Ill; Sparrow. D; Smith. TJ; Nie.
H; Weisskopf. MG: Hu. H; Wright. RO. (2008). Interaction of the delta-aminolevulinic acid
dehydratase polymorphism and lead burden on cognitive function: The VA Normative Aging Study. J
Occup Environ Med 50: 1053-1061. http://dx.doi.org/10.1097/JOM.Ob013e3181792463
Rajaraman. P; Stewart. PA; Samet. JM; Schwartz. BS: Linet. MS: Zahm. SH; Rothman. N; Yeager. M;
Fine. HA; Black. PM; Loeffler. J: Shapiro. WR; Selker. RG: Inskip. PP. (2006). Lead, genetic
susceptibility, and risk of adult brain tumors. Cancer Epidemiol Biomarkers Prev 15: 2514-2520.
http://dx.doi.org/10.1158/1055-9965.EPI-06-0482
Rezende. VB; Barbosa. F. Jr; Montenegro. MF; Sandrim. VC: Gerlach. RF; Tanus-Santos. JE. (2008).
Haplotypes of vitamin D receptor modulate the circulating levels of lead in exposed subjects. Arch
Toxicol 82: 29-36. http://dx.doi.org/10.1007/s00204-007-0231-4
Ris. MD; Dietrich. KN; Succop. PA; Berger. OG: Bornschein. RL. (2004). Early exposure to lead and
neuropsychological outcome in adolescence. J Int Neuropsychol Soc 10: 261-270.
http://dx.doi.org/10.1017/S1355617704102154
Rossi-George. A; Virgolini. MB; Weston. D; Corv-Slechta. DA. (2009). Alterations in glucocorticoid
negative feedback following maternal Pb, prenatal stress and the combination: A potential biological
unifying mechanism for their corresponding disease profiles. Toxicol Appl Pharmacol 234: 117-127.
http://dx.doi.0rg/10.1016/i.taap.2008.10.003
Rothenberg. SJ: Karchmer. S: Schnaas. L; Perroni. E; Zea. F; Alba. JF. (1994a). Changes in serial blood
lead levels during pregnancy. Environ Health Perspect 102: 876-880.
http://dx.doi.org/10.1289/ehp.94102876
Sarasua. SM; Vogt. RF; Henderson. LO; Jones. PA; Lybarger. JA. (2000). Serum immunoglobulins and
lymphocyte subset distributions in children and adults living in communities assessed for lead and
cadmium exposure. J Toxicol Environ Health A 60: 1-15. http://dx.doi.org/10.1080/009841000156556
Saxena. DK; Singh. C: Murthy. RC: Chandra. SV. (1991). Lead distribution in iron-deficient rats: I. Lead
exposure during the period of organogenesis. Nutr Res 11: 471-477.
http://dx.doi.org/10.1016/80271-5317(05)80009-0
Schell. LM; Denham. M; Stark. AD; Ravenscroft. J: Parsons. P: Schulte. E. (2004). Relationship between
blood lead concentration and dietary intakes of infants from 3 to 12 months of age. Environ Res 96:
264-273. http://dx.doi.0rg/10.1016/i.envres.2004.02.008
Schober. SB; Mirel. LB; Graubard. BI; Brodv. DJ: Flegal. KM. (2006). Blood lead levels and death from
all causes, cardiovascular disease, and cancer: Results from the NHANES III Mortality Study. Environ
Health Perspect 114: 1538-1541. http://dx.doi.org/10.1289/ehp.9123
Schuhmacher. M; Hernandez. M; Domingo. JL; Fernandez-Ballart JD: Llobet JM; Corbella. J. (1996). A
longitudinal study of lead mobilization during pregnancy: Concentrations in maternal and umbilical
cord blood. Trace Elem Electroly 13: 177-181.
Schwartz. BS: Lee. BK; Lee. GS: Stewart. WF; Simon. D; Kelsev. K; Todd. AC. (2000a). Associations of
blood lead, dimercaptosuccinic acid-chelatable lead, and tibia lead with polymorphisms in the vitamin
D receptor and delta-aminolevulinic acid dehydratase genes. Environ Health Perspect 108: 949-954.
http://dx.doi.org/10.2307/3435053
Schwartz. J. (1994). Low-level lead exposure and children's IQ: A meta-analysis and search for a
threshold. Environ Res 65: 42-55. http://dx.doi.org/10.1006/enrs.1994.1020
Scinicariello. F; Murray. HE; Moffett. DB; Abadin. HG: Sexton. MJ; Fowler. BA. (2007). Lead and delta-
aminolevulinic acid dehydratase polymorphism: Where does it lead? A meta-analysis. Environ Health
Perspect 115: 35-41. http://dx.doi.org/10.1289/ehp.9448
November 2012 6-52 Draft - Do Not Cite or Quote
-------
Scinicariello. F; Yesupriya. A; Chang. MH; Fowler. BA. (2010). Modification by ALAD of the
association between blood lead and blood pressure in the U.S. population: Results from the Third
National Health and Nutrition Examination Survey. Environ Health Perspect 118: 259-264.
http://dx.doi.org/10.1289/ehp.0900866
Selevan. SG: Rice. DC: Hogan. KA; Euling. SY; Pfahles-Hutchens. A; Bethel. J. (2003). Blood lead
concentration and delayed puberty in girls. N Engl J Med 348: 1527-1536.
http://dx.doi.org/10.1056/NEJMoa020880
Shaik. AP; Jamil. K. (2009). Individual susceptibility and genotoxicity in workers exposed to hazardous
materials like lead. J Hazard Mater 168: 918-924. http://dx.doi.Org/10.1016/i.jhazmat.2009.02.129
Sherry. B; Blanck. HM; Galuska. DA; Pan. L; Dietz. WH; Balluz. L. (2010). Vital signs: State-specific
obesity prevalence among adults - United States, 2009. MMWR Recomm Rep 59: 951-955.
Singh. C: Saxena. DK; Murthy. RC: Chandra. SV. (1993b). Embryo-fetal development influenced by lead
exposure in iron-deficient rats. Hum Exp Toxicol 12: 25-28.
http://dx.doi.org/10.1177/096032719301200105
Six. KM; Gover. RA. (1970). Experimental enhancement of lead toxicity by low dietary calcium. Transl
Res 76: 933-942.
Sobekova. A; Holovska. K; Lenartova. V: Legath. J: Javorsky. P. (2009). The alteration of glutathione
peroxidase activity in rat organs after lead exposure. Acta Physiol Hung 96: 37-44.
http://dx.doi.Org/10.1556/APhvsiol.96.2009.l.4
Sobin. C: Gutierrez. M; Alterio. H. (2009). Polymorphisms of delta-aminolevulinic acid dehydratase
(ALAD) and peptide transporter 2 (PEPT2) genes in children with low-level lead exposure.
Neurotoxicology 30: 881-887. http://dx.doi.Org/10.1016/i.neuro.2009.08.006
Songdej. N; Winters. PC: McCabe. MJ. Jr; van Wijngaarden. E. (2010). A population-based assessment of
blood lead levels in relation to inflammation. Environ Res 110: 272-277.
http://dx.doi.0rg/10.1016/i.envres.2009.12.008
SSDAN CensusScope (Social Science Data Analysis Network, CensusScope). (2010a). United States:
Age distribution [Database]. Ann Arbor, Michigan: Social Science Data Analysis Network. Retrieved
from http://www.censusscope.org/us/chart age.html
SSDAN CensusScope (Social Science Data Analysis Network, CensusScope). (2010b). United States:
Population by race [Database]. Ann Arbor, Michigan. Retrieved from
http://www.censusscope.org/us/chart race.html
SSDAN CensusScope (Social Science Data Analysis Network, CensusScope). (2010c). United States:
Poverty by age [Database]. Ann Arbor, Michigan. Retrieved from
http://www.censusscope.org/us/chart_povertv.html
Surkan. PJ; Schnaas. L; Wright. RJ: Tellez-Rojo. MM; Lamadrid-Figueroa. H; Hu. H; Hernandez-Avila.
M; Bellinger. DC: Schwartz. J: Perroni. E; Wright. RO. (2008). Maternal self-esteem, exposure to
lead, and child neurodevelopment. Neurotoxicology 29: 278-285.
http://dx.doi.0rg/10.1016/i.neuro.2007.ll.006
Theppeang. K; Glass. TA; Bandeen-Roche. K; Todd. AC: Rohde. CA; Schwartz. BS. (2008b). Gender and
race/ethnicity differences in lead dose biomarkers. Am J Public Health 98: 1248-1255.
http://dx.doi.org/10.2105/aiph.2007.118505
Theppeang. K; Schwartz. BS: Lee. BK; Lustberg. ME; Silbergeld. EK; Kelsev. KT; Parsons. PJ: Todd.
AC. (2004). Associations of patella lead with polymorphisms in the vitamin D receptor, delta-
aminolevulinic acid dehydratase and endothelial nitric oxide synthase genes. J Occup Environ Med
46: 528-537.
November 2012 6-53 Draft - Do Not Cite or Quote
-------
Tian. Y; Green. PG: Stamova. B; Hertz-Picciotto. I; Pessah. IN; Hansen. R; Yang. X; Gregg. JP;
Ashwood. P; Jickling. G: Van de Water. J; Sharp. FR. (2011). Correlations of gene expression with
blood lead levels in children with autism compared to typically developing controls. Neurotox Res 19:
1-13. http://dx.doi.org/10.1007/sl2640-009-9126-x
Tong. S: McMichael. AJ; Baghurst. PA. (2000). Interactions between environmental lead exposure and
sociodemographic factors on cognitive development. Arch Environ Health 55: 330-335.
http://dx.doi.org/10.1080/00039890009604025
Tsaih. SW: Korrick. S: Schwartz. J; Amarasiriwardena. C: Aro. A; Sparrow. D; Hu. H. (2004). Lead,
diabetes, hypertension, and renal function: The Normative Aging Study. Environ Health Perspect 112:
1178-1182. http://dx.doi.org/10.1289/ehp.7024
U.S. Census Bureau. (2010). U.S. population projections [Database]. Retrieved from
http://www.census.gov/population/www/projections/projectionsagesex.html
U.S. EPA (U.S. Environmental Protection Agency). (2006b). Air quality criteria for lead: Volume I of II
[EPA Report]. (EPA/600/R-05/144aF). Research Triangle Park, NC.
http ://cfpub. epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
U.S. EPA (U.S. Environmental Protection Agency). (2009a). Integrated science assessment for paniculate
matter [EPA Report]. (EPA/600/R-08/139F). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=216546
U.S. EPA (U.S. Environmental Protection Agency). (2010). Integrated science assessment for carbon
monoxide [EPA Report]. (EPA/600/R-09/019F). Research Triangle Park, NC.
http ://cfpub .epa. gov/ncea/cfm/recordisplav. cfm?deid=218686
Uzbekov. MG: Bubnova. NI; Kulikova. GV. (2007). Effect of prenatal lead exposure on superoxide
dismutase activity in the brain and liver of rat fetuses. Bull Exp Biol Med 144: 783-785.
http://dx.doi.org/10.1007/sl0517-007-0431-l
Virgolini. MB; Rossi-George. A; Lisek. R; Weston. DP; Thiruchelvam. M; Cory-Slechta. DA. (2008a).
CNS effects of developmental Pb exposure are enhanced by combined maternal and offspring stress.
Neurotoxicology 29: 812-827. http://dx.doi.Org/10.1016/i.neuro.2008.03.003
Virgolini. MB; Rossi-George. A; Weston. D; Cory-Slechta. DA. (2008b). Influence of low level maternal
Pb exposure and prenatal stress on offspring stress challenge responsivity. Neurotoxicology 29:
928-939. http://dx.doi.0rg/10.1016/i.neuro.2008.09.010
Wananukul. W: Sura. T; Salaitanawatwong. P. (2006). Polymorphism of delta-aminolevulinic acid
dehydratase and its effect on blood lead levels in Thai workers. Arch Environ Occup Health 61: 67-72.
http://dx.doi.Org/10.3200/AEOH.61.2.67-72
Wang. FT; Hu. H; Schwartz. J: Weuve. J: Spiro. AS. Ill; Sparrow. D: Nie. HL; Silverman. EK; Weiss. ST;
Wright. RO. (2007a). Modifying effects of the HFE polymorphisms on the association between lead
burden and cognitive decline. Environ Health Perspect 115: 1210-1215.
http://dx.doi.org/10.1289/ehp.9855
Wang. GS: Fowler. BA. (2008). Roles of biomarkers in evaluating interactions among mixtures of lead,
cadmium and arsenic [Review]. Toxicol Appl Pharmacol 233: 92-99.
http://dx.doi.0rg/10.1016/i.taap.2008.01.017
Wang. L; Wang. H; Hu. MZ; Cao. J: Chen. DW: Liu. Z. (2009c). Oxidative stress and apoptotic changes
in primary cultures of rat proximal tubular cells exposed to lead. Arch Toxicol 83: 417-427.
http://dx.doi.org/10.1007/s00204-009-0425-z
Wang. YY; Sui. KX; Hong. LI; Ma. HY. (2009e). The effects of lead exposure on placenta! NF-kappaB
expression and the consequences for gestation. Reprod Toxicol 27: 190-195.
http://dx.doi.0rg/10.1016/i.reprotox.2008.12.006
November 2012 6-54 Draft - Do Not Cite or Quote
-------
Weaver. VM; Ellis. LR; Lee. BK; Todd. AC: Shi. W: Ahn. KD; Schwartz. BS. (2008). Associations
between patella lead and blood pressure in lead workers. Am J Ind Med 51: 336-343.
http://dx.doi.org/10.1002/aiim.20573
Weaver. VM: Lee. BK: Todd. AC: Ahn. KD: Shi. W: Jaar. BG: Kelsev. KT: Lustberg. ME: Silbergeld.
EK; Parsons. PJ: Wen. J; Schwartz. BS. (2006). Effect modification by delta-aminolevulinic acid
dehydratase, vitamin D receptor, and nitric oxide synthase gene polymorphisms on associations
between patella lead and renal function in lead workers. Environ Res 102: 61-69.
http://dx.doi.0rg/10.1016/i.envres.2006.01.001
Weaver. VM: Schwartz. BS: Ahn. KD: Stewart. WF; Kelsev. KT; Todd. AC: Wen. J: Simon. DJ; Lustberg.
ME; Parsons. PJ: Silbergeld. EK; Lee. BK. (2003b). Associations of renal function with
polymorphisms in the "delta"-aminolevulinic acid dehydratase, vitamin D receptor, and nitric oxide
synthase genes in Korean lead workers. Environ Health Perspect 111: 1613-1619.
Weaver. VM: Schwartz. BS: Jaar. BG: Ahn. KD: Todd. AC: Lee. SS: Kelsev. KT: Silbergeld. EK:
Lustberg. ME; Parsons. PJ: Wen. J: Lee. BK. (2005b). Associations of uric acid with polymorphisms
in the "delta"-aminolevulinic acid dehydratase, vitamin D receptor, and nitric oxide synthase genes in
Korean lead workers. Environ Health Perspect 113: 1509-1515. http://dx.doi.org/10.1289/ehp.7927
Weiss. AL; Caravanos. J: Blaise. MJ; Jaeger. RJ. (2006). Distribution of lead in urban roadway grit and its
association with elevated steel structures. Chemosphere 65: 1762-1771.
http://dx.doi.0rg/10.1016/i.chemosphere.2006.04.079
Weuve. J: Kelsev. KT; Schwartz. J: Bellinger. D; Wright. RO: Raian. P; Spiro. A. Ill; Sparrow. D: Aro. A;
Hu. H. (2006). Delta-aminolevulinic acid dehydratase polymorphism and the relation between low
level lead exposure and the Mini-Mental Status Examination in older men: The Normative Aging
Study. Occup Environ Med 63: 746-753. http://dx.doi.org/10.1136/oem.2006.027417
Wright. JP: Dietrich. KN; Ris. MD; Hornung. RW: Wessel. SD; Lanphear. BP; Ho. M; Rae. MN. (2008).
Association of prenatal and childhood blood lead concentrations with criminal arrests in early
adulthood. PLoS Med 5: 732-740. http://dx.doi.org/10.1371/iournal.pmed.0050101
Wright. RO: Tsaih. SW: Schwartz. J: Spiro. A; McDonald. K; Weiss. ST; Hu. H. (2003). Lead exposure
biomarkers and mini-mental status exam scores in older men. Epidemiology 14: 713-718.
htrp://dx.doi.org/10.1097/01.EDE.0000081988.85964.db
Wu. J: Edwards. R; He. XE; Liu. Z; Kleinman. M. (2010). Spatial analysis of bioavailable soil lead
concentrations in Los Angeles, California. Environ Res 110: 309-317.
http://dx.doi.0rg/10.1016/i.envres.2010.02.004
Yang. Y; Ma. Y; Ni. L; Zhao. S: Li. L; Zhang. J: Fan. M; Liang. C: Cao. J: Xu. L. (2003). Lead exposure
through gestation-only caused long-term learning/memory deficits in young adult offspring. Exp
Neural 184: 489-495. http://dx.doi.org/10.1016/S0014-4886(03)00272-3
Yapici. G: Can. G: Kiziler. AR; Aydemir. B; Timur. IH; Kavpmaz. A. (2006). Lead and cadmium exposure
in children living around a coal-mining area in Yatagan, Turkey. Toxicol Ind Health 22: 357-362.
Zahran. S: Mielke. HW: Gonzales. CR; Powell. ET; Weiler. S. (2010). New Orleans before and after
hurricanes Katrina/Rita: A quasi-experiment of the association between soil lead and children's blood
lead. Environ Sci Technol 44: 4433-4440. http://dx.doi.org/10.1021/esl00572s
Zawia. NH; Basha. MR. (2005). Environmental risk factors and the developmental basis for Alzheimer's
disease [Review]. RevNeurosci 16: 325-337.
Zhang. A; Park. SK; Wright. RO: Weisskopf. MG: Mukherjee. B; Nie. H; Sparrow. D: Hu. H. (2010a).
HFE H63D polymorphism as a modifier of the effect of cumulative lead exposure on pulse pressure:
The Normative Aging Study. Environ Health Perspect 118: 1261-1266.
http://dx.doi.org/10.1289/ehp. 1002251
Zhao. Y; Wang. L; Shen. HB; Wang. ZX; Wei. QY; Chen. F. (2007). Association between delta-
aminolevulinic acid dehydratase (ALAD) polymorphism and blood lead levels: A meta-regression
analysis. J Toxicol Environ Health 70: 1986-1994. http://dx.doi.org/10.1080/15287390701550946
November 2012 6-55 Draft - Do Not Cite or Quote
-------
7 ECOLOGICAL EFFECTS OF LEAD
1 This chapter synthesizes and evaluates the most policy-relevant science to help form the
2 foundation for the review of the secondary (welfare-based) NAAQS for Pb. The Clean
3 Air Act definition of welfare effects includes, but is not limited to, effects on soils, water,
4 wildlife, vegetation, visibility, weather, and climate, as well as effects on materials,
5 economic values, and personal comfort and well-being. This chapter discusses the effects
6 of Pb on ecosystem components and processes and is organized into five sections. The
7 introduction (Section 7.1) presents the organizing principles of this chapter and several
8 important general ecology concepts. An overview of fate and transport of Pb in
9 ecosystems including measured concentrations of this metal in various environmental
10 media (i.e., soil, water, sediment) is presented in Section 7.2. Section 7.3 reviews the
11 effects of Pb on terrestrial ecosystems; how soil biogeochemistry affects Pb
12 bioavailability, biological effects of Pb exposure and subsequent vulnerability of
13 particular ecosystems. A similar discussion of the effects of Pb on freshwater and
14 saltwater ecosystems is presented in Section 7.4. including water-only exposures and
15 sediment-related effects. The terrestrial, freshwater and saltwater sections each conclude
16 with an integrative synthesis of new evidence for Pb effects and causal determinations,
17 based on the synthesis of new evidence and findings from previous Pb AQCDs. Section
18 7.5 summarizes the causal determinations. Areas not addressed here include literature
19 related to ingestion of Pb shot or pellets and studies that examine human health-related
20 endpoints which are described in other chapters of this document.
7.1 Introduction to Ecological Concepts
21 Metals, including Pb, occur naturally in the environment at measurable concentrations in
22 soils, sediments, and water. Organisms have developed adaptive mechanisms for living
23 with metals, some of which are required micronutrients (but not Pb). However,
24 anthropogenic enrichment can result in concentrations that exceed the capacity of
25 organisms to regulate internal concentrations, causing a toxic response and potentially
26 death. Differences in environmental chemistry may enhance or inhibit uptake of metal
27 from the environment, thus creating a spatial patchwork of environments that are at
28 greater risk than other environments. Similarly, organisms vary in their degree of
29 adaptation to, or tolerance of, the presence of metals. These fundamental principles of
30 how metals interact with organisms and ecosystems are described in detail in EPA's
31 Framework for Metals Risk Assessment (U.S. EPA. 2007c). This section introduces
32 critical concepts for understanding how Pb from atmospheric deposition may affect
November 2012 7-1 Draft - Do Not Cite or Quote
-------
1 organisms, communities, and ecosystems. The sections that follow provide more detail
2 for how aquatic and terrestrial ecosystems respond to Pb and how environmental
3 chemistry interacts with organisms to affect exposure and uptake.
7.1.1 Ecosystem Scale, Function, and Structure
4 For this assessment, an ecosystem is defined as the interactive system formed from all
5 living organisms (biota) and their abiotic (chemical and physical) environment within a
6 given area (IPCC. 2007).The boundaries of what could be called an ecosystem are
7 somewhat arbitrary, depending on the focus of interest or study. Thus, the extent of an
8 ecosystem may range from very small spatial scales to, ultimately, the entire Earth
9 (IPCC. 2007). Ecosystems cover a hierarchy of spatial scales and can comprise the entire
10 globe, biomes at the continental scale, or small, well-circumscribed systems such as a
11 small pond (U.S. EPA. 2008e). A pond may be a small but complex system with multiple
12 trophic levels ranging from phytoplankton to several feeding guilds offish plus fish-
13 eating birds or mammals. A large lake, on the other hand, may be a very simple
14 ecosystem, such as the Great Salt Lake in Utah that covers approximately 1,700 square
15 miles but contains only bacteria, algae, diatoms, and two invertebrate species. All
16 ecosystems, regardless of size or complexity, share the commonality of multiple
17 interactions between biota and abiotic factors, and a reduction in entropy through energy
18 flow from photosynthetic organisms to top predators. This includes both structural
19 (e-g-, soil type and food web trophic levels) and functional (e.g., energy flow,
20 decomposition, nitrification) attributes. Changes are often considered undesirable if
21 important structural or functional components of ecosystems are altered following
22 pollutant exposure (U.S. EPA. 1998V
23 Ecosystems are most often defined by their structure, and are based on the number and
24 type of species present. Structure may refer to a variety of measurements including the
25 species richness, abundance, community composition and biodiversity as well as
26 landscape attributes. Individual organisms of the same species are similar in appearance
27 and genetics, and can interbreed and produce fertile offspring. Interbreeding groups of
28 individual organisms within the same species that occupy some defined geographic space
29 form populations, and populations of different species form communities (Barnthouse et
30 al.. 2008). The community composition may also define an ecosystem type, such as a
31 pine forest or a tall grass prairie. Pollutants can affect the ecosystem structure at any of
32 these levels of biological organization (Suter et al., 2005). Individual plants or animals
33 may exhibit changes in metabolism, enzyme activities, hormone function, or overall
34 growth rates or may suffer gross lesions, tumors, deformities, or other pathologies.
35 Effects on the nervous system of animals may cause behavioral changes that alter
November 2012 7-2 Draft - Do Not Cite or Quote
-------
1 breeding behaviors or predator avoidance. However, only some organism-level endpoints
2 such as growth, survival and reproductive output have been definitively linked to effects
3 at the population level and above. Examples of organism-level endpoints with direct links
4 to population level effects include mass mortality, gross anomalies, survival, fecundity
5 and growth (Suteretal.. 2004). Population level effects of pollutants include changes
6 over time in abundance or density (number of individuals in a defined area), age or sex
7 structure, and production or sustainable rates of harvest (Barnthouse et al., 2008).
8 Community level attributes affected by pollutants include species richness and abundance
9 (also known as biodiversity), dominance of one species over another, or size (area) of the
10 community. Pollutants may affect communities in ways that are not observable in
11 organisms or populations (Bartell 2007). including: (1) effects resulting from interactions
12 between species, such as altering predation rates or competitive advantage; (2) indirect
13 effects, such as reducing or removing one species from the assemblage and allowing
14 another to emerge (Petraitis and Latham. 1999); and (3) alterations in trophic structure.
15 Alternatively, ecosystems may be defined on a functional basis. "Function" refers to the
16 suite of processes and interactions among the ecosystem components and their
17 environment that involve nutrient and energy flow as well as other attributes including
18 water dynamics and the flux of trace gases such as rates of photosynthesis,
19 decomposition, nitrification, or carbon cycling. Pollutants may affect abiotic conditions
20 (e.g., soil chemistry), which indirectly influences biotic structure and function (Bartell
21 2007). Feedback loops or networks influence the stability of the system, and can be
22 mathematically described through simplistic or complex process, or energy flow, models
23 (Bartell. 2007). For example, the Comprehensive Aquatic Systems Model (CASM) is a
24 bioenergetics-based multicompartment model that describes the daily production of
25 biomass (carbon) by populations of aquatic plants and animals over an annual cycle
26 (DeAngelis et al.. 1989). CASM, originally designed to examine theoretical relationships
27 between food web structure, nutrient cycling, and ecosystem stability, has since been
28 adapted for risk assessments and has been applied to numerous lakes with a variety of
29 pollutants (Bartell 2007). Likewise, other theoretical ecosystem models are being
30 modified for use in assessing ecological risks from pollutant exposures (Bartell. 2007).
31 Some ecosystems, and some aspects of particular ecosystems, are less vulnerable to long-
32 term consequences of pollutant exposure. Other ecosystems may be profoundly altered if
33 a single attribute is affected. Thus, spatial and temporal definitions of ecosystem structure
34 and function become an essential factor in defining impacted ecosystem services and
35 critical loads of particular pollutants, either as single pollutants or in combination with
36 other stressors. Both ecosystem services (Section 7.1.2) and critical loads (Section 7.1.3)
37 serve as benchmarks or measures of the impacts of pollutants on ecosystems.
November 2012 7-3 Draft - Do Not Cite or Quote
-------
7.1.2 Ecosystem Services
1 Ecosystem structure and function may be translated into ecosystem services (Daily.
2 1997). Ecosystem services are the benefits people obtain from ecosystems (UNEP, 2003).
3 Ecosystem services are defined as the varied and numerous ways that ecosystems are
4 important to human welfare and how they provide many goods and services that are of
5 vital importance for the functioning of the biosphere. This concept has gained recent
6 interest and support because it recognizes that ecosystems are valuable to humans, and
7 are important in ways that are not generally appreciated (Daily. 1997). Ecosystem
8 services also provide a context for assessing the collective effects of human actions on a
9 broad range of the goods and services upon which humans rely.
10 In general, both ecosystem structure and function play essential roles in providing goods
11 and services. Ecosystem processes provide diverse benefits including absorption and
12 breakdown of pollutants, cycling of nutrients, binding of soil, degradation of organic
13 waste, maintenance of a balance of gases in the air, regulation of radiation balance and
14 climate, and fixation of solar energy fWRI. 2000: Daily. 1997: Westman. 1977). These
15 ecological benefits, in turn, provide economic benefits and values to society (Costanza et
16 al.. 1997: Pimentel etal.. 1997). Goods such as food crops, timber, livestock, fish and
17 clean drinking water have market value. The values of ecosystem services such as flood
18 control, wildlife habitat, cycling of nutrients and removal of air pollutants are more
19 difficult to measure (Goulder and Kennedy. 1997).
20 Particular concern has developed within the past decade regarding the consequences of
21 decreasing biological diversity (Tilman. 2000: Ayensuetal.. 1999: Wall. 1999: Chapin et
22 al.. 1998: Hooper and Vitousek. 1997). Human activities that decrease biodiversity also
23 alter the complexity and stability of ecosystems and change ecological processes. In
24 response, ecosystem structure and function can be affected (Daily and Ehrlich. 1999:
25 Wall. 1999: Chapin etal. 1998: Levin. 1998: Peterson et al.. 1998: Tilman. 1996: Tilman
26 and Downing. 1994: Pimm. 1984). Biodiversity is an important consideration at all levels
27 of biological organization, including species, communities, populations, and ecosystems.
28 Human-induced changes in biotic diversity and alterations in the structure and
29 functioning of ecosystems are two of the most dramatic ecological trends of the past
30 century (U.S. EPA. 2004: Vitousek et al.. 1997).
November 2012 7-4 Draft - Do Not Cite or Quote
-------
1 Hassan et al. (2005) identified four broad categories of ecosystem services:
2 • Supporting services are necessary for the production of all other ecosystem
3 services. Some examples include biomass production, production of
4 atmospheric O2, soil formation and retention, nutrient cycling, water cycling and
5 provisioning of habitat. Biodiversity is a supporting service in that it is
6 increasingly recognized to sustain many of the goods and services that humans
7 enjoy from ecosystems. These supporting services provide a basis for an
8 additional three higher-level categories of services.
9 • Provisioning services such as products (Gitav et al.. 2001) i.e., food (including
10 game meat, roots, seeds, nuts, and other fruit, spices, fodder), water, fiber
11 (including wood, textiles) and medicinal and cosmetic products.
12 • Regulating services that are of paramount importance for human society such as
13 (1) carbon sequestration, (2) climate and water regulation, (3) protection from
14 natural hazards such as floods, avalanches, or rock-fall, (4) water and air
15 purification, and (5) disease and pest regulation.
16 • Cultural services that satisfy human spiritual and aesthetic appreciation of
17 ecosystems and their components.
November 2012 7-5 Draft - Do Not Cite or Quote
-------
7.1.3 Critical Loads as an Organizing Principle for Ecological Effects of
Atmospheric Deposition
1 A critical load is defined as, "a quantitative estimate of an exposure to one or more
2 pollutants below which significant harmful effects on specified sensitive elements of the
3 environment do not occur according to present knowledge" (Nilsson and Grennfelt.
4 1988). Critical loads are a powerful organizing principle for information that links
5 atmospheric deposition with ecological impairment. They allow for heterogeneity in
6 ecosystem sensitivity and exposure which often results in critical load values that vary by
7 ecosystem (e.g., aquatic-water; aquatic-sediment; terrestrial), and differ by endpoint of
8 concern. It is important to consider that critical loads are often calculated assuming
9 steady state conditions (i.e., how much input is required to balance the rate of output),
10 and there may be time required to reach the critical load (i.e., the lag time between onset
11 of exposure and induction of measurable effects). The following types of information are
12 required to calculate a critical load, each of which is discussed in more detail in the
13 subsequent sections of this chapter:
14 • Ecosystem at risk;
15 • Receptors of concern (plants, animals, etc.);
16 • Endpoints of concern (organism, population or community responses, changes
17 in ecosystem services or functions);
18 • Dose (concentration) - response relationships and threshold levels of effects;
19 • Bioavailability and bioaccumulation rates;
20 • Naturally occurring (background) Pb (or other metal) concentrations; and
21 • Biogeochemical modifiers of exposure.
22 There is no single "definitive" critical load for a pollutant, partly because critical load
23 estimates reflect the current state-of-knowledge and policy priorities, and also because of
24 local or regional differences among ecosystems (U.S. EPA, 2008e). Changes in scientific
25 understanding may include, for example, expanded information about dose-response
26 relationships, better understanding of bioavailability factors, and improved quantitative
27 models for effects predictions. Changes in policy may include new mandates for resource
28 protection, inclusion of perceived new threats that may exacerbate the effects of the
29 pollutant of concern (e.g., climate change), and a better understanding of the value of
30 ecosystem services.
November 2012 7-6 Draft - Do Not Cite or Quote
-------
7.2 Fate and Transport of Pb in Ecosystems
i
2
3
4
5
6
7
8
9
10
11
12
13
Fate and transport of Pb in ecosystems are difficult to assess because Pb detected in the
environment could have multiple sources and passes through various environmental
media within a watershed. These issues are described in detail in Section 3.3. Pb can be
emitted to air, soil, or water and then cycle through any or all of these media. In addition
to primary emission of particle-bound or gaseous Pb to the atmosphere, Pb can be
resuspended to the air from soil or dust (Section 7.2.2). Additionally, Pb-bearing PM can
be deposited from the air to soil or water through wet and dry deposition. The
complicated nature of Pb fate and transport in ecosystems is illustrated in Figure 7-1 in
which the Venn diagram depicts how Pb can cycle through multiple environmental media
that encompass both terrestrial and aquatic systems (see also Figure 3-9). The
"air/soil/water" arrows illustrate Pb exposures to plants and animals. Many of the studies
presented in the subsequent material focus on observations of Pb exposure via one
medium: air, soil, sediment, or water.
Newly Emitted Pb
Historically Emitted Pb
NATURAL WATERS
AND SEDIMENTS
OUTDOOR SOIL
AND DUST
Non-air Pb
eleases
AIR
SOIL
SEDIMENT
WATER
AIR
SOIL
SEDIMENT
WATER
PLANT
EXPOSURE
ANIMAL
EXPOSURE
Figure 7-1
Fate of atmospheric Pb in ecosystems.
November 2012
7-7
Draft - Do Not Cite or Quote
-------
7.2.1 Fate and Transport
1 This section provides a brief overview of the fate and transport of Pb in ecosystems. Fate
2 of Pb is determined by the chemical and physical properties of the medium in wet
3 deposition, bodies of water, or soil (e.g., pH, salinity, oxidation status, flow rate and the
4 suspended sediment load and its constituents). Desorption, dissolution, precipitation,
5 sorption and complexation processes can all occur concurrently and continuously, leading
6 to transformations and redistribution of Pb within a watershed. The pH of water is of
7 primary importance in determining the likely chemical fate of Pb in terms of solubility,
8 precipitation, or organic complexation. For more detailed information about the fate and
9 transport of Pb, please see Section 3.3.
10 Soluble Pb in air is mostly removed by wet deposition, and most of the insoluble Pb is
11 removed by dry deposition. As a result, dry deposition is the major removal mechanism
12 for Pb in coarse PM (which is mainly insoluble) and wet deposition is the most important
13 removal mechanism for fine PM and Pb halides (which were more soluble)
14 (Section 3.3.1). Recent research provides considerable evidence that appreciable amounts
15 of Pb can accumulate on coarse PM during transport, and that the physical and chemical
16 characteristics of Pb can be altered by this process due to accompanying transformations
17 (Section 3.3.1.1). Atmospheric removal of metals by wet or dry deposition is largely
18 controlled by solubility of Pb in rain water. The relative importance of wet and dry
19 deposition is highly variable with respect to location and season, probably reflecting both
20 variations in Pb speciation and variations in external factors such as pH and rain water
21 composition (Section 3.3.1.2).
22 Pb deposited to terrestrial ecosystems may remain in soils or eventually be transported in
23 runoff to streams, lakes or rivers in the watershed. Pb has a relatively long retention time
24 in the organic soil horizon, although its movement through the soil column also suggests
25 potential for contamination of groundwater (Section 3.3.3). Pb deposition to soils has
26 decreased since the phase-out of leaded on-road gasoline (Section 3.3.3.1). Recent studies
27 of metal concentrations in leaf litter and organic roadside debris suggest that the litter can
28 act as a temporary sink for metals from the soil around and below leaves on the ground
29 (Section 3.3.3.2). Leaching has been consistently observed to be a slower process for Pb
30 than for other contaminants because Pb is only weakly soluble in pore water, but
31 anthropogenic Pb is more available for leaching than naturally occurring Pb in soil
32 (Section 3.3.3.3). Overall, recent research confirms the generally low mobility of Pb in
33 soil. This limited mobility is strongly dependent on colloid amount and composition, as
34 well as pH, and may be greater in some contaminated soils. Low mobility allows soils to
35 act as a sink for atmospheric Pb potentially for decades or longer. Hence, atmospheric Pb
36 concentrations that peaked several decades ago may still be present in soil in the absence
37 of remediation.
November 2012 7-8 Draft - Do Not Cite or Quote
-------
1 Sources of Pb to surface waters include direct atmospheric deposition and indirect
2 deposition via runoff and industrial discharge (Section 3.3.2). Because dispersal in
3 waterways is a relatively rapid process, concentrations in surface waters are highest near
4 sources of pollution before substantial Pb removal by flushing, evaporation, and
5 sedimentation occurs. Transport in surface water is largely controlled by exchange with
6 sediments, and the cycling of Pb between water and sediments is governed by chemical,
7 biological, and mechanical processes that are affected by many factors, including salinity,
8 organic complexation, oxidation-reduction potential, and pH. Metals in waterways are
9 transported primarily as soluble chelates and ions, or adsorbed on colloidal surfaces,
10 including secondary clay minerals, Fe and Mn oxides or hydroxides, and organic matter,
11 and adsorption on organic or inorganic colloids is particularly important for Pb. The
12 extent of sorption strongly depends on particle size as smaller particles have larger
13 collective surface areas. Pb is relatively stable in sediments, with long residence times
14 and limited mobility (Section 3.3.2.1). As described in previous sections, Pb enters and is
15 distributed in bodies of water largely in PM form. In rivers, particle-bound metals can
16 often account for > 75% of the total load (Section 3.3.2.2). The flux of Pb in aquatic
17 ecosystems is therefore influenced by the dynamic physical and chemical interactions
18 within a watershed.
19 Particles associated with runoff are mostly PM, with a relatively small dissolved fraction,
20 and dissolution of carbonate and related compounds are important contributors to Pb
21 pollution in runoff waters. Pb release into runoff is dependent on storm intensity and
22 length of dry periods between rain events. A "first flush effect" occurs with highest
23 runoff concentrations observed at the beginning of a rain event. Most recent studies have
24 concluded that, during storm events, Pb is transported together with large PM. Some
25 studies, however, found that Pb was concentrated in the fine PM fraction and,
26 occasionally, Pb was found predominantly in the dissolved fraction. Since the
27 2006 Pb AQCD, snowmelt and rain-on-snow events are better understood, and it has
28 been observed that greater runoff occurs from snowmelt and in rain on snow events than
29 when snow is not present, and that metals, including Pb, are often associated with coarse
30 PM under these circumstances. Runoff in rural areas is strongly controlled by soil type
31 and the presence of vegetation, with less runoff and greater retention in mineral soils or
32 when grass is present, and more runoff for soils high in organic matter (OM).
33 Sediments can be either a source or a sink for metals in the aquatic environment
34 (Section 3.3.2). Release can be via re-suspension of the sediment bed via wind, wave, and
35 tidal action or by dissolution from sediment to the water column. Sediment resuspension
36 from marine environments is important, with disturbance of bed sediments by tidal action
37 in estuarine areas resulting in a general greater capacity for re-suspension of PM. Recent
38 research on Pb flux from sediments in natural waters has demonstrated that resuspended
November 2012 7-9 Draft - Do Not Cite or Quote
-------
1 Pb is largely associated with OM or Fe and Mn particles, but that anoxic or depleted
2 oxygen environments in sediments play an important role in Pb cycling. This newer
3 research indicated that resuspension and release from sediments largely occurs during
4 discrete events related to storms. It has also confirmed that resuspension is an important
5 process that strongly influences the lifetime of Pb in bodies of water. Finally, there have
6 been important advances in understanding and modeling of Pb partitioning in complex
7 aquatic environments.
7.2.2 Ecosystem Exposure, Lag Time and Re-entrainment of Historically
Deposited Pb
8 Ecosystem exposure from atmospheric emissions of Pb depends upon the amount of Pb
9 deposited per unit time. Ecosystem response will also depend upon the form in which the
10 Pb is deposited, the areal extent of such deposition, and modifying factors that affect Pb
11 bioavailability in soil, sediments, and water (e.g., pH, organic matter) (Sections 7.3.2.
12 7.4.2 and 7.4.3). However, there is frequently a lag in time between when metals are
13 emitted and when an effect is seen, particularly in terrestrial ecosystems and, to a lesser
14 extent, in aquatic sediments. This is because the buffering capacity of soils and sediments
15 permits Pb to become sequestered into organic matter, reducing its availability for uptake
16 by organisms. The lag time from start of emissions to achieving a critical load can be
17 calculated as the time to reach steady state after Pb was initially added to the system.
18 Excluding erosion processes, the time required to achieve 95% of steady state is about 4
19 half-lives (ti^)1 (Smolders et al.. 2007). Conversely, once emissions cease, the same
20 amount of time is required to reduce metal concentrations to background levels.
1 Time required to reduce the initial concentration by 50% if metal input is zero.
November 2012 7-10 Draft - Do Not Cite or Quote
-------
1 Time to steady state for metals in soils depends upon rates of erosion, uptake by plants,
2 and leaching or drainage from soils. Ignoring erosion, half-life of metals can be predicted
3 (Smolders et al.. 2007) for a soil as:
_ 0.69 xdx 10,000
1/2 y x TF + -§-
PKd
Equation 7-1
4 where:
5 d is the soil depth in meters (m)
6 y is the annual crop yield (tons/ha-yr)
7 TF is the ratio of the metal concentration in plant to that in soil
8 R is the net drainage loss from the soil depth of concern (m3/ha-yr)
9 Pis the bulk density of soil [kg(dry weight)/L]
10 Kd is the ratio of the metal concentration in soil to that in soil pore solution (L/kg)
11 Metals removed by crops (or plants in general) comprise a very small fraction of the total
12 soil metal and can be ignored for the purpose of estimating time to steady state. Thus,
13 equation 7-1 is simplified to:
0.69 xdx 10,000
Equation 7-2
14 and becomes a function of soil depth, the amount of rainfall, soil density, and soil
15 properties that affect Kd. Pb has a relatively long time to steady state compared to other
16 metals, as shown in Table 7-1.
November 2012 7-11 Draft - Do Not Cite or Quote
-------
Table 7-1 Comparison among several metals: Time to achieve 95% of steady
state metal concentration in soil; example in a temperate system.
Metal
Se
Cu
Cd
Pb
Cr
Loading rate (g/ha-yr)
100
100
100
100
100
Kd (L/kg)
0.3
480a
690a
19,000a
16,700a
Time (years)
1.3
1 ,860a
2,670a
73,300a
64,400a
aMean Kd (ratio of total metal concentrations in soils to that in soil pore water); and Time to achieve 95% of steady-state
concentration in soil. (49 Dutch soils) (de Groot et al.. 1998).
Note: Based on a soil depth of 25 cm, a rain infiltration rate of 3,000 m3/ha-yr, and the assumption that background was zero at the
start of loading.
Source: Reprinted with permission of CRC Press, Smolders et al. (2007)
1 In aquatic systems, ti/2 for Pb in the water column depends on the ratio of the magnitudes
2 of the fluxes coming from and going into the sediment, the ratio of the depths of the
3 water column and sediment, and the sediment ti/2. Sediment tU2 is dependent upon the
4 particulate and dissolved fractions and is calculated as for soils (Equation 7-2).
5 Re-entrainment of Pb particles via windblown dust from surface soils or dry sediments
6 may occur. Amount and distance of re-entrained particles and deposition rates are
7 dependent upon wind velocity and frequency; size, density, shape, and roughness of the
8 particle; soil or sediment moisture; and terrain features including openness (including
9 amount of vegetation), aspect relative to wind direction, and surface roughness.
10 Resuspension is defined in terms of a resuspension factor, K, with units of m"1, or a
11 resuspension rate (A), with units of sec"1 (Equation 7-3). The resuspension rate, A, is the
12 fraction of a surface contaminant that is released per time and is defined by:
R
A=c
Equation 7-3
13 where:
14 R is the upward resuspension flux (|ig/m2/sec)
15 C is the soil (or dry sediment) Pb concentration ((ig/m2)
16 Such emissions may have local impacts, but are not likely to have long-range effects, as
17 particles generally remain low to the ground and are not lifted into the atmosphere.
18 Although re-entrainment may alter the particle size distribution in a local area, it
19 generally does not alter the bioavailable fraction, and deposited particles will be subject
November 2012 7-12 Draft - Do Not Cite or Quote
-------
1 to the same biogeochemical forces affecting bioavailability. Therefore, exposure via
2 re-entrainment should be considered additive to exposure from atmospheric particulate
3 deposition in terrestrial and aquatic ecosystems.
7.2.3 Concentrations in Non-Air Media
4 Pb from multiple sources moves through environmental media as described in Section
5 7.2.1 and Figure 7-1 and has led to measurable Pb concentrations in soil, water, sediment
6 and biota in terrestrial and aquatic ecosystems (Table 7-2). The highest concentrations of
7 Pb in the environment are currently found near Pb sources, such as metal smelters and
8 industrial processing. After phase-out of Pb from on-road gasoline, Pb concentrations
9 have decreased considerably in rain, snowpack and surface waters. Declining Pb
10 concentrations in tree foliage, trunk sections, and grasses, as well as surface sediments
11 and soils in some locations, have also been observed (U.S. EPA. 2006b). In contrast, Pb
12 is retained in soils and sediments, where it may provide a historical record of deposition
13 and associated concentrations. In remote lakes, sediment profiles indicate higher Pb
14 concentrations in near surface sediment as compared to pre-industrial era sediment from
15 greater depth, with peak concentrations between 1960 and 1980 (when leaded on-road
16 gasoline was at peak use).
17 Atmospheric deposition has led to measurable Pb concentrations observed in rain,
18 snowpack, soil, surface waters, sediments, agricultural plants, livestock and wildlife.
19 Concentrations of Pb in moss, lichens, peat, and aquatic bivalves have been used to
20 understand spatial and temporal distribution patterns of air Pb concentrations. The
21 amount of Pb in ecosystems is influenced by numerous factors, however, and it is not
22 currently possible to determine the contribution of atmospherically-derived Pb from total
23 Pb. Food, drinking water, and inhalation are major routes of exposure for livestock and
24 terrestrial wildlife. Ingestion and water intake are the major routes of Pb exposure for
25 aquatic organisms. In these exposure pathways, the bioavailable Pb may be from multiple
26 sources. Information on ambient Pb concentrations in non-air media and biota is reported
27 in Section 3.6. and concentrations considered in the interpretation of the ecological
28 evidence are tabulated in Table 7-2.
November 2012 7-13 Draft - Do Not Cite or Quote
-------
Table 7-2 Ambient Pb Concentrations in Non-Air Media and Biota Considered
for Ecological Assessment.
Media
Soil
Freshwater
Sediment
Saltwater Sediment
Fresh Surface
Water
(Dissolved Pb)b
Pb Concentration
National Average:
18.9 mg Pb/kg (dry weight)
Range of state averages:
5-38.6 mg Pb/Kg (dry weight)
Median:
73 mg Pb/kg (dry weight)
Median:
28 mg Pb/kgb (dry weight)
Range:
0.6 to 1 ,050 mg Pb/kga
Median: 0.50 ug Pb/Lb; Max:
30 ug Pb/L, 95th percentile
1.1 ug Pb/L
Years Data
Obtained
1961-1997
1996-2001
1991-2003
Dates not available
1991-2003
References
U.S. EPA (2007d. 2006b.
Mahler etal. (2006)
U.S. EPA (2006b)
Sadiq (1992)
U.S. EPA (2006b)
2003b)
Range: 0.0003-0.075 ug Pb/L
(Set of National Parks in western
U.S.)
2002-2007
Field and Sherrell (2003).
U.S. National Park Service (2011)
Saltwater0
Vegetation
Vertebrate
Range: 0.01 - 27 ug Pb/L
Lichens: 0.3-5 mg Pb/kg (dry
weight) (Set of National Parks in
western U.S.)
Grasses: 31% (percent of soil Pb in
grass)
Fish:
Dates not available
2002-2007
1980s-2000s
1991-2001
Sadiq (1992)
U.S. National Park Service
Vandenhove et al. (2009)
U.S. EPA (2006b)
(2011)
Geometric Mean:
0.59 mg Pb/kg
(dry weight) (whole fish)
0.15 mg Pb/kg
(dry weight) (liver)
Range:
0.08-22.6 mg Pb/kg
(dry weight) (whole fish)
0.01-12.7 mg Pb/kg
(dry weight) (liver)
Fish (from a set of national parks
in western U.S.):
0.0033 (fillet) to 0.97 (liver)
mg Pb/kg (dry weight)
Moosed:
0.008-0.029 mg Pb/kg
(dry weight) (meat)
0.012-0.023 mg Pb/kg
(dry weight) (liver)
2002-2007
U.S. National Park Service (2011)
aNo information available regarding wet or dry weight
"Based on synthesis of NAWQA data reported in 2006 Pb AQCD (U.S. EPA. 2006b)
°Data from a combination of brackish and marine saltwater samples. In general, Pb in seawater is higher in coastal areas and
estuaries since these locations are closer to sources of Pb contamination and loading from terrestrial systems.
dThree moose in one Alaskan park
1
2
o
5
4
The most extensive survey of background soil Pb concentration in the contiguous U.S.
was conducted between 1961 and 1976, and comprised 1,319 non-urban, undisturbed
sampling locations, where 250 cm3 of soil was collected at a depth of 20 cm (Shacklette
and Boerngen. 1984). The lower detection limit was 10 mg Pb/kg, and 14% of the 1,319
November 2012
7-14
Draft - Do Not Cite or Quote
-------
1 samples were below it. The mean Pb concentration was 19.3 mg Pb/kg, the median 15 mg
2 Pb/kg, and the 95th percentile 50 mg/kg. Sixteen locations had Pb concentrations
3 between 100 and 700 mg Pb/kg. These results were in agreement with 3 previous
4 surveys. When creating the Ecological Soil Screening Level (Eco-SSL) guidance
5 document, the U.S. EPA (2007d, 2003b) augmented these data with observations from an
6 additional 13 studies conducted between 1982 and 1997, most of them limited to one
7 state. The resulting data were summarized using state means for each of the fifty states.
8 Those state means ranged between 5 and 38.6 mg Pb/kg, with an overall national mean of
9 18.9 mg Pb/kg. No new data on background concentrations of Pb in U.S. soils have been
10 published since 2005.
11 The 2006 Pb AQCD reported representative median and range of Pb concentrations in
12 surface waters (median 0.50 (ig Pb/L, range 0.04 to 30 (ig Pb/L) and sediments (median
13 28 mg Pb/kg dry weight, range 0.5 to 12,000 mg Pb/kg dry weight) in the U.S. based on a
14 synthesis of National Water Quality Assessment (NAWQA) data (U.S. EPA. 2006c). In
15 an additional study using data collected from 1996-2001 the median Pb concentration in
16 sediment was reported to be 73 mg Pb/kg dry weight (Mahler et al.. 2006). A range of
17 0.01 to 27 (ig Pb/L for saltwater was reported by Sadiq although the values are not
18 specific to the U.S. and include open sea areas as well as estuarine and coastal waters
19 (Sadiq, 1992). In general, Pb in seawater is higher in coastal areas and estuaries since
20 these locations are closer to sources of Pb contamination and loading from terrestrial
21 systems (Sadiq. 1992).
22 Measured concentrations of Pb in soils, sediment and water are not necessarily
23 representative of the amount of Pb that is bioavailable to plants, invertebrates and
24 vertebrates. Both bioaccessibility and bioavailability (Sections 7.3.3. 7.4.3, and 7.4.11) of
25 Pb are dependent upon the physical, chemical, and biological conditions under which an
26 organism is exposed at a particular geographic location. Experimental exposures may be
27 difficult to compare with exposures under natural field conditions in terrestrial and
28 aquatic systems where a variety of abiotic and biotic modifying factors affect Pb toxicity.
7.3 Terrestrial Ecosystem Effects
7.3.1 Introduction to Effects of Pb on Terrestrial Ecosystems
29 Numerous studies of the effects of Pb on components of terrestrial systems were
30 reviewed in the 1977 Pb AQCD, the 1986 Pb AQCD and the 2006 Pb AQCD. The focus
31 of the present document is on studies published since the last AQCD. Many of those
32 studies were conducted near stationary sources of atmospheric Pb such as metal
November 2012 7-15 Draft - Do Not Cite or Quote
-------
1 industries and mines, or using soil collected near those sources. Increasing proximity to
2 the source was often used to generate a gradient of increasing exposure. As may be
3 expected, concentrations found in close proximity to those sources are many times
4 greater than those found at most locations around the country (data on concentrations of
5 Pb in U.S. soils are reviewed in Section 7.2.3 and summarized in Table 7-2). and as
6 indicated in the present document's Preamble, concentrations within one to two orders of
7 magnitude of current conditions were considered. In addition, it is important to note that
8 in all studies where a gradient of multiple concentrations was used, effects increased with
9 increasing concentration. This is an important aspect in determining causality (see
10 Preamble), and therefore justifies inclusion of some studies with very high exposures.
11 Inclusion of those studies also provides potential data for establishing dose-response
12 relationships, and predicting effects at all concentrations, including those found away
13 from stationary sources. Finally, some studies at very high concentrations were used to
14 provide mechanistic information on Pb toxicity, allow for comparison of Pb uptake
15 across taxa, or demonstrate the wide range of sensitivity among closely-related species.
16 Concentrations used in studies where Pb was added to soil experimentally are difficult to
17 relate to concentrations found in natural environments that have been exposed to Pb
18 pollution. As reviewed in the following sections, there is ample evidence that multiple
19 factors, many of them known but not quantified, interact with Pb concentration to
20 produce responses of widely varying magnitude for similar concentrations, or similar
21 responses for varying concentrations of Pb. Thus, experimental concentrations that
22 appear relatively low may be most comparable to relatively high concentrations in natural
23 soils, and vice-versa. The various factors that interact with Pb concentration, and the
24 evidence for those interactions, are discussed in the following sections. However, the
25 same justifications for inclusion apply to added-Pb experiments as they do to studies
26 where proximity to sources is used to vary exposure: gradients of Pb concentrations
27 create gradients of response, and they often provide information on underlying
28 mechanisms of toxicity even if the concentrations cannot be easily compared to natural
29 ones.
30 The literature on terrestrial ecosystem effects of Pb published since the 2006 Pb AQCD,
31 is considered with brief summaries from the 1977 Pb AQCD, the 1986 Pb AQCD and the
32 2006 Pb AQCD, where relevant. Section 7.3 is organized to consider uptake of Pb and
33 effects at the species level, followed by community and ecosystem level effects. Recent
34 evidence for Pb effects on reproduction, growth and survival in terrestrial plants,
35 invertebrates and vertebrates is summarized in Table 7-4. Alterations to reproduction,
36 growth and survival of terrestrial organisms can lead to changes at the community and
37 ecosystem levels of biological organization such as decreased abundance, reduced taxa
38 richness, and shifts in species composition (Section 7.1). Soil biogeochemistry of Pb is
November 2012 7-16 Draft - Do Not Cite or Quote
-------
1 reviewed in Section 7.3.2. Section 7.3.3 considers the bioavailability and uptake of Pb by
2 plants, invertebrates, and vertebrates in terrestrial systems. Biological effects of Pb on
3 terrestrial ecosystem components including plants and lichen, invertebrates, and
4 vertebrates (Section 7.3.4) are followed by data on exposure and response of terrestrial
5 species (Section 7.3.5). Effects of Pb at the ecosystem level of biological organization are
6 discussed in Section 7.3.6. Section 7.3 concludes with a discussion of critical loads in
7 terrestrial systems (Section 7.3.7). soil screening levels (Section 7.3.8), characterization
8 of sensitivity and vulnerability of ecosystem components (Section 7.3.9). and effects on
9 ecosystem services (Section 7.3.10). Concentration of Pb in soil is expressed in mg Pb/kg
10 soil, and concentration in solutions applied to soil or extracted from soil is expressed in
11 mg Pb/L solution.
7.3.2 Soil Biogeochemistry and its Influence on Bioavailability
12 According to data presented in the 2006 Pb AQCD (U.S. EPA. 2006b). the fraction of
13 soil metal that is directly available to plants is the fraction found in soil pore water, even
14 though the concentration of metals in pore water is generally small relative to bulk soil
15 concentration. At any given bulk soil concentration, the amount of Pb dissolved in soil
16 solution is controlled by at least six variables: (1) solubility equilibria; (2) adsorption-
17 desorption relationship of total Pb with inorganic compounds (e.g., oxides of Al, Fe, Si,
18 Mn; clay minerals); (3) adsorption-desorption reactions of dissolved Pb phases on soil
19 organic matter; (4) pH; (5) cation exchange capacity (CEC); and (6) aging. Adsorption-
20 desorption of Pb to soil solid phases is largely controlled by total metal loading.
21 Therefore, areas with high Pb deposition will exhibit a lower fraction of total Pb
22 partitioned to inorganic and organic matter. Decreasing soil pH, CEC, and organic matter
23 have been strongly correlated to increases in the concentration of dissolved Pb species.
24 Aging of metals in soils results in decreased amounts of labile metal as the Pb becomes
25 incorporated into the soil solid phase (McLaughlin et al.. 2010). Data from recent studies
26 have further defined the impact of pH, CEC, organic matter (OM), and aging on Pb
27 mobilization and subsequent bioavailability in soils.
7.3.2.1 pH, CEC and Salinity
28 Models of metal bioavailability calibrated from 500+ soil toxicity tests on plants,
29 invertebrates, and microbial communities indicated that soil pH and CEC are the most
30 important factors governing both metal solubility and toxicity (Smolders et al.. 2009).
31 The variability of derived EC50 values was most closely associated with CEC. Smolders
32 et al. (2007) determined that 12 to 18 months of artificial aging of soils amended with
November 2012 7-17 Draft - Do Not Cite or Quote
-------
1 metal decreased the soluble metal fraction by approximately one order of magnitude.
2 Relatedly, lower soil pH in forest environments relative to adjacent agricultural land
3 resulted in higher solubility, and the mobility of smelter-produced metals was found to be
4 greater in forest than in agricultural lands (Douav et al.. 2009). Further, decreasing the
5 soil pH via simulated acid rain events increased naturally occurring Pb bioavailability in
6 field tests (Hu et al.. 2009b). Miretzky et al. (2007) also showed that the concentration of
7 mobile Pb was increased in acidic soils, and discovered that Pb adsorption to sandy loam
8 clay was a function of weak electrostatic bonds with charged soil surfaces and was
9 influenced by Fe and Mn oxide. Dayton et al. (2006) and Bradham et al. (2006) used path
10 analysis to help identify the main determinants of both organism Pb content and
11 responses from among multiple soil characteristics. In parallel studies with lettuce and
12 earthworms, they amended an array of 21 soils with varying characteristics with the same
13 amount of Pb (2,000 mg /kg as Pb nitrate), and found that in lettuce, the main
14 determinant of both accumulation and biological responses was OC, with contribution
15 from pH and Fe/Al oxides. These later characteristics only influenced accumulation and
16 responses through their own impact on CEC. In earthworms, the main determinant of
17 accumulation was pH, with contribution from CEC, but only through its association with
18 other variables including OC, Fe/Al oxides, and pH. The main determinant of
19 reproductive effects in earthworms was Fe/Al oxides, while pH drove differential
20 mortality between the various soils.
21 Salinity can also alter Pb mobility and bioavailability in soils. Application of CaCl2,
22 MgCl, or NaCl salts to field-collected soils containing 31 to 2,764 mg Pb/kg increased
23 the proportion of mobile metal. As the strength of the salt application was increased from
24 0.006 to 0.3 M, the proportion of released Pb increased from less than 0.5% to over 2%
25 for CaCl2 and from less than 0.5% to over 1% for MgCl (Acostaet al.. 2011). However,
26 the majority of salinity-induced effects occurred in soils containing less than 500 mg
27 Pb/kg, and the proportion of released Pb decreased with increasing total soil Pb
28 concentrations. In addition, the authors noted that Pb release from soils under increasing
29 salinity was reduced at higher carbonate concentrations, indicating that the effect of soil
30 salinity on Pb release is dependent on still other soil factors. A sequential extraction
31 procedure was employed by Ettler et al. (2005) to determine the relative bioavailability of
32 different Pb fractions present in soils collected from a mining and smelting area in the
33 Czech Republic. Five Pb fraction categories were identified: (Fraction A) exchangeable,
34 (Fraction B) acid extractable (bound to carbonates), (Fraction C) reducible (bound to Fe
35 and Mn oxides), (Fraction D) oxidizable (complexed with organic carbon), and (Fraction
36 E) residual (silicates). Tilled agricultural soils were found to have decreased Pb, likely as
37 a result of repeated cultivation, with the majority of Pb represented as the reducible
38 Fraction C. Pb concentration in undisturbed forest soils, however, was largely present as
39 the exchangeable fraction (A), weakly bound to soil OM. However, the validity of
November 2012 7-18 Draft - Do Not Cite or Quote
-------
1 associating sequentially extracted fractions with discrete geochemical components has
2 not been definitively established, and as a consequence, the association between
3 fractionation and bioavailability remains uncertain.
7.3.2.2 Organic Matter
4 Organic matter decreases bioavailability of Pb, but as it is turned over and broken down,
5 pedogenic minerals become more important in Pb sequestration (Schroth et al.. 2008).
6 Shaheen and Tsadilas (2009) noted that soils with higher clay content, organic matter,
7 total calcium carbonate equivalent, and total free sesquioxides also exhibited higher total
8 Pb concentration, indicating that less Pb had been taken up by resident plant species.
9 Huang et al. (2008) examined the re-mobilization potential of Pb in forest soils, and
10 determined that mobilization of total Pb was strongly associated with dissolved organic
11 matter (DOM). Groenenberg et al. (2010) used a non-ideal competitive adsorption
12 Donnan model to explain the variability of organic matter binding affinity and
13 uncertainties associated with metal speciation. They found that natural variations in fulvic
14 acid binding properties were the most important variable in predicting Pb speciation. Guo
15 et al. (2006b) determined that the -COOH and -OH groups associated with soil OM were
16 important factors in Pb sequestration in soil, and Pb sorption was increased as pH was
17 raised from 2 to 8. Because organic content increased the Pb sequestration efficiency of
18 soils, OM content had an inhibitory effect on Pb uptake by woodlouse species Oniscus
19 asellus and Porcellio scaber (Gal et al., 2008). Vermeulen et al. (2009) demonstrated that
20 invertebrate bioaccumulation of Pb from contaminated soils was dependent on pH and
21 OM, but that other unidentified habitat-dependent factors also contributed. The
22 relationship of bioaccumulation and soil concentration was modified by pH and OM, and
23 also by habitat type. Kobler et al. (2010) showed that the migration of atmospherically
24 deposited Pb in soil matrices was strongly influenced by soil type, indicating that certain
25 soil types may retain Pb for longer periods of time than others. In soils characterized by
26 well-drained substrate and limestone bedrock, Pb concentration decreased over time,
27 likely as a result of water drainage and percolation. The authors contrasted this
28 observation with reports of prolonged residence time in humic soils, particularly at the
29 lower depths of the humus layer. They theorized that the most significant Pb migration
30 route was transportation of particulate-bound Pb along with precipitation-related flow
31 through large soil pores.
32 A number of recent laboratory studies have further defined the relationship of soil
33 biogeochemical characteristics and Pb uptake by plants. As noted above, Dayton et al.
34 (2006) found through path analysis that the main determinant of both accumulation and
35 biological responses in lettuce grown on amended soil was OC. As part of a metal
November 2012 7-19 Draft - Do Not Cite or Quote
-------
1 partitioning study, Kalis et al. (2007) determined that not only did metal concentration in
2 the soil solution decrease as pH increased, but pH-mediated metal adsorption at the root
3 surface ofLolium perenne determined root Pb concentration, with concentration in the
4 shoot correlated with root concentration. Interestingly, Kalis et al. (2007) and Lock et al.
5 (2006) also observed that the influx of Pb in the water-soluble fraction had an impact on
6 soil pH. In addition, 1 (iM humic acid decreased root Pb concentration in L. perenne
1 plants grown in 0.1 and 1 (iM Pb solution, likely as a result of Pb complexation and
8 sequestration with the added OM (Kalis et al.. 2006). Ma et al. (2010) also reported that
9 long-term agricultural cultivation can decrease the rate of Pb desorption in soil through a
10 gradual OM-enrichment. Phosphorous soil amendments equivalent to 35 mg P/kg soil
11 were observed to reduce the quantity of DPTA-extractable Pb from an average of 19 and
12 24 mg Pb/kg in unamended soils to 12 to 15 mg Pb/kg in P-amended soils. As a result,
13 maize and soybean seedlings accumulated significantly less Pb: average concentrations in
14 soybean shoot and root ranged from 4.4 to 5.2 mg Pb/kg with P addition (versus 9.21 mg
15 Pb/kg without), while maize shoot concentrations average between 4.8 to 5.3 mg Pb/kg in
16 P-amended soils (as compared with 10.16 mg Pb/kg in controls) (Xie etal., 2011).
7.3.2.3 Aging
17 Smolders et al. (2007) defined aging as the process responsible for decreasing the
18 bioavailability of metals in soils independently of their persistence. Smolders et al. (2009)
19 reviewed the effects of aging of Pb in soils on the toxicity of Pb to plants and soil
20 invertebrates, with aging achieved in most studies primarily by leaching amended soil,
21 but also through natural binding and complexation. In nearly half of the Pb soil studies
22 reviewed, responses that were observed with freshly amended soil could no longer be
23 detected following soil leaching, indicating that aged soils likely contain less bioavailable
24 Pb. The authors concluded that competitive binding between soil ligands and biotic
25 ligands on plant roots or invertebrate guts can be used to model the relationship of
26 observed availability and toxicity of metals in soils. Because this concept is the basis of
27 the Biotic Ligand Model (BLM) (Section 7.3.3). the authors proposed a terrestrial BLM
28 approach to estimate the risk of metals to terrestrial organisms. However, Antunes et al.
29 (2006) noted that there were several key challenges involved in development of a
30 terrestrial BLM applicable to plants, particularly the reliable measurement of free ion
31 activities and ligand concentration in the rhizosphere, the identification of the organisms'
32 ligands associated with toxicity, and the possible need to incorporate kinetic dissolution
33 of metal-ligand complexes as sources of free ion. Further, Pb in aged field soils has been
34 observed to be less available for uptake into terrestrial organisms, likely as a result of
35 increased sequestration within the soil particles (Antunes et al.. 2006). Magrisso et al.
November 2012 7-20 Draft - Do Not Cite or Quote
-------
1 (2009) used a bioluminescent strain of the bacterium Cupriavidus metallidurans to detect
2 and quantify Pb bioavailability in soils collected adjacent to industrial and highway areas
3 in Jerusalem, Israel, and in individual simulated soil components freshly spiked with Pb.
4 The bacterium was genetically engineered to give off the bioluminescent reaction as a
5 dose-dependent response, and was inoculated in soil slurries for three hours prior to
6 response evaluation. Spiked soil components induced the bioluminescent response, and
7 field-collected components did not. However, the comparability of the simulated soils
8 and their Pb concentration with the field-collected samples was not entirely clear. Lock et
9 al. (2006) compared the Pb toxicity to springtails (Folsomia Candida) from both
10 laboratory-spiked soils and field-collected Pb-contaminated soils of similar Pb
11 concentrations. Total Pb concentrations of 3,877 mg Pb/kg dry weight and higher always
12 caused significant effects on F. Candida reproduction in the spiked soils. In field soils,
13 only the soil with the highest Pb concentration of 14,436 mg Pb/kg dry weight
14 significantly affected reproduction. When expressed as soil pore-water concentrations,
15 reproduction was never significantly affected at Pb concentrations of 0.5 mg Pb/L,
16 whereas reproduction was always significantly affected at Pb concentrations of 0.7 mg
17 Pb/L and higher, independent of the soil treatment. Leaching soils prior to use in
18 bioassays had only a slight effect on Pb toxicity to resident springtails, suggesting that
19 among the processes that constitute aging of Pb in field soils, leaching is not particularly
20 important with respect to bioavailability.
21 Red-backed salamanders (Plethodon cinereus) exposed to Pb-amended soils (553 mg
22 Pb/kg, 1,700 mg Pb/kg, 4,700 mg Pb/kg, and 9,167 mg Pb/kg) exhibited lowered appetite
23 and decreased white blood cell counts at the two highest concentrations, as compared to
24 controls (Bazar etal. 2010). However, salamanders tolerated field-collected, aged soils
25 containing Pb concentration of up to 16,967 mg Pb/kg with no significant deleterious
26 effects.
27 In summary, studies published during the past 5 years continue to substantiate the
28 important role that soil geochemistry plays in sequestration or release of Pb. Soil pH and
29 CEC have long been known to be the primary controlling factors of the amount of
30 bioavailable Pb in soils, and a recent review of more than 500 studies corroborates these
31 findings (Smolders et al.. 2009). Fe and Mn oxides are now known to also play an
32 important role in Pb sequestration in soils. Pb binds to OM, although relatively weakly,
33 and as the OM is broken down the Pb may be released into soil solution. Leaching of
34 metal through soil pores may be the primary route for loss of bioavailable soil Pb; OM
35 may reduce leaching and thus appear to be associated with Pb sequestration. Aging of Pb
36 in soils (through incorporation of the metal into the particulate solid-phase of the soil)
37 results in long term binding of the metal, and reduced bioavailability of Pb to plants and
38 soil organisms.
November 2012 7-21 Draft - Do Not Cite or Quote
-------
7.3.3 Bioavailability in Terrestrial Systems
1 Bioavailability was defined in the 2006 Pb AQCD as "the proportion of a toxin that
2 passes a physiological membrane (the plasma membrane in plants or the gut wall in
3 animals) and reaches a target receptor (cytosol or blood)" (U.S. EPA. 2006c). In 2007,
4 EPA took cases of bioactive adsorption into consideration and revised the definition of
5 bioavailability as "the extent to which bioaccessible metals absorb onto, or into, and
6 across biological membranes of organisms, expressed as a fraction of the total amount of
7 metal the organism is proximately exposed to (at the sorption surface) during a given
8 time and under defined conditions" (U.S. EPA. 2007c). The bioavailability of metals
9 varies widely depending on the physical, chemical, and biological conditions under
10 which an organism is exposed (U.S. EPA. 2007c). Characteristics of the toxicant itself
11 that affect bioavailability are: (1) chemical form or species, (2) particle size, (3) lability,
12 and (4) source. The bioavailability of a metal is also dependent upon the fraction of metal
13 that is bioaccessible. As stated in the Framework for Metals Risk Assessment (U.S. EPA.
14 2007c). the bioaccessible fraction of a metal is the portion (fraction or percentage) of
15 environmentally available metal that actually interacts at the organism's contact surface
16 and is potentially available for absorption or adsorption by the organism. The Framework
17 states that "the bioaccessibility, bioavailability, and bioaccumulation properties of
18 inorganic metals in soil, sediments, and aquatic systems are interrelated and abiotic
19 (e.g., organic carbon) and biotic (e.g., uptake and metabolism). Modifying factors
20 determine the amount of an inorganic metal that interacts at biological surfaces (e.g., at
21 the gill, gut, or root tip epithelium) and that binds to and is absorbed across these
22 membranes. A major challenge is to consistently and accurately measure quantitative
23 differences in bioavailability between multiple forms of organic metals in the
24 environment." A conceptual diagram presented in the Framework for Metals Risk
25 Assessment (U.S. EPA. 2007c) summarizes metals bioavailability and bioaccumulation
26 in aquatic, sediment and soil media (Figure 7-2).
November 2012 7-22 Draft - Do Not Cite or Quote
-------
Bioaccessible Fraction (BF) ":
Percent soluble metal ion
concentration relative to total
metal concentration (measured in
solution near biomembrane)
Relative Bioavai lability (RBA) b:
Percent adsorbed or absorbed
compared to reference material
(measure of membrane dynamics)
Absolute Bioavailability (ABA) c:
Percent of metal mass absorbed
internally corrpared to external
exposure (measures systerric
uptake/accumulation)
Bioaccessibility
X
io\vailability
Environmental availability \
Exposure ==
Bioaccumulation of metal
==============-» Efects
lembrane
uptake
\
Physiological
V^rnembrane
Total Metal Concentration
Predation
Foraging
Toxicoloical
accumulation
Detoxification
and Storage
Benign /
accumulation
Internal r
Transport
and
Distribution
aBF is most often measured using in vitro methods (e.g., artificial stomach), but it should be validated by in vivo methods.
bRBA is most often estimated as the relative absorption factor, compared to a reference metal salt (usually calculated on the basis of
dose and often used for human risk, but it can be based on concentrations).
°ABA is more difficult to measure and used less in human risk; it is often used in ecological risk when estimating bioaccumulation or
trophic transfer.
Source: ERG (2004) and U.S. EPA (2007c).
Figure 7-2 Conceptual diagram for evaluating bioavailability processes and
bioaccessibility for metals in soil, sediment, or aquatic systems.
i
2
3
4
5
6
7
8
9
10
The BLM attempts to integrate the principal physical and chemical variables that
influence Pb bioavailability. The model considers the reactions of Pb with biological
surfaces and membranes (the site of action) to predict the bioavailability and uptake of
the metal (Figure 7-3). and integrates the binding affinities of various natural ligands and
the biological uptake rates of organisms to predict both the bioaccessible and bioavailable
fraction of Pb in the environment, and to determine the site-specific toxicity of the
bioavailable fraction. In principle, the BLM can be used for determining toxicity in water,
sediment, and soil media, however, the parameter values that influence BLM are, in
general, characterized to a greater extent in aquatic systems than in terrestrial systems
(Section 7.4.4).
November 2012
7-23
Draft - Do Not Cite or Quote
-------
Organic Matter
Complexation
Inorganic Ligand
Complexation
Source: Reprinted with permission of John Wiley and Sons ; from Di Toro et al. (2001)
Figure 7-3 Schematic diagram of the biotic ligand model.
1 New information on sources of Pb in terrestrial ecosystems, and their influence on
2 subsequent bioavailability, was reviewed in Chapter_3, while new information on the
3 influence of soil biogeochemistry on speciation and chemical lability was presented in
4 Section 7.3.2. This section summarizes recent literature on uptake and subsequent
5 presence of Pb in tissues. The 2006 Pb AQCD (U.S. EPA. 2006b) extensively reviewed
6 the methods available for quantitative determination of the mobility, distribution, uptake,
7 and fluxes of atmospherically delivered Pb in ecosystems, and they are not reviewed in
8 this section. The 2006 Pb AQCD also reported bioaccumulation factors (BAF) and
9 bioconcentration factors (BCF). BAF is defined as the field measurement of metal
10 concentration in tissues, including dietary exposures, divided by metal concentration in
11 environmental media (Smolders et al., 2007). BCF is defined as the same measurement
12 carried out in artificial media in the laboratory that does not include dietary exposure
13 (Smolders et al., 2007). The EPA Framework for Metals Risk Assessment states that the
14 latest scientific data on bioaccumulation do not currently support the use of BCFs and
15 BAFs when applied as generic threshold criteria for the hazard potential of metals (U.S.
16 EPA. 2007c).
November 2012
7-24
Draft - Do Not Cite or Quote
-------
7.3.3.1 Terrestrial Plants
1 At the time of the 1977 Pb AQCD, it was understood that Pb uptake in plants was
2 influenced by plant species and by the available Pb pool in the soils (U.S. EPA. 1977).
3 The role of humic substances in binding Pb was better characterized by the 1986 Pb
4 AQCD where it was stated that most plants cannot survive in soil containing
5 10,000 (ig Pb/g (mg Pb/kg) dry weight if the pH is below 4.5 and the organic content is
6 below 5% (U.S. EPA. 1986b). At the time of the 1986 AQCD , it was thought that Pb can
7 be absorbed across the leaf surface into internal plant tissues, but that the vast majority of
8 uptake is via roots (U.S. EPA. 1986b). The 2006 Pb AQCD (U.S. EPA. 2006b) noted that
9 terrestrial plants accumulate atmospheric Pb primarily via two routes: direct stomatal
10 uptake into foliage, and incorporation of atmospherically deposited Pb from soil into root
11 tissue, followed by variable translocation to other tissues. Foliar Pb may include both
12 incorporated Pb (i.e., from atmospheric gases or particles) and surficial particulate Pb
13 deposition. Although the plant may eventually absorb the surficial component, its main
14 importance is its likely contribution to the exposure of plant consumers. This section will
15 first review recent studies on uptake of Pb by plants through foliar and soil routes, and
16 their relative contribution, followed by the consideration of translocation of Pb from roots
17 to shoots, including a discussion of variability in translocation among species. Data on
18 ambient Pb levels associated with vegetation are summarized in Section 3.6.6.
Leaf and Root Uptake
19 Although Pb is not an essential metal, it is taken up from soils through the symplastic
20 route, the same active ion transport mechanism used by plants to take up water and
21 nutrients and move them across root cell membranes (U.S. EPA. 2006c). As with all
22 nutrients, only the proportion of a metal present in soil pore water is directly available for
23 uptake by plants. In addition, soil-to-plant transfer factors in soils enriched with Pb have
24 been found to better correlate with bioavailable Pb soil concentration, defined as DTPA-
25 extractable Pb, than with total Pb concentration (U.S. EPA. 2006c). Since the publication
26 of the 2006 Pb AQCD, suggestive evidence has become available that a substantial
27 proportion of Pb accumulated in shoots of some species of trees originates in direct leaf
28 uptake of atmospheric Pb. Evidence for such direct uptake is weaker in herbaceous
29 plants, and all data came from near stationary sources.
30 Field studies carried out in the vicinity of Pb smelters have determined the relative
31 importance of direct foliar uptake and root uptake of atmospheric Pb deposited in soils.
32 Hu and Ding (2009) analyzed ratios of Pb isotopes in the shoots of commonly grown
33 vegetables and in soil at three distances from a smelter (0.1, 0.2, 5.0 km). Pb isotope
34 ratios in plants and soil were different at two of those locations, leading the authors to the
November 2012 7-25 Draft - Do Not Cite or Quote
-------
1 conclusion that airborne Pb was being assimilated via direct leaf uptake. Soil Pb
2 concentration in the rhizosphere at the three sites ranged between 287 and 379 mg Pb/kg
3 (Site I), 155 and 159 mg Pb/kg (Site II), and 58 and 79 mg Pb/kg (Site III, selected as the
4 control site). The median shoot and root Pb concentrations at each site were 36 and
5 47 mg Pb/kg, 176 and 97 mg Pb/kg, and 1.3 and 7 mg Pb/kg, respectively, resulting in
6 shootroot Pb ratios exceeding 1.0 in Site I (for Malabar spinach [Basella alba],
1 ratio = 1.6, and amaranth \Amaranthus spinosus], ratio = 1.1), and in Site II (for the
8 weeds Taraxacum mongolicum, ratio = 1.9, and Rostellariaprocumbens, ratio = 1.7).
9 However, the two species studied at Site II were not studied at Site I or Site III. In the
10 control site (Site III), no plant was found with a Pb shootroot ratio greater than 1.0. Hu
11 and Ding (2009) concluded that metal accumulation was greater in shoot than in root
12 tissue, which suggested both high atmospheric Pb concentration and direct stomatal
13 uptake into the shoot tissue.
14 Cui et al. (2007) studied seven weed species growing in the vicinity of an old smelter
15 (average soil Pb concentration of 4,020 mg Pb/kg) in Liaoning, China, to measure Pb
16 accumulation rates in roots and shoots. Cutleaf groundcherry (Physalis angulatd)
17 accumulated the most Pb, with root and shoot concentration of 527 and 331 mg Pb/kg,
18 respectively, and velvetleaf (Abutilon theophrasti) was the poorest absorber of Pb (root
19 and shoot concentration of 39 and 61 mg Pb/kg, respectively). In all cases, weed species
20 near the smelter accumulated more Pb than plants from non-polluted environments (5 mg
21 Pb/kg), indicating that aerially deposited Pb produced by smelting is bioavailable to
22 plants. However, the ratio of rootshoot Pb concentration varied by species, and the
23 authors presented no data to differentiate Pb taken up from soil from Pb incorporated via
24 foliar uptake. Angelova et al. (2010) examined Pb uptake by rapeseed plants (Brassica
25 napus) grown in heavy metal contaminated soils 0.5 km and 15 km from the Non-Ferrous
26 Metal Works, in Bulgaria. Average surface soil Pb concentration decreased with distance
27 from the plant (200.3 and 24.6 mg Pb/kg, respectively), as did average DTPA-extractable
28 Pb (69.7 and 4.9 mg Pb/kg, respectively). Pb content in stems and leaves in rapeseed
29 grown at 0.5 km from the plant averaged 1.73 and 8.69 mg Pb/kg ; average stem and leaf
30 Pb concentrations in rapeseed grown at the more distant location were reported as 0.72
31 and 1.42 mg Pb/kg, respectively (Angelova et al.. 2010).
32 Pb plant BAFs for plants grown in 70 actively cropped fields in California averaged
33 0.052 for vegetable crops and 0.084 for grains; the highest reported Pb BAF (0.577) was
34 found in onions. Authors compared the BAFs based on total Pb and Pb in solution and
35 determined that both were accurate predictors of plant uptake (Chen et al.. 2009b).
36 Likewise, Zhang et al. (20lib) compiled Pb uptake data for several crop species in
37 China, and reported an average BAF for grains (rice) of 0.009 (0.0009-0.03) and
38 0.41(0.0007-0.17) for leafy vegetables, such as spinach, Chinese cabbage and celery
November 2012 7-26 Draft - Do Not Cite or Quote
-------
1 (Zhang et al.. 20 lib). Chrastny et al. (2010) characterized the Pb contamination of an
2 agricultural soil in the vicinity of a shooting range. Pb was predominantly in the form of
3 PbO and PbCO3, and Pb was taken up by plants through both atmospheric deposition
4 onto the plant and by root uptake.
5 The Pb content of ripe date palm (Phoenix dactyliferd) fruit collected in Riyadh, Saudi
6 Arabia was determined to be indicative of areas of heavy industrialization and
7 urbanization; Pb concentrations in fruit flesh ranged from 0.34 to 8.87 mg Pb/kg dry
8 weight, with the highest Pb date concentrations detected near freeways and industrial
9 areas (Aldjain et al.. 2011). Likewise, Pb concentrations in rosemary (Rosmarinus
10 officinalis) flowers, stems, and leaves were significantly higher in the urban areas of Al-
11 Mafraq and Irbid, Jordan than in the smaller town of Ma'an, Jordan (53.6 to 86.5 mg
12 Pb/kg versus 16.2 to 16.7 mg Pb/kg). Authors noted a significant difference between Pb
13 concentrations in washed and unwashed rosemary samples, indicating that aerial
14 deposition and surface dust is likely a significant source of plant-associated Pb (El-Rjoob
15 et al.. 2008).
16 Bilberry (Vaccinium myrtillus), accumulated the highest amount of Pb out of four total
17 herbaceous species growing in Slovakian spruce ecosystems with variable soil Pb
18 concentrations, giving BAFs of 0.09 to 0.44, depending on location (Kuklova et al..
19 2010). Because of their long life spans, trees can provide essential information regarding
20 the sources of bioavailable Pb. A Scots pine forest in northern Sweden was found to
21 incorporate atmospherically derived Pb pollution directly from ambient air, accumulating
22 this Pb in bark, needles, and shoots (Klaminder et al.. 2005). Nearly 50% of total tree
23 uptake was estimated to be from direct adsorption from the atmosphere, as determined
24 using isotopic ratios and a binary mixing model. Further, Aznar et al. (2009a) found that
25 the Pb content of black spruce (Picea mariand) needles collected along a metal
26 contamination gradient emanating from a Canadian smelter in Murdochville, Quebec,
27 showed a significant decrease in Pb concentration with increasing distance from the
28 smelter. Interestingly, older needles were determined to accumulate larger quantities of
29 Pb than younger ones. Foliar damage and growth reduction were also observed in the
30 trees (Aznar et al.. 2009a). They were significantly correlated with Pb concentration in
31 the litter layer. In addition, there was no correlation between diminished tree growth and
32 Pb concentration in the deeper mineral soil layers, strongly suggesting that only current
33 atmospheric Pb was affecting trees (Aznar et al.. 2009b). Similarly, Kuang et al. (2007)
34 noted that the Pb concentration in the inner bark ofPinus massoniana trees growing
35 adjacent to a Pb-Zn smelter in the Guangdong province of China was much higher
36 (1.87 mg Pb/kg dry weight) than in reference-area trees. Because concentration in the
37 inner bark was strongly correlated with concentration in the outer bark, they concluded
38 that the origin of the Pb was atmospheric.
November 2012 7-27 Draft - Do Not Cite or Quote
-------
1 Dendrochronology (tree ring analysis) has become an increasingly important tool for
2 measuring the response of trees to Pb exposure (Watmough. 1999). Tree ring studies
3 reviewed in the 1977 Pb AQCD showed that trees could be used as indicators of
4 increasing environmental Pb concentrations with time. Additional studies in the 1986 Pb
5 AQCD indicated that Pb could be translocated from roots to the upper portions of the
6 plant and that the amounts translocated are in proportion to concentrations of Pb in soil
7 (U.S. EPA. 1986b). The advent of laser ablation inductively coupled plasma mass
8 spectrometry has made measurement of Pb concentration in individual tree rings possible
9 (Witte et al., 2004; Watmough. 1999). This allows for close analysis of the timing of Pb
10 uptake relative to smelter activity and/or changes in soil chemistry. For example, Aznar
11 et al. (2008a) measured Pb concentration in black spruce tree rings to determine the
12 extent and timing of atmospheric deposition near the Murdochville smelter. Variability in
13 tree-ring Pb content seemed to indicate that trees accumulated and sequestered
14 atmospheric Pb in close correlation with the rates of smelter emission, but that
15 sequestration lagged about 15 years behind exposure. However, the ability to determine
16 time of uptake from the location in growth rings is weakened in species that transfer Pb
17 readily from outer bark to inner bark. Cutter and Guyette (1993) identified species with
18 minimal radial translocation from among a large number of tree species, and
19 recommended the following temperate zone North American species as suitable for metal
20 dendrochronology studies: white oak (Quercus alba), post oak (Q. stellatd), eastern red
21 cedar (Juniperus virginiana), old-growth Douglas fir (Pseudotsuga menziesii), and big
22 sagebrush (Artemisia tridentata). In addition, species such as bristlecone pine (Pinus
23 aristata), old-growth redwood (Sequoia sempervirens), and giant sequoia (S. giganted)
24 were deemed suitable for local purposes. Patrick and Farmer (2006) determined that
25 European sycamore (Acer pseudoplatanus) are not suitable for this type of
26 dendrochronological analysis because of the formation of multiple annual rings.
27 Pb in sapwood and heartwood is more likely a result of soil uptake than of direct
28 atmospheric exposure (Guvette et al.. 1991). Differentiation of geogenic soil Pb in tree
29 tissue from Pb that originated in the atmosphere requires measurement of stable Pb
30 isotope ratios (Patrick. 2006). Tree bark samples collected from several areas of the
31 Czech Republic were subjected to stable Pb isotope analysis to determine the source and
32 uptake of atmospheric Pb (Conkova and Kubiznakova. 2008). Results indicated that
33 beech bark is a more efficient accumulator of atmospheric Pb than spruce bark. A
34 decrease in the 206Pb/207Pb ratio was measured in bark and attributed to increased usage of
35 leaded gasoline between 1955 and 1990; an increased 206Pb/207Pb ratio was ascribed to
36 coal combustion (Conkova and Kubiznakova. 2008). Similarly, Savard et al. (2006)
37 compared isotope ratios of 206Pb/207Pb and 208Pb/206Pb in tree rings from spruce trees
38 sampled at a control site near Hudson Bay, with those sampled near the Home smelter
39 active since 1928, in Rouyn-Noranda, Canada. The concentration of total Pb showed a
November 2012 7-28 Draft - Do Not Cite or Quote
-------
1 major increase in 1944 and a corresponding decrease of the 206Pb/207Pb ratios, suggesting
2 that the smelter was responsible for the increased Pb uptake (Savard et al.. 2006). The
3 authors suggested that the apparent delay of 14 years may have been attributable to the
4 residence time of metals in airborne particles the buffering effect of the soils and, to a
5 lesser extent, mobility of heavy metals in tree stems. Furthermore, through the use of the
6 two different isotope ratios, Savard et al. (2006) were able to differentiate three types of
7 Pb in tree rings: natural (derived from the mineral soil horizons), industrial (from coal
8 burning urban pollution), and mining (typical of the volcanogenic massive sulfide ore
9 deposits treated at the Home smelter).
10 Devall et al. (2006) measured Pb uptake by bald-cypress trees (Taxodium distichum)
11 growing in a swamp near a petroleum refinery and along a bank containing
12 Pb-contaminated dredge spoils. They measured Pb in tree cores and showed
13 greater uptake of Pb by trees in the swamp than by trees growing on the dredge spoil
14 bank, attributing the difference to exposure source (refinery versus dredge spoils) and
15 differences in soil chemistry between the swamp and the dredge spoil bank (Devall et al..
16 2006). Similarly, Gebologlu et al. (2005) found no correlation between proximity to
17 roadway and accumulated Pb in tomato and bean plants at sites adjacent to two state
18 roads in Turkey (average Pb concentration 5.4 and 6.0 mg Pb/kg), indicating that uptake
19 may be influenced by multiple factors, including wind direction, geography, and soil
20 chemistry. Average Pb levels in leaves were 0.6 and 0.5 mg Pb/kg for tomato and bean
21 plants, respectively, while fruit concentration averaged 0.4 mg Pb/kg for both species.
22 Conversely, if foliar contamination is due primarily to dust deposition, distance from a
23 source such as a road may be easily correlated with Pb concentration on the plants. For
24 example, Ai-Khlaifat and Al-Khashman (2007) collected unwashed date palm (Phoenix
25 dactyliferd) leaves at 3-meter trunk height from trees in Jordan to assess the extent of Pb
26 contamination from the city of Aqaba. Whereas relatively low levels of Pb were detected
27 in leaves collected at background sites (41 mg Pb/kg), leaves collected adjacent to
28 highway sites exhibited the highest levels of Pb (177 mg Pb/kg). The authors determined
29 that Pb levels in date palm leaves correlated with industrial and human activities
30 (e-g-, traffic density) (Ai-Khlaifat and Al-Khashman. 2007). Likewise, Pb concentrations
31 were significantly enriched in tree bark samples and road dust collected in highly
32 urbanized areas of Buenos Aires, Argentina (approximate average enrichment factors of
33 30 and 15 versus reference samples) (Fujiwara et al.. 2011). However, decreases in tissue
34 Pb concentration with increasing distance from stationary sources can also follow from
35 decreasing Pb in soil. Bindler et al. (2008) used Pb isotopes to assess the relative
36 importance of pollutant Pb versus natural Pb for plant uptake and cycling in Swedish
37 forested soils. The Pb isotopic composition of needles/leaves and stemwood of different
38 tree species and ground-cover plants indicated that the majority of Pb present in these
39 plant components was derived from the atmosphere, either through aerial interception or
November 2012 7-29 Draft - Do Not Cite or Quote
-------
1 actual uptake through the roots. For the ground-cover plants and the needles/leaves, the
2 206Pb/207Pb isotopic ratios (1.12 to 1.20) showed that the maj ority of Pb was of
3 anthropogenic origin. Stemwood and roots have higher 206Pb/207Pb ratio values (1.12 to
4 1.30) which showed the incorporation of some natural Pb as well as anthropogenic Pb.
5 For pine trees, the isotopic ratio decreased between the roots and the apical stemwood
6 suggesting that much of the uptake of Pb by trees is via aerial exposure. Overall, it was
7 estimated that 60-80% of the Pb in boreal forest vegetation originated from pollution; the
8 Pb concentrations were, however, quite low - not higher than 1 mg Pb/kg plant material,
9 and usually in the range of 0.01-0.1 mg Pb/kg plant material (while soils had a range of 5
10 to 10 mg Pb/kg in the mineral horizons and 50 to 150 mg Pb/kg in the O horizons).
11 Overall, the forest vegetation recycles very little of the Pb present in soils (and thus does
12 not play a direct role in the Pb biogeochemical cycle in boreal forest soils).
13 Fungal species, as represented by mushrooms, accumulate Pb from soils to varying
14 degrees. Based on the uptake of naturally occurring 210Pb, Guillen et al. (2009)
15 established that soil-associated Pb was bioavailable for uptake by mushrooms, and that
16 the highest 210Pb accumulation was observed in Fomes fomentarius mushrooms, followed
17 by Lycoperdon perlatum, Boletus aereus, and Macrolepiota procera, indicating some
18 species differences. Benbrahim et al. (2006) also showed species differences in uptake of
19 Pb by wild edible mushrooms, although they found no significant correlations between
20 Pb content of mushrooms and soil Pb concentration. Pb concentrations in mushroom
21 carpophores ranged from 0.4 to 2.7 mg Pb/kg from sites with soil concentrations ranging
22 from 3.6 and 7.6 mg Pb/kg dry soil. Likewise, Semreen and Aboul-Enein (2011).
23 reported the heavy metal uptake of wild edible mushrooms collected in various
24 mountainous regions of Jordan. Pb BCFs ranged between 0.05 (Russula delicd) and 0.33
25 (Bovistaplumbea) for six mushroom species. Pb BAFs for edible mushrooms collected
26 from quartzite acidic soils in central Spain (containing 19.2 mg Pb/kg) ranged from 0.07
27 (Macrolepiotaprocera) to 0.45 (Lepista nuda) (Campos and Tejera. 2011).
Translocation and Sequestration of Pb in Plants
28 In the 1977 Pb AQCD it was recognized that most Pb taken up from soil remains in the
29 roots and that distribution to other portions of the plant is variable among species (U.S.
30 EPA. 1977). The 2006 Pb AQCD (U.S. EPA. 2006b) stated that most of the Pb absorbed
31 from soil remains bound in plant root tissues either because (1) Pb may be deposited
32 within root cell wall material, or (2) Pb may be sequestered within root cell organelles.
33 More recent research largely confirms that Pb taken up from soil largely remains in roots,
34 but suggests that some species translocate meaningful amounts into shoot tissue.
35 Sequestration of Pb may be a protective mechanism for the plant. Recent findings have
36 been consistent with this hypothesis: Han et al. (2008) observed Pb deposits in the cell
November 2012 7-30 Draft - Do Not Cite or Quote
-------
1 walls and cytoplasm of malformed cells of Iris lactea exposed to 0 to 10 mM Pb (0 to
2 2,072 mg Pb/L) solution in sand culture for 28 days. They hypothesized that preferential
3 sequestration of Pb in a few cells, which results in damage to those cells, helps in
4 maintaining normal overall plant activities through the sacrifice of a small number of
5 active cells. Similarly, macroscopic analysis of the roots of broad bean (Viciafabd)
6 cultivated in mine tailings (average Pb concentration of 7,772 mg Pb/kg) by Probst et al.
7 (2009) revealed dark ultrastructural abnormalities that were demonstrated to be metal-
8 rich particles located in or on root cell walls. It is unclear whether the presence of these
9 structures had any effect on overall plant health.
10 Clark et al. (2006) investigated Pb bioavailability in garden soils in Roxbury and
11 Dorchester, MA. The sources of Pb were considered to be Pb from paints and from
12 leaded gasoline additives, with 40 to 80% coming from paint. The average Pb
13 concentration in foliar tissue of bean plants was 14 ± 5 mg Pb/kg while the concentration
14 in the bean pod was only 20.6 mg Pb/kg. For mustard plants, there was a linear
15 relationship (R2=0.85) between Pb concentration in plant tissues and Pb concentration in
16 the soil (both for plants grown in situ and those grown under greenhouse conditions).
17 Murray et al. (2009) investigated the uptake and accumulation of Pb in several vegetable
18 species (carrot [Daucus carota], radish [Raphanus sativus], lettuce [Lactuca sativa],
19 soybean [Glycine max], and wheat \Triticum aestivum]) from metal-contaminated soils,
20 containing 10 to 40 mg Pb/kg and demonstrated that most Pb remained in the roots. No
21 Pb was measured in the above-ground edible soybean and wheat tissues, while carrots,
22 the most efficient accumulator of Pb, contained a maximum Pb tissue concentration of
23 12 mg Pb/kg dry mass. Similarly, (Cho et al.. 2009) showed that green onion (Allium
24 fistulosum) plants also take up little Pb when planted in soil spiked with Pb nitrate. No
25 plant tissues contained a Pb concentration greater than 24 mg Pb/kg when grown for
26 14 weeks in soils of up to 3,560 mg Pb/kg, and the majority of bioavailable Pb was
27 determined to be contained within the roots. Chinese spinach (Amaranthus dublus) also
28 translocates very little Pb to stem and leaf tissue, and uptake from Pb-containing soils (28
29 to 52 mg Pb/kg) is minimal (Mellem et al.. 2009). Wang et al. (20lie) determined tissue-
30 specific BCFs for wheat grown in soils containing 93 to 1,548 mg Pb/kg. Although the
31 average calculated root BCF was 0.3, very little Pb was translocated to shoots (average
32 BCF=0.02), shells (0.006), and kernels (0.0007) (Wang et al.. 20lie). Sonmez et al.
33 (2008) reported that Pb accumulated by three weed species (Avena sterilis, Isatis
34 tinctoria, Xanthium strumarium) grown in Pb-spiked soils was largely concentrated in the
35 root tissues, and little was translocated to the shoots (Sonmez et al.. 2008).
November 2012 7-31 Draft - Do Not Cite or Quote
-------
1 The Pb BCFs for alfalfa (Medicago sativd) and crimson clover (Trifolium incarnatum)
1 grown in mixtures of heavy metals (Pb concentrations of 10 to 500 (ig Pb/kg) were
3 reportedly low. For alfalfa, BCFs ranged from 0.02 to 0.12, while for crimson clover,
4 these values were between 0.04 and 0.06 (Comino et al.. 2011). The low shoot-root
5 translocation factors reported for alfalfa (0.17 to 0.43) indicated that plant Pb content was
6 largely contained in root tissue. Businelli et al. (2011) calculated whole-plant Pb BAFs
7 for lettuce, radish, tomato and Italian ryegrass using Pb-spiked soils (average values of
8 0.025, 0.021, 0.032, and 0.65, respectively). Again, the majority of accumulated Pb was
9 stored in root tissue, with comparatively little translocated to above-ground tissues
10 (Businelli etal.. 2011).
11 Recent research has shown that Pb translocation to stem and leaf tissues does occur at
12 significant rates in some species, including the legume Sesbania drummondii (Peralta-
13 Videa et al., 2009) and buckwheat (Fagopyrum esculentum) (Tamura et al., 2005). Wang
14 et al. (2006b) noted that Pb soil-to-plant transfer factors were higher for leafy vegetables
15 (Chinese cabbage, pak-choi, and water spinach) than for the non-leafy vegetables tested
16 (towel gourd, eggplant, and cowpea). Tamura et al. (2005) demonstrated that buckwheat
17 is an efficient translocator of Pb. Buckwheat grown in Pb-containing soils collected from
18 a shooting range site (average 1M HC1 extractable Pb= 6,643 mg Pb/kg) preferentially
19 accumulated Pb in leaves (8,000 mg Pb/kg) and shoots (4,200 mg Pb/kg), over root
20 tissues (3,300 mg Pb/kg). Although plant growth was unaffected, this level of leaf and
21 shoot accumulation is likely to have significant implications for exposure of herbivores.
22 Similarly, Shaheen and Tsadilas (2009) reported that vegetables (pepper, okra, and
23 eggplant) grown in soils containing 24 to 30 mg Pb/kg total Pb were more likely to
24 accumulate Pb in leaves (range: undetected to 25 mg Pb/kg) rather than in fruits (range:
25 undetected to 19 mg Pb/kg); however, no significant correlation between soil Pb
26 concentration and plant tissue Pb concentration could be established (Shaheen and
27 Tsadilas. 2009). Tobacco plants were also observed to take up significant amounts of Pb
28 into leaf tissue. Field-grown plants in soils containing an average of 19.8 mg Pb/kg
29 contained average lower, middle and upper leaf Pb concentrations of 11.9, 13.3, and
30 11.6 mg Pb/kg respectively (Zaprianova et al.. 2010). Uptake by tobacco plants was
31 correlated with both total soil Pb concentrations and the mobile Pb fraction (average
32 3.8 mg Pb/kg soil).
33 There is broad variability in uptake and translocation among plant species, and
34 interspecies variability has been shown to interact with other factors such as soil type. By
35 studying multiple species in four Pb-Zn mining sites in Yunnan, China, Li et al. (2009d)
36 demonstrated not only significant differences in uptake and translocation among the
37 species studied, but also modification of the effect on species by type of soil. Plants
38 sampled represented nine species from four families—Caryophyllaceae, Compositae,
November 2012 7-32 Draft - Do Not Cite or Quote
-------
1 Cruciferae, and Pteridaceae. Overall, soil Pb concentration averaged 3,772 mg Pb/kg dry
2 weight, with the highest site average measured at the Minbingying site (5,330 mg Pb/kg),
3 followed by Paomaping (2,409 mg Pb/kg), Jinding (1,786 mg Pb/kg), and Qilinkeng
4 (978 mg Pb/kg). The highest average shoot Pb concentration (3,142 mg Pb/kg) was
5 detected in Stellaria vestita (Caryophyllaceae) collected at Paomaping, while Sinopteris
6 grevilloides (Pteridaceae) collected from Minbingying exhibited the lowest shoot Pb
7 concentration (69 mg Pb/kg). A similar trend was detected in root tissues. S. vestita root
8 collected from the Paomaping area contained the maximum Pb concentration measured
9 (7,457 mg Pb/kg), while the minimum root Pb levels were measured in Picris
10 hieracioides (Pteridaceae) tissues collected from Jinding. These results indicate
11 significant interspecies differences in Pb uptake, as well as potential soil-specific
12 differences in Pb bioavailability. S. vestita, in particular, was determined to be an
13 efficient accumulator of Pb, with a maximum enrichment coefficient of 1.3. Significant
14 correlations between soil Pb concentration and average shoot and root Pb levels were also
15 established (Li et al.. 2009d). Within plant species, the variability in uptake and
16 translocation of Pb may extend to the varietal level. Antonious and Kochhar (2009)
17 determined uptake of soil-associated Pb for 23 unique genotypes from four species of
18 pepper plants (Capsicum chinense, C.frutescens, C. baccatum, and C. annum). Soil Pb
19 concentration averaged approximately 0.6 mg Pb/kg dry soil. No Pb was detected in the
20 fruits of any of the 23 genotypes, except two out of seven genotypes of C. baccatum,
21 which had 0.9 and 0.8 mg Pb/kg dry weight Pb in fruit.
22 Recent studies substantiated findings from the 2006 Pb AQCD that plants store a large
23 portion of Pb in root tissue. Pb soil-to-plant transfer factors are higher for leafy
24 vegetables than for the non-leafy vegetables (Wang et al.. 2006b) and buckwheat has
25 recently been shown to be an efficient translocator of Pb from soil to above-ground
26 shoots (Tamura et al.. 2005).
27 Field studies carried out in the vicinity of Pb smelters (Hu et al.. 2009b) show that Pb
28 may accumulate in shoot tissue through direct stomatal uptake rather than by soil-root-
29 shoot translocation. For instance, Hovmand and Johnsen (2009) determined that about
30 98% of Pb sequestered in Norway spruce needles and twigs was derived from
31 atmospheric sources, and that less than 2% of Pb was translocated from the roots
32 (Hovmand et al.. 2009). Dendrochronology has become more advanced in recent years
33 and is a useful tool for monitoring historical uptake of Pb into trees exposed to
34 atmospheric or soil Pb. Trees accumulate and sequester atmospheric Pb in close
35 correlation with the rate of smelter emissions, although one study indicated that
36 sequestration can lag behind exposure from emissions by 15 years. Pb in the outer woody
37 portion of the tree is more likely the result of direct atmospheric exposure, while Pb in
November 2012 7-33 Draft - Do Not Cite or Quote
-------
1 sapwood is more likely a result of soil uptake. This difference provides an important tool
2 for analyzing source apportionment of Pb accumulation in plants (Guvette et al.. 1991).
7.3.3.2 Terrestrial Invertebrates
3 At the time of publication of the 2006 Pb AQCD (U.S. EPA. 2006b). little information
4 was available regarding the uptake of atmospheric Pb pollution (direct or deposited) by
5 terrestrial invertebrate species. Consequently, few conclusions could be drawn
6 concerning the Pb uptake rate of particular species although there was some evidence that
7 dietary or habitat preferences may influence exposure and uptake. Recent literature
8 indicates that invertebrates can accumulate Pb from consuming a Pb-contaminated diet
9 and from exposure via soil, and that uptake and bioaccumulation of Pb by invertebrates is
10 lower than that observed for other metals.
Snails
11 Pauget et al. (2011) reported that uptake of Pb from soil by the land snail (Cantareus
12 asperses) was most significantly influenced by soil pH and organic matter, as increases in
13 these variables were correlated to decreased Pb bioavailability. Cantareus asperses snails
14 exposed to dietary Pb at 3.3, 86, and 154 mg/kg of diet (spiked with Pb sulfate) for up to
15 64 days were found to assimilate a significant proportion of Pb, and feeding rates were
16 unaffected by the presence of the metal (Beebv and Richmond. 2010). While BCFs for
17 Cd were observed to increase over the 64-day study period, the rate of Pb assimilation
18 remained consistent over time and the authors inferred the absence of a regulatory
19 mechanism for uptake of Pb. The authors speculated that uptake is a function of growth
20 or cell turnover instead. Helix aspersa snails rapidly accumulated Pb from contaminated
21 soil (1,212 mg Pb/kg) and from eating contaminated lettuce (approximately 90 mg Pb/kg
22 after 16 weeks' growth on Pb-contaminated soil) during the first 2 weeks of exposure, at
23 which point snail body burdens reached a plateau (Scheifler et al., 2006b). There were no
24 observed effects of Pb exposure or accumulation on survival or growth in C. asperses or
25 H. aspersa. In another study (Ebenso and Ologhobo. 2009b). juvenile Achatina achatina
26 snails confined in cages on former Pb-battery waste dump sites were found to accumulate
27 Pb from both plant and soil sources. Soil Pb concentration averaged 20, 200, and
28 1,200 mg Pb/kg at the three main waste sites, while leaf tissues of radish (Raphanus
29 sativus) grown at these sites averaged 7, 30, and 68 mg Pb/kg dry weight, respectively.
30 Concentration of Pb in snail tissues rose with concentration in both soil and plants, and
31 the authors found that for both sources, a log-log relationship could be estimated with a
32 very close fit (r2 =0.94 and 0.95, respectively). Pb concentration in snail tissues averaged
November 2012 7-34 Draft - Do Not Cite or Quote
-------
1 12, 91, and 468 mg Pb/kg, respectively, at the three sites, which the authors stipulated
2 were above the maximum permissible concentration of Pb for human consumption of
3 mollusks, mussels, and clams (1.5 mg Pb/kg tissue) as determined by the U.K. Food
4 Standards Agency. Pb concentration in snail tissues generally is much lower than that of
5 the soil substrates upon which they were reared, but higher than in other soil-dwelling
6 organisms. De Vaufleury et al. (2006) exposed Helix aspersa snails to standardized
7 (International Organization for Standardization methodology [ISO 11267:1999])
8 artificial-substrate soils amended with sewer sludge containing 13, 26, 39, or 52 mg
9 Pb/kg for 28 days without supplemental food. After the exposure period, snail foot tissue
10 contained increased levels of Pb—1.9, 1.7, and 1.5 mg Pb/kg dry weight versus
11 concentration averaging 0.4 mg Pb/kg in control organisms. Viscera also exhibited
12 increased Pb levels at the two highest exposures, with measured tissue concentration of
13 1.2 and 1.1 mg Pb/kg, respectively, as compared with control tissue Pb levels of 0.4 mg
14 Pb/kg. However, there was no significant increase in snail-tissue Pb concentration when
15 natural soil was used in place of ISO medium, and there was no relationship between soil
16 Pb concentration and snail tissue concentration, strongly suggesting the presence of soil
17 variables that modify bioavailability. Notten et al. (2008) investigated the origin of Pb
18 pollution in soil, plants, and snails by means of Pb isotope ratios. They found that a
19 substantial proportion of Pb in both plants and snails was from current atmospheric
20 exposure.
21 Finally, a study by Coeurdassier et al. (2007) found that the presence of snails was
22 associated with higher Pb content in earthworms, suggesting that snails themselves may
23 have an effect on bioavailability.
Earthworms
24 Accumulation studies conducted with Eisenia sp. earthworms documented the difficulty
25 of extrapolating accumulation kinetic constants from one soil type to another, and
26 showed that many soil physiochemical properties, including pH, organic matter, and
27 CEC, among others, affect metal bioavailability (Nahmani et al.. 2009). Source of Pb,
28 and proportion of soil:leaf litter also affect Pb bioavailability. Bradham et al. (2006)
29 examined the effect of soil chemical and physical properties on Pb bioavailability.
30 Eisenia andrei earthworms were exposed to 21 soils with varying chemical and physical
31 properties that were freshly spiked with Pb to give a standard concentration of 2,000 mg
32 Pb/kg dry weight. At equivalent Pb exposure, the main determinants of both internal
33 earthworm Pb concentration and mortality were pH first (with lower pH resulting in
34 higher concentration and mortality), then CEC. However, the apparent importance of
35 CEC was due to its correlation with several other less important soil characteristics.
36 These data corroborate that Pb bioavailability and toxicity are increased in acidic soils
November 2012 7-35 Draft - Do Not Cite or Quote
-------
1 and in soils with a low CEC (Section 7.3.2). This finding was confirmed by Gandois et al.
2 (2010). who determined that the free-metal-ion fraction of total Pb concentration in field-
3 collected soils was largely predicted by pH and soil Fe content.
4 The role of soil profile and preferred depth was studied using eight species of earthworms
5 from 27 locations in Switzerland, representing three ecophysiological groups (Ernst et al..
6 2008): epigeic (surface-dwelling worms), endogeic (laterally burrowing worms that
7 inhabit the upper soil layers), and anecic (vertically burrowing worms that reach depths
8 of 6 inches). For epigeic and anecic earthworms, the total concentration of Pb in leaf litter
9 and in soil, respectively, were the most important drivers of Pb body burdens. By
10 contrast, the level of Pb in endogeic earthworms was largely determined by soil pH and
11 CEC. As a result of these differences, the authors suggested that atmosphere-sourced Pb
12 may be more bioavailable to epigeic than endogeic species, because it is less dependent
13 on modifying factors. Suthar et al. (2008). on the other hand, found higher Pb
14 bioaccumulation in the endogeic earthworm Metaphire posthuma than in the anecic
15 earthworm species Lampito mauritii, and speculated that differences in Pb tissue level
16 arose from differing life-history strategies, such as feeding behaviors, niche preferences,
17 and burrowing patterns, all of which exposed the endogeic species to greater Pb
18 concentration. Garg et al. (2009) reported that the smaller native earthworm
19 Allolobophoraparva accumulated significantly greater Pb concentrations than E. fetida.
20 Subsequently, it was concluded that native earthworm species may exhibit a higher Pb
21 accumulation potential as a result of increased tolerance to the heavy metal (Garg et al..
22 2009).
23 Earthworm activity can alter Pb bioavailability and subsequent uptake by earthworms
24 themselves and other organisms. Sizmur and Hodson (2009) speculated that earthworms
25 affect Pb mobility by modifying the availability of cations or anions. The concentration
26 of water-soluble Pb was observed to increase following earthworm (Lumbricus terrestris)
27 feeding activity in field-collected soils containing 132.7, 814.9, and 821.4 mg total Pb/kg
28 (calculated BAFs of 0.27, 0.33, and 0.13, respectively) (Alonso-Azcarate et al.. 2011).
29 However, Coeurdassier et al. (2007) found that snails did not have a higher Pb content
30 when earthworms were present, and that unexpectedly, Pb was higher in earthworm
31 tissue when snails were present.
32 Despite significant Pb uptake by earthworms, Pb in earthworm tissue may not be
33 bioavailable to predators. Pb in the earthworm (Aporrectodea caliginosa) was determined
34 to be contained largely in the granular fraction (approximately 60% of total Pb), while the
35 remaining Pb body burden was in the tissue, cell membrane, and intact cell fractions
36 (Vijver et al.. 2006). However, this may vary by species, as (Li et al.. 2008b) found that
37 more than half of the Pb accumulated by E. fetida was contained within earthworm tissue
November 2012 7-36 Draft - Do Not Cite or Quote
-------
1 and cell membranes. Regardless, Vijver et al. (2006) concluded that only a minority of
2 earthworm-absorbed Pb would be lexicologically available to cause effects in the
3 earthworms or in their predators.
Arthropods
4 Pb and other metals were analyzed in honeybees (Apis mellifera) foraging in sampling
5 sites that included both urban areas and wildlife reserves in central Italy. (Perugini et al..
6 2011). Pb in whole bees ranged from 0.28 to 0.52 mg Pb/kg with the highest
7 concentration in honeybees caught in hives near an airport. Cicadas pupating in
8 historically Pb-arsenate-treated soils accumulated Pb at concentrations similar to those
9 reported previously for earthworms (Robinson et al.. 2007). Likewise, tissue Pb levels
10 measured in Coleoptera specimens collected from areas containing average soil
11 concentration of 45 and 71 mg Pb/kg exhibited a positive relationship with soil Pb
12 content, although abundance was unaffected (Schipper et al.. 2008). By contrast, two
13 grasshopper species inhabiting Pb and Cd-contaminated areas near Zn smelting facilities
14 exhibited different Pb accumulation rates. Locust (Locusta migratoria) collected from
15 areas with an average Pb soil concentration of 540mg Pb/kg contained 47 mg Pb/kg,
16 while grasshoppers (Acrida chinensis) inhabiting the same area accumulated 93.9 mg
17 Pb/kg (Zhang etal.. 2012). This gives respective BAFs of 0.09 and 0.17. Similarly, the
18 Pb sequestration rates that were observed in two woodlouse species, O. asellus and
19 P. scaber, were species-dependent (Gal et al.. 2008). Both species were field collected at
20 Pb-contaminated sites (average concentration, 245 mg Pb/kg dry weight; range,
21 21-638 mg Pb/kg dry weight), with O. asellus Pb levels averaging 43 mg Pb/kg over all
22 sites, while P. scaber contained no detectable Pb residues. Pb concentration measured in
23 granivorous rough harvester ants (Pogonomyrmex rugosus), in the seeds of some plant
24 species they consume, and in surface soil, were all shown to decline with increasing
25 distance from a former Pb smelter near El Paso, Texas, where soil leachable Pb at the
26 three sites of ant collection ranged from 0.003 to 0.117 mg Pb/kg (Del Toro et al.. 2010).
27 Ants accumulated approximately twice as much Pb as was measured in seeds, but the
28 study did not separate the effects of dietary exposure from those of direct contact with
29 soil or respiratory intake.
7.3.3.3 Terrestrial Vertebrates
30 At the time of the 1977 Pb AQCD few studies of Pb exposure and effects in wild animals
31 other than birds had been conducted. A limited number of rodent trapping studies near
32 roadsides indicated general trends of species differences in Pb uptake and higher
November 2012 7-37 Draft - Do Not Cite or Quote
-------
1 concentrations of Pb in habitats adjacent to high-traffic areas (U.S. EPA. 1977). In the
2 1986 Pb AQCD concentration of Pb in bone tissue was reported for selected herbivore,
3 omnivore and carnivore species [Table 8-2 in (U.S. EPA, 1986b)1.
4 Tissue Pb residues in birds and mammals associated with adverse toxicological effects
5 were presented in the 2006 Pb AQCD. In general, avian blood, liver, and kidney Pb
6 concentrations of 0.2-3 (ig Pb/dL, 2-6 mg Pb/kg wet weight, and 2-20 mg Pb/kg wet
7 weight, respectively, were linked to adverse effects. A few additional studies of Pb
8 uptake and tissue residues in birds and mammals conducted since 2006 are reviewed
9 here.
10 In a study of blood Pb levels in wild Steller's eiders (Polysticta stelleri) and black scoters
11 (Melanitta nigra) in Alaska, the authors compiled avian blood Pb data from available
12 literature to develop reference values for sea ducks (Brown et al., 2006). The background
13 exposure reference value of blood Pb was <20 (ig Pb/dL, with levels between 20 and
14 59 (ig Pb/dL as indicative of Pb exposure. Clinical toxicity was in the range of
15 60-99 (ig Pb/dL in birds while >100 (ig Pb/dL results in acute, severe toxicity. In
16 measurement of blood Pb with a portable blood Pb analyzer, only 3% of birds had values
17 indicating exposure and none of the birds had higher blood Pb levels or clinical signs of
18 toxicity. Tissue distribution of Pb in liver, kidney, ovary and testes of rain quail (Coturnix
19 coramandelica) following oral dosing of 0.5 mg Pb/kg, 1.25 mg Pb/kg or 2.5 mg Pb/kg
20 Pb acetate for 21 days indicated that Pb uptake was highest in liver and kidney and low in
21 ovary and testes (Mehrotra et al.. 2008). Resident feral pigeons (Columba livid) captured
22 in the urban and industrial areas of Korea exhibited increased lung Pb concentration,
23 ranging from 1.6 to 1.9 mg Pb/kg wet weight (Nam and Lee. 2006). However, tissue
24 concentration did not correlate with atmospheric Pb concentration, so the authors
25 concluded that ingestion of particulate Pb (paint chips, cement, etc.) in the urban and
26 industrial areas was responsible for the pigeons' body burden. Similarly, 70% of
27 American woodcock (Scolopax minor) chicks and 43% of American woodcock young-of-
28 year collected in Wisconsin, U.S., exhibited high bone Pb levels of 9.6-93 mg Pb/kg dry
29 weight and 1.5-220 mg Pb/kg, respectively, even though radiographs of birds'
30 gastrointestinal tracts revealed no evidence of shot ingestion (Strom et al., 2005). Authors
31 hypothesized that unidentified anthropogenic sources may have caused the observed
32 elevated Pb levels.
33 In addition to birds, soil-dwelling mammals can also bioaccumulate atmospherically-
34 sourced Pb. Northern pocket gophers (Thomomys talpoides) trapped within the Anaconda
35 Smelter Superfund Site were shown to accumulate atmospherically deposited Pb. Gopher
36 liver and carcass Pb concentration averaged 0.3 and 0.4 mg Pb/kg wet weight on low Pb
37 soils (47 mg Pb/kg), 0.4 and 0.9 mg Pb/kg wet weight in medium Pb soils (95 mg Pb/kg)
November 2012 7-38 Draft - Do Not Cite or Quote
-------
1 and 1.6 and 3.8 mg Pb/kg wet weight in high Pb soils (776.5 mg Pb/kg) (Reynolds et al..
2 2006). Likewise, rats trapped in the vicinity of a Kabwe, Zambia Pb-Zn mine had
3 significantly elevated liver and kidney Pb concentrations. Soil Pb concentrations were
4 measured between 9 and 51,188 mg Pb/kg (approximate average of 200 mg Pb/kg dry
5 weight), while rat liver and kidney Pb concentrations ranged between 0.009 and 7.3 mg
6 Pb/kg dry weight and 0.3 and 22.1 mg Pb/kg dry weight, respectively. Consequently,
7 residence in the mining region was correlated to significantly increased Pb body burdens
8 for rats (Nakavama et al.. 2011). Angelova et al. (2010) reared rabbits on a fodder
9 mixture containing Pb-contaminated rapeseed grown adjacent to a metal works plant.
10 Following a four-week exposure, Pb was most heavily concentrated in rabbit kidney
11 tissue (3.9 mg Pb/kg and 1.9 mg Pb/kg, for high and low diet respectively), bone (1.0 and
12 0.3 mg Pb/kg, respectively), and liver (0.6 and 0.4 mg Pb/kg, respectively). Yucatan
13 micropigs (Sus scrofd) and Sprague-Dawley rats (Rattus norvegicus) reared on
14 Pb-contaminated soil (5% of 1,000 mg Pb/kg soil as dietary component) consumed
15 significantly different amounts of Pb. Over a 30-day period, rats consumed an average of
16 19.4 mg Pb, while micropig intake averaged 948 mg Pb (Smith et al.. 2009a). This
17 resulted in significantly higher Pb accumulation in micropigs, based on liver, blood,
18 kidney and bone Pb concentrations (average concentrations of 1.2, 25, 0.9, and 9 mg
19 Pb/kg for micropigs, and 0.2, 7, 0.5, and 1.5 mg Pb/kg for rats, respectively).
20 Casteel et al. (2006) found that bioavailability of Pb from environmental soil samples in
21 swine (Sus domesticd) depended on Pb form or type, with high absorption of cerussite
22 and Mn-Pb oxides and poor absorption of galena and anglesite. Juvenile swine
23 (approximately 5-6 weeks old and weighing 8-11 kg) were fed Pb-contaminated soils
24 collected from multiple sources for 15 days (concentration range of 1,270 to 14,200 mg
25 Pb/kg) to determine the relative bioavailability. While Pb concentrations were roughly
26 equivalent in blood, liver, kidney, and bone tissues, individual swine exhibited different
27 uptake abilities (Casteel et al.. 2006).
28 Consistent with observations in humans, dietary Ca2+ deficiency (0.45 mg Ca2+ daily
29 versus 4 mg under normal conditions) was linked to increased accumulation of Pb in
30 zebra finches (Taeniopygia guttatd) that were provided with drinking water containing
31 20 mg Pb/L (Dauwe et al.. 2006). Liver and bone Pb concentration were increased by an
32 approximate factor of three, while Pb concentration in kidney, muscle, and brain tissues
33 were roughly doubled by a Ca2+-deficient diet. However, it is not known whether this
34 level of dietary Ca2+ deficiency is common in wild populations of birds.
November 2012 7-39 Draft - Do Not Cite or Quote
-------
7.3.3.4 Food Web
1 In addition to the organism-level factors reviewed above, understanding the
2 bioavailability of Pb along a simple food chain is essential for determining risk to
3 terrestrial animals. While the bioavailability of ingested soil or particles is relatively
4 simple to measure and model, the bioavailability to secondary consumers of Pb ingested
5 and sequestered by primary producers and primary consumers is more complex. Kaufman
6 et al. (2007) caution that the use of total Pb concentration in risk assessments can result in
7 overestimation of risk to ecological receptors, and they suggest that the bioaccessible
8 fraction may provide a more realistic approximation of receptor exposure and effects.
9 This section reviews recent literature that estimates the bioaccessible fraction of Pb in
10 dietary items of higher order consumers, and various studies suggesting that Pb may be
11 transferred through the food chain but that trophic transfer of Pb results in gradual
12 attenuation, i.e., lower concentration at each successive trophic level.
13 Earthworm and plant vegetative tissue collected from a rifle and pistol range that
14 contained average soil Pb concentration of 5,044 mg Pb/kg were analyzed for Pb content
15 and used to model secondary bioavailability to mammals (Kaufman et al.. 2007).
16 Earthworms were determined to contain an average of 727 mg Pb/kg, and the Pb content
17 of unwashed leaf tissues averaged 2,945 mg Pb/kg. Canonical correspondence analysis
18 detected no relationship between earthworm and soil Pb concentration, but did show
19 correlation between unwashed vegetation and soil concentration. The authors noted that
20 the relatively high Pb concentration of unwashed as opposed to washed vegetation
21 indicated the potential importance of aerial deposition (or dust resuspension) in
22 determining total vegetative Pb concentration. Based on the mammalian gastric model,
23 they noted that 50% of vegetation tissue Pb and 77% of earthworm tissue Pb was
24 expected to be bioavailable to consumers. The avian gizzard model indicated that 53% of
25 soil Pb and 73% of earthworm Pb was bioaccessible to birds; and, for both mammals and
26 birds, the bioaccessible fraction of Pb was a function of total Pb concentration.
27 The transfer of Pb from soils contaminated by a Pb-Zn mine was limited along a soil-
28 plant-insect-chicken food chain (Zhuang et al.. 2009). In soils averaging 991 mg Pb/kg,
29 plants of the fodder plant Rumex patientia X tianschanicus sequestered an average of
30 1.6 mg Pb/kg wet weight in the shoot tissue, while larvae of the leafworm Spodoptera
31 litura accumulated an average Pb concentration of 3.3 mg Pb/kg wet weight S. litura-fed
32 chickens (Gallus gallus domesticus) accumulated 0.58 mg Pb/kg and 3.6 mg Pb/kg in
33 muscle and liver tissue, respectively, but only liver Pb burden was increased significantly
34 relative to controls. A large proportion of ingested Pb was excreted with the feces.
35 Likewise, an insectivorous bird species, the black-tailed godwit (Limosa limosa) was
36 shown to accumulate Pb from earthworms residing in Pb-contaminated soils (Roodbergen
November 2012 7-40 Draft - Do Not Cite or Quote
-------
1 et al.. 2008). Pb concentration in eggs and feathers was increased in areas with high soil
2 and earthworm Pb concentration (336 and 34 mg Pb/kg, respectively): egg Pb
3 concentration averaged 0.17 mg Pb/kg and feather concentration averaged 2.8 mg Pb/kg.
4 This suggests that despite a residence breeding time of only a few months, this bird
5 species could accumulate Pb when breeding areas are contaminated.
6 Rogival et al. (2007) showed significant positive correlations between soil Pb
7 concentration along a gradient (approximately 50 to 275 mg Pb/kg) at a metallurgical
8 plant, and Pb concentration in both acorns (from Quercus robur) and earthworms
9 (primarily Dendrodrilus rubidus and Lumbricus rubellus) collected on site. Acorn and
10 earthworm Pb contents were, in turn, positively correlated with the Pb concentration in
11 the liver, kidney, and bone tissues of locally trapped wood mice (Apodemus sylvaticus).
12 The uptake and transfer of Pb from soil to native plants and to red deer (Cervus elaphus)
13 was investigated in mining areas of the Sierra Madrona Mountains in Spain (Reglero et
14 al.. 2008). The authors reported a clear pattern between plant Pb concentration and the Pb
15 content of red deertissues with attenuation (i.e., decreasing concentration) of Pb up the
16 food chain. Interestingly, soil geochemistry likely was affected by mining activity as
17 Holm oak (Quercus ilex), gum rockrose (Cistus ladanifef), elm leaf blackberry (Rubus
18 ulmifolius), and grass (Graminae) tissues collected from mining areas exhibited increased
19 Pb levels (up to 98 mg Pb/kg in grasses and 21 mg Pb/kg in oak) despite the fact that total
20 soil Pb concentrations were not significantly greater than those of the non-mining areas.
21 Positive relationships were observed between Cepaea nemoralis snail tissue Pb levels
22 and Pb concentration measured in Urtica dioica leaves in field-collected samples from
23 areas characterized by metal soil contamination (approximately 200 to 400 mg Pb/kg)
24 (Notten et al.. 2005). Inouye et al. (2007) found that several invertebrate prey offence
25 lizards, including Acheta domestica crickets, Tenebrio molitor beetles, and P. scaber
26 isopods, accumulate Pb from dietary exposures (10, 50, 100, 250, 500, 750, and 1,000 mg
27 Pb/kg) lasting between 44 and 72 days. By day 44, Pb body burdens of crickets were 31,
28 50 and 68 mg Pb/kg (wet weight) at the three highest dietary exposures, respectively.
29 Isopods and beetle larvae accumulated significantly less Pb, with average body burdens
30 of 10, 15, and 14 mg Pb/kg following 56 days of exposure; and 12, 14, and 31 mg Pb/kg
31 following 77 days of exposure, respectively. For all invertebrates tested, Pb was
32 sequestered partly in the exoskeleton, and partly in granules. Exoskeleton Pb may be
33 available to predators, but returns to background level with each shedding, while granular
34 Pb is likely unavailable, at least to other invertebrates (Vijver et al.. 2004).
35 In a comparison of rural and urban blackbirds (Turdus meruld), Sheifler et al.(2006a)
36 found that while Pb concentration in unwashed tail feathers was equivalent in both
37 populations, urban birds had higher tissue concentrations. Pb content of urban
November 2012 7-41 Draft - Do Not Cite or Quote
-------
1 earthworms was also higher than that of rural earthworms. Hypothesizing that tail feather
2 Pb reflected deposition from air and resuspended dust, the authors suggested that elevated
3 Pb in the urban birds was mostly dietary in origin.
4 Overall, studies of Pb transfer in food webs have established the existence of pervasive
5 trophic transfer of the metal, but no consistent evidence of trophic magnification. It
6 appears that on the contrary, attenuation is common as Pb is transferred to higher trophic
7 levels. However, many individual transfer steps, as from particular plants to particular
8 invertebrates, result in concentration, which may then be undone when stepping to the
9 next trophic level. It is possible that whether trophic transfer is magnifying or attenuating
10 depends on Pb concentration itself. Kaufman et al. (2007) determined that, at low
11 concentrations of soil Pb, risk to secondary consumers (birds and mammals) was driven
12 by the bioavailability of Pb in worm tissues, while at high soil concentrations,
13 bioavailability of soil-associated Pb was more critical. The authors concluded that
14 incorporation of bioavailability/bioaccessibility measurements in terrestrial risk
15 assessments could lead to more accurate estimates of critical Pb levels in soil and biota.
16 Finally, while trophic magnification does greatly increase exposure of organisms at the
17 higher levels of the food web, these studies establish that atmospherically deposited Pb
18 reaches species that have little direct exposure to it. For those species, detrimental effects
19 are not a function of whether they accumulate more Pb than the species they consume,
20 but of the absolute amount they are exposed to, and their sensitivity to it.
7.3.4 Biological Effects of Pb in Terrestrial Systems
21 Various effects can be observed in exposed terrestrial species following uptake and
22 accumulation of Pb. While many of the responses are specific to organism type, induction
23 of antioxidant activities in response to Pb exposure has been reported in plants,
24 invertebrates, and vertebrates. In this section, the observed biological effects caused by
25 exposure to atmosphere-derived Pb will be discussed, while the results of dose-response
26 experimentation will be addressed in Section 7.3.5. Because environmental releases of Pb
27 often include simultaneous release of other metals, it can be difficult to identify
28 Pb-specific effects in field studies, with the exception of effects from leaded gasoline and
29 some Pb smelter deposition. Many laboratory studies that expose organisms to natural
30 soils (or to biosolids-amended soils) also include exposure to multiple metals. There is
31 some information about mechanisms of metal interactions, such as through competition
32 for binding locations on specific enzymes or on cellular receptors, but generally such
33 interactions (particularly of multiple metals) are not well understood (ATSDR. 2004).
34 Despite a few well-known examples of metal antagonism (e.g., Cu and Mo or Cd and
35 Zn), it is common practice to assume additivity of effects (Fairbrother et al.. 2007).
November 2012 7-42 Draft - Do Not Cite or Quote
-------
1 Because this review is focused on effects of Pb, studies reviewed for this section and the
2 following include only those for which Pb was the only, or primary, metal to which the
3 organism was exposed. All reported values are from exposures in which concentrations
4 of Pb were analytically verified unless nominal concentrations are stated.
7.3.4.1 Terrestrial Plants and Lichen
5 Pb exposure has been linked to decreased photosynthesis in affected plants, significant
6 induction of antioxidant activities, genetic abnormalities, and decreased growth.
7 Induction of antioxidant responses in plants has been shown to increase tolerance to
8 metal soil contamination, but at sufficiently high levels, antioxidant capacity is exceeded,
9 and metal exposure causes peroxidation of lipids and DNA damage, eventually leading to
10 accelerated senescence and potentially death (Stobrawa and Lorenc-Plucinska. 2008).
Effects on Photosystem and Chlorophyll
11 Photosynthesis and mitosis were recognized as targets of Pb toxicity in plants in the 1977
12 Pb AQCD and additional effects of Pb on these processes were reported in subsequent Pb
13 AQCDs (U.S. EPA. 2006c. 1986b. 1977). The effect of Pb exposure on the structure and
14 function of plant photosystem II was recently studied in giant duckweed, Spirodela
15 polyrrhiza (Ling and Hong. 2009). Although this is an aquatic plant, photosystem II is
16 present in all plants. This finding thus provides support for effects on photosystem II
17 being the cellular-level mechanism that leads to decreases photosynthesis observed in
18 other plants. The Pb concentration of extracted photosystem II particles was found to
19 increase with increasing environmental Pb concentration, and increased Pb concentration
20 was shown to decrease emission peak intensity at 340 nm, amino acid excitation peaks at
21 230 nm, tyrosine residues, and absorption intensities. This results in decreased efficiency
22 of visible light absorption by affected plants. The authors theorized that Pb2+ may replace
23 either Mg2+ or Ca2+ in chlorophyll or the oxygen-evolving center, inhibiting photosystem
24 II function through an alteration of chlorophyll structure. Consistently with these results,
25 Wu et al. (2008c) demonstrated that Pb exposure interfered with and decreased light
26 absorption by spinach (Spinacia oleracea) plants. Spinach seeds were soaked in 5, 12, or
27 25 mM Pb chloride (1036, 2486, or 5180 mg Pb/L) for 48 hours prior to germination, and
28 following 42 days of growth, plants were sprayed with Pb chloride solutions. Chloroplast
29 absorption peak intensity, fluorescence quantum yield at 680 nm, and whole-chain
30 electron transport rate all decreased with Pb exposure, as did photosystem II
31 photoreduction and oxygen evolution. Similarly, the photosynthetic rate of maize (Zea
32 mays) seedlings decreased over 21 days exposure to Pb, and measured leaf Pb
November 2012 7-43 Draft - Do Not Cite or Quote
-------
1 concentrations in photosynthetically-depressed seedlings ranged from approximately 100
2 to 300 mg Pb/kg dry weight (Ahmad et al.. 2011). Liu et al. (2010a) observed that
3 chlorophyll a and b content in wheat grown in soils spiked with Pb nitrate rose with
4 length of exposure until 14 days, at which point chlorophyll decreased. At nominal
5 exposures of 0.1 and 0.5 mM Pb (20.72 and 103.6 mg Pb/L) in hydroponic solution for
6 50 days, concentration of chlorophyll a and b was decreased in radish (R. sativus)
1 (Kumar and Tripathi. 2008). Changes in chlorophyll content in response to Pb were also
8 observed in lichen and moss species following exposures intended to simulate
9 atmospheric deposition (Carreras and Pignata. 2007). Usnae ambfyoclada lichen was
10 exposed to aqueous Pb solutions of 0.5, 1, 5, and 10 mM Pb nitrate (103.6, 207.2, 1,036,
11 and 10,360 mg Pb/L); chlorophyll a concentration was shown to decrease with increasing
12 Pb exposure. However, the ratio of lichen dry weight to fresh weight increased following
13 Pb exposures. It should be noted that highly productive Sphagnum mosses accumulated
14 atmospheric Pb at the same rate as slower growing mosses, indicating that moss growth
15 allowed for further Pb uptake, rather than a "dilution" effect (Kempter et al.. 2010). As
16 compared to other metals, however, Pb caused less physiological damage, which the
17 authors attributed to the metal's high affinity for binding to and sequestration within cell
18 walls (Carreras and Pignata. 2007).
19 The effect of Pb exposure on chlorophyll content of the moss and liverwort species
20 Thuidium delicatulum, T. sparsifolium, and Ptychanthus striatus was investigated
21 following immersion in six solutions of Pb nitrate containing from 10"10 to 10"2 M Pb
22 (0.00002 to 2,072 mg Pb/L) (Shakva et al.. 2008). Both chlorophyll a and total
23 chlorophyll content of the mosses (T. delicatulum and T. sparsifolium) decreased with
24 increasing Pb exposure. For the liverwort, increasing Pb exposure resulted in decreases in
25 content of chlorophyll a, chlorophyll b, and total chlorophyll. Further, the total
26 chlorophyll content of Hypnumplumaeforme mosses was decreased by 5.8% following
27 exposure to the highest concentration, while lower exposures slightly elevated
28 chlorophyll content.
29 These studies suggest that exposure to Pb has an impact on photosynthetic pigments, but
30 the exposure methods (seed soaking, spraying of Pb chloride solutions, hydroponic
31 growth systems) make it difficult to compare these effects to what might occur under the
32 uncontrolled conditions encountered in natural environments. These experiments bring to
33 light the presence of effects, and the underlying mechanisms, but strong uncertainties
34 remain regarding the natural concentrations at which theses effects would be observed.
November 2012 7-44 Draft - Do Not Cite or Quote
-------
Response of Antioxidants
1 Increased antioxidant activity is a common response to Pb exposure, although this
2 endpoint may not necessarily be an indication of deleterious effects on plant vitality.
3 Increases in reactive oxygen species with increasing exposure to Pb from 20 mg Pb/kg
4 soil to 2,000 mg Pb/kg have been demonstrated in broad bean (Viciafabd) (Wang et al..
5 2010c; Wang et al., 2010a; Wang et al., 2008b) and tomato (Lycopersicon esculentum)
6 (Wang et al.. 2008a). where they were accompanied up to approximately 500mg Pb/kg by
7 proportional increases in superoxide dismutase (SOD), glutathione, guaiacol peroxidase,
8 and lipid peroxidation, as well as decreases in catalase. Spinach seedlings grown in soil
9 containing six increasing concentrations of Pb from 20 to 520 mg Pb/kg exhibited higher
10 production of reactive oxygen species, increased rates of lipid peroxidation and increased
11 SOD concentrations. Many of these responses persisted for 50 days after germination and
12 growth in the Pb-contaminated soil (Wang etal. 201 la). Similarly, the bryophyte mosses
13 Hypnum plumaeforme, Thuidium cymbifolium, and Brachythecium piligerum exposed to
14 Pb solutions of greater than 0.1 mM Pb for 48 hours exhibited increased production of
15 «O2 and H2O2, although no single moss species could be identified as most sensitive to
16 Pb exposure (Sun et al.. 2011). Increased rates of lipid peroxidation were also observed in
17 Pb-exposed mosses; however, SOD and catalase activity was suppressed or unaffected by
18 Pb.
19 Reddy et al. (2005) found that horsegram (Macrotyloma uniflorum) and bengalgram
20 (Cicer arietinum) plants watered with Pb solutions containing 200, 500, and 800 mg Pb/L
21 exhibited increased antioxidant activity: at exposures of 800 mg Pb/L, root and shoot
22 SOD activity increased to 2-3 times that of controls, and induction was slightly higher in
23 M. uniflorum. Similarly, catalase, peroxidase, and glutathione-S-transferase activities
24 were elevated in Pb-stressed plants, but were again higher forM uniflorum. Antioxidant
25 activities were also markedly greater in the root tissues than the shoot tissues of the two
26 plants, and were very likely related to the higher Pb accumulation rate of the roots. The
27 effectiveness of the up-regulation of antioxidant systems in preventing damage from Pb
28 uptake was evidenced by lower malondialdehyde (MDA) (a chemical marker of lipid
29 peroxidation) concentration inM uniflorum versus C. arietinum, indicating a lower rate
30 of lipid peroxidation in response to M. uniflorum's higher antioxidant activity.
31 Gupta et al. (2010) contrasted responses of two ecotypes ofSedum alfredii (an Asian
32 perennial herb), one an accumulator of Pb collected from a Pb and Zn mining area, and
33 the other not. Glutathione level was increased in both, and root and shoot lengths were
34 decreased following long-term exposures to Pb up to 200 (iM (41.4 mg Pb/L) in
35 hydroponic solution. However, the accumulator plants exhibited greater SOD and
36 ascorbate peroxidase activity, likely as a result of greater Pb uptake and a concurrent
37 increased detoxification capacity. Similar results were reported by Islam et al. (2008):
November 2012 7-45 Draft - Do Not Cite or Quote
-------
1 following Pb exposures of 200 (iM (41.4 mg Pb/L), catalase, ascorbic acid, and
2 glutathione levels of another Chinese herb, Elsholtzia argyi, were increased, while SOD
3 and guaiacol peroxidase activities decreased. Microscopic analysis also showed that
4 affected plants exhibited abnormal chloroplast structures. The response of glutathione
5 was further confirmed in wheat (Liu et al., 2010a) grown in soils spiked with Pb nitrate.
6 Evidence of increasing lipid peroxidation (MDA accumulation) with increasing Pb
7 exposure was also found in mosses (Sun et al., 2009) and lichens. Lichens field-collected
8 from the trunks of poplar (Populus tremula) trees in eastern Slovakia were chemically
9 analyzed for metal concentration arising from exposure to smelter pollution (Dzubaj et
10 al., 2008). These concentrations (ranging from 13 to 1,523 mg Pb/kg dry weight) were
11 assessed in relation to physiological variables, including chlorophyll a and b, carotenoids,
12 photosystem II activity, CO2 gas exchange (respiration), and MDA content. Lichen Pb
13 levels were significantly correlated only with MDA content. Determination of plant
14 chitinase content following exposure to As, Cd and Pb indicated that while levels of these
15 defense proteins were elevated by As and Cd, chitinase levels were not increased
16 following exposure to Pb (Bekesiova et al., 2008). As in studies of effects on
17 photosynthesis, the methods used for exposure make it difficult to compare these effects
18 to what might occur under the uncontrolled conditions encountered in natural
19 environments.
Growth
20 Evidence of effects of Pb on higher growth processes in terrestrial plants was reported in
21 early NAAQS reviews. Impacts to growth can lead to effects at the population-level of
22 biological organization and higher (Section 7.1.1). Growth effects of Pb on plants in the
23 1977 Pb AQCD primarily included visible growth responses observed in laboratory
24 studies with plants grown in artificial nutrient culture (U.S. EPA. 1977). No Pb toxicity
25 was observed in plants growing under field conditions at the time of the 1977 Pb AQCD.
26 Indirect effects of Pb on plant growth (i.e., inhibition of uptake of other nutrients when
27 Pb is present in the plant) were also reported in the 1977 Pb AQCD. In the 1986 Pb
28 AQCD mechanisms of Pb effects on growth included reduction of photosynthetic rate,
29 inhibition of respiration, cell elongation, root development or premature senescence (U.S.
30 EPA. 1986b). All of these effects were observed to occur in isolated cells or in plants
31 grown hydroponically in solutions comparable to 1 to 2 mg Pb/kg soil or in soils with
32 10,000 mg Pb/kg or greater (U.S. EPA. 1986R Pb effects on other plant processes,
33 especially maintenance, flowering and hormone development had not been studied at the
34 time of the 1986 Pb AQCD and remain poorly characterized.
November 2012 7-46 Draft - Do Not Cite or Quote
-------
1 Recent evidence for growth effects in terrestrial plants available since the
2 2006 Pb AQCD is reviewed below and summarized in Table 7-4. Both growth and
3 carotenoid and chlorophyll content of Brassica juncea (mustard) plants were negatively
4 affected by Pb exposure (John et al.. 2009). Nominal Pb treatments of 1,500 (iM (311 mg
5 Pb/L) as Pb acetate solution decreased root lengths and stem heights by 50% after 60
6 days. Exposure to 600 (iM Pb (124 mg Pb/L) and greater decreased carotenoid content,
7 while chlorophyll a was decreased at Pb exposures of 450 (iM (93 mg Pb/L) and higher.
8 However, when smelter ash-spiked soils containing 1,466 mg Pb/kg (and 18.6 mg Cd/kg)
9 or 7,331 mg Pb/kg (98.0 mg/kg Cd) were used to grow maize (Zea mays), as well as other
10 metals in high concentrations, effects were seen in growth or chlorophyll production only
11 at the higher concentration (Komarek et al.. 2009). Given the low solubility of smelter
12 ash, these observations are consistent with solubility being a key determinant of
13 bioavailability. Similarly, wheat seedling growth was unaffected when exposed to soil
14 leachate containing up to 0.7 mg Pb/L for six weeks. Lettuce seedling root growth was
15 negatively correlated to leachate Pb concentration, but this correlation was only
16 significant for week 3 and week 6 measurements. Authors concluded that although the
17 total concentrations of multiple metals in tested soils and leachates exceeded Canadian
18 Environmental Quality Guidelines, no toxic or only slightly toxic effects occurred
19 following exposure to the metal mixture (Chapman et al.. 2010).
20 Chinese cabbage (Brassica pekinensis) exposed to Pb-containing soils exhibited
21 depressed nitrogen assimilation as measured by shoot nitrite content, nitrate reductase
22 activity, and free amino acid concentration (Xiong et al.. 2006). The authors planted
23 germinated cabbage seeds in soils spiked with Pb acetate to give final soil concentrations
24 of 0.2, 4, and 8 mM Pb/kg dry weight total Pb (41.4, 828.8 and 1,657.6 mg Pb/kg ) and
25 collected leaf samples for 11 days. At exposures of 4 and 8 mM Pb/kg (828.8 and 1,657.6
26 mg Pb/kg), leaf nitrite content was decreased by 29% and 20%, while nitrate content was
27 affected only at the highest Pb exposure (70% of control levels). Free amino acid content
28 in exposed plants was 81% and 82% of control levels, respectively. B. pekinensis shoot
29 biomass was observed to decrease with increasing Pb exposures, with biomass at the two
30 highest Pb exposures representing 91% and 84% of control growth, respectively.
31 Nitrogen, potassium, and phosphorus concentrations in the shoot and root tissues of four
32 canola cultivars (Brassica napus) also decreased as spiked soil Pb concentrations
33 increased from 0 to 90 mg/kg. At the highest soil Pb concentration, nitrogen
34 concentrations were reduced 56% in roots and 58% in shoots versus control levels, while
35 phosphorous concentrations were reduced 37% and 45%, respectively, and potassium
36 content decreased by 42% in both tissues (Ashraf et al.. 2011). Cultivation in Pb-spiked
37 soils was also linked to decreased shoot and root biomass (32% and 62%, respectively at
38 90 mg Pb/kg).
November 2012 7-47 Draft - Do Not Cite or Quote
-------
Genetic and Reproductive Effects
1 Exposure to Pb also resulted in genetic abnormalities, including bridges, condensed
2 bivalents, and laggards, in the meiotic cells of pea plants (Lathyrus sativus) (Kumar and
3 Tripathi, 2008). Seeds were germinated in soils amended with Pb nitrate at concentrations
4 of 25, 50, 100, 200, and 300 mg Pb/kg, and concentrations of 100 mg Pb/kg and greater
5 were found to be genotoxic or detrimental to pea viability. Cenkci et al. (2010) exposed
6 fodder turnip (B. rapa) to 0.5 to 5 mM of Pb nitrate (103.6 to 1036 mg Pb/L) for 6 days
7 and showed decreased genetic template stability (as quantified by random amplified
8 polymorphic DNA profiles) and decreased photosynthetic pigments.
9 Two genotypes of maize seedlings exhibited a significant and concentration-dependent
10 reduction in seed germination following 7 days of Pb treatment in nutrient solution of
11 0.01, 0.1 and 1 mg Pb/L as Pb sulfate (Ahmad et al.. 2011). Pb exposure also decreased
12 germination rate and growth, and increased pollen sterility in radish grown for 50 days in
13 hydroponic solutions containing 0.5 mM Pb (104 mg Pb/L) (Kumar and Tripathi. 2008).
14 Plants exposed to Pb exhibited decreased growth, curling and chlorosis of young leaves,
15 and decreased root growth. In addition, Gopal and Rizvi (2008) showed that Pb exposure
16 increased uptake of phosphorus and iron and decreased sulfur concentration in radish
17 tops.
18 Interestingly, as in zebra finch (Section 7.3.3.3) Ca2+ was found to moderate the effects of
19 Pb in both monocotyledon and dicotyledon plant seedlings, with tomato (Lycopersicon
20 esculentum), rye (Lolium sp.), mustard, and maize plants exhibiting increased tolerance
21 to Pb exposures of 5, 10, and 20 mg Pb/L in the presence of Ca2+ concentration of 1.2
22 mM (249 mg Pb/L) and higher (Antosiewicz. 2005).
7.3.4.2 Terrestrial Invertebrates
23 Exposure to Pb also causes antioxidant effects, reductions in survival and growth, as well
24 as decreased fecundity in terrestrial invertebrates as summarized in the 2006 Pb AQCD
25 (U.S. EPA. 2006b). Alterations in reproduction, growth and survival at the species level
26 can lead to effects at the population-level of biological organization and higher
27 (Section 7.1.1). In addition to these endpoints, recent literature also indicates that Pb
28 exposure can cause significant neurobehavioral aberrations, and in some cases,
29 endocrine-level impacts. Second-generation effects have been observed in some
30 invertebrate species.
31 The morphology of y-aminobutyric acid (GABA) motor neurons in Caenorhabditis
32 elegans nematodes was affected following exposure to Pb nitrate for 24 hours (Du and
November 2012 7-48 Draft - Do Not Cite or Quote
-------
1 Wang. 2009). The authors determined that exposures as low as 2.5 (iM Pb nitrate (0.5 mg
2 Pb/L) could cause moderate axonal discontinuities, and observed a significant increase in
3 the number of formed gaps and ventral cord gaps at Pb nitrate exposures of 75 and
4 200 (iM (6 and 41 mg Pb/L). Younger C. elegans larvae were more likely to exhibit
5 neurobehavioral toxicity symptoms in response to Pb exposure at 2.5 (iM (0.5 mg Pb/L)
6 (Xing et al.. 2009b). Neural degeneration, as demonstrated by dorsal and ventral cord
7 gaps and neuronal loss was also more pronounced in young larval C. elegans than in
8 older larvae and adults (Xing et al.. 2009c). C. elegans nematodes exposed to Pb
9 concentration as low as 2.5 (iM (0.5 mg Pb/L) for 24 hours also exhibited significantly
10 altered behavior characterized by decreased head thrashes and body bends. Exposures of
11 50 (iM Pb (10 mg Pb/L) and greater decreased the number of nematode forward turns
12 (Wang and Xing. 2008). Chemotaxis toward NaCl, cAMP, and biotin was also decreased
13 in C. elegans nematodes exposed to Pb concentration greater than 2.5 (iM (0.5 mg Pb/L)
14 (Xing et al., 2009a). This evidence suggests that Pb may exert neurotoxic action in
15 invertebrates as it does in vertebrates. However, it is unclear how these behavioral
16 aberrations would affect fitness or survival (Wang and Xing. 2008).
17 In a study of C. elegans exposed to 4 sub-lethal concentrations of Pb nitrate between 25
18 and 100 (iM (5 and 21 mg Pb/L), Vigneshkumar et al. (In Press) observed upregulation of
19 both catalase and antimicrobial response-related genes. When challenged with addition of
20 a pathogenic strain ofPseudomonas aeruginosa, exposed C. elegans showed greater
21 resistance to microbial colonization than controls.
22 Younger individuals also appear to be more sensitive to the reproductive effects of Pb
23 exposure. Guo et al. (2009) showed that concentrations of 2.5, 50, and 100 (iM Pb (0.5,
24 10, and 21 mg Pb/L) had greater significant adverse effects on reproductive output when
25 early-stage larval C. elegans were exposed. Adult C. elegans exhibited decreased brood
26 size only when exposed to the highest Pb concentration.
27 The progeny of C. elegans nematodes exposed nominally to 2.5, 75, and 200
28 Pb nitrate (0.5, 16, and 41 mg Pb/L) exhibited significant indications of multi-
29 generational toxicity (Wang and Peng. 2007). Life spans of offspring were decreased by
30 increasing parental Pb exposure, and were comparable to the reductions in parental life-
31 spans. Similarly, diminished fecundity was observed in the progeny of C elegans
32 exposed to Pb (9%, 19%, and 31% reductions of control fecundity, respectively),
33 although the decrease was smaller than in the exposed parental generation (reductions of
34 52%, 58%, and 65%, respectively). Significant behavioral defects affecting locomotion
35 were also observed in the offspring, but these impacts were not determined to be
36 concentration-dependent. Reproductive effects of Pb exposure were also observed in
37 springtails F. Candida following 10-day exposure to Pb-spiked soils. Egg hatch
November 2012 7-49 Draft - Do Not Cite or Quote
-------
1 significantly decreased at nominal concentrations of 1,600 mg Pb/kg dry soil and higher
2 and the EC50 for hatching was 2,361 mg Pb/kg dry soils (Xu et al. 2009b).
3 E. andrei earthworms exposed to 21 different soils, each containing 2,000 mg Pb/kg
4 freshly added Pb, for 28 days exhibited highly variable mortality, ranging from 0% to
5 100%, (Bradham et al.. 2006). Pb body burden of exposed worms ranged from 29 to
6 782 mg Pb/kg. Internal Pb concentration was also negatively correlated to reproductive
7 output. CEC and pH were found to be the principal soil characteristics determining the
8 differences in those effects, although the apparent role of CEC may only have been due to
9 its correlation with other soil characteristics. Low soil Pb concentration (5 mg Pb/kg) also
10 decreased the protein content of E. fetida earthworms during a 7-day exposure (Li et al..
11 2009b). Higher Pb concentration had no effect on protein production. However, cellulase
12 activity was increased by the 7-day exposures to Pb at all exposure concentrations (31%,
13 13%, and 23% of control activity at exposures of 5, 50, and 500 mg Pb/kg, respectively),
14 which the authors reported as an indication of detrimental effects on worm metabolism.
15 By contrast, Svendsen et al. (2007) found thatZ. rubellus earthworms exposed for 42
16 days to field-collected smelter-polluted soils containing average Pb concentration of 106,
17 309, and 514 mg Pb/kg dry weight exhibited normal survival and cocoon production
18 rates, even though they accumulated more Pb with increased environmental
19 concentration. The much smaller effect may be explained by the increased aging time
20 undergone by field soil, causing a larger fraction of the total Pb to be complexed and
21 sequestered by organic and inorganic compounds. Similarly, earthworms (E. fetida)
22 exposed to field-collected soils with concentrations of Pb and As up to 390 mg/kg and
23 128 mg/kg, respectively, due to historical treatments of Pb-arsenate pesticides, exhibited
24 no change in survival, behavior or morphology (Delistraty and Yokel. In Press). Soil
25 aging (e.g., from of the time of Pb-arsenate applications in 1942 to soil collection in
26 approximately 2009) likely reduced Pb bioavailability to earthworms.
27 As in plants, induction of metal chelating proteins and antioxidant activity in
28 invertebrates is affected by exposure to Pb. Metallothionein production in earthworms
29 (Lampito mauritif) was significantly induced following exposure to Pb-contaminated soil.
30 Tissue metallothionein levels increased after a two-week exposure to 75 to 300 mg Pb/kg
31 soil, although by 28 days levels had begun to decrease, perhaps as a result of Pb toxicity
32 (Maity et al., 2011). Further, the induction of antioxidant activity was correlated to
33 standard toxicity measurements in Thebapisana snails (Radwan et al.. 2010). Topical
34 application of Pb solutions (estimated to be 500 to 2,000 (ig Pb per animal) to snails
35 resulted in decreased survival, increased catalase and glutathione peroxidase activities,
36 and decreased glutathione concentration. The 48-hour LD50 concentration was
37 determined to be 653 (ig per snail, as measured in digestive gland tissue. Snail
38 glutathione content was decreased at exposures of 72.2% of the 48-hour LD50 value,
November 2012 7-50 Draft - Do Not Cite or Quote
-------
1 while Pb exposure at 40% of the 48-hour LD50 value induced catalase and glutathione
2 peroxidase activities.
3 Dietary exposure to Pb also affected T. pisana snail growth. After three weeks on
4 Pb-contaminated diet, snail feeding rates were depressed by all Pb exposures (nominal
5 concentration of 50 to 15,000 mg Pb/kg diet dry weight) (El-Gendv et al.. 2011). A five
6 week dietary exposure to 1,000 mg Pb/kg and greater resulted in reduced snail growth.
7 Food consumption, growth, and shell thickness were also observed to decrease with
8 increasing diet Pb in juvenile A. achatina snails (7 levels between 0 and 1,344 mg Pb/kg
9 feed, for 12 weeks) (Ebenso and Ologhobo. 2009a). A similar depression of growth was
10 observed in sentinel juvenile A. achatina snails deployed at Pb-polluted sites in the Niger
11 Delta region of Nigeria. Although snail mortality was not increased significantly by
12 exposure to soil Pb up to 1,200 mg Pb/kg, a concentration-dependent relationship was
13 established for growth, with significant reduction observed at 12-week exposures to
14 20 mg Pb/kg (Ebenso and Ologhobo. 2009b). However, consumption of field-collected
15 Pb-polluted U. dioica leaves containing 3 mg Pb/kg stopped all reproductive output in
16 C. nemoralis. Snails also exhibited diminished food consumption rates when offered
17 leaves with both low (1.5 mg Pb/kg) and high Pb content, but the mechanism of the
18 dietary aversion was not defined (Notten et al.. 2006).
19 Chronic dietary exposure to Pb was also examined in post-embryonic oribatid mites
20 (Archegozetes longisetosus) (Kohler et al., 2005). Both algae and bark samples were
21 soaked in 100 mg/L Pb as Pb nitrate and provided as diet and substrate, respectively, to
22 larval mites. In addition to elevated heat shock proteins (hsp70), 90.8% of the
23 protonymphs exhibited significant leg deformities, including abnormal claws, shortened
24 and thickened legs, and translocated setae. Although not specifically discussed, it is very
25 likely that these deformities would decrease mite mobility, prey capture, and reproductive
26 viability. While there is some evidence that oribatid mites exhibit Pb avoidance behavior,
27 this response may not significantly reduce Pb exposure and effects. Although soil-
28 inhabiting mites (Oppia nitens) were observed to avoid high Pb concentrations, the EC50
29 for this behavior was approximately five times higher than the chronic EC50 for
30 reproduction (8,317 and 1,678 mg Pb/kg, respectively) (Owojori et al., 2011).
31 Consequently, it is unlikely that oribatid mites will avoid soils containing toxic Pb
32 concentrations.
33 Lock et al. (2006) compared the toxicity of both laboratory-spiked soils and field-
34 collected Pb-contaminated soils to springtails (F. Candida). The 28-day EC50 values
35 derived for F. Candida ranged from 2,060 to 3,210 mg Pb/kg in leached and unleached
36 Pb-spiked soils, respectively, whereas field-collected soils had no significant effect on
37 springtail reproduction up to (but not including) 14,436 mg Pb/kg (Lock et al., 2006).
November 2012 7-51 Draft - Do Not Cite or Quote
-------
1 Consequently, leaching soils prior to use in bioassays had only a slight effect on Pb
2 toxicity to resident springtails, and did not provide an appropriate model for field-
3 weathered, Pb-contaminated soils. This indicates that physiochemical factors other than
4 leaching may be more important determinants of Pb bioavailability. A 4-week exposure
5 to Pb-amended soils containing up to 3,200 mg Pb/kg (nominal concentration) had no
6 significant effect on Sinella curviseta springtail survival or reproduction (Xu et al..
7 2009a).
8 Carabid beetles (Pterostichus oblongopunctatus) inhabiting soils contaminated by
9 pollution from a Pb-Zn smelter (containing 136 to 2,635 mg Pb/kg) were field-collected
10 and then laboratory-reared for two generations (Lagisz and Laskowski. 2008). While
11 fecundity was positively correlated to soil metal concentration (e.g., more eggs were
12 produced by females collected from contaminated areas), the hatching rate of eggs
13 diminished with increasing soil metal contamination. For the Fl generation, females
14 produced by parents inhabiting highly polluted areas exhibited decreased body mass. The
15 authors stated that these results indicate that invertebrates inhabiting metal- (or Pb-)
16 contaminated soils could face "significantly altered life-history parameters." Similarly,
17 aphids (Brevicoryne brassicae) reared on cabbage and radish plants exposed to 0.068 mg
18 Pb daily exhibited altered development and reproduction when compared to those reared
19 on non-exposed plants. Development time was increased by approximately two days,
20 which led to a reduction in relative fecundity (Gorur. 2007). Although the authors noted
21 that study exposures were greater than what would be expected in naturally polluted
22 areas, Pb exposure under field conditions could alter invertebrate life history patterns.
23 Several studies suggest that Pb may disrupt hormonal homeostasis in invertebrates. Shu
24 et al. (2009) reported that vitellogenin production in both male and female S. litura moths
25 was disrupted following chronic dietary exposure to Pb. Adult females reared on diets
26 containing 25, 50, 100, or 200 mg Pb/kg exhibited decreased vitellogenin mRNA
27 induction, and vitellogenin levels decreased with increasing Pb exposure. In addition,
28 vitellogenin mRNA induction was detected in males exposed to 12 and 25 mg Pb/kg, and
29 low levels of vitellogenin were found at those lower Pb exposures, when males normally
30 do not produce any. In the Asian earthworm (Pheretima guillelmi), sperm morphology
31 was found to be altered significantly following 2-week exposure to soils containing
32 nominal concentration of 1,000, 1,400, 1,800, and 2,500 mg Pb/kg (Zheng and Li. 2009).
33 Common deformities were swollen head and head helices, while head bending was also
34 recorded in some cases. These deformities were observed following exposures to
35 concentration below the 14-day LC50 (3,207 mg Pb/kg) and below the concentration at
36 which weight was diminished (2,800 mg Pb/kg). Experimentation with the model
37 organism Drosophila indicates that Pb exposure may increase time to pupation and
November 2012 7-52 Draft - Do Not Cite or Quote
-------
1 decrease pre-adult development, both of which are endocrine-regulated (Hirsch et al.,
2 2010).
7.3.4.3 Terrestrial Vertebrates
3 Pb poisoning is one of the earliest recognized toxicoses of terrestrial vertebrates,
4 occurring primarily through the ingestion of spent shot by birds. While the focus of the
5 ISA is on more environmentally relevant exposures, studies of Pb poisoning provide
6 historical context for the review. The widespread nature of this toxicosis was first noticed
7 in American waterfowl around the turn of the last century (see (Jones. 1939) for an
8 historical summary). Wetmore (1919) demonstrated that Pb shot caused the observed
9 effects and described in detail the species affected, associated symptoms, and additional
10 factors involved. By 1959, the estimated annual loss of waterfowl to Pb poisoning was
11 2-3 percent of the fall population (Bellrose. 1959). Smaller numbers of shorebirds and
12 upland game birds were also found poisoned by Pb (Locke and Thomas. 1996).
13 The first reported Pb poisoning of a bald eagle (Haliaeetus leucocephalus) was described
14 by Mulhern et al. (1970). and subsequently several hundred bald eagle Pb poisonings
15 were diagnosed throughout the U.S. prior to the ban on use of Pb shot for waterfowl
16 hunting (Kramer and Redig, 1997). Eagles and other raptors are poisoned by consuming
17 Pb pellets imbedded in the flesh of ducks or upland prey species and may also be exposed
18 to other sources of Pb, such as fishing sinkers and weights (Kramer and Redig. 1997).
19 The use of Pb shot for waterfowl hunting was banned in 1991 due to the poisoning of
20 bald eagles, which had been previously added to the endangered species list and were
21 specially protected under the Bald Eagle Protection Act of 1940.
22 Anderson et al. (2000) reported that by 1997, mallard (Anas platyrhynchus) deaths from
23 Pb poisoning in the Mississippi flyway were reduced by 64 percent, and ingestion of
24 toxic pellets had declined by 78 percent. They estimated the ban prevented approximately
25 1.4 million duck deaths in the first 6-year period. However, Pb exposure remains
26 widespread in bald eagles, although blood Pb concentrations have significantly decreased
27 (Kramer and Redig. 1997). The endangered California condor (Gymnogyps
28 californianus) also continues to have significantly elevated blood Pb levels as well as
29 Pb-associated mortality resulting from exposure to ammunition fragments contained in
30 food items (Cade. 2007; Church et al.. 2006). Although there is a significant amount of
31 information on Pb tissue residues of mammals, there are very few reports of Pb
32 poisoning; exceptions are reports of Pb poisoned bats in a cave in the southern U.S. and
33 small mammals in the vicinity of several smelters (Shore and Rattner. 2001).
November 2012 7-53 Draft - Do Not Cite or Quote
-------
1 At the time of the 1977 Pb AQCD few studies of the effects of exposure to Pb had been
2 conducted in wild animals other than birds, and the majority of those studies were of
3 direct poisoning (U.S. EPA. 1977). Several studies of domestic animals grazing near Pb
4 smelters indicated that horses are more susceptible than cattle to chronic Pb exposure
5 although the findings were not conclusive due to the presence of other metals. Delta-
6 aminolevulinic acid dehydratase (ALAD) was recognized as a sensitive indicator of Pb
7 exposure in rats and waterfowl. In the 1986 Pb AQCD, additional effects of Pb on small
8 mammals and birds were reported. According to the 2006 Pb AQCD (U.S. EPA. 2006b).
9 commonly observed effects of Pb on avian and mammalian wildlife include decreased
10 survival, reproduction, and growth, as well as effects on development and behavior. More
11 recent experimental data presented here expand and support these conclusions, and also
12 indicate that Pb can exert other effects on exposed terrestrial vertebrates, including
13 alteration of hormones and other biochemical variables.
14 Since the 2006 Pb AQCD, there is additional evidence for hematological effects of Pb
15 exposure in terrestrial vertebrates. Red-backed salamanders (Plethodon cinereus)
16 exposed to Pb-amended soils (553, 1,700, 4,700, and 9,167 mg Pb/kg) by Bazar et al.
17 (2010) exhibited lowered appetite and decreased white blood cell counts at the two
18 highest concentrations, but tolerated field-collected, aged soils containing Pb
19 concentrations of up to 16,967 mg Pb/kg with no significant deleterious effects. The
20 white blood cell count of adult South American toads, (Bufo arenarum) was also
21 decreased by weekly sublethal i.p. injections of Pb acetate at 50 mg Pb/kg body weight,
22 (Chiesa et al.. 2006). The toads also showed altered serum profiles and increased number
23 of circulating blast cells. Final toad blood Pb levels were determined to be 8.6 mg Pb/dL,
24 although it is unclear whether this is representative of Pb concentrations observed in field
25 B. arenarum populations exposed to Pb. The authors suggested that, based on these
26 findings, long-term environmental exposure to Pb could affect toad immune response. In
27 western fence lizards (S. occidentalis), sub-chronic (60-day) dietary exposure to 10 to
28 20 mg Pb/kg per day resulted in significant sublethal effects, including decreased cricket
29 consumption, decreased testis weight, decreased body fat, and abnormal posturing and
30 coloration (Salice et al.. 2009). Long-term dietary Pb exposures are thus likely to
31 decrease lizard fitness.
32 Even in cases of high environmental Pb exposures, however, linking Pb body burdens to
33 biological effects can be difficult. Pb concentration in the breast feathers, washed tail
34 feathers, and blood of field-collected blackbirds (Turdus meruld) were determined to be
35 3.2 mg Pb/kg, 4.9 mg Pb/kg, and 0.2 mg Pb/kg wet mass in urban birds, as opposed to
36 1.4 mg Pb/kg, 1 mg Pb/kg, and 0.05 mg Pb/kg in rural birds (Scheifler et al.. 2006a).
37 However, the elevated Pb tissue concentrations in urban birds were not significantly
38 correlated to any index of body condition.
November 2012 7-54 Draft - Do Not Cite or Quote
-------
1 The long-term effect of atmospheric Pb deposition on pied flycatcher (Ficedula
1 hypoleuca) nestlings was determined in native communities residing in the Laisvall
3 mining region of Sweden (Berglund et al.. 2010). Moss samples indicated that Pb
4 deposition in study areas ranged between 100 and 2,000 mg Pb/kg dry weight during
5 operations and 200 and 750 mg Pb/kg when operations ceased. A simultaneous slight
6 reduction was observed in pied flycatcher blood Pb levels, from 0.4 to 0.3 mg Pb/kg.
7 However, clutch size was decreased in pied flycatchers inhabiting the mining area both
8 during and after mining operations, and mean nestling mortality was 2.5 times higher in
9 the mining region than in reference areas during mining operations, and 1.7 higher five
10 years after cessation of mining operations. The authors noted that Pb deposition in the
11 mining region remained elevated even after mining operations ceased, and that stable Pb
12 isotope analysis suggested that smelter Pb remained available to pied flycatcher through
13 the transfer of historically deposited Pb in soil to prey items.
14 Berglund et al. (2010) also analyzed ALAD activity in pied flycatchers at the later period,
15 and found that it was 46% lower at the mine site. Beyer et al. (2004) observed that
16 elevated blood Pb levels in several types of birds inhabiting the Tri-State Mining District
17 (Oklahoma, Kansas, Missouri) were correlated with decreases in ALAD activity. Based
18 on reduction in ALAD activity, robins (Turdus migratorius) were most sensitive to Pb
19 exposure (35% reduction), followed by cardinals (Cardinalis cardinalis), waterfowl, and
20 bobwhite quail (Colinus virginianus) (40%, 41%, and 56% reductions, respectively).
21 Eagle owl (Bubo bubo) nestlings living in a historical mining area in Spain also exhibited
22 elevated blood Pb levels (average 8.61 (ig/dL as compared to an average reference area
23 value of 3.18 (ig/dL), and this was correlated to an approximate 60% reduction in ALAD
24 activity (Gomez-Ramirez et al.. 2011). Hansen et al. (2011 a) determined that ground-
25 feeding songbirds were frequently exposed to Pb within the Coeur d'Alene, ID mining
26 region. Robins, in particular, were significantly likely to exhibit blood Pb levels in the
27 clinical and severe clinical poisoning ranges (50 to 100 (ig/dL and >100 (ig/dL,
28 respectively). Ingested soil Pb accounted for almost all of the songbirds' exposure to Pb,
29 with Pb exposure correlated with estimated soil ingestion rates (20% for robins, 17% for
30 song sparrows, and 0.7% for Swainson's thrushes, Catharus ustulatus). More than half of
31 the robins and song sparrows from all contaminated sites and more than half of the
32 Swainson's thrushes from highly contaminated sites showed at least 50% inhibition of
33 ALAD. The highest hepatic Pb concentration of 61 mg/kg (dry weight) was detected in a
34 song sparrow (Hansen et al.. 201 la).
35 Blood Pb was significantly elevated in waterfowl in the Lake Coeur d'Alene areas of
36 Blackwell Island and Harrison Slough (mean sediment concentrations of 679 and
37 3,507 mg Pb/kg dry weight, respectively). Twenty-seven percent of the waterfowl
38 sampled in the Blackwell Island region had blood Pb concentrations suggestive of severe
November 2012 7-55 Draft - Do Not Cite or Quote
-------
1 clinical poisoning (average concentration=0.17 mg Pb/kg); in the Harrison Slough, 60%
2 of sampled waterfowl had highly elevated blood Pb levels that exceeded the severe
3 clinical poisoning threshold (average concentration=2,2 mg Pb/kg) (Spears et al., 2007).
4 The level of corticosteroid hormones in field populations of white stork nestlings
5 (Ciconia ciconid) in a mining area affected by Pb and other metals was positively
6 correlated with blood Pb levels (Baos et al.. 2006). The effect was more pronounced for
7 single nestlings than for multiple-chick broods. Surprisingly, average blood Pb levels in
8 chicks inhabiting reference areas was 910 (ig Pb/dL (± 51), which was higherthan blood
9 Pb levels from the mining area (440 ± 340 (ig Pb/dL). However, the correlation between
10 blood Pb levels and the corticosteroid stress response in white stork nestlings was
11 observed in both groups of birds. Burger and Gochfeld (2005) exposed herring gull
12 (Lams argentatus) chicks to Pb acetate via an i.p. injection of 100 mg Pb/kg body
13 weight, to produce feather Pb concentration approximately equivalent to those observed
14 in wild gulls. Pb-exposed gulls exhibited abnormal behaviors, including decreased
15 walking and food begging, erratic behavioral thermoregulation, and diminished
16 recognition of caretakers. Interestingly, subchronic exposure of Japanese quail (Coturnix
17 coturnixjaponica) to 5 and 50 mg Pb/L in drinking water caused an increase in their
18 immune response. Exposed quail exhibited significantly lower rates of death or health
19 effects (including septicemia, perihepatitis, and pericarditis among others) than control
20 animals following infection with Escherichia coli, and the incidence of infection-related
21 effects was dependent on Pb exposure (Nain and Smits. 2011). These observations
22 contrast with immunotoxicology results in mice reported in Section 5.6.5.1.
23 Again, dietary or other health deficiencies unrelated to Pb exposure are likely to
24 exacerbate the effects of Pb. Ca2+-deficient female zebra finches (T. guttatd) had a
25 suppressed secondary humoral immune response following 28-day exposures to 20 mg
26 Pb/L in drinking water (Snoeijs et al.. 2005). This response, however, was not observed
27 in birds fed sufficient Ca2+. Although a significant finding, these data are difficult to
28 interpret under field conditions where the overall health of avian wildlife may not be
29 easily determined.
30 Chronic Pb exposures were also demonstrated to affect several mammalian species.
31 Young adult rats reared on a diet containing 1,500 mg Pb/kg Pb acetate for 50 days
32 demonstrated less plasticity in learning than non-exposed rats (McGlothan et al.. 2008).
33 indicating that Pb exposure caused significant alteration in neurological function. Yu et
34 al. (2005) showed that dietary Pb exposure affected both the growth and endocrine
35 function of gilts (S. domestica). Consumption of 10 mg Pb/kg diet resulted in lower body
36 weight and food intake after 120 days of dietary exposure; Pb exposure decreased final
37 weight by 8.2%, and average daily food intake of Pb-exposed pigs was decreased by
38 6.8% compared to control intake. Additionally, concentration of estradiol, luteinizing
November 2012 7-56 Draft - Do Not Cite or Quote
-------
1 hormone, and pituitary growth hormone were decreased (by 12%, 14%, and 27% versus
2 controls, respectively), while blood Pb level was increased by 44% to an average
3 2.1 (ig/dL. In cattle grazing near Pb-Zn smelters in India, blood Pb levels were positively
4 correlated with plasma levels of the thyroid hormones thyroxine (T4) and tri-
5 iodothyronine (T3) and the hepatic biomarkers alanine transaminase and aspartate
6 transaminase (Swarup et al.. 2007). Total lipids, total protein and albumin levels were
7 decreased in the same animals. Rodriguez-Estival et al. (2011) determined that red deer
8 (Cervus elaphus) and wild boar (Sus scrofd) inhabiting a Pb-contaminated mining area in
9 Spain exhibited increased liver and bone Pb concentrations (geometric means of 0.35 and
10 0.46 mg Pb/kg for red deer, and 0.81 and 7.36 mg Pb/kg for wild boar, respectively).
11 These tissue concentrations were correlated to a significant decrease in red deer
12 glutathione production, but corresponded to an increase in wild boar glutathione
13 (Rodriguez-Estival et al.. 2011). Authors proposed that the different antioxidant
14 responses may be indicative of different Pb susceptibilities in the two species.
15 Following previous reports of in vivo follicle and oocyte damage in animals with
16 low-level Pb accumulation, Nandi et al. (2010) treated oocytes of buffalo (Bubalus
17 bubalis) in vitro with Pb at concentrations ranging from 0.005 to 10 mg/L in one-day
18 cultures indicated a significant decline in viability of oocytes at 1 mg/L. Dose-dependent
19 effects on oocyte viability, morphological abnormalities, cleavage, blastocyst yield and
20 blastocyst hatching were observed in Pb-treated oocytes with maturation significantly
21 reduced at 2.5 mg/L and 100% oocyte death at 32 mg/L. These results appear to confirm
22 previous reports, but the in vitro concentrations of Pb are difficult to relate to in vivo
23 exposures. On the other hand, the reproductive viability of wild red deer from the
24 Pb-contaminated mining area of Spain studied by Rodriguez-Estival et al. (2011) was
25 shown to be altered, with 11% and 15% reductions in spermatozoa and acrosome
26 integrity observed in male deer from the mining area compared with those residing in
27 reference areas (Reglero et al., 2009a).
7.3.5 Exposure and Response of Terrestrial Species
28 Evidence regarding exposure-response relationships and potential thresholds for Pb
29 effects on terrestrial populations can inform determination of standard levels that are
30 protective of terrestrial ecosystems. Given that exposure to Pb may affect plants,
31 invertebrates and vertebrates at the organism, population, or community level,
32 determining the rate and concentration at which these effects occur is essential in
33 predicting the overall risk to terrestrial organisms. This section updates available
34 information derived since the 2006 Pb AQCD, summarizing several dose-response
35 studies with soil invertebrates. As shown in the studies summarized in Table 7-4. several
November 2012 7-57 Draft - Do Not Cite or Quote
-------
1 experiments have been published that used multiple levels of Pb under controlled
2 conditions. However, none of them treated Pb concentration as a continuous variable,
3 i.e., none attempted to analyze results as a concentration-response relationship. In
4 addition, given the well-established presence of strong interactions with variables such as
5 pH, CEC, OC, or aging, applying exposure-response relationships from those
6 experiments to natural conditions with different values of those interacting variables
7 could be difficult.
8 Dose-dependent responses in antioxidant enzymes were observed in adult L. mauritii
9 earthworms exposed to soil-associated Pb contamination (75, 150, 300 mg Pb/kg) (Maitv
10 et al.. 2008). By day seven of exposure, glutathione-S-transferase activity and glutathione
11 disulfide concentration were positively correlated with increasing Pb exposures, while
12 glutathione concentration exhibited a negative dose-response relationship with soil Pb
13 concentration. However, these trends had become insignificant by the end of the total
14 exposure period (28 days), as a result of normalization of antioxidant systems following
15 chronic exposure. This strongly suggests that changes to earthworm antioxidant activity
16 are an adaptive response to Pb exposures.
17 Both survival and reproductive success of E. fetida earthworms showed concentration-
18 dependent relationships with soil Pb concentration during the course of standard 14- and
19 56-day toxicity tests (Jones et al.. 2009b). Five levels of Pb soil concentration were
20 prepared for the acute 14-day study via spiking with Pb nitrate—0, 300, 711, 1,687, and
21 2,249 mg Pb/kg, while soil concentration of 0, 355, 593, 989, and 1,650 mg Pb/kg were
22 used in chronic (56-day) earthworm bioassays. A 14-day acute LC50 of 2,490 mg Pb/kg
23 was determined from the dose-response relationship, while the approximate 56-day
24 NOEC (no observed effect concentration) and EC50 values were about 400 mg Pb/kg and
25 1,000 mg Pb/kg, respectively. Jones et al. (2009b)made use of continuous (regressional)
26 models to characterize the relationship between Pb soil concentration and Pb
27 accumulation in earthworms, but did not use continuous models for the relationship of
28 exposure and other responses. Currie et al. (2005) observed mortality of E. fetida after 7
29 and 14 days in spiked field soil at seven levels of Pb (0 to 10,000 mg Pb/kg). They
30 reported LC50 values of 2,662 mg Pb/kg at 7 days and 2,589 mg Pb/kg at 14 days or
31 2,827 mg Pb/kg at both 7 and 14 days, depending on the number of worms in the
32 experimental enclosure.
33 Other studies have shown no correlation between Pb concentration in either earthworm
34 tissue or soil, and earthworm survival rate. Although the Pb content of E. fetida held in
35 metal-contaminated soils containing between 9.7 and 8,600 mg Pb/kg was positively
36 correlated with Pb concentration of fully aged soil collected from disused mines, there
37 was no statistical relationship with earthworm survival during a 42-day exposure period
November 2012 7-58 Draft - Do Not Cite or Quote
-------
1 (Nahmani et al., 2007). However, Pb concentration in soil leachate solution was
2 significantly correlated with decreased earthworm survival and growth (linear regression:
3 R2= 0.64, p<0.0001). The 42-day Pb EC50 for E. fetida growth was 6,670 mg Pb/kg.
4 Langdon et al. (2005) exposed three earthworm species (E. andrei, L. rubellus, and
5 A. caliginosd) to Pb nitrate-amended soils at concentrations of 1,000 to 10,000 mg Pb/kg
6 to determine species variability in uptake and sensitivity. Twenty-eight-day LC50 values
7 for the three species were 5,824 mg Pb/kg, 2,867 mg Pb/kg, and 2,747 mg Pb/kg,
8 respectively, indicating thatZ. rubellus and A. caliginosa are significantly more
9 vulnerable to Pb contamination than E. andrei, a common laboratory species. This is
10 comparable to previous findings by Spurgeon et al. (1994) who reported 14-day LC50 of
11 4,480 mg Pb/kg and 50-day LC50 of 3,760 mg Pb/kg for E. fetida, another standard
12 laboratory test species. In the more recent study of E. fetida sensitivity summarized
13 above, Jones (2009b) reported LC50 values for E. fetida that are similar to those for
14 L. rubellus and A. caliginosa. It is likely that these apparent species differences are a
15 result of differential bioavailability of the Pb in test soils. However, the Pb body burden
16 of all three species in the study by Langdon et al. (2005) increased with increasing
17 environmental concentration, and there were no species differences in Pb tissue content.
18 When given a choice between treated and untreated soils, all worm species exhibited
19 significant avoidance of Pb-contaminated soils, and altering pH (and, consequently, Pb
20 bioavailability) had no impact on avoidance (Langdon et al.. 2005). Field earthworms
21 may thus be able to reduce their exposure to Pb through behavior.
22 Reproductive success of other soil invertebrates is impacted by Pb. The organismal and
23 population-level responses of the springtail Paronychiurus kimi to Pb were determined by
24 Son et al. (2007) using artificial soils, following the 1999 ISO methodology. The 7-day
25 Pb LC50 was determined to be 1,322 mg Pb/kg dry weight, while the 28-day reproduction
26 EC50 was established as 428 mg Pb/kg. The intrinsic rate of population increase was
27 lower at a Pb soil concentration of 1,312 mg Pb/kg, and the authors estimated that, at this
28 level, P. kimi populations would be extirpated. The authors noted that, in this case, the
29 reproductive endpoint overestimated the population-level risk for P. kimi springtails
30 exposed to Pb, and proposed that more specific measures of population-level endpoints
31 (such as the reduction in intrinsic rate of increase) be used to determine risk to
32 populations. Menta et al. (2006) showed that a nominal soil concentration of 1,000 mg
33 Pb/kg decreased the reproductive output of two collembolans, Sinella coeca and
34 F. Candida. Pb concentration of 50, 100, and 500 mg Pb/kg slightly but significantly
35 depressed S. coeca adult survival, while F. Candida survival was statistically unaffected
36 by Pb exposure. The hatching success ofF. Candida eggs was diminished by 10-day
37 exposure to Pb-spiked soils; the 10-day EC50 for hatching success was reported as
November 2012 7-59 Draft - Do Not Cite or Quote
-------
1 2,361 mg/kg Pb (Xu et al.. 2009b). However, authors noted that egg development was
2 more sensitive to Cu and Zn exposure, and by comparison, was less susceptible to Pb.
3 In addition to species variability, physical and chemical factors affecting Pb
4 bioavailability were also demonstrated to significantly influence the toxicity of Pb to
5 terrestrial species. As noted previously in Section 7.3.2. laboratory-amended artificial
6 soils provide a poor model for predicting the toxicity of Pb-contaminated field soils,
7 because aging and leaching processes, along with variations in physiochemical properties
8 (pH, CEC, OM), influence metal bioavailability. Consequently, toxicity values derived
9 from exposure-response experimentation with laboratory-spiked soils probably
10 overestimate true environmental risk, with the possible exception of highly acidic sandy
11 soils. Because toxicity is influenced by bioavailability of soil biogeological and chemical
12 characteristics, extrapolation of toxic concentrations between different field-collected
13 soils will be difficult. Models that account for those modifiers of bioavailability, such as
14 the terrestrial BLM proposed by Smolders et al. (2009). have proven difficult to develop
15 due to active physiological properties of soil organisms affecting either uptake (such as
16 root phytochelatins) or sequestration of Pb (such as granule formation in root tissues and
17 earthworms, or substitution of Pb for calcium in bones).
7.3.6 Terrestrial Community and Ecosystem Effects
18 A study reviewed in the 1977 Pb AQCD provided evidence for Pb effects on forest-
19 nutrient cycling and shifts in community composition. Reduced arthropod density,
20 biomass and richness were observed in the vicinity of a smelting complex in southeastern
21 Missouri where Pb, Cd, Zn and Cu were measured in the litter layer and soil (U.S. EPA.
22 1977; Watson et al.. 1976). In the 1986 Pb AQCD it was reported that Pb at
23 environmental concentrations occasionally found near roadsides and smelters (10,000 to
24 40,000 mg Pb/kg dry weight) can eliminate populations of bacteria and fungi on leaf
25 surfaces and in soil. At soil concentrations of 500 to 1,000 mg Pb/kg or higher,
26 populations of plants, microorganisms, and invertebrates may shift toward Pb-tolerant
27 populations of the same or different species (U.S. EPA. 1986b).
28 According to the 2006 Pb AQCD (U.S. EPA. 2006b). natural terrestrial ecosystems near
29 significant Pb stationary sources (such as smelters and mines) exhibited a number of
30 ecosystem-level effects, including decreased species diversity, changes in floral and
31 faunal community composition, and decreasing vigor of terrestrial vegetation. These
32 findings are summarized in Table AX7-2.5.2 of the Annex to the 2006 Pb AQCD (U.S.
33 EPA. 2006c). More recent literature explored the interconnected effects of Pb
November 2012 7-60 Draft - Do Not Cite or Quote
-------
1 contamination on soil bacterial and fungal community structure, earthworms, and plant
2 growth, in addition to impacts on soil microbial community function.
3 Inoculation of maize plants with Glomus intraradices arbuscular mycorrhizal fungi
4 isolates decreased Pb uptake from soil, resulting in lower shoot Pb concentration and
5 increased plant growth and biomass (Sudova and Vosatka. 2007). Similarly, Wong et al.
6 (2007) showed that the presence of arbuscular mycorrhizal fungi improved vetiver grass
7 (Vetiveria zizanioides) growth, and while Pb uptake was stimulated at low soil
8 concentration (10 mg Pb/kg), it was depressed at higher concentration (100 and 1,000 mg
9 Pb/kg). Bojarczuk and Kieliszewska-Rokicka (2010) found that the abundance of
10 ectomycorrhizal fungi was negatively correlated with the concentration of metals,
11 including Pb, in the leaves of silver birch seedlings. Arbuscular mycorrhizal fungi may
12 thus protect plants growing in Pb-contaminated soils. Microbes too may dampen Pb
13 uptake and ameliorate its deleterious effects: biomass of plants grown in metal-
14 contaminated soils (average Pb concentration 24,175 mg Pb/kg dry weight) increased
15 with increasing soil microbial biomass and enzymatic activity (Epelde et al.. 2010).
16 However, above certain Pb concentration, toxic effects on both plants and microbial
17 communities may prevent these ameliorating effects. R.Y. Yang et al. (2008b) found that
18 both the mycorrhizal colonization and the growth ofSolidago canadensis were negatively
19 affected by soil Pb contamination. They suggested that, more generally, Pb-mediated
20 alterations in plant-fungal dynamics may be the cause of ecological instability in
21 terrestrial vegetative communities exposed to metals.
22 The presence of both earthworms and arbuscular mycorrhizal fungi decreased the
23 mobility of Pb in mining soils undergoing phytoremediation (Maet al.. 2006).
24 Inoculation with both earthworms and fungi increased plant growth at sites contaminated
25 with mine tailings compared to that observed at sites with 75% less Pb contamination.
26 Most likely, this was a result of the decrease in bioavailable (DTPA-extractable and
27 ammonium acetate-extractable) Pb to 17% to 25% of levels in areas without the
28 earthworm and arbuscular mycorrhizal fungi amendments. The presence of earthworms
29 in metal-contaminated soils decreased the amount of water-soluble Pb (Sizmur and
30 Hodson. 2008). but despite this decrease, ryegrass accumulated more Pb from
31 earthworm-worked soils than soils without worms present. Sizmur and Hodson
32 speculated that increased root dry biomass may explain the increased uptake of Pb in the
33 presence of earthworms. However, Sizmur et al. (2011) found that the presence of anecic
34 (deep-burrowing) earthworms (L. terrestris) increased soil leachate Pb concentrations by
35 190%. The authors observed that worms promoted a faster breakdown of organic matter,
36 which caused a decrease in soil pH and a concurrent increase in Pb solubility. As a result,
37 ryegrass (L. perenne) accumulated a greater amount of Pb in systems with earthworms
38 (Sizmur et al.. 2011). Further, the presence of earthworms (Lumbricus terrestris) was
November 2012 7-61 Draft - Do Not Cite or Quote
-------
1 found to increase Pb concentrations in both maize and barley, although growth of these
2 species was unaffected (Ruiz et al.. 2011). Authors noted that worm activity increased Pb
3 extraction yields by factors of 4.4 and 7.6, for barley and maize. By contrast,
4 Coeurdassier et al. (2007) found that Pb was higher in earthworm tissue when snails were
5 present, but that snails did not have a higher Pb content when earthworms were present.
6 Microbial communities of industrial soils containing Pb concentrations of 61, 456, 849,
7 1,086, and 1,267 mg Pb/kg dry weight were also improved via revegetation with native
8 plants, as indicated by increased abundances of fungi, actinomycetes, gram-negative
9 bacteria, and protozoa, as well as by enhanced fatty acid concentration (Zhang et al..
10 2006). Increased plant diversity ameliorated the effects of soil Pb contamination (300 and
11 600 mg Pb/kg) on the soil microbial community (Yang et al.. 2007).
12 The effect of Pb on microbial community function has been quantified previously using
13 functional endpoints such as respiration rates, fatty acid production, and soil acid
14 phosphatase and urease activities, which may provide an estimate of ecological impacts
15 separate from microbial diversity and abundance measurements. Most studies of metal-
16 induced changes in microbial communities have been conducted using mixtures of
17 metals. However, Akerblom et al. (2007) tested the effects of six metals (Cr, Zn, Mo, Ni,
18 Cd, and Pb) individually. All tested metals had a similar effect on the species
19 composition of the microbial community. Exposure to a high Pb concentration (52 mg
20 Pb/kg) also negatively affected respiration rates. Total phospholipid fatty acid content
21 was determined to negatively correlate with increasing Pb exposure, indicating alteration
22 of the microbial community. When Yang et al. (2006) compared the microbial properties
23 of metal-contaminated urban soils to those of rural soils, significant differences were
24 detected in basal community respiration rates and microbial abundance. The urban soils
25 studied contained multiple metal contaminants, but microbial biomass was the only
26 measured endpoint to be significantly and negatively correlated to Pb concentration.
27 Similarly, the fungal community in a naturally Pb-enriched forest in Norway exhibited
28 differences in community composition and abundance when compared with other, low Pb
29 sites. The number of colony-forming fungal units was diminished by soil Pb, and was
30 approximately 10 times lower in the highest Pb soil concentration (-4300 mg Pb/kg).
31 Further, only one fungus species was isolated from both high Pb and control soils,
32 indicating highly divergent communities; species diversity was also reduced by high soil
33 Pb concentrations (Baath et al.. 2005). These studies suggest that anthropogenic Pb
34 contamination may affect soil microbial communities, and alter their ecological function.
35 However, (Khan et al.. 2010c) reported that it is possible for indicators of microbial
36 activity to recover after an initial period depression. (Khan et al.. 2010c) found that
37 following a 2-week exposure to three levels of Pb (150, 300, and 500 mg Pb/kg), the
38 number of culturable bacteria at the highest exposure concentration tested was decreased.
November 2012 7-62 Draft - Do Not Cite or Quote
-------
1 Acid phosphatase and urease levels (measures of soil microbial activity) decreased
2 significantly, but they had recovered by the ninth week. Another study (Bamborough and
3 Cummings. 2009) reported that no changes in bacterial and actinobacterial diversity in
4 metallophytic soils containing 909 to 5,280 mg Pb/kg (43 to 147 mg Pb/kg bioavailable
5 Pb (as defined by the study authors)). Soil bacteria community structure and basal
6 respiration rates were examined in natural soils with pH values ranging from 3.7 to 6.8
7 (Lazzaro et al., 2006). Six soil types of differing pH were treated with Pb nitrate
8 concentrations of 0.5, 2, 8, and 32 mM (104, 414, 1,658, and 6,630 mg Pb/L). Basal
9 respiration was decreased in two soil types tested at the highest Pb treatment (32 mM,
10 =6,630 mg Pb/L), and in a third at the two highest Pb treatments (8 and 32 mM, =1658
11 and 6,630 mg Pb/L). Terminal Restriction Fragment Length Polymorphism analysis
12 indicated that bacterial community structure was only slightly altered by Pb treatments.
13 While pH was correlated with the amount of water-soluble Pb, these increases were
14 apparently not significant enough to affect bacterial communities, because there were no
15 consistent relationships between soil pH and respiration rate or microbial community
16 structure at equivalent soil Pb concentration. Pb contamination was also demonstrated to
17 reduce phenol oxidase activity in several type of soils; concentrations between 5 and 50
18 nM Pb (0.001 and 0.01 mg Pb/L) significantly decreased phenol oxidase activity in all
19 soils tested, while 400 nM (0.08 mg Pb/L) and greater completely arrested phenol
20 oxidase activity in one soil tested (a high pH sandy loam) (Carine et al., 2009). Carine et
21 al. (2009) suggested that the decreased soil enzymatic activity resulted from changes in
22 the microbial community following Pb exposure. Pb concentrations between 50 and
23 500 mg Pb/kg significantly reduced microbial abundance and diversity, and also resulted
24 in lower soil phosphatase, urease, and dehydrogenase activities (Gao etal.. 201 Ob).
25 Further, the weekly soil carbon dioxide evolution rate was significantly reduced by
26 concentrations of 5, 10, and 50 mg Pb/g, which also indicated decreased microbial
27 respiration and adverse effects on the microbial community (Nwachukwu and Pulford.
28 2011). Gai et al. (2011) examined the microbial activity of three soils via
29 microcalorimetric methods following Pb exposure. They noted an increase in activity
30 immediately following Pb application (giving 10, 20, 40, 80, and 160 mg Pb/kg), and
31 theorized that this was a result of rapid mortality of sensitive microbial species, followed
32 by a concurrent proliferation of Pb-tolerant microorganisms. As Pb concentrations
33 increased, however, the calculated microbial growth rate constant decreased, indicating a
34 suppression of microbial activity (Gai et al.. 2011). Authors also noted a strong
35 correlation between microcalorimetry estimates and the number of colony forming units
36 isolated from soil samples.
37 Pb exposure negatively affected the prey capture ability of certain fungal species.
38 Nematophagous fungi are important predators of soil-dwelling nematodes, collecting
39 their prey with sticky nets, branches, and rings. The densities of traps they constructed
November 2012 7-63 Draft - Do Not Cite or Quote
-------
1 decreased in soils treated with 0.15 mM Pb chloride (31 mg Pb/L) (Mo et al., 2008). This
2 suppression caused a subsequent reduction in fungal nematode capturing capacity, and
3 could result in increased nematode abundance.
4 In a study of microbial communities and enzyme activity, Vaisvalavicius et al. (2006)
5 observed that high concentration of soil metals were linked to a significant reduction in
6 soil microorganism abundance and diversity. Soil columns spiked with Cu, Zn, and
7 Pb acetate (total Pb concentration of 278 to 838 mg Pb/kg, depending on depth) exhibited
8 a 10- to 100-fold decrease in microbial abundance, with specific microbe classes
9 (e.g., actinomycetes) seemingly more affected than others (Vaisvalavicius et al.. 2006).
10 Concurrently, decreases in soil enzymatic activity were also observed, with saccharase
11 activity decreased by 57-77%, dehydrogenase activity by 95-98%, and urease activity
12 65-97%. Although this suggests that Pb contamination may alter the nutrient cycling
13 capacity of affected soil communities, it is difficult to separate the impact of Pb in this
14 study from the contributions of Cu and Zn that were also added. In contrast, Zeng et al.
15 (2007) reported that soil concentrations of 300 mg Pb/kg and less stimulated soil
16 enzymatic activity. Both urease and dehydrogenase levels were increased and rice dry
17 weight was unaffected by concentrations of 100 and 300 mg Pb/kg. However, at 500 mg
18 Pb/kg, both rice and soil enzyme activities and microbial biomass were decreased
19 suggesting impacts at the community level for the soil-rice system. The authors proposed
20 that these concentrations could be considered the critical Pb concentration in rice paddy
21 systems (Zeng et al.. 2007).
22 The microbial communities of soils collected from a Pb-Zn mine and a Pb-Zn smelter
23 were significantly affected by Pb and other metals (e.g., Cd) (Hu et al.. 2007b). At a mine
24 site, Pb concentration of 57 to 204 mg Pb/kg and Cd concentration of 2.4 to 227 mg
25 Cd/kg decreased the number of bacteria-forming colonies extracted from soils. Principal
26 component analysis of microbial community structure demonstrated that different
27 communities were associated with different metal soil concentration. Similarly, soil
28 microbial communities exposed to metal contamination from a smelter site (soil Pb
29 concentration ranging from 30 to 25,583 mg Pb/kg dry weight) showed decreased
30 bacterial functional diversity (although fungal functional diversity increased) and no
31 effects on soil respiration rates were observed (Stefanowicz et al.. 2008). This led the
32 authors to conclude that bacterial diversity is a more sensitive endpoint and a better
33 indicator of metal exposure than fungal diversity or microorganism activity. In a similar
34 study, Kools et al. (2009) showed that soil ecosystem variables measured after a 6-month
35 exposure to metal-contaminated soil indicated that Pb concentration (536 or 745 mg
36 Pb/kg) was an important driver of soil microbial species biomass and diversity.
November 2012 7-64 Draft - Do Not Cite or Quote
-------
1 Pb-resistant bacterial and fungal communities were extracted regularly from soil samples
2 at a shooting range site in southern Finland (Hui et al.. 2009). While bioavailable Pb
3 concentration averaged 100 to 200 mg Pb/kg as determined by water extraction, the total
4 Pb concentrations measured on site were 30,000 to 40,000 mg Pb/kg. To determine Pb
5 tolerance, bacterial colonies extracted and cultured from shooting range and control soils
6 were grown on media containing either 0.4 or 1.8 mM Pb (83 or 373 mg Pb/L). While
7 bacteria isolated from control soil did not proliferate on high-Pb media, shooting-range
8 soil microbe isolates grew on high-Pb media and were deemed Pb tolerant. The authors
9 noted that bacterial species common in control samples were not detected among the
10 Pb-tolerant species isolated from shooting-range soils. They speculated that if long-term
11 exposure to minimally bioavailable Pb can alter the structure of soil decomposer
12 communities, decomposition rates could be altered. However, this would require that the
13 microbial ecosystem decomposing function be altered along with structure, and the
14 authors provided no evidence for alteration of function.
15 Microbial communities associated with habitats other than soils are also affected by
16 exposure to atmospherically deposited Pb. Alder (Alnus nepalensis) leaf microorganism
17 populations were greater in number at non-affected sites than at sites adjacent to a major
18 Indian highway with increased Pb pollution (Joshi. 2008). The density, species richness,
19 and biomass of testate amoebae communities grown on Sphagnum fallax mosses were
20 significantly decreased following moss incubation in Pb solutions of either 0.6 or 2.5 mg
21 Pb/L (Nguyen-Viet et al.. 2008). More importantly, species richness and density were
22 negatively correlated with Pb concentration accumulated within the moss tissue. The
23 structure of microbial communities associated with lichen surfaces was affected by lichen
24 trace-element accumulation, including Pb content. Lichens collected from industrial areas
25 had elevated Pb concentration (10 to 20 mg Pb/kg versus 5 to 7 mg Pb/kg in urban and
26 rural areas, respectively) and housed bacterial communities characterized by increased
27 cyanobacteria biomass (Meyer et al.. 2010).
28 Following a 28-day exposure to field-collected soils contaminated with metals (including
29 Pb at 426 mg Pb/kg), both population growth and individual growth of the earthworm
30 L. rubellus were diminished (Klok et al.. 2006). The authors proposed that, although
31 these reductions were unlikely to result in extirpation, avian predators such as the godwit
32 (Limosa limosa) that feed heavily on earthworms may be affected by a reduction of
33 available earthworm biomass.
34 During the past 5 years, there has been increasing interest in the effects of Pb and other
35 metals on the functional aspects of soil microbial communities. Most studies show that
36 Pb decreases diversity and function of soil microorganisms. However, in an example of
37 ecological mutualism, plant-associated arbuscular mycorrhizal fungi were found to
November 2012 7-65 Draft - Do Not Cite or Quote
-------
1 protect the host plant from Pb uptake, while fungal viability is protected by the host
2 plants. Similarly, soil microbial communities (bacterial species as well as fungi) in
3 Pb-contaminated soils are improved by revegetation. A few studies have reported on
4 effects of Pb to populations of soil invertebrates. They demonstrated that Pb can decrease
5 earthworm population density, although not to levels that would result in local extinction.
6 There have been no recently reported studies on the potential effects of Pb on terrestrial
7 vertebrate populations or communities, or possible indirect effects through reduction of
8 prey items such as earthworms.
7.3.7 Critical Loads in Terrestrial Systems
9 The general concept and definition of critical loads is introduced in Section 7.1.3 of this
10 chapter [also see Section 7.4 of the 2006 Pb AQCD (U.S. EPA. 2006c)1. An international
11 workshop was conducted in 2005 on the development of critical loads for metals and
12 other trace elements (Lofts et al., 2007). Among the findings of the workshop it was
13 reported that soil transport and transformation processes are key in controlling the fate of
14 metals and trace elements, thus their importance in the input-output mass balance needs
15 to be considered. The degree to which these processes are understood and can be
16 quantified varies. Complexation, sorption, ion exchange and precipitation are well
17 understood under laboratory conditions, but to a lesser extent in the field (Lofts et al..
18 2007). Slower processes of weathering and fixation are less well understood or studied
19 than leaching (Lofts et al., 2007).
20 As noted in previous sections, soil pH and organic matter influence Pb availability. De
21 Vries et al. (2007) demonstrate that critical limits, measured as critical reactive metal
22 content, can significantly vary between soil types that differ in pH and organic matter.
23 Critical limits of Pb increased from 30 to 64 (mg Pb/kg) over a pH range of 4-7 when soil
24 organic matter content was 5%, while these limits increased from 187 to 400 (mg Pb/kg)
25 over the same pH range when organic content was 80%. These implications suggest that
26 critical limits increase with increasing soil organic matter. This has important
27 consequences for forest soils because many are covered by an organic layer where roots,
28 fungi and other microorganisms are located. Baath (1989) evaluated the effects of organic
29 matter on critical limits for microorganisms, measured via enzyme synthesis, litter
30 decomposition and soil respiration. Results indicate critical limits are up to four times
31 higher in the organic (135 to 976 mg Pb/kg) than the mineral soil layer (32 to 690 mg
32 Pb/kg) at hazardous concentration ranging from 5-50% of species. In general, De Vries et
33 al. (2007) found support that ecotoxicological critical limits in European soils for Pb
34 decrease with increasing pH.
November 2012 7-66 Draft - Do Not Cite or Quote
-------
1 Several methods are routinely used for Pb risk assessment of terrestrial animals. Buekers
2 et al. (2009) proposed the use of a Tissue Residue Approach as a risk estimation method
3 for terrestrial vertebrates that eliminates the need for quantitative estimation of food
4 intake or Pb species bioavailability. Blood Pb no observed effect concentration (NOEC)
5 and lowest observed effects concentration (LOEC) data derived from 25 studies
6 examining the effects of Pb exposure on growth, reproduction, and hematological
7 endpoints were used to construct a series of species sensitivity distributions for mammals
8 and birds. They also used the HC5 criterion (5th percentile of species NOEC values for
9 collection of species) proposed by Aldenberg and Slob (1993). For mammals, the HC5
10 values obtained ranged from 11 to 18 (ig Pb/dL blood; HC5 values for birds ranged from
11 65 to 71 (ig Pb/dL. The authors proposed the use of 18 and 71 (ig Pb/dL as critical
12 threshold values for mammals and birds respectively, which are below the lowest NOEC
13 for both data sets used, and are above typical background Pb values. It is difficult to
14 determine environmental Pb toxicity given the variation of physiochemical and soil
15 properties that alter bioavailability and toxicity. This variability makes it difficult to
16 extrapolate between areas. Furman et al. (2006) proposed the use of a physiologically
17 based extraction test to predict risks posed to waterfowl from environmental Pb
18 contamination. The extraction process was modeled after gastric and intestinal conditions
19 of waterfowl, and was used to gauge the bioavailability of Pb from freshly amended and
20 aged contaminated soils. The concentration of Pb extracted through the use of the
21 physiologically based extraction test was demonstrated to be significantly correlated to
22 Pb tissue concentration in waterfowl exposed via in vivo studies of the same soils.
23 There are few critical loads for Pb reported for terrestrial ecosystems in the U.S.;
24 however, work has been conducted in Europe. Given that local conditions (including
25 historic loading, soil transport and transformation processes) are key elements to critical
26 load calculation the utility of critical loads that are developed from other countries for
27 application to U.S. ecosystems is unclear. The most recent European publications on Pb
28 critical loads include assessments of the U.K., Netherlands and Italy. Hall et al. (2006)
29 used the critical load approach to conduct a national risk assessment of atmospheric Pb
30 deposition for the U.K. While specific regions were determined to have low critical load
31 values for Pb (central England, the Pennines, and southern Wales), the authors noted that
32 this approach can be significantly biased, as available ecotoxicological data used in the
33 modeling were from studies that were not conducted in soils representative of all U.K.
34 soils. De Vries et al. (2009) similarly observed that the uncertainty inherent in a critical
35 load approach to Pb risk assessment is influenced by the critical concentration of
36 dissolved metal and the absorption coefficients of exposed soils. However, this approach
37 did indicate that for forest soils in the Netherlands, 29% of the areas would be expected
38 to exceed the critical load, based on currently available toxicity data and Pb pollution
39 data (de Vries and Groenenberg, 2009). Similarly, although Pb soil concentrations in the
November 2012 7-67 Draft - Do Not Cite or Quote
-------
1 Bologna Province of Italy were far below concentrations harmful to soil organisms,
2 current atmospheric Pb deposition rates suggest that critical load exceedances are likely
3 in the future, unless annual Pb emissions are decreased (Morselli et al., 2006).
4 Given the heterogeneity of ecosystems affected by Pb, and the differences in expectations
5 for ecosystem services attached to different land uses, it is expected that there will be a
6 range of critical load values for Pb for soils within the U.S. In the short term, metal
7 emissions generally have greater effects on biota in freshwater systems than in terrestrial
8 systems because metals are more readily immobilized in soils than in sediment. However,
9 over the longer term, terrestrial systems may be more affected particularly by those
10 metals with a long soil residence time, such as Pb.
7.3.8 Soil Screening Levels
11 Developed by EPA, ecological soil screening levels (Eco-SSLs) are maximum
12 contaminant concentrations in soils that are predicted to result in little or no quantifiable
13 effect on terrestrial receptors. These conservative values were developed so that
14 contaminants that could potentially present an unacceptable hazard to terrestrial
15 ecological receptors are reviewed during the risk evaluation process while removing from
16 consideration those that are highly unlikely to cause significant effects. The studies
17 considered for the Eco-SSLs for Pb and detailed consideration of the criteria for
18 developing the Eco-SSLs are provided in the 2006 Pb AQCD (U.S. EPA. 2006c).
19 Preference is given to studies using the most bioavailable form of Pb, to derive
20 conservative values. Soil concentration protective of avian and mammalian diets are
21 calculated by first converting dietary concentration to dose (mg/kg body weight per day)
22 for the critical study, then using food (and soil) ingestion rates and conservatively derived
23 uptake factors to calculate soil concentration that would result in unacceptable dietary
24 doses. This frequently results in Eco-SSL values below the average background soil
25 concentration [19 mg Pb/kg dry weight (U.S. EPA. 2005b. 2003b)1. as is the case with Pb
26 for birds. The Pb Eco-SSL was completed in March 2005 and has not been updated since.
27 Values for terrestrial birds, mammals, plants, and soil invertebrates are 11, 56, 120, and
28 1,700 mg Pb/kg soil (dry weight), respectively.
November 2012 7-68 Draft - Do Not Cite or Quote
-------
7.3.9 Characterization of Sensitivity and Vulnerability
1 Research has long demonstrated that Pb affects survival, reproduction, growth,
2 metabolism, and development in a wide range of species. The varying severity of these
3 effects depends in part upon species differences in metabolism, sequestration, and
4 elimination rates. Dietary factors also influence species sensitivity to Pb. Because of
5 effects of soil aging and other bioavailability factors discussed above (Section 7.3.2). in
6 combination with differing species assemblages and biological accessibility within prey
7 items, ecosystems may also differ in their sensitivity and vulnerability to Pb. The
8 2006 Pb AQCD reviewed many of these factors which are updated herein by reference to
9 recent literature.
7.3.9.1 Species Sensitivity
10 There is wide variation in sensitivity of terrestrial species to Pb exposure, even among
11 closely related organisms. Langdon et al. (2005) showed a two-fold difference in LC50
12 values among three common earthworm species, with the standard laboratory species,
13 E. andrei, being the least sensitive. Mammalian NOEC values expressed as blood Pb
14 levels were shown to vary by a factor of 8, while avian blood NOECs varied by a factor
15 of 50 (Buekers et al.. 2009). Age at exposure, in particular, may affect sensitivity to Pb.
16 For instance, earlier instar C. elegans were more likely than older individuals to exhibit
17 neurobehavioral toxicity following Pb exposure (Xing et al.. 2009b). and also
18 demonstrated more pronounced neural degeneration than older larvae and adults (Xing et
19 al.. 2009c).
7.3.9.2 Nutritional Factors
20 Dietary factors can exert significant influence on the uptake and toxicity of Pb in many
21 species of birds and mammals. The 2006 Pb AQCD (U.S. EPA. 2006b) describes how
22 Ca2+, Zn, Fe, vitamin E, Cu, thiamin, P, Mg, fat, protein, minerals, and ascorbic acid
23 dietary deficiencies increase Pb absorption and its toxicity. For example, vitamin E
24 content was demonstrated to protect against Pb-induced lipid peroxidation in mallard
25 ducks. Generally, Pb exposure is more likely to produce behavioral effects in conjunction
26 with a nutrient-deficient diet. As previously reported in the 2006 Pb AQCD, Ca2+
27 deficiencies may increase the susceptibility of different terrestrial species to Pb, including
28 plant (Antosiewicz. 2005). avian (Dauwe et al.. 2006; Snoeijs et al.. 2005) and
29 invertebrate species. Antosiewicz (2005) determined that, for plants, Ca2+ deficiency
30 decreased the sequestration capacity of several species (tomato, mustard, rye, and maize),
November 2012 7-69 Draft - Do Not Cite or Quote
-------
1 and that this likely resulted in an increased proportion of Pb at sites of toxic action.
2 Because Pb ions can interact with plant Ca2+ channel pores, in the presence of low Ca2+
3 and high Pb concentration, a higher proportion of Pb can interact with these channels and
4 be taken up by plants. A similar phenomenon has been observed in invertebrates, where
5 the metabolic pathway of metals mimics the metabolic pathway of Ca2+ [Simkiss et al.
6 (1982). as cited in Jordaens et al. (2006)1. Hence, in environments with
7 disproportionately high Pb versus Ca2+ concentration, accumulation of Pb may be
8 accelerated, as in plants. Ca2+ deficiency in birds was demonstrated to stimulate the
9 production of Ca2+-binding proteins in the intestinal tract, which extract more Ca2+ from
10 available diet; however, this response also enhances the uptake and accumulation of Pb
11 from diet and drinking water [Fullmer (1997). as cited in Dauwe et al. (2006)].
7.3.9.3 Soil Aging and Site-Specific Bioavailability
12 Total soil Pb concentration is a poor predictor of hazards to avian or mammalian wildlife,
13 because site-specific biogeochemical and physical properties (e.g., pH, OM, metal oxide
14 concentration) can affect the sequestration capacity of soils. Additionally, soil aging
15 processes have been demonstrated to decrease the bioavailable Pb fraction; as such,
16 laboratory toxicity data derived from spiked soils often overestimate the environmental
17 risk of Pb. Smolders et al. (2009) compared the toxicity of freshly Pb-spiked soils to
18 experimentally aged spiked soils and field-collected Pb-contaminated soils. Experimental
19 leaching and aging was demonstrated to increase invertebrate Pb EC50 values by factors
20 of 0.4 to greater than 8; in approximately half the cases, the proportionality of toxicity to
21 Pb content disappeared following experimental aging of freshly spiked soils through
22 leaching. The leaching-aging factor for Pb was determined to be 4.2, and represented the
23 ratio of EDi0 values derived in aged soils to freshly spiked soils (factors greater than one
24 indicate decreased toxicity in aged field soils relative to laboratory spiked soils).
25 Consequently, the sensitivity of terrestrial vertebrates to environmental Pb exposures will
26 be heavily dependent on the relative rate of aging and site-specific bioavailability.
7.3.9.4 Ecosystem Vulnerability
27 Relative vulnerability of different terrestrial ecosystems to effects of Pb can be inferred
28 from the information discussed above on species sensitivity and how soil geochemistry
29 influences the bioavailability and toxicity of Pb. Soil ecosystems with low pH,
30 particularly those with sandy soils, are likely to be the most sensitive to the effects of Pb.
31 Examples of such systems are forest soils, including oak, beech, and conifer forests.
32 The Pine Barrens in southern New Jersey (also known as the Pinelands) is an example of
November 2012 7-70 Draft - Do Not Cite or Quote
-------
1 a highly vulnerable ecosystem: it is a dense coniferous (pine) forest with acidic, sandy,
2 nutrient poor soil. As agricultural areas are taken out of production and revert to old
3 fields and eventually forests, their vulnerability to Pb is likely to increase as a result of
4 decreasing OM and acidification of soils (from discontinuation of fertilizing and liming).
5 On the other hand, increasing density of native or invasive plants with associated
6 arbuscular mycorrhizal fungi will likely act to ameliorate some of the effects of Pb (see
7 previous discussion of studies by Sudova and Vostka (2007) and Wong et al. (2007). It is,
8 however, difficult to categorically state that certain plant or soil invertebrate communities
9 are more vulnerable to Pb than others, as the available toxicity data have not yet been
10 standardized for differences in bioavailability (because of use of different Pb salts,
11 different soil properties, and different lengths of aging of soil prior to testing), nutritional
12 state, or organism age, or other interacting factors. Data from field studies are
13 complicated by the co-occurrence of other metals and alterations of pH, such as
14 acidification from SO2 in smelter emissions, which are almost universal at sites of high
15 Pb exposure, especially at mine or smelter sites. However, because plants primarily
16 sequester Pb in the roots, uptake by soil invertebrates is the most likely pathway for Pb
17 exposure of higher trophic level organisms. Invertivores are likely at higher risk than
18 herbivores. In fact, estimations of Pb risk at a former Pb smelter in northern France
19 indicated that area Pb concentration presented the greatest threat to insectivorous bird and
20 mammal species, but only minimal risk to soil invertebrate and herbivorous mammals
21 (Fritsch et al.. 2010). By extension, birds and mammals in ecosystems with a richer
22 biodiversity of soil invertebrates may be more vulnerable to Pb than those in ecosystems
23 with fewer invertebrates (e.g., arid locations). Regardless, the primary determinant of
24 terrestrial ecosystem vulnerability is soil geochemistry, notably pH, CEC, and amount of
25 OM.
November 2012 7-71 Draft - Do Not Cite or Quote
-------
7.3.10 Ecosystem Services Associated with Terrestrial Systems
1 Pb deposited on the surface of, or taken up by organisms has the potential to alter the
2 services provided by terrestrial biota to humans. There are no publications at this time
3 that specifically focus on the ecosystem services affected by Pb in terrestrial systems and
4 the directionality of impacts is not always clear. For example, terrestrial soils provide a
5 service to aquatic ecosystems by sequestering Pb through sorption and precipitation. At
6 the same time, the sequestration of Pb by soils may result in a degredation in the quality
7 of soil and may result in decreased crop productivity. The evidence reviewed in the
8 present document illustrates that Pb can cause ecological effects in each of the four main
9 categories of ecosystem services (Section 7.1.2) as defined by Hassan et al. (2005). These
10 effects are sorted into ecosystem services categories and summarized here:
11 • Supporting: altered nutrient cycling, decreased biodiversity, decline of
12 productivity, food production for higher trophic levels
13 • Provisioning: plant yields
14 • Regulating: decline in soil quality, detritus production
15 • Cultural: ecotourism and cultural heritage values related to ecosystem integrity
16 and biodiversity, impacts to terrestrial vertebrates.
17 A few studies since the 2006 Pb AQCD, consider the impact of metals in general on
18 ecosystem services. Honeybees are important for provisioning services such as
19 pollination and production of honey. They can be exposed to atmospheric Pb by direct
20 deposition or through Pb associated with plants, water or soil. In a study of heavy metals
21 in honeybees in central Italy, there was a statistically significant difference in Pb between
22 bees collected in wildlife reserves compared to bees collected in urban areas with the
23 highest concentration of Pb detected from bees caught in hives near an airport (Perugini
24 et al.. 2011). In a review of the effects of metals on insect behavior, ecosystem services
25 provided by insects such as detritus reduction and food production for higher trophic
26 levels were evaluated by considering changes in ingestion behavior and taxis (Mogren
27 and Trumble. 2010). Pb was shown in a limited number of studies to affect ingestion by
28 insects. Crickets (Chorthippus spp) in heavily contaminated sites reduced their
29 consumption of leaves in the presence of increasing Cd and Pb concentrations (Migula
30 and Binkowska. 1993). Decreased feeding activity in larval and adult Colorado potato
31 beetle (Leptinotarsa decemlineata) were observed as a result of dietary exposures of Pb
32 and Cu (Kwartirnikov et al.. 1999). while no effects were found in ingestion studies of Pb
33 with willow leaf beetle, Lochmaea caprae (Rokvtova et al.. 2004) mottled water hyacinth
November 2012 7-72 Draft - Do Not Cite or Quote
-------
1 weevil, Neochetina eichhorniae (Kay and Haller. 1986) and hairy springtail, Orchesella
1 cincta (Van Capelleveen et al.. 1986J.
3 Soil health for agricultural production and other soil-associated ecosystem services is
4 dependent upon the maintenance of four major functions: carbon transformations,
5 nutrient cycles, soil structure maintenance, and the regulation of diseases and pests and
6 these parameters may be altered by metal deposition (Kibblewhite et al., 2008). Pb
7 impacts to terrestrial systems reviewed in the previous sections provide evidence for
8 impacts to supporting, provisioning, and regulating ecosystem services provided by soils.
9 For example, earthworms were shown to impact soil metal mobility and availability,
10 which in turn resulted in changes to microbial populations (biodiversity), pH, dissolved
11 organic carbon, and metal speciation (Sizmur and Hodson, 2009). all of which may
12 directly affect soil fertility.
13 Pb is bioaccumulated in plants, invertebrates and vertebrates inhabiting terrestrial and
14 aquatic systems that receive Pb from atmospheric deposition. This represents a potential
15 route for Pb mobilization into the food web or into food products. For example, Pb
16 bioaccumulation in leaves and roots of an edible plant may represent an adverse impact to
17 the provisioning of food, an essential ecosystem service. Although there is no consistent
18 evidence of trophic magnification there is substantial evidence of trophic transfer. It is
19 through consumption of Pb-exposed prey or Pb-contaminated food that atmospherically
20 deposited Pb reaches species that may have very little direct exposure to it.
21 There is limited evidence of Pb impacts to plant productivity. Productivity of gray birch
22 (Betula populifolid) was impaired in soils with elevated As, Cr, Pb, Zn and V (Gallagher
23 et al.. 2008). Tree growth measured in both individuals and at the assemblage level using
24 satellite imagery and field spectrometry was significantly decreased with increasing metal
25 load in soil.
7.3.11 Synthesis of New Evidence for Pb Effects in Terrestrial Systems
26 This synthesis of the effects of Pb on terrestrial ecosystems covers information from the
27 publication of the 2006 Pb AQCD to present. It is followed in Section 7.5 by
28 determinations of causality that take into account evidence dating back to the 1977 Pb
29 AQCD.
High concentrations of Pb
30 The state-level mean concentration of Pb in U.S. soils ranges from 5 to 39 mg Pb/kg.
31 Studies of the effects of Pb use much higher concentrations, whether they use soils that
November 2012 7-73 Draft - Do Not Cite or Quote
-------
1 have been exposed to Pb pollution, or experimental amendment with salts of Pb (Table
2 7-4). Studies that were conducted in situ or used soil collected from natural environments
3 all took place near stationary sources, i.e., in highly contaminated areas. All of them took
4 advantage of gradients of exposure produced by distance from the source to create
5 several levels of Pb, sometimes with only two sampling locations-control and elevated-
6 rather a larger set of levels representative of the whole gradient. In most cases, the
7 highest concentration of Pb in the study is very high relative to those found anywhere
8 except heavily exposed sites. In amendment experiments, variation in Pb was generated
9 by addition of Pb salts to either natural or artificial soils. These experiments often
10 included concentrations that were even higher than those found in heavily exposed
11 natural environments. In either type of study, however, effects gradually increased with
12 increasing exposure, and studies that include very high exposures were thus informative
13 for lower exposures as well. This would not be true if there was clear evidence of the
14 presence of discontinuity (breakpoint) in the relationship of exposure and effects, but
15 without evidence of discontinuity, the presence of effects at elevated exposures implies
16 effects at lower exposures. Using concentration-response models where concentration is
17 taken as a continuous variable to analyze data with multiple values of Pb concentration
18 would allow better estimation of the size of effects at any value of exposure, including
19 low ones, and also better estimation of uncertainties around the size of effects. However,
20 none of studies with multiple Pb concentrations used a continuous model to characterize
21 the relationship between concentration and effects.
Comparability of effect concentrations
22 Strong interactions of Pb concentration and several other soil variables, including pH,
23 CEC, OC, and Fe/Al oxides have been amply demonstrated with respect to various
24 biological responses. For example, Dayton et al.(2006) and Bradham et al.(2006) tested
25 an array of different soils to which the same amount of Pb was added, using lettuce and
26 earthworms, respectively. They found differences in biological effects that were as large
27 as 27, 35, or even 72-fold between soils.
28 In studies where Pb was introduced through amendment, those interacting variables can
29 be changed experimentally in a controlled way, or held constant. In studies where natural
30 soils were used in which Pb originated from pollution, they are left to vary freely. In
31 either case, the presence and magnitude of those interactions make calculations of
32 expected responses under other sets of conditions particularly difficult, as well as
33 comparisons between studies conducted under different conditions.
34 In addition, the amount of Pb dissolved in soil pore water determines the impact of soil
35 Pb on terrestrial ecosystems to a much greater extent than the total amount present. It has
November 2012 7-74 Draft - Do Not Cite or Quote
-------
1 long been established that the amount of Pb dissolved in soil solution is controlled by at
2 least six variables: (1) solubility equilibria; (2) adsorption-desorption relationship of total
3 Pb with inorganic compounds; (3) adsorption-desorption reactions of dissolved Pb phases
4 on soil OM; (4) pH; (5) CEC; and (6) aging. Since 2006, further details have been
5 contributed to the understanding of the role of pH, CEC, OM, and aging. Smolders et al.
6 (2009) demonstrated that the two most important determinants of solubility (and also
7 toxicity) in soils are pH and CEC. However, they had previously shown that aging,
8 primarily in the form of initial leaching following deposition, decreases soluble metal
9 fraction by approximately one order of magnitude (Smolders et al.. 2007). Since 2006,
10 OM has been confirmed as an important influence on Pb sequestration, leading to longer-
11 term retention in soils with higher OM content, and also creating the potential for later
12 release of deposited Pb. Aging, both under natural conditions and simulated through
13 leaching , was shown to substantially decrease bioavailability to plants, microbes, and
14 vertebrates. However, most studies report some measure of total extracted Pb, or total
15 added Pb, rather than pore water or soluble fraction.
Plants
16 Recent studies with herbaceous species growing at various distances from smelters added
17 to the existing strong evidence that atmospherically transported Pb is taken up by plants.
18 These studies did not establish the relative proportion that originated from atmospheric
19 Pb deposited in the soil, as opposed to that taken up directly from the atmosphere through
20 the leaves. Studies found that in trees, Pb that is taken up through the roots is then
21 generally translocated from the roots to other parts. However, multiple recent studies
22 showed that in trees, the proportion of Pb that is taken up through the leaves is likely to
23 be very substantial. One study attempted to quantify it, and suggested that 50% of the Pb
24 contained in Scots Pine in Sweden is taken up directly from the atmosphere
25 (Section 7.3.3.1). Studies with herbaceous plants found that in most species tested, soil
26 Pb taken up by the roots is not translocated into the stem and leaves, but when growth
27 and survival were reported, growth of the whole plant decreased with increasing Pb, and
28 mortality increased (Table 7-4). Experimental studies have added to the existing evidence
29 of photosynthesis impairment in plants exposed to Pb, and have found damage to
30 photosystem II due to alteration of chlorophyll structure, as well as decreases in
31 chlorophyll content in diverse taxa, including lichens and mosses. A substantial amount
32 of evidence of oxidative stress in response to Pb exposure has also been produced.
33 Reactive oxygen species were found to increase in broad bean and tomato plants exposed
34 to increasing concentrations of soil Pb, and a concomitant increase in superoxide
35 dismutase, glutathione, peroxidases, and lipid peroxidation, as well as decreases in
36 catalase were observed in the same plants. Monocot, dicot, and bryophytic taxa grown in
November 2012 7-75 Draft - Do Not Cite or Quote
-------
1 Pb-contaminated soil or in experimentally spiked soil all responded to increasing
2 exposure with increased antioxidant activity. In addition, genotoxicity, decreased
3 germination, and pollen sterility were observed in some experiments. All effects were
4 small outside of contaminated areas (Section 7.3.4.1).
5 Invertebrates
6 Since the 2006 Pb AQCD, various species of terrestrial snails have been found to
7 accumulate Pb from both diet and soil, although effects on growth, survival and
8 reproduction are inconsistent. Recent studies with earthworms have found that both
9 internal concentration of Pb and mortality increase with decreasing soil pH and CEC. In
10 addition, tissue concentration differences have been found in species of earthworms that
11 burrow in different soil layers. The rate of accumulation in each of these species could
12 result from layer differences in interacting factors such as pH and CEC (Section 7.3.3.2).
13 Because earthworms often sequester Pb in granules, some authors have suggested that
14 earthworm Pb is not bioavailable to their predators. There is some evidence that
15 earthworm activity increases Pb availability in soil, but it is inconsistent. In arthropods
16 collected at contaminated sites, recent studies found gradients in accumulated Pb that
17 corresponded to gradients in soil with increasing distance from stationary sources.
18 Recently published studies have shown neuronal damage in nematodes exposed to low
19 concentrations of Pb (2.5 uM = 0.5 mg Pb/L), accompanied by behavioral abnormalities.
20 Reproductive effects were found at lower exposure in younger nematodes, and effects on
21 longevity and fecundity were shown to persist for several generations. Increased
22 mortality was found in earthworms, and was strongly dependent on soil characteristics
23 including pH, CEC, and aging. Snails exposed to Pb through either topical application or
24 through consumption of Pb-exposed plants had increased antioxidant activity and
25 decreased food consumption, but effects on growth and survival were inconsistent.
26 Effects on arthropods exposed through soil or diet varied with species and exposure
27 conditions, and included diminished growth and fecundity in springtails, endocrine and
28 reproductive anomalies, and body deformities. Increasing concentration of Pb in the
29 exposure medium generally resulted in increased effects within each study, but the
30 relationship between concentration and effects varied between studies, even when the
31 same medium, e.g., soil, was used (Section 7.3.4.2). Evidence suggested that aging and
32 pH are important modifiers.
33 Vertebrates
34 There were few recent studies of Pb bioavailability and uptake in vertebrates since the
35 2006 Pb AQCD. A study of two species of sea ducks in Alaska found that 3% of the birds
36 had tissue levels of Pb that indicated exposure above background. Urban pigeons in
37 Korea were found to accumulate 1.6 to 1.9 mg Pb/kg wet weight Pb in the lungs, while in
November 2012 7-76 Draft - Do Not Cite or Quote
-------
1 Wisconsin 70% of American woodcock chicks and 43% of young-of-year had elevated
2 bone Pb (9.6 to 93 mg Pb/kg dry weight in chicks, 1.5 to 220 mg Pb/kg dry weight in
3 young-of-year). None of the locations for these studies was in proximity to stationary
4 sources of heavy contamination, and none was able to identify the origin of the Pb
5 (Section 7.3.3.3). Effects on amphibians and reptiles included decreased white blood cell
6 counts, decreased testis weight, and behavioral anomalies. However, large differences in
7 effects were observed at the same concentration of Pb in soil, depending on whether the
8 soil was freshly amended, or field-collected from contaminated areas. As in most studies
9 where the comparison was made, effects were smaller when field-collected soils were
10 used. A study at the Anaconda Smelter Superfund site found increasing Pb accumulation
11 in gophers with increasing soil Pb around the location of capture. Effects of dietary
12 exposure were studied in several mammalian species, and cognitive, endocrine,
13 immunological, and growth effects were observed. Pigs fed various Pb-contaminated
14 soils showed that the form of Pb determined accumulation, and another study showed
15 lower feed efficiency and weight in pigs with 2,08 versus 1.44 ug Pb/dl in blood,
16 originating in Pb-sulfate feed supplement. In some birds, maternal elevated blood Pb
17 level was associated in recent studies with decreased hatching success, smaller clutch
18 size, high corticosteroid level, and abnormal behavior. Some species show little or no
19 effect of elevated blood Pb level. A study of Japanese quail found that Pb added to the
20 diet could improve survival and incidence of several pathologies, and a long term study
21 of pied flycatchers at a mine site produced mixed evidence for the effects of Pb
22 (Section 7.3.4.3).
23 Food web
24 Recent studies were able to measure Pb in the components of various food chains that
25 included soil, plants, invertebrates, arthropods and vertebrates. They confirmed that
26 trophic transfer of Pb is pervasive, but no consistent evidence of trophic magnification
27 was found (Section 7.3.3.4).
28 Community and Ecosystem Effects
29 New evidence of effects of Pb at the community and ecosystem levels of biological
30 organization include several studies of the ameliorative effects of mycorrhizal fungi on
31 plant growth, attributed to decreased uptake of Pb by plants, although both mycorrhizal
32 fungus and plant were negatively affected. The presence of both earthworms and
33 mycorrhizal fungi decreased solubility and mobility of Pb in soil in one study, but the
34 presence of earthworms was associated with higher uptake of Pb by plants in another.
35 The presence of snails increased uptake of Pb by earthworms, but not vice-versa. Most
36 recently published research on community and ecosystem effects of Pb has focused on
37 soil microbial communities, which have been shown to be impacted in both composition
November 2012 7-77 Draft - Do Not Cite or Quote
-------
1 and activity. Many recent studies have been conducted using mixtures of metals, but have
2 tried to separate the effects of individual metals when possible. One study compared the
3 effects of 6 metals individually (Akerblom et al.. 2007). and found that their effects on
4 community composition were similar. In studies that included only Pb, or where effects
5 of Pb could be separated, soil microbial activity was generally diminished, but in some
6 cases recovered overtime. Species and genotype composition were consistently altered,
7 and those changes were long-lasting or permanent (Section 7.3.6) .
8 Exposure-Response
9 Several studies with various organisms have included gradients of Pb exposure. None has
10 characterized the exposure-relationship using a continuous model of exposure-response.
11 However, evidence indicates clearly that increased exposure to Pb is associated with
12 increases in observed effects in terrestrial ecosystems. Evidence also demonstrates that
13 many factors, including species and various soil physiochemical properties, interact
14 strongly with Pb concentration to modify those effects. In terrestrial ecosystems, where
15 soil is generally the main component of the exposure route, Pb aging is a particularly
16 important factor, and one that may be difficult to reproduce experimentally. Without
17 adequate quantification of those interactions, characterizations of exposure-response
18 relationships may be difficult to transfer outside of experimental settings.
7.3.12 Causal Determinations for Pb in Terrestrial Systems
19 In the following sections, organism-level effects on reproduction and development,
20 growth and survival are considered first since these endpoints can lead to effects at the
21 population level or above and are important in ecological risk assessment.
22 Neurobehavioral effects are considered next followed by sub-organismal responses
23 (hematological effects, physiological stress) for which Pb has been shown to have an
24 impact in multiple species and across taxa, including humans. Causal determinations for
25 terrestrial, freshwater and saltwater ecological effects are summarized in Table 7-3.
7.3.12.1 Reproductive and Developmental Effects-Terrestrial Biota
26 In terrestrial invertebrates and vertebrates, evidence assessed for the present document
27 and in Pb AQCDs indicates an association between reproductive effects and Pb exposure.
28 Impaired fecundity at the organism level of biological organization can result in a decline
29 in abundance and/or extirpation of populations, decreased taxa richness, and decreased
30 relative or absolute abundance at the community level (Suter et al.. 2005; U.S. EPA.
31 2003a). Evaluation of the literature on Pb effects in terrestrial species indicates that
November 2012 7-78 Draft - Do Not Cite or Quote
-------
1 exposure to Pb is associated with reproductive effects. Various endpoints have been
2 measured in various taxa of terrestrial organisms to assess the effect of Pb on fecundity,
3 development, and hormone homeostasis. Although reproductive effects were
4 demonstrated, no single endpoint in a single taxon has been extensively studied. Recent
5 evidence available since the 2006 Pb AQCD for effects of Pb on reproductive endpoints
6 in terrestrial species is summarized in Table 7-4.
7 In terrestrial plants, few studies were available to the 2006 Pb AQCD (U.S. EPA. 2006b).
8 and few are available more recently that specifically address reproductive effects of Pb
9 exposure. Two genotypes of maize seedlings exhibited a significant and concentration-
10 dependent reduction in seed germination following 7 days of Pb treatment in nutrient
11 solution of nominal concentration of 0, 0.007, 0.7 and 7 mg Pb/L as Pb sulfate (Ahmad et
12 al.. 2011). Germination inhibition and chromosomal abnormalities also increased in a
13 concentration-dependent manner in Grass pea grown in soil irrigated with solutions
14 containing nominal concentration of 0 to 188 mg Pb/L (Kumar and Tripathi. 2008).
15 However, germination increased in a broad sample of soils when amended with 2,000 mg
16 Pb/kg (Davton et al.. 2006).
17 In terrestrial invertebrates, Pb can alter developmental timing, hatching success, sperm
18 morphology and hormone homeostasis. The number of species studied has been small,
19 but reproductive effects consistently increase with increasing exposure. The
20 2006 Pb AQCD reported effects on reproduction in collembolans and earthworms, with
21 LOECs and NOECs typically well above Pb soil concentrations observed away from
22 stationary sources of contamination, more recently, an increase in development time
23 (approximately two days) and a reduction in relative fecundity were observed in aphids
24 feeding on plants exposed to high concentrations of Pb (Goriir. 2007). Hatching success
25 of the collembolan F. Candida was decreased following 10 day nominal exposure to
26 Pb-spiked soils (EC50 2,361 mg Pb/kg dry soils) (Xu et al., 2009b). Sperm morphology in
27 Asian earthworms was significantly altered following 2-week exposures to soils
28 containing nominal concentration of 1,000, 1,400, 1,800 and 2,500 mg Pb/kg soil (Zheng
29 and Li. 2009). Pb may also disrupt hormonal homeostasis in invertebrates as studies with
30 moths have suggested (Shu et al.. 2009). Adult female moths reared on diets containing
31 25, 50, 100, or 200 mg Pb/kg exhibited decreased vitellogenin mRNA induction, and
32 vitellogenin levels were demonstrated to decrease with increasing Pb exposure. Evidence
33 of multi-generational toxicity effects of Pb is also present in terrestrial invertebrates,
34 specifically springtails, mosquitoes, carabid beetles and nematodes where decreased
35 fecundity in progeny of Pb-exposed individuals was observed. The magnitude of effects
36 is variable, but they are present in multiple phyla, and increase with increasing exposure
37 within studies. Reproductive effects in terrestrial invertebrates are also coherent with
38 similar effects observed in aquatic invertebrates.
November 2012 7-79 Draft - Do Not Cite or Quote
-------
1 In terrestrial vertebrates, there is evidence for reproductive effects associated with Pb
2 exposure in recent evidence and Pb AQCDs. The 2006 Pb AQCD (U.S. EPA. 2006c)
3 concluded that exposure to affects reproduction and development in terrestrial
4 vertebrates. Effects reported in that document included declines in clutch size, number of
5 young hatched, number of young fledged, decreased fertility, and decreased eggshell
6 thickness observed in birds near areas of Pb contamination and in birds with elevated Pb
7 tissue concentration regardless of location. More recently, decreased testis weight was
8 observed in lizards administered a sublethal dose of 10 or 20 mg Pb/kg day by oral
9 gavage for 60 days (Salice et al. 2009). Few studies in the field have addressed
10 reproductive effects of Pb specifically in mammals, due to most available data in wild or
11 grazing animals being from near smelters, where animals are co-exposed to other metals.
12 For example, the reproductive viability of red deer (C. elaphus) inhabiting a
13 Pb-contaminated mining area of Spain was shown to be altered, with 11% and 15%
14 reductions in spermatozoa and acrosome integrity observed in male deer from the mining
15 area compared with those residing in reference areas (Reglero et al.. 2009aX but multiple
16 other metals were present at high concentrations. Evidence from AQCDs and the present
17 document for terrestrial vertebrates is coherent with evidence from freshwater
18 amphibians, and fish (Section 7.4.12.1). However, experimental evidence obtained using
19 mammals in the context of human health research demonstrates a consistency of adverse
20 effects of Pb on sperm (Section 5.8.4.1) and the onset of puberty in males and females
21 (Sections 5.8.1.1 and 5.8.1.2) with strong evidence from both toxicology and
22 epidemiology studies. Other reproductive endpoints including spontaneous abortions,
23 pre-term birth, embryo development, placental development, low birth weight,
24 subfecundity, hormonal changes, and teratology were also affected, but less consistently
25 (Section 5.8).
26 For reproductive and developmental effects in terrestrial ecosystems, the current body of
27 evidence is inadequate to conclude that exposure to Pb is causal in plants, and is
28 sufficient to conclude that there is a causal relationship in invertebrates and vertebrates.
7.3.12.2 Growth Effects-Terrestrial Biota
29 Alterations in growth at the organism level of biological organization can have impacts at
30 the population, community and ecosystem levels. In terrestrial ecosystems, evidence for
31 effects of Pb on growth is strongest in terrestrial plants, although these effects are
32 typically observed in laboratory studies with high exposure concentrations or in field
33 studies near stationary sources. In terrestrial plants, there is evidence over several decades
34 of research that Pb inhibits photosynthesis and respiration, all of which can reduce the
35 growth of the plant (U.S. EPA. 2006c. 1986a. 1977).Decreases in chlorophyll a and b
November 2012 7-80 Draft - Do Not Cite or Quote
-------
1 content have been observed in various algal and plant species. Many laboratory toxicity
2 studies report effects on the growth of plants in synthetic growing media, but observed
3 effects typically occur at concentrations higher than the average background
4 concentrations in U.S. soils (19 mg Pb/kg dry weight) (U.S. EPA. 2005b) and there are
5 few field studies. Effects on plant growth can result in reduced productivity and
6 decreased biomass. The 2006 Pb AQCD relied principally on evidence assembled in the
7 Ecological Soil Screening Levels for Lead document (U.S. EPA. 2005b), which
8 concluded that growth (biomass) was the most sensitive and ecologically relevant
9 endpoint for plants.
10 Evidence for growth effects in terrestrial fauna is sparse. In the 1986 Pb AQCD, a study
11 was reviewed in which the Fl and F2 generations of the springtail Onychiurus armatus
12 fed a diet of Pb-exposed fungi (0.008 to 3.1 mg Pb/g) experienced a delay in achieving
13 maximum length (Bengtsson et al., 1983). The authors suggested that the reduced growth
14 may be accompanied by delayed sexual maturity. The 2006 Pb AQCD (U.S. EPA.
15 2006b) reported that growth effects observed in both terrestrial invertebrates and
16 vertebrates were more pronounced in juvenile organisms, underscoring the importance of
17 lifestage to overall Pb susceptibility. Recent evidence available since the 2006 Pb AQCD
18 for effects of Pb on growth endpoints in terrestrial species is summarized in Table 7-4:
19 reduced growth of garden snail T. pisana, increasing with increasing exposure, was
20 observed following a five week dietary exposure to eight nominal concentrations of Pb
21 (El-Gendy et al., 2011). Studies also show concentration-dependent inhibition of growth
22 in earthworms raised in Pb-amended soil (Zheng and Li. 2009; Currie et al.. 2005;
23 Langdon et al., 2005). In AQCDs, growth effects of Pb have been reported in birds
24 (changes in juvenile weight gain), at concentrations typically higher than currently found
25 in the environment away from heavily exposed sites. The current body of evidence is
26 sufficient to conclude that there is a causal relationship between Pb exposures and growth
27 effects in terrestrial plants, and that a causal relationship is likely to exist between Pb
28 exposure and growth effects in terrestrial invertebrates. Evidence is inadequate to
29 establish causal relationship between Pb exposures and growth effects in terrestrial
30 vertebrates.
7.3.12.3 Survival-Terrestrial Biota
31 The relationship between Pb exposure and survival has been well demonstrated in
32 terrestrial species as presented in the Pb AQCDs and in Section 7.3.5 of the present
33 document. Exposure can be either lethal, or produce sublethal effects that diminish
34 survival probabilities. In the 1977 Pb AQCD, deaths from Pb poisoning in domestic
35 animals caused by emissions from stationary sources were reported (U.S. EPA. 1977).
November 2012 7-81 Draft - Do Not Cite or Quote
-------
1 Additional studies in the 1986 and 2006 Pb AQCDs and current ISA provide evidence for
2 a concentration-dependent response of mortality in terrestrial biota. Recent evidence
3 available since the 2006 Pb AQCD for effects of Pb on survival in terrestrial species is
4 summarized in Table 7-4.
5 Survival is a biologically important response that can have direct impact on population
6 size. Survival is often quantified using LC50 (the concentration of toxicant where 50%
7 mortality is observed or modeled), which may be a better metric for acute toxicity than
8 for typical environmental exposure, which is more often comparatively low, and
9 cumulative or chronic. From the LC50 data on Pb in this review and previous Pb AQCDs,
10 a wide range of sensitivity to Pb is evident across taxa and within genera. As expected,
11 reported LC50 are usually much higher than current environmental levels of Pb in the U.S
12 away from heavily exposed sites, even though physiological dysfunction that adversely
13 impacts the fitness of an organism often occurs well below concentrations that result in
14 mortality. When available, LCio, NOEC or LOEC have been reported in the present
15 document.
16 Pb is generally not phytotoxic to plants at concentrations found in the environment away
17 from heavily exposed sites, probably due to the fact that plants often sequester large
18 amounts of Pb in roots, and that translocation to other parts of the plant is limited. No
19 data have become available to change this assessment since the 2006 Pb AQCD.
20 Survival of soil-associated organisms is adversely affected by Pb exposure. In the 1986
21 Pb AQCD it was reported that Pb at the high extreme of concentrations found near
22 roadsides and smelters at the time (10,000 to 40,000 mg Pb/kg dry weight) can eliminate
23 populations of bacteria and fungi on leaf surfaces and in soil. Severe impairment of
24 decomposition has long been accepted to be one of the most apparent results of soil
25 contamination with Pb and other metals. In nematodes, the 2006 Pb AQCD reported LC50
26 values varying from 10 to 1,550 mg Pb/kg dry weight dependent upon soil OM content
27 and soil pH (U.S. EPA. 2006c). In earthworms, 14 and 28 day LC50 values typically fell
28 in the range of 2,400-5,800 mg Pb/kg depending upon the species tested. More recent
29 evidence has been consistent with these values, and also showed concentration-dependent
30 decreases in survival in collembolans and earthworms under various experimental
31 conditions. The evidence in terrestrial invertebrates is coherent with evidence in
3 2 fire shwater invertebrate s.
33 In terrestrial avian and mammalian species, toxicity is observed in laboratory studies over
34 a wide range of doses (<1 to > 1,000 mg Pb/kg body weight-day) as reviewed for the
35 development of Eco-SSLs (U.S. EPA. 2005b). and subsequently reported in the
36 2006 Pb AQCD. The NOAELs for survival ranged from 3.5 to 3,200 mg Pb/kg-day.
37 Surprisingly, the only study to have reported survival data following exposure to Pb in an
November 2012 7-82 Draft - Do Not Cite or Quote
-------
1 avion species since the 2006 Pb AQCD, found that survival was greater than in controls
2 in quail exposed to 50 mg Pb/L in drinking water for 7 weeks (Nain and Smits. 2011).
3 Evidence for association of Pb exposure with mortality in terrestrial vertebrates is
4 coherent with observations in freshwater vertebrates (Section 7.4.12.3). The evidence is,
5 therefore, sufficient to conclude that a causal relationship is likely to exist between Pb
6 exposures and survival in terrestrial vertebrates and that there is a causal relationship
7 between Pb exposures and survival in terrestrial invertebrates. The evidence is inadequate
8 to conclude that there is a causal relationship between Pb exposures and survival in
9 terrestrial plants.
7.3.12.4 Neurobehavioral Effects-Terrestrial Biota
10 The central nervous system of animals was recognized as a target of Pb toxicity in the
11 1977 Pb AQCD (U.S. EPA. 1977). and subsequent Pb reviews have provided additional
12 supporting evidence of Pb as a neurotoxicant in terrestrial invertebrates and vertebrates.
13 Effects of Pb on neurological endpoints in terrestrial animal taxa include changes in
14 behaviors that may decrease the overall fitness of the organism such as food
15 consumption, prey capture ability and avoidance behaviors.
16 Some organisms exhibit behavioral avoidance while others do not seem to detect the
17 presence of Pb (U.S. EPA. 2006c). Decreased food consumption of Pb-contaminated diet
18 has been demonstrated in some invertebrates (snails) and vertebrates (lizards, pigs).
19 Decreased food consumption were observed in juvenile A. achatina snails exposed to
20 Pb-contaminated (concentration greater than 134 mg Pb/kg) diet for 12 weeks (Ebenso
21 and Ologhobo. 2009a). Similarly, feeding rate in T. pisana snails was depressed in 3
22 week dietary nominal exposures of 50 to 15,000 mg Pb/kg (El-Gendy et al.. 2011). while
23 other snails exposed to Pb at similar concentrations have shown no effects on feeding rate
24 (Beeby and Richmond. 2010). Consumption of 10 mg/Pb kg diet resulted in lower food
25 intake after 120 days of dietary exposure in pigs (S. domestied) (Yu et al.. 2005).
26 In limited studies available on nematodes there is evidence that Pb may affect the ability
27 to escape or avoid predation (Wang and Xing. 2008). Additional new evidence of
28 changes in the morphology of GABA motor neurons was also found in nematodes
29 (C. elegans) (Du and Wang. 2009).
30 Gull chicks experimentally exposed to Pb exhibited abnormal behaviors such as
31 decreased walking, erratic behavioral thermoregulation and food begging that could make
32 them more vulnerable in the wild (Burger and Gochfeld. 2005). Pb was administered via
33 injection to reach a Pb concentration in feathers equivalent to Pb levels in feathers of wild
November 2012 7-83 Draft - Do Not Cite or Quote
-------
1 gull populations. Lizards exposed to Pb through diet of 10 to 20 mg Pb/kg per day for 60
2 days in the laboratory exhibited abnormal coloration and posturing behaviors.
3 These findings in terrestrial invertebrates and vertebrates are coherent with findings from
4 studies in aquatic biota that showed neurobehavioral alterations in various species offish,
5 and also in some aquatic invertebrates (Section 7.4.12.4). They are also coherent with
6 findings in laboratory animals that show that Pb induces changes in learning and memory
7 (Section 5.3.2.3). as well as attention and motor skills (Section 5.3.3.1). New behaviors
8 induced by exposure to Pb reviewed in Chapter_5 that are relevant to effects of Pb
9 observed in terrestrial systems include hyperactivity and mood disorders, effects on
10 visual and auditory sensory systems, changes in structure and function of neurons and
11 supporting cells in the brain, and effects on the blood brain barrier. Mechanisms that
12 include the displacement of physiological cations, oxidative stress and changes in
13 neurotransmitters and receptors are also reviewed. Data from ecological studies are
14 highly coherent with these data from animal experiments, especially neurobehavioral
15 findings and evidence of structural changes. Overall, the evidence from aquatic and
16 terrestrial systems is sufficient to conclude that a causal relationship is likely to exist
17 between Pb exposures and neurobehavioral effects in invertebrates and vertebrates in
18 terrestrial ecosystems.
7.3.12.5 Hematological Effects-Terrestrial Biota
19 Hematological responses are commonly reported effects of Pb exposure in vertebrates in
20 terrestrial systems. In the 1977 Pb AQCD, ALAD was recognized as the most sensitive
21 indicator of Pb exposure in rats (U.S. EPA. 1977). Furthermore, inhibition of ALAD was
22 associated with death of waterfowl following ingestion of Pb shot. In the 1986 Pb AQCD,
23 decreases in red blood cell ALAD activity were documented in birds and mammals near a
24 smelter (Beyer et al.. 1985). Additional evidence for effects on blood parameters and
25 their applicability as biomarkers of Pb exposure in terrestrial birds and mammals were
26 presented in the 2005 Ecological Soil Screening Levels for Lead, the 2006 Pb AQCD and
27 the current ISA (U.S. EPA. 2006c. 2005b). Field studies available since the
28 2006 Pb AQCD, include evidence for elevated blood Pb levels correlated with decreased
29 ALAD activity in songbirds and owls living in historical mining areas (Gomez-Ramirez
30 etal., 2011; Hansen et al., 201 la).
31 This evidence is strongly coherent with evidence from freshwater invertebrates and
32 vertebrates (Section 7.4.12.5) and observations from human epidemiologic and animal
33 toxicology studies showing that exposure to Pb induces effects on hematological
34 endpoints, including altered heme synthesis mediated through decreased ALAD and
November 2012 7-84 Draft - Do Not Cite or Quote
-------
1 ferrochelatase activities, decreased red blood cell survival and function, and increased red
2 blood cell oxidative stress. Taken together, the overall weight of human epidemiologic
3 and animal toxicological evidence is sufficient to conclude that a causal relationship
4 exists between Pb exposure and decreased RBC survival and function, and altered heme
5 synthesis in humans and in laboratory animals (Section 5.7). Based on observations in
6 terrestrial birds and mammals and additionally supported by observations in aquatic
7 organisms, and toxicological and epidemiological findings in laboratory animals and
8 humans evidence is sufficient to conclude that there is a causal relationship between Pb
9 exposures and hematological effects in terrestrial vertebrates. The evidence is inadequate
10 to conclude that there is a causal relationship between Pb exposures and hematological
11 effects in terrestrial invertebrates.
7.3.12.6 Physiological Stress-Terrestrial Biota
12 Induction of enzymes associated with oxidative stress response in terrestrial plants,
13 invertebrates and vertebrates is a recognized effect of Pb exposure (U.S. EPA. 2006c).
14 Several studies from the 2006 Pb AQCD in birds and plants provide evidence that Pb
15 induces lipid peroxidation, however, exposures in these studies were higher than would
16 be found generally in the environment (U.S. EPA. 2006c). Building on the body of
17 evidence presented in the 2006 Pb AQCD, recent studies provide evidence of
18 upregulation of antioxidant enzymes and increased lipid peroxidation associated with Pb
19 exposure in many species of plants, invertebrates and vertebrates. In plants, increases of
20 antioxidant enzymes with Pb exposure occur in some terrestrial species at concentrations
21 approaching the average Pb concentrations in U.S. soils (18.9 mg Pb/kg). For example, in
22 a series of studies Wang et al. observed increases in reactive oxygen species with
23 increasing exposure to Pb from 20 mg Pb/kg soil to 2,000 mg Pb/kg in broad bean (V.
24 fabd) (Wangetal..2010c: Wang etal.. 201 Oa: Wang et al.. 2008b) and tomato (L.
25 esculentum) (Wang et al., 2008a). where they were accompanied up to approximately
26 500mg Pb/kg by proportional increases in SOD, glutathione, guaiacol peroxidase, and
27 lipid peroxidation, as well as decreases in catalase. Spinach seedlings grown in soil
28 containing six increasing concentrations of Pb from 20 to 520 mg Pb/kg exhibited higher
29 production of reactive oxygen species, increased rates of lipid peroxidation and increased
30 SOD concentrations. (Wang et al.. 201 la). Markers of oxidative damage are also
31 observed in terrestrial invertebrates, including snails and earthworms, and in terrestrial
32 mammals. Across these biota, there are differences in the induction of antioxidant
33 enzymes that appear to be species-dependent.
November 2012 7-85 Draft - Do Not Cite or Quote
-------
1 The oxidative stress responses associated with Pb exposure in terrestrial plants,
2 invertebrates and vertebrates are consistent with responses in freshwater
3 (Section 7.4.12.6) and saltwater organisms (Section 7.4.21.6). and in humans
4 (Section 5.2.4). This oxidative stress is characterized by increased presence of reactive
5 oxygen species and membrane and lipid peroxidation that can promote tissue damage,
6 cytotoxicity, and dysfunction. Increases in reactive oxygen species are often followed by
7 a compensatory and protective upregulation in antioxidant enzymes, such that this
8 upregulation is itself indicative of oxidative stress conditions. Continuous production of
9 reactive oxygen species may overwhelm this defensive process, leading to further
10 oxidative stress and injury.
11 Upregulation of antioxidant enzymes and increased lipid peroxidation are considered
12 reliable biomarkers of stress, and provide evidence that Pb exposure induces a stress
13 response in those organisms which may itself increase susceptibility to other stressors and
14 reduce individual fitness. Evidence is sufficient to conclude that there is a causal
15 relationship between Pb exposures and physiological stress in terrestrial plants, and that a
16 causal relationship is likely to exist between Pb exposure and physiological stress in
17 terrestrial invertebrates and vertebrates.
7.3.12.7 Community and Ecosystem Level Effects-Terrestrial Biota
18 Most direct evidence of community and ecosystem level effects is from near stationary
19 sources where Pb concentrations are higher than typically observed environmental
20 concentrations for this metal. Impacts of Pb on terrestrial ecosystems near smelters,
21 mines, and other industrial sources have been studied for several decades. Emissions of
22 Pb from smelting and other industrial activities are accompanied by other trace metals
23 (e-g-, Zn, Cu, Cd) and SO2 that may cause toxic effects independently or in concert with
24 Pb. Those impacts include decreases in species diversity and changes in floral and faunal
25 community composition. Ecosystem-level field studies are complicated by the
26 confounding of Pb exposure with other factors such as the presence of trace metals and
27 acidic deposition and the inherent variability in natural systems. In natural systems, Pb is
28 often found co-existing with other stressors, and observed effects may be due to
29 cumulative toxicity.
30 In laboratory and microcosm studies where it is possible to isolate the effect of Pb, this
31 metal has been shown to alter competitive behavior of species, predator-prey interactions
32 and contaminant avoidance. These dynamics may change species abundance and
33 community structure at higher levels of ecological organization. Uptake of Pb into
34 aquatic and terrestrial organisms and subsequent effects on mortality, growth,
November 2012 7-86 Draft - Do Not Cite or Quote
-------
1 physiological stress, blood, neurobehavioral and developmental and reproductive
2 endpoints at the organism level are expected to have ecosystem-level consequences, and
3 thus provide consistency and plausibility for causality in ecosystem-level effects.
4 In the 1977 Pb AQCD the potential for Pb to interfere with ecosystem level processes
5 was explored in a detailed review of a study on the effects of Pb on relationships between
6 arthropods and leaf litter decomposition (U.S. EPA. 1977). Reduced arthropod density,
7 biomass and richness were observed in the vicinity of a Pb smelting complex in Missouri.
8 There were also several studies correlating feeding habits, habitat, and Pb concentrations
9 in body tissues reported in the 1977 Pb AQCD, specifically in insects and small
10 mammals indicating that species differences in Pb concentrations are determined in part
11 by trophic position and habitat preference.
12 In the 1986 Pb AQCD it was reported that Pb at environmental concentrations
13 occasionally found near roadsides and smelters (10,000 to 40,000 mg Pb/kg dry weight
14 [mg Pb/kg]) can eliminate populations of bacteria and fungi on leaf surfaces and in soil
15 (U.S. EPA. 1986b). Some key populations of soil microorganisms and invertebrates die
16 off at 1,000 mg Pb/kg soil interrupting the flow of energy through decomposition
17 processes and altering community structure. At soil concentrations of 500 to 1,000 mg
18 Pb/kg or higher, populations of plants, microorganisms, and invertebrates may shift
19 toward Pb-tolerant populations of the same or different species (U.S. EPA. 1986b).
20 The 2006 Pb AQCD reported that decreased species diversity, changes in floral and
21 faunal community composition and decreased vigor of terrestrial vegetation were
22 observed in ecosystems surrounding former smelters including the Anaconda smelter in
23 southwestern Montana (U.S. EPA. 2006c). Several studies in the 2006 Pb AQCD
24 documented reduced organic matter decomposition rates and decreased microbial
25 biomass in areas heavily polluted by metals. Lower abundance and reduced biodiversity
26 of soil invertebrate communities were observed in field surveys in proximity to Pb
27 stationary sources.
28 Recent evidence published since the 2006 Pb AQCD (summarized in Table 7-4) supports
29 previous findings of a link between high concentration of soil metals and substantial
30 changes in soil microorganism community composition, as well as decreased abundance
31 and diversity. In a naturally Pb-enriched forest in Norway, The number of fungal colony
32 forming units was approximately 10 times lower in the highest Pb soil concentration
33 (~4.5 mg Pb/g dry weight) than in control soils (Baath et al.. 2005). The composition of
34 the fungal community was drastically altered, with only one species common to both
35 soils, and the number of species present was substantially lower.
November 2012 7-87 Draft - Do Not Cite or Quote
-------
1 The effect of Pb on microbial community function has been quantified previously using
2 functional endpoints such as respiration rates, fatty acid production, and soil acid
3 phosphatase and urease activities. These may provide estimate of ecological impacts that
4 emphasize functionality irrespective of microbial diversity or abundance measurements.
5 Studies available since the 2006 Pb AQCD provide further evidence of Pb effects on
6 microbial processes. Pb contamination reduced phenol oxidase activity in several types of
7 soils; concentrations between 5 and 50 nM Pb 0.001 and 0.01 mg Pb/L significantly
8 decreased phenol oxidase activity in all soils tested, while 400 nM (0.08 mg Pb/L) and
9 greater completely arrested phenol oxidase activity in one soil tested (a high pH sandy
10 loam) (Carine et al., 2009). Pb concentrations between 50 and 500 mg Pb/kg significantly
11 reduced microbial abundance and diversity, and also resulted in lower soil phosphatase,
12 urease, and dehydrogenase activities (Gao et al., 2010b). When the microbial properties
13 of metal-contaminated urban soils were compared to those of rural soils, significant
14 differences (Sudova and Vosatka. 2007) were detected in basal community respiration
15 rates and microbial abundance (Yang et al.. 2006). Gai et al. (2011) examined the
16 microbial activity of three soils via microcalorimetric methods following Pb exposure.
17 They noted an increase in activity immediately following Pb application (giving 10, 20,
18 40, 80, and 160 mg Pb/kg), and theorized that this was a result of rapid mortality of
19 sensitive microbial species, followed by a concurrent proliferation of Pb-tolerant
20 microorganisms. As Pb concentrations increased, however, the calculated microbial
21 growth rate constant decreased, indicating a suppression of microbial activity (Gai et al..
22 2011). Akerblom et al. (2007) tested the effects of six metals (Cr, Zn, Mo, Ni, Cd, and
23 Pb) individually. All tested metals had a similar effect on the species composition of the
24 microbial community. Exposure to a high Pb concentration (52 mg Pb/kg) negatively
25 affected respiration rates.
26 In addition to microbial communities, there is new evidence for effects of Pb on other
27 terrestrial ecosystem components. Increased plant diversity was shown to ameliorate
28 effects of Pb contamination on a microbial community (Yang et al.. 2007). The presence
29 of arbuscular mycorrhizal fungi may protect plants growing in Pb-contaminated soils
30 (Bojarczuk and Kieliszewska-Rokicka. 2010; Sudova and Vosatka. 2007). Invertebrates
31 affected by Pb in terrestrial systems may be altering community structure. Recent
32 evidence since the 2006 Pb AQCD, indicates that some species of worms avoid
33 Pb-contaminated soils (Langdon et al., 2005). Reductions in microbial and detritivorous
34 populations can affect the success of their predators (U.S. EPA. 2006c). Following a
35 28-day exposure to field-collected soils contaminated with metals (including Pb at
36 426 mg Pb/kg), both population growth and individual growth of the earthworm
37 L. rubellus were diminished (Klok et al.. 2006). The authors proposed that, although
38 these reductions were unlikely to result in extirpation, avian predators such as the godwit
39 (Limosa limosa) that feed heavily on earthworms may be affected by a reduction of
November 2012 7-88 Draft - Do Not Cite or Quote
-------
1 available earthworm biomass. Furthermore, the presence of earthworms increased Pb
2 uptake by plants (Ruiz etal.. 2011; Sizmur et al.. 2011).
3 In terrestrial ecosystems, most studies show decreases in microorganism abundance,
4 diversity, and function with increasing soil Pb concentrations in areas near point-sources.
5 Specifically, shifts in nematode communities, bacterial species, and fungal diversity have
6 been observed. Most evidence for Pb toxicity to terrestrial plants, invertebrates and
7 vertebrates is from single-species assays in laboratory studies. Although the evidence is
8 strong for effects of Pb on growth (Section 7.3.12.2), reproduction (Section 7.3.12.1) and
9 survival (Section 7.3.12.3) in certain species, considerable uncertainties exist in
10 generalizing effects observed under small-scale, particular conditions up to predicted
11 effects at the ecosystem level of biological organization. In many cases it is difficult to
12 characterize the nature and magnitude of effects and to quantify relationships between
13 ambient concentrations of Pb and ecosystem response due to existence of multiple
14 stressors, variability in field conditions, and to differences in Pb bioavailability at that
15 level of organization. However, the cumulative evidence for Pb effects at higher levels of
16 ecological organization is sufficient to conclude that a causal relationship is likely to exist
17 between Pb exposures and the alteration of species richness, species composition and
18 biodiversity in terrestrial ecosystems.
7.4 Aquatic Ecosystem Effects
7.4.1 Introduction to Effects of Pb on Aquatic Ecosystems
19 This section of the Pb ISA reviews the recent literature published since the
20 2006 Pb AQCD (U.S. EPA. 2006c). on the effects of Pb on freshwater and saltwater
21 ecosystems. Freshwater and marine/estuarine systems are considered separately due to
22 differences in Pb speciation, bioavailability of Pb, salinity, and physiological adaptations
23 of organisms in freshwater versus saltwater environments, as modifying factors for Pb
24 toxicity. The focus is on the effects of Pb to aquatic organisms including algae, aquatic
25 plants, invertebrates, vertebrates, and other biota with an aquatic lifestage
26 (e.g., amphibians). In the freshwater and saltwater sections, aqueous concentrations of Pb
27 are reported as (ig Pb/L and sediment concentrations are in mg Pb/kg.
28 In the present document, studies in some freshwater and saltwater organisms are included
29 where responses are observed at very high Pb concentrations that might not be expected
30 in most environmental scenarios or where the relevance of the exposure method to
31 atmospherically-deposited Pb is unknown. These studies can provide mechanistic
32 information on Pb toxicity, allow for comparison of Pb uptake across taxa, or
November 2012 7-89 Draft - Do Not Cite or Quote
-------
1 demonstrate the wide range of sensitivity among closely-related species. Furthermore,
2 although exposure to Pb in natural systems is likely characterized as a chronic, low dose
3 exposure, it is not always feasible to conduct long-term experiments under natural
4 conditions. Observations from short-term experiments in which high concentrations are
5 used can help to elucidate the shape of concentration-response relationships and provide
6 evidence for a gradient of response to Pb exposure but the extent to which effects would
7 be observed at concentrations of Pb typically found in the environment is uncertain.
8 There are a few studies in the following sections for which effects are reported at very
9 low concentrations of Pb that appear to be below analytical detection limits. These
10 studies are included to the extent that they provide information on responses to Pb.
11 However, the difficulty in maintaining low concentrations of Pb and the potential for
12 contamination limits the interpretation of the reported observations and consideration of
13 the observed effects in the absence of analytical verification. In these cases, less weight is
14 placed on study findings in drawing conclusions regarding the effects of Pb exposure.
15 In the following sections, the literature on aquatic ecosystem effects of Pb, published
16 since the 2006 Pb AQCD, is considered with brief summaries from the 1977 Pb AQCD,
17 the 1986 Pb AQCD and the 2006 Pb AQCD where relevant. Biogeochemistry of Pb in
18 aquatic systems is reviewed in Section 7.4.2. Sections 7.4.3 and 7.4.4 consider the
19 bioavailability and uptake of Pb by freshwater plants, invertebrates, and vertebrates.
20 Biological effects of Pb on freshwater ecosystem components (plants, invertebrates, and
21 vertebrates) are discussed in Section 7.4.5. In this section, effects are generally presented
22 from sub-organismal responses (i.e., enzymatic activities, changes in blood parameters)
23 to endpoints relevant to the population-level and higher (growth, reproduction and
24 survival; summarized in Table 7-5). Biological effects are followed by data on exposure
25 and response of freshwater species (Section 7.4.6). Effects of Pb at the freshwater
26 ecosystem level of biological organization are discussed in Section 7.4.7. Section 7.4
27 includes a discussion of critical loads in freshwater systems (Section 7.4.8).
28 characterization of sensitivity and vulnerability of freshwater ecosystem components
29 (Section 7.4.9) and a discussion of Pb effects on ecosystem services (Section 7.4.10). A
30 synthesis of the new evidence for Pb effects on freshwater organisms (Section 7.4.11) is
31 followed by causal determinations based on evidence dating back to the 1977 Pb AQCD
32 (Section 7.4.12). Corresponding sections on saltwater systems introduced in
33 Section 7.4.13 include bioavailability of Pb in saltwater (Section 7.4.14). biological
34 effects of Pb in saltwater (Section 7.4.15). exposure and response of saltwater species
35 (Section 7.4.16). community and ecosystem level effects (Section 7.4.17) and
36 characterization of sensitivity and vulnerability in saltwater species (Section 7.4.18) and
37 ecosystem services (Section 7.4.19). The saltwater ecosystem section concludes with a
38 synthesis of new evidence for Pb effects in marine/estuarine systems (Section 7.4.20) and
November 2012 7-90 Draft - Do Not Cite or Quote
-------
1 causal determinations based on evidence dating back to earlier AQCDs when available
2 (Section 7.4.21V
7.4.2 Biogeochemistry and Chemical Effects of Pb in Freshwater and Saltwater
Systems
3 Quantifying Pb speciation in aquatic environments is critical for determining the toxicity
4 of the metal to aquatic organisms. As reviewed in the 2006 Pb AQCD (U.S. EPA. 2006b)
5 and discussed in detail in Sections 3.3 and 7.2 of this assessment (Fate and Transport),
6 the speciation process is controlled by many environmental factors. Although aerially
7 deposited Pb largely consists of the labile Pb fraction, once the atmospherically-derived
8 Pb enters surface waters its fate and bioavailability are influenced by Ca2+ concentration,
9 pH, alkalinity, total suspended solids, and dissolved organic carbon (DOC), including
10 humic acids. In sediments, Pb is further influenced by the presence of sulfides and Fe and
11 Mn oxides. For instance, in neutral to acidic aquatic environments, Pb is typically present
12 as PbSO4, PbCl4, Pb2+, cationic forms of Pb hydroxide, and ordinary hydroxide
13 [Pb(OH)2], while in alkaline waters, common forms of Pb include Pb carbonates
14 [Pb(CO3)] and hydroxides [Pb(OH)2]. In addition to these inorganic forms, Pb humate is
15 present in the solid phase and Pb fulvate is present in solution. In freshwater systems, Pb
16 complexes with inorganic OH" and CO32 and forms weak complexes with Cl;
17 conversely, Pb speciation in seawater is a function of chloride concentration and the
18 primary species are PbCl3, PbCO3, PbCl2, and PbCl+. In many, but not all aquatic
19 organisms, Pb dissolved in water can be the primary exposure route to gills or other biotic
20 ligands. The toxicity associated with Pb in the water column or sediment pore waters is
21 directly affected by the competitive binding of Pb to the anions listed above.
22 Currently, national and state ambient water quality criteria for Pb attempt to adjust
23 measured concentrations to better represent the bioavailable free ions, and express the
24 criteria value as a function of the hardness (i.e., amount of Ca2+ and Mg ions) of the water
25 in a specific aquatic system. Models such as the BLM (Figure 7-3) (Paquin et al., 2002;
26 Pi Toro etal.. 2001) include an aquatic speciation model (WHAM V; see below)
27 combined with a model of competitive binding to gill surfaces, and provides a more
28 comprehensive method for expressing Pb concentrations at specific locations in terms of
29 the bioavailable metal. Sediment quality criteria have not been established, although the
30 EPA has developed methods based on equilibrium partitioning theory to estimate
31 sediment benchmarks for Pb and a few other metals (U.S. EPA. 2005d). The approach is
32 based on the ratio of the sum of simultaneously extracted metals and amount of AVS,
33 adjusted for the fraction of organic carbon present in the sediments, and is reviewed in
34 detail in the 2006 Pb AQCD (U.S. EPA. 2006c). It is important to note that this method
November 2012 7-91 Draft - Do Not Cite or Quote
-------
1 cannot accurately predict which sediments are toxic or which metal is the primary risk
2 driver.
3 A more detailed understanding of the biogeochemistry of Pb in aquatic systems (both the
4 water column and sediments) is critical to accurately predicting toxic effects of Pb to
5 aquatic organisms. It should be recognized, however, that in addition to exposure via
6 sediment and water, chronic exposures to Pb also include dietary uptake, even though the
7 toxicokinetics of this exposure pathway are not yet well understood in aquatic organisms
8 and the influence of the bioavailability factors described above is unknown. Furthermore,
9 changes in environmental factors that reduce the bioaccessible Pb fraction can result in
10 either sequestration in sediments or subsequent release as mobile, bioaccessible forms.
11 This section provides updated information about the influence of chemical parameters
12 that affect Pb bioaccessibility in the aquatic environment (in sediments and the water
13 column).
14 Several models are available for estimating the speciation of dissolved Pb. These models
15 were tested by Balistrieri and Blank (2008) by comparing the speciation of dissolved Pb
16 in aquatic systems affected by historical mining activities with that predicted by several
17 models, including Windermere humic aqueous model (WHAM VI), non-ideal
18 competitive absorption Donnan-type model (NICA-Donnan), and Stockholm humic
19 model (SHM). Accurate prediction of labile Pb concentrations was achieved only with
20 SHM, although other metal concentrations were better described by the WHAM model.
21 Whereas both WHAM VI and NICA-Donnan predicted that the bulk of Pb contamination
22 would be complexed with Fe, SHM predicted Pb speciation predominantly characterized
23 by Fe and inorganic Pb complexes. Predicted dynamic Pb concentrations developed with
24 the WHAM VI and NICA-Donnan methods overestimated Pb concentrations measured
25 using diffusive gradients in thin-films in Lake Greifen (Switzerland), but underestimated
26 concentrations in Furbach stream (located in both the Coeur d'Alene and Spokane River
27 Basins in Idaho), indicating that such models may not be able to accurately describe
28 metal speciation under all environmental conditions (Balistrieri and Blank. 2008).
29 Quantification of different sediment metal-binding phases, including sulfide, organic
30 carbon (OC), Fe, and Mn phases, is important to fully understand the bioaccessible
31 fraction of Pb and the toxicity to benthic organisms (Simpson and Batley. 2007).
32 However, physical disturbance, pH change, and even the biota themselves also alter
33 sediment binding or release of Pb. Atkinson et al. (2007) studied the effects of pH on
34 sequestration or release of Pb from sediments. Although high and circumneutral water pH
35 (8.1 and 7.2) did not affect the release of sequestered Pb from sediments, lowering the pH
36 to 6 increased the concentration of Pb in overlying waters from less than 100 (ig Pb/L to
37 200-300 (ig Pb/L. Physical sediment disturbance also increased the amount of sediment-
November 2012 7-92 Draft - Do Not Cite or Quote
-------
1 bound Pb released into the aqueous phase. When Pb-contaminated sediment was
2 physically disturbed, the dissolved oxygen content of the overlying water was observed
3 to significantly impact Pb mobilization, with greater Pb mobilization at lower dissolved
4 oxygen levels (3 to 9 mg/L O2) (Atkinson et al. 2007). In addition, although Pb
5 concentrations in the sediments of a mine-impacted wetland in Hezhang, China, were
6 determined to be strongly associated with organic/sulfide and residual fractions (e.g., 34
7 to 82% of total Pb), the presence of aquatic macrophytes altered the Pb speciation,
8 increasing the fraction of Pb bound to Fe-Mn oxides (42% to 47% of total Pb) (Bi et al..
9 2007). This phenomenon was investigated in greater depth by Sundby et al. (2005). who
10 determined that release of oxygen from macrophyte roots resulted in the oxidation of
11 sediment-bound Pb, leading to the release of bioaccessible Pb fractions (Sundby et al..
12 2005).
7.4.2.1 Other Metals
13 Multiple metals are present simultaneously in many aquatic environments and may
14 interact with one another influencing Pb uptake and toxicity. Interactions of Pb with other
15 metals were reviewed in the 2006 Pb AQCD, and more recent evidence supports previous
16 findings of altered bioavailability associated with metal mixtures. Komjarova and Blust
17 (2008) looked at the effect of the presence of Cd2+ on the uptake of Pb by the freshwater
18 cladoceran Daphnia magna. While Pb uptake rates were not affected by Cu, Ni or Zn,
19 enhanced Pb accumulation was observed in the presence of 0.2 (iM Cd. The highest Pb
20 concentration, 0.25 (iM (51.8 (ig Pb/L) in turn facilitated Cu uptake. Area-specific and
21 whole organism Pb transport rates were greatest in the mid-intestine. It was concluded
22 that Pb-induced disruptions of ion homeostasis and metal absorption processes might be a
23 possible explanation of stimulated Pb uptake in the presence of Cd, as well as the
24 increase in Cu uptake rates provoked by presence of Pb at its highest studied
25 concentration. Komjarova and Blust (2009b) then considered the effect of Na, Ca2+ and
26 pH on simultaneous uptake of Cd, Cu, Ni, Pb and Zn. Cd and Pb showed increased
27 uptake rates at high Na concentration. It was thought that increased Na uptake rates
28 promoted Pb entrance to the cell. With respect to the effect of pH, reduced proton
29 competition begins to influence Pb uptake in waters with high pH. A clear suppression of
30 Cd, Ni, Pb and Zn uptake was observed in the presence of Ca2+ (2.5 mM). Ca2+ has been
31 reported to have a protective effect in other studies (involving other organisms). The
32 presence of other metals may also affect the uptake of Pb by fish. At low concentrations,
33 Cd in a Pb-Cd mixture out-competed Pb at gill tissue binding sites in rainbow trout
34 (Oncorhynchus mykiss), resulting in a less-than additive toxicity when fish were exposed
35 to both metals in tandem (Birceanu et al.. 2008). Evidence for the presence of Pb
November 2012 7-93 Draft - Do Not Cite or Quote
-------
1 influencing the uptake of other metals was observed in the marine bivalves Macomona
1 hliana and Austrovenus stutchburyi. Significantly, more Zn bioaccumulated in the
3 presence of Pb in these mussels than with Zn alone following a 10-day exposure to
4 spiked sediments (Fukunaga and Anderson. 2011).
7.4.2.2 Biofilm
5 Farag et al. (2007) measured Pb concentrations in various media (water, colloids,
6 sediment, biofilm) as well as invertebrates and fish collected within the Boulder River
7 watershed, MT, U.S. They concluded that the fraction of Pb associated with Fe-oxides
8 was most frequently transferred to biofilms and the other biological components of the
9 sampled systems (Farag et al.. 2007). Consequently, an increase in the Pb Fe-oxide
10 fraction could signify a potential increase in the bioaccessible pool of Pb. The authors
11 also noted that this fraction may promote downstream transport of Pb contamination.
12 Ancion et al. (2010) investigated whether urban runoff metal contaminants could modify
13 biofilm bacterial community structure and diversity and therefore potentially alter the
14 function of biofilms in stream ecosystems. They found that accumulation rates for metals
15 in biofilm were maximal during the first day of exposure and then decreased with time.
16 Equilibrium between metal concentrations in the water and in the biofilm was reached for
17 all metals after 7-14 days of exposure. The affinity of the biofilm for Pb was, however,
18 much greater than for Cu and Zn. With respect to recovery, the release of metals was
19 slow and after 14 days in clean water 35% of Pb remained in the biofilm. By retaining
20 and releasing such metal pollutants, biofilms may play a key role in determining both the
21 concentration of the dissolved metals in the water column and the transfer of the metals
22 to invertebrates and fish grazing on them. An enrichment factor of 6,000:1 for Pb
23 between the biofilm and the water was measured after 21 days exposure to synthetic
24 urban runoff. The relatively slow release of such metal may greatly influence the transfer
25 of Pb to organisms feeding on the biofilms. This may be of particular importance during
26 storm events when large amounts of Pb are present in the urban runoff. It was suggested
27 that biofilms constitute an integrative indicator of metal exposure over a period of days to
28 weeks.
November 2012 7-94 Draft - Do Not Cite or Quote
-------
7.4.2.3 Carbonate
1 An investigation of heavy metal concentrations in an industrially impacted French canal
2 (Deule canal) indicated that total extractable Pb in sediments ranged from 27 to
3 10,079 mg Pb/kg, with 52.3% present in Fe-Mn oxide fractions, 26.9% as organic sulfide
4 fraction, 10.7% in carbonates, and 10.1% in the residual fraction (Boughriet et al.. 2007).
5 The relatively high fraction of Pb associated with carbonates was not observed at other
6 sites, as sediments in these areas contained low proportions of carbonates. Hence,
7 addition of carbonates (either from anthropogenic or natural sources) can significantly
8 impact Pb speciation in sediments, and potential bioavailability to resident organisms. In
9 addition, increased surface water carbonate concentrations also reduced the bioaccessible
10 Pb fraction as measured by chronic Pb accumulation in the fathead minnow, (Pimephales
11 promelas) (Mager etal.. 2010). and by Pb toxicity to fathead minnow and the cladoceran
12 (Ceriodaphnia dubid) (Mager et al.. 201 Ib).
7.4.2.4 Dissolved Organic Matter (DOM)
13 Uptake of Pb by water-column organisms is affected by the concentration of DOM
14 (Mager etal.. 201 la; Mager etal.. 2010). In a 7-day chronic study with C. dubia, DOM
15 protected against toxicity while water hardness was not protective (Mager et al., 201 la).
16 The specific composition of DOM has been shown to affect the bioaccessibility of
17 environmental Pb. Humic acid-rich DOM resulted in decreased free Pb ion concentration
18 when compared to systems containing DOM with high concentrations of polysaccharides
19 (Lamelas and Slavevkova. 2008). When the sequestering abilities of various components
20 of DOM were compared, humic acid again was shown to be most efficient at reducing the
21 Pb free ion concentration, followed by fulvic acid, alginic acid, polygalacturonic acid,
22 succinoglycan, and xanthan (Lamelas et al., 2005). Lamelas et al. (2009) considered the
23 effect of humic acid on Pb(II) uptake by freshwater algae taking account of kinetics and
24 cell wall speciation. The uptake flux was described by a Michaelis-Menten type equation.
25 Comparison of Cu(II), Cd(II) and Pb(II) uptake by green freshwater algae, (Chlorella
26 Kessleri), in the presence of either citric acid or humic acid was made. The uptake fluxes,
27 percentage adsorbed and percentage internalized for Cu and Cd were identical in the
28 presence of either citric or humic acid. In contrast, however, there was a ten-fold increase
29 in the respective values for Pb. The increase in adsorbed Pb was attributed to the increase
30 in adsorption sites from the adsorbed humic acid on the surface of the algae. Two
31 hypotheses were considered to explain the increase in internalized Pb and the
32 internalization flux: (1) direct interaction of Pb-humic acid complexes with the
33 internalization sites, and (2) uptake of Pb(II) after dissociation from the Pb-humic acid
34 complex. The authors favor the former hypothesis but no evidence is presented for the
November 2012 7-95 Draft - Do Not Cite or Quote
-------
1 proposed ternary Pb-humic acid-internalized site complexes, nor is there an explanation
2 as to why this behavior is not observed for Cd or Cu.
3 There is evidence, however, that DOC/DOM does not have the same effect on free Pb ion
4 concentration in marine systems as in freshwater systems. No correlation was observed
5 between DOM concentration or composition and Pb toxicity when examined using the
6 sea urchin (Paracentrotus lividus) embryo-larval bioassay (Sanchez-Marin et al.. 2010a).
7 For marine invertebrates, the presence of humic acid increased both the uptake and
8 toxicity of Pb, despite the fact that a larger fraction of Pb is complexed with humic acid
9 (25 to 75%). Although the authors could not provide a precise explanation for this, they
10 theorized that in marine environments, addition of humic acid could induce and enhance
11 uptake of Pb via membrane Ca2+ channels (Sanchez-Marin et al., 2010b). This
12 mechanism was observed in the marine diatom (Thalassiosira weissflogii), in that humic
13 acids absorbed to cell surfaces increased metal uptake; however, water column Pb-humic
14 acid associations did appear to reduce free Pb ion concentrations (Sanchez-Marin et al..
15 201 Ob). Formation of a ternary complex that is better absorbed by biological membranes
16 was another proposed mechanism that could describe the increased bioaccessibility to
17 marine invertebrates of Pb bound to humic acid (Sanchez-Marin et al.. 2007).
18 Sanchez-Marin et al. (2011) subsequently have shown that different components of DOM
19 have different effects on Pb bioavailability in marine systems. Their initial research using
20 Aldrich humic acid found that increasing humic acid concentrations increased Pb uptake
21 by mussel gills and increased toxicity to sea urchin larvae in marine environments
22 (Sanchez-Marin et al., 2007). In contrast, a subsequent investigation found that fulvic
23 acid reduced Pb bioavailability in marine water (Sanchez-Marin et al.. 2011). The
24 contradictory effects of different components of DOM on marine bioavailability likely
25 reflect their distinct physico-chemical characteristics. More hydrophobic than fulvic acid,
26 humic acid may adsorb directly with cell membranes and enhance Pb uptake through
27 some (still unidentified) mechanism (Sanchez-Marin et al.. 2011).
28 As little as 1 (iM of humic acid introduced into surface waters was sufficient to reduce Pb
29 uptake by perennial ryegrass, Lolium perenne, grown in nutrient solution. This resulted
30 from a decrease in the concentration of the free Pb fraction by several orders of
31 magnitude following complexation with the OM. Pb content on the root surface was
32 reduced to 1,658 mg Pb/kg from 4,144 mg Pb/kg following humic acid addition, and
33 relative Pb absorption (absorption in the presence of humic acid divided by absorption in
34 the absence of humic acid) was determined to be approximately 0.2 (Kalis et al.. 2006).
35 Conversely, humic acid may increase the bioaccessible Pb fraction for green algae
36 through formation of a ternary complex that promotes algal uptake of the metal. Lamelas
37 and Slaveykova (2007) found that aqueous Pb formed complexes with humic acid, which
November 2012 7-96 Draft - Do Not Cite or Quote
-------
1 in turn would become adsorbed to C. kesslerii algal surfaces, and that the presence of Pb
2 sorbed to humic acid did not interfere with humic acid-algae complexation. The authors
3 concluded that humic acids bound to algae acted as additional binding sites for Pb, thus
4 increasing the concentrations associated with the algal fraction (Lamelas and Slavevkova.
5 2007).
6 Based on the above, the recent literature indicates the existence of a number of deviations
7 from current models used to predict bioaccessibility of Pb. In marine aquatic systems, for
8 instance, surface water DOM was found to increase (rather than decrease) uptake of Pb
9 by fish gill structures, potentially through the alteration of membrane Ca2+-channel
10 permeability. This phenomenon would not be accurately predicted by a BLM developed
11 using data from freshwater organisms. Further, in both freshwater and marine
12 environments, algal biosorption of labile Pb fraction was also increased by humic acid
13 and DOM, likely through the formation of ternary complexes that increase Pb binding
14 sites on the algal surface. Although it is unclear whether Pb in this form is available for
15 toxic action on algae, it is likely to comprise a significant source of dietary Pb for
16 primary consumers. Moreover, the attempted field verification of freshwater
17 bioaccessibility models was conducted at sites with distinct point-sources of Pb
18 contamination, and only one model (SHM) adequately predicted Pb bioaccessibility.
7.4.2.5 Sulfides
19 In sediments, Pb bioavailability is further influenced by sulfides. In the presence of
20 sulfides, most of the reactive metal in sediments will form insoluble metal sulfide that is
21 not bioavailable for uptake by benthic organisms. Acid volatile sulfide (AVS) has been
22 used to predict the toxicity of Pb and other metals in sediments (Ankley etal. 1996; Di
23 Toro et al.. 1992) and in the development of sediment quality criteria (Section 7.4.3). The
24 role of sulfides in the flux of Pb from sediments is discussed further in Section 3.3.2.3.
7.4.3 Introduction to Bioavailability and Biological Effects of Pb in Freshwater
Ecosystems
25 Freshwater ecosystems across the U.S. encompass many habitats including ponds,
26 streams, rivers, wetlands and lakes. Concentrations of Pb available for fresh surface-
27 water and freshwater sediments are reported in Section 7.2.3 and Table 7-2 and are
28 summarized here. Representative median and range of Pb concentrations in surface
29 waters (median 0.50 (ig Pb/L, range 0.04 to 30 (ig Pb/L), sediments (median 28 mg Pb/kg
30 dry weight, range 0.5 to 12,000 mg Pb/kg dry weight) and fish tissues (geometric mean
November 2012 7-97 Draft - Do Not Cite or Quote
-------
1 0.54 mg Pb/kg dry weight, range 0.08 to 23 mg Pb/kg dry weight [whole body]) in the
2 U.S. based on a synthesis of NAWQA data reported in the previous 2006 Pb AQCD
3 (U.S. EPA. 2006c). Additional information on ambient Pb levels in waters, sediments and
4 biota is presented in Section 3.6.5 and Table 7-2 including new data from the Western
5 Airborne Contaminants Assessment Project (WACAP) on Pb in environmental media and
6 biota from remote ecosystems in the western U.S. WACAP assessed concentrations of
7 semi-volatile organic compounds and metals in up to seven ecosystem components (air,
8 snow, water, sediment, lichen, conifer needles and fish) in watersheds of eight core
9 national parks during a multi-year project conducted from 2002-2007 (Landers et al.,
10 2008). The goals of the study were to assess where these contaminants were
11 accumulating in remote ecosystems in the western U.S., identify ecological receptors for
12 the pollutants, and to determine the source of the air masses most likely to have
13 transported the contaminants to the parks.
14 The 2006 Pb AQCD (U.S. EPA. 2006b) provided an overview of regulatory
15 considerations for water and sediments in addition to consideration of biological effects
16 and major environmental factors that modify the response of aquatic organisms to Pb
17 exposure. Regulatory guidelines for Pb in water and sediments have not changed since
18 the 2006 Pb AQCD, and are summarized below with consideration of limited new
19 information on these criteria since the last review. This section is followed by new
20 information on biogeochemistry, bioavailability and biological effects of Pb since the
21 2006 Pb AQCD.
22 The most recent ambient water quality criteria for Pb in freshwater were released in 1985
23 (U.S. EPA. 1985) by the EPA Office of Water which employed empirical regressions
24 between observed toxicity and water hardness to develop hardness-dependent equations
25 for acute and chronic criterion. These criteria are published pursuant to Section 304(a) of
26 the Clean Water Act and provide guidance to states and tribes to use in adopting water
27 quality standards for the protection of aquatic life and human health in surface water. The
28 ambient water quality criteria for Pb are expressed as a criteria maximum concentration
29 (CMC) for acute toxicity and criterion continuous concentration (CCC) for chronic
30 toxicity (U.S. EPA. 2009b). In freshwater, the CMC is 65 (ig Pb/L and the CCC is
31 2.5 (ig Pb/L at a hardness of 100 mg/L.
32 The 2006 Pb AQCD summarized two approaches for establishing sediment criteria for Pb
33 based on either bulk sediment or equilibrium partitioning (Section 7.2.1 and
34 Section AX7.2.1.4). The first approach is based on empirical correlations between metal
35 concentrations in bulk sediment and associated biological effects to derive threshold
36 effect concentrations (TEC) and probable effects concentrations (PEC) (MacDonald et
37 al., 2000). The TEC/PEC approach derives numeric guidelines to compare against bulk
November 2012 7-98 Draft - Do Not Cite or Quote
-------
1 sediment concentrations of Pb. The other approach in the 2006 Pb AQCD was the
2 equilibrium partitioning procedure published by the EPA for developing sediment criteria
3 for metals (U.S. EPA. 2005d). The equilibrium partitioning approach considers
4 bioavailability by relating sediment toxicity to pore water concentration of metals. The
5 amount of simultaneously extracted metal (SEM) is compared with the metals extracted
6 via AVS since metals that bind to AVS (such as Pb) should not be toxic in sediments
7 where AVS occurs in greater quantities than SEM.
8 Since the 2006 Pb AQCD, both of these methods for estimating sediment criteria for
9 metals, have continued to be used and refined. The SEM approach was further refined in
10 the development of the sediment BLM (Di Toro et al.. 2005). The BLM is discussed
11 further in Sections 7.3.3 and 7.4.4. Comparison of empirical approaches with AVS-SEM
12 in metal contaminated field sediments shows that samples where either method predicted
13 there should be no toxicity due to metals, no toxicity was observed in chronic amphipod
14 exposures (Besser et al.. 2009; MacDonald et al.. 2009). However, when the relationship
15 between invertebrate habitat (epibenthic and benthic) and environmental Pb
16 bioaccumulation was investigated, De Jonge et al. (2010) determined that different
17 environmental fractions of Pb were responsible for invertebrate uptake and exposure. Pb
18 uptake by benthic invertebrate taxa was not significantly correlated to AVS Pb levels, but
19 rather to total sediment concentrations (De Jonge et al.. 2009). Conversely, epibenthic
20 invertebrate Pb body burdens were better correlated to AVS concentrations, rather than
21 total Pb sediment concentrations (De Jonge et al.. 2010).
22 In the following sections, recent information since the 2006 Pb AQCD on Pb in
23 freshwater ecosystems will be presented. Throughout the sections, brief summaries of
24 conclusions from the 1977 Pb AQCD, the 1986 Pb AQCD and 2006 Pb AQCD are
25 included where appropriate. The sections are organized to consider uptake of Pb and
26 effects at the species level, followed by community and ecosystem level effects. New
27 research on the bioavailability and uptake of Pb into freshwater organisms including
28 plants, invertebrates and vertebrates is presented in Section 7.4.4. Effects of Pb on the
29 physiology of freshwater flora and fauna (Section 7.4.5) are followed with data on
30 exposure and response of freshwater organisms (Section 7.4.6). Responses at the
31 community and ecosystem levels of biological organization are reviewed in Section 7.4.7
32 followed by a brief consideration of critical loads in freshwater systems (Section 7.4.8),
33 characterization of sensitivity and vulnerability of ecosystem components (Section 7.4.9)
34 and a discussion of ecosystem services (Section 7.4.10). The freshwater ecosystem
35 section concludes with a synthesis of new evidence (Section 7.4.11) and causal
36 determinations based on evidence dating back to the 1977 Pb AQCD (Section 7.4.12).
November 2012 7-99 Draft - Do Not Cite or Quote
-------
7.4.4 Bioavailability in Freshwater Systems
1 Bioavailability was defined in the 2006 Pb AQCD as "the proportion of a toxin that
2 passes a physiological membrane (the plasma membrane in plants or the gut wall in
3 animals) and reaches a target receptor (cytosol or blood)." In 2007, EPA took cases of
4 bioactive adsorption into consideration and revised the definition of bioavailability as
5 "the extent to which bioaccessible metals absorb onto, or into, and across biological
6 membranes of organisms, expressed as a fraction of the total amount of metal the
7 organism is proximately exposed to (at the sorption surface) during a given time and
8 under defined conditions" (U.S. EPA. 2007c). See Section 7.3.3 for additional discussion
9 of bioavailability.
10 The bioavailability of metals varies widely depending on the physical, chemical, and
11 biological conditions under which an organism is exposed (U.S. EPA. 2007c). The
12 bioavailability of a metal is also dependent upon the bioaccessible fraction of metal. The
13 bioaccessible fraction of a metal is the portion (fraction or percentage) of
14 environmentally available metal that actually interacts at the organism's contact surface
15 and is potentially available for absorption or adsorption by the organism (U.S. EPA.
16 2007c). The processes for evaluating bioavailability and bioaccessibility are presented in
17 Figure 7-2 and in Section 7.3.3. In brief, trace metals, and their complexes, must first
18 diffuse from the external medium to the surface of the organism (mass transport). Metal
19 complexes may dissociate and re-associate in the time that it takes to diffuse to the
20 biological surface. These processes are considered further in Chapter_3. To have an effect
21 on the organism, metals must then react with a sensitive site on the biological membrane
22 (adsorption/desorption processes), often but not necessarily followed by biological
23 transport (internalization). Any of these processes may be the rate limiting step for the
24 overall biouptake process. Internalization is, however, the key step in the overall
25 biouptake process. Although the transport sites often have a high affinity for required
26 metals they do not always have high selectivity and so a toxic metal may bind to the site
27 of an essential metal with a similar ionic radius or co-ordination geometry, e.g., Pb2+,
28 Cd2+ and Zn2+ are similar to Ca2+. At the molecular level, there are three major classes of
29 transition metal transporter: P-type ATPases, Zn regulated transporter/iron-regulated
30 transporter, and natural resistance associated macrophage proteins (Worms et al.. 2006).
31 Of these, natural resistance associated macrophage proteins have been shown to promote
32 the uptake of various metals including Pb. This type of trace metal transport can be
33 described by Michaelis-Menten uptake kinetics and equilibrium considerations.
November 2012 7-100 Draft - Do Not Cite or Quote
-------
Routes of Exposure
1 According to the 2006 Pb AQCD (U.S. EPA. 2006b). Pb adsorption, complexation,
2 chelation, etc., are processes that alter its bioavailability to different aquatic species, and
3 it was suggested that multiple exposure routes may be important in determining overall
4 bioavailability of Pb. Given its low solubility in water, bioaccumulation of Pb by aquatic
5 organisms may preferentially occur via exposure routes other than direct absorption from
6 the water column, including ingestion of contaminated food and water, uptake from
7 sediment pore waters, or incidental ingestion of sediment. If uptake and accumulation are
8 sufficiently faster than depuration and excretion, Pb tissue levels may become sufficiently
9 high to result in physiological effects (Luoma and Rainbow. 2005). Pb accumulation rates
10 are controlled, in part, by metabolic rate. Other factors that influence bioavailability of Pb
11 to organisms in aquatic systems are reviewed in Section 7.4.2. As summarized in the
12 2006 Pb AQCD, organisms exhibit three Pb accumulation strategies: (1) accumulation of
13 significant Pb concentrations with low rate of loss resulting in substantial accumulation;
14 (2) balance between excretion and bioavailable metal in the environment; and (3) very
15 low metal uptake rate without significant excretion, resulting in weak net accumulation
16 (Rainbow. 1996). Uptake experiments with aquatic plants, invertebrates and vertebrates
17 reviewed in the 2006 Pb AQCD showed increases in Pb uptake with increasing Pb in
18 solution. The 2006 Pb AQCD findings included consideration of bioaccumulation in
19 different trophic levels. Pb concentrations were found to be typically higher in algae and
20 benthic organisms and lower in higher trophic-level consumers.
21 In this section:
22 1) Recent information on bioavailability and uptake in algae, plants,
23 invertebrates and vertebrates from freshwater systems are reviewed with
24 summary material from the 2006 Pb AQCD and earlier Pb AQCDs where
25 appropriate.
26 2) An overview of the BLM is presented as the most widely used method for
27 predicting both the bioaccessible and bioavailable fractions of Pb in the
28 aquatic environment. This is followed by a discussion of
29 3) Bioavailability in algae, plants, invertebrates and vertebrates. As reviewed by
30 Wang and Rainbow (2008), aquatic organisms exhibit distinct patterns of
31 metal bioaccumulation. The authors suggest that the observed differences in
32 accumulation, body burden, and elimination between species are due to metal
33 biogeochemistry and physiological and biological responses of the organism.
34 The studies presented below generally support the observations of Wang and
November 2012 7-101 Draft - Do Not Cite or Quote
-------
1 Rainbow (2008) that closely related species can vary greatly in
2 bioaccumulation of Pb and other non-essential metals.
3 The bioaccumulation and toxicity of Pb to aquatic organisms are closely linked to the
4 environmental fate of the metal under variable environmental conditions (Section 3.3) as
5 they are highly dependent upon the relative proportion of free metal ions in the water
6 column. However, information is lacking on the uptake of Pb through ingestion of
7 Pb-sorbed particles or dietary exposure to biologically-incorporated Pb. Such routes of
8 exposure are not included in models such as the BLM that predict toxicity as a function
9 of Pb concentration in the water column. This uncertainty may be greater for Pb than for
10 other more soluble metals (such as Cu) as a greater proportion of the total mass of Pb in
11 an aquatic ecosystem is likely to be bound to particulate matter. Therefore, estimating
12 chronic toxicity of Pb to aquatic receptors may have greater uncertainty than predicting
13 acute effects.
14 BLM Models
15 In addition to the biogeochemical effects that govern the environmental pool of
16 accessible Pb, reactions of Pb with biological surfaces and membranes determines the
17 bioavailability and uptake of the metal by aquatic organisms. The BLM (Figure 7-3)
18 predicts both the bioaccessible and bioavailable fraction of Pb in the aquatic
19 environment, and can be used to estimate the importance of environmental variables such
20 as DOC in limiting uptake by aquatic organisms (Alonso-Castro et al.. 2009). The BLM
21 integrates the binding affinities of various natural ligands in surface waters and the
22 biological uptake rates of aquatic organisms to determine the site-specific toxicity of the
23 bioavailable fraction.
24 In the 2006 Pb AQCD, limitations of the use of BLM in developing air quality criteria
25 were recognized including the focus of this model on acute endpoints and the absence of
26 consideration of dietary uptake as a route of exposure. Atmospheric deposition of Pb to
27 aquatic systems and subsequent effects on ecosystem receptors is likely characterized as a
28 chronic, cumulative exposure rather than an acute exposure. Recommendations from the
29 2006 Pb AQCD included developing both chronic toxicity BLMs and BLMs that
30 consider the dietary route of Pb uptake. The EPA recently incorporated the BLM into the
31 Framework for Metal Risk Assessment (U.S. EPA. 2007c) and has published an ambient
32 freshwater criteria document for Cu based on the BLM model (U.S. EPA. 2007a). This
33 section reviews the literature from the past 5 years on applications of the BLM to
34 predicting bioavailability of Pb to aquatic organisms. However, the primary focus of
35 initial BLMs has been acute toxicity endpoints for fish and invertebrates following gill or
36 cuticular uptake of metals.
November 2012 7-102 Draft - Do Not Cite or Quote
-------
1 Di Toro et al. (2005) constructed BLMs for metals exposure in sediments, surface water,
2 and sediment pore water to determine how to most accurately predict the toxicity of
3 metals-contaminated sediments. Results from models were compared with literature-
4 derived acute toxicity values for benthic and epibenthic invertebrates to establish the
5 accuracy of the developed models. Although the models tended to overestimate the
6 toxicity of aqueous and sediment-bound Pb in freshwater environments, it was
7 determined that the model significantly underestimated Pb toxicity to marine
8 invertebrates (Di Toro et al.. 2005). This may be because pore water metal concentrations
9 were not modeled. Consequently, these results may suggest that either 1) mobilization of
10 Pb concentrations from sediments into pore water is greater in marine environments, or 2)
11 marine invertebrates are significantly more sensitive to Pb exposures than are freshwater
12 species.
13 A number of deviations from results predicted by Pb exposure models (such as the BLM)
14 were documented by Ahlf et al. (2009). They highlighted that uptake of metals by
15 sediment-dwelling bivalves was significantly greater than predicted, because bivalves
16 accumulate Pb from multiple sources not included in the model, such as ingestion of
17 algae, bacteria, and colloidal matter. Species-specific dietary assimilation of ingested
18 particulate-bound metals is also likely to play a role in the toxicity of Pb to aquatic
19 organisms, yet insufficient data are available to permit modeling of this additional factor
20 (Ahlf etal. 2009). The authors outlined the need for additional data in developing
21 bioavailability models for chronic metal exposures. As recent evidence suggests that the
22 hydrophobic DOC fraction (e.g., humic and fulvic acids) sequesters the greatest fraction
23 of Pb in aquatic systems (Pernet-Coudrier et al.. 2011), understanding the influence of
24 this adsorption on Pb toxicity is critical for the prediction of chronic aquatic Pb toxicity.
25 For instance, although the presence of humic acid is considered to reduce the bioavailable
26 fraction of metals in surface water, green algae uptake and biosorption of metals,
27 including Pb, was actually increased by humic acid. The authors determined that humic
28 acid bound to algal surfaces served to increase the total number of metal binding sites
29 over those afforded solely by the algal surface (Lamelas and Slavevkova. 2007). This
30 highlights the complexity of modeling chronic metals bioavailability through multiple
31 exposure routes, as humic acid would decrease gill or cuticular uptake of metals from the
32 water column, but could potentially enhance dietary exposure by increasing algal metal
33 content. Slaveykova and Wilkinson (2005) also noted that humic acid is likely to interact
34 with other biological membranes and alter their permeability to metals, especially in
35 acidic environments. Further, they observed that increased surface water temperatures
36 can not only increase membrane permeability but also change metabolic rates, both of
37 which can enhance metals uptake and assimilation; however, this factor is not included in
38 bioavailability models such as the BLM (Slaveykova and Wilkinson. 2005). Despite this,
November 2012 7-103 Draft - Do Not Cite or Quote
-------
1 the authors noted that, in most cases, the BLM could predict acute metals toxicity with a
2 reasonable degree of accuracy.
7.4.4.1 Freshwater Plants and Algae
3 In the 1977 Pb AQCD, the root system of plants was recognized as the major route of
4 uptake for Pb (U.S. EPA. 1977). Uptake and translocation studies of Pb in plants and
5 algae reviewed in the 1977 Pb AQCD and the 2006 Pb AQCD indicated that plants tend
6 to sequester larger amounts of Pb in their roots than in their shoots. Recent studies on
7 bioavailability of Pb to plants support the findings of the previous Pb AQCDs and
8 provide additional evidence for species-dependent differences in responses to Pb in water
9 and sediments.
10 Most biouptake studies in aquatic plants and algae available since the 2006 Pb AQCD,
11 were typically conducted at very high concentrations of Pb that are not representative of
12 current levels of Pb typically encountered in freshwater. However, most of these
13 exposures included a series of increasing concentrations of Pb and generally, Pb was
14 accumulated in a dose-dependent manner. Studies in which high concentrations of Pb are
15 used and an exposure-response relationship is observed may imply effects at lower
16 concentrations but uncertainty remains to the extent to which effects would be observed
17 at concentrations of Pb typically found in the environment. The role of modifying factors,
18 such as the presence of other metals, on uptake rates as well as species differences in Pb
19 uptake rates can be determined from experimental Pb concentrations that are higher than
20 measured Pb in the environment. Plants that are hyperaccumulators of Pb and other
21 metals may be used for phytoremediation at highly contaminated sites and there is a large
22 body of literature on uptake of very high concentrations of metals by different species.
23 This chapter focuses on environmentally relevant concentrations of Pb and also those
24 studies with doses or exposures in the range of one or two orders of magnitude above
25 current or ambient conditions, as described in the Preamble. In freshwater ecosystems in
26 the U.S., the average Pb concentration in surface water is 0.5 (ig Pb/L (Table 7-2).
27 however, total Pb in water has been measured as high as 2,000 (ig Pb/L where mining
28 and smelting operations have affected streams (Table 3-11).Studies with freshwater algae
29 available since the 2006 Pb AQCD, are primarily limited to nominal media exposures at
30 high concentrations of Pb with metal quantified in tissues. For example, the microalgae
31 Spirulina platensis was demonstrated to accumulate Pb from Zarrouk culture medium in
32 a concentration-dependent manner with nominal initial concentrations of 5,000, 10,000,
33 30,000, 50,000 and 100,000 jig Pb/L (Pb in medium was measured every two days
34 thereafter), following a 10-day incubation period (Arunakumara et al., 2008). Pb
35 concentrations accumulated by algae appeared to decrease when culture time increased
November 2012 7-104 Draft - Do Not Cite or Quote
-------
1 from 2 to 10 days. This may have occurred as a result of a gradual recovery of growth
2 and an addition of biomass that would have reduced the concentration of Pb in algal
3 tissue. An aquatic moss, Fontinalis antipyretica, accumulated up to an average of 622 mg
4 Pb/kg dry weight over a 7-day nominal exposure to 20,700 (ig Pb/L despite saturation of
5 intracellular Pb concentrations after 5 days of exposure (Rau et al., 2007). Interestingly,
6 experimentation with concurrent Cu and Pb exposure indicated that the presence of Cu
7 increased the uptake of Pb by the green algae Chlamydomonas reinhardtii (Chen et al.,
8 201 Oc). The authors noted that, in the case of Cu-Pb binary exposures, uptake rates of Pb
9 exhibited complex non-linear dynamics in other aquatic organisms as well.
10 Additional uptake studies conducted since the 2006 Pb AQCD, include new information
11 for freshwater macrophytes. When exposed to nominal water concentrations of up to
12 20,700 (ig Pb/L, floating (non-rooted) coontail plants (Ceratophyllum demersum)
13 accumulated an average Pb concentration of 1,748 mg Pb/kg after 7 days, although this
14 was not significantly higher than levels accumulated in the first day of exposure (Mishra
15 et al., 2006b). Induction of the antioxidant system improved the tolerance of the aquatic
16 plant Najas indica for bioaccumulated Pb, allowing for increased biomass and the
17 potential to accumulate additional Pb mass. High Pb accumulation (3,554 mg Pb/kg dry
18 weight tissue following a 7-day exposure to 20,720 (ig Pb/L) was considered to be a
19 function of plant morphology; as a submerged, floating plant, N. indica provides a large
20 surface area for the absorption of Pb (Singh etal.. 2010).
21 Given that atmospherically-derived Pb is likely to become sequestered in sediments
22 (Section 7.2). uptake by aquatic macrophytes is a significant route of Pb removal from
23 sediments, and a potential route for Pb mobilization into the aquatic food web. The rooted
24 aquatic macrophyte Eleocharis acicularis was determined to be a hyperaccumulator of
25 Pb in an 11-month bioaccumulation experiment with mine tailings. When grown in
26 sediments containing 1,930 mg Pb/kg, the maximum concentration of Pb in E. acicularis
27 was determined to be 1,120 mg Pb/kg dry weight. However, calculated BCFs for Pb were
28 all less than one, indicating that Pb uptake, although high, was less efficient than for other
29 metals present (Ha et al.. 2009).
30 Aquatic plants inhabiting a wetland containing an average sediment Pb concentration of
31 99 mg Pb/kg exhibited variable Pb tissue concentrations, but these do not appear to be
32 related to macrophyte type (e.g., submerged, floating, emergent, etc.). Consequently, the
33 authors concluded that uptake of Pb by aquatic plants appears to be dependent on species,
34 at the exclusion of habitat or type. For instance, among the submerged plant species,
35 Ceratophyllum demersum accumulated the greatest amount of Pb (22 mg Pb/kg dry
36 weight), while Potamogeton malainus tissue contained the least amount of Pb, 2.4 mg
37 Pb/kg dry weight (Bi et al.. 2007). Tissues of the floating plants Azolla imbricata and
November 2012 7-105 Draft - Do Not Cite or Quote
-------
1 Spirogyra communis were found to contain 12 and 20 mg Pb/kg dry weight, respectively,
2 while emergent macrophytes Scirpus triqueter and Alternantheraphiloxeroides
3 accumulated 1.4 and 10 mg Pb/kg dry weight. Fritioff and Greger (2006) determined that
4 anywhere from 24-59% of the total Pb taken up by Potamogeton natans aquatic plants
5 was sequestered in the cell wall fraction, depending on plant tissue and environmental Pb
6 concentration. More importantly, no translocation of Pb was observed when plant tissues
7 (leaf, stem, root) were exposed to Pb solutions separately (Fritioff and Greger, 2006).
8 Dwivedi et al. (2008) reared nine different species of aquatic plants in a fly-ash
9 contaminated medium containing approximately 7 mg Pb/kg dry weight. Not only did
10 species exhibit different Pb accumulation efficiencies but they also compartmentalized
11 sequestered Pb differently. The submerged macrophyte Hydrilla verticillata accumulated
12 the greatest amount of Pb (approximately 180 mg Pb/kg dry weight tissue), but Pb was
13 sequestered solely in the shoot tissue. In contrast, other plant species accumulated
14 between 15 and 100 mg Pb/kg dry weight (Ranunculus scloralus andMarsilea
15 quadrifolia) with the majority compartmentalizing the metal in root tissue, except for
16 C. demersum andM quadrifolia, which also utilized shoot tissue for Pb storage (Dwivedi
17 et al.. 2008).
18 Pb concentrations in the root, leaf, and stem tissues of three aquatic plant species were
19 found to correlate most closely with the concentration of the exchangeable Pb fraction
20 (e-g-, the fraction of Pb that is easily and freely leachable from the sediment). Authors
21 noted that seasonal variations can alter the amount of Pb present in the exchangeable
22 fraction, and that Pb was more likely than Cd or Cu to remain tightly bound to sediments,
23 and therefore the relationship between total sediment Pb and Pb in aquatic plant tissues
24 was weaker (Ebrahimpour and Mushrifatu 2009).
25 Lemna sp., a free floating macrophyte, incubated in a water extract of waste ash
26 containing 19 (ig Pb/L accumulated 3.5 mg Pb/kg dry weight over 7 days of exposure.
27 Slight toxic effects, including suppression of growth, were observed over this exposure
28 period, but this may have been a result of exposures to multiple metals in the water
29 extract, including Cr, Mn, Cu, and Zn (Horvat et al., 2007). Lemna sp. was also
30 demonstrated to be effective in the biosorption of Pb from solution, even in the presence
31 of sediments (1 g per 700 mL water). Over 7 days of exposure to 3,600 and
32 7,000 (ig Pb/L, plant biomass was found to contain an average of 2,900 and 6,600 mg/kg
33 (wet weight) Pb, respectively, versus 200 and 300 mg/kg (dry weight) in sediment (Kurd
34 and Sternberg. 2008).
35 Young Typha latifolia, another rooted macrophyte, were grown in analytically verified
36 concentrations of 5,000 and 7,500 (ig/L Pb-spiked sediment for 10 days to determine
37 their value as metal accumulators. Within the exposure period, plants exposed to the
November 2012 7-106 Draft - Do Not Cite or Quote
-------
1 lower concentration were able to remove 89% of Pb, while 84% of the Pb present in the
2 higher treatment was taken up by T. latifolia. Pb concentrations measured in root and leaf
3 tissue ranged from 1,365 to 4,867 mg Pb/kg and 272 to 927 mg Pb/kg, respectively, and
4 were higher at the greater Pb exposure (Alonso-Castro et al. 2009).
5 Uptake studies available for aquatic macrophytes since the 2006 Pb AQCD, include some
6 studies where Pb was measured in field collected plants growing in metal-contaminated
7 areas. Common reeds (Phragmites australis) grown in metal-impacted aquatic
8 environments in Sicily, Italy, preferentially accumulated Pb in root and rhizome tissues
9 (Bonanno and Lo Giudice. 2010). Pb concentrations in water and sediment averaged
10 0.4 (ig Pb/L and 2.7 mg Pb/kg. These levels yielded root and rhizome concentrations of
11 17 and 15 mg Pb/kg, respectively, whereas stem and leaf Pb concentrations were lower
12 (9.9 and 13 mg Pb/kg). These tissue concentrations were significantly correlated to both
13 water and sediment concentrations (Bonanno and Lo Giudice. 2010). Conversely, the
14 semi-aquatic plant Ammania baccifera, grown in mine tailings containing 35 to 78 mg
15 Pb/kg, did not accumulate analytically detectable levels of Pb in either root or shoot
16 tissues, despite the fact that other metals (Cu, Ni, Zn) were bioaccumulated (Das and
17 Maiti. 2007). This would indicate that at low/moderate environmental Pb concentrations,
18 some plant species may not bioaccumulate significant (or measurable) levels of Pb.
19 The average concentration of Pb in the tissues of rooted aquatic macrophytes (Callitriche
20 verna, P. natans, C. demersum, Polygonum amphibium, Veronica beccabunga) collected
21 from two metals-polluted streams in Poland (average sediment concentration 38 to 58 mg
22 Pb/kg) was less than 30 mg Pb/kg. Pb bioaccumulation in plants was significantly
23 correlated with sediment Pb concentrations (Samecka-Cymerman and Kempers. 2007). A
24 similar significant correlation was established between reed sweet grass root Pb
25 concentration and sediment Pb concentrations (Skorbiowicz. 2006).
26 Pb tissue concentrations of aquatic plants P. australis and Ludwigia prostrata collected
27 from wetlands containing an average of 52 mg Pb/kg in surficial sediments were
28 predominantly in root tissues, indicating poor translocation of Pb from roots. In the
29 former, Pb decreased from an average of 37 mg Pb/kg in roots to 17, 14, and
30 12 mg Pb/kg in rhizome, stem and leaf tissues, respectively, while L. prostrata Pb tissue
31 concentrations decreased from 77 mg Pb/kg in fibrous root to 7 and 43 mg Pb/kg in stem
32 and leaf tissues (Yang et al.. 2008a). The authors proposed that this diminished transfer
33 ability explained the relatively low BCFs for Pb uptake in these two species, when
34 compared with those of other metals.
35 Despite no significant seasonal effect on surface water Pb concentrations, shining
36 pondweed (Potamogeton lucens), a rooted aquatic macrophyte grown in an urbanized
37 metal-contaminated lake in Turkey, exhibited seasonal alterations in Pb tissue
November 2012 7-107 Draft - Do Not Cite or Quote
-------
1 concentrations. Average water Pb concentrations were 28 (ig Pb/L in spring, 27 (ig Pb/L
2 in summer, and 30 (ig Pb/L in autumn. Over this same time period, root tissue Pb
3 concentrations significantly increased from 6 mg Pb/kg dry weight in spring, to 9 mg
4 Pb/kg dry weight in summer, and to 10 mg Pb/kg dry weight in autumn (Duman et al..
5 2006). No differences were detected in stem Pb concentrations between spring and
6 summer (approximately 4 mg Pb/kg dry weight), but stem Pb concentrations were found
7 to be significantly higher in autumn (6 mg Pb/kg dry weight). In the same system,
8 P. australis plants accumulated the most Pb during winter: 103, 23, and 21 mg Pb/kg dry
9 weight in root, rhizome, and shoot tissue, respectively, in sediments containing 13 mg
10 Pb/kg dry weight. By contrast, Schoenoplectus lacustris accumulated maximum rhizome
11 and stem Pb concentrations of 5.1 and 7.3 mg Pb/kg dry weight in winter, but sequestered
12 the greatest amount of Pb in root tissues during the spring (30 mg Pb/kg dry weight) at a
13 comparable sediment concentration, 18 mg Pb/kg dry weight (Duman et al.. 2007). The
14 authors suggest that this indicated that metal uptake was regulated differently between
15 species.
16 Tree species that inhabit semi-aquatic environments have also been shown to absorb Pb
17 from Pb-contaminated sediments. Bald-cypress trees (Taxodium distichum) growing in
18 sediments of a refinery-impacted bayou in Louisiana accumulated significantly greater
19 amounts of Pb than did trees of the same species growing in bankside soil, despite the
20 lower Pb concentrations of sediments. Bankside soils contained greater than 2,700 mg
21 Pb/kg versus concentrations of 10 to 424 mg Pb/kg in sediments, yet Pb concentrations in
22 trees averaged 4.5 and 7.8 mg Pb/kg tissue, respectively (Devall et al.. 2006). The authors
23 theorized that Pb was more readily released from sediments and that soil dispersion to the
24 swamp sediments provides additional, if periodic, loads of Pb into the system. Willow
25 seedlings planted in Pb-contaminated sediment were more effective at removing Pb from
26 the media than a diffusive gradient in thin film technique predicted (Jakl et al.. 2009).
27 The authors proposed that the plant's active mobilization of nutrients from soil during
28 growth also resulted in increased Pb uptake and sequestration.
29 Given that sediments are a significant sink for Pb entering aquatic systems, it is not
30 surprising that rooted macrophytes bioaccumulate significant quantities of the metal.
31 Although there are some similarities to Pb accumulation observed in terrestrial plants
32 (e-g-, preferential sequestration of the metal in root tissue), Pb appears to be more
33 bioavailable in sediment than it is in soil. This may be a result of differences in plant
34 physiology between aquatic and terrestrial plants (e.g., more rapid growth or more
35 efficient assimilation of nutrients and ions from a water-saturated medium). While rooted
36 macrophytes are likely to be chronic accumulators of Pb sequestered in sediments, aerial
37 deposition of Pb into aquatic systems may result in pulsed inputs of labile Pb that would
38 be available for uptake by floating macrophytes and algae.
November 2012 7-108 Draft - Do Not Cite or Quote
-------
7.4.4.2 Freshwater Invertebrates
1 Uptake and subsequent bioaccumulation of Pb in freshwater invertebrates varies greatly
2 between species and across taxa as previously characterized in the 2006 Pb AQCD. This
3 section expands on the findings from the 1986 Pb AQCD and 2006 Pb AQCD on
4 bioaccumulation and sequestration of Pb in aquatic invertebrates. In the case of
5 invertebrates, Pb can be bioaccumulated from multiple sources, including the water
6 column, sediment, and dietary exposures, and factors such as proportion of bioavailable
7 Pb, lifestage, age, and metabolism can alter the accumulation rate. In this section, new
8 information on Pb uptake from freshwater and sediments by invertebrates will be
9 considered, followed by a discussion on dietary and water routes of exposure and factors
10 that influence species-specific Pb tissue concentrations such as invertebrate habitat and
11 functional feeding group.
12 In a recent uptake study in freshwater mussels available since the 2006 Pb AQCD, the
13 Eastern elliptic mussel (Elliptic complanatd) was shown to accumulate Pb rapidly from
14 water and then reach an equilibrium with exposure level and tissue concentration by two
15 weeks following average daily exposures of 1, 4, 14, 57 or 245 (ig Pb/L as Pb nitrate
16 (Mosher et al. 2012). Tissue concentrations of Pb increased at an exposure-dependent
17 rate for the first 14 days and then did not change significantly for the remainder of the 28-
18 day exposure although mussels continued to accumulate Pb. At the end of the exposure
19 period, average Pb in tissue ranged from 0.33 to 898 mg Pb/kg. The authors concluded
20 that the mussels were likely eliminating Pb via pseudo feces and through storage of Pb in
21 shell.
22 The 2006 Pb AQCD (U.S. EPA. 2006b) summarized studies of uptake of Pb from
23 sediment by aquatic invertebrates and noted that sediment pore water, rather than bulk
24 sediment, is the primary route of exposure. However, a recent study suggests that in the
25 midge, Chironomus riparius, total metal concentrations in bulk sediment are better
26 predictors of metal accumulation than dissolved metal concentrations in sediment pore
27 water based on bioaccumulation studies using contaminated sediments from six different
28 sites (Roulier et al.. 2008a). Vink (2009) studied six river systems and found that, for a
29 range of metals, uptake by benthic organisms (the oligochaete, Limnodrilus (Family
30 Tubificidae) and the midge, C. riparius) from the sediment pore water (as compared with
31 surface water) was observed only occasionally, and solely for Pb. The physiological
32 mechanisms of Pb uptake are still unclear but it is suggested that uptake and elimination
33 of Pb obey different mechanisms than for other heavy metals.
34 The 2006 Pb AQCD recognized the potential importance of the dietary uptake pathway
35 as a source of Pb exposure for invertebrates. Specifically, in a study with the freshwater
36 amphipod Hyalella azteca, dietary exposure was found to contribute to the chronic
November 2012 7-109 Draft - Do Not Cite or Quote
-------
1 toxicity of Pb, while acute toxicity was unaffected (Besser et al., 2004). Since the
2 2006 Pb AQCD, additional studies have considered the relative importance of water and
3 dietary uptake of Pb in aquatic invertebrates. A stable isotope technique was used to
4 simultaneously measure uptake of environmentally relevant concentrations of Pb
5 (10.4 (ig Pb/L) in the water column by the freshwater cladoceran D. magna directly from
6 water and through food, the green algae Pseudokirchneriella subcapitata. (Komjarova
7 and Blust 2009a). D. magna accumulated the metal from both sources, but the relative
8 proportion of uptake from each source changed over the exposure period. After the first
9 day of exposure, 12% of accumulated Pb was determined to have been absorbed from
10 dietary (algal) sources, but this percentage decreased by day four of exposure to 4%. Pb
11 absorbed from water exposure only resulted in Daphnia body burdens of approximately
12 62.2 mg Pb/kg dry weight (300 (imol Pb/kg dry weight), and was similar to the amount
13 absorbed by algae (Komjarova and Blust. 2009a). In a comparison of dietary and
14 waterborne exposure as sources of Pb to aquatic invertebrates, no correlation between Pb
15 uptake and dietary exposure was observed in the amphipod H. azteca fBorgmann et al..
16 2007).
17 Stable isotope analysis was to used measure uptake and elimination simultaneously in
18 net-spinning caddisfly larvae (Hydropsyche sp.) exposed to aqueous Pb concentrations of
19 0.2 (control) or 0.6 (ig Pb/L for 18 days (Evans et al., 2006). The measured uptake
20 constant for Pb in this study was 7.8 g/dry weight per day, and the elimination rate
21 constant of 0.15/day for Pb-exposed larvae was similar in both presence and absence of
22 the metal in the water. Tissue concentrations ranged from approximately 15 to 35 mg
23 Pb/kg. Hydropsychid Pb BCFs ranged from 41 to 65, and averaged 54, indicating a
24 relatively high accumulation when compared to other metals tested (average BCF of 17
25 for Cd, 7.7 for Cu, and 6.3 for Zn) (Evans et al.. 2006).
26 Recent reports on Pb distribution in freshwater organisms generally support the findings
27 of the 2006 Pb AQCD that Pb is primarily sequestered in the gills, hepatopancreas, and
28 muscle. Uptake of Pb by the crayfish Cher ax destructor exposed to nominal
29 concentration of 5,000 (ig Pb/L as Pb nitrate for 21 days resulted in accumulation at the
30 highest concentration in gill, followed by exoskeleton >mid-gut gland >muscle
31 >hemolymph (Morris et al.. 2005). Body burden analysis following 96 hour nominal
32 exposure to 50, 100 and 500 (ig Pb/L as Pb nitrate in the freshwater snail Biomphalaria
33 glabrata indicated that bioaccumulation increased with increasing concentrations of Pb
34 and the highest levels were detected in the digestive gland (Ansaldo et al.. 2006).
35 When the relationship between invertebrate habitat (epibenthic and benthic) and
36 environmental Pb bioaccumulation was investigated, De Jonge et al. (2010) determined
37 that different environmental fractions of Pb were responsible for invertebrate uptake and
November 2012 7-110 Draft - Do Not Cite or Quote
-------
1 exposure. Pb uptake by benthic invertebrate taxa was not significantly correlated to AVS
2 Pb levels, but rather to total sediment concentrations (De Jonge et al.. 2009). Conversely,
3 epibenthic invertebrate Pb body burdens were better correlated to AVS concentrations,
4 rather than total Pb sediment concentrations (De Jonge et al.. 2010). For instance, the
5 biologically available Pb (e.g., bound to metal-rich granules or metallothioneins)
6 accumulated by the oligochaete Tubifex tubifex was determined to correlate with
7 sediment SEM-AVS Pb concentrations (De Jonge et al., 2011). Similarly, Desrosiers et
8 al. (2008) reported that Pb accumulation by chironomid larvae from St. Lawrence river
9 sediments was significantly correlated to both total Pb and reactive Pb sediment
10 concentrations.
11 Both inter- and intra-specific difference in Pb uptake and bioaccumulation may occur in
12 macroinvertebrates of the same functional-feeding group. Cid et al. (2010) reported
13 significant differences in Pb bioaccumulation between field collected Ephoron virgo
14 mayflies and Hydro psyche sp, caddisflies, with only the mayfly exhibiting increased Pb
15 tissue concentrations when collected from Pb-contaminated sites; the caddisfly Pb tissue
16 concentrations were similar between reference and Pb-contaminated areas. The authors
17 also examined the lifestage specific accumulation of Pb for E. virgo mayflies, and
18 although there was no statistical difference in Pb tissue concentrations between different
19 lifestages, Pb bioaccumulation did change as mayflies aged (Cid et al., 2010).
20 Reported BAF values for Pb in aquatic invertebrates from the 2006 Pb AQCD ranged
21 from 499 to 3,670 [Table AX7-2.3.2 (U.S. EPA. 2006c)1. Since the 2006 Pb AQCD,
22 additional BAF values have been established for invertebrates in field studies which tend
23 to be higher than BCF values calculated in laboratory exposures (Casas et al.. 2008;
24 Gagnon and Fisher. 1997). A complicating factor in establishing BAF values is that
25 laboratory studies usually assess uptake in water-only or sediment only exposures while
26 field studies take into account dietary sources of Pb as well as waterborne Pb resulting in
27 BAF values that are frequently 100-1,000 times larger than BCF values for the same
28 metal and species (DeForest et al., 2007). The EPA Framework for Metals Risk
29 Assessment states that the latest scientific data on bioaccumulation do not currently
30 support the use of BCFs and BAFs when applied as generic threshold criteria for the
31 hazard potential of metals (U.S. EPA. 2007c). See Section 7.3.3 for further discussion.
32 As reviewed by Wang and Rainbow (2008) and supported by additional studies reviewed
33 in the present document, there are considerable differences between species in the
34 amount of Pb taken up from the environment and in the levels of Pb retained in the
35 organism. The bioaccumulation and subsequent toxicity of Pb to aquatic organisms
36 (Section 7.4.5) are closely linked to the environmental fate of the metal under variable
November 2012 7-111 Draft - Do Not Cite or Quote
-------
1 environmental conditions (Sections 3.3 and 7.2) as they are highly dependent upon the
2 proportion of free metal ions in the water column.
7.4.4.3 Freshwater Vertebrates
3 Uptake of Pb by vertebrates considered here includes data from fish species as well as a
4 limited amount of new information on amphibians and aquatic mammals. The
5 bioaccessibility and bioavailability of Pb is affected by abiotic and biotic modifying
6 factors considered in Sections 7.4.2 and 7.4.4. In fish, Pb is taken up from water via the
7 gills and from food via ingestion. Amphibians and aquatic mammals are exposed to
8 waterborne Pb primarily through dietary sources. In the 2006 Pb AQCD, dietary Pb was
9 recognized as a potentially significant source of exposure to all vertebrates since Pb
10 adsorbed to food, particulate matter and sediment can be taken up by aquatic organisms.
11 Since the 2006 Pb AQCD, tissue accumulation of Pb via gill and dietary uptake has been
12 further characterized in freshwater fish and new techniques such as the use of stable
13 isotopes have been applied to further elucidate bioaccumulation of Pb. For example,
14 patterns of uptake and subsequent excretion of Pb in fish as measured by isotopic ratios
15 of Pb in each tissue can determine whether exposure was due to relatively long term
16 sources (which favor accumulation in bone) or short term sources (which favors
17 accumulation in liver) (Miller et al.. 2005). Recent information since the 2006 Pb AQCD,
18 on uptake of Pb by fish from freshwater is reviewed below, followed by studies on
19 dietary uptake as a route of Pb exposure. Next, tissue accumulation patterns in fish
20 species are reported with special consideration of the anterior intestine as a newly
21 identified target of Pb from dietary exposures. Finally, studies that report Pb tissue
22 concentrations in amphibians, reptiles and freshwater mammals are considered.
Freshwater Fish
23 Pb uptake in freshwater fish is accomplished largely via direct uptake of dissolved Pb
24 from the water column through gill surfaces and by ingestion of Pb-contaminated diets.
25 According to the data presented in the 2006 Pb AQCD (U.S. EPA. 2006b). accumulation
26 rates of Pb are influenced by both environmental factors, such as water pH, DOC, and
27 Ca2+ concentrations, and by species-dependent factors, such as metabolism, sequestration,
28 and elimination capacities. The effects of these variables on Pb bioaccumulation in fish
29 are largely identical to the effects observed for invertebrates (discussed above).
30 Pb in fish is primarily found in bone, gill, blood, kidney and scales (Spry and Wiener.
31 1991). Since the 2006 Pb AQCD, multiple studies on uptake of Pb from water by fathead
November 2012 7-112 Draft - Do Not Cite or Quote
-------
1 minnow and subsequent tissue distribution have been conducted. Spokas et al. (2006)
2 showed that Pb accumulates to the highest concentration in gill when compared to other
3 tissues over a 24-day exposure. This pattern was also observed in larval fathead minnows
4 exposed to 26 (ig Pb/L for 10-30 days, where gill exhibited the highest Pb concentration
5 compared to carcass, intestine, muscle and liver (Grosell et al., 2006a). In the larval
6 minnows, Pb concentration in the intestine exhibited the highest initial accumulation of
7 all tissues on day 3 but then decreased for the remainder of the experiment while
8 concentrations in the other organs continued to increase. By day 30, gill tissue exhibited
9 the highest Pb concentration (approximately 120 mg Pb/kg), followed by whole fish and
10 carcass (whole fish minus gill, liver, muscle and intestine) Pb concentrations
11 (approximately 70 to 80 mg Pb/kg). However, in considering overall internal Pb body
12 burden, nearly 80% was largely concentrated in the bone tissue, while gill contributed
13 <5%.
14 In another study with fathead minnow, chronic (300 day) exposure to 120 (ig Pb/L
15 resulted in accumulation of approximately 41 mg Pb/kg tissue, although this number was
16 decreased from initial body burdens of greater than 104 mg Pb/kg at test initiation (Mager
17 et al.. 2010). Tissue distribution at 300 days was consistent with Grosell et al. (2006a)
18 with highest concentration in gill, followed by kidney, anterior intestine, and carcass.
19 Addition of humic acid and carbonate both independently reduced uptake of Pb in these
20 fish over the exposure time period. Interestingly, fathead minnow eggs collected daily
21 during 21 day breeding assays that followed the chronic exposure described above
22 accumulated similar levels of Pb from the test solutions regardless of Pb concentration or
23 water chemistry (e.g., addition of humic acid and carbonate) (Mager et al., 2010). Direct
24 acute exposure from water rather than parental transfer accounted for the majority of the
25 Pb accumulation in eggs. Similarly, exposure offish to 32.5 (ig Pb/L in base water for
26 150 days resulted in fathead minnow whole body concentrations of approximately 31 mg
27 Pb/kg, with the most rapid accumulation rate occurring within the first 10 days of
28 exposure, followed by an extended period of equilibrium (Mager etal.. 2008). In this
29 same study, fish were tested in two additional treatments: 36.7 (ig Pb/L in hard water
30 (Ca2+ 500 (iM) or 38.7 (ig Pb/L in humic acid supplemented water (4 mg/L). While the
31 addition of humic acid significantly reduced Pb bioaccumulation in minnows (to
32 approximately 10.4 mg Pb/kg on a whole body basis), Ca2+ sulfate did not alter uptake.
33 Despite the fact that Ca2+-mediated Pb toxicity occurred in larval fathead minnow, there
34 was no concurrent effect on whole body Pb accumulation.
35 Uptake studies in other freshwater teleosts have generally followed the pattern of Pb
36 uptake described above for fathead minnow. In the cichlid, Nile tilapia (Oreochromis
37 niloticus), Pb accumulated significantly in gill (45.9 ±34.4 (ig/g dry weight at
38 2,070 (ig Pb/L), 57.4 ±26.1 (ig/g dry weight at 4,100 (ig Pb/L) and liver (14.3 (ig/g dry
November 2012 7-113 Draft - Do Not Cite or Quote
-------
1 weight at2,070 jig Pb/L) and 10.2 (ig/g dry weight at 4,100 jig Pb/L) during a 14-day
2 nominal exposure (as Pb nitrate) (Atli and Canli. 2008). In rainbow trout exposed to
3 100 (ig Pb/L (as Pb acetate) for 72 hours, the accumulation in tissues was gill >kidney
4 >liver and this same pattern was observed in all concentrations tested
5 (100-10,000 ng Pb/L) (Suicmez et al.. 2006). In contrast to uptake in teleosts, in
6 Pb-uptake studies with the Chondrostei fish Chinese Sturgeon (Acipenser sinensis),
1 muscle tissue accumulated higher levels of Pb than gills (Hou et al., 2011).
8 Sloman et al. (2005) investigated the uptake of Pb in dominant-subordinate pairings of
9 rainbow trout exposed to 46 (ig/L or 325 \ig Pb/L (as Pb nitrate) for 48 hours. Significant
10 Pb accumulation in gill, liver and kidney was only observed in the highest concentration.
11 Pb accumulated preferentially in liver of subordinate trout when compared to dominant
12 trout. Brown trout (Salmo truttd) exposed to aqueous Pb concentrations ranging from 15
13 to 46 (ig Pb/L for 24 days accumulated 6 mg Pb/kg dry weight in gill tissue and Pb
14 concentrations in liver tissue reached 14 mg Pb/kg dry weight. Interestingly, Pb in gill
15 tissue peaked on day 11 and decreased thereafter, while liver Pb concentrations increased
16 steadily over the exposure period, which may indicate translocation of Pb in brown trout
17 from gill to liver (Heier et al., 2009).
18 Zebrafish (Danio rerio) Pb uptake rates from media containing 5.2 (ig Pb/L was
19 significantly increased by neutral pH (versus a pH of 6 or 8) and by Ca2+ concentrations
20 of 0.5 mM; uptake rate of Pb was increased from 10 L/kg-h to 35 L/kg-h by increasing pH
21 from 6 to 7, and from 20 L/kg-h to 35 L/kg-h by increasing Ca2+ concentration from 0.1 to
22 0.5 mM (Komjarova and Blust 2009c). This study also demonstrated that zebrafish gill
23 tissue is the main uptake site for the metal, as Pb concentrations in these tissues were up
24 to eight times as high as that in other tissues.
25 The Eurasian silver crucian carp (Carassius auratus) collected from a pond containing an
26 average of 1,600 mg Pb/kg in the sediments exhibited increased average Pb whole body
27 burden of 36.5 mg Pb/kg dry weight (range 12 to 68 mg Pb/kg dry weight) (Khozhina
28 and Sherriff. 2008). Pb was primarily sequestered in skin, gill, and bone tissues, but was
29 also detected at elevated levels in muscle and liver tissues, as well as in eggs. Two fish
30 species (Labeo rohita and Ctenopharyngodon idella) collected from the Upper Lake of
31 Bhopal, India with average Pb concentration 30 (ig Pb/L in the water column contained
32 elevated Pb tissue concentrations (Malik et al.. 2010). However, while liver and kidney
33 Pb concentrations were similar between the two species (1.5 and 1.1 mg Pb/kg tissue and
34 1.3 and 1.0 mg Pb/kg tissue for C. idella and L. rohita, respectively), they accumulated
35 significantly different amounts of Pb in gill and muscle tissues. C. idella accumulated
36 more than twice the Pb in these tissues (1.6 and 1.3 mg Pb/kg) than did L. rohita (0.5 and
37 0.4 mg Pb/kg).
November 2012 7-114 Draft - Do Not Cite or Quote
-------
1 The studies reviewed above generally support the conclusions of the 2006 Pb AQCD
2 (U.S. EPA. 2006b) that the gill is a major site of Pb uptake in fish and that there are
3 species-dependent differences in the rate and pattern of Pb accumulation. As indicated in
4 the 2006 Pb AQCD, exposure duration can be a factor in Pb uptake from water. In a
5 30-day exposure study, Nile tilapia fmgerlings had a three-fold increase in Pb uptake at
6 the gill on day 30 compared to Pb concentration in gill at day 10 and 20 (Kamaruzzaman
7 et al., 2010). In addition to uptake at the gill, a time-dependent uptake of Pb into kidney
8 in rainbow trout exposed to 570 (ig Pb/L for 96 hours (Patel et al.. 2006) was observed.
9 Pb was accumulated preferentially in the posterior kidney compared to the anterior
10 kidney. A similar pattern was observed by Alves and Wood (2006) in a dietary exposure.
11 In catla (Catla catld) fmgerlings, the accumulation pattern of Pb was kidney >liver >gill
12 >brain >muscle in both 14 day and 60 day Pb exposures (Palaniappan et al., 2009). In
13 multiple studies with fathead minnow at different exposure durations, tissue uptake
14 patterns were similar at 30 days (Grosell et al., 2006a) and 300 days (Mager et al., 2010).
15 In the larval minnows, Pb concentration in the intestine exhibited the highest initial
16 accumulation of all tissues on day 3 but then decreased for the remainder of the
17 experiment while concentrations in the other organs continued to increase (Grosell et al..
18 2006a). By day 30, gill tissue exhibited the highest Pb concentration followed by whole
19 fish and carcass (whole fish minus gill, liver, muscle and intestine). The most rapid rate
20 of Pb accumulation in this species occurs within the first 10 days of exposure (Mager et
21 al.. 2008). African catfish (Clarias gariepinus) exposed to nominal Pb concentrations of
22 50 to 1,000 (ig Pb/L (as Pb nitrate) for 4 weeks accumulated significant amounts of Pb in
23 heart (520-600 mg Pb/kg), liver (150-242 mg Pb/kg), and brain (120-230 mg Pb/kg)
24 tissues (Kudirat 2008). Doubling the exposure time to 8 weeks increased sequestration of
25 Pb in these tissues as well as in skin (125-137.5 mg Pb/kg) and ovaries (30-60 mg Pb/kg).
26 Since the 2006 Pb AQCD, several studies have focused on dietary uptake of Pb in
27 teleosts. Metals have been shown to assimilate differently in tissues depending on the
28 exposure route (Rozon-Ramilo et al.. 2011; Meyer etal.. 2005). Alves et al. (2006)
29 administered a diet of three concentrations of Pb (7, 77 and 520 mg Pb/kg dry weight) to
30 rainbow trout for 21 days. Doses were calculated to be 0.02 (ig Pb/day (control),
31 3.7 (ig Pb/day (low concentration), 39.6 (ig Pb/day (intermediate concentration) and
32 221.5 (ig Pb/day (high concentration). Concentrations in the study were selected to
33 represent environmentally relevant concentrations in prey. After 21 days exposure to the
34 highest concentration, Pb accumulation was greatest in the intestine, followed by carcass,
35 kidney and liver leading the authors to hypothesize that the intestine is the primary site of
36 exposure in dietary uptake of Pb. All tissues, (gill, liver, kidney, intestine, carcass)
37 sequestered Pb in a dose-dependent manner. The gills had the greatest concentration of
38 Pb on day 7(8.0 mg Pb/kg tissue wet weight) and this accumulation decreased to
39 2.2 mg Pb/kg tissue wet weight by the end of the experiment suggesting that the Pb was
November 2012 7-115 Draft - Do Not Cite or Quote
-------
1 excreted or redistributed (Alves et al., 2006). Furthermore, with increasing dietary
2 concentrations, the percentage of Pb retained in the fish decreased. Additionally, in this
3 study red blood cells were identified as a reservoir for dietary Pb. Plasma did not
4 accumulate significant Pb (0.012 mg Pb kg wet weight in the high dose), however, Pb
5 was elevated in blood cells (1.5 mg Pb kg wet weight in the high dose) (Alves et al..
6 2006).
7 Additional studies have supported the anterior intestine as a target for Pb in fish. Nile
8 tilapia exposed to dietary Pb for 60 days (105, 418, and 803 mg Pb/kg dry weight)
9 accumulated the greatest concentration of Pb in the intestine, followed by the stomach
10 and then the liver (Dai et al.. 2009a). The amount of Pb in tissue increased with
11 increasing dietary Pb concentration. In a 42 day chronic study of dietary uptake in
12 rainbow trout, fish fed 45 or 480 mg Pb/kg, accumulated Pb preferentially in anterior
13 intestine (Alves and Wood. 2006). Pb accumulation in the gut was followed by bone,
14 kidney, liver, spleen, gill, carcass, brain and white muscle (Alves and Wood. 2006). Ojo
15 and Wood (2007) investigated the bioavailability of ingested Pb within different
16 compartments of the rainbow trout gut using an in vitro gut sac technique. Although a
17 significant increase in Pb uptake was observed in the mid-intestines, this was determined
18 to be much lower than Pb uptake rates via gill surfaces. However, given that intestinal
19 uptake rate for Pb did not significantly differ from those derived for essential metals
20 (e.g., Cu, Zn, and Ni), this uptake route is likely to be significant when aqueous Pb
21 concentrations are low and absorption via gill surfaces is negligible (Ojo and Wood.
22 2007).
23 Following a chronic 63-day dietary exposure to Pb, male zebrafish had significantly
24 increased Pb body burdens, but did not exhibit any significant impairment when
25 compared with controls. Fish were fed diets consisting of field-collected Nereis
26 diversicolor oligochaetes that contained 1.7 or 33 mg Pb/kg dry weight. This resulted in a
27 daily Pb dose of either 0.1 or 0.4 mg Pb/kg (Boyle etal.. 2010). At the end of the
28 exposure period, tissue from male fish reared on the high-Pb diet contained
29 approximately 0.6 mg Pb/kg wet weight, as compared with approximately 0.48 mg Pb/kg
30 wet weight in the low-Pb dietary exposure group. Pb level was elevated in female fish fed
31 the high-Pb diet, but not significantly so.
32 Ciardullo et al. (2008) examined bioaccumulation of Pb in rainbow trout tissues
33 following a 3-year chronic dietary exposure to the metal. Diet was determined to contain
34 0.19 mg Pb/kg wet weight. Fish skin accumulated the greatest Pb concentrations (0.02 to
35 0.05 mg Pb/kg wet weight), followed by kidney, gills, liver, and muscle. Pb accumulation
36 in muscles (.005 mg Pb/kg) remained constant over all sampled growth stages (Ciardullo
37 et al., 2008). The authors concluded that dietary Pb was poorly absorbed by rainbow
November 2012 7-116 Draft - Do Not Cite or Quote
-------
1 trout. Comparison of dietary and water-borne exposures suggest that although
2 accumulation of Pb can occur from dietary sources, toxicity does not correlate with
3 dietary exposure, but does correlate with gill accumulation from waterborne exposure
4 (Alves et al.. 2006). Comparison of uptake rates across the gut and gill have shown that
5 transporter pathways in the gill have a much higher affinity for Pb than do similar
6 pathways in the gut (Ojo and Wood. 2007).
7 Since the 2006 Pb AQCD, several field studies have considered Pb uptake and
8 bioaccumulation in fish as a tool for environmental assessment. Pb tissue concentrations
9 were elevated in several species offish exposed in the field to Pb from historical mining
10 waste, and blood Pb concentrations were highly correlated with elevated tissue
11 concentrations, suggesting that blood sampling may be a useful and potentially non-lethal
12 monitoring technique (Brumbaugh et al.. 2005).
13 This review of the recent literature indicates that the primary and most efficient mode of
14 Pb absorption for freshwater fish is assimilation of labile Pb via gill surfaces; recent
15 research indicates that chronic dietary Pb exposure may result in some Pb
16 bioaccumulation although it is not the predominant route of exposure. Nevertheless, if
17 benthic invertebrates comprise a large portion offish diets in chronically contaminated
18 systems, assimilated Pb loads may be significant. This was demonstrated by Boyle et al.
19 (2010). who showed that laboratory diets consisting of less than one third field-collected
20 Pb-contaminated invertebrates were sufficient to raise fish tissue Pb levels. However,
21 data from field sites suggest that fish accumulation of Pb from dietary sources is highly
22 variable and may be strongly dependent on the physiology of individual species and
23 absorption capacities.
Amphibians
24 Since the 2006 Pb AQCD, there are a few recent field measurements and laboratory-
25 based studies that consider uptake of Pb in amphibians. Whole body Pb measured in three
26 species of field-collected tadpoles in the Mobile-Tensaw River Delta in Alabama
27 averaged 1.19 mg Pb/kg dry weight in Rana clamitans, 0.65 mg Pb/kg dry weight in
28 Rana catesbeiana and 1.32 mg Pb/kg dry weight in Hyla cinerea fAlbrecht et al.. 2007 j.
29 Blood-Pb levels in Ozark hellbender salamanders (Cryptobranchus alleganiensis
30 bishopi), a candidate species for the Endangered Species Act, ranged from 0.044 to
31 0.055 mg/kg dry whole blood weight, in three rivers in Missouri (Huang et al., 2010). In
32 the same study, Pb-blood levels were measured from Eastern hellbenders
33 (Cryptobranchus alleganiensis alleganiensis), a species of concern, collected from four
34 rivers and ranged from 0.075 to 0.088 mg Pb/kg dry whole blood weight.
November 2012 7-117 Draft - Do Not Cite or Quote
-------
1 In a chronic laboratory-based study with tadpoles of the Northern Leopard frog (Rana
1 pipiens), Pb tissue concentrations were evaluated following exposures to 3, 10, and
3 100 (ig Pb/L from embryo to metamorphosis. The tadpole tissue concentrations ranged
4 from 0.1 to 224.5 mg Pb/kg dry mass and were positively correlated to Pb concentrations
5 in the water (Chen et al., 2006b). Dose-dependent bioaccumulation of Pb was observed in
6 the livers of tadpoles of the African clawed frog (Xenopus laevis) exposed to nominal
7 concentrations ranging from 1.0 to 30,000 (ig Pb/L (3 to 115 mg Pb/kg wet weight) for
8 12 days (Mouchet et al.. 2007). Pb concentrations were measured in livers, bodies
9 without liver and whole bodies in Southern leopard frog (Rana sphenocephald) tadpoles
10 exposed to Pb in sediment (45 to 7,580 mg Pb/kg dry weight) with corresponding pore
11 water concentrations of 123 to 24,427 (ig Pb/L from embryonic stage to metamorphosis
12 (Sparling et al.. 2006). There was 100% mortality at 3,940 mg Pb/kg and higher. In all
13 body residues analyzed there was a significant positive correlation between Pb in
14 sediment and Pb in sediment pore water. Concentrations of Pb in liver were similar to
15 results with whole body and bodies without liver indicating that Pb is not preferentially
16 sequestered in liver.
Reptiles
17 Recent field surveys of Pb in water snakes since the 2006 Pb AQCD, indicate that Pb is
18 bioaccumulated in several species. Water snakes spend time in terrestrial and aquatic
19 habitats and could potentially be exposed to atmospherically deposited-Pb in both
20 environments. Average Pb levels in whole body samples of Eastern Ribbon Snakes
21 (Thamnophis sauntus) collected from the Mobile-Tensaw River, a large watershed that
22 drains more than 75% of Alabama were 0.35 ± 0.12 mg Pb/kg dry weight) (Albrecht et
23 al.. 2007). Burger et al. (2007) measured Pb levels in blood, kidney, liver, muscle and
24 skin from water snakes, (Nerodia sepedori) collected from an urban/suburban canal in
25 New Jersey. Pb was highest in skin (0.467 mg Pb/kg wet weight) followed by kidney
26 (0.343 mg Pb/kg wet weight) blood (0.108 mg Pb/kg wet weight), muscle (0.103 mg
27 Pb/kg wet weight) and liver (0.063 mg Pb/kg wet weight). No interspecies differences
28 were observed in blood Pb (range 0.04 to 0.1 mg Pb/kg) from field-collected banded
29 water snakes (Nerodia fasciata), brown water snakes (N. taxispilota) and cottonmouth
30 (Agkistrodon piscivorus) from a reference area and an area contaminated by chemical and
31 radiation releases from the 1950's to the 1980's at the Department of Energy's Savannah
32 River site in South Carolina (Burger et al.. 2006). Cottonmouth and brown water snake
33 from the exposed site had significantly higher levels of Pb in tail muscle when compared
34 to the reference creek.
November 2012 7-118 Draft - Do Not Cite or Quote
-------
Mammals
1 Pb bone levels in Eurasian otters (Lutra lutrd) measured in dead individuals collected in
2 southwest England fell by 73% between 1992 and 2004 (Chadwicket al.. 2011). Annual
3 mean bone Pb levels were 446 (ig Pb/kg in 1992 and 65 (ig Pb/kg in 2004. The 73%
4 decline of Pb in otter bones from 1992 to 2004 was found to coincide with legislative
5 controls on Pb emissions implemented in the U.K. starting in 1986. A positive correlation
6 with stream sediment Pb and bone Pb was also observed in this study. The strength of
7 this correlation decreased with increasing Ca2+ in streams.
7.4.4.4 Food Web
8 In the 2006 Pb AQCD, trophic transfer of Pb through aquatic food chains was considered
9 to be negligible (U.S. EPA. 2006c). Concentrations of Pb in the tissues of aquatic
10 organisms were found to be generally higher in algae and benthic organisms and lower in
11 higher trophic-level consumers indicating that Pb was bioaccumulated but not
12 biomagnified (U.S. EPA, 2006c; Eisler. 2000). Recent literature since the
13 2006 Pb AQCD, provides evidence of the potential for Pb to be transferred in aquatic
14 food webs. Other studies indicate Pb is decreased with increasing trophic level. This
15 section incorporates recent literature on transfer of Pb through freshwater aquatic food
16 chains including the application of stable isotope techniques to trace the accumulation
17 and dilution of metals through producers and consumers.
18 Pb was transferred through at least one trophic level in El Niagara reservoir,
19 Aguascalientes, Mexico, a freshwater ecosystem that lacks fishes (Rubio-Franchini et al..
20 2008). Pb was quantified in sediment (0.55 mg Pb/kg to 21 mg Pb/kg), water (5.8 to
21 39 (ig Pb/L), and zooplankton samples of this freshwater system. BAFs were calculated
22 for predatory and grazing zooplanktonic species. The BAF of the rotifer A. brightwellii
23 (BAF 49,300) was up to four times higher than the grazing cladocerans D. similis (BAF
24 9,022) andM micrura (BAF 8,046). According to the authors, since M. micrura are prey
25 for A. brightwellii this may explain the biomagnifications of Pb observed in the predatory
26 rotifer and provides evidence that Pb biomagnifies at intermediate trophic levels.
27 The relative contribution of water and food as source of trace metals including Pb was
28 investigated in the larvae of the alderfly Sialis velata fCroisetiere et al. 2006). Its prey,
29 the midge (C. riparius) was reared in the laboratory and then exposed to trace elements in
30 a metal-contaminated lake for one week prior to being fed to S. velata. During the one-
31 week exposure period of C. riparius to the contaminated water, five of six trace elements,
32 including Pb, reached steady state within C. riparius. Alderfly larvae were held in the lab
33 in uncontaminated lake water and feed one of the treated C. riparius per day for up to six
November 2012 7-119 Draft - Do Not Cite or Quote
-------
1 days to measure Pb uptake via prey. A separate group of alderfly larvae were exposed
2 directly to the contaminated lake water for six days and fed uncontaminated C. riparius
3 while a third group was exposed to Pb via prey and water. Trace metal concentrations in
4 S. velata that consumed contaminated C. riparius increased significantly compared to
5 S. velata in water-only exposures. Food was concluded to be the primary source of Pb
6 (94%) to these organisms, not Pb in the water.
7 The trophic transfer of Pb from the sediment dwelling polychaete worm N. diversicolor
8 to the invertebrate polychaete predator Nereis virens provides additional evidence for
9 assimilation of Pb by a predator and the potential for further transport up the food chain
10 (Rainbow et al.. 2006). N. virens significantly accumulated Pb from a diet of
11 N. diversicolor and there was a significant inverse linear relationship between the trophic
12 transfer coefficient and prey Pb concentration. In the same study, another predator, the
13 decapod Palaemonetes varians, did not significantly accumulate Pb from N. diversicolor
14 indicating that trophic transfer is dependent on species-specific differences in metal
15 assimilation efficiencies and accumulation patterns.
16 In a recent dietary metal study, field-collected invertebrates representing ecologically
17 relevant sources of Pb were fed to zebrafish, to assess bioavailability of this metal via
18 food. The polychaete worm N. diversicolor was collected from two sites; an estuary
19 contaminated with Pb and a reference site with low metal concentrations (Boyle et al..
20 2010). Male zebrafish fed Pb-enriched N. diversicolor had significant increases in whole-
21 body Pb burden when compared to zebrafish fed prey from the reference site, brine
22 shrimp or flake food diets. There was a trend toward increased Pb levels in females under
23 the same dietary regimen. In this study, deposit feeding invertebrates were shown to
24 mobilize sediment-bound metals in the food chain since zebrafish were exposed only to
25 biologically incorporated metal.
26 The concentration of Pb in the tissues of various aquatic organisms was measured during
27 the biomonitoring of mining-impacted stream systems in Missouri. Generally, Pb
28 concentrations decreased with increasing trophic level: detritus contained 20 to 60 mg
29 Pb/kg dry weight, while periphyton and algae contained 1 to 30 mg Pb/kg dry weight;
30 invertebrates and fish collected from the same areas exhibited Pb tissue concentrations of
31 0.1 to 8 mg Pb/kg dry weight (Besser et al.. 2007). In addition, Pb concentrations in
32 invertebrates (snails, crayfish, and other benthos) were negatively correlated with Pb
33 concentrations in detritus, periphyton, and algae. Fish tissue concentrations, however,
34 were consistently correlated only with detritus Pb concentrations (Besser et al.. 2007).
35 Other studies have traced Pb in freshwater aquatic food webs and have found no evidence
36 of biomagnification of Pb with increasing trophic level. Watanabe et al. (2008) observed
37 decreasing Pb concentrations through a stream macroinvertebrate food web in Japan from
November 2012 7-120 Draft - Do Not Cite or Quote
-------
1 producers to primary and secondary consumers. In a Brazilian freshwater coastal lagoon
2 food chain, Pb was significantly higher in invertebrates than in fishes (Pereira et al..
3 2010).
4 Introduction of exotic species into an aquatic food web may alter Pb concentrations at
5 higher tropic levels. In Lake Erie, the invasive round goby (Neogobius melanostomus)
6 and the introduced zebra mussel (Dreissena polymorphd) have created a new benthic
7 pathway for transfer of Pb and other metals (Southward Hogan et al.. 2007). The goby is
8 a predator of the benthic zebra mussel, while the endemic smallmouth bass (Micropterus
9 dolomieui) feed on goby. Since the introduction of goby into the lake, total Pb
10 concentrations have decreased in bass. The authors attribute this decrease of Pb in bass to
11 changes in food web structure, changes in prey contaminant burden or declines in
12 sediment Pb concentrations.
7.4.5 Biological Effects of Pb in Freshwater Systems
13 This section focuses on the studies of biological effects of Pb on freshwater algae, plants,
14 invertebrates, fish and other biota with an aquatic lifestage (e.g., amphibians) published
15 since the 2006 Pb AQCD. Key studies from the 1977 Pb AQCD, the 1986 Pb AQCD and
16 the 2006 Pb AQCD on biological effects of Pb are summarized where appropriate.
17 Waterborne Pb is highly toxic to aquatic organisms with bioavailability and subsequent
18 toxicity varying depending upon the species and lifestage tested, duration of exposure,
19 the form of Pb tested, and water quality characteristics (e.g., pH, alkalinity, DOC)
20 (Sections 7.4.2 and 7.4.3).
21 The 2006 Pb AQCD (U.S. EPA. 2006c) noted that the physiological effects of Pb in
22 aquatic organisms can occur at the biochemical, cellular, and tissue levels of biological
23 organization and include inhibition of heme formation, alterations of blood chemistry,
24 and decreases in enzyme levels. A review of the more recent literature corroborated these
25 findings, and added information about induction of oxidative stress by Pb, alterations in
26 chlorophyll, and changes in production and storage of carbohydrates and proteins. Recent
27 studies available since the 2006 Pb AQCD further consider effects of Pb on reproduction
28 and development, growth and survival of aquatic organisms. Alterations to these
29 endpoints can lead to changes at the community and ecosystem levels of biological
30 organization such as decreased abundance, reduced taxa richness, and shifts in species
31 composition (Section 7.1). Effects on reproduction, growth and survival are reported in
32 additional species with some effects occurring in sensitive freshwater organisms at or
33 near ambient levels of Pb (Table 7-2). Because this review is focused on effects of Pb,
34 studies reviewed for this section include only those for which Pb was the only, or
November 2012 7-121 Draft - Do Not Cite or Quote
-------
1 primary, metal to which the organism was exposed. Areas of research not addressed here
2 include literature related to exposure to Pb from ingestion of shot or pellets. Biological
3 effects of Pb on freshwater algae and plant species are considered below, followed by
4 information on effects on freshwater invertebrates and vertebrates. All reported values are
5 from exposures in which concentrations of Pb were analytically verified unless nominal
6 concentrations are stated.
7.4.5.1 Freshwater Plants and Algae
7 The toxicity of Pb to algae and plants has been recognized in earlier agency reviews of
8 this metal. In the 1977 Pb AQCD, differences in sensitivity to Pb among different species
9 of algae were observed and concentrations of Pb within the algae varied among genera
10 and within a genus (U.S. EPA. 1977). The 1986 Pb AQCD (U.S. EPA. 1986b) reported
11 that some algal species (e.g., Scenedesmus sp.) were found to exhibit physiological
12 changes when exposed to high Pb concentrations in situ. The observed changes included
13 increased numbers of vacuoles, deformations in cell organelles, and increased autolytic
14 activity. Effects of Pb on algae reported in the 2006 Pb AQCD included decreased
15 growth, deformation and disintegration of algae cells, and blocking of the pathways that
16 lead to pigment synthesis, thus affecting photosynthesis. Observations in additional algal
17 species since the 2006 Pb AQCD, support these findings and indicate that Pb exposure is
18 associated with oxidative stress. All of these effects were observed at concentrations of
19 Pb that exceed those found currently in most surface waters (Table 7-2).
20 Recent studies available since the 2006 Pb AQCD, report additional mechanistic
21 information on Pb toxicity to freshwater macrophytes as well as further evidence for
22 effects on oxidative stress and growth endpoints. However, many of these studies were
23 conducted at nominal concentrations of Pb, complicating the comparisons to Pb
24 quantified in surface waters. Furthermore, their relevance to conditions encountered in
25 natural environments is difficult to establish since modifying factors of bioavailability,
26 such as DOC, are often absent from controlled exposures.
27 The effect of Pb exposure on the structure and function of plant photosystem II was
28 studied in giant duckweed, S. polyrrhiza fLing and Hong. 2009J. The Pb concentration of
29 extracted photosystem II particles was found to increase with increasing Pb
30 concentration, and increased Pb concentration was shown to decrease emission peak
31 intensity at 340 nm, amino acid excitation peaks at 230 nm, tyrosine residues, and
32 absorption intensities. This results in decreased efficiency of visible light absorption by
33 affected plants. The authors theorized that Pb2+ may replace either Mg2+ or Ca2+ in
November 2012 7-122 Draft - Do Not Cite or Quote
-------
1 chlorophyll or the oxygen-evolving center, inhibiting photosystem II function through an
2 alteration of chlorophyll structure.
3 Pb exposure in microalgae species has been linked to several effects, including disruption
4 of thylakoid structure and inhibition of growth in both Scenedesmus quadricauda and
5 Anabaena flos-aquae fArunakumara and Zhang. 2008J. Arunakumara et al. (2008)
6 determined the effect of aqueous Pb on the algal species S. platensis using solutions of
7 Pb nitrate. Exposures at 3,440 (ig Pb/L stimulated 10-day algal growth, growth was
8 inhibited at higher concentrations of 6,830, 21,800, 32,800 and 44,500 (ig Pb/L by 5, 40,
9 49, and 78%, respectively. In addition to growth inhibition, algal chlorophyll a and b
10 content were significantly diminished at the three highest Pb exposures (Arunakumara et
11 al., 2008). Although no specific morphological abnormalities were linked to Pb exposure,
12 filament breakage was observed in S. platensis at Pb concentrations >50,000 (ig Pb/L.
13 Since the 2006 Pb AQCD, the production of reactive oxygen species following Pb
14 exposure has been measured directly in cells of the freshwater algae Chlamydomonas
15 reinhardtii at nominal concentrations of Pb as Pb nitrate (0.02 to 52 (ig Pb/L) with the
16 greatest response at 3.15 times more stained cells compared to the control sample
17 following an exposure of 2.5 hours (Szivak et al., 2009). Although this study provides
18 direct evidence for a mechanism of Pb-toxicity at the sub-organism level of biological
19 organization, the relevance of the exposure method to conditions encountered in natural
20 environments is unknown. The concentration data are not reliable in this case since Pb
21 concentrations were not quantified and the lowest reported values are below the
22 analytical detection limit for Pb.
23 At the time of the 1977 Pb AQCD, there was limited information available on Pb effects
24 on aquatic macrophytes. For plants in general, Pb was recognized to affect
25 photosynthesis, mitosis, and growth, however, the majority of studies reporting Pb
26 toxicity were not conducted with plants grown under field conditions (U.S. EPA. 1977).
27 The mechanism for Pb inhibition of photosynthesis was further elucidated in the 1986 Pb
28 AQCD. Additional evidence of Pb effects on plant growth was also observed, however,
29 the available studies were conducted under laboratory conditions at concentrations that
30 exceeded Pb levels in the environment except near smelters or roadsides (U.S. EPA.
31 1986b). In the 1986 Pb AQCD, EC50 values for plant growth were available for several
32 aquatic plants with the lowest EC50 of 1,100 (ig Pb/L in Azolla pinnata exposed to
33 Pb nitrate for 4 days. Effects of Pb on metabolic processes in aquatic plants reviewed in
34 the 2006 Pb AQCD (U.S. EPA. 2006b) included nitrate uptake, nitrogen fixation,
35 ammonium uptake and carbon fixation at concentrations of 20,000 (ig Pb/L and higher.
36 New information is available on Pb effects on oxidative stress endpoints such as changes
37 in antioxidant enzymes, lipid peroxidation and reduced glutathione in aquatic plant,
November 2012 7-123 Draft - Do Not Cite or Quote
-------
1 algae, and moss species exposed to Pb, however most evidence is from studies with high
2 concentrations where Pb was not quantified in the exposure media. An aquatic moss,
3 F. antipyretica, exhibited increased SOD and ascorbate levels following a 2-day exposure
4 to nominal Pb chloride solutions of concentrations of 20, 200, 2,070, 20,700 and
5 207,200 (ig Pb/L. When exposure duration was increased to 7 days, only SOD activity
6 remained significantly increased by Pb exposure (Dazy et al.. 2009). Bell-shaped
7 concentration-response curves were commonly observed for the induction of antioxidant
8 enzymes in F. antipyretica. The chlorophyll, carotenoid, and protein contents of the
9 aquatic macrophyte Elodea canadensis were significantly reduced following Pb
10 accumulation at nominal exposures of 1,000 10,000 and 100,000 (ig Pb/L (Dogan et al..
11 2009). This, along with the induction of some antioxidant systems and the reduction of
12 growth at the highest two exposures, indicated that exposure to the metal caused
13 significant stress, and that toxicity increased with exposure. In addition, native
14 Myriophyllum quitense exhibited elevated antioxidant enzyme activity (glutathione-S-
15 transferase, glutathione reductase, peroxidase) following transplantation in
16 anthropogenically polluted areas containing elevated Pb concentrations. These were
17 correlated with sediment Pb concentrations in the range of 5 to 23 mg Pb/g dry weight
18 (Nimptsch et al.. 2005).
19 Since the 2006 Pb AQCD, toxicity and oxidative stress were also observed in coontail
20 (C. demersum) rooted aquatic macrophytes following 7-day nominal exposures to
21 aqueous Pb 200 to 20,700 (ig Pb/L ,with increasing effects observed with greater
22 exposure concentrations and times. Chlorosis and leaf fragmentation were evident
23 following a 7-day exposure to the highest concentration, while induction of antioxidant
24 enzymes (glutathione, superoxide dismutase, peroxidases, and catalase) was observed at
25 lower exposure concentrations and times. However, as the duration and concentration of
26 Pb exposure was increased, activities of these antioxidant enzymes decreased (Mishra et
27 al.. 2006b).
28 Sobrino et al. (2010) observed reductions in soluble starch stores and proteins with
29 subsequent increases in free sugars and amino acids in Lemna gibba plants exposed
30 nominally to Pb (50,000 to 300,000 (ig Pb/L); total phenols also increased with
31 increasing Pb exposure. Authors noted that this species exhibited similar responses under
32 extreme temperatures, drought, and disease. According to Odjegba and Fasidi (2006).
33 nominal exposure to 18,600 (ig Pb/L as Pb nitrate for 21 days was sufficient to induce a
34 gradual reduction of both chlorophyll and protein content in the macrophyte Eichhornia
35 crassipes. Decreased proteins were theorized to be related to inefficient protein formation
36 following disruption of nitrogen metabolism after Pb exposure (Odjegba and Fasidi.
37 2006). Foliar proline (which is thought to act as an antioxidant) concentrations were
November 2012 7-124 Draft - Do Not Cite or Quote
-------
1 found to increase in a concentration-dependent manner as Pb concentrations increase
2 from 20,720 to 1,036,000 ng Pb/L.
3 Following 72-hour aqueous exposure to 8,495 (ig Pb/L as Pb nitrate, phytochelatin and
4 glutathione concentrations in the freshwater algae Scenedesmus vacuolatus were
5 significantly increased over that of non-exposed algal cultures (Le Faucheur et al.. 2006).
6 The 72-hour Pb exposure also significantly reduced S. vacuolatus growth, and of all the
7 metals tested (Cu, Zn, Ni, Pb, Ag, As, and Sb), Pb was determined to be the most toxic to
8 the algae species. In the algae Chlamydomonas reinhardtii, phytochelatin concentrations
9 were lower than intracellular Pb and not sufficient to bind to accumulated metal
10 following 72-hour exposure (Scheidegger et al.. 2011).
11 In addition to oxidative stress responses, there is new information since the
12 2006 Pb AQCD on growth effects observed at high concentrations of Pb summarized in
13 Table 7-5. Growth effects at the species level can lead to effects at the population-level of
14 biological organization and higher (Section 7.1.1). Root elongation was significantly
15 reduced in a number of wetland plant species (Beckmannia syzigachne, Juncus effusus,
16 Oenanthe javanica, Cyperusflabelliformis, Cyperus malaccensis, and Neyraudia
17 reynaudiana) following nominal Pb exposures of 20,000 (ig Pb/L as Pb nitrate for 21
18 days (Deng et al., 2009). Further, while both Zn and Fe exposures exerted some selective
19 pressure on plants, the authors did not observe the same with Pb, leading them to theorize
20 that concentrations of bioavailable Pb were not present in high enough quantities to have
21 such an effect. Lemna sp. aquatic plants were determined to effectively sequester aqueous
22 Pb at nominal exposures of 5,000 and 10,000 (ig Pb/L in a 7-day experiment, however,
23 15,000 (ig Pb/L resulted in plant mortality (Hurd and Sternberg. 2008). In another study
24 with duckweed, Paczkowska et al. (2007) observed that nominal Pb exposures of 2,070 to
25 20,700 (ig Pb/L for 9 days stimulated the growth of Lemna minor cultures, although there
26 was concurrent evidence of chlorosis and induction of antioxidant enzymes. Additionally,
27 Cd was found to be more toxic than Pb, although the authors determined that this resulted
28 from poor uptake of Pb by L. minor (Paczkowska et al., 2007). Pb exposure (as
29 Pb nitrate) caused oxidative damage, growth inhibition, and decreased biochemical
30 parameters, including photosynthetic pigments, proteins, and monosaccharides, in Wolffia
31 arrhiza plants. Fresh weight of plants was reduced following both 7- and 14-day
32 exposures to Pb concentrations greater than 2,120 (ig Pb/L while chlorophyll a content
33 was decreased at 210 (ig Pb/L and higher (Piotrowska et al.. 2010).
34 Effects of Pb on algae reported in the 2006 Pb AQCD (U.S. EPA. 2006b) included
35 decreased growth, deformation and disintegration of algae cells, and blocking of the
36 pathways that lead to pigment synthesis, thus affecting photosynthesis. Observations in
37 additional algal species since the 2006 Pb AQCD support these findings. Effects on
November 2012 7-125 Draft - Do Not Cite or Quote
-------
1 plants supported by additional evidence in this review and evidence from previous
2 reviews include oxidative damage, decreased photosynthesis and reduced growth.
3 Elevated levels of antioxidant enzymes are commonly observed in aquatic plant, algae,
4 and moss species exposed to Pb. All of the observed effects on aquatic macrophytes and
5 algae occur at concentrations not typically encountered in surface waters of the U.S.
7.4.5.2 Freshwater Invertebrates
6 Few studies on biological effects of Pb in freshwater invertebrates had been conducted at
7 the time of the 1977 Pb AQCD. One study reported an effect on reproduction in Daphnia
8 magna at 30 ng Pb/L (U.S. EPA. 1977). In the 1986 Pb AQCD (U.S. EPA. 1986b).
9 increased mortality was observed in the freshwater snail Lymnaea palustris as low as
10 19 (ig Pb/L and reproductive impairment was reported as low as 27 (ig Pb/L for
11 Daphnia sp. Population-level endpoints of Pb reviewed in the 2006 Pb AQCD included
12 reproduction, growth, and survival. Pb was recognized to be more toxic in longer-term
13 exposures than shorter-term exposures with chronic toxicity thresholds for reproduction
14 in water fleas (D. magna) ranging as low as 30 (ig Pb/L. In aquatic invertebrates, Pb has
15 also been shown to affect stress responses and osmoregulation (U.S. EPA. 2006c). Recent
16 evidence that supports previous findings of Pb effects on reproduction and growth in
17 invertebrates is reviewed here as well as limited studies on behavioral effects associated
18 with Pb exposure. Some of these effects are observed in the range of Pb values found in
19 surveys of U.S. surface waters (median 0.50 (ig Pb/L, range 0.04 to 30 (ig Pb/L), in the
20 U.S. based on a synthesis of NAWQA data reported in the previous 2006 Pb AQCD
21 (U.S. EPA. 2006c) (Table 7-2). The studies are generally presented in this section from
22 responses at the sub-organismal level of biological organization to consideration of
23 endpoints relevant to ecological risk assessment (growth, reproduction, survival).
24 Recent literature strengthens the evidence indicating that Pb affects enzymes and
25 antioxidant activity in aquatic invertebrates. These alterations at the sub-organismal level
26 may serve as biomarkers for effects at the organism level and higher. In invertebrate
27 species that have hemoglobin, ALAD activity can be measured as a biomarker for Pb
28 exposure. In the freshwater gastropod B. glabrata and the freshwater oligochaete
29 Lumbriculus variegatus a significant negative correlation between whole body tissue
30 ALAD enzyme activity and increasing Pb was observed following 48-hour exposure to
31 varying nominal concentrations of the metal (Aisemberg et al.. 2005). The concentration
32 at which 50% of enzyme inhibition was measured was much lower in B. glabrata (23 to
33 29 (ig Pb/L) than in L. variegatus (703 (ig Pb/L). A significant negative correlation was
34 also observed between ALAD activity and metal accumulation by the organisms. Sodium
35 and potassium ATPase (Na+/K+ATPase) activity in gills of Eastern elliptic mussels was
November 2012 7-126 Draft - Do Not Cite or Quote
-------
1 significantly reduced following a 28-day exposure to 57 (ig Pb/L and 245 (ig Pb/L
2 (Mosher et al.. 2012). A significant reduction in Na+ and significant increase in Ca2+ in
3 hemolymph was only observed at the highest concentration.
4 Studies of stress responses to Pb in invertebrates, conducted since the 2006 Pb AQCD,
5 include induction of heat shock proteins and depletion of glycogen reserves. Although
6 these stress responses are correlated with Pb exposure, they are non-specific and may be
7 altered with exposure to any number of environmental stressors. Induction of heat shock
8 proteins in zebra mussel exposed to an average concentration of 574 (ig Pb/L for 10
9 weeks exhibited a 12-fold higher induction rate as compared to control groups (Singer et
10 al.. 2005). Energetic reserves in the freshwater snail B. glabrata in the form of glycogen
11 levels were significantly decreased by 20%, 57% and 78% in gonads compared to control
12 animals following 96-hour exposures to nominal concentrations of 50, 100 and
13 500 (ig Pb/L, respectively (Ansaldo et al.. 2006). Decreases in glycogen levels were also
14 observed in the pulmonary and digestive gland region at 50 and 100 (ig Pb/L treatment
15 levels. Pb did not exacerbate the effects of sustained hypoxia in the crayfish (C.
16 destructor) exposed to 5,000 (ig Pb/L for 14 days while being subjected to decreasing
17 oxygen levels in water (Morris et al.. 2005). The crayfish appeared to cope with Pb by
18 lowering metabolic rates in the presence of the metal.
19 The effect of Pb on osmoregulatory response has been studied since the 2006 Pb AQCD.
20 The combined effect of Pb and hyperosmotic stress on cell volume regulation was
21 analyzed in vivo and in vitro in the freshwater red crab, Dilocarcinus pagei (Amado et
22 al.. 2006). Crabs held in either freshwater or brackish water lost 10% of their body weight
23 after one day when exposed to 2,700 (ig Pb2+/L as Pb nitrate. This weight loss was
24 transient and was not observed during days 2-10 of the exposure. In vitro, muscle from
25 red crabs exposed to hyperosmotic saline solution had increased ninhydrin-positive
26 substances and muscle weight decreased in isosmotic conditions upon exposure to Pb
27 indicating that this metal affects tissue volume regulation in crabs although the exact
28 mechanism is unknown.
29 Behavioral responses of aquatic invertebrates to Pb reviewed in the 2006 Pb AQCD (U.S.
30 EPA. 2006b) included avoidance. A limited number of recent studies have considered
31 additional behavioral endpoints. Feeding rate of the blackworm L. variegatus was
32 significantly suppressed by day 6 of a 10 day sublethal test in Pb-spiked sediments
33 (Penttinen et al.. 2008) as compared to feeding rates at the start of the experiment.
34 However, this decrease of approximately 50% of the initial feeding rate was also
35 observed in the controls; therefore it is likely caused by some other factor other than Pb
36 exposure. Aqueous soil leachates containing multiple metals, including Pb, had no effect
37 on D. magna mobility. Authors noted that although some concentrations (13 to
November 2012 7-127 Draft - Do Not Cite or Quote
-------
1 686 (ig Pb/L) exceeded Canadian Environmental Quality Guidelines, no significant
2 correlation could be established between Pb exposure and D. magna mobility; in fact, the
3 cladocerans were more sensitive to Fe and Al in the leachate than to Pb (Chapman et al.,
4 2010).
5 Alterations in reproductive and developmental endpoints at the species level can lead to
6 effects at the population-level of biological organization and higher (Section 7.1.1). For
7 example, reduced fecundity may lead to a decreased population size and developmental
8 defects can compromise the ability of an organism to escape predation. Recent evidence
9 of reproductive and developmental effects of Pb on freshwater invertebrates available
10 since the 2006 Pb AQCD, include data from previously untested species as well as
11 further characterization of reproductive effects in commonly tested organisms such as
12 Daphnia sp (Table 7-5). However, many of these studies are conducted at nominal Pb
13 concentration complicating direct comparison to Pb quantified in freshwater
14 environments. Sublethal concentrations of Pb negatively affected the total number of
15 eggs, hatching success and embryonic survival of the freshwater snail B. glabrata
16 exposed to nominal concentrations of 50, 100, or 500 (ig Pb/L as Pb nitrate (Ansaldo et
17 al., 2009). Following exposure of adult snails for 96 hours, adults were removed and the
18 eggs were left in the Pb solutions. The total number of eggs was significantly reduced at
19 the highest concentration tested (500 (ig Pb/L). Time to hatching was doubled and
20 embryonic survival was significantly decreased at 50 and 100 (ig Pb/L, while no embryos
21 survived in the highest concentration. Theegala et al. (2007) observed that the rate of
22 reproduction was significantly impaired in Daphnia pulex at >500 (ig Pb/L in 21-day
23 exposures at nominal concentrations of Pb. In a 21-day reproductive test in D. magna the
24 number of neonates born per female was significantly reduced at nominal concentrations
25 of 25, 250, and 2,500 (ig Pb/L (Ha and Choi. 2009). C. dubia reproduction was also
26 impacted by a seven-day exposure to 50 to 500 (ig Pb/L. Both DOC, and, to a lesser
27 degree, alkalinity were observed to ameliorate the effects of Pb on C. dubia reproduction.
28 As DOC increased from 100 (imol C/L to 400 and 600 (imol C/L, the calculated mean
29 EC50 values for C. dubia reproduction increased from approximately 25 \ig Pb/L to
30 200 (ig Pb/L and greater than 500 jig Pb/L, respectively (Mager et al.. 2011 a).
31 Reproductive variables including average lifespan, rate of reproduction, generation time
32 and rate of population increase were adversely affected in the rotifer Brachionus patulus
33 under conditions of increasing turbidity and Pb concentration (Garcia-Garcia et al..
34 2007).
35 In larvae of the mosquito, Culex quinquefasciatus, exposed to 50 (ig Pb/L, 100 (ig Pb/L
36 or 200 (ig Pb/L (as Pb nitrate), exposure was found to significantly reduce hatching rate
37 and egg-production at all concentrations and larval emergence rate at 200 (ig Pb/L
38 (Kitvatanachai et al.. 2005). Larval emergence rates of 78% (FO), 86% (Fl) and 86% (F2)
November 2012 7-128 Draft - Do Not Cite or Quote
-------
1 were observed in the control group while emergence rates decreased in each generation
2 46% (FO), 26% (Fl) and 58% (F2) in mosquitoes reared in a concentration of
3 200 (ig Pb/L. The time to first emergence also increased slightly to 10 days in the
4 Pb-exposed group as compared to the control group where emergence was first observed
5 on day 9. In the F2 generation of parents exposed to 200 (ig Pb/L, the ratio of female to
6 male offspring was 3.6:1.0. No effects were observed on oviposition preference of adult
7 females, larval weight or larval deformation.
8 Impacts to growth can lead to effects at the population-level of biological organization
9 and higher (Section 7.1.1). As noted in the 2006 Pb AQCD (U.S. EPA. 2006b). Pb
10 exposure negatively affects the growth of aquatic invertebrates. Some studies reviewed in
11 the previous Pb AQCD suggested that juveniles do not discriminate between the uptake
12 of essential and non-essential metals (Arai et al.. 2002). In recent literature (summarized
13 in Table 7-5). the freshwater pulmonate snail Lymnaea stagnalis has been identified as a
14 species that is extremely sensitive to Pb exposure. Growth of juveniles was inhibited at
15 EC20 <4 (ig Pb/L. (Grosell and Brix. 2009; Grosell et al.. 2006b). In L. stagnalis exposed
16 to 18.9 (ig/L Pb for 21 days, Ca2+ influx was significantly inhibited and model estimates
17 indicated 83% reduction in growth of newly hatched snails after 30 days at this exposure
18 concentration (Grosell and Brix. 2009). The authors speculate that the high Ca2+ demand
19 of juvenile L. stagnalis for shell formation and interference of the Ca2+uptake pathway by
20 Pb result in the sensitivity of this species.
21 In a study of the combined effects of temperature (22 °C or 32 °C), nominal Pb
22 concentration (50, 100 and 200 (ig Pb/L as Pb chloride) and presence of a competitor, the
23 population growth rate of two freshwater rotifer species, Brachionus havanaensis and
24 B. rubens, as measured by quantifying the number of live rotifers for 15 days, responded
25 to presence of stressors (Montufar-Melendez et al.. 2007). At the lowest temperature,
26 B. rubens suppressed population growth of B. havanaensis at 50 (ig Pb/L and higher and
27 B. rubens population growth did not increase at any Pb concentration at 32 °C, a
28 temperature more suited for B. havanaensis. In situ toxicity testing with the woodland
29 crayfish (Orconectes hylas) indicated that crayfish survival and biomass were
30 significantly lower in streams impacted by Pb mining and that concentrations of Pb and
31 other metals in water, detritus, macroinvertebrates, fish and crayfish were significantly
32 higher at mining sites (Allert et al., 2009a).
33 Although Pb is known to cause mortality when invertebrates are exposed at sufficiently
34 high concentrations, species that are tolerant of Pb may not exhibit significant mortality
35 even at high concentrations of Pb. Odonates are highly tolerant of Pb with no significant
36 differences in survival of dragonfly larvae Pachydiplax longipennis and Erythemis
37 simplicicollis exposed for 7 days to nominal concentrations of Pb as high as
November 2012 7-129 Draft - Do Not Cite or Quote
-------
1 185,000 (ig Pb/L (Tollett et al. 2009). This apparent tolerance to Pb may be even more
2 pronounced in natural environments where the presence of multiple modifying factors
3 (e-g-, pH, alkalinity, hardness, DOC) influences Pb bioavailability. Other species are
4 more sensitive to Pb in the environment and these responses are reviewed in
5 Section 7.4.6.
7.4.5.3 Freshwater Vertebrates
6 The 1977 Pb AQCD reported on Pb effects to domestic animals, wildlife and aquatic
7 vertebrates. The available Pb studies were from exposure to Pb via accidental poisoning
8 or ingestion of Pb shot (U.S. EPA. 1977). Studies on aquatic vertebrates reviewed in the
9 1986 Pb AQCD were limited to hematological, neurological and developmental
10 responses in fish (U.S. EPA. 1986b). In the 2006 Pb AQCD, effects on freshwater
11 vertebrates included recent data for fish specifically considering the effects of water
12 quality parameters on toxicity, as well as limited information on sensitivity of turtles and
13 aquatic stages of frogs to Pb (U.S. EPA. 2006c). Biological effects of Pb on freshwater
14 fish that have been studied since the 2006 Pb AQCD are reviewed here, and limited
15 recent evidence of Pb effects on amphibians are considered. This section presents recent
16 information available on the mechanism of Pb as a neurotoxicant in fish and effects of
17 this metal on blood chemistry. Additional mechanisms of Pb toxicity have been
18 elucidated in the gill and the renal system offish since the 2006 Pb AQCD. Further
19 supporting evidence of reproductive effects of Pb on fish is discussed along with limited
20 new information on behavioral effects of Pb.
Freshwater Fish
21 Evidence of toxicity of Pb and other metals to freshwater fish goes back to early
22 observations whereby contamination of natural areas by Pb mining lead to extirpation of
23 fish from streams (U.S. EPA. 1977). At the time of the 1977 Pb AQCD, documented
24 effects of Pb on fish included anemia, mucous secretion, functional damage to inner
25 organs, physical deformities and growth inhibition. Additionally, the role of temperature,
26 pH, hardness and other water quality parameters on Pb toxicity was discussed in the 1977
27 Pb AQCD. The 1986 Pb AQCD reported that hematological and neurological responses
28 were the most commonly observed effects in fish and the lowest exposure concentration
29 causing either hematological or neurological effects was 8 (ig Pb/L. These findings were
30 additionally supported in the 2006 Pb AQCD, where observed effects of Pb on fish
31 included inhibition of heme formation, alterations in brain receptors, effects on blood
32 chemistry, and decreases in some enzyme activities (U.S. EPA. 2006c). Functional
November 2012 7-130 Draft - Do Not Cite or Quote
-------
1 responses resulting from Pb exposure included increased production of mucus, changes in
2 growth patterns, and gill binding affinities. According to Eisler (2000) and reviewed in
3 the 2006 Pb AQCD, the general symptoms of Pb toxicity in fish include production of
4 excess mucus, lordosis, anemia, darkening of the dorsal tail region, degeneration of the
5 caudal fin, destruction of spinal neurons, ALAD inhibition, growth inhibition, renal
6 pathology, reproductive effects, growth inhibition and mortality.
7 Evidence of Pb effects on fish available since the 2006 Pb AQCD generally supports the
8 findings in previous Pb reviews and further elucidates the mechanisms of Pb-associated
9 toxicity on some physiological responses. At the sub-organism level, new information on
10 Pb effects on DNA, specific enzymes, ionoregulation and other biochemical responses is
11 presented followed by a discussion of new information on population-level endpoints
12 (i.e., growth reproduction summarized in Table 7-5).
13 Since the 2006 Pb AQCD evidence of direct interaction of Pb with fish DNA has become
14 available as well as additional studies on the genotoxic effects of Pb exposure to fish.
15 Hong et al. (2007a) observed covalent binding of Pb with kidney DNA from silver
16 crucian carp (Carassius auratus gibelio) though extended X-ray absorption fine structure
17 spectroscopy. This study suggests that exposure to Pb results in effects to DNA but the
18 exposure method (in vitro) makes it difficult to estimate the natural environmental
19 conditions that would be equivalent to the experimental one. In the freshwater fish
20 Prochilodus lineatus, blood, liver, and gill cells were sampled from fish treated with
21 nominal concentration of 5,000 (ig Pb/L as Pb nitrate for 6, 24 and 96-hours and then
22 DNA damage was assessed by comet assay (Monteiro et al., 2011). DNA breaks were
23 observed in all cell types after 96-hour exposure. The concentrations used in this study
24 were high compared to Pb concentrations currently encountered in freshwater (Table
25 7-2). however, it presents supporting evidence for a possible mechanism of Pb toxicity to
26 fish.
27 Upregulation of antioxidant enzymes in fish is a well-recognized response to Pb
28 exposure. Since the last review, additional studies demonstrating antioxidant activity as
29 well as evidence for production of reactive oxygen species following Pb exposure are
30 available. Silver crucian carp injected with nominal concentration of 10, 20 or 30 mg
31 Pb/kg wet weight Pb chloride showed a significant increase in the rate of production of
32 superoxide ion and hydrogen peroxide in liver (Ling and Hong. 2010). In the same fish,
33 activities of liver SOD, catalase, ascorbate peroxidase, and glutathione peroxidase were
34 significantly inhibited. Both glutathione and ascorbic acid levels decreased and
35 malondialdehyde content increased with increasing Pb dosage, suggesting that lipid
36 peroxidation was occurring and the liver was depleting antioxidants. Although this
37 exposure pathway is unlikely to be relevant for air related deposition of Pb, it provides
November 2012 7-131 Draft - Do Not Cite or Quote
-------
1 evidence for the mechanism of toxicity (production of reactive oxygen species) and the
2 responses of antioxidant enzymes observed in this study are supported by findings in
3 studies from fish from nominal water-only exposures. For example, in the freshwater fish
4 Nile tilapia, liver catalase, liver alkaline phosphatase, Na+/K+ATPase, and muscle
5 Ca2+ATPase activities were quantified in various tissues following a 14-day exposure to
6 nominal concentrations (1,000, 2,000 and 4,000 jig Pb/L )of Pb nitrate (Atli and Canli.
7 2007). Liver catalase activity significantly increased in the 1,000 and 4,000 (ig Pb/L
8 concentrations while liver alkaline phosphatase activity was significantly increased only
9 at the 4,000 (ig Pb/L concentration. No significant change in alkaline phosphatase
10 activity was observed in intestine or serum. Ca2+ATPase activity was significantly
11 decreased in muscle. Na+/K+ATPase was elevated in gill in the highest concentration of
12 Pb while all concentrations resulted in significant decreases of this enzyme in intestine.
13 Serum alanine aminotransferase and aspartate aminotransferase activities were elevated
14 in Nile tilapia exposed to 50 (ig Pb/L in 4 and 21 day aqueous exposures while elevations
15 in alkaline phosphatase and lactate dehydrogenase were only observed at 21 days (Firat et
16 al.. 2011). In another study with Nile tilapia, Pb had no effect on glutathione measured in
17 liver, gill, intestine, muscle and blood and liver metallothionein levels following a 14-day
18 exposure to 1,000, 2,000 and 4,000 jig Pb/L concentrations of Pb as Pb nitrate (Atli and
19 Canli. 2008V
20 Metabolic enzyme activity in teleosts has also been measured following dietary
21 exposures. Alves and Wood (2006) in a 42 day chronic dietary Pb study with 45 and
22 480 mg Pb/kg found that gill Na+/K+ATPase activity was not affected in rainbow trout
23 while increased Na+/K+ATPase was observed in the anterior intestine. Metabolic
24 activities measured in liver and kidney of Nile tilapia following 60 day dietary
25 administration of 100, 400, and 800 mg Pb/kg indicated that alanine transaminase,
26 aspartate transaminase, and lactate dehydrogenase activities significantly decreased in
27 kidney in a concentration-dependent manner (Dai et al., 2009b) and increased in liver
28 with increasing concentration of dietary Pb. In a subsequent study using the same
29 exposure paradigm, the digestive enzymes amylase, trypsin and lipase in tilapia were
30 inhibited by dietary Pb in a concentration-dependent manner (Dai et al.. 2009a). Lesions
31 were also evident in histological sections from livers of Pb-exposed fish from this study
32 and included irregular hepatocytes, cell hypertrophy, and vacuolation although no
33 quantification of lesions by dose-group was presented.
34 There is also evidence for Pb exposure leading to changes in hepatic CYP450 content
35 although relevance of these in vitro and injection studies to air related exposures to Pb is
36 unknown. Pb was shown to inhibit hepatic cytochrome P450 in vitro in carp (C. carpio),
37 silver carp (Hypothalmichtys molitrix) and wels catfish (Silurus glanis) in a
38 concentration-dependent manner from 0 to 4 (ig/mL (Pb2+) (Henczova et al., 2008). The
November 2012 7-132 Draft - Do Not Cite or Quote
-------
1 concentrations of Pb that resulted in 50% inhibition of EROD and 7-ethoxycoumarin-o-
2 deethylase (ECOD) isoenzymes varied with the fish species. Silver carp was the least
3 sensitive to the inhibitory effects of Pb (EROD 1.21, ECOD 1.52 (ig Pb/mL) while carp
4 EROD activity was inhibited at 0.76 (ig Pb/mL. Interaction of Pb with cytochrome P450
5 was verified by spectral changes using Fourier Transform Infrared (FTIR) spectroscopy.
6 In the same study, CYP450 content was elevated and EROD isoenzyme activities were
7 decreased in vivo in silver carp for two days following an injection of 2 mg Pb/kg as
8 Pb acetate and returned to control values by 6 days. Liver damage to African catfish
9 exposed to nominal concentrations of Pb (50-1,000 (ig Pb/L) for 4 or 8 weeks included
10 hepatic vacuolar degeneration followed by necrosis of hepatocytes (Adeyemo. 2008b).
11 The severity of observed histopathological effects in the liver was proportional to the
12 duration of exposure and concentration of Pb.
13 In environmental assessments of metal-impacted habitats, ALAD is a recognized
14 biomarker of Pb exposure (U.S. EPA. 2006c). For example, lower ALAD activity has
15 been significantly correlated with elevated blood Pb concentrations in wild caught fish
16 from Pb-Zn mining areas although there are differences in species sensitivity (Schmitt et
17 al., 2007b; Schmitt et al., 2005). Suppression of ALAD activity in brown trout
18 transplanted to a metal contaminated stream was linked to Pb accumulation on gills and
19 in liver in a 23-day exposure (Heier et al., 2009). Alves Costa et al. (2007) observed
20 inhibition of ALAD in hepatocytes of the neotropical traira (Hoplias malabaricus)
21 following dietary dosing of 21 mg Pb/kg every 5 days for 70 days. Cytoskeletal and
22 cytoplasmic disorganization were observed in histopathological examination of affected
23 hepatocytes. In fathead minnow exposed to Pb in either control water (33 (ig Pb/L),
24 CaSO4 (37 (ig Pb/L) or (39 (ig Pb/L) humic acid-supplemented water for 30 days and
25 subsequently analyzed by quantitative PCR analysis there were no significant changes in
26 ALAD mRNA gene response leading the authors to speculate that water chemistry alone
27 does not influence this gene response (Mager et al., 2008). In the same study, glucose-6-
28 phosphate dehydrogenase, glutathione-S-transferase and ferritin were upregulated, in
29 microarray analysis, however, no changes in whole body ion concentrations were
30 observed (Mager etal.. 2008).
31 In fish, changes in blood chemistry associated with Pb exposure were noted in the
32 2006 Pb AQCD (U.S. EPA. 2006b), however, only limited recent studies consider effects
33 on blood parameters. In a 70-day feeding study with traira exposed to dietary doses
34 (21 mg Pb/kg as Pb nitrate via prey [Astyanax sp.]) each five days (corresponding to
35 daily nominal doses of approximately 4 mg Pb/kg), there were no significant changes to
36 leukocytes or hemoglobin concentration and volume (Oliveira Ribeiro et al., 2006).
37 Significant differences in area, elongation and roundness of erythrocytes were observed
38 in the Pb-exposed individuals using light microscopy image analysis. Other studies
November 2012 7-133 Draft - Do Not Cite or Quote
-------
1 available since the 2006 Pb AQCD have only shown effects on blood chemistry at high
2 aqueous concentrations of Pb that are not representative of Pb concentrations in U.S.
3 surface waters. For example, in the African catfish packed cell volume decreased with
4 increasing nominal concentration of Pb (25,000 to 200,000 (ig Pb/L as Pb nitrate) and
5 platelet counts increased in a 96-hour exposure (Adevemo. 2007). Red blood cell counts
6 also decreased in some of the treatments when compared to controls, although the
7 response was not dose-dependent and so may not have been caused by Pb exposure.
8 Disruption of ionoregulation is one of the major modes of action of Pb toxicity. The gill
9 has long been recognized as a target of Pb in teleosts. Acute Pb toxicity at the fish gill
10 primarily involves disruption of Ca2+ homeostasis as previously characterized in the
11 2006 Pb AQCD (Rogers and Wood. 2004; Rogers and Wood. 2003). In addition to this
12 mechanism, Pb was found to induce ionoregulatory toxicity at the gill of rainbow trout
13 through a binding of Pb with Na+/K+ATPase and rapid inhibition of carbonic anhydrase
14 activity thus enabling noncompetitive inhibition of Na+ and Cl" influx (Rogers et al..
15 2005). Alves et al. (2006) administered a diet of three concentrations of Pb (7, 77 and
16 520 mg Pb/kg dry weight) to rainbow trout for 21 days, and measured physiological
17 parameters including Na+ and Ca2+ influx rate from water. Dietary Pb had no effect on
18 brachial Na+ and Ca2+ rates except on day 8 where Na+ influx rates were significantly
19 elevated. These studies suggest that Pb is intermediate between purely Ca2+ antagonists
20 such as Zn2+ and Cd2+ and disrupters of Na+ and Cl" balance such as Ag+ and Cu2+. This
21 finding has implications for BLM modeling since it suggests that both Ca2+ and Na+ need
22 to be considered as protective cations for Pb toxicity. Indeed, protection from Pb toxicity
23 by both Na+ and Ca2+ has been documented in freshwater fish (Komjarova and Blust
24 2009b).
25 Additional experiments conducted since the 2006 Pb AQCD provide supporting evidence
26 for underlying mechanisms of Pb toxicity. It was previously established that long-term
27 exposures of Pb can impact gill structure and function. Histopathological observations of
28 gill tissue in the catfish (C. gariepinus) following an 8-week aqueous exposure to
29 nominal concentrations of Pb nitrate revealed focal areas of epithelial hyperplasia and
30 necrosis at the lower exposure concentrations (50 (ig Pb/L and 100 (ig Pb/L) (Adeyemo.
31 2008a). Hyperplasia of mucous cells and epithelial cells were apparent in the tissue from
32 fish exposed the highest concentrations of Pb in the study (500 (ig Pb/L and
33 1,000 (ig Pb/L). In vitro incubation of gill tissue from fathead minnow with Pb
34 concentrations of 2,500, 12,500 and 25,000 (ig Pb/L for 60 minutes decreased the ratio of
35 reduced glutathione to oxidized glutathione, indicating that lipid peroxidation at the gill
36 likely contributes to Pb toxicity at low water hardness (Spokas et al.. 2006). It is difficult
37 to extrapolate these observations to natural environments due to the methods used for
38 exposure and the use of nominal exposure concentrations.
November 2012 7-134 Draft - Do Not Cite or Quote
-------
1 In addition to recent evidence of Pb interruption of Na+ and Cl" at the gill (Rogers et al.,
2 2005). Pb can interfere with the ionoregulation of Na+ and Cl" and tubular reabsorption of
3 Ca2+, Mg2+, glucose, and water in the teleost kidney (Patel et al., 2006). Renal parameters
4 including urine flow rate, glomerular filtration rate, urine pH, and ammonia excretion
5 were monitored in a 96-hour exposure of rainbow trout to analytically verified
6 concentration of 1,200 (ig Pb/L as Pb nitrate. Rates of Na+ and Cl" excretion decreased by
7 30% by 48 hours while Mg excretion increased two-to-three fold by 96 hours. Urine flow
8 rate was not altered by Pb exposure, although urinary Pb excretion rate was significantly
9 increased. After 24 hours of Pb exposure, the urine excretion rate of Ca2+ increased
10 significantly by approximately 43% and remained elevated above the excretion rate in the
11 control group for the duration of the exposure. Glomerular filtration rate significantly
12 decreased only during the last 12 hours of the exposure. Ammonia excretion rate
13 increased significantly at 48 hours as urine pH correspondingly decreased. At the end of
14 the experiment glucose excretion was significantly greater in Pb-exposed fish. Although
15 the exposures in this study approached the 96-hour LC50, nephrotoxic effects of Pb
16 indicate the need to consider additional binding sites for this metal in the development of
17 biotic ligand modeling (Patel et al.. 2006). Additional evidence for Pb effects on ion
18 levels were observed in serum of Nile tilapia; Na+ and Cl" were decreased and K+ levels
19 were elevated following a 21 day nominal exposure to 50 (ig Pb/L as Pb nitrate (Tirat et
20 al..2011).
21 Neurological responses offish to Pb exposure were reported in the 1986 Pb AQCD (U.S.
22 EPA. 1986b). Additional evidence of the neurotoxic effects of Pb on teleosts has become
23 available since the 2006 Pb AQCD. The mitogen-activated protein kinases (MAPK),
24 extracellular signal-regulated kinase (ERK)l/2 and p38MAPK were identified for the first
25 time as possible molecular targets for Pb neurotoxicity in a teleost (Leal et al., 2006). The
26 phosphorylation of ERK1/2 and p38MAPK by Pb was determined in vitro and in vivo in the
27 catfish (Rhamdia queleri). R. quelen exposed to a nominal concentration of 1,000 (ig Pb/L
28 (as Pb acetate) for two days showed a significant increase in phosphorylation of ERK1/2
29 and p38MAPK in the nervous system. Incubation of cerebellar slices for 3 hours in 1,035
30 and 2,070 (ig Pb/L as Pb acetate also showed significant phosphorylation of MAPKs. The
31 observed effects of Pb on the MAPK family of signaling proteins have implications for
32 control of brain development, apoptosis and stress response. In the neotropical fish traira,
33 muscle cholinesterase was significantly inhibited after 14 dietary doses of 21 mg Pb/kg
34 wet weight (Rabitto et al.. 2005). Histopathological observations of brains of African
35 catfish exposed to nominal concentrations of 500 (ig Pb/L or 1,000 (ig Pb/L Pb as
36 Pb nitrate for 4 weeks included perivascular edema, focal areas of malacia, and diffuse
37 areas of neuronal degeneration (Adevemo. 2008b). As in the observed effects of Pb on
38 gill function and ionoregulation, it is difficult to assess the significance of these findings
39 to fish in natural environments due to the methods used for exposure.
November 2012 7-135 Draft - Do Not Cite or Quote
-------
1 Evidence from the 2006 Pb AQCD (U.S. EPA. 2006b) and earlier Pb reviews indicate
2 that Pb can impair both cognitive and motor function in fish. Reduced locomotion and
3 foraging ability were observed in Chinese sturgeon juveniles exhibiting abnormal body
4 curvature following nominal exposure to either 800 or 1,600 (ig Pb/L for 112 days (Hem
5 et al., 2011). Actual concentrations of Pb were quantified at the end of the 112-day
6 exposure period (30 to 50% of test media was renewed daily): 129 (ig Pb/L (200 (ig Pb/L,
7 nominal), 458 (ig Pb/L (800 (ig Pb/L nominal), and 1,276 (ig Pb/L (1,600 (ig Pb/L,
8 nominal). These chondrostean fish gradually recovered from deformities during a
9 depuration period and were able to swim and forage effectively 6 weeks after transfer
10 into clean water
11 Since the 2006 Pb AQCD, several studies integrating behavioral and physiological
12 measures of Pb toxicity have been conducted on fish. Some of these observations are
13 reported to occur at concentrations of Pb reported in freshwater. Zebrafish embryos
14 exposed nominally to low concentrations of Pb as Pb chloride (2.0 and 6.0 (ig Pb/L
15 prepared from serial dilutions of a stock solution) until 24 hours post-fertilization and
16 then subsequently tested as larvae or adult fish exhibited behavioral disruptions in
17 response to mechanosensory and visual stimuli (Rice et al., 2011) Although Pb was not
18 measured in the water, Pb uptake in the embryos was quantified during the first 24 hours
19 post-fertilization (approximately 0.08 nM/100 embryos at 2.0 jig Pb/L and 0.32 nM
20 Pb/100 embryos at 6.0 (ig Pb/L). Startle response time in larvae measured as maximum
21 head turn velocity and escape time decreased in a concentration-dependent pattern
22 following a directional, mechanical stimulus (tapping). The pattern of escape swimming
23 was altered in larvae of Pb-exposed embryos compared to controls. In the adult fish
24 hatched from Pb-exposed embryos (6.0 \ig Pb/L), visual response to a rotating black bar
25 against a white background (ability to detect contrast) was significantly degraded. These
26 findings provide evidence for behavioral effects of Pb at concentrations lower than
27 previously reported in fish (U.S. EPA. 2006c). however, aqueous exposure
28 concentrations were not analytically verified.
29 Sloman et al. (2005) investigated the effect of Pb on hierarchical social interactions and
30 the corresponding monoaminergic profiles in rainbow trout. Trout were allowed to
31 establish dominant-subordinate relationships for 24 hours, and then were exposed to
32 46 (ig Pb/L or 325 (ig Pb/L (Pb nitrate) for 48 hours to assess effects on behavior and
33 brain monoamines. In non-exposed fish, subordinate individuals had higher
34 concentrations of circulating plasma cortisol and telencephalic 5-hydroxyindoleacetic
35 acid/5-hydroxytryptamine (serotonin) (5-HIAA/5-HT) ratios. In the high concentration of
36 Pb, there was significant uptake of Pb into gill, kidney and liver when compared with the
37 control group and dominant fish appeared to have elevated hypothalamic 5-HIAA/5HT
38 ratios. Uptake of Pb into the liver was higher in subordinate fish when compared to the
November 2012 7-136 Draft - Do Not Cite or Quote
-------
1 dominant fish. No significant differences were observed in cortisol levels or behavior
2 after metal exposure.
3 Mager et al. (2010) conducted prey capture assays with 10 day old fathead minnow
4 larvae born from adult fish exposed to 120 (ig Pb/L for 300 days, then subsequently
5 tested in a breeding assay for 21 days. The time interval between 1st and 5th ingestion of
6 10 prey items (Artemia nauplif) was used as a measure of behavior and motor function of
7 offspring of Pb-exposed fish. Larvae were offered 10 Artemia and the number ingested
8 within 5 minutes was scored. The number of larvae ingesting 5 Artemia decreased within
9 the time period in offspring of Pb-exposed fish as compared to the control group, leading
10 the authors to suggest this behavior is indicative of motor/behavioral impairment. In
11 another study with fathead minnows, swimming performance measured as critical aerobic
12 swim speed was significantly impaired in minnows in 24-hour acute (139 (ig Pb/L) and
13 chronic 33 to 57 day (143 (ig Pb/L) exposures, however, no significant difference in
14 swim speed was observed in chronic exposures to 33 (ig Pb/L (Mager and Grosell. 2011).
15 Alterations in reproductive and developmental endpoints at the species level can lead to
16 effects at the population-level of biological organization and higher (Section 7.1.1). For
17 example, reduced fecundity may lead to a decreased population size and developmental
18 effects may decrease the ability of a fish to escape predators or reduce spawning
19 mobility. Reproductive and developmental effects of Pb in fish have been reported for
20 several decades. In the 1977 Pb AQCD, second generation brook trout (Salvelinus
21 fontinalis) exposed to 235 or 474 (ig Pb/L were shown to develop severe spinal
22 deformities (lordoscoliosis) (U.S. EPA. 1977). Pb concentration of 120 (ig Pb/L produced
23 spinal curvature in rainbow trout (Oncorhynchus mykiss) and spinal curvatures were
24 observed in developing eggs of killifish as reviewed in the 1986 Pb AQCD (U.S. EPA.
25 1986b). Recent studies on reproductive effects of Pb in fish from oocyte formation to
26 spawning are summarized in Table 7-5.
27 Reproductive performance of zebrafish as measured by incidence of spawning, numbers
28 of eggs per breeding pair or hatch rate of embryos was unaffected following a 63 day diet
29 of field-collected Pb-contaminated polychaetes that were representative of a daily dose of
30 0.3-0.48 mg Pb/kg-day (dry weight diet/wet weight fish) through food (Boyle et al..
31 2010). Mager et al. (2010) conducted 21 day breeding exposures at the end of chronic
32 300 day toxicity testing with fathead minnow. Non-exposed breeders were switched to
33 water containing Pb and Pb-exposed breeders were moved to control tanks and effects on
34 egg hatchability and embryo Pb accumulation were assessed. Fish in the high Pb
3 5 concentration with HCO3 (113 (ig Pb/L) and DOC (112 (ig Pb/L) and the low Pb
36 concentration with HCO3" (31 (ig Pb/L) reduced total reproductive output, while a
37 significant increase in average egg mass was observed in the high Pb HCO3" and DOC
November 2012 7-137 Draft - Do Not Cite or Quote
-------
1 treatments as compared to egg mass size in controls and in low HCO3" and DOC
2 treatments with Pb. No significant differences were present between treatments in egg
3 hatchability.
4 The effects of metals on embryonic stage offish development in C. carpio and other
5 species were reviewed in Jezierska et al. (2009) and included developmental
6 abnormalities during organogenesis as well as embryonic and larval malformations. The
7 authors concluded that the initial period of embryonic development, just after
8 fertilization, and the period of hatching are the times at which developing embryos are
9 most sensitive to metals. Additional nominal exposure studies provide supporting
10 evidence for embryo malformations associated with Pb-exposure. A significant
11 concentration-dependent increase in morphological malformations was observed in
12 African catfish embryos exposed to nominal concentrations of 100 (ig Pb/L, 300 (ig Pb/L
13 or 500 (ig Pb/L Pb nitrate from 6 hours post-fertilization to 168 hours post-fertilization
14 (Osman et al.. 2007b). Hatching was delayed with increasing Pb concentration and hatch
15 success of the embryos decreased from 75% in the controls to 40% in the group exposed
16 to 500 (ig Pb/L. Chinese sturgeon exposed to nominal concentrations of 200 (ig Pb/L,
17 800 (ig Pb/L or 1,600 (ig Pb/L for 112 days (96 hour post-fertilized eggs through juvenile
18 stages) exhibited body curvatures in the two highest concentrations (Hou et al.. 2011).
19 During a 42 day depuration period in clean water following exposure, the degree of
20 curvature in affected individuals decreased with decreasing tissue concentrations of Pb.
21 Reproductive and endocrine effects of Pb have also been reported at the cellular level in
22 fish, including alterations in gonadal tissue and hormone secretions that are associated
23 with Pb-exposure, however, recent studies that report these effects are limited to
24 experiments where only nominal concentrations of Pb were tested. Histopathological
25 observations of ovarian tissue in the African catfish following an 8-week aqueous
26 exposure to Pb nitrate indicated necrosis of ovarian follicles at the lowest concentration
27 tested (50 (ig Pb/L) (Adevemo. 2008a). Severe degeneration of ovarian follicles was
28 observed in the highest concentrations of 500 (ig Pb/L and 1,000 (ig Pb/L. Chaube et al.
29 (2010) considered the effects of Pb on steroid levels through 12 and 24 hour in vitro
30 exposures of post-vitellogenic ovaries from the catfish (Heteropneustes fossilis) to
31 nominal concentrations of Pb as Pb nitrate (0, 10, 100, 1,000, 3,000 and 10,000 (ig Pb/L).
32 Progesterone, 17-hydroxyprogesterone, 17, 20 beta-dihydroxyprogesterone,
33 corticosterone, 21-deoxycortisol and deoxycorticosterone were inhibited in a dose-
34 dependent manner. Pb was stimulatory on the steroids estradiol-17-(3, testosterone and
35 cortisol at low concentrations, and inhibitory at higher concentrations. The authors
36 propose that the disruption of steroid production and altered hormone secretion patterns
37 observed at the lower concentrations of Pb in this study are suggestive of the potential for
38 impacts to fish reproduction (Chaube et al.. 2010).
November 2012 7-138 Draft - Do Not Cite or Quote
-------
1 There is also evidence for alterations in steroid levels associated with Pb exposure in
2 other species offish although these studies were all conducted with nominal
3 concentrations of Pb and the actual exposure concentrations were not verified. Carp
4 (Cyprinus carpio) exposed for 35 days to nominal concentration of 410 (ig Pb/L
5 experienced altered plasma cortisol and prolactin levels. Plasma cortisol levels
6 significantly increased throughout the study period while plasma prolactin increased up
7 to day 14 and then declined and was not significantly different from controls by the end
8 of the experiment (Ramesh et al.. 2009). Cortisol levels were significantly decreased in
9 Nile tilapia exposed to 50 (ig Pb/L (nominal) for 4 days but were followed by a return to
10 control levels at 21 days of exposure (Firat et al., 2011). In a comparative study between
11 in vitro and in vivo estrogenic activity of Pb, vitellogenin was reported to be significantly
12 induced in juvenile goldfish (Carassius auratus) following 96-hour exposure to nominal
13 concentration of 0.2 and 0.02 (ig Pb/L when compared to control fish (Isidori et al..
14 2010). In the same study, estrogenicity of Pb was detected in vitro using a proliferation
15 assay with estrogen receptor-positive human MCF-7 cells. The estrogenic effects of Pb
16 reported by the authors were observed at concentrations at or below that of Pb typically
17 encountered in freshwaters, however, actual concentrations of Pb were not measured and
18 the reported concentrations were at or below analytical detection limits for Pb. The
19 observations of effects of Pb on vitellogenin are interesting; however, additional studies
20 are warranted considering the difficulty in maintaining these low concentrations of Pb.
21 The relevance of the observed in vitro activity to air related exposure to Pb in natural
22 environments is unknown.
23 Reduction of growth in fish was noted as an effect of Pb exposure in the 2006 Pb AQCD.
24 Recent studies available since the 2006 Pb AQCD do not present consistent evidence of
25 growth reduction in fish associated with Pb (Table 7-5). In a series of exposures in which
26 Ca2+, DOC and pH were varied to assess effects on Pb toxicity to fathead minnows,
27 Grosell et al. (2006a) observed a significant increase in growth in some groups exposed
28 to higher concentrations, however, the increase in body mass was noted to have occurred
29 in tanks with high mortality earlier in the exposure (Grosell et al.. 2006a). Fathead
30 minnows exposed to 33 (ig Pb/L to test swimming performance had significantly greater
31 body length and body mass compared to control fish following a mean Pb exposure
32 duration of 41 days (range 33 to 57 days) (Mager and Grosell 2011). In 30 day chronic
33 tests in which a range of pH values (6.4, 7.5 and 8.3) were tested with low
34 (25-32 (ig Pb/L), intermediate (82-156 (ig Pb/L) and high (297-453 (ig Pb/L)
35 concentrations of Pb, Mager et al. (20 lib) did not observe growth impairment in fathead
36 minnows at environmentally relevant concentrations of Pb. However, two 60-day early
37 lifestage tests with rainbow trout showed differences in LOEC for reduced growth
38 (Mebane et al.. 2008). In the first test, a 69-day exposure, the LOECs for mortality and
November 2012 7-139 Draft - Do Not Cite or Quote
-------
1 reduced growth were the same (54 (ig Pb/L). In the second test, a 62-day exposure of Pb
2 to rainbow trout, the LOEC for fish length was 18 (ig Pb/L with an EC2o of >87 (ig Pb/L.
3 No effects on growth were observed in recently conducted feeding studies with fish.
4 Growth and survival were not significantly affected in juvenile rainbow trout, fathead
5 minnow and channel catfish (Ictalurus punctatus) fed a live diet of L. variegatus
6 contaminated with Pb (846-1,000 (ig Pb/L-g dry mass for 30 days). (Erickson et al..
7 2010). No effects on growth rates were observed in rainbow trout administered a diet
8 containing three concentrations of Pb (7, 77 and 520 mg Pb/kg dry weight) for 21 days
9 (Alves et al.. 2006) or in Nile tilapia fed diets with nominal concentration of 100, 400, or
10 800 mg/kg Pb dry weight for 60 days (Dai et al.. 2009b).
11 In one recent field study, faster growth rates were associated with lower whole-body trace
12 element concentrations in salmon (Salmo salar) across several streams in New
13 Hampshire and Massachusetts, U.S., regardless of whether accumulation was from prey
14 items or from water (Ward etal. 2010). In sites where conditions in the streams were
15 conducive to rapid salmon growth, Pb concentrations were 86% lower than in streams
16 where salmon were smaller.
Amphibians
17 Amphibians move between terrestrial and aquatic habitats and can therefore be exposed
18 to Pb both on land and in water. The studies reviewed here are all aquatic or sediment
19 exposures. Biological effects of Pb on amphibians in terrestrial exposure scenarios are
20 reviewed in Sections 7.3.3.3 and 7.3.4.3. Amphibians lay their eggs in or around water
21 making them susceptible to water-borne Pb during swimming, breeding and
22 development. In the 2006 Pb AQCD amphibians were considered to be relatively tolerant
23 to Pb. Observed responses to Pb exposure included decreased enzyme activity
24 (e.g., ALAD reduction) and changes in behavior summarized in Table AX7-2.4.3 (U.S.
25 EPA. 2006c). Since the 2006 Pb AQCD, studies conducted within two orders of
26 magnitude of the range of published Pb concentrations for surface waters and sediments
27 of the U.S. (Section 7.2.3) have indicated sublethal effects on tadpole endpoints including
28 growth, deformity, and swimming ability. Genotoxic and enzymatic effects of Pb
29 following chronic exposures have been assessed in laboratory bioassays, however, these
30 studies were limited to nominal exposures.
31 The genotoxic potential of Pb to larvae of the frog (X. laevis) was assessed by
32 determining the number of micronucleated erythrocytes per thousand (MNE) following a
33 12-day exposure to nominal concentrations of Pb as Pb nitrate (Mouchet et al.. 2007).
34 The lowest Pb concentrations withX laevis (10 and 100 (ig Pb/L) did not exhibit
November 2012 7-140 Draft - Do Not Cite or Quote
-------
1 genotoxic effects while both 1,000 and 10,000 (ig Pb/L significantly increased MNE to
2 14 and 202, respectively compared to the control (6 MNE). In another chronic genotoxic
3 study, erythrocytic micronuclei and erythrocytic nuclear abnormalities were significantly
4 increased with increasing Pb concentrations (700 (ig Pb/L, 1,400 (ig Pb/L,
5 14,000 ng Pb/L, 70,000 ng Pb/L) during 45, 60, and 75-day exposures of tadpoles Bufo
6 raddei fZhang et al.. 2007bj. The authors noted that the erythrocytic micronuclei and
7 erythrocytic nuclear abnormalities frequencies generally decreased with increasing
8 exposure time and that this may be indicative of regulation of genotoxic factors by
9 tadpoles.
10 Endpoints of oxidative damage were measured in testes of the black-spotted frog (Rana
11 nigromaculata) treated with nominal concentrations of 100 (ig Pb/L, 200 (ig Pb/L,
12 400 (ig Pb/L, 800 (ig Pb/L or 1,600 ng Pb/L Pb nitrate by epidermal absorption for 30
13 days (Wang and Jia. 2009). All doses significantly increased MDA, a product of
14 oxidative stress, and glutathione levels were elevated in all but the lowest treatment
15 group. In the same study, damage to DNA assessed by DNA tail length showed effects at
16 >200 (ig Pb/L and DNA tail movement showed effects at >400 (ig Pb/L. The authors
17 concluded that the effects on endpoints of oxidative stress and DNA damage detected in
18 testes indicated a possible reproductive effect of Pb to black-spotted frogs. The exposure
19 method and use of nominal concentration in this study make it difficult to determine the
20 relevance of this study to exposure scenarios under natural environmental conditions.
21 Various sublethal endpoints (growth, deformity, swimming ability, metamorphosis) were
22 evaluated in northern leopard frog (R. pipiens) tadpoles exposed to nominal
23 concentrations of 3, 10, and 100 (ig Pb/L as Pb nitrate from embryonic stage to
24 metamorphosis (Chen et al., 2006b). In this chronic study, the concentrations represent
25 the range of Pb found in surface freshwaters across the U.S. The lowest concentration of
26 3 (ig Pb/L approaches the EPA chronic criterion for Pb of 2.5 (ig Pb/L at a hardness of
27 100 mg/L or 4.5 (ig Pb/L at a hardness of 170 mg/L (U.S. EPA. 2002b). No effects were
28 observed in the lowest concentration. In the 100 (ig Pb/L treatment, tadpole growth rate
29 was slower (Gosner stages 25-30), 92% of tadpoles had lateral spinal curvature
30 (compared with 6% in the control) and maximum swimming speed was significantly
31 slower than the other treatment groups. In this study, Pb concentrations in the tissues of
32 tadpoles were quantified and the authors reported that they were within the range of
33 reported tissue concentrations from wild-caught populations.
34 The effects of Pb-contaminated sediment on early growth and development were assessed
35 in the southern leopard frog (Sparling et al., 2006). Tadpoles exposed to Pb in sediment
36 (45, 75, 180, 540, 2,360, 3,940, 5,520, and 7,580 mg Pb/kg dry weight) with
37 corresponding sediment pore water concentrations of 123, 227, 589, 1,833, 8,121, 13,579,
November 2012 7-141 Draft - Do Not Cite or Quote
-------
1 19,038 and 24,427 (ig Pb/L from embryonic stage to metamorphosis exhibited sublethal
2 responses to Pb in sediment at levels below 3,940 mg Pb/kg. There was 100% mortality
3 in the 3,940, 5,520 and 7,580 mg Pb/kg exposures by day 5. The authors noted that the
4 most profound effects of Pb on the tadpoles were on skeletal development. At 75 mg
5 Pb/kg, subtle effects on skeletal formation such as clinomely and brachydactyly were
6 observed. Skeletal malformations increased in severity at 540 mg Pb/kg and included
7 clinodactyly, brachymely and spinal curvature and these effects persisted after
8 metamorphosis. At the highest concentration with surviving tadpoles (2,360 mg Pb/kg)
9 all individuals displayed severe skeletal malformations that impacted mobility. Other
10 sublethal effects of Pb observed in this study were reduced rates of early growth of
11 tadpoles at concentrations < 540 mg Pb/kg and increased time to metamorphosis in the
12 2,360mg Pb/kg (8,121(ig Pb/L sediment pore water) treatment.
Birds
13 As reviewed in Koivula and Eeva (2010) measurement of enzymes associated with
14 oxidative stress in birds is a well-established biomarker of exposure to metals, however,
15 little is known about the effects of this stress response in wild populations or at higher
16 levels of ecological organization. Changes in ALAD activity and other oxidative stress
17 biomarkers at low levels of Pb exposure were recently documented in mallards and coots
18 (Fulica atra) from a lagoon in Spain impacted by Pb shot (Martinez-Haro et al., 2011).
19 ALAD ratio in mallards decreased linearly with blood Pb levels between 6 and
20 40 (ig Pb/dL, and at Pb levels of <20 (ig Pb/dL effects on several antioxidant enzymes
21 were observed in coots. Although the primary route of exposure to the birds was via
22 ingestion of Pb shot, effects were observed lower than 20 (ig Pb/dL, the background level
23 frequently applied to Pb exposures in birds (Martinez-Haro et al., 2011; Brown et al.,
24 2006).
25 Consideration of toxicity of Pb to vertebrate embryos that develop surrounded by a
26 protective egg shell has been expanded since the 2006 Pb AQCD. Pb treatment of
27 mallard duck (Anas platyrhynchos}, eggs by immersion in an analytically verified
28 concentration of 100 (ig Pb/L for 30 minutes on day 0 of development did not increase
29 malformations or mortality of embryos (Kertesz and Fancsi. 2003). However, immersion
30 of eggs in 2,900 (ig Pb/L under the same experimental conditions resulted in increased
31 rate of mortality and significant malformations including hemorrhages of the body,
32 stunted growth, and absence of yolk sac circulatory system (Kertesz et al. 2006). The
33 second study was conducted to emulate environmental levels of Pb following a dam
34 failure in Hungary.
November 2012 7-142 Draft - Do Not Cite or Quote
-------
7.4.6 Exposure and Response of Freshwater Species
1 Evidence regarding exposure-response relationships and potential thresholds for Pb
2 effects on aquatic populations can inform determination of standard levels that are
3 protective of aquatic ecosystems. The Annex of the 2006 Pb AQCD (U.S. EPA. 2006c)
4 summarized data on exposure-response functions for invertebrates (Table AX7-2.4.1) and
5 fish (Table AX7-2.4.2). The recent exposure-response studies in this section expand on
6 the findings from the 2006 Pb AQCD with information on newly-tested organisms
7 (including microalgae, invertebrate, amphibian and fish species). Overall, new data for
8 freshwater invertebrates generally support the previous finding of sensitivity of juvenile
9 lifestages and indicates some effects of Pb observed in some species at concentrations of
10 Pb reported in freshwater environments. (Table 7-2). All reported values are from
11 exposures in which concentrations of Pb were analytically verified unless nominal
12 concentrations are stated.
13 The aquatic macrophyte Lemna minor (duckweed) exhibited a EC50 for growth inhibition
14 of 6,800 (ig Pb/L in a 4-day exposure and 5,500 (ig Pb/L for a 7-day exposure to a range
15 of Pb concentrations from 100 to 9,970 (ig Pb/L (Dirilgen. 2011). Growth (measured as
16 biomass) was slightly increased at 100 and 200 (ig Pb/L and then decreased in subsequent
17 concentrations. In an assay using nominal concentrations of Pb the aquatic freshwater
18 microalgae Scenedesmus obliquus was significantly more sensitive to Pb exposure than
19 Chlorella vulgaris algae, although these authors stated that both appeared to be very
20 tolerant of the heavy metal. Laboratory 48-hour standard toxicity tests were performed
21 with both of these species and respective EC50 values of 4,040 and 24,500 (ig Pb/L for
22 growth as measured by cell division rate were derived (Atici et al., 2008).
23 Exposure-response data for freshwater invertebrates provide evidence for effects of Pb at
24 concentrations of Pb encountered in U.S. surface waters. In the 2006 Pb AQCD, effects
25 of Pb-exposure in amphipods (H. aztecd) and water fleas (D. magnd) were reported at
26 concentrations as low as 0.45 (ig Pb/L. Effective concentrations for aquatic invertebrates
27 were found to range from 0.45 to 8,000 (ig Pb/L. Since the 2006 Pb AQCD, recent
28 studies have identified the freshwater snail L. stagnalis as a species that is extremely
29 sensitive to Pb exposure (Grosell and Brix. 2009; Grosell et al., 2006b). Growth of
30 juvenile L. stagnalis was inhibited below the lowest concentration tested resulting in an
31 EC20 of <4 (ig Pb/L. In the same study, the NOEC was 12 jig Pb/L and the LOEC was
32 16 (ig Pb/L. In contrast, freshwater juvenile ramshorn snails M. cornuarietis were less
33 sensitive to Pb with the same LOEC for hatching rate and LC50, calculated to be about
34 10,000 (ig Pb/L based on nominal exposure data (Sawasdee and Kohler. 2010).
35 Additional studies on Pb effects in aquatic invertebrates published since the
36 2006 Pb AQCD provide further evidence for differences in sensitivity of different
November 2012 7-143 Draft - Do Not Cite or Quote
-------
1 lifestages of aquatic organisms to Pb. In the freshwater mussel, L. siliquoidea (fatmucket)
2 a Pb concentration response was observed in which newly transformed (5-day-old)
3 juveniles were the most sensitive lifestage in a 96-hour toxicity test when compared to
4 acute and chronic results with other lifestages (Wang et al.. 2010e). The 96-hour EC50
5 values for the 5-day-old L. siliquoidea in two separate toxicity tests were 142 and
6 298 (ig Pb/L (mean EC50 220 (ig Pb/L) in contrast to older juveniles (2 months old) with
7 an EC50 >426 (ig/L. The 24-hour median effect concentration for glochidia (larvae) of
8 L. siliquoidea in 48-hour acute toxicity tests was >299 (ig/L. A 28-day exposure chronic
9 value of 10 (ig Pb/L was obtained from 2-month-old L. siliquoidea juveniles, and was the
10 lowest genus mean chronic value ever reported for Pb (Wang et al., 2010e). A 96-hour
11 test on newly transformed juveniles was also conducted on Lampsilis rafmesqueana
12 (Neosho mucket), a mussel that is a candidate for the endangered species list. The EC50
13 for this species was 188 (ig Pb/L.
14 Different lifestages of chironomids have been shown to have varying sensitivity to Pb
15 exposure in several studies available since the 2006 Pb AQCD. The acute toxicity of Pb
16 to first-instar C. riparius larvae was tested in soft water, with hardness of 8 mg/L as
17 CaCO3 (Bechard et al.. 2008). The 24-hour LC50 of 610 ng Pb/L for first instar
18 C. riparius larvae was much lower than previous values reported for later instars in
19 harder water. In a chronic test with Chironomus tentans, (8 day-old larvae exposed to Pb
20 until emergence [approximately 27 days]), the NOEC was 109, and the LOEC was
21 497 (ig Pb/L (Grosell et al., 2006b). The EC2o for reduced growth and emergence of the
22 midge Chironomus dilutus was 28 (ig Pb/L, observed in a 55-day exposure, while the
23 same species had a 96-hour LC50 of 3,323 (ig Pb/L (Mebane et al., 2008). In fourth instars
24 of the freshwater midge larvae Chironomus javanus the 24, 48, 72 and 96 hour LC50's
25 were 20,490, 6,530, 1,690 and 720 jig Pb/L, respectively (Shuhaimi-Othman et al.,
26 201 Ic). This was comparable to the 96-hour LC50 (400 (ig Pb/L) in the midge larvae
27 Culicoides furens (Vedamanikam and Shazilli. 2008a). In the same study, the 96-hour
28 LC50 for Chironomus plumosus ranged from 8,300 (ig Pb/L to 16,210 (ig Pb/L under
29 different temperatures indicating the role of environmental factors in modulation of
30 toxicity and differences in sensitivity to Pb even among related species.
31 Cladocerans are commonly tested aquatic organisms, with data from three species:
32 D. magna, D. pulex and Ceriodaphnia dubia, representing approximately 70% of
33 available metal toxicological literature on this group (Wong et al.. 2009). Recent studies
34 have been conducted with C. dubia and acute toxicity values for other cladocerans as
35 well as sublethal endpoints for D. magna are available. In a series of 48 hour acute
36 toxicity tests with C. dubia conducted in a variety of natural waters across North
37 America, LC50 values ranged from 29 to 1,180 (ig Pb/L and were correlated with DOC
38 (Esbaugh et al., 2011). Median lethal concentrations forMoina micrura (LC50
November 2012 7-144 Draft - Do Not Cite or Quote
-------
1 690 (ig Pb/L), Diaphanosoma birgei (LC50 3,160 (ig Pb/L), andAlona rectangular (LC50
2 7,060 (ig Pb/L) indicate differences in sensitivity to Pb in these freshwater cladocerans
3 from Mexico (Garcia-Garcia et al.. 2006). Several additional studies available since the
4 2006 Pb AQCD report exposure response values for Daphnia that are based an nominal
5 data: an acute study of Pb with D. pulex identified a 48-hour LC50 of 4,000 (ig/L for this
6 species (Theegala et al.. 2007) and the EC50 for swimming inhibition in neonate
7 D. magna exposed to Pb nitrate for 24 hours was 18,153 (ig Pb/L (Ha and Choi. 2009).
8 Rotifers are among the most sensitive aquatic genera to Pb with wide variation in LC50
9 values reported between species (Perez-Legaspi and Rico-Martinez. 2001). For example,
10 in the rotifer genus Lecane, a 22-fold difference in LC50 values was observed in 48-hour
11 exposure to Pb between L. hamata, L. luna andZ. quadridentata. (Perez-Legaspi and
12 Rico-Martinez. 2001). L. luna was most sensitive to Pb toxicity with a 48-hour LC50 of
13 140 (ig Pb/L. In a 48-hour toxicity test with the rotifer Brachionus calyciflorus, an NOEC
14 (194 (ig Pb/L), an LOEC (284 (ig Pb/L), and an EC20 of 125 (ig Pb/L was established for
15 this species (Grosell et al., 2006b). The freshwater rotifer Euchlanis dilatata 48 hour
16 LC50 was 35 (ig Pb/L using neonates hatched from asexual eggs (Arias-Almeida and
17 Rico-Martinez. 2011). In this study the authors estimated the NOEC to be 0.1 jig Pb/L
18 and the LOEC to be 0.5 (ig Pb/L. In contrast, for rotifer Brachionus patulus neonates, the
19 24-hour LC50 was 6,150 (ig Pb/L, however, this value was based on nominal exposures
20 (Garcia-Garcia et al.. 2007).
21 Exposure-response assays on other freshwater species have been conducted since the
22 2006 Pb AQCD. The 24-hour LC50 for larvae of C. quinquefasciatus mosquitoes was
23 180 (ig Pb/L (Kitvatanachai et al.. 2005). A 48-hour LC50 of 5,200 (ig Pb/L was observed
24 in water-only exposures of the blackworm Lumbriculus variegatus fPenttinen et al.,
25 2008J. In the mayfly Baetis tricaudatus, the 96-hour LC50 was 664 (ig Pb/L (Mebane et
26 al.. 2008). An EC2o value of 66 (ig Pb/L was derived for B. tricaudatus by quantifying the
27 reduction in the number of molts over a 10-day exposure to Pb (Mebane et al.. 2008). The
28 number of molts was significantly less than the control (average of 14 molts over 10
29 days) at concentrations of 160 (ig Pb/L and higher with the lowest number of molts
30 (average of 5.3 molts over 10 days) observed in the highest concentration (546 \ig Pb/L).
31 In the freshwater ostracod Stenocypris major, the 96-hour LC50 was 526 (ig Pb/L
32 (Shuhaimi-Othman et al.. 201 Ib). In another freshwater crustacean, the prawn
33 Macrobrachium lancesteri, the 96-hour LC50 was 35 (ig Pb/L in soft water (<75 mg/L as
34 CaCO3) (Shuhaimi-Othman et al.. 201 la).
35 In the studies reviewed for the 2006 Pb AQCD, freshwater fish demonstrated adverse
36 effects at concentrations ranging from 10 to >5,400 (ig Pb/L, generally depending on
37 water quality parameters (e.g., pH, hardness, salinity) (U.S. EPA. 2006c). Pb tended to be
November 2012 7-145 Draft - Do Not Cite or Quote
-------
1 more toxic in longer-term exposures and correlated to Pb-uptake in tissues. Table
2 AX7-2.4.2 of the 2006 Pb AQCD summarizes effects of Pb to fish. A series of studies
3 published since the 2006 Pb AQCD have been conducted and have further elucidated the
4 influence of water chemistry parameters on Pb uptake and toxicity in fathead minnow
5 resulting in additional dose-response data for this species. Grosell et al. (2006b)
6 conducted a series of 30-day exposures with larval fathead minnow in which varying
7 concentrations of Ca2+ (as CaSO4) and DOC were tested. The effects of reduced pH (6.7)
8 and increased pH (8.1) compared to a control pH of 7.4 on Pb toxicity were also assessed
9 in this study. DOC, CaSO4 and pH influenced Pb toxicity considerably over the range of
10 water parameters tested. The 30-day LC50 for low hardness (19 mg CaSO4/L) in basic test
11 water was 39 (ig dissolved Pb/L and the highest LC50 value (obtained from the protection
12 from increased concentrations of DOC and CaSO4) was 1,903 (ig dissolved Pb/L (Grosell
13 et al.. 2006a). This range in LC50 values for larval fathead minnows for differing pH and
14 concentrations of DOC and hardness clearly demonstrates the importance of the
15 chemistry of the exposure medium to Pb toxicity.
16 Mager et al. (2010) conducted 300-day chronic toxicity tests at 35 and 120 (ig Pb/L with
17 fathead minnow under conditions of varied DOC and alkalinity to assess the effects of
18 these water quality parameters on fish growth and Pb-uptake. In additional tests with
19 fathead minnow, Mager et al. (20lib) conducted both 96-hour acute and 30-day chronic
20 tests to further characterize Ca2+, DOC, pH, and alkalinity values on Pb toxicity.
21 Increased Ca2+, DOC and NaHCO3 concentration afforded protection to minnows in acute
22 studies. The role of pH in Pb toxicity is complex and likely involves Pb speciation and
23 competitive interaction of fT with Pb2+ (Mager et al.. 201 Ib). In a series of 96-hour acute
24 toxicity tests with fathead minnow conducted in a variety of natural waters across North
25 America, LC50 values ranged from 41 to 3,598 (ig Pb/L and no Pb toxicity occurred in
26 three highly alkaline waters (Esbaugh et al.. 2011).
27 In the 2006 Pb AQCD, fish lifestage was recognized as an important variable in
28 determining the sensitivity of these organisms to Pb. Recent data available since the
29 2006 Pb AQCD (U.S. EPA. 2006c) support the findings of increased sensitivity of
30 juvenile fish to Pb when compared to adults. Acute (96-hour) and chronic (60-day) early-
31 lifestage test exposures were conducted with rainbow trout to develop acute-chronic
32 ratios (ACR's) for this species (Mebane et al., 2008). Two early-lifestage chronic tests
33 were conducted, the first with an exposure range of 12-384 (ig Pb/L (69 days) at 20 mg
34 CaCO3/L water hardness and the second with an exposure range of 8 to 124 (ig Pb/L
35 (62 days) and a water hardness of 29 mg CaCO3/L. In the 69-day test, the following
36 chronic values were observed for survival: NOEC=24 (ig Pb/L, maximum acceptable
37 toxicant concentration (MATC)=36 (ig Pb/L, EC10=26 (ig Pb/L, EC20=34 (ig Pb/L, and
38 LC50=55 (ig Pb/L. Results from the 62-day test, with fish length as the endpoint, were
November 2012 7-146 Draft - Do Not Cite or Quote
-------
1 NOEC=8 ng Pb/L, MATC=12 ng Pb/L, EC10=7(ig Pb/L, EC20=102 ng Pb/L and
2 LC50=120 (ig Pb/L. In acute tests with rainbow trout run concurrently with the chronic
3 tests, 96-hour LC50 values were 120 and 150 (ig Pb/L, respectively. Data from this study
4 resulted in ACR's for trout lower than previously reported. The low ACR values were
5 due to the acute tests which produced LC50 values that were 10 to 25 times lower than
6 earlier studies with trout (Mebane et al.. 2008). The authors speculated that the lower
7 LC50 values were due to the age of the fish used in the study (two to four week old fry)
8 and that testing with larger and older fish may not be protective of more sensitive
9 lifestages.
10 There have been only a few recent exposure-response studies in amphibians since the
11 2006 Pb AQCD. Southern leopard frog tadpoles exposed to Pb in sediment (45 to
12 7,580 mg Pb/kg dry weight) with corresponding sediment pore water concentrations from
13 123 to 24,427 (ig Pb/L from embryonic stage to metamorphosis exhibited concentration-
14 dependent effects on survival (Sparling et al.. 2006). The LC50 value for Pb in sediment
15 was 3,738 mg Pb/kg, which corresponds to 12,539 (ig Pb/L in sediment pore water. In the
16 same study, concentration-dependent effects on skeletal development were observed. The
17 40 day-ECso for deformed spinal columns in the tadpoles was 1,958 mg Pb/kg
18 (corresponding to 6,734 (ig Pb/L sediment pore water) and the 60 day-EC50 was 579 mg
19 Pb/kg (corresponding to 1,968 (ig Pb/L sediment pore water) (Sparling et al., 2006).
7.4.7 Freshwater Community and Ecosystem Effects
20 As discussed in the 1986 Pb AQCD (U.S. EPA. 1986b) and the 2006 Pb AQCD (U.S.
21 EPA. 2006b). exposure to Pb is likely to have impacts in aquatic environments via effects
22 at several levels of ecological organization (organisms, populations, communities, or
23 ecosystems). These effects resulting from toxicity of Pb would be evidenced by changes
24 in species composition and richness, in ecosystem function, and in energy flow. The
25 2006 Pb AQCD concluded that, in general, there was insufficient information available
26 for single materials in controlled studies to permit evaluation of specific impacts on
27 higher levels of organization (beyond the organism). Furthermore, Pb rarely occurs as a
28 sole contaminant in natural systems making the effects of Pb difficult to ascertain. New
29 information on effects of Pb at the population, community, and ecosystem level is
30 reviewed below.
31 In laboratory studies reviewed in the 2006 Pb AQCD and in more recent studies, Pb
32 exposure has been demonstrated to alter predator-prey interactions, as well as feeding and
33 avoidance behaviors. In aquatic ecosystems there are field studies reviewed in the 1977
34 Pb AQCD (U.S. EPA. 1977). the 1986 Pb AQCD (U.S. EPA. 1986b), the
November 2012 7-147 Draft - Do Not Cite or Quote
-------
1 2006 Pb AQCD (U.S. EPA. 2006b) and more recent studies that report reductions of
2 species abundance, richness or diversity particularly in benthic macroinvertebrate
3 communities coexisting with other metals where the sources of Pb were from mining or
4 urban effluents. Additionally, field studies have linked Pb contamination to reduced
5 primary productivity and respiration, and to altered energy flow and nutrient cycling.
6 However, because of the complexity inherent in defining such effects, there are relatively
7 few available population, community, or ecosystem level studies that conclusively relate
8 Pb exposure to aquatic ecosystem effects. In addition, most of the available work is
9 related to point-source Pb contamination, with very few studies considering the effects of
10 diffuse Pb pollution. Both plant species and habitat type were determined to be factors
11 affecting the rate of Pb accumulation from contaminated sediments. While the rooted
12 aquatic plant E. canadensis was observed to accumulate the highest concentrations of Pb,
13 the authors concluded that submerged macrophytes (versus emergent plants) as a group
14 were the most likely to accumulate Pb and other heavy metals (Kurilenko and
15 Osmolovskaya. 2006). This would suggest that certain types of aquatic plants, such as
16 rooted and submerged species, may be more susceptible to aerially-deposited Pb
17 contamination, resulting in shifts in plant community composition as a result of Pb
18 pollution.
19 Alteration of macrophyte community composition was demonstrated in the presence of
20 elevated surface water Pb concentrations at three lake sites impacted by mining effluents
21 (Mishra et al., 2008). A total of 11 species of macrophytes were collected. Two sites
22 located 500 meters and 1,500 meters downstream from the mine discharge point (study
23 sites 2 and 3) exhibited similar dissolved Pb concentrations (78 to 92 (ig Pb/L, depending
24 on season) and contained six and eight unique macrophyte species, respectively. The site
25 nearest the discharge point of the mine effluent (study site 1) had the highest Pb
26 concentrations (103 to 118 (ig Pb/L) and the lowest number of resident macrophyte
27 species; these included E. crassipes, L. minor, Azolla pinnata and S. polyrrhiza. Based on
28 analysis of plant tissue Pb concentrations, the authors theorized that certain species may
29 be more able to develop Pb tolerant eco-types that can survive at higher Pb
30 concentrations (Mishra et al.. 2008). In field studies available for certain freshwater
31 habitats, exposure to Pb has been shown to result in significant alterations of invertebrate
32 communities. Macroinvertebrate community structure in mine-influenced streams was
33 determined to be significantly correlated to Pb sediment pore water concentrations.
34 Multiple invertebrate community indices, including Ephemeroptera, Plecoptera,
35 Trichoptera (EPT) taxa richness, Missouri biotic index, and Shannon-Wiener diversity
36 index, were integrated into a macroinvertebrate biotic condition score (Poulton et al..
37 2010). These scores were determined to be significantly lower at sample sites
38 downstream from mining sites where Pb pore water and bulk sediment concentrations
39 were elevated. Sediment Pb, Cd, and Zn levels were inversely correlated to mussel taxa
November 2012 7-148 Draft - Do Not Cite or Quote
-------
1 richness in the Spring River basin encompassing sites in Kansas, Missouri and Oklahoma
2 overlapping a former Pb and Zn mining and processing area (Angelo et al.. 2007). In sites
3 upstream of the mining area, 21 to 25 species of mussels were present whereas in sites
4 downstream, only 6 to 8 species were observed.
5 Rhea et al. (2006) examined the effects of multiple heavy metals in the Boulder River,
6 Montana, watershed biofilm on resident macroinvertebrate assemblages and community
7 structure, and determined that, among all the metals, biofilm Pb concentrations exerted
8 the greatest influence on the macroinvertebrate community indices. Pb biofilm
9 concentrations were significantly correlated with reduced EPT taxa richness, reduced
10 EPT abundance, and an increase in Diptera species abundance. Interestingly, Pb
11 concentrations in invertebrate tissues were correlated to an increase in Hydropsychidae
12 caddisfly abundance, but this may have resulted from the intrinsically high variability in
13 tissue Pb concentrations. The authors concluded that Pb-containing biofilm represented a
14 significant dietary exposure for impacted macroinvertebrate species, thus altering
15 invertebrate community metrics (Rhea et al.. 2006).
16 Kominkova and Nabelkova (2005) examined ecological risks associated with metal
17 contamination (including Pb) in small urban streams. Although surface water Pb
18 concentrations in monitored streams were determined to be very low, concentrations of
19 the metal in sediment were high enough to pose a risk to the benthic community (e.g., 34
20 to 101 mg Pb/kg). These risks were observed to be linked to benthic invertebrate
21 functional feeding group, with collector-gatherer species exhibiting larger body burdens
22 of heavy metals than other groups (Kominkova and Nabelkova. 2005). In contrast,
23 benthic predators and collector-filterers accumulated significantly lower metals
24 concentrations. Consequently, it is likely that sediment-bound Pb contamination would
25 differentially affect members of the benthic invertebrate community, potentially altering
26 ecosystems dynamics.
27 Invertebrate functional feeding group may also affect invertebrate Pb body burdens in
28 those systems where Pb bioconcentration occurs. The predaceous zooplanktonic rotifer,
29 A. brightwellii collected from a Pb-impacted freshwater reservoir in Mexico, contained
30 384 ng Pb/mg and exhibited a water-to-tissue BCF of 49,344. The authors theorized that
31 Pb biomagnification may have been observed in this case because the cladoceran
32 M. micrura is both a known Pb accumulator and a favorite prey item of the rotifer
33 (Rubio-Franchini et al.. 2008). They showed thatM micrura had twice the Pb body
34 burden of D. similis, another grazing cladoceran species present in the reservoir. These
35 two species exhibited average Pb tissue concentrations of 57 and 98 ng Pb/mg,
36 respectively, with respective water column BCFs of 9,022 and 8,046. Conversely, an
37 examination of the simultaneous uptake of dissolved Pb by the algae P. subcapitata and
November 2012 7-149 Draft - Do Not Cite or Quote
-------
1 the cladoceran D. magna suggests that the dietary exposure route for the water column
2 filter-feeder is minor. Although Pb accumulated in the algal food source, uptake directly
3 from the water column was determined to be the primary route of exposure for D. magna
4 flComiarova and Blust. 2009c).
5 For many invertebrate species, sediment Pb concentrations may be the most important
6 driver in determining Pb uptake. For instance, while Hg and Cd body burdens in lentic
7 invertebrates were affected by lake ecological processes (e.g., eutrophication), a similar
8 effect was not observed for Pb concentrations in crayfish tissue, despite a high variability
9 between sites (Larsson et al.. 2007). Although this may be a result of differing
10 bioaccumulation tendencies, the authors suggested that other factors, including the
11 potential for sediment exposures, may be responsible for Pb uptake in lentic
12 invertebrates.
13 A field survey of fishes in the Viburnum Trend Pb-Zn mining district in southeast
14 Missouri available since the 2006 Pb AQCD, found that species richness and species
15 density of riffle-swelling benthic fishes were negatively correlated with metal
16 concentrations in pore water and in fish in mining impacted streams (Allert et al., 2009b).
17 Density of Ozark sculpin (Coitus hypselurus) and banded sculpin (Coitus carolinae) were
18 positively correlated with distance from mining sources.
19 In addition to the ecological effects discussed above, there is additional evidence that Pb
20 exposure could alter bacterial infection (and potentially disease transmission) in certain
21 fish species. Following 96-hour exposures to 4,000 (ig Pb/L, bacterial density in Channa
22 punctatus fish was observed to be significantly altered when compared to non-exposed
23 fish. Bacteria population densities in fish spleen, gills, liver, kidneys and muscle tissues
24 were higher following Pb exposure, with bacterial abundance in the gills too numerous to
25 quantify (Pathak and Gopal. 2009). In addition, bacteria inhabiting Pb-exposed fish were
26 more likely to exhibit antibacterial resistance than colonies isolated from non-exposed
27 fish. Although the mechanism remains unknown, this study suggests that Pb exposure
28 may increase the likelihood of infection in fish, potentially affecting fish abundance and
29 recruitment.
30 In summary, despite the fact that alterations of macrophyte communities may be highly
31 visible effects of increased sediment Pb concentrations, several recently published papers
32 propose that ecological impacts on invertebrate communities are also significant, and can
33 occur at environmental Pb concentrations lower than those required to impact plant
34 communities. High sediment Pb concentrations were linked to shifts in amphipod
35 communities inhabiting plant structures, and potentially to alterations in ecosystem
36 nutrient processing through selective pressures on certain invertebrate functional feeding
37 groups (e.g., greater bioaccumulation and toxic effects in collector-gatherers versus
November 2012 7-150 Draft - Do Not Cite or Quote
-------
1 predators or filter-feeders). Increased sediment pore water Pb concentrations were
2 demonstrated to likely be of greater importance to invertebrate communities, as well.
3 Interestingly, recent research also suggests that Pb exposure can alter bacterial
4 infestations in fish, increasing both microbial density and resilience, and potentially
5 increasing the likelihood of serious disease outbreak.
7.4.8 Critical Loads in Freshwater Systems
6 The general concept and definition of critical loads is introduced in Section 7.1.3 of this
7 chapter [also see Section 7.4 of the 2006 Pb AQCD (U.S. EPA. 2006c)1. Critical load
8 values are linked to critical limits of Pb for endpoints/receptors of interest in the
9 ecosystems, such as blood Pb. Some important critical limits for Pb in aquatic ecosystems
10 are discussed in this section along with information on aquatic critical loads for Pb.
11 Unit World Models (UWM) have been used to calculate critical loads for metals in
12 aquatic ecosystems. These models couple an ecotoxicity model, the BLM, to a
13 speciation/complexation model, the Windermere Humic Adsorption Model (WHAM),
14 then to the multi-species fate model, TRANsport-SPECiation (TRANSPEC). Gandhi et
15 al. apply the UWM to estimate speciation/complexation, fate and critical loads using
16 lakes of three different trophic status. A high percentage of colloidal-bound Pb was found
17 in the eutrophic and mesotrophic lakes (75-80%) versus the oligotrophic lakes (2%),
18 owing the high affinity of Pb to DOM. Pb concentrations were lowest for mesotrophic
19 and highest for oligotrophic systems. Critical loads were not calculated for Pb; however,
20 for the other metals tested the critical load was lowest in the oligotrophic and highest in
21 the eutrophic systems.
22 A critical load of 39.0 g Pb/m2-yr (0.19 mol Pb/ m2-yr) was calculated for a generalized
23 lake in the Sudbury area of the Canadian Shield using TICKET-UWM based on acute
24 toxicity data for D. magna. (Farley et al.. 2011). The model was set up to calculate
25 critical loads of metals by specifying free metal ion activity or the critical biotic ligand
26 concentration. This critical load for Pb was much higher than for Cu, Ni and Zn and the
27 authors attribute this difference to the strong binding of Pb to particulate organic matter
28 and the sequestration of PbCO3 in sediment.
29 As stated previously in Section 7.3.7. in the short term, metal emissions generally have
30 greater effects on biota in freshwater systems than in terrestrial systems because metals
31 are more readily immobilized in soils than in sediment. However, over the longer term,
32 terrestrial systems may be more affected particularly by those metals with a long soil
33 residence time, such as Pb. Thus, for a particular locale, either the terrestrial or the
34 aquatic ecosystem at that site may have the lower critical load. Given the heterogeneity of
November 2012 7-151 Draft - Do Not Cite or Quote
-------
1 ecosystems affected by Pb, and the differences in expectations for ecosystem services
2 attached to different land uses, it is expected that there will be a range of critical load
3 values for Pb for soils and waters within the U.S. Refer to Section 7.4.6 of the
4 2006 Pb AQCD for additional discussion of critical loads of Pb in aquatic systems.
7.4.9 Characterization of Sensitivity and Vulnerability in Freshwater Systems
5 Data from the literature indicate that exposure to Pb may affect survival, reproduction,
6 growth, metabolism, and development in a wide range of freshwater aquatic species.
7 Often, species differences in metabolism, sequestration, and elimination rates control
8 relative sensitivity and vulnerability of exposed organisms. Diet and lifestage at the time
9 of exposure also contribute significantly to the determination of sensitive and vulnerable
10 populations and communities. Further, environmental conditions in addition to those
11 discussed as affecting bioavailability (Sections 7.4.3 and 7.4.4) may also alter Pb toxicity.
12 The 2006 Pb AQCD (U.S. EPA. 2006b) reviewed the effects of genetics, age, and body
13 size on Pb toxicity. While genetics appears to be a significant determinant of Pb
14 sensitivity, effects of age and body size are complicated by environmental factors that
15 alter metabolic rates of aquatic organisms. A review of the more recent literature
16 corroborated these findings, and identified seasonally-affected physiological changes and
17 lifestage as other important determinants of differential sensitivity to Pb.
7.4.9.1 Seasonally-Affected Physiological Changes
18 A study by Duman et al. (2006) identified species and seasonal effects of Pb uptake in
19 aquatic plants. P. australis accumulated higher root Pb concentrations than S. lacustris.
20 Additionally, the P. australis Pb accumulation factor was significantly higher during the
21 winter versus other seasons, while the Pb accumulation factor for S. lacustris was greatest
22 in spring and autumn. The Pb accumulation factor for a third species, P. lucens, was
23 greatest in autumn (Duman et al.. 2006). Most significantly, these changes in
24 bioaccumulation were not linked with biomass increases, indicating that species-
25 dependent seasonal physiological changes may control Pb uptake in aquatic macrophytes
26 (Duman et al.. 2007). Significant interspecies differences in Pb uptake were observed for
27 plants representing the same genus (Sargassum), indicating that uptake of Pb by aquatic
28 plants also may be governed by highly species-dependent factors (Jothinayagi and
29 Anbazhagan. 2009).
30 Heier et al. (2009) established the speciation of Pb in water draining from a shooting
31 range in Norway and looked at the time dependent accumulation in brown trout. They
November 2012 7-152 Draft - Do Not Cite or Quote
-------
1 found that high molecular weight (>10 kDa) cationic Pb species correlated with high flow
2 episodes and accumulation of Pb on gills and in the liver. Thus, high flow episodes can
3 remobilize metals from a catchment and induce stress to aquatic organisms.
7.4.9.2 Increased Nutrient Uptake
4 Singh et al. (2010) proposed that metal-resistant plants have the capacity to not only up-
5 regulate antioxidant synthesis, but also have the ability to increase nutrient consumption
6 and uptake to support metal sequestration and detoxification via production of
7 antioxidants (Singh et al.. 2010). Therefore, it is likely that such plant species would be
8 significantly less susceptible to Pb exposure than those species without those abilities.
7.4.9.3 Temperature and pH
9 Water temperature also appears to affect the toxicity of Pb to aquatic organisms, with
10 higher temperatures leading to greater responses. Pb toxicity to crayfish increased 7 to
11 10% when the water temperature was increased by 4 °C, and by 14% when the
12 temperature increased by 7 °C. The authors determined that the increased toxicity was a
13 result of the negative impact of Pb on crayfish respiration, which was exacerbated by the
14 lower dissolved oxygen concentrations at higher water temperatures (Khan et al.. 2006).
15 In a study of the combined effects of temperature and Pb concentration on two freshwater
16 rotifer species, Brachionus havanaensis and B. rubens, population growth was measured
17 in three nominal concentrations of Pb as Pb chloride (50, 100 and 200 (ig Pb/L) for 15
18 days at either 22°C or 32°C (Montufar-Melendez et al.. 2007). At 22°C, population
19 growth of B. havanaensis was suppressed by B. rubens regardless of Pb treatment. At the
20 higher temperature, there was no population increase of B. rubens at any Pb
21 concentration. In the controls, population growth rates of B. havanaensis, but not
22 B. rubens, increased with an increase in temperature. These studies highlight the role of
23 temperature in Pb toxicity in organisms adapted to low temperatures.
24 The sequestration ability of L. minor macrophytes was similarly impacted by increased
25 surface water temperature; plants absorbed a maximum Pb concentration of 8.6 mg /g at
26 30 °C, while uptake at 15 °C was only 0.3 mg/g (Uvsal and Taner. 2009). Decreased pH
27 was also demonstrated to increase the uptake of environmental Pb in aquatic plants
28 (Wang et al.. 2010b: Uysal and Taner. 2009). Lower pH was shown to result in increased
29 sensitivity to Pb in juvenile fathead minnows in 30-day exposure to Pb of varying
30 concentrations (Grosell et al.. 2006a). Additionally, Birceanu et al. (2008) determined
31 that fish (specifically rainbow trout) were more susceptible to Pb toxicity in acidic, soft
November 2012 7-153 Draft - Do Not Cite or Quote
-------
1 waters characteristic of sensitive regions in Canada and Scandinavia. Hence, fish species
2 endemic to such systems may be more at risk from Pb contamination than fish species in
3 other habitats.
7.4.9.4 Lifestage
4 It is clear that certain stages of a life cycle are more vulnerable to Pb. A comparison of
5 C. riparius Pb LC50 values derived from toxicity tests with different instars indicates a
6 significant effect of lifestage on Pb sensitivity for aquatic invertebrates. Bechard et al.
7 (2008) calculated a first instar C. riparius 24-hour LC50 value of 613 (ig Pb/L, and
8 contrasted this value with the 24-hour and 48-hour LC50 values derived using later instar
9 larvae—350,000 and 200,000 (ig Pb/L, respectively. This disparity would suggest that
10 seasonal co-occurrence of aquatic Pb contamination and sensitive early instars could have
11 significant population-level impacts (Bechard et al.. 2008). Similarly, Wang et al. (2010e)
12 demonstrated that the newly transformed juvenile mussels, L. siliquoidea and
13 L. rafmesqueana, at 5 days old were more sensitive to Pb exposure than were glochidia
14 or two to six month- old juveniles, suggesting that Pb exposure at particularly sensitive
15 lifestages could have a significant influence on population viability (Wang et al., 2010e).
16 Evidence for differences in susceptibility to Pb at distinct lifestages is also available for
17 freshwater fish. In chronic (60-day) early-lifestage test exposures conducted with
18 rainbow trout to develop ACR's for this species the study resulted in ACR's for rainbow
19 trout lower than previously reported due to the acute tests which produced LC50 values
20 that were 10 to 25 times lower than earlier studies with trout. (Mebane et al., 2008). The
21 authors speculated that the lower LC50 values were due to the age of the fish used in the
22 study (two to four week old fry) and that testing with larger and older fish may not be
23 protective of more sensitive lifestages. Post-hatching stages of the African catfish were
24 more sensitive than the embryonic stage to Pb-exposure and the authors attributed this
25 apparent protective effect to the presence of a hardened chorion in embryos (Osman et
26 al.. 2007a).
7.4.9.5 Species Sensitivity
27 Species-specific Ca2+ requirements have been shown to affect the vulnerability of aquatic
28 organisms to Pb. The snail, L. stagnalis, exhibits an unusually high Ca2+ demand due to
29 CaCO3 formation required for shell production and growth, and exposure to Pb prevents
30 the uptake of needed Ca2+, leading to toxicity. Consequently, aquatic species that require
31 high assimilation rates of environmental Ca2+ for homeostasis are likely to be more
November 2012 7-154 Draft - Do Not Cite or Quote
-------
1 sensitive to Pb contamination (Grosell and Brix. 2009). Grosell and colleagues also noted
2 that reduced snail growth following chronic Pb exposure was likely a result of reduced
3 Ca2+ uptake (Grosell et al.. 2006b).
4 There is some indication that molting may comprise an additional sequestration and
5 excretion pathway for aquatic animals exposed to Pb (Soto-Jimenez et al.. 201 la:
6 Mohapatra et al.. 2009; Tollett et al.. 2009; Bergev and Weis. 2007). Libellulidae
7 dragonfly nymphs (Tollett et al.. 2009) have been shown to preferentially sequester Pb in
8 exoskeleton tissue. Consequently, aquatic arthropod species and those species that shed
9 their exoskeleton more frequently may be able to tolerate higher environmental Pb
10 concentrations than non-arthropods or slow-growing molting species, as this pathway
11 allows them to effectively lower Pb body burdens.
12 In contrast, the effect of Pb exposure on fish bacterial loads demonstrated by Pathak and
13 Gopal (2009) suggest that infected fish populations may be more at risk to the toxic
14 effects of Pb than healthier species. Aqueous Pb was demonstrated to both increase
15 bacteria density in several fish organs and to improve the likelihood of antibacterial
16 resistance (Pathak and Gopal. 2009).
17 Tolerance to prolonged Pb exposure may develop in aquatic invertebrates and fish. Multi-
18 generational exposure Pb appears to confer some degree of metal tolerance in
19 invertebrates such as C. plumosus larvae; consequently, previous population Pb
20 exposures may decrease species' susceptibility to Pb contamination fVedamanikam and
21 Shazilli, 2008b). However, the authors noted that metal tolerant larvae were significantly
22 smaller than larvae reared under clean conditions, and that transference of Pb-tolerant
23 C. plumosus larvae to clean systems resulted in a subsequent loss of tolerance. Evidence
24 of acclimation to elevated Pb in fathead minnow was suggested in the variations in
25 ionoregulatory parameters that were measured on day 10 and 30 in fish exposed to
26 115 (ig Pb/L for 30 days. At the end of the experiment, whole body Ca2+ was elevated
27 while Na+ and K+ recovered from elevated levels at 30 days (Grosell et al., 2006a).
28 A series of species sensitivity distributions constructed by Brix et al. (2005) in freshwater
29 systems indicated that sensitivity to Pb was greatest in crustacean species, followed by
30 coldwater fish, and warmwater fish and aquatic insects, which exhibited a similar
31 sensitivity. Further, analysis of both acute and chronic mesocosm data sets indicated that
32 Pb-contaminated systems exhibited diminished species diversity and taxa richness
33 following both types of exposure (Brix et al.. 2005). Wong et al. (2009) constructed Pb
34 species sensitivity distributions for both cladoceran and copepod freshwater species. A
35 comparison of the two curves indicated that cladoceran species, as a group, were more
36 sensitive to the toxic effects of Pb than were copepods, with respective hazardous
37 concentration values for 5% of the species (HC5) values of 35 and 77 (ig Pb/L. This
November 2012 7-155 Draft - Do Not Cite or Quote
-------
1 difference in sensitivities would indicate that cladoceran species are more likely to be
2 impacted at lower environmental Pb concentrations than copepods, potentially altering
3 community structures or ecosystem functions (Wong et al., 2009).
7.4.9.6 Ecosystem Vulnerability
4 Relative vulnerability of different freshwater ecosystems to effects of Pb can be inferred
5 from the information discussed above on species sensitivity and the influence of water
6 quality variables on the bioavailability and toxicity of Pb. It is, however, difficult to
7 categorically state that certain freshwater plant, invertebrate or vertebrate communities
8 are more vulnerable to Pb than others, since toxicity is dependent on many variables and
9 data from field studies are complicated by co-occurrence of other metals and alterations
10 of pH, such as in mining areas. Aquatic ecosystems with low pH and low DOM are likely
11 to be the most sensitive to the effects of atmospherically-deposited Pb. Examples of such
12 systems are acidic, soft waters such as sensitive regions in Canada and Scandinavia
13 (Birceanu et al.. 2008). In the U.S., aquatic systems that may be more sensitive to effects
14 of Pb include habitats that are acidified due to atmospheric deposition of pollutants,
15 runoff from mining activities or lakes and streams with naturally occurring organic acids.
16 Hence, fish and invertebrate species endemic to such systems may be more at risk from
17 Pb contamination than corresponding species in other habitats.
7.4.10 Ecosystem Services Associated with Freshwater Systems
18 Pb deposited on the surface of, or taken up by organisms has the potential to alter the
19 services provided by freshwater biota to humans although the directionality of impacts is
20 not always clear. For example, aquatic macrophytes provide a service by sequestering Pb.
21 At the same time, the uptake of Pb by plants may result in toxicological effects associated
22 with Pb exposure and decreased capacity of wetland species to remove contaminants. At
23 this time, few publications address Pb impacts on ecosystem services associated with
24 freshwater systems and most studies focus on wetlands rather than lakes and streams. Pb
25 can affect the ecological effects in each of the four main categories of ecosystem services
26 (Section 7.1.2) as defined by Hassan et al. (2005). These effects are sorted into ecosystem
27 services categories and summarized here:
28 • Supporting: food for higher trophic levels, biodiversity
29 • Provisioning: clean drinking water, contamination of food by heavy metals,
30 decline in health offish and other aquatic species
November 2012 7-156 Draft - Do Not Cite or Quote
-------
1 • Regulating: water quality
2 • Cultural: ecosystem and cultural heritage values related to ecosystem integrity
3 and biodiversity, wildlife and bird watching, fishing
4 Freshwater wetlands are sinks for atmospheric Pb as well as Pb from terrestrial runoff
5 (Landre etal.. 2010; Watmough and Dillon. 2007). Several studies have addressed the
6 response of natural wetlands to Pb (Odum. 2000; Gambrell. 1994). Recent reviews of
7 pollution control (Mander and Mitsch. 2009) or removal of metals (Marchand et al.,
8 2010) by constructed wetlands and phytoremediation of metals by wetland plants (Rai.
9 2008) indicate that these systems can remove Pb from the aquatic environment and are
10 important for water quality, sediment stabilization, nutrient cycling and shelter for
11 aquatic biota. The use of plants as a tool for immobilization of Pb and other metals from
12 the environment is not limited to wetland species. Recent advances in the
13 phytoremediation of metals are reviewed in Dickinson et al. (2009). The impact of Pb on
14 ecological services provided by specific components of aquatic systems has been
15 considered in a limited number of studies. For example, Theegala et al. (2007) discuss the
16 high uptake rate of Pb by D. pulex as the basis for a possible Daphnia-based remediation
17 for aquatic systems.
18
7.4.11 Synthesis of New Evidence for Pb Effects in Freshwater Ecosystems
19 This synthesis of the effects of Pb on freshwater ecosystems covers information from the
20 publication of the 2006 Pb AQCD to present. It is followed in Section 7.4.12 by
21 determinations of causality that take into account evidence dating back to the 1977 Pb
22 AQCD.
23 Evidence assessed in the present document supports the findings of the previous Pb
24 AQCDs that waterborne Pb is highly toxic to freshwater organisms, with toxicity varying
25 with species and lifestage, duration of exposure, form of Pb, and water quality
26 characteristics. The studies that are available for freshwater plants, invertebrates and
27 vertebrates include studies where Pb concentration was analytically verified and those
28 that reported nominal concentrations (Table 7-5). Many of the studies that report nominal
29 concentrations in media are uptake studies that subsequently quantify Pb in tissues,
30 however, measurement of Pb in water or sediment at the beginning of an exposure is
31 desirable when comparing laboratory studies to concentrations of Pb in freshwater
32 systems. As reported in Section 7.2.3 and Table 7-2. the median and range of Pb
33 concentrations in surface waters (median 0.50 ug Pb/L, range 0.04 to 30 ug Pb/L) and
November 2012 7-157 Draft - Do Not Cite or Quote
-------
1 sediments (median 28 mg Pb/kg dry weight, range 0.5 to 12,000 mg Pb/kg dry weight) in
2 the U.S. based on a synthesis of NAWQA data was reported in the previous
3 2006 Pb AQCD (U.S. EPA. 2006c).
4 Recent studies provide further evidence for the role of modifying factors such as pH,
5 DOC and hardness on the effects of Pb on plants, invertebrates and vertebrates. The same
6 Pb concentration added to water or sediment produces far greater effects under some
7 conditions, than others. Many studies reviewed in the ISA included concentrations that
8 were higher than Pb found near contaminated areas. However, when multiple
9 concentrations were used, effects gradually increased with increasing Pb exposure.
10 Effects at lower concentrations can be implied from many studies since an exposure-
11 response relationship to Pb was observed, although uncertainty remains in relating these
12 findings to reported concentrations of Pb in freshwater. Many studies only report an LC50
13 value when an LOEC or LCio would be more relevant for consideration of effects on
14 organisms since an effect occurring at the LC50 would most likely not maintain a stable
15 population. Most available studies only report acute toxicity and are conducted at higher
16 concentrations of Pb than found in sampling from U.S. surface waters (Table 7-2).
17 however, exposure to Pb in freshwater systems is most likely characterized as a chronic
18 low dose exposure.
Plants
19 Most recent studies on effects of Pb in freshwater algal species reviewed in Section
20 7.4.5.1 were conducted with nominal media exposures and effect concentrations greatly
21 exceeded Pb reported in surface water. In studies where Pb was quantified, effect
22 concentrations for growth (EC50) for aquatic macrophytes were much higher than
23 currently reported ambient Pb, however, some sublethal endpoints such as effects on
24 chlorophyll were observed at lower concentrations. For example, chlorophyll a content
25 was significantly inhibited at 210 (ig Pb/L and higher in W. arrhiza (Piotrowska et al..
26 2010). An increase in biomass was reported in L. minor exposed to 100 or 200 (ig Pb/L
27 with inhibition observed at higher concentrations (Dirilgen. 2011). There were also
28 numerous studies conducted at nominal Pb concentration that report effects on enzyme
29 activities and protein content in freshwater aquatic plant species. Exposure-response
30 relationships in which increasing concentrations of Pb lead to increasing effects were
31 consistently observed for freshwater aquatic plants.
32 Recent studies on bioavailability of Pb in aquatic plants and algae support the findings of
33 previous Pb AQCDs that plants tend to sequester larger amounts of Pb in their roots than
34 in their shoots and provide additional evidence for species differences in
35 compartmentalization of sequestered Pb and responses to Pb in water and sediments.
November 2012 7-158 Draft - Do Not Cite or Quote
-------
1 Given that atmospherically-derived Pb is likely to become sequestered in sediments,
2 uptake by aquatic plants is a significant route of Pb removal from sediments, and a
3 potential route for Pb mobilization into the aquatic food web. Although there are some
4 similarities to Pb accumulation observed in terrestrial plants (e.g., preferential
5 sequestration of the metal in root tissue), Pb appears to be more bioavailable in sediment
6 than it is in soil.
Invertebrates
7 The largest body of evidence for effects of Pb at or near concentrations of this metal
8 found in surveys of surface waters of the U.S. is for invertebrates and recent studies
9 reviewed in Sections 7.4.5.2 and 7.4.6 further support this observation. Exposure-
10 response relationships in which increasing concentrations of Pb lead to increasing effects
11 were consistently observed for freshwater invertebrates. Among the most sensitive
12 species, growth of juvenile freshwater snails L. stagnalis was inhibited at an EC20 of
13 <4 (ig Pb/L. (Grosell and Brix. 2009: Grosell et al. 2006b). A chronic value of
14 10 (ig Pb/L obtained in 28-day exposures of 2-month-old fatmucket mussel,
15 L. siliquoidea juveniles was the lowest genus mean chronic value ever reported for Pb
16 (Wang etal.. 2010e). The 96-hour EC50 values for 5-day-old juveniles in two separate
17 toxicity tests with this species were 142 and 298 (ig Pb/L (mean EC50 220 (ig Pb/L).
18 Recent studies (Sections 7.4.5.2 and 7.4.6) have further elucidated the role of water
19 quality on Pb toxicity. In freshwater invertebrates some effects were observed at
20 concentrations occasionally encountered in U.S. surface waters (Table 7-2). In a 7-day
21 exposure of the cladoceran C. dubia to 50 to 500 (ig Pb/L, increased DOC leads to an
22 increase in mean EC50 for reproduction ranging from approximately 25 (ig Pb/L to
23 >500 (ig Pb/L (Mager et al.. 201 la). The 48-hour LC50 values for the cladoceran C. dubia
24 tested in eight natural waters across the U.S. varied from 29 to 1,180 (ig Pb/L and were
25 correlated with DOC (Esbaugh etal.. 2011).
26 Additional new evidence reviewed in Sections 7.4.5.2 and 7.4.6 for effects near the upper
27 range of concentrations of Pb available from surveys of U.S. surface waters include
28 studies with rotifer, midge and mayfly species. The freshwater rotifer E. dilatata 48 hour
29 LC50 was 35 (ig Pb/L using neonates hatched from asexual eggs (Arias-Almeida and
30 Rico-Martinez. 2011). An EC2o for reduced growth and emergence of the midge
31 C. dilutus was reported to be 28 (ig Pb/L, observed in a 55-day exposure, while the same
32 species had a 96-hour LC50 of 3,323 (ig Pb/L (Mebane et al.. 2008) The ECio for molting
33 in the mayfly B. tricaudatus was 37 (ig Pb/L (Mebane et al., 2008). All of these effect
34 concentrations provide additional evidence for Pb effects on freshwater invertebrates.
November 2012 7-159 Draft - Do Not Cite or Quote
-------
Vertebrates
1 For freshwater fish (Sections 7.4.5.3 and 7.4.6). most recent studies available since the
2 2006 Pb AQCD, were conducted with fathead minnow, P. promelas, or rainbow trout,
3 O. mykiss. In a series of 96-hour acute toxicity tests with fathead minnow conducted in a
4 variety of natural waters across North America, LC50 values ranged from 41 to
5 3,598 (ig Pb/L (Esbaugh et al.. 2011). Reproductive effects associated with water quality
6 parameters were also noted with this species (Mager et al.. 2010). In trout, no effects of
7 Pb were observed in dietary studies. In chronic aqueous exposures with trout the
8 following endpoints were reported: NOEC=24 (ig Pb/L, ECi0=26 (ig Pb/L,
9 EC2o=34 (ig Pb/L, and LC50=55 (ig Pb/L. In a separate test with the same species an
10 NOEC=8 ng Pb/L, EC10=7(ig Pb/L, EC20=102 ng Pb/L and LC50=120 ng Pb/L were
11 reported. In acute tests with rainbow trout run concurrently with the chronic tests,
12 96-hour LC50 values were 120 and 150 \ig Pb/L, respectively (Mebane et al.. 2008).
13 These reported effects provide additional evidence for toxicity of Pb to fish and chronic
14 NOEC and ECio values reported for trout, a sensitive species, are within the upper range
15 of Pb currently reported in U.S. surface waters (Table 7-2).
16 In Section 7.4.5.3. a study with the frog R. pipiens exposed nominally to Pb, tissue
17 concentrations were quantified at the end of the study and found to be in the range of Pb
18 tissue concentrations in wild-caught tadpoles. Growth rate was significantly slower in the
19 100 (ig Pb/L nominal concentration and more than 90% of tadpoles developed lateral
20 spinal curvature. Time to metamorphosis was also delayed at this treatment level.
Food Web
21 In the 2006 Pb AQCD, trophic transfer of Pb through aquatic food chains was considered
22 to be negligible. Concentrations of Pb in the tissues of aquatic organisms were generally
23 higher in algae and benthic organisms than in higher trophic-level consumers indicating
24 that Pb was bioaccumulated but not biomagnified (U.S. EPA. 2006c: Eisler. 2000). Some
25 studies published since the 2006 Pb AQCD, (see Section 7.4.4.4) support the potential for
26 Pb to be transferred in aquatic food webs, while other studies indicate that Pb
27 concentration decreases with increasing trophic level (biodilution).
Community and Ecosystem Effects
28 New evidence of effects of Pb at the community and ecosystem levels of biological
29 organization reviewed in Section 7.4.7 include shift in community composition in
30 macrophytes. Effects on reproduction, growth or survival (summarized in Table 7-5) may
31 lead to effects at the population-level of biological organization and higher. Additional
November 2012 7-160 Draft - Do Not Cite or Quote
-------
1 evidence for community and ecosystem level effects of Pb have been observed primarily
2 in microcosm studies or field studies with other metals present.
7.4.12 Causal Determinations for Pb in Freshwater Systems
3 In the following sections, organism-level effects on reproduction and development,
4 growth and survival are considered first since these endpoints can lead to effects at the
5 population level or above and are important in ecological risk assessment.
6 Neurobehavioral effects are considered next followed by sub-organismal responses
7 (hematological effects, physiological stress) for which Pb has been shown to have an
8 impact in multiple species and across taxa, including humans. Causal determinations for
9 terrestrial, freshwater and saltwater ecological effects are summarized in Table 7-3.
7.4.12.1 Reproductive and Developmental Effects-Freshwater Biota
10 Evaluation of the findings in previous Pb AQCDs and recent literature on Pb effects in
11 aquatic fauna indicates that exposure to Pb is associated with reproductive effects at or
12 near ambient concentrations of this metal (Table 7-2) in some freshwater species.
13 Impaired fecundity at the organismal level can result in a decline in abundance and/or
14 extirpation of populations, decreased taxa richness, and decreased relative or absolute
15 abundance at the community level (Suter etal. 2005; U.S. EPA. 2003a). Various
16 endpoints have been measured in freshwater organisms to assess the effect of Pb on
17 fecundity, development and hormone homeostasis. However, there are typically only
18 limited studies available from different taxa. Recent evidence available since the
19 2006 Pb AQCD for effects of Pb on reproductive endpoints in freshwater invertebrates
20 and vertebrates is summarized in Table 7-5.
21 There are no studies reviewed in the ISA or previous Pb AQCDs on development and
22 reproductive effects of Pb in freshwater aquatic algae or macrophytes.
23 Experimental data from freshwater invertebrates provide evidence for increasing
24 reproductive effects associated with increasing exposure to Pb. The exposure-response
25 relationship is used in judging causality (Table I Preamble). Reproductive effects of Pb in
26 freshwater aquatic invertebrates are well-characterized in previous Pb AQCDs, the draft
27 Ambient Water Quality Criteria for Pb (U.S. EPA. 2008b) and in the current ISA and
28 have been observed at or near current ambient concentrations (median 0.5 (ig Pb/L, range
29 0.04 to 30 (ig Pb/L) (U.S. EPA. 2006c) in some species in laboratory exposures. In the
30 1986 Pb AQCD reproductive effects were reported to begin at 19 (ig Pb/L for the
31 freshwater snail Lymnaeapalustris and 27 (ig Pb/L for Daphnia sp. (U.S. EPA. 1986b).
November 2012 7-161 Draft - Do Not Cite or Quote
-------
1 In the 2006 Pb AQCD (U.S. EPA. 2006b) the number of neonates per surviving adult was
2 significantly decreased in the amphipod H. azteca during chronic 42-day exposures to Pb
3 (Besser et al., 2005). In the group exposed to Pb in water-only exposures, the LOEC for
4 reproductive effects was 16 (ig Pb/L while in amphipods receiving both water-borne and
5 dietary Pb the LOEC for reproduction was 3.5 (ig Pb/L.
6 New evidence in freshwater invertebrates (Table 7-5 and Section 7.4.5.2) show
7 consistency of the observed association between reproductive endpoints and Pb exposure.
8 In the freshwater rotifer B. calyciflorus, reproductive output was measured as total
9 number of individuals and intrinsic growth rate. The EC2o for number of rotifers was
10 125 (ig Pb/L and the 48 hour EC2o for intrinsic rate of population increase was
11 307 (ig Pb/L with an NOEC of 194 (ig Pb/L (Grosell et al.. 2006b). In a 7-day exposure
12 of the cladoceran C. dubia to 50 to 500 (ig Pb/L, increased DOC leads to an increase in
13 mean EC50 for reproduction ranging from approximately 25 (ig Pb/L to >500 (ig Pb/L
14 (Mager et al.. 201 la). Additional reproductive impairment endpoints for freshwater
15 cladocerans are reported in Table 6 of the draft Ambient Aquatic Life Water Quality
16 Criteria for Pb (U.S. EPA. 2008b). It is not clear how these laboratory-derived values for
17 freshwater invertebrates compare to Pb exposures in natural systems due to the role of
18 modifying factors (i.e., pH, hardness, and DOC) which affect Pb speciation and
19 bioavailability, however, results under controlled conditions have consistently shown
20 reproductive effects of Pb in sensitive taxa (amphipods, cladocerans) at concentrations at
21 or near Pb quantified in freshwater environments.
22 In freshwater aquatic vertebrates there is evidence for reproductive and developmental
23 effects of Pb. Pb exposure in frogs has been demonstrated to delay metamorphosis,
24 decrease larval size and produce skeletal malformations. For example, in northern
25 leopard frog R. pipiens exposed to nominal concentrations of 100 \ig Pb/L from
26 embryonic stage to metamorphosis, maximum swimming speed was significantly slower
27 than other treatment groups and 92% of tadpoles exposed to 100 (ig Pb/L had lateral
28 spinal curvature (compared with 6% in the control) (Chen et al., 2006b). Pb tissue
29 concentrations were quantified in frogs following exposure and fell within the range of
30 tissue concentrations in wild-caught tadpoles.
31 The weight of evidence for reproductive and developmental effects in freshwater
32 vertebrates is from studies with fish. Pb AQCDs have reported developmental effects in
33 fish, specifically spinal deformities in brook trout (Salvelinus fontinalis) exposed to
34 119 ng Pb/L for three generations (U.S. EPA. 1977). and in rainbow trout as low as
35 120 ng Pb/L (U.S. EPA. 1986b). Reproductive behaviors of fathead minnows including
36 reduced time spent in nest preparation by males, increased interspawn periods and
37 reduced oviposition by females was observed following a 4-week exposure to
November 2012 7-162 Draft - Do Not Cite or Quote
-------
1 500 ng Pb/L fWeber. 1993). In the 2006 Pb AQCD (U.S. EPA. 2006b). decreased
2 spermatocyte development in rainbow trout was reported at 10 (ig Pb/L and, in fathead
3 minnow testicular damage occurred at 500 (ig Pb/L. In a recent study, reproductive
4 effects in fathead minnows were influenced by water chemistry parameters (alkalinity
5 and DOC) in breeding exposures following 300 day chronic toxicity testing with Pb
6 (Maaeretal.. 2010). Specifically, in fish treated in both 35 and 120 jig Pb/L with HCO3"
7 and with 120 (ig Pb/L with DOC, total reproductive output was decreased and average
8 egg mass production increased as compared to egg mass size in controls and in low
9 HCO3" and DOC treatments with Pb. No significant differences were present between
10 treatments in egg hatchability. In a feeding study, Reproductive performance was
11 unaffected in zebrafish exposed to Pb-via consumption of contaminated prey (Boyle et
12 al., 2010). In fish, there is evidence for alteration of steroid profiles and additional
13 reproductive parameters although most of the available studies were conducted using
14 nominal concentrations of Pb.
15 Observations of Pb toxicity to reproductive and developmental endpoints in freshwater
16 fauna are further supported by evidence in terrestrial invertebrates and vertebrates
17 (Section 7.3.12.1). marine invertebrates (Section 7.4.21.1) and from laboratory animals
18 (Section 5.8). Pb appears to affect multiple endpoints associated with reproduction and
19 development in aquatic invertebrates and vertebrates. A few sensitive invertebrate taxa
20 (amphipods, cladocerans) have been identified where effects are observed in laboratory
21 studies at concentrations of Pb that occur in the environment. Overall, there is a dearth of
22 information on reproductive effects of Pb in natural environments, however, the weight
23 of evidence is sufficient to conclude that there is a causal relationship between Pb
24 exposures and developmental and reproductive effects in freshwater invertebrates and
25 vertebrates. In freshwater plants, the evidence is inadequate to conclude that there is a
26 causal relationship between Pb exposures and plant developmental and reproductive
27 effects.
7.4.12.2 Growth Effects-Freshwater Biota
28 Alterations in the growth of an organism can impact population, community and
29 ecosystem level variables. Growth is a commonly measured endpoint in aquatic plants,
30 however, reported effects typically occur at concentrations that exceed Pb quantified in
31 freshwater habitats. Growth effects of Pb on plants include visible growth responses and
32 reduction of photosynthetic rate, inhibition of respiration, cell elongation, root
33 development or premature senescence (U.S. EPA. 1986b). In the 2006 Pb AQCD (U.S.
34 EPA. 2006b). both freshwater algae and plants had EC50 values for growth in the range of
35 1,000 to >100,000 (ig Pb/L with minimal inhibition of growth observed as low as
November 2012 7-163 Draft - Do Not Cite or Quote
-------
1 100 (ig Pb/L (U.S. EPA. 2006c). The most sensitive aquatic macrophyte reported in the
2 2006 Pb AQCD was A. pinnata with an EC50 for relative growth rate of 1,100 (ig Pb/L
3 following a 4-day exposure to Pb (Gauretal. 1994). An LOEC of 25 jig Pb/L for
4 reduced chlorophyll in Coontail (Ceratophyllum demersum), and 50 (ig Pb/L in Cattail
5 (71. latifolid) following 12-day exposure to Pb (as Pb acetate) were the lowest reported
6 concentrations of growth-related effects in freshwater plants in the draft Ambient Aquatic
7 Life Water Quality Criteria for Pb (U.S. EPA. 2008b) and were near the upper range of
8 Pb values reported from sampling of U.S. surface waters (Table 7-2). Additional growth
9 studies in freshwater algae and plants summarized in Table 6 of the draft Ambient
10 Aquatic Life Water Quality Criteria for Pb and Table 7-5 of the present document report
11 growth effects in laboratory studies at concentrations that exceed measured levels of Pb
12 in the aquatic environment (U.S. EPA. 2008b).
13 Most of the evidence for growth effects of Pb in freshwater biota is for invertebrates.
14 Some of these studies report inhibition of growth in sensitive species occurring at or near
15 the current upper range of Pb in surface waters (median 0.50 (ig Pb/L, range 0.04 to
16 30 (ig Pb/L) (U.S. EPA. 2006c). Growth effects of Pb on aquatic invertebrates are
17 reviewed in the draft Ambient Aquatic Life Water Quality Criteria for Pb (U.S. EPA.
18 2008b) and the 2006 Pb AQCD. The lowest reported LOEC for growth in the
19 2006 Pb AQCD (16 (ig Pb/L) was in amphipods (H. azteca) in a 42-day chronic exposure
20 (Besser et al. 2005).
21 Recent studies provide additional evidence for effects on growth of freshwater aquatic
22 invertebrates at < 10 (ig Pb/L. Growth effects observed in invertebrates underscores the
23 importance of lifestage to overall Pb sensitivity. In general, juvenile organisms are more
24 sensitive than adults. Growth of juvenile freshwater snails L. stagnalis was inhibited
25 below the lowest concentration tested resulting in an EC2o <4 (ig Pb/L (Grosell and Brix.
26 2009; Grosell et al.. 2006b). In the same study, the NOEC was 12 jig Pb/L and the LOEC
27 was 16 (ig Pb/L. The authors indicated that the observed effect level for Pb was very
28 close to the current U.S. EPA water quality criteria for Pb (3.3 (ig Pb/L normalized to test
29 water hardness) (Grosell and Brix. 2009). In the freshwater mussel, fatmucket (L.
30 siliquoided) juveniles were the most sensitive lifestage (Wang et al., 2010e). In this
31 study, growth of juvenile mussels at the end of a 28-day exposure in 17 (ig Pb/L was
32 significantly reduced from growth in the controls. A chronic value of 10 (ig Pb/L in 2-
33 month-old fatmucket juveniles was the lowest genus mean chronic value ever reported
34 for Pb. The ECi0 and EC20 for reduced growth and emergence of the midge C. dilutus in a
35 55-day exposure were 28 (ig Pb/L and 55 (ig Pb/L, respectively, while the same species
36 had a 96-hour LC50 of 3,323 (ig Pb/L (Mebane et al.. 2008) The EC10 and EC20 for
37 molting in the mayfly B. tricaudatus were 37 (ig Pb/L and 66 (ig Pb/L, respectively
38 (Mebane et al.. 2008). In natural freshwater systems the effects of Pb are influenced by
November 2012 7-164 Draft - Do Not Cite or Quote
-------
1 additional factors (i.e., pH, hardness, and DOC) which may modulate the toxicity of Pb
2 observed under laboratory conditions.
3 Evidence for growth effects of Pb in freshwater aquatic vertebrates is limited to a few
4 studies in amphibians and fish. In the 2006 Pb AQCD growth effects of Pb were reported
5 in frogs at concentrations typically higher than currently found in the environment. A
6 recent study supports findings of growth effects in frogs and suggests that these effects
7 may be occurring at lower concentrations: the growth rate of tadpoles of the northern
8 leopard frog exposed nominally to 100 (ig Pb/L from embryo to metamorphosis was
9 slower than the growth rate of the controls (Chen et al.. 2006b). In this study, Pb
10 concentrations in the tissues of tadpoles were quantified and the authors reported that
11 they were within the range of reported tissue concentrations reported in wild-caught
12 populations.
13 Reports of Pb-associated growth effects in freshwater fish are inconsistent (Mager. 2012).
14 In a review cited in the 2006 Pb AQCD, general symptoms of Pb toxicity in fish included
15 growth inhibition (Eisler. 2000) however, other studies with Pb have shown no effects on
16 growth (Mager. 2012). In the studies reviewed for the current ISA no growth effects were
17 observed in fish exposed to Pb via dietary intake. Recent aqueous exposure studies with
18 fathead minnows showed significant increases in body length and body mass following
19 chronic low Pb exposure, however, the authors noted that some effects were observed in
20 tanks with high mortality early in the exposure (Mager and Grosell. 2011; Grosell et al.,
21 2006a). Other studies with fathead minnows have shown growth reductions with Pb
22 exposure, however, concentrations of observed effects typically exceeded the 96-hour
23 LC50 value (Mager. 2012: Mager etal.. 2010: Grosell et al.. 2006a). Two 60-day early
24 lifestage tests with rainbow trout showed differences in LOEC for reduced growth
25 (Mebane et al.. 2008). In the first test, a 69-day exposure, the LOECs for mortality and
26 reduced growth were the same (54 (ig Pb/L). In the second test, a 62-day exposure of Pb
27 to rainbow trout, the LOEC for fish length was 18 (ig Pb/L with an EC2o of >87 (ig Pb/L.
28 Evidence of effects of Pb exposure on growth in terrestrial plants (Section 7.3.12.2) is
29 highly coherent with evidence from freshwater plants. Although there is a lack of
30 evidence in freshwater plants for growth effects at concentrations of Pb typically
31 encountered in U.S. surface waters, several studies suggest that minimal growth
32 inhibition can occur within one to two orders of magnitude of the reported range for
33 freshwater. Due to the concentration-response relationship observed between Pb exposure
34 and freshwater plants, growth is likely impacted at lower, more ecologically relevant
35 ECio or LOEC values, than the typically reported EC50 values which may not be suitable
36 for a maintaining a sustainable population.
November 2012 7-165 Draft - Do Not Cite or Quote
-------
1 There is a large body of evidence to support growth effects of Pb on aquatic plants at
2 concentrations that greatly exceed those typically found in U.S. surface waters. Less
3 evidence is available at current concentrations of Pb measured in U.S. surface waters and
4 within one to two orders of magnitude above the range of these measured values. The
5 available evidence is, however, sufficient to conclude that a causal relationship is likely
6 to exist between Pb exposures and growth effects in freshwater plants. The evidence is
7 sufficient to conclude that there is a causal relationship between Pb exposures and growth
8 effects in aquatic invertebrates. Available evidence is inadequate to conclude that there is
9 a causal relationship between Pb exposures and growth effects in aquatic vertebrates.
7.4.12.3 Survival-Freshwater Biota
10 The relationship between Pb exposure and survival has been well demonstrated in
11 freshwater species as presented in Section 7.4.5 and Table 7-5 of the present document
12 and in the previous Pb AQCDs. Pb exposure can either result in direct lethality or
13 produce sublethal effects that diminish survival probabilities. Survival is a biologically
14 important response that can have a direct impact on population size. However, the
15 concentration typically reported at which there is 50% mortality of test organisms (LC50)
16 is a poor measure for consideration of effects at ecologically-relevant concentrations.
17 LC50 is a measure for acute toxicity whereas Pb effects on ecosystem receptors are likely
18 characterized as a chronic, cumulative exposure rather than acute exposure. Furthermore,
19 a scenario in which 50% of a population does not survive is likely not a sustainable
20 population. From the LC50 data on Pb in this review and previous Pb AQCDs, a wide
21 range of sensitivity to Pb is evident across taxa and within genera. However, the LC50 is
22 usually much higher than current environmental levels of Pb in the U.S, even though
23 physiological dysfunction that adversely impacts the fitness of an organism often occurs
24 at concentrations well below lethal ones. When available, LCio, NOEC or LOEC are
25 therefore reported.
26 There are no studies reported in the previous Pb AQCDs or the current ISA for aquatic
27 plants that indicate phytotoxicity at current concentrations of Pb in freshwater
28 environments.
29 There are considerable data available on toxicity of Pb to aquatic invertebrates as
30 reviewed in the previous Pb AQCDs and Ambient Water Quality Criteria for Lead (U.S.
31 EPA. 1985) (U.S. EPA. 2008b). Table AX7-2.4.1 from the 2006 Pb AQCD summarizes
32 LC50 data and other endpoints for freshwater and marine invertebrates (U.S. EPA.
33 2006c). Recent studies available since the 2006 Pb AQCD and draft Aquatic Life Water
34 Quality Criteria for Pb that report mortality data are summarized in Table 7-5. Freshwater
November 2012 7-166 Draft - Do Not Cite or Quote
-------
1 invertebrates are generally more sensitive to Pb exposure than other taxa, with survival
2 impacted in a few species at or near concentrations that are encountered in aquatic
3 environments (Table 7-2). These impacted species may include candidate or endangered
4 species. For example, the freshwater mussel L. rafinesqueana (Neosho mucket), is a
5 candidate for the endangered species list. The EC50 for foot movement (a measure of
6 viability) for newly transformed juveniles of this species was 188 (ig Pb/L. (Wang et al..
7 2010e).
8 Most of the evidence for Pb effects on survival in freshwater invertebrates is from
9 sensitive species of gastropods, amphipods, cladocerans and rotifers (Sections 7.4.5.2 and
10 7.4.6). In some of these organisms, increased mortality is observed in the upper range of
11 Pb concentration values found in surveys of U.S. surface waters (median 0.50 (ig Pb/L,
12 range 0.04 to 30 (ig Pb/L) (U.S. EPA. 2006c). although the toxicity of Pb is highly
13 dependent upon water quality variables such as DOC, hardness and pH. In the 1986 Pb
14 AQCD, increased mortality was reported in the freshwater gastropod Lymnaeapalutris at
15 Pb concentration as low as 19 (ig Pb/L effectively reducing total biomass production
16 (Borgmann et al.. 1978). Toxicity testing with amphipods under various water conditions
17 indicate these organisms are sensitive to Pb at <10 (ig Pb/L (U.S. EPA. 2006c) and the
18 present document). A 7 day LC50 of 1 (ig Pb/L was observed in soft water with the
19 amphipod H. azteca ^Borgmann et al.. 2005,). In this same species, the 96-hour LC50 for
20 Pb at pH of 5 was 10 (ig Pb/L (Mackie. 1989). In 42-day chronic exposures ofH. azteca
21 exposed to Pb via water and diet, the LC50 was 16 (ig Pb/L (Besser et al.. 2005). At
22 higher pH and water hardness, amphipods are less sensitive to Pb (U.S. EPA. 2006c). In a
23 series of 48 hour acute toxicity tests with the cladoceran C. dubia conducted in a variety
24 of natural waters across North America, LC50 values ranged from 29 to 1,180 (ig Pb/L
25 (NOEC range 18 to <985 (ig Pb/L) and were most significantly influenced by DOC and
26 water ionic strength (Esbaugh et al.. 2011). In the 2006 Pb AQCD the range of 48 hour
27 LC50 values for C. dubia were reported from 280 to >2,700 (ig Pb/L when tested at
28 varying pH levels (U.S. EPA. 2006c). In the rotifer genus Lecane, a 22-fold difference in
29 LC50 values was observed in 48-hour exposure to Pb between L. hamata, L. luna and
30 L. quadridentata. (Perez-Legaspi and Rico-Martinez. 2001). L. luna was most sensitive to
31 Pb toxicity with a 48-hour LC50 of 140 (ig Pb/L. In neonate rotifers, E. dilatata the 48-
32 hour LC50 was 35 (ig Pb/L (Arias-Almeida and Rico-Martinez. 2011). A wide range of
33 LC50 values were reported for chironomid species (Table 7-5). however, the available
34 evidence suggests these freshwater invertebrates are less sensitive to Pb than amphipods,
35 cladocerans and rotifers. Other freshwater invertebrates such as odonates may be tolerant
36 of Pb concentrations that greatly exceed concentrations of Pb reported in environmental
37 media. Some invertebrates are able to detoxify Pb such as through sequestration of Pb in
38 the exoskeleton which is subsequently molted.
November 2012 7-167 Draft - Do Not Cite or Quote
-------
1 There is considerable historic information on Pb toxicity to freshwater fish. Early
2 observations from mining areas where Pb and other metals were present indicated local
3 extinction offish from streams (U.S. EPA. 1977). The lowest LC50 for fish reported in the
4 1977 Pb AQCD was 1,000 (ig Pb/L in soft water for rainbow trout O. mykiss (reclassified
5 from Salmo gairdneri} following 96-hour exposure to Pb (U.S. EPA. 1977). Additional
6 LC50 values for freshwater fish are summarized in the 1985 Ambient Water Criteria for
7 Pb (U.S. EPA. 1985) and the draft Ambient Aquatic Life Water Quality Criteria for Pb
8 (U.S. EPA. 2008b). An LC50 of 236 jig Pb/L adjusted to a total hardness of 50 mg/L
9 CaCO3 was reported for O. mykiss in the draft Ambient Aquatic Life Water Quality
10 Criteria for Pb.
11 More recently reviewed studies using fish have considered the role of water quality
12 variables and bioavailability on Pb toxicity. Higher toxicity tends to occur in acidic
13 waters where more free-Pb ion is available for uptake. The interactive effects of Pb
14 concentration and water quality variables on toxicity may result in equivalent toxicity for
15 a broad range of Pb concentrations. In a series of 96-hour acute toxicity tests with
16 juvenile fathead minnow conducted in a variety of natural waters across North America,
17 LC50 values ranged from 41 to 3,598 (ig Pb/L and no Pb toxicity occurred in three highly
18 alkaline waters (Esbaugh et al.. 2011). In the 2006 Pb AQCD, the 96-hr LC50 values in
19 fathead minnow ranged from 810->5,400 (ig Pb/L in varying pH and hardness (U.S.
20 EPA. 2006c).
21 Decreased survival is also a function of age of the fish. Thirty day LC50 values for larval
22 fathead minnows ranged from 39 to 1,903 (ig Pb/L in varying concentrations of DOC,
23 CaSO4 and pH (Grosell et al.. 2006b). In a recent study of rainbow trout fry at 2 to 4
24 weeks post swim-up, the 96-hour LC50 was 120 (ig Pb/L at a hardness of 29 mg/L as
25 CaCO3, a value much lower than in testing with older fish (Mebane et al.. 2008). In the
26 same study, two chronic (>60 day) tests were conducted with rainbow trout and the
27 NOECs for survival were 24 and 87 (ig Pb/L and the LOECs were 54 and 125 (ig Pb/L,
28 respectively. In contrast to aqueous exposures, 30 day dietary studies with rainbow trout
29 fathead minnow, and channel catfish fed a live diet of L. variegatus contaminated with Pb
30 showed no statistically significant effects on survival (Erickson et al.. 2010).
31 Freshwater fish are less sensitive to Pb than freshwater invertebrates, however, recent
32 studies have highlighted the importance of considering pH, hardness and additional
33 modifying factors in assessing toxicity since effects can vary over several orders of
34 magnitude. Fish mortalities occur above the concentrations of Pb encountered in U.S.
35 surface waters although, in some cases, the observed effects may be just above the upper
36 measured range of Pb in some aquatic environments (Table 7-2). Furthermore, although
37 LC50 values are the most commonly reported, effects are occurring at lower
November 2012 7-168 Draft - Do Not Cite or Quote
-------
1 concentrations. A more relevant indication of exposure impacts would be an LCi0 or
2 LOEC, however, these values are not always provided. The evidence is sufficient to
3 conclude that there is a causal relationship between Pb exposures and survival in
4 freshwater invertebrates and vertebrates. The evidence is inadequate to conclude that
5 there is a causal relationship between Pb exposures and survival in freshwater plants.
7.4.12.4 Neurobehavioral Effects-Freshwater Biota
6 Evidence from laboratory studies and limited data from field studies reviewed in this
7 chapter, in the draft Ambient Aquatic Criteria document for Pb which updates the 1985
8 Ambient Water Quality Criteria for Pb (U.S. EPA. 1985). and in previous Pb AQCDs
9 have shown effects of Pb on neurological endpoints in aquatic animal taxa. These include
10 changes in behaviors that may decrease the overall fitness of the organism such as
11 avoidance responses, decreased ability of an organism to capture prey or escape
12 predators, and alterations in feeding behaviors. Evidence of alteration in behaviors at the
13 level of the organism is a potential endpoint for effects at population or community levels
14 of biological organization (U.S. EPA. 2003a).
15 In the 1977 Pb AQCD behavioral impairment of a conditioned response (avoidance of a
16 mild electric shock) in goldfish was observed as low as 70 (ig Pb/L (Weir and Hine.
17 1970). In the 2006 Pb AQCD several studies were reviewed in which Pb was shown to
18 affect predator-prey interactions, including alteration in prey size choice and delayed prey
19 selection in juvenile fathead minnows following 2-week pre-exposure to 500 (ig Pb/L
20 (Weber. 1996). In limited studies available on worms, snails, tadpoles, hatchling turtles
21 and fish there is evidence that Pb may affect the ability to escape or avoid predation. For
22 example, in the tubificid worm T. tubifex the 96 hour EC50 for immobilization was
23 42 (ig Pb/L (Khangarot. 1991). Some organisms exhibit behavioral avoidance while
24 others do not seem to detect the presence of Pb (U.S. EPA. 2006c). Additional behavioral
25 endpoints reported in the Draft Ambient Aquatic Life Quality Criteria for Pb include an
26 EC50 of 140 (ig Pb/L for feeding inhibition in the freshwater cladoceran C. dubia and
27 deceased learning acquisition in bullfrogs at 500 (ig Pb/L (31.51 (ig Pb/L adjusted to a
28 total hardness of 50 mg/L CaCO3). All of these effects occur at concentrations that
29 exceed Pb concentration values found in surveys of U.S. surface waters although within
30 the range of Pb detected near some mining-disturbed areas (Table 3-11).
31 Recent information since the 2006 Pb AQCD provides evidence for Pb impacts on
32 behaviors that may affect feeding and predator avoidance in freshwater environments at
33 concentrations near the range of Pb detected in U.S. surface waters (Table 7-2 and
34 Section 7.4.5.3). Prey capture ability was decreased in 10 day old fathead minnow larvae
November 2012 7-169 Draft - Do Not Cite or Quote
-------
1 born from adult fish exposed to 120 (ig Pb/L for 300 days, then subsequently tested in a
2 21-day breeding assay (Mager et al.. 2010). Another study in fish reported effects at
3 low (ig Pb/L concentration, however, the findings are not considered as strong evidence
4 for causality since exposure concentrations in water were not analytically verified.
5 Specifically, zebrafish embryos exposed nominally to concentrations of Pb (2.0 to
6 6.0 (ig Pb/L) until 24 hours post-fertilization and then subsequently tested as larvae
7 exhibited decreased startle response time and altered pattern of escape swimming (Rice et
8 al.. 2011). In adult fish raised from the exposed embryos (6.0 (ig Pb/L), the ability to
9 detect visual contrast was degraded. Although this study was conducted with nominal
10 concentration of Pb in media, uptake of Pb by embryos was quantified and more Pb was
11 measured in tissues of embryos exposed to the higher concentration of Pb when
12 compared to the lower exposure concentration. Additional studies are needed in fish to
13 support these initial findings of effects on ecologically relevant behavioral impairments.
14 Findings in laboratory animals support the limited evidence for neurobehavioral effects
15 of Pb in freshwater invertebrates and vertebrates. In animal toxicological studies Pb
16 induced changes in learning and memory (Section 5.3.2.3). as well as attention and motor
17 skills (Section 5.3.3.1). New behaviors induced by exposure to Pb reviewed in Chapter_5
18 that are relevant to effects of Pb observed in freshwater systems include effects on visual
19 and auditory sensory systems and changes in structure and function of neurons and
20 supporting cells in the brain. Mechanisms that include the displacement of physiological
21 cations, oxidative stress and changes in neurotransmitters and receptors are also
22 reviewed. Central nervous system effects in fish recognized in previous Pb AQCDs
23 include effects on spinal neurons and brain receptors. New evidence from this review
24 identifies the MAPKs ERK1/2 and pSS^15 as possible molecular targets for Pb
25 neurotoxicity in catfish (Leal et al.. 2006). Evidence in terrestrial ecosystems
26 (Section 7.3.12.4) is not as extensive, but it is highly coherent with findings in aquatic
27 ecosystems. Overall, the evidence from available studies on neurobehavioral effects of Pb
28 in aquatic systems is limited, but sufficient to conclude that a causal relationship is likely
29 to exist between Pb exposures and neurobehavioral effects in aquatic invertebrates and
30 vertebrates.
7.4.12.5 Hematological Effects-Freshwater Biota
31 Hematological responses are commonly reported effects of Pb exposure in aquatic
32 invertebrates and vertebrates. Anemia was recognized as a symptom of chronic Pb
33 poisoning in fish in the 1977 Pb AQCD and has been subsequently reported in various
34 fish species using common hematological endpoints (e.g., red blood cell counts,
35 hematocrit, Hb concentrations) (Mager. 2012). In the 1986 Pb AQCD, hematological
November 2012 7-170 Draft - Do Not Cite or Quote
-------
1 effects of Pb exposure on fish included decrease in red blood cells and inhibition of
2 ALAD (U.S. EPA. 1986b). Inhibition of ALAD activity under various test conditions is
3 reported in Table 6 of the draft Ambient Aquatic Life Water Quality Criteria for Pb for
4 freshwater fish species (O. mykiss (Rainbow Trout), S. fontinalis (Brook Trout),
5 C. auratus (Goldfish) and Lepomis gibbosus (Pumpkinseed)) (U.S. EPA. 2008b). In these
6 studies, Rainbow Trout was the most sensitive with inhibition of ALAD activity reported
7 in multiple studies within the upper range of Pb in surface waters of the U.S. (median
8 0.5 ng Pb/L, range 0.04 to 30 jig Pb/L) (U.S. EPA. 2006c).
9 Laboratory studies with freshwater invertebrates have also indicated considerable species
10 differences in ALAD activity in response to Pb. For example, the concentration at which
11 50% ALAD inhibition was measured in the freshwater gastropod B. glabrata (23 to
12 29 (ig Pb/L) was much lower than in the freshwater oligochaete L. variegatus
13 (703 (ig Pb/L) based on nominal exposure data (Aisemberg et al.. 2005).
14 Findings in laboratory studies are additionally supported by evidence from field-collected
15 organisms providing coherence to the observations of Pb effects on ALAD activity. In
16 environmental assessments of metal-impacted habitats, ALAD is a recognized biomarker
17 of Pb exposure (U.S. EPA. 2006c). ALAD activity is negatively correlated with total Pb
18 concentration in freshwater bivalves, and lower ALAD activity has been correlated with
19 elevated blood Pb levels in field-collected fish as well. Further evidence from the
20 2006 Pb AQCD and this review of Pb effects on ALAD enzymatic activity, including
21 effects in bacteria, amphibians and additional field and laboratory studies on freshwater
22 fish, confirms that the decreased activity in this enzyme is an indicator for Pb exposure
23 across a wide range of taxa and that a common mode of action is likely for invertebrates
24 and vertebrates. The finding that the hematological system is a target for Pb in natural
25 systems is also supported by some evidence of Pb-induced alterations of serum profiles
26 and changes in white blood cell counts in fish (U.S. EPA. 2006c) and amphibians. This
27 evidence is strongly coherent with evidence from terrestrial vertebrates (Section
28 7.3.12.5). It is also coherent with observations from human epidemiologic and animal
29 toxicology studies (Section 5.7) where there is consistent evidence that exposure to Pb
30 induces adverse effects on hematological endpoints, including altered heme synthesis
31 mediated through decreased ALAD and ferrochelatase activities, decreased red blood cell
32 survival and function, and increased red blood cell oxidative stress. The overall weight of
33 epidemiologic and toxicological evidence for humans was sufficient to conclude that a
34 causal relationship exists between exposure to Pb and hematological effects (Section 5.7).
35 Based on observations in freshwater organisms and additionally supported by findings in
36 terrestrial systems, saltwater invertebrates (Section 7.4.21.5). and by toxicological and
37 epidemiologic evidence on human health effects, evidence is sufficient to conclude that
November 2012 7-171 Draft - Do Not Cite or Quote
-------
1 there is a causal relationship between Pb exposures and hematological effects in
2 freshwater vertebrates. Evidence is sufficient to conclude that a causal relationship is
3 likely to exist between Pb exposures and hematological effects in freshwater
4 invertebrates.
7.4.12.6 Physiological Stress-Freshwater Biota
5 Building on the body of evidence presented in the 2006 Pb AQCD (U.S. EPA. 2006c)
6 recent studies provide consistent and coherent evidence of upregulation of antioxidant
7 enzymes and increased lipid peroxidation associated with Pb exposure within one or two
8 orders of magnitude above current or ambient conditions in many species of freshwater
9 plants, invertebrates and vertebrates. A few studies provide evidence of effects at
10 concentrations of Pb encountered in some sediments of the U.S. (Table 7-2). In aquatic
11 plants, increases of antioxidant enzymes with Pb exposure occur in algae, mosses, and
12 floating and rooted aquatic macrophytes. Most available evidence for antioxidant
13 responses in aquatic plants are from laboratory studies lasting from 2 to 7 days and at
14 concentrations higher than typically found in the environment. However, data from
15 transplantation experiments from non-polluted to polluted sites indicate that elevated
16 enzyme activities are associated with Pb levels measured in sediments. For example, the
17 freshwater macrophyte Myriophyllum quitense exhibited elevated antioxidant enzyme
18 activity (glutathione-S-transferase, glutathione reductase, peroxidase) following
19 transplantation in anthropogenically polluted areas containing elevated Pb concentrations.
20 These were correlated with sediment Pb concentrations in the range of 5 to 23 mg Pb/g
21 dry weight (Nimptsch et al. 2005).There is evidence for antioxidant activity in response
22 to Pb exposure in freshwater invertebrates (i.e., bivalves). Markers of oxidative damage
23 are also observed in fish, amphibians and mammals in laboratory studies. Across all
24 organisms, there are differences in the induction of antioxidant enzymes that appear to be
25 species-dependent.
26 Additional stress responses to Pb in a few aquatic invertebrates have been reported since
27 the 2006 Pb AQCD, and included elevated heat shock proteins, osmotic stress, lowered
28 metabolism and decreased glycogen levels associated with Pb exposure. Although these
29 stress responses are correlated with Pb exposure, they are non-specific and may be altered
30 with exposure to any number of environmental stressors. Heat shock protein induction
31 has been observed in zebra mussels exposed to 500 (ig Pb/L for 10 weeks (Singer et al..
32 2005). Crayfish exposed for 14 days to 500 (ig Pb/L exhibited a Pb-induced
33 hypometabolism under conditions of environmental hypoxia in the presence of the metal
34 (Morris et al., 2005). Glycogen levels in the freshwater snail B. glabrata were
November 2012 7-172 Draft - Do Not Cite or Quote
-------
1 significantly decreased following 96-hour exposures at 50 (ig/L and higher (Ansaldo et
2 al.. 2006V
3 Evidence for stress responses observed in freshwater plants, invertebrates and vertebrates
4 is coherent with findings in terrestrial species (Section 7.3.12.6) and saltwater
5 invertebrates (Section 7.4.21.6). It is also coherent with evidence from human and
6 experimental animal studies of oxidative stress following impairment of normal metal ion
7 functions (Section 5.2.4). Upregulation of antioxidant enzymes and increased lipid
8 peroxidation are considered to be reliable biomarkers of stress, and provide evidence that
9 Pb exposure induces a stress response in those organisms which may increase
10 susceptibility to other stressors and reduce individual fitness. Evidence is sufficient to
11 conclude that a causal relationship is likely to exist between Pb exposures and
12 physiological stress in freshwater aquatic plants, invertebrates and vertebrates.
7.4.12.7 Community and Ecosystem Level Effects-Freshwater Biota
13 Most direct evidence of community- and ecosystem-level effects in freshwater systems is
14 from heavily contaminated sites where Pb concentrations are higher than typically
15 observed environmental concentrations for this metal. Impacts of Pb on aquatic habitats
16 that receive runoff from contaminated areas have been studied for several decades. For
17 aquatic systems, the literature focuses on evaluating ecological stress from Pb originating
18 from urban and mining effluents rather than atmospheric deposition. Ecosystem-level
19 field studies are complicated by the confounding of Pb exposure with other factors such
20 as the presence of trace metals and acidic deposition and by the variability inherent in
21 natural systems. In natural systems, Pb is often found co-existing with other stressors, and
22 observed effects may be due to cumulative toxicity.
23 In laboratory studies and simulated ecosystems, where it is possible to isolate the effect
24 of Pb, this metal has been shown to alter competitive behavior of species, predator-prey
25 interactions and contaminant avoidance. These dynamics may change species abundance
26 and community structure at higher levels of ecological organization. Uptake of Pb into
27 aquatic and terrestrial organisms and subsequent effects on mortality, growth,
28 developmental and reproductive endpoints at the organism level are expected to have
29 ecosystem-level consequences, and thus provide consistency and plausibility for causality
30 in ecosystem-level effects.
31 In aquatic ecosystems, field studies reviewed in the 2006 Pb AQCD (summarized in
32 Table AX7-2.5.2) and more recent studies report reductions of species abundance,
33 richness or diversity. This is particularly the case for benthic macroinvertebrate
34 communities where sources of Pb were mining or urban effluents, and Pb coexisted with
November 2012 7-173 Draft - Do Not Cite or Quote
-------
1 other metals. The results often indicate a correlation between the presence of one or more
2 metals and the negative effects observed. For example, in the 2006 Pb AQCD, the Coeur
3 d'Alene River watershed in Idaho, U.S. was used as a case study for Pb effects at the
4 population and community level. Significant negative correlations were observed
5 between Pb in water column and total taxa richness and EPT taxa richness in the river. In
6 a simulated aquatic microcosm a reduction in abundance and richness of protozoan
7 species was observed with increasing Pb concentration from 50 to 1,000 (ig Pb/L
8 (Fernandez-Leborans and Antonio-Garcia. 1988).
9 Since the 2006 Pb AQCD, there is further evidence for effects of Pb in sediment-
10 associated communities. Sediment-bound Pb contamination appears to differentially
11 affect members of the benthic invertebrate community, potentially altering ecosystems
12 dynamics in small urban streams (Kominkova and Nabelkova. 2005). Although surface
13 water Pb concentrations in monitored streams were determined to be very low,
14 concentrations of the metal in sediment were high enough to pose a risk to the benthic
15 community (e.g., 34 to 101 mg Pb/kg). These risks were observed to vary with benthic
16 invertebrate functional feeding group, with collector-gatherer species exhibiting larger
17 body burdens of heavy metals than benthic predators and collector-filterers.
18 In a recent study conducted since the 2006 Pb AQCD, changes to aquatic plant
19 community composition have been observed in the presence of elevated surface water Pb
20 concentrations at three lake sites impacted by mining effluents. The site with highest Pb
21 concentration (103-118 (ig Pb/L) had lowest number of aquatic plant species when
22 compared to sites with lower Pb concentrations (78-92 (ig Pb/L) (Mishraet al.. 2008).
23 Certain types of plants such as rooted and submerged aquatic plants may be more
24 susceptible to aerially deposited Pb resulting in shifts in Pb community composition.
25 High Pb sediment concentrations are linked to shifts in amphipod communities inhabiting
26 plant structures.
27 Avoidance response to Pb exposure varies widely in different species and this could
28 affect community composition and structure and species abundance. For example, frogs
29 and toads lack avoidance response while snails and fish avoid higher concentrations of Pb
30 (U.S. EPA. 2006c).
31 In the Annex to the 2006 Pb AQCD, the Coeur d'Alene River basin in Idaho was
32 presented as a case study for a watershed impacted by Pb and other metals. A significant
33 negative correlation was observed between Pb in water column (0.5 to 30 (ig Pb/L) and
34 total taxa richness, EPT taxa richness, and the number of metal-sensitive mayfly species
35 (Maret et al.. 2003). Additional lines of evidence including mine density, metal
36 concentrations, and bioaccumulation in caddisfly tissue were included. Since the
37 2006 Pb AQCD, additional research at this site and model development has resulted in
November 2012 7-174 Draft - Do Not Cite or Quote
-------
1 further characterization of the effects of Pb on waterfowl and other aquatic organisms in
2 this heavily contaminated ecosystem. Mean Pb concentrations in Coeur d'Alene sediment
3 range from 14 to 5,009 mg Pb/kg dry weight (Spears et al., 2007). Modeling of sediment
4 and Pb levels in waterfowl predict a sediment Pb effects range of 147-944 mg Pb/kg dry
5 weight and a mortality effects level of 1,652 mg/kg dry weight (Spears et al. 2007). In a
6 6-week feeding study with mallard (Anas platyrhynchos) ducklings, ingestion of
7 Pb-contaminated sediments from the Coeur d' Alene basin was shown to result in
8 decreased brain growth and altered brain chemistry (Douglas-Stroebel et al. 2004). These
9 findings support previous observations of altered behavior and hematological,
10 hepatotoxic, and histopathological endpoints in waterfowl from Lake Coeur d'Alene that
11 ingest Pb contaminated sediments and vegetation during feeding.
12 In addition to the evidence from microcosm and field studies presented above, effects on
13 reproduction (Section 7.4.12.1). growth (Section 7.4.12.2) and survival (Section 7.4.12.3)
14 have been clearly demonstrated in freshwater species. These endpoints can have effects at
15 the population-level and community-level of biological organization which may lead to
16 ecosystem-level impacts. Although the evidence is strong for effects of Pb on growth,
17 reproduction and survival in certain species in experimental settings at or near the range
18 of Pb concentrations reported in surveys of U.S. freshwater systems, considerable
19 uncertainties exist in generalizing effects observed under small-scale, particular
20 conditions up to predicted effects at the ecosystem level of biological organization. In
21 many cases it is difficult to characterize the nature and magnitude of effects and to
22 quantify relationships between ambient concentrations of Pb and ecosystem response due
23 to presence of multiple stressors, variability in field conditions and to differences in Pb
24 bioavailability at that level of organization. Bioavailability of Pb is influenced by pH,
25 alkalinity, total suspended solids, and DOC among other factors and can vary greatly in
26 natural environments. Nevertheless, evidence of ecosystem effects in aquatic systems is
27 coherent with similar evidence in terrestrial systems, and based on the cumulative
28 evidence from laboratory studies and field observations, a causal relationship is likely to
29 exist between Pb exposures and the alteration of species richness, species composition
30 and biodiversity in freshwater aquatic ecosystems.
7.4.13 Introduction to Bioavailability and Biological Effects of Pb in Saltwater
Ecosystems
31 Saltwater ecosystems include salt marsh, estuaries, embayments, beaches, and other
32 coastal areas; and encompass a range of salinities from just above that of freshwater to
33 that of seawater. These ecosystems may receive Pb contributions from direct atmospheric
34 deposition and/or via runoff from terrestrial systems. A range of 0.01 to 27 (ig Pb/L
November 2012 7-175 Draft - Do Not Cite or Quote
-------
1 including coastal areas, estuaries and open ocean was reported by Sadiq (1992) with the
2 higher values associated with sites involving human activity, however, these values are
3 not specific to the U.S. (Table 7-2). In an earlier publication, levels of Pb in the North
4 Atlantic and North Pacific surface waters ranged from 0.005 to 0.4 (ig Pb/L but the range
5 of values in coastal waters and estuaries were approximately equal to the range of Pb in
6 freshwater (Leland and Kuwabara. 1985). Additional information on Pb levels in water is
7 available in Sections 7.2.3 and 3.6. The 2006 Pb AQCD provided an overview of
8 regulatory considerations for water and sediments in addition to consideration of
9 biological effects and major environmental factors that modify the response of marine
10 organisms to Pb exposure. Regulatory guidelines for Pb in saltwater have not changed
11 since the 2006 Pb AQCD and are summarized below. This section is followed by new
12 information on bioavailability and biological effects of Pb in saltwater since the
13 2006 Pb AQCD.
14 The most recent ambient water quality criteria for Pb in saltwater were released in 1985
15 (U.S. EPA. 1985) by the EPA Office of Water which employed empirical regressions
16 between observed toxicity and water hardness to develop hardness-dependent equations
17 for acute and chronic criteria. These criteria are published pursuant to Section 304(a) of
18 the Clean Water Act and provide guidance to states and tribes to use in adopting water
19 quality standards for the protection of aquatic life and human health in surface water. The
20 ambient water quality criteria for Pb are currently expressed as a criteria maximum
21 concentration (CMC) for acute toxicity and criterion continuous concentration (CCC) for
22 chronic toxicity (U.S. EPA. 2009b). In saltwater, the CMC is 210 (ig Pb/L and the CCC
23 is 8.1 (ig Pb/L. The 2006 Pb AQCD summarized two approaches for establishing
24 sediment criteria for Pb based on either bulk sediment or equilibrium partitioning as
25 reviewed in the present document in Section 7.4.2.
26 In the following sections, recent information available since the 2006 Pb AQCD on Pb in
27 marine and estuarine ecosystems will be presented. Throughout the sections, brief
28 summaries of conclusions from the 1977 Pb AQCD (U.S. EPA. 1977). the 1986 Pb
29 AQCD (U.S. EPA. 1986b) and the 2006 Pb AQCD (U.S. EPA. 2006b) are included
30 where appropriate. Recent research on the bioavailability and uptake of Pb into saltwater
31 organisms including plants, invertebrates and vertebrates is presented in Section 7.4.14.
32 Toxicity of Pb to marine flora and fauna including growth, reproductive and
33 developmental effects (Section 7.4.15) are followed with data on exposure and response
34 of saltwater organisms (Section 7.4.16). Responses at the community and ecosystem
35 levels of biological organization are reviewed in Section 7.4.17 followed by
36 characterization of sensitivity and vulnerability of saltwater ecosystem components
37 (Section 7.4.18) and a discussion of ecosystem services (Section 7.4.19). The saltwater
38 sections conclude with a synthesis of the new data for Pb effects on saltwater plants,
November 2012 7-176 Draft - Do Not Cite or Quote
-------
1 invertebrates and vertebrates (Section 7.4.20) and causal determinations based on
2 evidence from previous Pb AQCDs and recent studies (Section 7.4.21).
7.4.14 Bioavailability of Pb in Saltwater Systems
3 Bioavailability was defined in the 2006 Pb AQCD as "the proportion of a toxin that
4 passes a physiological membrane (the plasma membrane in plants or the gut wall in
5 animals) and reaches a target receptor (cytosol or blood)". In 2007, EPA took cases of
6 bioactive adsorption into consideration and revised the definition of bioavailability as
7 "the extent to which bioaccessible metals absorb onto, or into, and across biological
8 membranes of organisms, expressed as a fraction of the total amount of metal the
9 organism is proximately exposed to (at the sorption surface) during a given time and
10 under defined conditions" (U.S. EPA. 2007c)
11 Factors affecting bioavailability of Pb to marine organisms are the same as those in
12 freshwater systems (Sections 7.4.2 and 7.4.4). However, although routes of exposure and
13 physiological mechanisms for storage and excretion influence uptake of metals by all
14 organisms, they may be different in marine organisms, particularly for ion transport
15 mechanisms (Niyogi and Wood. 2004). Marine environments are characterized by higher
16 levels of ions, such as Na+, Ca2+, and Mg2+, which compete for potential binding sites on
17 biotic ligands such as gills, thereby generally reducing the effective toxicity of metal ions
18 as compared to freshwater environments. However, because the concentrations of these
19 ions are relatively constant, bioavailability may be more predictable in marine systems
20 that are little influenced by freshwater than in freshwater systems, varying mostly with
21 amount and type of dissolved organic matter. In estuaries and embayments, changing
22 salinities and proximity to anthropogenic loading of pollutants add to the complexity of
23 predicting Pb speciation in these dynamic systems. BLMs (Figure 7-3) now being
24 developed for marine organisms are functionally similar to those applied to freshwater
25 organisms (Section 7.4.4).
26 Although in freshwater systems the presence of humic acid is considered to
27 reduce the bioavailable fraction of metals in freshwater, there is evidence that
28 DOC/DOM does not have the same effect on free Pb ion concentration in marine systems
29 (see Section 7.4.2.4 for detailed discussion). For the sea urchin P. lividus, the presence of
30 humic acid increased both the uptake and toxicity of Pb possibly by enhancing uptake of
31 Pb via membrane Ca2+ channels (Sanchez-Marin et al., 2010b). This also was observed in
32 the marine diatom Thalassiosira weissflogii, where humic acids absorbed to cell surfaces
33 increased metal uptake (Sanchez-Marin et al., 2010b). Formation of a ternary complex
34 that is better absorbed by biological membranes was another proposed mechanism that
November 2012 7-177 Draft - Do Not Cite or Quote
-------
1 could describe the increased bioavailability to marine invertebrates of Pb bound to humic
2 acid (Sanchez-Marin et al.. 2007).
3 Sanchez-Marin et al. (2011) subsequently have shown that different components of DOM
4 have different effects on Pb bioavailability in marine systems. Their initial research using
5 Aldrich humic acid found that increasing humic acid concentrations increased Pb uptake
6 by mussel gills and increased toxicity to sea urchin larvae in marine environments
7 (Sanchez-Marin et al.. 2007). In contrast, a subsequent investigation found that fulvic
8 acid reduced Pb bioavailability in marine water (Sanchez-Marin et al.. 2011). The
9 contradictory effects of different components of DOM on marine bioavailability likely
10 reflect their distinct physico-chemical characteristics. More hydrophobic than fulvic acid,
11 humic acid may adsorb directly with cell membranes and enhance Pb uptake through
12 some (still unidentified) mechanism (Sanchez-Marin et al.. 2011). Pb AVS-measurements
13 were also determined to accurately predict uptake by mussels (Mytilus sp.) in the
14 presence of 2.5 to 20 mg/L fulvic acid (Sanchez-Marin et al.. 2011). However, the effects
15 of DOM on Pb bioavailability to mussels were underpredicted by AVS Pb concentration
16 measurements, potentially as a result of adsorption of DOM-Pb complexes.
17 Based on the above, BLMs (see Section 7.4.4 and Figure 7-3) used to predict
18 bioavailability of Pb to aquatic organisms (Pi Toro et al.. 2005). may require
19 modifications for application to marine organisms. Of particular importance is the finding
20 that in marine aquatic systems, surface water DOM was found to increase (rather than
21 decrease) uptake of Pb by fish gill structures, potentially through the alteration of
22 membrane Ca2+ channel permeability. Veltman et al. (2010) proposed integrating BLM
23 and bioaccumulation models in order to more accurately predict metal uptake by fish and
24 invertebrates, and calculated metal absorption efficiencies for marine fish species from
25 both types of models. They noted that affinity constants for Ca2+, Cd, Cu, Na, and Zn
26 were highly similar across different aquatic species, including fish and invertebrates
27 (Veltman et al.. 2010). These findings suggest that the BLM can be integrated with
28 bioaccumulation kinetics to account for both environmental chemical speciation and
29 biological and physiological factors in both marine and freshwater systems.
7.4.14.1 Saltwater Plants and Algae
30 In the 1977 Pb AQCD, the cordgrass Spartina alterniflora was found to reduce by a small
31 amount the quantity of Pb in sediments (U.S. EPA. 1977). Limited data on marine algal
32 species reviewed in the 1986 Pb AQCD and 2006 Pb AQCD provided additional
33 evidence for Pb uptake. Recent data available since the 2006 Pb AQCD includes Pb
34 bioaccumulation studies conducted with five species of marine algae, (Tetraselmis chuii,
November 2012 7-178 Draft - Do Not Cite or Quote
-------
1 Rhodomonas satinet, Chaetoceros sp., Isochrysis galbana and Nannochloropsis
1 gaditana). In this study it was demonstrated that bioaccumulation rates varied with
3 species following 72-hour exposure to Pb. /. galbana accumulated the lowest
4 concentrations of Pb (0.01 and 0.6 pg Pb/cell at water concentrations of 51 and
5 6,348 (ig Pb/L), while Chaetoceros sp. was observed to be the most efficient Pb
6 bioaccumulator, adsorbing 0.04 and 54 pg Pb/cell at 51 and 6,348 (ig Pb/L (Debelius et
7 al.. 2009).
8 Recent uptake studies of Pb in plants associated with marine environments are also
9 available. The roots of two salt marsh species, Sarcocornia fruticosa and Spartina
10 maritima, significantly accumulated Pb, to maximum concentrations of 2,870 mg Pb/kg
11 and 1,755 mg Pb/kg, respectively (Caetano et al., 2007). Roots had similar isotopic
12 signature to those of sediments in vegetated zones indicating that Pb uptake by plants
13 reflects the input in sediments. BCFs for Pb in root tissue from mangrove tree species
14 range between 0.09 and 2.9, depending on the species and the habitat, with an average
15 BCF of 0.84. The average BCF for mangrove species leaf tissue was considerably less
16 (0.11), as these species are poor translocators of Pb (MacFarlane et al.. 2007).
7.4.14.2 Saltwater Invertebrates
17 Uptake and subsequent bioaccumulation of Pb in marine invertebrates varies greatly
18 between species and across taxa as previously characterized in the 2006 Pb AQCD. This
19 section expands on the findings from the 2006 Pb AQCD on bioaccumulation and
20 sequestration of Pb in saltwater invertebrates. In the case of invertebrates, Pb can be
21 bioaccumulated from multiple sources, including the water column, sediment, and dietary
22 exposures, and factors such as proportion of bioavailable Pb, lifestage, age, and
23 metabolism can alter the accumulation rate. In this section, new information on Pb uptake
24 and subsequent tissue and subcellular distribution will be considered, followed by a
25 discussion on dietary and water routes of exposure and strategies for detoxification of Pb
26 in marine invertebrates.
27 In marine invertebrates, sites for Pb accumulation include the gill and digestive
28 gland/hepatopancreas. The gills were the main sites of Pb accumulation in pearl oyster,
29 Pinctada fucata followed by mantle, in 72-hour exposures to 103.5 (ig Pb/L (Jing et al..
30 2007). Following a 10-day exposure to 2,500 (ig Pb/L as Pb nitrate, accumulation of Pb
31 was higher in gill than digestive gland ofMytilus edulis: after a 10 day depuration, Pb
32 content was decreased in the gills and digestive gland of these mussels (Einsporn et al..
33 2009). In blue crabs, Callinectes sapidus, collected from a contaminated and a clean
November 2012 7-179 Draft - Do Not Cite or Quote
-------
1 estuary in New Jersey, U.S., the hepatopancreas was found to be the primary organ for Pb
2 uptake (Reichmuth et al.. 2010).
3 There is more information now on the cellular and subcellular distribution of Pb in
4 invertebrates than there was at the time of writing the 2006 Pb AQCD. Specifically,
5 localization of Pb at the ultrastructural level has been assessed in the marine mussel (M
6 edulis) through an antibody-based detection method (Einsporn et al.. 2009; Einsporn and
7 Koehler. 2008). Dissolved Pb was detected mainly within specific lysosomal structures in
8 gill epithelial cells and digestive gland cells and was also localized in nuclei and
9 mitochondria. Transport of Pb is thought to be via lysosomal granules associated with
10 hemocytes (Einsporn et al.. 2009). In the digestive gland of the variegated scallop
11 (Chlamys varia), Pb was also mainly bound to organelles, (66% of the total metal burden)
12 (Bustamante and Miramand. 2005). In the digestive gland of the cephalopod Sephia
13 officinalis, (cuttlefish) most of the Pb was found in the organelles (62%) (Bustamante et
14 al.. 2006). In contrast, only 7% of Pb in the digestive gland of the octopus (Octopus
15 vulgaris) was associated with the fraction containing nuclei, mitochondria, lysosome and
16 microsomes: the majority of Pb in this species was found in cytosolic proteins (Raimundo
17 et al.. 2008).
18 Metian et al. (2009) investigated the uptake and bioaccumulation of 210Pb in variegated
19 scallop and king scallop to determine the major accumulation route (seawater or food)
20 and then assess subsequent tissue distribution. Dietary Pb from phytoplankton in the diet
21 was poorly assimilated (<20%) while more than 70% of Pb in seawater was retained in
22 the tissues. In seawater, 210Pb was accumulated more rapidly in variegated scallop than
23 king scallop and soft tissue distribution patterns differed between the species. Variegated
24 scallop accumulated Pb preferentially in the digestive gland (50%) while in king scallop,
25 Pb was equally distributed in the digestive gland, kidneys, gills, gonad, mantle, intestine,
26 and adductor muscle with each tissue representing 12-30% of 210Pb body load. An
27 additional test with Pb-spiked sediment in king scallop showed low bioaccumulation
28 efficiency of Pb from spiked sediment.
29 Recently, several studies have attempted to establish biodynamic exposure assessments
30 for various contaminants. In an in situ metal kinetics field study with the mussel
31 M. galloprovincialis, simultaneous measurements of metal concentrations in water and
32 suspended particles with mussel biometrics and physiological indices were conducted to
33 establish uptake and excretion rates in the natural environment (Casas et al., 2008). The
34 mean logarithmic ratio of metal concentration in mussels (ng/kg of wet-flesh weight) to
35 metal concentration in water (ng/L) was found to be 4.3 inM galloprovincialis, based on
36 the rate constants of uptake and efflux in a series of transplantation experiments between
November 2012 7-180 Draft - Do Not Cite or Quote
-------
1 contaminated and clean environments. Equilibrium concentrations of Pb in mussels
2 leveled out at approximately 30 days with a concentration of 6.7 mg Pb/kg.
3 The protective barrier against Pb toxicity formed by the egg structure in some
4 invertebrates was recognized in the 2006 Pb AQCD. Consideration of toxicity of Pb to
5 embryos that develop surrounded by a protective egg shell has been expanded since the
6 2006 Pb AQCD. In a study with cuttlefish (S. officinalis) eggs, radioisotopes were used to
7 assess the permeability of the egg to Pb at low exposure concentrations (210Pb activity
8 concentration corresponding to 512 (ig/L Pb) (Lacoue-Labarthe et al., 2009). Retention
9 and diffusion properties of the cuttlefish egg change throughout the development of the
10 embryo and since the eggs are fixed on substrata in shallow coastal waters they may be
11 subject to both acute and chronic Pb exposures. In the radiotracer experiments, 210Pb was
12 never detected in the internal compartments of the egg during the embryonic
13 development stage, while concentrations in the eggshell increased throughout the 48-day
14 exposure. These results are consistent with a study of cuttlefish eggs collected from
15 natural environments in which Pb was only detected in the eggshell. These studies
16 indicate that the cuttlefish egg provides a protective barrier from Pb toxicity (Miramand
17 et al.. 2006).
18 Aquatic invertebrate strategies for detoxifying Pb were reviewed in the 2006 Pb AQCD
19 and include sequestration of Pb in lysosomal-vacuolar systems, excretion of Pb by some
20 organisms, and deposition of Pb to molted exoskeleton. Molting of the exoskeleton can
21 result in depuration of Pb from the body (see Knowlton et al. (1983) and Anderson et al.
22 (1997). as cited in the 2006 Pb AQCD). New research has provided further evidence of
23 depuration of Pb via molting in invertebrates. Mohapatra et al. (2009) observed that Pb
24 concentrations in body tissues were lower in the newly molted mud crabs (Scylla serrata)
25 than in the pre-molt, hard-shelled crabs. However, the carapace of hard shelled crabs had
26 lower concentrations of Pb than the exuvium of the soft shell crabs, leading the authors to
27 speculate that some of the metal might be partially excreted during the molting process,
28 rather than entirely through shedding of the previous exoskeleton. Bergey and Weis
29 (2007) showed that differences in the proportion of Pb stored in exoskeleton and soft
30 tissues changed during intermolt and immediate postmolt in two populations of fiddler
31 crabs (Ucapugnax) collected from New Jersey. One population from a relatively clean
32 estuary eliminated an average of 56% of Pb total body burden during molting while
33 individuals from a site contaminated by metals eliminated an average of 76% of total Pb
34 body burden via this route. Pb distribution within the body of crabs from the clean site
35 shifted from exoskeleton to soft tissues prior to molting. The authors observed the
36 opposite pattern of Pb distribution in fiddlers from the contaminated site where larger
37 amounts of Pb were depurated in the exoskeleton. The exact dynamics of Pb depuration
38 through molting in crabs are thus still not completely characterized.
November 2012 7-181 Draft - Do Not Cite or Quote
-------
7.4.14.3 Saltwater Vertebrates
Saltwater Fish
1 In comparison to freshwater fish, fewer studies have been conducted on Pb uptake in
2 marine fish. Since marine fish drink seawater to maintain osmotic homeostasis, Pb can be
3 taken up via both gills and intestine (Wang and Rainbow. 2008). Pb was significantly
4 accumulated in gill, liver, plasma, kidney, rectal gland, intestine, skin, and muscle of the
5 elasmobranch spotted dogfish (Scyliorhinus canicula) exposed to 2,072 (ig Pb/L for one
6 week (De Boeck et al., 2010). In contrast to Pb distribution patterns in freshwater
7 teleosts, high Pb concentrations were present in this species in the skin and rectal gland.
8 Egg cases of the spotted dogfish exposed to 210Pb in seawater for 21 days, accumulated
9 radiolabeled Pb rapidly and the metal was subsequently detected in embryos indicating
10 the permeability of shark eggs to Pb in coastal environments (Jeffree et al., 2008). A
11 study of Pb bioaccumulation in five marine fish species (Chloroscombrus chrysurus,
12 Sardinella aurita, Ilisha africana, Galeoides decadactylus, Caranx latus) found that
13 C. chrysurus was an especially strong bioaccumulator, yielding Pb concentrations of 6 to
14 10 mg Pb/kg (Gnandi et al., 2006). However, C. chrysurus metal content was not
15 correlated to the Pb concentrations along the mine tailings gradient from which they were
16 collected (8.5 and 9.0 (ig Pb/L for minimum and maximum tissue concentrations,
17 respectively). This lack of correlation was also observed for fish species that were
18 considered to be weaker Pb bioaccumulators, indicating that unidentified sources of Pb
19 (e.g., in sediments or in dietary sources) may be contributing to Pb uptake by marine fish.
20 In grunt fish H. scudderi, exposed to Pb via dietary uptake through a simulated marine
21 food chain, mean total Pb body burden increased from 0.55 to 3.32 mg Pb/kg in a 42-day
22 feeding study (Soto-Jimenez et al.. 201 Ib). Pb was accumulated to the highest relative
23 concentration in liver with less than 3% of total Pb accumulated in gills. Most of the Pb
24 based on total body mass was accumulated in skeleton, skin, scales and muscle.
25 The 2006 Pb AQCD considered detoxification mechanisms in fish including mucus
26 production and Pb removal by shedding of scales in which Pb is chelated with keratin.
27 Since the 2006 Pb AQCD review, additional Pb detoxification mechanisms in marine fish
28 have been further elucidated. Mummichog (Fundulus heteroclitus) populations in metal-
29 polluted salt marshes in New York exhibited different patterns of intracellular
30 partitioning of Pb although body burden between sites was not significantly different
31 (Goto and Wallace. 2010). Mummichogs at more polluted sites stored a higher amount of
32 Pb in metal rich granules as compared to other detoxifying cellular components such as
33 heat-stable proteins, heat-denaturable proteins and organelles.
November 2012 7-182 Draft - Do Not Cite or Quote
-------
Marine Mammals
1 Studies that consider uptake of Pb in aquatic mammals are limited. Kannan et al. (2006)
2 compared trace element concentrations in livers of free-ranging sea otters (Enhydra lutris
3 nereis) found dead along the California coast. They detected Pb in all individuals
4 sampled (N=80) in a range of 0.019 to 1.06 mg Pb/kg. The otters were classified by cause
5 of death (infectious causes, non-infectious causes, emaciated condition) and trace element
6 patterns of tissue distribution were compared. Livers from emaciated otters had
7 significantly elevated levels of Pb compared to non-diseased individuals.
7.4.14.4 Marine Food Web
8 As discussed in Section 7.4.4.4 trophic transfer of Pb through aquatic food chains was
9 considered to be negligible in the 2006 Pb AQCD (U.S. EPA. 2006c). Measured
10 concentrations of Pb in the tissues of aquatic organisms were found to be generally higher
11 in algae and benthic organisms and lower in higher trophic-level consumers, indicating
12 that Pb was bioaccumulated but not biomagnified (U.S. EPA. 2006c; Eisler. 2000).
13 Recent literature since the 2006 Pb AQCD, provides evidence of the potential for Pb to
14 be transferred in marine food webs while other studies indicate Pb is decreased with
15 increasing trophic level. This section incorporates recent literature on transfer of Pb
16 through marine food chains.
17 In a dietary study using environmentally realistic concentrations of Pb in prey through
18 four levels of a simplified marine food chain, biological responses including decreased
19 growth and survival and changes in behavior were observed at different trophic levels.
20 However, the concentration of Pb did not increase along the trophic gradient (Soto-
21 Jimenez et al.. 201 Ib; Soto-Jimenez et al.. 201 la). The base of the simulated food chain
22 was the microalgae Tetraselmis suecica (phytoplankton) grown in 20 (ig Pb/L.
23 Pb-exposed cultures of T. suecica had significantly less cell divisions per day (growth),
24 biomass and total cell concentrations than control microalgae at 72 hours of exposure.
25 The microalgal cultures were then fed to Artemia franciscana (crustacean, brine shrimp)
26 which were then fed to Litopenaeus vannamei (crustacean, whiteleg shrimp) and finally
27 to Haemulon scudderi (fish, grunt). Effects on behavior, growth and survival were
28 observed in shrimp and in grunt fish occupying the intermediate and top levels of the
29 simulated marine food chain. The authors speculate that the species used in the simulated
30 food chain were able to regulate and eliminate Pb (Soto-Jimenez et al.. 201 Ib).
31 Partial evidence for biomagnification was observed in a subtropical lagoon in Mexico
32 with increases of Pb concentration occurring in 14 of the 31 (45.2%) of trophic
33 interactions considered (Ruelas-Inzunza and Paez-Osuna. 2008). The highest rate of
November 2012 7-183 Draft - Do Not Cite or Quote
-------
1 transference of Pb as measured in muscle tissue occurred between the prey species
2 whiteleg shrimp (Litopenaeus vannamei) and mullet (Mugil cephalus) to pelican
3 (Pelecanus occidentalis).
4 Other studies have traced Pb in marine food webs and have found no evidence of
5 biomagnification of Pb with increasing trophic level. In the southeastern Gulf of
6 California, Mexico, Pb was not positively transferred (biomagnification factor <1)
7 through primary producers (seston, detritus) and 14 consumer species in a lagoon food
8 web (Jara-Marini et al., 2009). In a planktonic food web in Bahia Blanca estuary,
9 Argentina, Pb levels in macrozooplankton and mesozooplankton exhibited temporal
10 fluctuations, however no biomagnification was observed between mesozooplankton and
11 macrozooplankton (Fernandez Severini et al., 2011). It is important to note, however, that
12 even in the absence of biomagnification, aquatic organisms can bioaccumulate relatively
13 large amounts of metals and become a significant source of dietary metal to their
14 predators (Fairbrother et al.. 2007: Reinfelder et al.. 1998).
7.4.15 Biological Effects of Pb in Saltwater Systems
15 This section focuses on the studies of biological effects of Pb on marine and estuarine
16 algae, plants, invertebrates, fish and mammals published since the 2006 Pb AQCD. Key
17 studies from the 1977 Pb AQCD, the 1986 Pb AQCD and the 2006 Pb AQCD on
18 biological effects of Pb are summarized where appropriate. Biological effects of Pb on
19 saltwater algae and plant species are considered below, followed by information on
20 effects on marine invertebrates and vertebrates. Alterations to reproduction, growth and
21 survival of saltwater organisms can lead to changes at the community and ecosystem
22 levels of biological organization such as decreased abundance, reduced taxa richness, and
23 shifts in species composition (Section 7.1). New evidence for Pb effects on reproduction,
24 growth and survival in saltwater plants, invertebrates and vertebrates is summarized in
25 Table 7-6. In general, Pb toxicity to saltwater organisms is less well characterized than
26 toxicity of Pb in freshwater ecosystems due to the fewer number of available studies on
27 marine species. Because this review is focused on effects of Pb, studies reviewed for this
28 section include only those for which Pb was the only, or primary, metal to which the
29 organism was exposed. All reported values are from exposures in which concentrations
30 of Pb were analytically verified unless nominal concentrations are stated.
November 2012 7-184 Draft - Do Not Cite or Quote
-------
7.4.15.1 Saltwater Algae and Plants
1 New evidence on toxicity of Pb to marine algae indicates that species exhibit varying
2 sensitivities to Pb in saltwater. The lowest 72-hour EC50 for growth inhibition reported for
3 marine algae was 105 (ig Pb/L in Chaetoceros sp (Debelius et al.. 2009). The microalgae
4 T. suecica, had statistically significant decreased biomass yield, growth rate and cell
5 count following 72 hours nominal exposure to 20 (ig Pb/L (Soto-Jimenez et al.. 201 Ib).
6 Pb tested at nominal concentrations up to -2,000 (ig Pb/L over a 14-day period did not
7 affect photosynthetic activity in seven species of marine macroalgae (Ascophyllum
8 nodosum, Fucus vesiculosus, Ulva intestinalis, Cladophora rupestris, Chondrus crispus,
9 Palmaria palmate, Polysiphonia lanosa) as measured by pulse amplitude modulation
10 chlorophyll fluorescence yield although Pb was readily accumulated by these species
11 (Baumann et al.. 2009). In a recent review of the production of phytochelatins and
12 glutathione by marine phytoplankton in response to metal stress, Kawakami et al. (2006)
13 included several studies in which Pb exposure was shown to induce glutathione and
14 phytochelatin at high concentrations in a few species.
7.4.15.2 Saltwater Invertebrates
15 No studies with marine invertebrates were reviewed in the 1977 Pb AQCD or the 1986
16 Pb AQCD. Effects of Pb on marine invertebrates reported in the 2006 Pb AQCD included
17 impacts on embryo development in bivalves with an EC50 of 221 (ig Pb/L for
18 embryogenesis, gender differences in sensitivity to Pb in copepods and increasing
19 toxicity with decreasing salinity in mysids. Survival, growth and reproduction are
20 affected by Pb in marine organisms. Pb has also been shown to affect stress responses,
21 antioxidant activity and osmoregulation.
22 Recent literature strengthens the evidence indicating that Pb affects enzymes and
23 antioxidant activity in marine invertebrates. Most of these studies only report nominal
24 concentrations of Pb. Activity of enzymes associated with the immune defense system in
25 the mantle of pearl oyster were measured at 0, 24, 48 and 72 hour nominal exposure to
26 100 (ig Pb/L (Jing et al.. 2007). Activity of AcPase, a lysosomal marker enzyme, was
27 detected at 24 hours and subsequently decreased. Phenoloxidase activity was depressed
28 compared with controls and remained significantly lower than control after 72 hours of
29 exposure to Pb. Increased SOD activity was observed in the mantle but decreased with
30 time, although always remaining higher than in the control animals (Jing et al.. 2007).
31 Activity of Se-dependent glutathione peroxidase did not change with Pb exposure. SOD,
32 catalase, and glutathione peroxidase were significantly reduced at environmentally
33 relevant concentrations of Pb (2 (ig Pb/L as measured in Bohai Bay, China) in the
November 2012 7-185 Draft - Do Not Cite or Quote
-------
1 digestive gland of the bivalve Chlamys farreri fZhang et al., 20101x). In contrast,
2 Einsporn et al. (2009) observed no change in catalase activity in the digestive gland and
3 gill of blue mussel M edulis following nominal exposure to 2,500 (ig Pb/L as Pb nitrate
4 for 10 days and again following a 10 day depuration period. However, in this same
5 species, glutathione-S-transferase activity was elevated in the gills after Pb exposure and
6 remained active during depuration while no changes to glutathione-S-transferase activity
7 were observed in the digestive gland. In black mussel (M galloprovincialis) exposed 10
8 days to sublethal nominal concentrations of Pb, fluctuations in SOD activity were
9 observed over the length of the exposure and MDA levels were increased in mantle and
10 gill (Vlahogianni and Valavanidis. 2007). Catalase activity was decreased in the mantle
11 of these mussels but fluctuated in their gills, as compared with the control group. In the
12 bivalve C. farreri exposed to Pb, there was induction of lipid peroxidation measured as
13 MDA of 24% and a 37% reduction in 7-ethoxyresorufm-o-deethylase (EROD) activity
14 when compared to controls (Zhang et al., 2010b). In red fingered marsh crab,
15 Parasesarma erythrodactyla, collected from sites along an estuarine lake in New South
16 Wales, Australia, elevated glutathione peroxidase activity was correlated with individuals
17 with higher metal body burdens (MacFarlane et al.. 2006).
18 ALAD is a recognized biomarker of exposure across a wide range of taxa including
19 bacteria (Korean et al., 2007). invertebrates and vertebrates. Since the 2006 Pb AQCD,
20 there are additional studies measuring changes in ALAD activity in field-collected
21 bivalves and crustaceans from saltwater habitats. In the bivalve Chamelea gallina
22 collected from the coast of Spain, ALAD inhibition was greater with higher
23 concentrations of Pb measured in whole tissue (Kalman et al., 2008). In another study
24 conducted in Spain, ALAD activity was negatively correlated with total Pb concentration
25 in seven marine bivalves (C. gallina, Mactra corallina, Donax trunculus, Cerastoderma
26 edule, M. galloprovincialis, Scrobicularia plana and Crassostrea angulata). However,
27 the authors of this study indicated the need to consider variability of responses between
28 species when using ALAD as a biomarker for Pb (Company et al.. 2011). Pb content
29 varied significantly among species and was related to habitat (sediment versus substrate)
30 and feeding behavior.
31 Behavioral responses of aquatic invertebrates to Pb reviewed in the 2006 Pb AQCD
32 included avoidance. A limited number of recent studies have considered additional
33 behavioral endpoints in marine organisms. Valve closing speed was used as a measure of
34 physiological alterations due to Pb exposure in the Catarina scallop (Sobrino-Figueroa
35 and Caceres-Martinez. 2009). The average valve closing time increased from under one
36 second in the control group to 3 to 12 seconds in juvenile scallops exposed to analytically
37 verified concentrations of Pb as Pb nitrate (40 (ig/L to 400 (ig/L) for 20 days. Damage to
November 2012 7-186 Draft - Do Not Cite or Quote
-------
1 sensory cilia of the mantle was observed following microscopic examination of
2 Pb-exposed individuals.
3 Since the 2006 Pb AQCD, limited studies on marine invertebrates have indicated effects
4 of Pb on reproduction. In a long term (approximately 60 days) sediment
5 multigenerational bioassay with the estuarine-sediment dwelling amphipod Elasmopus
6 laevis, onset to reproduction was significantly delayed at 118 mg Pb/kg compared to
7 controls. In the higher concentrations, start of offspring production was delayed further; 4
8 days in 234 mg Pb/kg and 8 days in 424 mg Pb/kg (Ringenary et al., 2007). Fecundity
9 and time of first offspring production was also reduced with increasing Pb concentration
10 in sediment above 118 mg Pb/kg. The authors indicate that this concentration is below
11 the current marine sediment regulatory guideline for Pb (218 mg Pb/kg sediment)
12 (NOAA. 1999) and that reproductive endpoints are more sensitive than survival in this
13 species. Exposure of gametes to Pb prior to fertilization resulted in a decrease of the
14 fertilization rates of the marine polychaete Hydroides elegans (Gopalakrishnan et al..
15 2008). In sperm pretreated in 97 (ig Pb/L filtered seawater for 20 minutes, fertilization
16 rate decreased by approximately 70% compared to controls. In a separate experiment,
17 eggs were pretreated with Pb prior to addition of an untreated sperm suspension. The
18 fertilization rate of eggs pretreated in 48 (ig Pb/L filtered seawater decreased to 20% of
19 the control. In another test with H. elegans in which gametes were not pre-treated, but
20 instead added directly to varying concentrations of Pb for fertilization, there appears to be
21 a protective effect following fertilization due to the formation of the fertilization
22 membrane during the first cell division that may prevent Pb from entering the oocytes
23 (Gopalakrishnan et al., 2007).
24 As noted in the 2006 Pb AQCD and supported by recent studies, Pb exposure negatively
25 affects the growth of marine invertebrates. Wang et al., (2009d_) observed growth of
26 embryos of the Asian Clam (Meretrix meretrix) was significantly reduced by Pb with an
27 EC50 of 197 (ig/L. In juvenile Catarina scallop, Argopecten ventricosus, exposed to Pb for
28 30 days, the EC50 for growth was 4,210 (ig Pb/L (Sobrino-Figueroa et al., 2007). Rate of
29 growth of the deposit feeding Capitella sp. polychaetes decreased significantly from the
30 controls in 3 and 6-day exposures, however, the observed changes did not exhibit a clear
31 dose response with increasing Pb concentration (Horng et al.. 2009).
32 Although Pb is known to cause mortality when invertebrates are exposed to sufficiently
33 high concentrations, some species may not exhibit significant mortality even at high
34 concentrations. In a 10-day Pb-spiked sediment exposure (1,000 mg Pb/kg and
35 15 (ig Pb/L dissolved Pb in pore water), 100% of individuals of the Australian estuarine
36 bivalve Tellina deltoidalis survived (King et al.. 2010). In the deposit feeding Capitella
37 sp., polychaetes, exposure to varying concentrations of Pb associated with spiked
November 2012 7-187 Draft - Do Not Cite or Quote
-------
1 sediment up to 870 mg Pb/kg had no effect on survival (Horng et al., 2009). No
2 differences in adult survival were observed in 28 and 60 day sediment exposures to a
3 range of Pb concentrations from 58 mg Pb/kg to 424 mg Pb/kg in the amphipod E. laevis
4 (Ringenary et al. 2007). Other species are more sensitive to Pb and these responses are
5 reviewed in Section 7.4.16.
7.4.15.3 Saltwater Vertebrates
Saltwater Fish
6 There is a dearth of information in previous Pb AQCDs on Pb effects in saltwater fish.
7 Recent data available since the 2006 Pb AQCD include a study with a marine
8 elasmobranch. De Boeck et al. (2010) exposed the spotted dogfish (S. canicula) to
9 2,072 (ig Pb/L for one week and measured metallothionein induction in gill and liver
10 tissue, and the electrolytes Na, K, Ca2+ and Cl, in plasma. No effects were observed in
11 Pb-exposed fish for any of the physiological variables measured in this study, although
12 Pb was detected in all organs (De Boeck et al.. 2010).
13 Since the 2006 Pb AQCD, several studies integrating behavioral and physiological
14 measures of Pb toxicity have been conducted on marine fish. The ornate wrasse
15 (Thalassoma pavo) was exposed nominally to sublethal (400 (ig Pb/L) or a maximum
16 acceptable toxicant concentration (1,600 (ig Pb/L) dissolved in seawater for one week to
17 assess the effects of Pb on feeding and motor activities (Giusi et al.. 2008). In the
18 sublethal concentration group, hyperactivity was elevated 36% over controls. In the high
19 concentration, a 70% increase in hyperactivity was observed and hyperventilation
20 occurred in 56% of behavioral observations. Elevated expression of heat shock protein
21 70/90 orthologs was detected in the hypothalamus and mesencephalic areas of the brains
22 of Pb-treated fish and neuronal damage was observed in the posterior hypothalamic area
23 and optic tectum. No changes in feeding activity were noted between non-treated and
24 treated fish.
25 Additional behavioral studies in fish consider effects of dietary Pb. The grunt fish
26 H. scudderi, occupying the top level of a simulated marine food chain, exhibited lethargy
27 and decreased food intake during the last week of a 42-day feeding study (Soto-Jimenez
28 et al., 20 lib). The fish were fed white shrimp exposed to Pb via brine shrimp that were in
29 turn fed microalgae cultured at a nominal concentration of 20 (ig Pb/L. Pb was quantified
30 in shrimp and fish. The authors noted a few of the fish exposed to Pb via dietary transfer
31 through the food chain were observed surfacing and speculated that this behavior was air
32 breathing as a response to stress.
November 2012 7-188 Draft - Do Not Cite or Quote
-------
1 Evidence for reproductive effects of Pb in saltwater fish is limited to a field study in
2 which decreased oocyte diameter and density in the toadfish (Tetractenos glaber) were
3 associated with elevated levels of Pb in the gonad offish collected from contaminated
4 estuaries in Sydney, Australia (Alquezar et al.. 2006). The authors state this is suggestive
5 of a reduction in egg size which ultimately may lead to a decline in female reproductive
6 output.
Mammals
7 Although Pb continues to be detected in tissues of marine mammals in U.S. coastal
8 waters (Bryan et al., 2007; Stavros et al., 2007; Kannan et al.. 2006) few studies exist that
9 consider biological effects associated with Pb exposure. Pb effects on immune variables,
10 including cell viability, apoptosis, lymphocyte proliferation, and phagocytosis were tested
11 in vitro on phagocytes and lymphocytes isolated from the peripheral blood of bottlenose
12 dolphin (Tursiops truncates} (Camara Pellisso et al., 2008). No effects on viability of
13 immune cells, apoptosis, or phagocytosis were observed in 72-hour exposure to nominal
14 concentrations of 1,000, 10,000, 20,000 and 50,000 (ig Pb/L. Proliferative response of
15 bottlenose dolphin leukocytes was significantly reduced at 50,000 (ig Pb/L, albeit by only
16 10% in comparison to the control. This in vitro exposure with nominal concentrations of
17 Pb is likely not relevant for assessing effects of atmospherically-deposited Pb on marine
18 mammals, however, no additional studies were available for review on the effects on Pb
19 on these organisms.
7.4.16 Exposure and Response of Saltwater Species
20 Evidence regarding exposure-response relationships and potential thresholds for Pb
21 effects on saltwater populations can inform determination of standard levels that are
22 protective of marine ecosystems. The Annex of the 2006 Pb AQCD (U.S. EPA. 2006c)
23 summarized data on exposure-response functions for invertebrates (Table AX7-2.4.1)
24 (Table AX7-2.4.2). The recent exposure-response studies reviewed in this section expand
25 on earlier findings with information on microalgal and invertebrate species. Studies
26 specific to growth, reproduction and survival endpoints are summarized in Table 7-6. All
27 reported values are from exposures in which concentrations of Pb were analytically
28 verified unless nominal concentrations are stated.
29 A series of 72-hour Pb toxicity tests were conducted with five marine microalgae species
30 (T. chuii, R. salina, Chaetoceros sp., /. galbana and N. gaditana) to determine the relative
31 Pb sensitivities as measured by growth inhibition. The respective 72-hour EC50 values
32 derived were 2,640, 900, 105, 1,340, and 740 (ig Pb/L (Debelius et al.. 2009). The
November 2012 7-189 Draft - Do Not Cite or Quote
-------
1 authors noted that species cellular size, sorption capacity, or taxonomy did not explain
2 differences in sensitivity to Pb, leaving the mechanism of response still open to question.
3 In the deposit feeding polychaete, Capitella sp. an LOAEL of 85 mg Pb/kg sediment was
4 established in 3 day and 6 day growth experiments (Horng et al., 2009). Other studies of
5 marine invertebrates published since the 2006 Pb AQCD (U.S. EPA. 2006c) have
6 indicated differences in sensitivity of different lifestages of aquatic organisms to Pb. In a
7 series of seawater and sediment exposures using adult and juvenile amphipods Melita
8 plumulosa, juveniles were more sensitive to Pb than adults (King et al., 2006). In the
9 seawater-only exposures, the 96-hour LC50 was 1,520 (ig Pb/L for juveniles and
10 3,000 (ig Pb/L for adults. In comparison,10 day juvenile sediment test results were LC50
11 1,980, NOEC 580 and LOEC 1,020 mg Pb/kg dry weight compared to the LC50, NOEC,
12 and LOEC value for the adults exposed in sediment (3,560 mg Pb/kg dry weight). A 24-
13 hour LC50 of 4,500 (ig Pb/L for adult black mussel (M galloprovincialis) suggests that, in
14 general, juvenile bivalves are more sensitive to Pb exposure than adults although this
15 value was based on nominal exposure data (Vlahogianni and Valavanidis. 2007).
16 Since the 2006 Pb AQCD , Pb toxicity to larval stages of marine species has been
17 assessed at sublethal and lethal concentrations. The effective concentrations at which Pb
18 resulted in 50% of abnormal embryogenesis of the Asian clam (M meretrix) was
19 297 (ig Pb/L. The 96-hour LC50 for larvae of the same species was 353 (ig Pb/L (Wang et
20 al., 2009d). In comparison, juvenile Catarina scallop (A. ventricosus) had a LC50 of
21 830 (ig Pb/L in a 96-hour exposure (Sobrino-Figueroa et al.. 2007). In the marine
22 polychaete H. elegans, EC50 values of gametes, embryos, larvae (blastula to trochophore
23 and larval settlement), and adults, exhibited dose-responses to Pb that reflected the
24 differential sensitivity of various lifestages of this organism (Gopalakrishnan et al..
25 2008). The EC50 values for sperm and egg toxicity were 380 and 690 (ig Pb/L
26 respectively. Larval settlement measured as the metal concentration causing 50%
27 reduction in attachment was most sensitive to Pb with an EC50 of 100 (ig Pb/L, while the
28 EC50 for abnormal development of embryos was 1,130 (ig Pb/L. The LC50 values for
29 adult worms in 24-hour and 96-hour tests were 25,017 and 946 (ig Pb/L, respectively.
30 Manzo et al. (2010) established a LOEC of 500 (ig Pb/L and a maximum effect at
31 3,000 (ig Pb/L in an embryotoxicity assay with sea urchin P. lividus exposed to nominal
32 concentrations of Pb. The EC50 for developmental defects in this species was
33 1,250 (ig Pb/L with a NOEL of 250 (ig Pb/L. In a study using nominal concentrations of
34 Pb, morphological deformities were observed in 50% of veliger larvae of blacklip
35 abalone (Haliotis rubra) at 4,100 (ig Pb/L following a 48-hour exposure, suggesting this
36 species is not as sensitive to Pb as other marine invertebrate larvae (Gorski and
37 Nugegoda. 2006).
November 2012 7-190 Draft - Do Not Cite or Quote
-------
7.4.17 Community and Ecosystem Effects in Saltwater Systems
1 As discussed in the 1986 Pb AQCD and the 2006 Pb AQCD (U.S. EPA. 2006c).
2 exposure to Pb is likely to have impacts in aquatic environments via effects at several
3 levels of ecological organization (organisms, populations, communities, or ecosystems).
4 But fewer studies explicitly consider community and ecosystem-level effects in marine
5 and brackish waters than in freshwater. Reduced species abundance and biodiversity of
6 protozoan and meiofauna communities were observed in laboratory microcosm studies
7 with marine water and marine sediments reviewed in the 2006 Pb AQCD as summarized
8 in Table AX7-2.5.2 (U.S. EPA. 2006c). In a laboratory study with larval mummichogs
9 reviewed in the 2006 Pb AQCD, feeding and predator avoidance behaviors were altered
10 in this marine fish species following a 4-week exposure to Pb. Observations from field
11 studies reviewed in the 2006 Pb AQCD included findings of a negative correlation
12 between Pb and species richness and diversity indices of macroinvertebrates associated
13 with estuary sediments and changes in species distribution and abundance in fish,
14 crustaceans and macroinvertebrates correlated with Pb levels in marine sediments. The
15 2006 Pb AQCD concluded that, in general, information from controlled studies for single
16 pollutants was insufficient to permit evaluation of specific impacts on higher levels of
17 organization (beyond the organism). In studies from natural saltwater ecosystems, Pb
18 rarely occurs as a sole contaminant making its effects difficult to ascertain. New
19 information on effects of Pb at the population, community and ecosystem level in coastal
20 ecosystems is reviewed below.
21 The faunal composition of seagrass beds in a Spanish coastal saltwater lagoon was found
22 to be impacted by Pb in sediment, plants, and biofilm (Marin-Guirao et al., 2005).
23 Sediment Pb concentrations ranged from approximately 100 to 5,000 mg Pb/kg and
24 corresponding biofilm concentrations were 500 to 1,600 mg Pb/kg, with leaf
25 concentrations up to 300 mg Pb/kg. Although multiple community indices (abundance,
26 Shannon-Wiener diversity, Simpson dominance index) did not vary from site to site,
27 multivariate analysis and similarity analysis indicated significant differences in
28 macroinvertebrate communities between sites with different sediment, biofilm, and leaf
29 Pb concentrations. Differences were largely attributable to three amphipod species
30 (Microdeutopus sp., Siphonoecetes sabatieri, Gammarus sp.). This indicates that,
31 although seagrass abundance and biomass were unaffected by Pb exposure, organisms
32 inhabiting these plants still may be adversely impacted.
33 Caetano et al. (2007) investigated the mobility of Pb in salt marshes using total content
34 and stable isotope signature. They found that roots had similar isotopic signature to
35 sediments in vegetated zones indicating that Pb uptake by plants reflects the input in
36 sediments. At one site, there was a high anthropogenic Pb content while at the other
November 2012 7-191 Draft - Do Not Cite or Quote
-------
1 natural mineralogical sources dominated. The roots of S. fruticosa and S. maritima
1 significantly accumulated Pb, having maximum concentrations of 2,870 mg Pb/kg and
3 1,755 mg Pb/kg, respectively, indicating that below-ground biomass played an important
4 role in the biogeochemical cycling of Pb.
5 Exposure to three levels of sediment Pb contamination (322, 1,225, and 1,465 mg Pb/kg
6 dry weight) had variable effects on different species within a marine nematode
7 community (Mahmoudi et al.. 2007). Abundance, taxa richness, and species dominance
8 indices were altered at all Pb exposures when compared with unexposed communities.
9 Further, while the species Oncholaimellus mediterraneus dominated control communities
10 (14% of total abundance), communities exposed to low and medium Pb concentrations
11 were dominated by Oncholaimus campylocercoides (36%) andMarylynnia stekhoveni
12 (32%), and O. campylocercoides (42%) and Chromadorina metulata (14%), respectively.
13 Communities exposed to the highest Pb sediment concentrations were dominated by
14 Spirinia gerlachi (41%) and Hypodontolaimus colesi (29%). Given this, the authors
15 concluded that exposure to Pb significantly reduced nematode diversity and resulted in
16 profound restructuring of the community structure.
17 In another laboratory microcosm experiment with nematodes, nematode diversity and
18 community structure was altered with a mean number of 8 genera present in microcosms
19 contaminated with Pb compared to the control with 20 genera. The spiked sediments used
20 in the study were collected from the Swartkop River estuary, South Africa. Pb (3 to
21 6,710 mg Pb/kg sediment dry weight) was tested alone and in combination with Cu, Fe,
22 and Zn (Gyedu-Ababio and Baird. 2006). The synergistic effect of the four metals on
23 nematode community structure was greater than the individual metals and the effects of
24 Pb could not be distinguished from Cu, Fe and Zn.
November 2012 7-192 Draft - Do Not Cite or Quote
-------
7.4.18 Characterization of Sensitivity and Vulnerability in Saltwater Species
1 Species differences in metabolism, sequestration, and elimination rates have been shown
2 to control relative sensitivity and vulnerability of exposed organisms and effects on
3 survival, reproduction, growth, metabolism, and development. Diet and lifestage at the
4 time of exposure also contribute significantly to the determination of sensitive and
5 vulnerable populations and communities. Further, environmental conditions in addition to
6 those discussed as affecting bioavailability may also alter Pb toxicity. The
7 2006 Pb AQCD (U.S. EPA. 2006c) reviewed the effects of genetics, age, and body size
8 on Pb toxicity. While genetics appears to be a significant determinant of Pb sensitivity,
9 effects of age and body size are complicated by environmental factors that alter metabolic
10 rates of saltwater organisms. A review of the more recent literature corroborated these
11 findings, and identified seasonal physiological changes and lifestage as other important
12 determinants of differential sensitivity to Pb.
7.4.18.1 Seasonally Affected Physiological Changes
13 Couture et al. (2010) investigated seasonal and decadal variations in Pb sources to
14 mussels (M. edulis) from the French Atlantic shoreline. Pb concentrations in the mussels
15 were 5-66 times higher than the natural background value for the north Atlantic. The
16 206Pb/207Pb signature indicated that the bioaccumulated Pb was anthropogenic in origin.
17 The signature was not, however, the same as that emitted in western Europe, as a result of
18 leaded gasoline combustion, although that was a major emission source to the atmosphere
19 during a large part of the study period (1985-2005). Instead, it was most similar to that of
20 Pb released into the environment from wastewater treatment plants, municipal waste
21 incinerators and industries such as metal refineries and smelters. Thus continental runoff
22 rather than atmospheric deposition was identified as the main source of Pb to the French
23 coastal area. The strong seasonal variations in 206Pb/208Pb were used to conclude that
24 re suspension of Pb triggered by high river runoff events was a key factor affecting
25 bioaccumulation of Pb inM edulis.
26 In another monitoring study, Pearce and Mann (2006) investigated variations in
27 concentrations of trace metals in the U.K. including Pb in the shells of pod razor shell
28 (Ensis siliqud). Pb concentration varied from 3.06-36.2 mg Pb/kg and showed a regional
29 relationship to known sources, e.g., former metal mining areas such as Cardigan Bay,
30 Anglesey, and industrial activity in Liverpool Bay. Seasonal variations were also found
31 for Pb in both Cardigan Bay and Liverpool Bay, relating to increased winter fluxes of Pb
32 (and other metals) into the marine environment. In contrast, levels of Pb and other metals
November 2012 7-193 Draft - Do Not Cite or Quote
-------
1 were highest in summer and lowest in winter in oysters Crassostrea corteziensis collected
2 from Sonora, Mexico (Garcia-Rico et al.. 2010).
3 Carvalho et al. (2011) quantified 210Pb inM galloprovincialis sampled at coastal
4 locations in Portugal and noted that the apparent seasonal fluctuation in radionuclide
5 concentrations in mussel soft tissues was mostly attributable to changes in physiological
6 condition (i.e., fat content, gonadal development) and not to radionuclide body burden
7 fluctuation. The authors caution that since concentrations of contaminants are dependent
8 upon tissue composition, corrections for mussel physiological condition are need to
9 compare results from different seasons and different locations.
7.4.18.2 Lifestage
10 Lifestages of the marine polychaete H. elegans including embryogenesis, sexual
11 maturation, and offspring development were shown to be differentially affected by Pb
12 exposure. Pb water concentrations of 91 (ig Pb/L and greater significantly affected
13 fertilization and embryonic development, but the greatest effects were exhibited by 24-
14 hour-old larvae (Gopalakrishnan et al.. 2007). The authors suggested that timing of Pb
15 exposure may have different impacts on marine polychaete populations, if life cycles are
16 offset (Gopalakrishnan et al., 2007). Further, given that the adult lifestage is sedentary,
17 reduction of the mobile early lifestage as a result of Pb exposures may disproportionally
18 affect sessile polychaetes. For instance, larval settlement was significantly reduced at Pb
19 exposures of 48 (ig Pb/L and greater (Gopalakrishnan et al.. 2008).
7.4.18.3 Species Sensitivity
20 Both inter- and intra-specific differences in Pb uptake and bioaccumulation may occur in
21 macroinvertebrates of the same functional feeding group. Data from 20 years of
22 monitoring of contaminant levels in filter-feeding mussels of the Mytilus species and
23 Crassostrea virginica oysters in coastal areas of the U.S. through the National Oceanic
24 and Atmospheric Administration (NOAA) Mussel Watch program indicate that Pb is on
25 average three times higher in mussels than in oysters (Kimbrough et al., 2008). Limpet
26 (Patella sp.) from the Lebanese Coast had Pb BAF values ranging from 2,500 to 6,000
27 and in the same field study Pb BAF values for a mussel (Brachidontes variabilis) ranged
28 from 7,500-8,000 (Nakhle et al.. 2006).
29 There is some indication that molting may comprise an additional sequestration and
30 excretion pathway for aquatic animals exposed to Pb (Soto-Jimenez et al.. 201 la;
31 Mohapatra et al.. 2009: Tollett et al.. 2009: Bergev and Weis. 2007). Crab species
November 2012 7-194 Draft - Do Not Cite or Quote
-------
1 U. pugnax (Bergey and Weis. 2007) and Scylla serrata fMohapatra et al., 2009A and
2 white shrimp L. vannamei (Soto-Jimenez et al.. 201 la) have been shown to sequester Pb
3 preferentially in exoskeleton tissue, where it is later shed along with other tissue.
4 Consequently, aquatic arthropod species and those species that shed their exoskeleton
5 more frequently may be able to tolerate higher environmental Pb concentrations than
6 non-arthropods or slow-growing molting species, as this pathway allows them to
7 effectively lower Pb body burdens.
8 Some tolerant species offish (e.g., mummichog) have the ability to sequester
9 accumulated Pb in metal-rich granules or heat-stable proteins (Goto and Wallace. 2010).
10 Fish with such abilities are more likely to thrive in Pb-contaminated environments than
11 other species.
7.4.19 Ecosystem Services Associated with Saltwater Systems
12 Pb deposited on the surface of (or taken up by) organisms has the potential to alter the
13 services provided by saltwater biota to humans although the directionality of impacts is
14 not always clear. For example, oysters and mussels provide a service by sequestering Pb.
15 At the same time, the uptake of Pb by these bivalves may result in toxicological effects
16 associated with Pb exposure and decreased value of shellfish as a commodity. At this
17 time, a few publications address Pb impacts on ecosystem services associated with
18 saltwater ecosystems. Pb can affect the ecological effects in each of the four main
19 categories of ecosystem services (Section 7.1.2) as defined by Hassan et al. (2005). These
20 effects are sorted into ecosystem services categories and summarized here:
21 • Supporting: food for higher trophic levels, biodiversity
22 • Provisioning: contamination of food by heavy metals, decline in health offish
23 and other aquatic species
24 • Regulating: water quality
25 • Cultural: ecosystem and cultural heritage values related to ecosystem integrity
26 and biodiversity, wildlife and bird watching, fishing
27 A few recent studies explicitly consider the impact of Pb and other heavy metals on
28 ecosystem services provided by salt marsh (Gedan et al., 2009) and estuaries (Smith et
29 al.. 2009b). These systems are natural sinks for metals and other contaminants. Pb can be
30 toxic to salt marsh plant species and decaying plant detritus may result in resuspension of
31 Pb into the aquatic food chain (Gedan et al.. 2009). Salt marsh and estuaries provide
32 habitat and breeding areas for both terrestrial and marine wildlife and are locations for
November 2012 7-195 Draft - Do Not Cite or Quote
-------
1 bird watching. Using a modeling approach designed to assess the degree of risk of Pb and
2 Hg to wading birds in estuarine habitats in the U.K., the authors found a high probability
3 that Pb poses an ecologically relevant risk to dunlin, Calidris alpina fSmith et al., 2QQ9b).
4 However, the authors noted that a major source of uncertainty in this study was the
5 NOAEL values for Pb.
6 The impact of Pb on ecological services provided by specific components of aquatic
7 systems has been considered in a limited number of studies. Recent research has
8 suggested that dietary Pb (i.e., Pb adsorbed to sediment, particulate matter, and food) may
9 contribute to exposure and toxicity in primary and secondary order consumers (including
10 humans). Aquatic fauna can take up and bioaccumulate metals. If the bioaccumulating
11 species is a food source, the uptake of metals may make it toxic or more dangerous for
12 people or other wildlife to consume. For example, oysters and mussels bioaccumulate Pb
13 from anthropogenic sources, including atmospheric deposition, and are a food source that
14 is widely consumed by humans and wildlife (Couture et al.. 2010). Their capacity to
15 bioaccumulate Pb makes them good bioindicators of environmental contamination and
16 they have been used as monitors of coastal pollutants by the NOAA Mussel Watch
17 program since 1986. Although bioaccumulation may render aquatic fauna toxic to
18 consumers, bioaccumulation is a way to sequester the metals and remove them from
19 waters and soils. Sequestration for this purpose is itself an ecosystem service and has
20 been quantified. For example, the total ecological services value of a constructed
21 intertidal oyster (Crassostrea sp.) reef in improving water quality and sequestering metals
22 including Pb was calculated in the Yangtze River estuary to be about $500,000 per year
23 (Quan et al.. 2009).
7.4.20 Synthesis of New Evidence for Pb Effects in Saltwater Systems
24 This synthesis of the effects of Pb on saltwater ecosystems covers information from the
25 publication of the 2006 Pb AQCD (U.S. EPA. 2006c) to present. It is followed in
26 Section 7.4.21 by determinations of causality that take into account evidence dating back
27 to the 1977 Pb AQCD. In general, evidence for toxicity to saltwater organisms is less
28 well characterized than toxicity of Pb in freshwater ecosystems due to the fewer number
29 of available studies on marine species. The studies that are available for marine plants,
30 invertebrates and vertebrates include studies where Pb concentration was analytically
31 verified and those that reported nominal concentrations (Table 7-6). Many of the studies
32 that report nominal concentrations in media are uptake studies that subsequently quantify
33 Pb in tissues; however, measurement of Pb in water or sediment at the beginning of an
34 exposure is desirable when comparing laboratory studies to concentrations of Pb in
35 marine systems. In Section 7.2.3 and Table 7-2. a range of 0.01 to 27 (ig Pb/L was
November 2012 7-196 Draft - Do Not Cite or Quote
-------
1 reported for saltwater, including estuaries and open ocean, with the higher values
2 associated with sites involving human activity (Sadiq. 1992).
3 Most studies on marine organisms reviewed in the present document included
4 concentrations that were higher than Pb encountered in seawater. However, when
5 multiple concentrations were used, effects generally increased with increasing Pb
6 exposure. Effects at lower concentrations can be implied from many reported studies
7 since an exposure response relationship to Pb was observed. In marine and estuarine
8 systems, exposure to Pb from air is most likely characterized as a chronic low dose
9 exposure, however, most studies only report an acute LC50 value when an LOEC or LCio
10 would be more appropriate measurement for consideration of effects on organisms since
11 an effect occurring at the LC50 value would most likely not maintain a stable population.
Plants
12 Only a few studies were available since the 2006 Pb AQCD, that consider effects of Pb
13 on marine algae (Section 7.4.15.1). A 72-hour EC50 for growth inhibition was reported in
14 the marine algae Chaetoceros sp. at 105 (ig Pb/L (Debelius et al.. 2009). A study with the
15 green alga T. suecica reports a statistically significant decease in growth rate, total dry
16 biomass and final cell concentration between control cultures and algae cultured in
17 20 (ig Pb/L (Soto-Jimenez et al., 201 Ib). Both of these studies suggest growth effects at
18 or near the highest recorded values of Pb in seawater (27 (ig Pb/L), however, effects are
19 likely to occur at lower concentrations since only EC50 values are reported.
Invertebrates
20 In saltwater invertebrates (Section 7.4.15.2 and 7.4.16) there are studies that consider
21 Pb-effects on supporting endpoints (stress responses, hematological effects and
22 neurobehavior) as well as studies that assess Pb impacts to reproduction, growth, and
23 survival; endpoints that have the potential to alter population, community and
24 ecosystem—levels of biological organization. Many studies, especially those that
25 consider enzymatic responses to Pb exposure, were conducted with nominal Pb
26 concentrations. Two of these studies; Jing et al. (2007) and Zhang et al., (201 Ob) consider
27 Pb nominal exposures at 100 (ig Pb/L or lower and reported significant decreases in
28 antioxidant enzyme activity. The Zhang et al. (201 Ob) study observed effects on
29 enzymatic activity at a nominal exposure of 2 (ig Pb/L. Although these effects are near
30 reported Pb concentrations in seawater they were not analytically verified.
31 Other studies that report sub-organismal responses in saltwater organisms have quantified
32 Pb exposure. Field studies with bivalves collected off the coast of Spain correlated
November 2012 7-197 Draft - Do Not Cite or Quote
-------
1 ALAD activity with measured levels of Pb in tissue (Company et al.. 2011; Kalman et al..
2 2008). An increase in valve closing time with increasing Pb exposure in the range of 40
3 to 400 (ig Pb/L was observed in the scallop, A.ventricosus (Sobrino-Figueroa and
4 Caceres-Martinez. 2009). Although the concentrations in this study exceed reported
5 levels of Pb in seawater, the lower range is near 27 (ig Pb/L reported by Sadiq (1992).
6 Evidence for effects on reproduction, growth and survival in marine invertebrates (Table
7 7-6) are primarily from studies in which Pb in the exposure media was quantified. In the
8 amphipod, E. laevis, onset to reproduction was significantly delayed at 118 mg/Pb kg
9 sediment; a concentration that the authors indicate is below the current marine sediment
10 regulatory guideline for Pb (218 mg Pb/kg sediment) (Ringenary et al.. 2007; NOAA.
11 1999). In the same study, no effects of Pb on adult survival in 28 and 60 day sediment
12 exposures were observed. In another study with amphipods, juvenile M. plumosa were
13 more sensitive than adults in 10-day sediment exposures with an NOEC of 580 mg Pb/kg
14 dry weight compared to an NOEC of 3,560 mg Pb/kg dry weight for adults (King et al..
15 2006). Effects of Pb on gametes of the marine polycheate H. elegans were observed at
16 48 (ig Pb/L (Gopalakrishnan et al.. 2008). a concentration near the upper range of Pb in
17 seawater reported by Sadiq (1992). Specifically, fertilization rate of eggs pretreated with
18 48 (ig Pb/L decreased to 20% of control. Life stages ofH. elegans varied in their
19 sensitivity to Pb with the most sensitive period being larval settlement with an EC50 of
20 100 (ig Pb/L.
21 There are only a few recent studies that considered effects of Pb on growth of marine
22 invertebrates (Sections 7.4.15.2 and 7.4.16). In the polychaete Capitella sp. growth was
23 decreased significantly from controls, however, there was not a clear dose-response
24 relationship between increasing Pb concentrations and observed effects (Horng et al..
25 2009). The authors reported a LOAEL of 85 mg Pb/kg in the sediment exposure. In the
26 Asian Clam M. meretrix, an EC50 of 197 (ig Pb/L was reported for growth (Wang et al..
27 2009d). Other marine invertebrate growth effects were observed at much higher Pb
28 concentrations (Table 7-6).
29 Survival was a less sensitive endpoint in marine invertebrates than reproduction or
30 growth with no effects reported at concentrations typically observed in seawater (Table
31 7-6). In the amphipodM plumulosa an NOEC of 400 (ig Pb/L for juveniles and an
32 NOEC of 850 (ig Pb/L was reported for adults in 96-hour seawater only exposures (King
33 et al.. 2006). In 10 day sediment tests with the same species, juveniles were also more
34 sensitive than adults. Other concentrations at which survival effects were reported in
35 marine invertebrates also greatly exceeded concentrations of Pb typically found in
36 seawater.
November 2012 7-198 Draft - Do Not Cite or Quote
-------
Vertebrates
1 There is not sufficient new evidence for saltwater vertebrates especially for reproductive,
2 growth and survival endpoints that may have relevance to the population-level of
3 biological organization and higher.
Food Web
4 Some studies published since the 2006 Pb AQCD (see Section 7.4.14.4) support the
5 potential for Pb to be transferred in saltwater food webs, while other studies have found
6 no evidence for biomagnification.
Ecosystem Level Effects
7 Evidence for effects at higher levels of biological organization in saltwater habitats is
8 primarily supported by observations in a small number of microcosm and field studies
9 where shifts in community structure are the most commonly observed effects of Pb
10 (Section 7.4.17). Effects on reproduction, growth or survival (summarized in Table 7-6)
11 may lead to effects at the population-level of biological organization and higher.
7.4.21 Causal Determinations for Pb in Saltwater Systems
12 In the following sections, organism-level effects on reproduction and development,
13 growth and survival are considered first since these endpoints can lead to effects at the
14 population level or above and are important in ecological risk assessment.
15 Neurobehavioral effects are considered next followed by sub-organismal responses
16 (hematological effects, physiological stress) for which Pb has been shown to have an
17 impact in multiple species and across taxa, including humans. Causal determinations for
18 terrestrial, freshwater and saltwater ecological effects are summarized in Table 7-3.
7.4.21.1 Reproductive and Developmental Effects-Saltwater Biota
19 Reproductive effects of Pb have been reported in a few marine organisms and the
20 majority of the available studies are with invertebrate species. In a study reviewed in the
21 2006 Pb AQCD (U.S. EPA. 2006c). embryo development in two commercial bivalves
22 Ruditapes decussatus and M. galloprovincialis was inhibited by Pb (Beiras and
23 Albentosa. 2003). In R. decussatus an EC50 range of 156 to 312 (ig Pb/L and LOEC of
24 156 (ig Pb/L were observed for inhibition of embryonic development while in
November 2012 7-199 Draft - Do Not Cite or Quote
-------
1 M. galloprovincialis the EC50 was 221 (ig Pb/L and the LOEC was 50 (ig Pb/L. Larvae of
2 the mussel M. edulis were sensitive to Pb exposure with an EC50 of 476 (ig Pb/L for
3 abnormal development of embryos following 48-hour exposure to Pb during
4 embryogenesis (Martin et al.. 1981). The LOEC for embryogenesis in the marine bivalve
5 M. galloprovincialis was 50 (ig Pb/L with an EC50 for embryogenesis of 221 (ig Pb/L
6 (Beiras and Albentosa. 2003).
7 Recent evidence for reproductive effects of Pb on marine invertebrates is summarized in
8 Table 7-6. In the marine polychaete H. elegans an EC50 of 261 (ig Pb/L was observed for
9 unhatched or abnormal larvae following 20 hour incubation with Pb (Gopalakrishnan et
10 al.. 2008). The EC50 for the metal concentration causing 5% reduction in larval
11 attachment was 100 (ig Pb/L. The EC50 values for sperm and egg toxicity were 380 and
12 692 (ig Pb/L, respectively. The EC50 for embryogenesis in the clamM meretrix was
13 297 (ig Pb/L (Wang et al., 2009d). In a multigenerational bioassay with the marine
14 amphipod E. laevis, statistically significant delays in onset of reproduction (4 to 8 days),
15 sexual maturation and first offspring were observed at concentrations of 188 mg Pb/kg
16 sediment and higher (Ringenary et al.. 2007). The authors indicate that this concentration
17 is below the current sediment regulatory guideline for Pb (218 mg Pb/kg sediment)
18 (TSfOAA. 1999) and that reproductive effects are a more sensitive endpoint than lethality.
19 Although LC50 values are typically reported for Pb effects on reproductive endpoints in
20 saltwater invertebrates, a concentration dependent relationship between reproductive
21 impairment and increasing concentration of Pb is reported in most studies. This exposure-
22 response relationship implies that effects on reproduction are occurring at concentrations
23 lower than the LC50 value.
24 Reproductive effects are only characterized in a few species and endpoints for marine
25 systems. The weight of the current evidence for reproductive effects is limited to
26 laboratory-based studies with saltwater invertebrates in which observed effects occur at
27 Pb concentrations that are higher than Pb concentrations encountered in the marine
28 environment. Evidence for reproductive effects of Pb on marine plant species is limited to
29 one study on the red alga (Champia parvuld) reviewed in the draft Ambient Aquatic Life
30 Water Quality Criteria for Pb (U.S. EPA. 2008b). In one study from a saltwater fish,
31 field-collected smooth toadfish (T. glaber) from metal contaminated estuaries in Sydney,
32 Australia had elevated Pb levels in gonad and decreased oocyte diameter and density.
33 Evidence is, therefore, inadequate to conclude that there is a causal relationship for
34 reproductive effects in saltwater plants, and vertebrates. The available studies on marine
35 invertebrates are suggestive that there is a causal relationship between Pb exposure and
36 reproductive effects.
November 2012 7-200 Draft - Do Not Cite or Quote
-------
7.4.21.2 Growth Effects-Saltwater Biota
1 There are few studies that measure growth effects of Pb on marine organisms; available
2 information is limited to marine flora and invertebrates. Growth studies in saltwater plant
3 species are summarized in Table 4 and Table 6 of the draft Ambient Aquatic Life Water
4 Quality Criteria for Pb (U.S. EPA. 2008b) and Table 7-6 of the present document.
5 Diatoms are among the most sensitive algae; however, growth effects are typically
6 observed at concentrations of Pb higher than the range of values available from saltwater
7 locations [0.01 to 27 (ig Pb/L, (Sadiq. 1992)1. In studies available since the draft Ambient
8 Aquatic Life Water Quality Criteria for Pb, the lowest 72-hour EC50 for growth inhibition
9 reported in marine diatoms was 105 (ig Pb/L in Chaetoceros sp (Debelius et al., 2009)
10 and the growth of the green alga T. suecica exposed nominally to 20 (ig Pb/L was 40%
11 lower than control cultures (Soto-Jimenez et al., 201 Ib). The microalgae was the base of
12 a simulated marine food chain including primary, secondary and tertiary level consumers
13 and effects on survival were observed at the higher trophic levels that originated from Pb
14 exposure via consumption of the primary producer. The majority of growth effects
15 reported in saltwater algae exceed concentrations of Pb in seawater by several orders of
16 magnitude. Effects of Pb on growth in two species of brown algae, Fucus vesiculosus and
17 Fucus serratus are summarized in Table 6 of the draft Ambient Aquatic Life Water
18 Quality Criteria for Pb (U.S. EPA. 2008b). Concentrations where growth impairment was
19 observed in these species greatly exceed available values for Pb measured in seawater.
20 In saltwater invertebrates, evidence for growth effects is limited to a few species at
21 concentrations that exceed Pb concentrations reported in seawater. Growth inhibition in
22 the bivalve Macoma balthica (EC50=453.4 (ig Pb/L) is reported in Table 6 of the draft
23 Ambient Aquatic Life Water Quality Criteria for Pb (U.S. EPA. 2008b). Recent studies
24 include Wang et al., (2009d_) in which observed growth of embryos of the Asian Clam
25 (M meretrix) was significantly reduced by Pb with an EC50 of 197 (ig Pb/L. In juvenile
26 Catarina scallop, A. ventricosus, exposed to Pb for 30 days, the EC50 for growth was
27 4,210 (ig Pb/L (Sobrino-Figueroa et al., 2007). Rate of growth of the deposit feeding
28 polychaete Capitella sp. exposed to Pb-spiked sediments from polluted estuaries
29 decreased significantly from the control; however, changes were inconsistent with
30 increasing concentration of Pb (Horng et al.. 2009). Evidence is therefore inadequate to
31 conclude that there is a causal relationship between Pb exposure and growth effects in
32 saltwater plants, invertebrates and vertebrates.
November 2012 7-201 Draft - Do Not Cite or Quote
-------
7.4.21.3 Survival-Saltwater Biota
1 There are no studies reported in the previous Pb AQCDs or the current ISA for aquatic
2 plants that indicate phytotoxicity at or near current concentrations of Pb in saltwater [0.01
3 to 27 ng Pb/L, (Sadiq. 1992)1.
4 Mortality data for saltwater invertebrate species are summarized in the draft Ambient
5 Aquatic Life Water Quality Criteria for Pb (U.S. EPA. 2008b) and reported LC50 values
6 greatly exceed Pb concentrations encountered in seawater. Recent studies available since
7 the 2006 Pb AQCD, and draft Aquatic Life Water Quality Criteria for Pb that report
8 mortality data are summarized in Table 7-6. In general, marine fauna are less sensitive to
9 this metal than freshwater fauna and the highest toxicity is observed in juveniles. A 144-
10 hour LC50 of 680 (ig Pb/L was reported for juvenile scallop A. ventricosus (Sobrino-
11 Figueroa et al., 2007) and a 96-hour LC50 of 353 (ig Pb/L for embryos of the clam
12 M. meretrix (Wang et al.. 2009d). In the amphipod M. plumulosa, juveniles were more
13 sensitive to Pb than adults in 96 hour seawater-only exposures and 10 day sediment
14 exposures (King et al.. 2006). The 96-hour LC50 was 1,520 jig Pb/L and the NOEC was
15 400 (ig Pb/L for juveniles in comparison to adults (96-hour LC50 =3,000 (ig Pb/L;
16 NOEC=1,680 (ig Pb/L). In the 10-day sediment exposures, the NOEC for juveniles was
17 580 mg Pb/kg dry weight compared to an adult NOEC of 3,560 mg Pb/kg dry weight. In
18 10-day exposures to Pb nitrate spiked sediment, all individuals of the bivalve
19 T. deltoidalis survived at 1,000 mg/Pb kg with 15 (ig Pb/L dissolved in pore water (King
20 et al., 2010). No effects on survival were observed in either the amphipod E. laevis
21 exposed 60 days to Pb-spiked sediment up to 424 mg Pb/kg (Ringenary et al.. 2007). or
22 in the polychaete Capitella sp. exposed to sediment for 3 or 6 days up to 871 mg Pb/kg
23 (Horng et al.. 2009).
24 Effects of Pb on survival have been demonstrated though a simulated marine food chain
25 in which the primary producer, the microalgae T. suecica, was exposed nominally to
26 20 (ig Pb/L and subsequently fed to brine shrimp A. franciscana, (mean Pb content 12 to
27 15 mg Pb/kg) which were consumed by white-leg shrimp L. vannamei, itself consumed
28 by grunt fish H. scudderi representing the top of the marine food chain (Soto-Jimenez et
29 al., 201 Ib). Survival of brine shrimp was 25 to 35% lower than the control and both
30 white shrimp and grunt fish had significantly higher mortalities than controls.
31 Data on Pb toxicity to eight species of marine fishes are summarized in Table 1 of the
32 draft Ambient Aquatic Life Water Quality Criteria for Pb (U.S. EPA. 2008b). All of the
33 LC50 values for these fish (range 1,500 to 315,000 (ig Pb/L) greatly exceed
34 concentrations of Pb reported in seawater. Additionally, in the 2006 Pb AQCD (U.S.
35 EPA. 2006c) the acute toxicity of Pb to plaice (Pleuronectes platessa) was reported to
36 range from 50 (ig Pb/L to 300,000 (ig Pb/L depending on the form of Pb (Eisler. 2000).
November 2012 7-202 Draft - Do Not Cite or Quote
-------
1 The existing evidence on toxicity of Pb to marine vertebrates is limited to laboratory-
2 based studies conducted under different salinities and exposure conditions. Considerable
3 uncertainties exist in applying laboratory observations to actual conditions in the field
4 where other modulating factors can affect Pb bioavailability and toxicity.
5 Although evidence exists for increased mortality of marine fish at very high
6 concentrations of Pb, the focus of the causal determinations are on studies where effects
7 were observed within one to two orders of magnitude of Pb measured in the environment
8 (Preamble Table II). Evidence is therefore inadequate to conclude that there is a causal
9 relationship between Pb and survival in saltwater plants, invertebrates and vertebrates.
7.4.21.4 Neurobehavioral Effects-Saltwater Biota
10 In marine organisms evidence for neurobehavioral effects of Pb is limited to a few studies
11 on bivalves and fish. In a study reviewed in the 2006 Pb AQCD (U.S. EPA. 2006c). prey
12 capture rate and predator avoidance was affected in mummichogs starting at 300 (ig Pb/L
13 (Weis and Weis. 1998). Recent studies support previous findings of decreased ability to
14 escape predation associated with Pb exposure. In juvenile Catarina scallops exposed to
15 Pb (40 (ig/L to 400 (ig/L) for 20 days, the average valve closing time increased from
16 under one second in the control group to 3 to 12 seconds in juvenile scallops A decrease
17 in valve closing speed in these bivalves may impact escape swimming behaviors
18 important for predator avoidance (Sobrino-Figueroa and Caceres-Martinez. 2009).
19 Behavioral effects in grunt fish H. scudderi, occupying the top level of a simulated
20 marine food chain included lethargy and decreased food intake in a 42-day feeding study
21 (Soto-Jimenez et al.. 201 Ib). These fish were fed white shrimp exposed to Pb via brine
22 shrimp that were initially fed microalgae cultured at a nominal concentration of
23 20 (ig Pb/L. In the same study, surfacing, reduction of motility, and erratic swimming
24 were observed in the white shrimp after 30 days of exposure to Pb via diet. The ornate
25 wrasse, T. pavo, was exposed nominally to sublethal (400 (ig Pb/L) or a maximum
26 acceptable toxicant concentration (1,600 (ig Pb/L) dissolved in seawater for one week to
27 assess the effects of Pb on feeding and motor activities (Giusi et al.. 2008). In the
28 sublethal concentration group, hyperactivity was elevated 36% over controls. In the high
29 concentration, a 70% increase in hyperactivity was observed and hyperventilation
30 occurred in 56% of behavioral observations, however, no changes in feeding activity
31 were noted between non-treated and treated fish.
32 Most of the evidence for neurobehavioral changes in marine organisms is observed with
33 concentrations of Pb that exceed the range of Pb values available for saltwater of 0.01 to
34 27 (ig Pb/L (Sadiq. 1992)1. with the exception of the food chain study discussed above in
November 2012 7-203 Draft - Do Not Cite or Quote
-------
1 which behavioral effects were observed in shrimp and their fish predators following
2 ingestion of microalgae cultured in nominal concentration of 20 (ig Pb/L and then
3 quantified in prey (Soto-Jimenez et al.. 201 Ib). Marine species are typically
4 underrepresented in toxicity testing of behavioral endpoints with metals. There are
5 considerable uncertainties in applying observations from laboratory-based studies to field
6 scenarios including the role of environmental factors such as salinity and DOM on Pb
7 bioavailability. Evidence is inadequate to conclude that there is a causal relationship
8 between Pb exposures and neurobehavioral endpoints in saltwater invertebrates and
9 vertebrates.
7.4.21.5 Hematological Effects-Saltwater Biota
10 Evidence for hematological effects of Pb on saltwater organisms is limited primarily to
11 field monitoring studies on bivalves. Several recent field studies using a multi-biomarker
12 approach to study the sources and impacts of Pb in marine environments have measured
13 ALAD activity in bivalve species and found positive correlations between increased
14 tissue Pb levels and ALAD inhibition (e.g., Company et al. (2011). Kalman et al (2008)).
15 Generally, these studies have noted that Pb content varies significantly among species
16 and is related to habitat and feeding behavior. There is precedent, especially in Europe,
17 for the inclusion of ALAD as a biomarker of exposure to Pb in marine invertebrates. The
18 mechanism of ALAD inhibition in response to Pb exposure is likely mediated through a
19 common pathway in both marine and freshwater invertebrates (Section 7.4.12.5) as well
20 as in terrestrial species (Section 7.3.12.5) and humans (Section 5.7). Evidence is
21 therefore, suggestive of a causal relationship between Pb exposure and hematological
22 effects in saltwater invertebrates. Evidence is inadequate to conclude that there is a causal
23 relationship between hematological effects and saltwater vertebrates.
7.4.21.6 Physiological Stress-Saltwater Biota
24 Most studies on physiological stress responses in marine invertebrates are laboratory-
25 based exposures where effects are observed at Pb concentrations that exceed those known
26 to occur in seawater [0.01 to 27 (ig Pb/L (Sadiq. 1992). Table 7-21. However, some
27 recent evidence for invertebrate antioxidant response in bivalves and crustaceans
28 indicates effects may occur at Pb concentrations that are detected in the marine
29 environment. For example, SOD, catalase, and glutathione peroxidase activities were
30 significantly reduced in the digestive gland of the marine bivalve C. farreri at 2 (ig Pb/L
31 (as measured in Bohai Bay, China) (Zhang et al.. 2010b). In red fingered marsh crabs,
32 P. erythrodactyla collected from an estuarine lake in Australia, elevated glutathione
November 2012 7-204 Draft - Do Not Cite or Quote
-------
1 peroxidase activity was correlated with individuals with higher metal body burdens
2 (MacFarlane et al.. 2006).
3 Additional evidence from environmental monitoring studies that compared biomarker
4 responses between reference and contaminated sites indicated a correlation between the
5 amount of Pb with changes in antioxidant enzyme activity [e.g., (Serafim etal. 2011;
6 Cravo et al.. 2009)1. Marine bivalves are the organisms typically sampled for these
7 biomonitoring studies since both metals and enzymatic activities can be readily measured
8 in these invertebrates. Although these studies show clear evidence of alterations in
9 antioxidant stress markers in response to marine pollution, these effects cannot be
10 attributed solely to Pb in the environment due to the presence of other metals and
11 contaminants. Evidence for stress responses in marine organisms is typically limited to
12 invertebrates, however, elevated expression of heat shock protein orthologs were reported
13 for the first time in the hypothalamic and mesencephalic brain regions of Pb-treated fish
14 (Giusi et al.. 2008).
15 Evidence for physiological stress responses in saltwater invertebrates are supported by
16 evidence in freshwater species (Section 7.4.12.6) and terrestrial species (Section 7.3.12.6)
17 as well as in humans and experimental animal studies of oxidative stress following
18 impairment of normal metal ion functions (Section 5.2.4). Stress responses may increase
19 susceptibility to other stressors and reduce individual fitness. Evidence is suggestive of a
20 causal relationship between Pb exposures and physiological stress in saltwater
21 invertebrates. The evidence is inadequate to conclude that there is a causal relationship
22 between Pb exposure and physiological stress in saltwater plants and vertebrates.
7.4.21.7 Community and Ecosystem Level Effects-Saltwater Biota
23 No studies on community and ecosystem level effects of Pb in marine systems were
24 reviewed in the 1977 Pb AQCD (U.S. EPA. 1977). or the 1986 Pb AQCD (U.S. EPA.
25 1986a). Observations from field studies reviewed in the 2006 Pb AQCD (U.S. EPA.
26 2006c) included findings of a negative correlation between Pb and species richness and
27 diversity indices of macroinvertebrates associated with estuary sediments (summarized in
28 Table AX7-2.5.2 of the 2006 Pb AQCD). Additional findings in marine environments
29 included changes in species distribution and abundance in fish, crustaceans and
30 macroinvertebrates correlated with Pb levels in marine sediments.
31 New evidence for community and ecosystem level effects of Pb in saltwater ecosystems
32 includes laboratory microcosm studies as well as observations from field-collected
33 sediments, biofilm and plants in which changes in community structure were observed. In
34 a recent study, significant differences in macroinvertebrate communities associated with
November 2012 7-205 Draft - Do Not Cite or Quote
-------
1 seagrass beds were reported between sites with different sediment, biofilm, and leaf Pb
2 concentrations (Marin-Guirao et al.. 2005). Sediment Pb concentrations ranged from
3 approximately 100 to 5,000 mg Pb/kg and corresponding biofilm concentrations were
4 500 to 1,600 mg Pb/kg, with leaf concentrations up to 300 mg Pb/kg. In a laboratory
5 microcosm experiment conducted with estuarine sediments from South Africa, total
6 meiofauna density decreased (range 3 to 5 taxa) after 32 days in Pb-treated (1,886 to
7 6,710 (ig/Pb g sediment dry weight) sediments compared to 9 taxa in the control (3 (ig/Pb
8 g sediment dry weight) (Gvedu-Ababio and Baird. 2006). In a microcosm experiment,
9 exposure to three levels of sediment Pb contamination (322, 1,225, and 1,465 mg Pb/kg
10 dry weight) significantly reduced marine nematode diversity and resulted in profound
11 restructuring of the community structure (Mahmoudi et al.. 2007).
12 There is not sufficient information at this time to characterize and to quantify
13 relationships between ambient concentrations of Pb and response in saltwater
14 communities and ecosystems. Fewer studies are available for saltwater organisms when
15 compared to freshwater systems. There are likely differences in uptake and
16 bioaccumulation in marine species due to physiological characteristics for adaptation in
17 salt water. Additional uncertainties in evaluating the effects of Pb in marine environments
18 include the presence of multiple stressors, inherent natural variability, and differences in
19 Pb bioavailability across saltwater ecosystems. Evidence is inadequate to establish if
20 there is a causal relationship between Pb exposures and the alteration of species richness,
21 species composition and biodiversity in saltwater ecosystems.
7.5 Causal Determinations for Ecological Effects of Pb
22 This section summarizes the key conclusions regarding causality for welfare effects of
23 Pb. Causal determinations for reproductive, growth, survival, neurobehavioral,
24 hematological and physiological stress endpoints are presented separately for terrestrial,
25 freshwater and saltwater organisms (Sections 7.3.12, 7.4.12. and 7.4.21). In Section 2.7.3.
26 causal determinations for the same endpoints are further integrated across terrestrial,
27 freshwater and saltwater taxa. Evidence considered in establishing causality was drawn
28 from findings presented in the 1977 (U.S. EPA. 1977). 1986 (U.S. EPA. 1986a) and
29 2006 Pb AQCDs (U.S. EPA. 2006c). integrated with an exhaustive review of more recent
30 evidence. The causal statements for terrestrial, freshwater and saltwater effects are
31 divided into two categories: (1) endpoints that are commonly used in ecological risk
32 assessment (reproduction, growth and survival) because they clearly can lead to
33 population-level (e.g., abundance, production, extirpation), community-level (taxa
34 richness, relative abundance) and ecosystem-level effects (Anklev etal.. 2010; Suter et
35 al.. 2005). and (2) organism and sub-organism responses such as neurobehavioral effects,
November 2012 7-206 Draft - Do Not Cite or Quote
-------
1 hematological effects and physiological stress. There are many different effects at the
2 molecular and cellular levels, and chronic toxicity of Pb in ecosystems is thus likely
3 attained through multiple modes of action. Furthermore, the effects of Pb on ecosystems
4 necessarily begin with some initial effects at the molecular level of specific organisms
5 within the ecosystem (U.S. EPA. 1986b).
6 Experimental settings for studies used in making causal determinations for the ecological
7 effects of Pb include controlled exposures in the laboratory, microcosm experiments and
8 field observations. Controlled exposure studies in laboratory or small-to medium-scale
9 field settings provide the most direct evidence for causality, but their scope of inference
10 may be limited. In contrast, microcosms and field studies where exposure is not
11 controlled include potentially confounding factors (e.g., other metals) or factors known to
12 interact with exposure (e.g., pH), thus increasing the uncertainty in associating effects
13 with exposure to Pb specifically. A large majority of the available studies of Pb
14 exposures are laboratory toxicity tests on single species, in which an organism is exposed
15 to a known concentration of Pb and the effect on a specific endpoint is evaluated. These
16 studies provide evidence for a temporal sequence between Pb exposure and an effect, an
17 aspect important in judging causality (Table I Preamble). As detailed in the Framework
18 for Causality (see Preamble), coherence between different types of studies also provides
19 strong support to a determination of causality. Evidence from laboratory studies
20 conducted under controlled conditions provides the largest amount of information used in
21 the causal determinations summarized in Table 7-3. but their coherence with microcosm
22 and field-based studies plays an important role in those determinations. Biological
23 gradients (Table I Preamble) are often found in studies of the effects of Pb, and add
24 support to causality where present. For some ecological endpoints, support for causal
25 determinations is additionally supported by toxicological findings reviewed in the
26 chapters of the ISA that evaluate evidence for human health effects associated with Pb
27 exposure, particularly when a common mode of action is documented.
28 The amount of Pb in ecosystems is a result of a number of inputs and it is not currently
29 possible to determine the contribution of atmospherically-derived Pb from total Pb in
30 terrestrial, freshwater or saltwater systems. The causal determinations are, therefore, not
31 specific to Pb from atmospheric deposition since atmospherically-derived Pb may
32 ultimately be present in water, sediments, soils and biota (Section 7.2 and Figure 7-1).
33 The causal determinations encompass findings of studies at concentrations of Pb reported
34 from environmental media (Table 7-2). and up to one to two orders of magnitude above
35 the range of these values (Preamble Table II). Studies at the upper range of Pb
36 concentrations are generally conducted at and near heavily exposed sites such as mining
37 and metal industries-disturbed areas. Studies at those higher concentrations were used
38 only when they were part of a range of concentrations that also included more typical
November 2012 7-207 Draft - Do Not Cite or Quote
-------
1 values, or when they informed understanding of modes of action and illustrated the wide
2 range of sensitivity to Pb across taxa.
3 The exposure values at which Pb elicits a specific effect in terrestrial and aquatic systems
4 are difficult to establish, due the influence of other environmental variables on Pb
5 bioavailability and toxicity and to substantial differences among biological species in
6 their sensitivity to Pb. In the 1977 Pb AQCD (U.S. EPA. 1977). no correlation could be
7 established between toxic effects in invertebrates, fish, birds or small mammals and
8 environmental concentrations of Pb. At the time of the 1986 Pb AQCD additional data
9 were available on toxicity but there was still little information on the exposure values that
10 can cause toxic effects in small mammals or birds (U.S. EPA. 1986b). In the
11 2006 Pb AQCD (U.S. EPA, 2006c) several studies on effects of Pb exposure on natural
12 ecosystem structure and function advanced the characterization of Pb levels in the
13 environment that occur near contaminated sites (i.e., smelters, mining, industry).
14 According to the 2006 Pb AQCD, natural terrestrial ecosystems near significant Pb
15 sources exhibited a number of ecosystem-level effects, including decreased species
16 diversity, changes in floral and faunal community composition, and decreasing vigor of
17 terrestrial vegetation. These findings were summarized in Table AX7-2.5.2 of the Annex
18 to the 2006 Pb AQCD (U.S. EPA. 2006c). The 2006 Pb AQCD concluded that, in
19 general, there was insufficient information available for single materials in controlled
20 studies to permit evaluation on higher levels of biological organization (beyond the
21 organism). Furthermore, Pb rarely occurs as a sole contaminant in natural systems
22 making the effects of Pb difficult to ascertain. Recent information available since the
23 2006 Pb AQCD, includes additional field studies in both terrestrial and aquatic
24 ecosystems, but the connection between air concentration and ecosystem exposure
25 continues to be poorly characterized for Pb and the contribution of atmospheric Pb to
26 specific sites is not clear.
November 2012 7-208 Draft - Do Not Cite or Quote
-------
Table 7-3 Summary of Pb causal determinations for plants, invertebrates and
vertebrates.
Level
Effect
Terrestrial3 Freshwater3 Saltwater3
Community
and
Ecosystem
1 Population-Level Endpoints
Sub-organismal Organism-Level Responses
Responses
Community and Ecosystem Effects
Reproductive and Developmental Effects-Plants
Reproductive and Developmental Effects-
Invertebrates
Reproductive and Developmental Effects-
Vertebrates
Growth-Plants
Growth-Invertebrates
Growth-Vertebrates
Survival-Plants
Survival- Invertebrates
Survival- Vertebrates
Neurobehavioral Effects-Invertebrates
Neurobehavioral Effects- Vertebrates
Hematological Effects-Invertebrates
Hematological Effects-Vertebrates
Physiological Stress-Plants
Physiological Stress-Invertebrates
Physiological Stress-Vertebrates
Likely Causal
Inadequate
Causal
Causal
Causal
Likely Causal
Inadequate
Inadequate
Causal
Likely Causal
Likely Causal
Likely Causal
Inadequate
Causal
Causal
Likely Causal
Likely Causal
Likely Causal
Inadequate
Causal
Causal
Likely Causal
Causal
Inadequate
Inadequate
Causal
Causal
Likely Causal
Likely Causal
Likely Causal
Causal
Likely Causal
Likely Causal
Likely Causal
Inadequate
Inadequate
Suggestive
Inadequate
Inadequate
Inadequate
Inadequate
Inadequate
Inadequate
Inadequate
Inadequate
Inadequate
Suggestive
Inadequate
Inadequate
Suggestive
Inadequate
aBased on the weight of evidence for causal determination in Table II of the ISA Preamble. Ecological causal
determinations are based on doses or exposures generally within one to two orders of magnitude of the range of Pb
currently measured in the environment (Table 7-2).
November 2012
7-209
Draft - Do Not Cite or Quote
-------
7.6 Supplemental Material
Table 7-4
Species
Terrestrial plants, invertebrates and vertebrates; growth, reproduction and survival.
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
Exposure
Method
Modifying
factors
Reference3
(Published since
Effect the 2006 Pb
Effects on Endpoint Concentration AQCD)
Plants
Buckwheat
(Fagopyrum
esculentum)
Canola
(Brassica
napus)
Contaminated
soil:
HCI extractable:
6,643 mg Pb/kg
Acetate
extractable:
832 mg Pb/kg
Water leachate:
0.679 mg Pb/kg
Control soil:
HCI extractable:
5 mg Pb/kg
Acetate
extractable: ND
Water leachate:
ND
0; 22; 45; and 67
mg Pb/kg
Plants were
grown for 8
weeks in
contaminated soil
collected from a
shooting range,
and control soil.
Plants of four
cultivars were
grown for 40
days in soil
amended with
Pb chloride.
Contaminated soil
Sand: 62.3%
Silt: 36.7%
Clay: 1 .0%
pH:6.0
CECM3.0
Control soil
Sand: 87.7%
Silt: 12.3%
Clay: ND
pH: 6.3
CEC:7.6
Growth: Tamura et al.
No effect on growth (2005)
Survival:
No effect on survival
Growth: Ashraf et al. (201 1 )
Shoot and root dry weight
decreased with increasing
Pb
Zn, Cu, Fe, Mn content
decreased with increasing
Pb.
N, P, K, and Ca^+content
decreased to a lesser
degree.
November 2012
7-210
Draft - Do Not Cite or Quote
-------
Species
Chinese
cabbage
(Brassica
pekinensis)
Corn (Zea
mays)
Grass pea
(Lathyrus
sativus)
Exposure Exposure
Concentration Concentration Exposure Modifying
(Nominal) (Measured) Method factors
46; 874; 1 ,703 Plants were
mg Pb/kg dry soil grown for 12
days in soil
amended with
Pb acetate.
0; 0.007; 0.7; 7 Seeds were
mg Pb/L germinated on
paper soaked in
P-sulfate.
Plants were
grown for 21
days in washed
sand with
Pb-sulfate
amended nutrient
solution.
16; 31; 63; 125; Plants were
188mgPb/L grown in soil
amended with
Pb nitrate.
Effect
Effects on Endpoint Concentration
Growth:
Shoot biomass decreased
with increasing Pb (91% and
84% of lowest exposure).
Reproduction:
Germination%, germination
index, plant decreased with
increasing Pb.
Growth:
Shoot length, plant dry
weight, water use efficiency
decreased with increasing
Pb.
Reproduction:
Germination decreased with
increasing Pb (control 100%,
highest exposure 30%).
Chromosomal abnormalities
increased with increasing Pb
(control 0%, highest
exposure 72%).
Growth:
Shoot length decreased with
increasing Pb (highest
exposure was 50% of
control).
Reference3
(Published since
the 2006 Pb
AQCD)
Xiong et al. (2006)
Ahmad et al.
(201 1 )
Kumar and Tripathi
(2008)
November 2012
7-211
Draft - Do Not Cite or Quote
-------
Species
Lettuce
(Lactuca
sativa)
Mustard
(Brassica
juncea)
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
2,000 mg Pb/kg,
but soil was
mixed with 50%
V/V vermiculite
following
amendment with
Pb nitrate.
Tissue Pb:
3.22-233
n-l« DK/U«
mg ru/kg
0;31;62; 124;
186; 249; 311
mg Pb/L
Exposure
Method
Plants were
grown for 40
days in 21 soils
with varying
native CEC, OC,
pH, and
amorphous Fe
and Al oxides,
which were then
all amended with
Pb nitrate and
mixed with 50%
V/V vermiculite.
Plants were
grown for 60
days in field soil
amended with
Pb acetate.
Modifying
factors
After amendment:
pH: 3.8-7.8
CEC:
3.01-32.04cmolc/kg
OC:
5 - 30 g/kg
Fe/AI oxides:
0.009-0.1 95 mol/kg
Effect
Effects on Endpoint Concentration
Reproduction:
Germination 50 - 92%
Germination was greater in
amended soils.
Growth:
2.5-88.5% of control
In the presence of the same
amount of Pb(NO3)2, OC was
the main determinant of
effects, although CEC had a
strong influence, but
mediated by its effect on pH
and Fe/AI oxides
Growth:
Root and shoot length
decreased with increasing
Pb, and the decrease was
greater with time.
Reference3
(Published since
the 2006 Pb
AQCD)
Dayton et al.
(2006)
John et al. (2009)
After 60 days, roots were two
times longer in controls than
in the highest Pb exposure
shoot length was 75%
greater.
Radish 0; 21; 105
(Raphanus mg Pb/L
sativus)
Plants were
grown for 35
days in sand with
a full nutrient
solution
amended with
Pb nitrate.
Growth:
Leaf area, root volume,
shoot and root dry weight
decreased with
increasing Pb.
(total dry weight at
91 mn Ph/l was 3D% smaller
Gopal et al. (2008)
than control, 52% smaller at
105mg Pb/L).
November 2012
7-212
Draft - Do Not Cite or Quote
-------
Species
Wheat
(Triticum
aestivum)
Lettuce
(Lactuca
sativa)
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
69-9,714
mg Pb/kg
Exposure Modifying
Method factors
Wheat plants pH: 4.25-7.26
were grown for 6 oc. 6 2.47 Q%
weeks in
undisturbed core
samples from
four locations in
each two
Pb-contaminated
sites
Lettuce seeds
were germinated
in the leachate.
Reference3
(Published since
Effect the 2006 Pb
Effects on Endpoint Concentration AQCD)
Growth: Chapman et al.
No effects were found on (201 0)
germination or growth of
either species.
Invertebrates
Cabbage 0.87 mg Pb/L in
aphid watering solution
(Brevicoryne used for plants
brassicae)
Aphids were
reared for several
generations on
radish and
cabbage plants
grown in soil
amended with
Pb nitrate.
Reproduction:
In aphids fed
Pb-contaminated plants,
development time was
longer, and relative fecundity
and rate of population
increase were lower than in
control aphids.
Survival:
Mortality was higher in
exposed aphids, both adults
and offspring.
November 2012
7-213
Draft - Do Not Cite or Quote
-------
Species
Collembolan
(Folsomia
Candida)
Collembolan
(Folsomia
Candida)
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
Approximate
range was 12 mg
Pb/kg soil to
1 5,000 mg Pb/kg
soil;
Pore water
approximately
0.002 Pb/L to
1 ,000 mg Pb/L
(Concentrations
were measured,
but not reported).
0; 100; 200; 400;
800; 1,600; 3,200
mg Pb/kg dry soil
Exposure
Method
Springtails were
reared for 28
days in soil
collected at
seven locations
along each of
three transects
with increasing
Pb concentration
s within each
transect
(21 locations).
Lowest
concentration
soils from each of
the three
transects were
then amended
with Pb nitrate to
match the
gradient, and one
set of the
amended
samples were
then leached, for
a total of
57 concentration
sofPb.
Springtails were
reared for 1 0
days in field soil
amended with
Pb chloride.
Modifying
factors
pH was constant in
transects, but
decreased with
increasing addition
of Pb(NO3)2 in both
amended and
amended-and-
leached soils.
pH decreased by 3
units in the highest
addition, regardless
of subsequent
leaching.
Effects on Endpoint
Reproduction:
Reproduction decreased by
up to 50% in transect soils
Amended soils
Pb concentrations 2,207 mg
Pb/kg or lower never had a
significant effect on
reproduction.
Reproduction:
Hatching success decreased
with increasing Pb.
Effect
Concentration
Transect A 28 day
EC50 in mg Pb/kg dry
weight:
native: >5,690
amended: 2,570
amended and
leached: 2,060
Transect B 28 day
EC50 in mg Pb/kg dry
weight:
native: > 14, 400
amended: 3,210
amended and
leached: 2,580
Transect C 28 day
EC50 in mg Pb/kg dry
weight:
native: >5,460
amended: 2,160
amended and
leached: 2,320
EC50 (hatching):
2,361
mg Pb/kg dry soil
Reference3
(Published since
the 2006 Pb
AQCD)
Lock et al. (2006)
Xu et al. (2009b)
November 2012
7-214
Draft - Do Not Cite or Quote
-------
Species
Collembolan
(Sinella
curviseta)
Collembolan
(Paronychiuru
s kimi)
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
0; 100; 200; 400;
800; 1,600; 3,200
mg Pb/kg dry soil
Toxicity run: 100;
500; 1,000; 2,000
mg Pb/kg
Reproduction run:
0; 250; 500;
1,000; 2,000;
3,000 mg/kg Pb
Exposure Modifying
Method factors
Springtails were
reared for 28
days in field soil
amended with
Pb chloride.
Springtails were
reared for 28
days on artificial
soil amended
with Pb chloride
in two separate
runs.
Effects on Endpoint
There was a small effect of
Pb on survival and growth,
and a stronger effect on
reproduction.
Survival:
Survival decreased with
increasing Pb.
Reproduction:
Offspring production and
instantaneous rate of
increase values decreased
with increasing Pb.
Effect
Concentration
Survival:
1 ,838 mg Pb/kg
Reproduction:
642 mg Pb/kg
EC50:
3,21 2 mg/kg Pb
Body Size:
4,094 mg Pb/kg
Survival LC50:
7 day:
1 ,322 mg Pb/kg
28 day:
1 ,299 mg Pb/kg
EC50
28 day: 428 mg Pb
/kg
NOEC
reproduction: EC50
28 day: 428 mg
Pb/kg
NOEC: 250 mg
Pb/kg
LOEC: 500 mg
Pb/kg
Reference3
(Published since
the 2006 Pb
AQCD)
Xu et al. (2009a)
Son et al. (2007)
November 2012
7-215
Draft - Do Not Cite or Quote
-------
Exposure
Concentration
Species (Nominal)
Collembolans 10; 50; 100; 500;
(Sinella coeca, 1 ,000 mg Pb/kg
Folsomia
Candida)
Earthworm
(Eisenia
andrei)
Exposure
Concentration
(Measured)
2,000 mg Pb/kg in
soil
Internal
concentration of
Pb in earthworms
varied among the
amended soils,
between 29 and
782 mg Pb/kg dry
weight.
Exposure
Method
Springtails were
reared for 42 or
45 days in
artificial soil
amended with
Pb nitrate.
Earthworms were
reared for 28
days in 21 soils
with varying
native CEC ,OC,
„[! nn/J
pn, ana
amorphous
Fe and Al oxides,
which were then
all amended with
Pb nitrate.
Modifying
factors
After amendment
pH:
3.8-7.8
CEC:
3.01 - 32.04
cmolc/kg
OC:
5 - 30 g/kg
Fe/AI oxides:
0.009 -0.1 95 mol/kg
Effects on Endpoint
S. coeca:
Survival:
Mortality significantly
increased with increasing
concentration in adult
population.
Reproduction:
Juvenile production not
significantly compromised at
10-500 mg Pb/kg, reduced at
1 ,000 mg Pb/kg
F. Candida:
Survival:
Increase in mortality with
increasing concentration
Reproduction:
Juvenile production not
significantly reduced
between 10-500 mg Pb/kg,
significant effect at 1 ,000 mg
Pb/kg
Survival:
Mortality ranged between 0
and 100%.
In the presence of potentially
lethal amounts of Pb, the
main determinant of mortality
was pH, with little or no
effect from OC, CEC, or
Fe/AI oxides.
Reproduction:
Reproduction relative to
controls ranged between 0
and 167%.
Effects of Pb on reproduction
are dependent principally on
Fe/AI oxides, with some
influence of CEC
Effect
Concentration
LC50:
S. coeca: Could not
be determined
F. Candida: Could
not be determined
EC50 reproduction:
S. coeca:
490 mg Pb/kg Pb on
dry soil
F. Candida:
Could not be
calculated with
accuracy; Ranged
from 500-1 ,000
mg Pb/kg
Reference3
(Published since
the 2006 Pb
AQCD)
Mentaetal. (2006)
Bradham et al.
(2006)
November 2012
7-216
Draft - Do Not Cite or Quote
-------
Species
Earthworm
(Eisenia fetida)
Earthworm
(Eisenia fetida)
Earthworm
(Eisenia fetida)
Exposure
Concentration
(Nominal)
0; 300; 711;
1 ,687; 2,249
mg Pb/kg
Soil!:
0; 355; 593; 989;
1 ,650 mg Pb/kg
Soil 2:
59; 297; 593;
2,965 mg Pb/kg
Soil 3:
386; 771 ; 1 ,929;
3,857 mg Pb/kg
Exposure
Concentration Exposure
(Measured) Method
18-9,311 Earthworms were
mg Pb/kg reared for 1 4
days in OECD-
standard toxicity
testing soil with
7 concentrations
of Pb, either one
or 10 earthworms
per container.
Mean: 79% of Earthworms were
nominal reared for 14
days in soil
amended with
five levels of
Pb nitrate and
artificially aged.
Earthworms were
reared for 28
days in three
soils amended
with five levels of
Pb nitrate without
aging, after which
they were
removed from the
containers.
Containers were
then kept in the
same conditions
for another 28
days, after which
cocoons were
extracted.
Modifying
factors
pH decreased with
increasing Pb nitrate
addition
NH3 concentration
increased with
Pb concentration
and time
pH 6.72 prior to
amendment
OC 0.7%
CEC 11 meq/100g
PH
6.72; 5.48; 6.75
(prior to
amendment)
OC
0.7; 1.2; 5.2%
CEC
11;8;27meq/100g
Reference3
(Published since
Effect the 2006 Pb
Effects on Endpoint Concentration AQCD)
Survival: LC50 (multiple- Currie et al. (2005)
Mortality increased from 0 to occupancy):
1 00% with increasing Pb, 2 662 mg Pb/kg at
with 1 00% reached at 4,500 7'd a9nd 9
mg Pb/kg after 7 days and
1 4 days. Number of worms 2>589 m9 Pb/k9 at
per container had no effect 14 days or
on mortality. 2,827 mg Pb/kg at
Growth. both 7 and 1 4 days
Worm weight decreased with
increasing Pb, and faster in
multiple-worm containers.
Survival: Jones etal.
Mortality was only observed (2009b)
at the highest exposure.
Reproduction: Jones etal.
Soil!: (200*)
Juvenile and cocoon count
decreased from 19 and 45,
respectively, to near 0 with
increasing Pb.
Soil 2:
Cocoon count decreased to
40% of control at highest Pb.
Soil 3:
Cocoon count was 0 at all
concentrations.
November 2012
7-217
Draft - Do Not Cite or Quote
-------
Species
Earthworm
(Pheretima
guillelmi)
Earthworms
(Eisenia
andrei,
Lumbricus
ru bell us,
Aporrectodea
caliginosa)
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
Toxicity run:
0, 1,000; 1,400;
2,000; 2,800;
3,800; 5,400;
7,500 mg Pb/kg
dry weight (form
of Pb not
reported).
Sublethal toxicity
run:
1,000; 1,400;
1,800; 2,500
mg Pb/kg dry
weight (form of Pb
not reported).
0; 1,000; 3,000;
4,000; 5,000;
7,500; 10,000
mg Pb/kg
Exposure
Method
Earthworms were
reared for 1 4
days in OECD-
standard soil
amended with Pb
(form of Pb
unreported).
There were two
runs with
different
concentrations.
Earthworms were
reared for 28
days in sterilized
Kettering Loam
watered with
Pb nitrate
solution.
Modifying
factors
Temperature:
20 °C (E. andrei);
15°C(L rubellus
and A. caliginosa)
pH
Day 7 : 4.57-5.83,
Day 28: 4.71-5.83,
increasing with
decreasing Pb
Effects on Endpoint
Survival:
Mortality increased with
increasing Pb.(0% in control,
100% at 7,500 mg Pb/kg
after 14 days).
Growth:
Weight decreased with
increasing Pb(2.5 g in
control, 1 .4 g at 5.400 mg
Pb/kg after 14 days).
Reproduction:
Sperm abnormalities
increased with increasing Pb
(10% of control, 21% at
2,500 mg Pb/kg after 14
days).
Growth:
Weight decreased with
increasing concentration and
time, severity of weight
decrease varied with
species.
Survival:
Mortality increased with
increasing concentration and
time, and varied with species
100% mortality for all
species at higher
concentrations after 28 days.
Reference3
(Published since
Effect the 2006 Pb
Concentration AQCD)
Zheng and Li
(2009)
LC50: Langdon et al.
E. andrei: (2005)
5,824 mg Pb/kg
L. rubellus:
2,867 mg Pb/kg
A. caliginosa:
2,747mg Pb/kg
EC50:
E. andrei:
2,841 mg Pb/kg
L. rubellus:
1 ,303 mg Pb/kg
A. caliginosa:
1 ,208 mg Pb/kg
November 2012
7-218
Draft - Do Not Cite or Quote
-------
Exposure
Concentration
Species (Nominal)
Nematode
(Caenorhabditi
s elegans)
Nematode 5; 10; 16; 21
(Caenorhabditi mg Pb/L
s elegans)
Nematode 0.5; 16; 41 mg
(Caenorhabditi Pb/L
s elegans)
Exposure
Concentration Exposure Modifying
(Measured) Method factors
0.5; 10; 21 Nematodesat
mg Pb/L various
developmental
stages were
exposed to
Pb(NO3)2 for four
hours.
Late larval
nematodes (L4)
were exposed for
one or three
days.
Nematodes were
placed for 48
hours in growing
medium with
4 concentrations
ofPb.
Nematodes were
placed for three
days in growth
medium
amended with
Pb nitrate.
Effect
Effects on Endpoint Concentration
Reproduction:
Brood size decreased with
increasing Pb , but the
decrease was smaller with
increasing developmental
age.
Generation time increased
with increasing Pb, and the
increase was smaller with
increasing developmental
age.
These effects were greater in
late larval nematodes when
exposure duration increased
from four hours to one and
three days.
Survival:
No effect
Growth:
Life span, body size
decreased with increasing
Pb.
Reproduction:
Generation time and brood
size increased with
increasing Pb.
All effects were present and
of comparable magnitude in
progeny of exposed
nematodes.
Reference3
(Published since
the 2006 Pb
AQCD)
Guo et al. (2009)
Vigneshkumar et
al. (In Press)
Wang and Peng
(2007)
November 2012
7-219
Draft - Do Not Cite or Quote
-------
Species
Snail
(Achatina
achatina)
Exposure
Concentration
(Nominal)
Exposure
Concentration
(Measured)
1.33;
70.98;
134.61;
339.40;
674. 86;
1,009.22;
1 ,344.39 mg
Pb/kg
Exposure Modifying
Method factors
Snails were
reared for 1 2
weeks on a diet
amended with
Pb chloride.
Effect
Effects on Endpoint Concentration
Survival:
no effect
Growth:
Small decrease in feeding at
highest exposure, small
decrease in weight gain with
Reference3
(Published since
the 2006 Pb
AQCD)
Ebenso and
Ologhobo (2009a)
increasing Pb (over 12
weeks, snails in the highest
exposure gained 12% less
weight than in the lowest
exposure).
Snail
(Achatina
achatina)
Snail (Theba 0; 50; 100; 500;
pisana) 1 ,000; 5,000;
10,000; 15,000
mg Pb/Kg
Snails
(Cantareus
aspersus,
Helix aspersa)
0.56;
20.37;
200.42;
1,200.30
mg Pb/Kg
Total Soil Pb:
1 740-2060 mg
Pb/kg
CaCI2extractable;
4-80 mg Pb/kg
Dissolved
(estimated):
0.007-0.09 mg
Pb/L
Snails from
laboratory source
were reared for
12 weeks in
bottomless
enclosures at
four locations
within the
grounds of an
abandoned
battery factory.
Snails were
reared for 5
weeks on
Pb-amended
diet.
Snails were
reared for 7 - 9
weeks in field soil
amended with
varying amounts
of Pb-sulfate,
clay, peat, and
CaCO3.
pH: 4.42 -6.29,
decreasing with
increasing Pb
OC: 1.39-3.45%,
decreasing with
increasing Pb
CEC: 3.32 -5.37
cmol/kg, increasing
with increasing Pb.
Clay content
11 - 1 6%
Organic matter
1.2-1 0%
pH
AC ~7 AC,
4.D - / .4y
Growth:
Feeding, weight gain and
shell thickness all decreased
with increasing Pb (13; 17;
and 19% lower in highest
exposure than in lowest).
Growth:
Feeding and weight gain
decreased with increasing
Pb and time (snails in 0
added Pb gained 45% more
weight than in highest Pb).
Survival:
No effect
Growth:
No effect
Survival:
No effect
Ebenso and
Ologhobo (2009b)
EI-Gendy et al.
(201 1 )
Pauget et al.
(201 1 )
November 2012
7-220
Draft - Do Not Cite or Quote
-------
Exposure
Concentration
Species (Nominal)
Exposure
Concentration
(Measured)
Exposure Modifying
Method factors
Reference3
(Published since
Effect the 2006 Pb
Effects on Endpoint Concentration AQCD)
Vertebrates
Japanese
quail (Coturnix
coturnix
japonica)
Pied
flycatchers
(Ficedula
hypoleuca)
Drinking water
Pb:
0; 5; 50 mg Pb/L
Tissue Pb:
0;1.1; 10.7
mg/kg wet weight
Blood Pb in
nestlings at
mining site
while active:
41 mg Pb/100kg
wet weight;
after closing:
29 mg Pb/100kg
wet weight.
Blood Pb in
nestlings at
reference site
while active:
2 mg Pb/1 00 kg
wet weight;
after closing:
0.4 mg Pb/1 00 kg
wet weight.
Quails were
given
Pb-amended
water for 7
weeks.
Data were
collected in wild
flycatchers near
a Pb mine and at
a reference site
for three years
while the mine
was active, and
for three years
five years after
mine closing.
Growth: Main and Smits
Feed intake and growth rate (201 1 )
not affected.
Survival:
Morbidity/mortality was lower
in highest exposure than in
control.
Incidence of pericarditis,
airsacculitis, perihepatitis,
and arthritis was lower in
highest exposure than in
control.
Reproduction: Berglund et al.
Clutch size and breeding (201 0)
success were lower at the
mine site, but did not change
after closure of the mine
(clutch size
5.6 reference,
4.9 mining site;
breeding success
80% reference site,
76% mining site).
Nestling mortality was higher
at the mine site , and
increased after closure
(5% reference site,
1 1 % mining site while active;
1 1 % reference site,
26% mining site after
closure).
November 2012
7-221
Draft - Do Not Cite or Quote
-------
Species
Pig (Sus
domestica)
Exposure Exposure
Concentration Concentration
(Nominal) (Measured)
Feed Pb
control:
unreported
exposed:
1 0 mg Pb/kg
Blood Pb
control:
1 .44 |jg/dL
exposed:
2.08 |jg/dL
Exposure Modifying
Method factors
Pigs were reared
for 120 days with
Pb-sulfate-
amended feed.
Effect
Effects on Endpoint Concentration
Growth:
Significant decrease in body
weight, average day gain,
average day feed intake, and
feed efficiency.
Increase in feed conversion
ratio.
Reproduction:
No effect on ovary and
uterus weight
Reference3
(Published since
the 2006 Pb
AQCD)
Yu et al. (2005)
References included are those which were published since the 2006 Pb AQCD.
November 2012
7-222
Draft - Do Not Cite or Quote
-------
Table 7-5 Freshwater plants, invertebrates and vertebrates; growth, reproduction and survival.
Species
Concentration
Exposure Method
Modifying
factors
Effects on Endpoint
Effect Concentration
Reference3
(Published
since the
2006 Pb
AQCD)
Algae/Plants
Blue-green algae
(Spirulina
(Arthrospira)
platensis)
Microalgae
(Scenedesmus
obliquus)
Microalgae
(Chlorella vulgaris)
Duckweed
(Lemna minor)
Duckweed
(Lemna minor)
5,000; 10,000;
30,000; 50,000;
and
1 00,000 ug Pb/L
(nominal
concentration at
day 0 and then Pb
in media was
measured every
two days
thereafter).
5,000to
300,000 ug Pb/L
(nominal)
100; 200; 490;
900; 2,000; 5,020;
7,990; and
9,970 ug Pb/L
(measured)
2,070; 10,360;
20,700; and
1 03,600 ug Pb/L
(nominal)
10-day exposure to
Pb nitrate in Zarrouk
liquid medium.
48 and 96-hour acute
toxicity test with
Pb nitrate in BG11
medium.
4 day or 7-day
exposures to
Pb chloride in static
test conditions with
Jacob culture medium
under continuous
illumination.
9-day exposures to
Pb nitrate in a growth
chamber on Knopp's
medium under a 14-
hour photoperiod.
Temperature:
25 ± 1 °C
pH:
7.0
Light dark cycle of
14:10 hours.
Plants were
incubated at
normal room
temperature
(not provided)
Temperature:
25 ± 2 °C
PH
6.0
Growth:
10-day algal growth (measured
turbidimetrically at 560 nm) was
stimulated by 3.7% in the lowest
concentration, growth was inhibited
at higher concentrations of 30,000;
50,000; and 100,000 ug Pb/L by 40;
49; and 78%, respectively.
Chlorophyll a and b content were
significantly diminished at the three
highest exposures.
Growth:
In growth studies (measured as cell
division rate) S. obliquus was
significantly more sensitive to Pb
exposure than C. vulgaris
Growth:
Growth (measured as biomass) of
the duckweed was promoted up to
103% at 100 ug Pb/L and
200 ug Pb/L. However, growth was
inhibited monotonically at all other
test levels with increasing
concentrations. Overall, the relative
growth rate was reduced to 37-38%
at the highest concentration.
Growth: At lower Pb doses, growth
was slightly stimulated. Fresh weight
was lower by 65% at the highest
dose. Pb-induced chlorosis occurred
and the enzymes of the antioxidative
system were modified due to Pb
exposure in all concentrations.
LC50:
75,340 ug/L
48 hour EC50:
4,040 ug Pb/L
S. obliquus
48 hour EC50:
24,500 ug/L
C. vulgaris
4 day EC50:
6,800 ug Pb/L
7 day EC50:
5,500 ug/L
Arunakumara
et al. (2008)
Atici et al.
(2008)
Dirilgen (2011)
Paczkowska
et al. (2007)
November 2012
7-223
Draft - Do Not Cite or Quote
-------
Species
Duckweed
(Woffia arrhiza)
Waterweed
(Elodea canadensis)
Wetland plants
(Beckmannia
syzigachne,
Alternanthera
philoxeroides,
Juncus effusus,
Oenanthe
javanica,
Cyperus
flabelliformis,
Cvoerus
\SJ/ftl,l UVJ
malaccensis,
Polypogon fugax,
Leersia hexandra,
Panicum
paludosum,
Neyraudia
reynaudiana)
Concentration
210; 2,120;
20,720; and
207,200 ug Pb/L
(analytically
verified)
1,000; 10,000;
and 100,000 ug/L
(nominal)
20,000 ug/L
(nominal)
Metals were
analyzed in plant
samples.
Exposure Method
14-day exposure to Pb
as Pb nitrate in sterile
1/50 dilution of
Hutner's medium, day
to night cycle of 16:8
hours.
Plants were exposed 5
days to Pb as
Pb acetate in a 10%
nutrient solution and
then assayed for
pigment content, total
ascorbic acid, and
protein content.
Field-collected tillers or
seedlings (from various
locations in China) for
each species were
used in 21 -day
experiments to
determine Pb tolerance
as inferred from
measuring the
elongation of the
longest root in a
hydroponic system in a
Pb nitrate solution.
Modifying
factors
Temperature:
25 ± 0.5 °C
pH'
r
7.0
Temperature:
25-27°C
pH
6.5-6.7
10% Hoagland &
Arnon nutrient
solution
Reference3
(Published
since the
2006 Pb
Effects on Endpoint Effect Concentration AQCD)
Growth: Piotrowska et
Biomass decreased proportionally al. (2010)
with increasing Pb concentration;
Chlorophyll a content was
significantly inhibited at 210 ug Pb/L
and greater; carotenoid,
monosaccharide and protein content
significantly decreased in higher
concentrations.
Growth: Dogan et al.
The chlorophyll, carotenoid, and (2009)
protein contents of £. canadensis
were significantly reduced following
Pb accumulation.
Growth: Deng et al.
Root elongation was significantly (2009)
reduced in a number of wetland
species (6. syzigachne, J. effusus,
O. javanica, C. flabelliformis,
C. malaccensis, and
N. reynaudiana). Metal tolerance was
related to root anatomy and spatial
pattern of radical oxygen loss.
November 2012
7-224
Draft - Do Not Cite or Quote
-------
Species
Concentration
Exposure Method
Modifying
factors
Effects on Endpoint
Effect Concentration
Reference3
(Published
since the
2006 Pb
AQCD)
Invertebrates
Rotifer
(Brachionus
calyciflorus)
Rotifer
(Brachionus patulus)
Rotifer
(Euchlanis dilatata)
67; 194; 284; 390;
and 700 |jg Pb/L
(measured)
1,250; 2,500;
4,000; 5,000; and
8.000 ug Pb/L
(nominal) for
acute toxicity
tests.
Chronic
exposures used
nominal
concentration of
60 and
600 ug Pb/L with
varying turbidity
levels.
0.1; 0.5; 50; 100;
250; 1 ,000;
2,500 ug Pb/L
(analytically
verified, actual
concentrations not
reported)
Cysts of rotifers were
obtained from Florida
Aqua farms in Dade
City, Florida, U.S.
Tests with Pb nitrate
were performed in total
darkness for 48 hours.
24-hour exposures to
Pb chloride in the
presence and absence
of sediments using
rotifers originally
isolated from the
Chimaliapan wetland,
Toluca, Mexico.
Three week chronic
toxicity tests were also
conducted.
48-hour acute toxicity
tests with rotifer
neonates exposed to
Pb nitrate in synthetic
moderately hard water.
Adult rotifers were
collected in a reservoir
in Aguascalientes,
Mexico.
Temperature:
25 ± 1 °C
pH:
8.19
Temperature:
20 °C
Temperature:
25 ± 2 °C
pH:
7.5
Hardness:
80-100mg/L
/>_/>PI
oauUs
Reproduction:
The total number of rotifers and the
intrinsic rate of population increase
exhibited concentration-dependent
responses at the end of the 48 hour
incubation period.
Reproduction:
In chronic tests, net reproductive rate
and rate of population increase
decreased under conditions of
increasing turbidity and Pb
concentration.
Survival:
24-hour LC50 reported for this
species. In chronic tests, average life
span and life expectancy at birth
decreased under conditions of
increasing turbidity and Pb
concentration.
Survival:
Based on 48-hour LC50 E. dilatata is
among the most sensitive rotifer
species to Pb. E. dilatata may be a
more suitable test organism for
ecotoxicology in Mexico, where this
study was conducted, instead of
D. magna, a species that is not been
found in Mexico reservoirs.
EC2o for number of
rotifers: 125 ug Pb/L
48-hour EC2o for intrinsic
rate of population
increase: 307 ug Pb/L
NOEC:194ugPb/L
LOEC: 284 ug Pb/L
24-hour LC50:
6, 150 ug/L
48-hour
NOEC: 0.1 ug/L
LOEC: 0.5 ug/L
LC 50: 35 ug/L
(estimated from
analytically verified
concentrations)
Grosell et al.
(2QQ6b)
Garcia-Garcia
etal. (2007)
Arias-Almeida
and Rico-
Martinez (2011)
November 2012
7-225
Draft - Do Not Cite or Quote
-------
Species
Cladoceran
(Ceriodaphnia dubia)
Cladoceran
(Ceriodaphnia dubia)
Cladoceran
(Diaphanosoma
birgei)
Cladoceran
(Moina micrura)
Cladoceran
(Nona rectangular)
Concentration
Measured but not
reported.
Predicted
concentration of
major Pb
chemical species
in the natural
water bioassays is
provided in Table
4 of Esbaugh et
al. (2011).
A range of 5 to 6
Pb concentrations
(measured but not
reported) were
prepared with
varying pH,
hardness and
alkalinity.
2,000to
5,500 ug Pb/L
(analytically
verified)
1 ,000 to
8,000 ug Pb/L
(analytically
verified)
3,000to
1 0,000 ug Pb/L
(analytically
verified)
Exposure Method
Acute toxicity of Pb to
C. dubia was assessed
in 48-hour exposures
to two lab generated
reference waters and
eight natural waters
from across North
America selected to
include a range of
water quality
parameters. Waters
were spiked with
varying concentrations
of Pbas Pb nitrate.
Chronic 7-day static
renewal 3 times per
week in 2:1
dechlorinated, aerated
tap waterdeionized
water to determine the
effects of hardness (as
CaSO4 and MgSO4),
alkalinity, pH, and
DOM on Pb toxicity.
24-hour exposure to
Pb chloride in
moderately hard water.
24-hour exposure to
Pb chloride in
moderately hard water
24-hour exposure to
Pb chloride in
moderately hard water
Modifying
factors
Temperature: 26°C
Water chemistry of
the field-collected
waters are reported
in Table 1 of
Esbaugh et al.
(201 1 ) including pH
(range 5.5 to 8.5),
Ca2+ (range 24 to
1,934uM), DOC
(range 36 to 1 ,244
uM) and hardness
(range 4 to 298
mg/L)
Temperature:
25 °C
pH:
6.4-8.2
Hardness:
22-524 mg/L
Temperature:
23 °C
pH:
7.0-7.5
Temperature:
23 °C
pH:
7.0-7.5
Temperature:
23 °C
pH:
7.0-7.5
Effects on Endpoint
Survival:
LC50 values ranged from 29 to
1,180 ug Pb/L. Sensitivity to Pb
varied greatly with water chemistry.
DOC was correlated with protection
from acute toxicity.
Survival:
DOM and alkalinity have a protective
effect against chronic toxicity of Pb.
CaSO4 and MgSO4 do not have a
protective influence of water
hardness; Pb toxicity increased at
elevated Ca2+ and Mg2+. Low pH
increases the toxicity of Pb.
Reproduction:
Increased DOC leads to an increase
in mean EC50 for reproduction
ranging from approximately
25 ug Pb/L to >500 ug Pb/L.
Survival:
LC50 reported
Survival:
M. micrura was more sensitive to Pb
than D. birgei and A. rectangular.
Survival:
A. rectangular was more resistant to
Pb than the other species tested
Effect Concentration
48 -hour LC50 range:
29 to 1,1 80 ug Pb/L
NOEC range: 18 to
<985 ug Pb/L.
LOEC range: 52 to
1 ,039 ug Pb/L
Control base water EC20:
45 ug Pb/L
5.0 mM CaSO4 EC20:
22 ug Pb/L
32 mg/L DOM EC20:
523 ug Pb/L
2.5 mM NaHCO3 EC20:
73 ug Pb/L
Additional EC20 and all
EC50 values were reported
in the study.
24 hour LC50
3,160ug Pb/L
24 hour LC50
690 ug Pb/L
24 hour LC50
7,000 ug Pb/L
Reference3
(Published
since the
2006 Pb
AQCD)
Esbaugh et al.
(2011)
Mageret al.
(2011 a)
Garcia-Garcia
et al. (2006)
Garcia-Garcia
et al. (2006)
Garcia-Garcia
et al. (2006)
November 2012
7-226
Draft - Do Not Cite or Quote
-------
Species
Cladoceran
(Daphnia magna)
Cladoceran
(Daphnia pulex)
Ostracod
(Stenocypris major)
Concentration
Acute test:
Concentrations
not provided
Chronic test:
25 |jg Pb/L
250 |jg Pb/L
2,500 |jg Pb/L
(nominal)
Acute test:
250; 500; 1 ,000;
2,000;
5,000 |jg Pb/L
(nominal)
Chronic test:
250; 500;
1 ,000 |jg Pb/L
(nominal).
475; 1,160; 3,410;
4, 829;
8,972 |jg Pb/L
(measured)
Exposure Method
24 hour acute toxicity
test and 21 day toxicity
test with Pb nitrate,
static renewal every
two days.
48-hour acute toxicity
test and two 21 -day
exposures to Pb nitrate
in dechlorinated tap
water.
96-hour static renewal
with Pb nitrate in
dechlorinated tap
water. Ostracods were
collected from a filter
system of a fish pond
in Bangi, Selangor,
Malaysia.
Modifying
factors
Temperature:
20 ± 1 °C
Temperature:
28-30 °C
pH:
6.5 ±0.01
Conductivity:
244.3 ± 0.6 uS/cm
DO: 6.3 ± 0.06
mg/L
Total hardness:
15.6 mg/L as
CaCO3
Light dark cycle of
12:12 hours.
Effects on Endpoint
Reproduction:
Significant concentration-dependent
decrease in number of neonates per
female; Significant long-term effects
on reproduction. Negative correlation
between hemoglobin gene
expression and reproduction
outcomes.
Reproduction:
Reproduction rates (cumulative
neonates) significantly decreased at
1 ,000 ug Pb/L in the first chronic
toxicity test and at 500 ug Pb/L in a
second test.
Survival:
LC50 values reported
Survival:
LC50 values reported
Reference3
(Published
since the
2006 Pb
Effect Concentration AQCD)
24-hour EC50 (immobility): Ha and Choi
1 8,1 53 ug Pb/L (2009)
48-hour LC50: Theegala et al.
4,000 ug Pb/L (2QQ7J
24-hour LC50: Shuhaimi-
6 583 ua Pb/L Othman et al.
,ou ,„ (201 1b)
48-hour LC50:
2,886 ug Pb/L
72-hour LC50:
1,491 ug Pb/L
96-hour LC50:
526 ug Pb/L
November 2012
7-227
Draft - Do Not Cite or Quote
-------
Species
Midge
(Chironomus dilutus,
formerly C. tentans)
Midge
(Chironomus
riparius)
Concentration Exposure Method
29; 57; 75; 1 1 5; 96-hour static renewal
128; 152 ug Pb/L test
(measured) 20 day midge life cycle
test in Pb chloride
spiked water, flow
through, and
emergence at 55 days.
The C. dilutus culture
was initially started with
egg cases from Aquatic
Biosystems, Fort
Collins, CO, USA.
Logarithmic range 24-hour acute toxicity
from 0 to tests with first-instar
25,000 ug Pb/L larvae exposed to
(nominal) Pb nitrate in synthetic
soft water. C. riparius
culture was from egg
masses from
Environment Canada.
Modifying
factors
Average ± SD
(range):
Temperature:
22.2 ± 1.0 °C
(1 9.7-24.4 °C)
pH:
7.26 ±0.21
(6.9 - 7.7)
Hardness:
32 ± 3.2 mg/L
as CaCO3
Alkalinity:
31 ± 3.0 mg/L
as CaCO3
Conductivity:
76 ± 4.9 us
DO:
7.8 ± 0.8 mg/L
Temperature:
20 °C
Water hardness:
8 mg/Lof CaCO3,
Effects on Endpoint
Growth:
Growth and emergence decreased
as concentration increased.
Reproduction:
No effect
Survival:
No effect
Survival:
Concentration-dependent decrease
in survival with increasing Pb.
Of the five metals tested in the study
(Cd, Cu, Pb, Ni, Zn), Pb was most
toxic to first instar C. riparius.
Effect Concentration
96-hour LC50:
3,323 ug Pb/L
Survival
NOEC: 152 ug Pb/L
LOEC: >152 ug Pb/L
MATC: >1 52 ug Pb/L
Weight
NOEC: 57 ug Pb/L
LOEC: 75 ug Pb/L
MATC: 65 ug Pb/L
EC10(95%):15ugPb/L
EC20(95%):28ugPb/L
Fecundity
NOEC: 152 ug Pb/L
LOEC: >1 52 ug Pb/L
MATC: >1 52 ug Pb/L
EC10(95%):>152ugPb/L
EC20(95%): >152ug Pb/L
Emergence
NOEC: 1 15 ug Pb/L
LOEC: 128ugPb/L
MATC: 121 ug Pb/L
EC10: 28 ug Pb/L
EC20: 55 ug Pb/L
24-hour LC 50:
613 ug Pb/L
Reference3
(Published
since the
2006 Pb
AQCD)
Mebane et al.
(2008)
Bechard et al.
(2008)
November 2012
7-228
Draft - Do Not Cite or Quote
-------
Species
Midge
(Chironomus
javanus)
Midge
(Culicoides furens)
Concentration
430; 580,; 1,330;
2,460;
7,670 |jg Pb/L
(measured)
100; 290; 510;
800;
2,800 ug Pb/L
(measured)
Exposure Method
4-day exposure with
fourth instar larvae to
Pb nitrate in aerated,
filtered, dechlorinated
tap water with static
aerated renewal at
2 days. Larvae were
collected from a filter
system of a fish pond
in Bangi, Selangor,
Malaysia.
Series of 96-hour
exposures to
Pb chloride in
dechlorinated water
under several
temperature ranges.
Modifying
factors
Temperature:
28-30 °C
pH:
6.51 ± 0.01
Conductivity:
244.3 ± 0.6 uS/cm
DO:
6.25 ± 0.06 mg/L
Total hardness
(Mg2+ and Ca2+):
15.63 ±2.74 mg/L
as CaCO3
1st experiment:
Temperature:
25-28 °C
2nd experiment:
Temperature:
20-26 °C
3rd experiment:
Temperature:
10, 15,20,23,25,
28, 30, 35,
40 ± 0.5 °C
Effects on Endpoint
Survival:
LC50 values reported for this species.
Survival:
Higher and lower temperatures
brought about increased toxicities.
LC50 values generally increased in
10-25°C and decreased in 28-40°C.
40°C temperature produced 100%
mortality.
Effect Concentration
24 hour LC50:
20,490 ug Pb/L
48 hour LC50:
6,530 ug Pb/L
72-hour LC50:
1 ,690 ug Pb/L
96 hour LC50:
790 ug Pb/L
96 hour LC50 values:
28-25 °C*: 400 ug Pb/L
26-20 °C*: 300 ug Pb/L
25 °C: 400 ug Pb/L
35 °C-25 °C*: 500 ug Pb/L
23 °C: 700 ug Pb/L
20 °C: 400 ug Pb/L
15 °C: 400 ug Pb/L
10 °C: 357 ug Pb/L
'temperature decreased
over the duration of the
experiment
Reference3
(Published
since the
2006 Pb
AQCD)
Shuhaimi-
Othman et al.
(2011c)
Vedamanikam
and Shazilla.
(2008a)
November 2012
7-229
Draft - Do Not Cite or Quote
-------
Species
Midge
(Chironomus
plumosus)
Oligochaete worm
(Lumbriculus
variegatus)
Mayfly
(Baetis tricaudatus)
Concentration
3,000; 5,400;
8,200; 30,000;
54,000 |jg Pb/L
(measured)
1,300; 3,200;
8,000; 20,000;
50,000 ug Pb/L
(nominal)
69; 103; 160; 222;
350; 546 ug Pb/L
(measured)
Exposure Method
Series of 96-hour
exposures to
Pb chloride in
dechlorinated water
under several
temperature ranges.
24 and 48-hour
exposures to Pb nitrate
spiked water from
Lake Vesijarvi, Finland
96-hour static renewal
test and 10 day chronic
study with aerated
Pb chloride spiked
water, static renewal
every 48 hours.
Mayflies were collected
from the South Fork
Coeurd'Alene River,
Idaho.
Modifying
factors
1st experiment:
Temperature:
25-28°C
2nd experiment:
20-26°C
3rd experiment:
10, 15, 20, 23, 25,
28, 30, 35,
40 ± 0.5 °C
Temperature:
20 °C
Mean ± SD
Temperature:
9.3 ± 0.67 °C
nU-l-
pH.
6.64 ±0.18
Hardness:
20.7 ± 0.58 mg/L
as CaCOS
Alkalinity:
19.8± 1.04 mg/L
as CaCOS
Conductivity:
47.7 ± 1 .72 us
DO:
10.1 ±0.45 mg/L
Effects on Endpoint
Survival:
Higher and lower temperatures
brought about increased toxicities
40°C temperature produced 100%
mortality
LC50 values generally increased in
10-25°C and decreased in 28-40°C
Survival:
48-hour LC50 reported
Growth:
Consistent dose-dependent
reductions in mayfly growth (as
number of molts); growth decreased
with increased Pb exposure.
Survival:
96-hour EC50 reported for this
species. Reduced molting endpoint
more sensitive than mortality
endpoint.
Reference3
(Published
since the
2006 Pb
Effect Concentration AQCD)
96 hour LC50 values: Vedamanikam
28-25°C*' 16 200 ua Pb/L and Snazilla
' ' (2008a)
26-20°C*: 8,300 ug Pb/L
25 °C: 9,500 ug Pb /L
35 °C: 700 ug Pb/L
30 °C: 700 ug Pb/L
28 °C: 900 ug Pb/L
25 °C: 900 ug Pb/L
23 °C: 700 ug Pb/L
20 °C: 600 ug Pb/L
15 °C: 600 ug Pb/L
10 °C: 500 ug Pb/L
'temperature decreased
over the duration of the
experiment
48-hour LC50: Penttinen et al.
5,200 ug Pb/L (2008)
96-hour EC50 Mebane et al.
664 ug Pb/L (2008)
Survival:
NOEC: 222 ug Pb/L
LOEC: 350 ug Pb/L
MATC: 279 ug Pb/L
EC1() (95%): 169ugPb/L
EC20(95%):23ugPb/L
Molting :
NOEC: 103ugPb/L
LOEC: 160ugPb/L
MATC: 130ug Pb/L
EC10(95%):37ugPb/L
EC20(95%):66 ug Pb/L
November 2012
7-230
Draft - Do Not Cite or Quote
-------
Species
Mosquito
(Culex
quin quefascia tus)
Neosho mucket
(Lampsilis
rafinesqueana)
Concentration
Acute test:
100; 150; 200;
250 ug Pb/L
(analytically
verified)
50; 150; and
200 ug Pb/L
for reproductive
studies
Concentrations
were measured
and used to
calculate EC50
values, reported
in supplemental
data.
Exposure Method
24-hour acute toxicity
test and several tests
to assess reproductive
endpoints. All tests
were conducted with
Pb nitrate in distilled
water
24 and 48-hour
exposure with 5 day
old juveniles obtained
from adults collected
from Spring River, KS,
U.S.
Modifying
factors
Temperature:
25 ± 2 °C
pH:
7
Temperature:
20 ± 1 °C
pH:
7.2-7.6
DOC:
>7.0 mg/L
Hardness:
40-48 mg/L
as CaCO3
Alkalinity:
30-35 mg/L
as CaCO3
Effects on Endpoint
Reproduction:
Hatching rate significantly decreased,
lower emergence rates, larval
development from L1 to adults took
longer.
Survival:
24 hour LC50 reported
Survival:
Neosho mucket is a candidate
species for U.S. federal endangered
and threatened status. Toxicity
testing with newly transformed
juveniles indicated that this species is
sensitive to Pb exposure.
Reference3
(Published
since the
2006 Pb
Effect Concentration AQCD)
Survival: Kitvatanachai et
24 hour LC50: aL ^^>
1 80 ug Pb/L
24 hour EC50: Wang et al.
5 day old juveniles (201 Oe)
>507 ug Pb/L
48 hour EC50:
5 day old juveniles:
1 88 ug Pb/L
November 2012
7-231
Draft - Do Not Cite or Quote
-------
Species
Fatmucket mussel
(Lampsilis
siliquoidea)
Concentration Exposure Method
For 28-day 24 and 48-hour
exposure: exposure with
0.04; 2.9; 6.1; 17; glochidia, 96-hour
36; 83 ug Pb/L exposure with 5 day
(measured) old, and 2 or 6 month
old juveniles and
28-day exposure with 2
or 4 month old mussels
in reconstituted soft
water. Tests were
conducted with
glochidia and juveniles
obtained from adults
collected from the
Silver Fork of Perche
Creek, MO, U.S.
Modifying
factors
Temperature:
20 ± 1 °C
pH'
r
7.2-7.6
DOC:
>7.0 mg/L
Hardness:
40-48 mg/L
as CaCO3
Alkalinity:
30-35 mg/L
as CaCO3
Effects on Endpoint
Growth:
Growth of juvenile mussels in the
17 ug Pb/L concentration was
statistically significantly reduced
compared to growth in the controls at
the end of 28 days. Growth was not
assessed in the higher
concentrations due to mortality.
Survival:
The 24-hour EC50 values for glochidia
and 96-hour EC50 values for 2 and 6
month old juveniles were much
higher than 96 hour LC50 value for 5
day old newly transformed juveniles.
Genus mean chronic value was the
lowest value ever reported for Pb.
Survival was based on foot
movement within a 5-minute
observation period.
Effect Concentration
24 and 48 hour EC50:
glochidia
>400 ug Pb/L (test 1 )
>299 ug Pb/L (test 2)
48 hour EC50:
5 day old juveniles
465 ug Pb/L (test 1)
392 ug Pb/L (test 2)
96 hour EC50:
5 day old juveniles
142 ug Pb/L (test 1)
298 ug Pb/L (test 2)
24 and 48 hour EC50:
2 month old juveniles
>426 ug Pb/L
4 day EC50: >83 ug Pb/L
10 day EC50: >83 ug Pb/L
21 day EC50: 29 ug Pb/L
28 day EC50: 20 ug Pb/L
Reference3
(Published
since the
2006 Pb
AQCD)
Wang et al.
(201 Oe)
28-day NOEC
Juvenile fatmucket:
6.1 ug/L
28-day LOEC
Juvenile fatmucket:
17 ug/L
Genus mean chronic
value
10 ug Pb/L
Snail
(Lymnaea stagnalis)
4; 12; 16; 42; 113;
and 245 ug Pb/L
(measured)
30-day exposure with
newly hatched snails
(<24-hour old) in
artificial fresh water
with Pb nitrate.
Temperature: Growth:
23 ± 1 °C Newly hatched snails exhibited
greatly reduced growth in response
to Pb exposure
Survival:
No Pb-induced mortality was
observed.
EC20 <4 ug Pb/L
NOEC: 12 ug Pb/L
LOEC: 16 ug Pb/L
Grosell et al.
(2006b)
November 2012
7-232
Draft - Do Not Cite or Quote
-------
Species
Snail
(Lymnaea stagnalis)
Snail
(Marisa cornuarietis)
Snail
(Biomphalaria
glabrata)
Concentration
1st Experiment:
<0.5 (control),
2.7 and
18.9 |jg Pb/L
(measured)
2nd Experiment:
1.3 and
7.5 ug Pb/L
(measured)
5,000;1 0,000; and
15,000 ug Pb/L
(nominal)
50; 100; and
500 ug Pb/L
(nominal)
Exposure Method
Pb exposures were
performed with juvenile
snails (~ 1 g) for 21
days and then 1 4 days
in dechlorinated tap
water under flow-
through conditions.
5-day, 6-day, and
10-day exposure to
Pb chloride in
deionized or double-
distilled water. Snail
strain used for egg
production was from
the Zoological Institute
in Frankfurt, Germany.
96-hour acute
laboratory bioassays
Modifying
factors
Dechlorinated City
of Miami tap water
([Na+] -1.1 mmol/L
[Ca2+] -0.31
mmol/L
[Cf] -1.03 mmol/L
[HCO3~] -0.68
mmol/L,
[DOC]~200 umol/L
pH~7.7 at room
temperature
Temperature:
24 ± 1 °C
pH:
-7.5
Conductivity:
-800 uS/cm
Dechlorinated
continuously
aerated tap water:
Temperature:
22 °C
pH:
7.1 ±0.2
Total hardness:
65 ± 3 mg CaCO3/L
Alkalinity:
29 ± 2 mg CaCO3/L
Conductivity"
230 ± 17 uS
Effects on Endpoint Effect Concentration
Growth:
In juveniles exposed to 1 8.9 ug/L Pb
for 21 days, Ca2+ influx was
significantly inhibited and model
estimates indicated 83% reduction in
growth of newly hatched snails after
30 days at this exposure
concentration
Survival:
No Pb-induced mortality was
observed
Reproduction: LOEC: 10,000 ug Pb/L
Significant delay in hatching at
1 0,000 ug Pb/L
Growth:
Significantly delayed development
(reduced visible tentacles, eye
formation) at 15,000 ug Pb/L.
No effect on fresh weight.
Survival:
Significantly increased mortality at
15,000 ug Pb/L
Reproduction:
Significant decrease in number of
eggs laid at 500 ug Pb/L.
Survival:
Embryonic survival was 12% of the
number of eggs laid by the control
group at 100 ug Pb/L. Time to
hatching increased 3 fold from the
control. No embryos survived the
highest concentration.
Reference3
(Published
since the
2006 Pb
AQCD)
Grosell and Brix
(2009)
Sawasdee and
Kohler (201 0)
Ansaldo et al.
(2009)
November 2012
7-233
Draft - Do Not Cite or Quote
-------
Species
Prawn
(Macrobrachium
lancesteri)
Crayfish
(Orconectes hylas)
Concentration
22; 31; 48; 126;
170|jgPb/L
(measured)
Reference sites:
0.03 ug Pb/L
Mining sites:
0.12 to
1 .59 ug Pb/L
Downstream sites
0.03 to
0.04 ug Pb/L
Exposure Method
4-day exposures in
Pb chloride in aerated,
filtered, dechlorinated
tap water, static
aerated, renewal at
2 days. Prawns were
purchased from
aquarium shops in
Bani, Selangor,
Malaysia.
in situ 28-day exposure
with juvenile crayfish in
streams impacted by
Pb mining and
reference sites in
Missouri, USA.
Modifying
factors
Temperature:
28-30 °C
pH:
6.51 ±0.01
Conductivity:
244.3 ± 0.6 uS/cm
DO:
6.25 ± 0.06 mg/L
Total hardness
(Mg2+ and Ca2+):
15.63 ±2.74 mg/L
as CaCO3
Water quality
parameters were
measured at each
site
Temperature
23 to 26 °C
pH:
7.9 to 8.1
Conductivity:
282 to 858
DO:
6.3 to 8.4 mg/L)
Alkalinity 141 to
182 mg/L as
CaCO3
Turbidity: 0.4 to 0.6
NTU
Sulfate:
0.3 to 304 mg/L
Other metals were
present
downstream of
mining sites
Effects on Endpoint
Survival:
LC50 increased with decrease in
mean exposure concentration
Survival:
Crayfish survival and biomass were
significantly lower in streams
impacted by Pb mining Metal
concentrations were negatively
correlated with caged crayfish
survival.
Effect Concentration
24 hour LC50
85.9 ug Pb/L
48 hour LC50:
58.5 ug Pb/L
72 hour LC50:
45.5 ug Pb/L
96 hour LC50
35.0 ug Pb/L
Reference3
(Published
since the
2006 Pb
AQCD)
Shuhaimi-
Othman et al.
(2011 a)
Allertetal.
(2009a)
November 2012
7-234
Draft - Do Not Cite or Quote
-------
Species
Concentration
Exposure Method
Modifying
factors
Reference3
(Published
since the
2006 Pb
Effects on Endpoint Effect Concentration AQCD)
Vertebrates
Fathead minnow
(Pimephales
promelas)
Measured:
Mean ± SEM
Tap low Pb:
28 ± 1.1 |jg Pb/L
Tap high Pb:
105 ±4.8 |jg Pb/L
HCOs'lowPb:
31 ± 1 .2 |jg Pb/L
HC03"highPb:
113 ±4.6 |jg Pb/L
Humic low Pb:
30 ± 1 .4 |jg Pb/L
Humic high Pb:
112 ±4.5 ug Pb/L
4-day, 10-day, 30-day,
150-day, and 300-day
exposures in Pb nitrate
spiked dechlorinated
tap water with static
renewal to study the
effects of DOC and
alkalinity on Pb toxicity.
Breeding assays (21
days) were also
performed.
Temperature:
22 ±1 °C
Tap H2O:
Hardness:
91 mg/L
pH:
8.1
+500 |JM NaHC03:
DOC:
257 uM
Hardness:
93 mg/L
pH:
8.3
NaHCO3:
Hardness:
93 mg/L
pH:
8.3
+4 mg/L HA:
Hardness:
(-JQ n-i/N/l
yo mg/L
pH:
8.0
Humic:
Hardness:
93 mg/L
pH:
8.0
Growth: Mager et al.
No statistically significant growth (29-1Q)
differences observed at any age due
to water chemistry alone; DOC
addition strongly protected against
Pb accumulation; increased alkalinity
reduced whole body Pb burdens;.
Growth inhibited at 4 days, but
recovered by 30 days in high Pb
concentration.
Reproduction:
HCO3- reduced 21 day total
reproductive output (reduced clutch
size and number of clutches
produced); addition of HCO3" alone
actually increased reproductive
output; significantly higher fecundity
in HCO3" treatment; Egg attachment
low in both tap water and HCO3"
treatments; HA promoted
attachment; No statistically significant
differences in egg hatchability. HCO3"
and humic acid treatments increased
average egg mass; no effects on
hatchability in the HCO3"and humic
acid treatment;
November 2012
7-235
Draft - Do Not Cite or Quote
-------
Species
Fathead minnow
(Pimephales
promelas)
Concentration Exposure Method
<0.1 - 3,605 30 day flow through
ug Pb/L exposure to Pb nitrate
(measured) to determine to the
Pb was quantified effects of Ca*' '- numic
in fish tissues in a acid and PH (6-3 and
separate set of 8-3) on pb
experiments. accumulation and
toxicity in juvenile
fathead minnows.
Modifying
factors
Exposure media
were made up from
a base-water
consisting of 2:1
deionized water:
dechlorinated tap
water.
Temperature: 23
°C and had various
levels of Ca2+,
humics, and pH
values:
0.5; 1 ; 2 mM Ca2+
2; 4; 8; 16mg
humic
6.3; 8.3 pH
Effects on Endpoint
Growth:
No growth inhibition was observed in
any treatment. An increase in growth
was observed in groups exposed to
higher Pb concentrations where there
were high initial mortalities.
Survival:
For most treatments, mortalities
occurred during the first 5 to 7 days
of exposure. The lowest tolerance
was observed at low pH (6.8).
Addition of DOC or CaSO4
decreased Pb toxicity.
Effect Concentration
30 day LC50, EC20,
and LOEC values*:
LC 50 in ug Pb/L:
0.5 mM Ca2+: 91
1.0 mM Ca2+: 104
2 mg humic: 255
4 mg humic: 443
8 mg humic: 832
16 mg humic: 1903
pH 6.3: 4.5
pH 8.3: 13
EC 20 in ug Pb/L:
0.5 mM Ca2+: 47
1.0 mM Ca2+251
2 mg humic: 189
4 mg humic: 319
8 mg humic: 736
16 mg humic: 1729
pH 6.3: 2.1
pH 8.3: 8.7
LOEC in ug Pb/L:
0.5 mM Ca?+: 40
1.0mMCa2+:107
2 mg humic: 199
4 mg humic: 475
8 mg humic: 919
16 mg humic: 1751
pH 6.3: 6.2
pH8.3:15
*4 day and 1 0 day LC50,
LC2o, and LOEC values also
reported in the paper.
Reference3
(Published
since the
2006 Pb
AQCD)
Grosell et al.
(2006a)
November 2012
7-236
Draft - Do Not Cite or Quote
-------
Species
Fathead minnow
(Pimephales
promelas)
Fathead minnow
(Pimephales
promelas)
Concentration
Measured but not
reported. Figure 1
of Esbaugh et al.
(2011) plots the
relationship
between
dissolved and
nominal Pb
concentrations in
three waters with
low Pb solubility.
Predicted
concentration of
major Pb
chemical species
in the natural
water bioassays is
provided in
Table 4 of
Esbaugh et al.
(2011)
0.2 ±0.1 ugPb/L
(control)
33 ± 4 ug/L
(chronic low)
143 ± 14 ug/L
(chronic high)
(measured)
Exposure Method
Acute toxicity of Pb to
juvenile P. promelas
(<24 hours old) was
assessed in 96-hour
static renewal
exposures to two lab
generated reference
waters and seven
natural waters from
across North America
selected to include a
range of water quality
parameters. Waters
were spiked with
varying concentrations
of Pbas Pb nitrate.
33 to 57-day exposures
in dechlorinated tap
water and deionized
water to Pb nitrate to
assess swimming
performance.
Modifying
factors
Temperature: 26
°C
Water chemistry of
the field-collected
waters are reported
in Table 1 of
Esbaugh et al.
(201 1 ) including pH
(range 5.5 to 8.5),
Ca2+ (range 24 to
1 ,934 uM), DOC
(range 36 to 1 ,244
uM) and hardness
(range 4 to 298
mg/L)
Temperature:
21 ± 1 °C
pH:
7.50 ±0 .03
Total CO2:
543 ± 69 umol/L
DOC:
108± 4umol
carbon/L
Hardness:
26 ± 3 mg/L
Effects on Endpoint Effect Concentration
Survival: 96-hour LC50 range:
LC50 values ranged from 41 to 41 to 3.598 U9 pb/L
3,598 ug Pb/L. NOEC: range
DOC had the strongest protective 1 4 to 2,271 ug Pb/L.
effect. LOEC: range
The lowest LC50 occurred in the pH 42 to 5>477 U9 pb/L
5.5 water.
No Pb toxicity was observed in three
alkaline natural waters.
Growth:
Fish from the chronic low exposure
were significantly larger than control
fish (increased mass and body
length).
Reference3
(Published
since the
2006 Pb
AQCD)
Esbaugh et al.
(2011)
Magerand
Grosell (201 1 )
November 2012
7-237
Draft - Do Not Cite or Quote
-------
Species
Fathead minnow
(Pimephales
promelas)
Zebra fish
(Dan/o rerio)
Concentration
Fish were feed
L. variegatus
exposed via water
to 628 |jg Pb/L.
Mean daily dietary
dose of 0.417
(0.3-0.48)
or 0.1
(0.07-0.14)
mg Pb/kg/day
(measured)
Exposure Method
Juvenile fish fed a live
diet of the oligochaete
L. variegatus for
30 days contaminated
with Pb.
63 day dietary
exposure with
Pb-enriched
polychaete, Nereis
diversicolor.
Adult zebrafish were
fed a daily dose of 1 %
flake food (dry wet
diet/wet weight fish),
1% brine shrimp, and
1 % N. diversicolor
collected from either
Gannel Estuary,
Cornwall, U.K., an
estuary with legacy Pb
contamination, or
Blackwater Estuary,
Essex, U.K., (reference
site)
Modifying
factors
Water from Lake
Superior that was
subsequently
filtered and UV-
treated.
Temperature:
25 °C
pH:
7.5-8.0,
Hardness:
45-50 mg CaCO3,
Alkalinity :
40-45 mg CaCO3/L
Temperature:
29 ± 1 °C
Reference3
(Published
since the
2006 Pb
Effects on Endpoint Effect Concentration AQCD)
Growth: Erickson et al.
Not significantly affected (201 0)
Survival:
Not significantly affected
Reproduction: Boyle et al.
No impairment observed to incidence (2P-1PJ
of spawning, numbers of eggs per
breeding pair or hatch rate of
embryos compared with pre-
exposure levels. Metal analyses
revealed significant increases in
whole-body Pb burdens of male fish
fed polychaetes from the
contaminated estuary.
November 2012
7-238
Draft - Do Not Cite or Quote
-------
Species
Tilapia
(Oreochromis
niloticus)
Channel catfish
(Ictalurus
Punctatus)
African catfish
(Clarias gariepinus)
Concentration
Mean
concentration of
Pb in food pellets:
100; 400; and 800
mg Pb/kg dry
weight (nominal)
Fish were fed
L. variegatus
exposed via water
to 576 |jg Pb/L
100; 300; and
500 |jg Pb/L
(nominal)
Pb was quantified
in tissues
following
exposure.
Exposure Method
Fish obtained from an
unpolluted fish farm in
Hangzhou, China were
held in tanks with
dechlorinated tap water
and were fed diets with
Pb nitrate twice daily
for 60 days.
Juvenile fish fed a live
diet of the oligochaete
L. variegatus for
30 days contaminated
with Pb.
24; 42; 90; 138; and
162 hour embryo
exposure to Pb nitrate
in dechlorinated tap
water.
These intervals
corresponded to 30;
48,; 96; 144; and 168
hours post-fertilization.
Modifying
factors
Temperature:
25 ± 1 °C
„. i_| .
pH.
7.1-7.5
DO:
7.5-7.8 mg/L
Alkalinity:
109 mg CaCOs/L
Hardness:
118 mg CaCO3/ L
Water from Lake
Superior that was
subsequently
filtered and UV-
treated.
Temperature: 25
°C
pH:
7.5-8.0,
Hardness:
45-50 mg CaCOs/L
Alkalinity:
40-45 mg CaCO3/L
Temperature:
24 °C
pH:
8.0
Conductivity:
700 us/cm
Oxygen:
90-95% saturation
Reference3
(Published
since the
2006 Pb
Effects on Endpoint Effect Concentration AQCD)
Growth: Dai et al.
No effects on growth were found. (2009b)
Survival:
Exposure to Pb-contaminated diets
did not result in mortality.
Growth: Erickson et al.
Not significantly affected (201 0)
Survival:
Not significantly affected
Growth: Osmanetal.
Malformations observed in exposed (2007b)
embryos (malformed embryos only
survived shortly after hatching), delay
in development.
Reproduction:
Concentration-dependent delay in
hatching, reduced percentage of
embryos completing egg stage
period from 75% in control to 40% in
500 ug Pb/L.
November 2012
7-239
Draft - Do Not Cite or Quote
-------
Species
Rainbow trout
(Oncorhynchus
mykiss)
Rainbow trout
(Oncorhynchus
mykiss)
Concentration
In diet:
6.9 mg Pb-(g dry
mass)
(L. variegatus
exposed via
sediments)
Control Pb-free
diet of 0.06 mg
Pb/kg dry weight,
and three different
diets of 7; 77; and
520 mg Pb dry
weight (0.02
(control), 3.7;
39.6; and 221 .5
mg Pb/day dry
weight calculated
for food
consumption)
Pb also quantified
in tissues
Exposure Method
Juvenile fish fed a live
diet of the oligochaete
L. variegatus for
30 days contaminated
with Pb of varying
concentrations of
Pb nitrate
21 -day exposure to
juvenile rainbow trout
via diet amended with
Pb nitrate. Fish were
held in aerated tanks
with dechlorinated
water
Modifying
factors
Water from Lake
Superior that was
subsequently
filtered and UV-
treated.
Temperature:
11 °C
pH:
7.5-8.0
Hardness:
45-50 mg CaCOs/L
Alkalinity :
40-45 mg CaCO3/L
Temperature:
11-13°C
pH
7.5-8.0
Hardness:
140 ppm
as CaCO3
Reference3
(Published
since the
2006 Pb
Effects on Endpoint Effect Concentration AQCD)
Growth: Erickson et al.
Not significantly affected (201 0)
Survival:
Not significantly affected
Growth: Alvesetal.
No effects on growth rates were (2006)
observed in rainbow trout
administered a diet containing three
concentrations of Pb.
Dietary Pb was poorly absorbed.
Comparison of dietary and water-
borne exposures suggest that toxicity
does not correlate with dietary
exposure, but does correlate with gill
accumulation from waterborne
exposure.
Survival:
Not significantly affected by dietary
Pb
November 2012
7-240
Draft - Do Not Cite or Quote
-------
Species
Rainbow trout
(Oncorhynchus
mykiss)
Concentration Exposure Method
ELS (early life 96-hour static renewal
stage) test 1 : acute toxicity test with
1 2; 24; 54; 1 43; swim-up stage fry (2 to
and 384 ug Pb/L 4 weeks post-hatch)
(measured) reared from eggs used
ELS test 2- in the chronic studies.
8; 18; 37; 87; and Two 60+ day ELS
124 ug Pb/L exposures were
(measured) conducted in a flow-
through system using
temperature controlled
water from Little North
Fork of the South Fork
Coeurd'Alene river in
ELS 1 (69 days) and
water from the South
Fork in ELS 2 (62
days).
Modifying
factors
ELS 1:
Temperature
9.8 ± 0.6 °C
pH:
6.75 ±0.4
Hardness:
19.7± 1.5mg/L
as CaCO3
Conductivity:
45.8 ±2.2 us
Alkalinity:
19.6± 2.2mg/Las
PaPPl
OdO*— '3
DO:
10.2±0.7mg/L
ELS 2:
Temperature
\ O C 4. ri Q op
i z.o x u.y o
pH:
7.19 ± 0.3
Hardness:
29.4 ± 3.6 mg/L
as CaCO3
Conductivity:
69.1 ±7.4 us
Alkalinity:
27 mg/L ± 2.1 as
CaCO3
DO:
9.2 ± 0.9 mg/L
Effects on Endpoint
Growth:
In ELS 1 , growth generally
decreased as concentration
increased, with fish in the highest
surviving treatment (143 ug Pb/L )
exhibiting severely stunted growth
that was statistically different from
the control.
In ELS 2, growth increased in the
highest treatment with a slight
reduction in length and wet weight in
intermediate exposures.
Survival:
In ELS 1 Survival decreased as
concentration increased with
complete mortality before the end of
the test in the highest treatment
(384 ug Pb/L).
In ELS 2, survival decreased
significantly in the highest treatment
with high survival in intermediate
exposure. In both tests, the greatest
number of mortalities occurred
around or shortly after the swim-up
stage.
Reference3
(Published
since the
2006 Pb
Effect Concentration AQCD)
96-hour LC50: Mebane et al.
120ugPb/L (2008)
ELS 1: Survival
NOEC: 24 ug Pb/L
LOEC: 54 ug Pb/L
MATC: 36 ug Pb/L
EC10: 26 ug Pb/L
EC20 : 34 ug Pb/L
ELS 1: Weight
NOEC: 24 ug Pb/L
LOEC: 54 ug Pb/L
MATC: 36 ug Pb/L
EC10: 39 ug Pb/L
EC20: 55 ug Pb/L
ELS 1: Length
NOEC: 54 ug Pb/L
LOEC: 143ug Pb/L
MATC: 88 ug Pb/L
EC10: 64 ug Pb/L
EC20: 98 ug Pb/L
ELS 2: Survival
NOEC: 87 ug Pb/L
LOEC: 1 25 ug Pb/L
MATC: 104ug Pb/L
EC10: 108ug Pb/L
EC20: 113ug Pb/L
ELS 2: Weight
NOEC: 37 ug Pb/L
LOEC: 87 ug Pb/L
MATC: 57 ug Pb/L
EC10: 7 ug Pb/L
EC20: >87 ug Pb/L
ELS 2: Length
NOEC: 8 ug Pb/L
LOEC: 18 ug Pb/L
MATC: 12ugPb/L
EC10: >87 ug Pb/L
EC20: >87 ug Pb/L
November 2012
7-241
Draft - Do Not Cite or Quote
-------
Species
Southern leopard
frog
(Rana
sphenocephala)
Northern leopard
frog
(Rana pipiens)
Concentration
Sediment:
45; 75; 180; 540;
2,360; 3,940;
5,520; 7,580
mg Pb/kg
dry weight
Corresponding
sediment pore
water:
123; 227; 589;
1,833; 8,121;
13,579; 19,038;
24,427 ug Pb/L
(measured)
3; 10; and
100 ug Pb/L
(nominal)
(Pb was
measured in
tissues at the end
of the study. Pb
tissue
concentrations
ranged from 0.1 to
224.5 mg Pb/kg
dry mass and fell
within the range of
tissue
concentrations in
wild-caught
tadpoles).
Exposure Method
20; 40; 61 ; and 82-day
exposures in
Pb acetate spiked
sediment collected
from wetland, static
renewal twice per
week.
Northern leopard frogs
were exposed to Pb as
Pb nitrate in
dechlorinated water
from the embryonic
stage to
metamorphosis
(>66 days post-
hatching)
Modifying
factors
Mean (SD)
Temperature:
21.6°C
pH:
6.92
DOC:
6.08 mg/L
Conductivity:
168uS/L
Hardness:
7.30 mg Ca2+/L
Ammonia:
0.39 mg/L
Sediment:
Organic carbon:
8.25%
Sand: 22.4%
Silt: 38.4%
Clay: 39.1%
Temperature:
21 to 22 °C
PH
7.9
Hardness:
1 70 mg/L
as CaCO3
Effects on Endpoint
Growth:
Snout-vent length and body mass
increased through time in all
treatments; skeletal deformations
(spinal deformations, digits truncated
and twisted, long bones curved and
truncated) increased with Pb content
and length of exposure
Survival:
Exposure to a 3,940 mg/kg sediment
Pb (13,579 ug/L) pore water) killed all
tadpoles within 5 days; tadpoles that
reached climax stage (Gosner 42)
had no difference in survival among
treatments through the completion of
metamorphosis.
Growth:
Tadpole growth was significantly
slower in the early stages in
100 ug Pb/L treatment. More than
90% of tadpoles in the 100 ug Pb/L
treatment developed lateral spinal
curvature, whereas almost all the
tadpoles in the other groups were
morphologically normal. No
significant effect of Pb exposure was
found on percentage metamorphosis,
snout-vent length, mortality, or sex
ratio. Time to metamorphosis was
delayed in 100 mg/L treatment.
Reference3
(Published
since the
2006 Pb
Effect Concentration AQCD)
40 day EC50 Sparling et al.
(for deformed spinal (2006)
columns in sediment):
1 ,958 mg/kg Pb
Corresponding EC50
(for deformed spinal
columns in pore water):
6,734 ug/L
40 day EC50
(for deformed spinal
columns in sediment):
579 mg Pb/kg,
and
1 ,968 ug Pb/L (in pore
water).
Sediment: LC50:
3,728 mg Pb/kg
(Corresponding to a
Pore water LC50 of:
12,539 ug Pb/L).
Chen et al.
(2006b)
"References included are those which were published since the 2006 Pb AQCD.
November 2012
7-242
Draft - Do Not Cite or Quote
-------
Table 7-6 Saltwater plants, invertebrates, and vertebrates: growth, reproduction, and survival.
Species
Concentration
Exposure
Method
Modifying
factors
Effects on Endpoint
Effect Concentration
Reference3
(Published
since the
2006 Pb
AQCD)
Algae
Microalgae
(Tetraselmis chuii,
Rhodomonas salina,
Chaetocerossp.,
Isochrysis galbana,
Nannochloropsis
gaditana)
Microalgae
(Tetraselmis
suecica)
50; 100; 250; 500;
800; 1,000; 1,600;
3,000; and
6,000 |jg Pb/L,
(nominal). Five
nominal
concentrations
were analytically
verified: 51 ; 225;
824; 1 ,704;
6,348 |jg Pb/L
(measured)
20 ug Pb/L
(nominal)
T. suecica in this
study was then
fed to Artemia
franciscana
(mean Pb content
12 to 15 mg
Pb/kq)
y/
Populations of
each microalgal
species were
exposed for 72
hours to ten
progressively
increasing
nominal
concentrations of
Pb in filtered
seawater
72-hour
exposure to
Pb nitrate in
filtered natural
seawater from
Mazatlan Bay,
Mexico. This
was the first step
in a four-level
food chain.
Temperature:
20 ± 1 °C
pH
8.0
Temperature:
28 ± 2 °C
Salinity:
34.6 ± 1 .2 ppt
nU-l-
pH.
-T Q Q 0
/ .y-o.z
DO saturation:
90-95%
(>7 mg/L)
Growth:
Growth inhibition (as measured
by flow cytometry) was reported
for each species. Species cellular
size, sorption capacity, or
taxonomy did not explain
differences in sensitivity to Pb.
Growth:
Mean final cell concentrations,
growth rate and total dry biomass
were significantly reduced (40%
lower than control cultures).
Effects on primary, secondary
and tertiary consumers were
observed following Pb-exposure
via T. suecica at the base of a
simulated marine food chain.
EC50:
T. chuii:
2,640 ug Pb/L
R. salina:
900 ug Pb/L
Cftaefocerossp.:
105ug Pb/L
/. galbana:
1 ,340 ug Pb/L
N. gaditana:
740 ug Pb/L
Debeliuset al.
(2009)
Soto-Jimenez
et al. (201 1 b)
Invertebrates
Polychaete
(Hydroides elegans)
91 ; 245; 451 ;
4,443; 9,210; and
41 ,060 ug Pb/L
(measured)
24-hour
exposure of
fertilized eggs to
Pb chloride.
Assay was
stopped at
2 hours to
assess effects
on blastula.
Temperature:
27 ± 1 °C
DO
(86.5%)
Salinity
(34 ± 1 ppt)
pH(8.1 ±0.1)
Carbonate
24.5 mg/L.
Reproduction:
Exposure to Pb caused a
significant decrease in the
number of embryos developing
normally to blastula after 2 to 3
hours of exposure to Pb.
EC50
Fertilization
membrane stage :
30,370 ug Pb/L
Blastula
1 ,429 ug Pb/L
24 hour trochophore
larva:
231 ug Pb/L.
Gopalakrishnan
et al. (2007)
November 2012
7-243
Draft - Do Not Cite or Quote
-------
Species Concentration
Polychaete 48; 97; 201 ; 407;
(Hydroides elegans) 803;
' ' w ' 1,621 |jgPb/L
(measured)
Polychaete 85; 137; 251;
(Capitellasp.) 392; 487; 738;
871 mg Pb/kg
(measured)
Exposure
Method
A series of
experiments
were performed
from 20 minutes
to 4 days in
Pb chloride
using
polychaetes
collected from
seawater in
Chennai, India.
3 and 6-day
exposure for
growth
experiments,
96-hour
exposure to
Pb chloride
spiked sediment
from
Chi-kou Estuary,
Taiwan
Modifying
factors
Temperature:
27 ± 1 °C
DO:
7-9 mg/L
Salinity
34 ± 1 ppt
pH:
8.1 ±0.1
Carbonate:
22.5 mg/L
Aerating
circulating
seawater
Temperature:
20 ± 2 °C
Salinity:
30%
Effects on Endpoint
Reproduction:
Fertilization rate decreased by
70% in sperm pretreated with
97 ug Pb/L for 20 minutes.
Fertilization rate of eggs
pretreated in 48 ug Pb/L
decreased to 20% of control. Life
stages of H. elegans varied in
their sensitivity to Pb. Gametes,
embryo and larvae were more
sensitive than adults with the
larval settlement period being
most sensitive to Pb exposure.
Survival:
LC50 reported for adults
Growth:
Significant differences among
growth rates of Capitella sp. in
different levels of
Pb-contaminated sediments, with
the exception of 251 mg/kg
treatment in the 6-day
experiment. Growth rates
deceased significantly from the
control in the 3-day experiment
but changes were inconsistent
with increasing Pb concentration.
Survival:
No effect
Effect Concentration
EC50
Sperm toxicity:
380 ug Pb/L
Egg toxicity:
692 ug Pb/L
Embryo toxicity:
1,130 ug Pb/L
Blastula to trochophore:
261 ug Pb/L
Larval settlement:
100 ug Pb/L
Adult 96-hour LC50 :
946 ug/L
Growth LOAEL:
85 mg Pb/kg
Reference3
(Published
since the
2006 Pb
AQCD)
Gopalakrishnan
et al. (2008)
Horng et al.,
(2009)
November 2012
7-244
Draft - Do Not Cite or Quote
-------
Species
Amphipod
(Elasmopus laevis)
Amphipod
(Melita plumulosa)
Concentration
30-mg Pb/kg
(control whole-
sediment),
58 mg Pb/kg;
118mg Pb/kg;
234 mg Pb/kg;
424 mg Pb/kg
(measured)
Water-only tests
ranged from 0 to
4,000 ug Pb/L
(analytically
verified)
Adult Sediment
Test:
500; 1,000; 2,000;
4,000 mg Pb/kg
dry weight
(analytically
verified)
Juvenile
Sediment Test:
500; 1,000; 2,000
mg Pb/kg dry
weight
(analytically
verified)
Exposure
Method
Multi-
generational
bioassay with
amphipods
collected in
Jamaica Bay,
New York
exposed
60+ days to
sediment spiked
with Pb acetate
in filtered
seawater. 10-
day and 28-day
bioassays were
also conducted.
Juveniles and
adults were
tested in 96-hour
seawater only or
1 0 day static-
non-renewal
exposure spiked
sediment
collected from
intertidal mud
flats, Woronora
River, New
South Wales,
Australia. Adults
were also tested
in 10-day
seawater only
exposures.
Modifying
factors
Temperature:
19-24°C
Salinity:
27-29 g/L
DO:
>6.57 mg/L
Temperature
21 ± 1 °C,
Salinity
30 ± 1 %,
PH
7.2-8.2,
Ammonia (total)
<3 mg N/L
Effects on Endpoint
Reproduction:
Fecundity was reduced as
sediment Pb concentration
increased.
Onset of reproduction and
reproduction were delayed as Pb
concentration increased.
Survival:
No differences in adult survival
among the Pb concentrations
tested in 28-day and 60-day
exposures.
Survival:
Juvenile amphipods were more
sensitive to Pb than adults in
seawater and sediment
exposures.
Effect Concentration
Fecundity and time of first
offspring was significantly
reduced with increasing
sediment concentration
above 118 mg Pb/kg. Onset
to reproduction significantly
delayed at 1 1 8 mg Pb/kg
and delayed further at
higher tested
concentrations.
96-hour seawater-only
Adults:
LC50 3,000 ug Pb/L
NOEC 850 ug Pb/L
LOEC 1 ,680 ug Pb/L
Juveniles:
LC50 1 ,530 ug Pb/L
NOEC 400 ug Pb/L
LOEC 600 ug Pb/L
Seawater-only 10 days:
Adults:
LC50 1 ,270 ug Pb/L
NOEC 1 90 ug Pb/L
LOEC 390 ug Pb/L
10 days Sediment-only
Adults:
LC50 NOEC, LOEC
>3,560, mg Pb/kg
Juveniles:
LC50 1 ,980 mg Pb/kg
NOEC 580 mg Pb/kg
LOEC 1 ,020 mg Pb/kg
Reference3
(Published
since the
2006 Pb
AQCD)
Ringenary et al.
(2007)
King et al.
(2006)
November 2012
7-245
Draft - Do Not Cite or Quote
-------
Species
Brine shrimp
(Artemia
franciscana)
Shrimp
(Litopenaeus
vannamei)
Sea urchin
(Paracentrotus
lividus)
Concentration
Mean Pb content
12 to 15mg Pb/kg
from dietary
exposure to Pb.
Pb in exoskeleton,
hepatopancreas,
muscle, and
remaining tissues
was quantified on
daysO; 15; 28;
and 42 of the
dietary study
50 to
5,000 ug Pb/L
(nominal)
Exposure
Method
This was the
second step in a
four-level food
chain.
A. franciscana
feeding on
Tetraselmis
suecica cultured
in 20 ug/L Pb, as
Pb nitrate.
This was the
third step in a
four-level food
chain.
L. vannamei, fed
A. franciscana
(mean Pb
content 12 to 15
mg Pb/kg)
feeding on
T. suecica
cultured in
20 ug/L Pb as
Pb nitrate.
Gametes and
embryos
exposed 48 to
50 hours in
filtered seawater
to Pb nitrate
from adults
collected from
the Bay of
Naples, Italy.
Modifying
factors
Temperature:
28 ± 2°C
Salinity:
34.6 ± 1 .2 ppt
nU-l-
pH.
7.9-8.2
DO saturation:
90-95%
(>7 mg/L)
Temperature:
28 ± 2°C
Salinity:
34.6 ± 1 .2 ppt
pH:
7.9-8.2
DO saturation:
90-95%
(>7 mg/L)
Filtered sea
water
Temperature:
18 ± 1 °C
Salinity: 38%
pH: 8 ±0.2
Effects on Endpoint
Growth:
A tendency toward lower biomass
yields was reported (significant
only on day of final harvest).
Survival:
A tendency toward lower survival
was reported (significant only on
day of final harvest).
Growth:
Tendency toward decreased total
length and weight was reported
(significant only on day of final
harvest)
Survival:
Tendency toward lower survival
was reported (significant only on
day of final harvest)
Growth:
Up to LOEC concentration,
defects observed in plutei were
mainly reduction in size (20%);
above LOEC concentration,
developmental defects were
mainly larvae affected in skeletal
or gut differentiation up to
2,000 ug Pb/L, where arrest of
development started to increase.
Effect Concentration
Dry biomass was 1 95 mg/L
in control cultures and 153
mg/L in cultures fed Pb
exposed T. suecica. Mean
cell count (individuals/L) on
days 19-23 (harvest) was
320 in control and 255 in
A. franciscana cultures fed
Pb-exposed T. suecica.
Total mean length in the
shrimp fed the experimental
diet was 13 mm and wet
weight was 7.1 g compared
to mean total length (14.8
mm) and wet weight (8.5 g)
at day 42.
Mean survival in the shrimp
fed the experimental diet
ranged from 67 to 78%
compared to control survival
(84 to 90%) at day 42.
EC50
1 ,250 ug Pb/L
NOEL
250 ug Pb/L
LOEC
500 ug Pb/L
Reference3
(Published
since the
2006 Pb
AQCD)
Soto-Jimenez
et al. (201 1 b)
Soto-Jimenez
et al. (201 1 b)
Manzo et al.
(201 0)
November 2012
7-246
Draft - Do Not Cite or Quote
-------
Species
Mussel
(Mytilus
galloprovincialis)
Clam
(Meretrix meretrix)
Scallop
(Argopecten
ventricosus)
Concentration
3,500; 4,500;
5,500;
6,000 ug Pb/L
(nominal)
2; 20; 197; 1,016;
7,158ug Pb/L
(measured)
For growth:
10; 100; 1,000;
and 10,000 ug/L
(analytically
verified)
For survival:
280; 560; 1120;
2,250; and
5,000 ug/L
(analytically
verified)
Exposure
Method
24-hour static
aerated
exposure
in seawaterwith
Pb acetate with
mussels
collected from a
mussel farm in
Greece.
24-hour and 96-
hour toxicity test
with Pb nitrate in
filtered sea water
using gametes
from adults
collected from
Wenzhou, China
and held under
laboratory
conditions.
144 hour
(survival) or 30
day (growth)
exposure to
Pb nitrate (static
renewal every
48 hours) with
juvenile
A. ventricosus
hatched from
laboratory
cultures held at
Universidad
Autonoma de
Baja California
Sur, Mexico.
Modifying
factors
Temperature:
25 ± 2°C
Salinity:
35%
DO:
7-8 mg/L
Temperature:
28 ± 1 °C
Salinity:
ono/.
ZU fO
pH:
7.8
Temperature:
20°C
Salinity:
36 ± 1 %
HO-
uu.
>4 mg/L
Effects on Endpoint
Survival:
Mortality at high Pb
concentrations
Reproduction:
Embryo development inhibited by
increasing Pb concentrations
Growth:
Significant concentration-
dependent growth inhibition in
larvae. Larval metamorphosis
rate decreased, no adverse effect
on larval settlement at
20.4 ug Pb/L
Survival:
Significant concentration-
dependent survival inhibition of
embryos over time
Growth:
Juvenile growth rates and weight
were significantly reduced at high
concentrations of Pb
Survival:
Juvenile mortality was
significantly different than control
at 96 hour LC50
Effect Concentration
24-hour LC50
4,500 ug Pb/L
Embryogenesis EC50:
297 ug Pb/L
Growth:
EC50:
197ug Pb/L
Metamorphosis:
>7,160ugPb/L
48-hour LC50:
>7,160ugPb/L
96-hour LC50:
353 ug Pb/L
EC50 for growth:
4,210 ug Pb/L
72-hour LC50:
4,690 ug Pb/L
96-hour LC50:
830 ug Pb/L
120-hour LC50:
680 ug Pb/L
144-hour LC50:
680 ug Pb/L
Reference3
(Published
since the
2006 Pb
AQCD)
Vlahogianni
and Valavanidis
(2007)
Vlahogianni
and Valavanidis
(2007)
Sobrino-
Figueroa et al.
(2007)
November 2012
7-247
Draft - Do Not Cite or Quote
-------
Species
Bivalve
(Tellina deltoid alls)
Exposure
Concentration Method
1 ,000 mg Pb/kg 1 0 day direct
(analytically exposure to
verified) Pb nitrate spiked
sediment
collected from
Woronora River,
New South
Wales, Australia.
Adults used in
the test were
collected from
Lane Cove
River, Sydney,
Australia and
held in filtered
seawater.
Modifying
factors
Temperature:
21 ± 1 °C
Salinity
30 ± 1%,
PH
7.2-8.2
Ammonia (total)
<3 mg N/L
Effects on Endpoint Effect Concentration
Survival: 10DayNOEC
All individuals survived. Porewater: 1 5 ug Pb/L
dissolved Pb
Sediment: 1 ,000 mg Pb/kg
Reference3
(Published
since the
2006 Pb
AQCD)
King et al.
(201 0)
Vertebrates
Toadfish
(Tetractenos glaber)
Measured but not Field-collected
reported fish-Pb sampled
from sediments
collected in two
reference and
two metal
contaminated
estuaries near
Sydney,
Australia
Temperature :
15 to 16°C
Salinity:
29 to 31 %,
pH:
8.4 to 8.6,
amongst
estuaries.
Sediment pH:
7.0-7.5
Organic matter:
1.5-2.1%
Reproduction: N/A
Decreased oocyte diameter and
density in the toadfish were
associated with elevated levels of
Pb in the gonads of field collected
fish; authors state that this is
suggestive of a reduction in egg
size.
Alquezaret al.
(2006)
November 2012
7-248
Draft - Do Not Cite or Quote
-------
Species
Grunt fish
(Haemulon scudderi)
Concentration
Mean total Pb
body burden
increased from
0.55 to 3.32 mg
Pb/kg during the
feeding
experiment.
Exposure
Method
42-day dietary
exposure from
simulated marine
food chain-
shrimp,
L. vannamei, fed
A franciscana
(mean Pb
content
12-1 5 mg Pb/kg)
feeding on
T. suecica
cultured in
20 ug/L Pb as
Pb nitrate.
Modifying
factors
Temperature:
28 ± 2 °C
Salinity:
34.6 ± 1 .2 ppt
nU-l- 7 Q R O
pn. /.y-o.z
DO saturation:
90-95%
(>7 mg/L)
Effects on Endpoint
Survival:
No significant differences
observed in intermediate and
final length, mean wet weight, or
Fulton's condition factor: Final
survival significantly lower; mean
total Pb body burden increased.
Effect
Concentration
Mean survival in the fish fed
the experimental diet ranged
from 65 to 75% compared to
control diet survival (88 to
91 %).
Reference3
(Published
since the
2006 Pb
AQCD)
Soto-Jimenez
et al. (201 1 b)
References included are those which were published since the 2006 Pb AQCD.
November 2012
7-249
Draft - Do Not Cite or Quote
-------
References for Chapter 7
Acosta. JA; Jansen. B; Kalbitz. K; Faz. A; Martinez-Martinez. S. (2011). Salinity increases mobility of
heavy metals in soils. Chemosphere 85: 1318-1324.
http://dx.doi.0rg/10.1016/i.chemosphere.2011.07.046
Adevemo. OK. (2007). Haematological profile of Clarias gariepinus (Burchell, 1822) exposed to lead.
Turkish Journal of Fisheries and Aquatic Sciences 7: 163-169.
Adevemo. OK. (2008a). Histological alterations observed in the gills and ovaries of Clarias gariepinus
exposed to environmentally relevant lead concentrations. J Environ Health 70: 48-51.
Adevemo. OK. (2008b). Histological alterations observed in the liver and brain of Clarias gariepinus
exposed to chronic sublethal dose of lead. Bulletin of the European Association of Fish Pathologists
28: 105-114.
Ahlf. W: Drost. W: Heise. S. (2009). Incorporation of metal bioavailability into regulatory frameworks:
Metal exposure in water and sediment. Journal of Soils and Sediments 9: 411-419.
http://dx.doi.org/10.1007/sll368-009-0109-6
Ahmad. MS: Ashraf. M; Tabassam. Q; Hussain. M; Firdous. H. (2011). Lead (Pb)-induced regulation of
growth, photosynthesis, and mineral nutrition in maize (Zea mays L.) plants at early growth stages.
Biol Trace Elem Res 144: 1229-1239. http://dx.doi.org/10.1007/sl2011-011-9099-5
Ai-Khlaifat. AL; Al-Khashman. OA. (2007). Atmospheric heavy metal pollution in Aqaba City, Jordan,
using Phoenix dactylifera L. leaves. Atmos Environ 41: 8891-8897.
http://dx.doi.0rg/10.1016/i.atmosenv.2007.08.028
Aisemberg. J; Nahabedian. DE; Wider. EA; Verrengia Guerrero. NR. (2005). Comparative study on two
freshwater invertebrates for monitoring environmental lead exposure. Toxicology 210: 45-53.
http://dx.doi.0rg/10.1016/i.tox.2005.01.005
Akerblom. S: Baath. E; Bringmark. L; Bringmark. E. (2007). Experimentally induced effects of heavy
metal on microbial activity and community structure of forest mor layers. Biol Fertil Soils 44: 79-91.
http://dx.doi.org/10.1007/s00374-007-0181-2
Albrecht. J: Abalos. M; Rice. TM. (2007). Heavy metal levels in ribbon snakes (Thamnophis sauritus)
and anuran larvae from the Mobile-Tensaw River Delta, Alabama, USA. Arch Environ Contam
Toxicol 53: 647-654. http://dx.doi.org/10.1007/s00244-006-0175-3
Aldenberg. T; Slob. W. (1993). Confidence limits for hazardous concentrations based on logistically
distributed NOEC toxicity data. Ecotoxicol Environ Saf 25: 48-63.
http://dx.doi.org/10.1006/eesa.1993.1006
Aldjain. I.. M.: Al-Whaibi. M.. H.; Al-Showiman. S.. S.: Siddiqui. M.. H. (2011). Determination of
heavy metals in the fruit of date palm growing at different locations of Riyadh. Saudi J Biol Sci 18:
175-180. http://dx.doi.0rg/10.1016/i.sibs.2010.12.001
Allert. AL; Fairchild. JF; DiStefano. RJ; Schmitt. CJ: Brumbaugh. WG: Besser. JM. (2009a). Ecological
effects of lead mining on Ozark streams: In-situ toxicity to woodland crayfish (Orconectes hylas).
Ecotoxicol Environ Saf 72: 1207-1219. http://dx.doi.Org/10.1016/i.ecoenv.2008.08.005
Allert. AL; Fairchild. JF; Schmitt. CJ: Besser. JM; Brumbaugh. WG: Olson. SJ. (2009b). Effects of
mining-derived metals on riffle-dwelling benthic fishes in Southeast Missouri, USA. Ecotoxicol
Environ Saf 72: 1642-1651. http://dx.doi.Org/10.1016/i.ecoenv.2009.02.014
Alonso-Azcarate. J: Ruiz. E; Rodriguez. L. (2011). Changes in heavy metal speciation induced by
earthworm feeding activity. Fresen Environ Bull 20: 26-35.
November 2012 7-250 Draft - Do Not Cite or Quote
-------
Alonso-Castro. AJ; Carranza-Alvarez. C: Alfaro-De la Torre. MC: Chavez-Guerrero. L; Garcia-De la
Cruz. RF. (2009). Removal and accumulation of cadmium and lead by Typha latifolia exposed to
single and mixed metal solutions. Arch Environ Contam Toxicol 57: 688-696.
http://dx.doi.org/10.1007/s00244-009-9351-6
Alquezar. R; Markich. SJ: Booth. DJ. (2006). Effects of metals on condition and reproductive output of
the smooth toadfish in Sydney estuaries, south-eastern Australia. Environ Pollut 142: 116-122.
http://dx.doi.0rg/10.1016/i.envpol.2005.09.009
Alves Costa. JRM; Mela. M; da Silva de Assis. HC: Pelletier. E; Randi. MAP; de Oliveira Ribeiro. CA.
(2007). Enzymatic inhibition and morphological changes in Hoplias malabaricus from dietary
exposure to lead(II) or methylmercury. Ecotoxicol Environ Saf 67: 82-88.
http://dx.doi.0rg/10.1016/i.ecoenv.2006.03.013
Alves. LC: Glover. CN; Wood. CM. (2006). Dietary Pb accumulation in juvenile freshwater rainbow trout
(Oncorhynchus mykiss). Arch Environ Contam Toxicol 51: 615-625.
http://dx.doi.org/10.1007/s00244-005-0212-7
Alves. LC: Wood. CM. (2006). The chronic effects of dietary lead in freshwater juvenile rainbow trout
(Oncorhynchus mykiss) fed elevated calcium diets. Aquat Toxicol 78: 217-232.
http://dx.doi.0rg/10.1016/i.aquatox.2006.03.005
Amado. EM; Freire. CA; Souza. MM. (2006). Osmoregulation and tissue water regulation in the
freshwater red crab Dilocarcinus pagei (Crustacea, Decapoda), and the effect of waterborne inorganic
lead. Aquat Toxicol 79: 1-8. http://dx.doi.Org/10.1016/i.aquatox.2006.04.003
Ancion. PY; Lear. G: Lewis. GD. (2010). Three common metal contaminants of urban runoff (Zn, Cu &
Pb) accumulate in freshwater biofilm and modify embedded bacterial communities. Environ Pollut
158: 2738-2745. http://dx.doi.Org/10.1016/i.envpol.2010.04.013
Anderson. MB; Preslan. JE; Jolibois. L; Bollinger. JE; George. WJ. (1997). Bioaccumulation of lead
nitrate in red swamp crayfish (Procambrus clarkii). J Hazard Mater 54: 15-29.
http://dx.doi.org/10.1016/S0304-3894(96)01852-3
Anderson. WL; Havera. SP; Zercher. BW. (2000). Ingestion of lead and nontoxic shotgun pellets by ducks
in the Mississippi flyway. J Wildl Manag 64: 848-857.
Angelo. RT; Cringan. MS: Chamberlain. PL; Stahl. AJ: Haslouer. SG: Goodrich. CA. (2007). Residual
effects of lead and zinc mining on freshwater mussels in the Spring River Basin (Kansas, Missouri,
and Oklahoma, USA). Sci Total Environ 384: 467-496.
http://dx.doi.0rg/10.1016/i.scitotenv.2007.05.045
Angelova. VR; Ivanova. RV: Todorov. JM; Ivanov. KI. (2010). Lead, cadmium, zinc, and copper
bioavailability in the soil-plant-animal system in a polluted area. ScientificWorldJournal 10: 273-285.
http://dx.doi.org/10.1100/tsw.2010.33
Anklev. GT; Bennett. RS: Erickson. RJ; Hoff. DJ: Hornung. MW: Johnson. RD; Mount. PR; Nichols.
JW: Russom. CL; Schmieder. PK; Serrrano. JA; Tietge. JE: Villeneuve. PL. (2010). Adverse outcome
pathways: a conceptual framework to support ecotoxicology research and risk assessment [Review].
Environ Toxicol Chem 29: 730-741. http://dx.doi.org/10.1002/etc.34
Anklev. GT; Di Toro. DM; Hansen. DJ: Berry. WJ. (1996). Assessing the ecological risk of metals in
sediments [Editorial]. Environ Toxicol Chem 15: 2053-2055.
http://dx.doi.org/10.1002/etc.562Q151201
Ansaldo. M; Nahabedian. DE; Di Fonzo. C: Wider. EA. (2009). Effect of cadmium, lead and arsenic on
the oviposition, hatching and embryonic survival of Biomphalaria glabrata. Sci Total Environ 407:
1923-1928. http://dx.doi.0rg/10.1016/i.scitotenv.2008.12.001
Ansaldo. M; Nahabedian. DE; Holmes-Brown. E; Agote. M; Ansav. CV: Guerrero. NR; Wider. EA.
(2006). Potential use of glycogen level as biomarker of chemical stress in Biomphalaria glabrata.
Toxicology 224: 119-127. http://dx.doi.Org/10.1016/i.tox.2006.04.037
November 2012 7-251 Draft - Do Not Cite or Quote
-------
Antonious. G: Kochhar. T. (2009). Mobility of heavy metals from soil into hot pepper fruits: Afield study.
Bull Environ Contam Toxicol 82: 59-63. http://dx.doi.org/10.1007/s00128-008-9512-8
Antosiewicz. DM. (2005). Study of calcium-dependent lead-tolerance on plants differing in their level of
Ca-deficiency tolerance. Environ Pollut 134: 23-34. http://dx.doi.Org/10.1016/i.envpol.2004.07.019
Antunes. PM; Berkelaar. EJ; Boyle. D; Hale. BA; Hendershot. W: Voigt A. (2006). The biotic ligand
model for plants and metals: Technical challenges for field application. Environ Toxicol Chem 25:
875-882. http://dx.doi.Org/10.1897/04-586R.l
Arai. T; Maeda. M; Yamakawa. H; Kamatani. A; Miyazaki. N. (2002). Growth effect on the uptake and
elimination of trace metals in the abalones Haliotis. Fish Sci 68: 1094-1098.
http://dx.doi.0rg/10.1046/i.1444-2906.2002.00537.x
Arias-Almeida. JC: Rico-Martinez. R. (2011). Toxicity of cadmium, lead, Mercury and methyl parathion
on euchlanis dilatata ehrenberg 1832 (Rotifera: Monogononta). Bull Environ Contam Toxicol 87:
138-142. http://dx.doi.org/10.1007/s00128-011-0308-x
Arunakumara. K; Zhang. X. (2008). Heavy metal bioaccumulation and toxicity with special reference to
microalgae. J Ocean Univ China 7: 60-64. http://dx.doi.org/10.1007/sll802-008-0060-y
Arunakumara. K; Zhang. X; Song. X. (2008). Bioaccumulation of Pb(2+) and its effects on growth,
morphology and pigment contents of Spirulina (Arthrospira) platensis. J Ocean Univ China 7:
397-403. http://dx.doi.org/10.1007/sll802-008-0397-2
Ashraf. MY; Azhar. N; Ashraf. M; Hussain. M; Arshad. M. (2011). Influence of lead on growth and
nutrient accumulation in canola (Brassica napus L.) cultivars. J Environ Biol 32: 659-666.
Atici. T; Katircioglu. H; Akin. B. (2008). Sensitivity of freshwater microalgal strains (Chlorella vulgaris
Beijerinck and Scenedesmus obliquus (Turpin) Kutzing) to heavy metals. Fresen Environ Bull 17:
268-274.
Atkinson. CA; Jollev. DF; Simpson. SL. (2007). Effect of overlying water pH, dissolved oxygen, salinity
and sediment disturbances on metal release and sequestration from metal contaminated marine
sediments. Chemosphere 69: 1428-1437. http://dx.doi.Org/10.1016/i.chemosphere.2007.04.068
Atli. G: Canli. M. (2007). Enzymatic responses to metal exposures in a freshwater fish Oreochromis
niloticus. Comp Biochem Physiol C Toxicol Pharmacol 145: 282-287.
http://dx.doi.0rg/10.1016/i.cbpc.2006.12.012
Atli. G: Canli. M. (2008). Responses of metallothionein and reduced glutathione in a freshwater fish
Oreochromis niloticus following metal exposures. Environ Toxicol Pharmacol 25: 33-38.
http://dx.doi.0rg/10.1016/i.etap.2007.08.007
ATSDR (Agency for Toxic Substances and Disease Registry). (2004). Guidance manual for the
assessment of joint toxic action of chemical mixtures. Atlanta, GA: U.S. Department of Health and
Human Services, Agency for Toxic Substances and Disease Registry.
http://www.atsdr.cdc.gov/interactionproFiles/IP-ga/ipga.pdf
Ayensu. E; van Claasen. PR; Collins. M; Dearing. A; Fresco. L; Gadgil. M; Gitav. H; Glaser. G: Juma. C:
Krebs. J: Lenton. R; Lubchenco. J: McNeely. JA; Moonev. HA; Pinstrup-Andersen. P; Ramos. M;
Raven. P; Reid. WV: Samper. C: Sarukhan. J: Schei. P; Tundisi. JG: Watson. RT; Guanhua. X; Zakri.
AH. (1999). International ecosystem assessment. Science 286: 685-686.
http://dx.doi.org/10.1126/science.286.5440.685
Aznar. JC: Richer-Lafleche. M; Begin. C: Rodrigue. R. (2008a). Spatiotemporal reconstruction of lead
contamination using tree rings and organic soil layers. Sci Total Environ 407: 233-241.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.09.044
Aznar. JC: Richer-Laflechea. M; Begin. C: Begin. Y. (2009a). Lead exclusion and copper translocation in
black spruce needles. Water Air Soil Pollut 203: 139-145.
http://dx.doi.org/10.1007/sll270-009-9997-8
November 2012 7-252 Draft - Do Not Cite or Quote
-------
Aznar. JC: Richer-Laflechea. M; Paucar-Munoza. H; Bordeleaua. M; Begina. Y. (2009b). Is tree growth
reduction related to direct foliar injuries or soil chemistry modifications? Chemosphere 76:
1366-1371. http://dx.doi.0rg/10.1016/i.chemosphere.2009.06.023
Baath. E. (1989). Effects of heavy metals in soil on microbial processes and populations (a review)
[Review]. Water Air Soil Pollut 47: 335-379. http://dx.doi.org/10.1007/BF00279331
Baath. E; Diaz-Ravina. M; Bakken. LR. (2005). Microbial biomass, community structure and metal
tolerance of a naturally Pb-enriched forest soil. Microb Ecol 50: 496-505.
http://dx.doi.org/10.1007/s00248-005-0008-3
Balistrieri. LS: Blank. RG. (2008). Dissolved and labile concentrations of Cd, Cu, Pb, and Zn in the South
Fork Coeur d'Alene River, Idaho: Comparisons among chemical equilibrium models and implications
for biotic ligand models. Appl Geochem 23: 3355-3371.
http://dx.doi.0rg/10.1016/i.apgeochem.2008.06.031
Bamborough. L; Cummings. SP (2009). The impact of zinc and lead concentrations and seasonal
variation on bacterial and actinobacterial community structure in a metallophytic grassland soil. Folia
Microbiol (Praha) 54: 327-334. http://dx.doi.org/10.1007/sl2223-009-0042-5
Baos. R; Bias. J: Bortolotti. GR: Marchant. TA; Hiraldo. F. (2006). Adrenocortical response to stress and
thyroid hormone status in free-living nestling white storks (Ciconia ciconia) exposed to heavy metal
and arsenic contamination. Environ Health Perspect 114: 1497-1501.
http://dx.doi.org/10.1289/ehp.9099
Barnthouse. LW: Munns. RM. Jr; Sorensen. MT. (2008). Population-level ecological risk assessment.
Pensacola, FL: Taylor & Francis, http://www.crcpress.com/product/isbn/9781420053326
Bartell. SM. (2007). Ecosystem effects modeling. In GW Suterll (Ed.), Ecological risk assessment (2nd
ed., pp. 413-432). Boca Raton, FL: CRC Press.
Baumann. HA; Morrison. L; Stengel. DB. (2009). Metal accumulation and toxicity measured by PAM-
Chlorophyll fluorescence in seven species of marine macroalgae. Ecotoxicol Environ Saf 72:
1063-1075. http://dx.doi.0rg/10.1016/i.ecoenv.2008.10.010
Bazar. MA; Quinn. MJ; Mozzachio. K; Bleiler. JA; Archer. CR; Phillips. CT; Johnson. MS. (2010).
Toxicological responses of red-backed salamander (plethodon cinereus) exposed to aged and amended
soils containing lead. Arch Environ Contam Toxicol 58: 1040-1047.
http://dx.doi.org/10.1007/s00244-010-9471-z
Bechard. KM; Gillis. PL; Wood. CM. (2008). Acute toxicity of waterborne Cd, Cu, Pb, Ni, and Zn to
first-instar Chironomus riparius larvae. Arch Environ Contam Toxicol 54: 454-459.
http://dx.doi.org/10.1007/s00244-007-9048-7
Beebv. A; Richmond. L. (2010). Magnesium and the regulation of lead in three populations of the garden
snail Cantareus aspersus. Environ Pollut 158: 2288-2293.
http://dx.doi.0rg/10.1016/i.envpol.2010.02.002
Beiras. R; Albentosa. M. (2003). Inhibition of embryo development of the commercial bivalves Ruditapes
decussatus and Mytilus galloprovincialis by trace metals; implications for the implementation of
seawater quality criteria. Aquaculture 230: 205-213.
http://dx.doi.org/10.1016/S0044-8486(03)00432-0
Bekesiova. B; S. H; Libantova. J: Moravcikova. J: Matusikova. I. (2008). Heavy-metal stress induced
accumulation of chitinase isoforms in plants. Mol Biol Rep 35: 579-588.
http://dx.doi.org/10.1007/sll033-007-9127-x
Bellrose. FC. (1959). Lead poisoning as a mortality factor in waterfowl populations. Bull 111 Nat Hist
Surv 27: 235-288.
Benbrahim. M; Denaix. L; Thomas. AL; Balet. J: Carnus. JM. (2006). Metal concentrations in edible
mushrooms following municipal sludge application on forest land. Environ Pollut 144: 847-854.
http://dx.doi.0rg/10.1016/i.envpol.2006.02.014
November 2012 7-253 Draft - Do Not Cite or Quote
-------
Bengtsson. G: Nordstrom. S: Rundgren. S. (1983). Population density and tissue metal concentration of
lumbricids in forest soils near a brass mill. Environ Pollut Ecol Biol 30: 87-108.
http://dx.doi.org/10.1016/0143-1471(83)90007-7
Sergey. LL; Weis. JS. (2007). Molting as a mechanism of depuration of metals in the fiddler crab, Uca
pugnax. Mar Environ Res 64: 556-562. http://dx.doi.Org/10.1016/i.marenvres.2007.04.009
Berglund. AMM; Ingvarsson. PK; Danielsson. H; Nyholm. NEI. (2010). Lead exposure and biological
effects in pied flycatchers (Ficedula hypoleuca) before and after the closure of a lead mine in northern
Sweden. Environ Pollut 158: 1368-1375. http://dx.doi.Org/10.1016/j.envpol.2010.01.005
Besser. JA; Brumbaugh. WG: Hardestv. DK; Hughes. JP; Ingersoll. CG. (2009). Assessment of metal-
contaminated sediments from the Southeast Missouri (SEMO) mining district using sediment toxicity
tests with amphipods and freshwater mussels. (08-NRDAR-02). Columbia, MO: U.S. Geological
Survey, Columbia Environmental Research Center.
Besser. JM; Brumbaugh. WG: Brunson. EL; Ingersoll. CG. (2005). Acute and chronic toxicity of lead in
water and diet to the amphipod Hyalella azteca. Environ Toxicol Chem 24: 1807-1815.
http://dx.doi.Org/10.1897/04-480R.l
Besser. JM; Brumbaugh. WG: Kemble. NE; May. TW: Ingersoll. CG. (2004). Effects of sediment
characteristics on the toxicity of chromium(III) and chromium(VI) to the amphipod, Hyalella azteca.
Environ Sci Technol 38: 6210-6216. http://dx.doi.org/10.1021/es049715i
Besser. JM: Brumbaugh. WG: May. TW: Schmitt. CJ. (2007). Biomonitoring of lead, zinc, and cadmium
in streams draining lead-mining and non-mining areas, Southeast Missouri, USA. Environ Monit
Assess 129: 227-241. http://dx.doi.org/10.1007/sl0661-006-9356-9
Beyer. WN; Dalgarn. J; Pudding. S: French. JB; Mateo. R; Miesner. J; Sileo. L; Spann. J. (2004). Zinc
and lead poisoning in wild birds in the Tri-State Mining District (Oklahoma, Kansas, and Missouri).
Arch Environ Contam Toxicol 48: 108-117. http://dx.doi.org/10.1007/s00244-004-0010-7
Beyer. WN; Pattee. OH; Sileo. L; Hoffman. DJ; Mulhern. BM. (1985). Metal contamination in wildlife
living near two zinc smelters. Environ Pollut Ecol Biol 38: 63-86.
http://dx.doi.org/10.1016/0143-1471(85)90094-7
Bi. XY: Feng. XB: Yang. YG: Li. XD: Sin. GPY: Oiu. GL: Oian. XL: Li. FL: He. TR: Li. P: Liu. TZ: Fu.
ZY. (2007). Heavy metals in an impacted wetland system: Atypical case from Southwestern China.
Sci Total Environ 387: 257-268. http://dx.doi.Org/10.1016/j.scitotenv.2007.07.059
Bindler. R; Renberg. I; Klaminder. J. (2008). Bridging the gap between ancient metal pollution and
contemporary biogeochemistry. J Paleolimnol 40: 755-770.
http://dx.doi.org/10.1007/sl0933-008-9208-4
Birceanu. O; Chowdhury. MJ; Gillis. PL; McGeer. JC: Wood. CM; Wilkie. MP (2008). Modes of metal
toxicity and impaired branchial ionoregulation in rainbow trout exposed to mixtures of Pb and Cd in
soft water. Aquat Toxicol 89: 222-231. http://dx.doi.Org/10.1016/i.aquatox.2008.07.007
Bojarczuk. K; Kieliszewska-Rokicka. B. (2010). Effect of ectomycorrhiza on Cu and Pb accumulation in
leaves and roots of silver birch (Betula pendula Roth.) seedlings grown in metal-contaminated soil.
Water Air Soil Pollut 207: 227-240. http://dx.doi.org/10.1007/sll270-009-0131-8
Bonanno. G: Lo Giudice. R. (2010). Heavy metal bioaccumulation by the organs of Phragmites australis
(common reed) and their potential use as contamination indicators. Ecol Indicat 10: 639-645.
http://dx.doi.0rg/10.1016/i.ecolind.2009.ll.002
Borgmann. U; Couillard. Y; Doyle. P; Dixon. DG. (2005). Toxicity of sixty-three metals and metalloids to
Hyalella azteca at two levels of water hardness. Environ Toxicol Chem 24: 641-652.
http://dx.doi.Org/10.1897/04-177R.l
Borgmann. U; Couillard. Y; Grapentine. LC. (2007). Relative contribution of food and water to 27 metals
and metalloids accumulated by caged Hyalella azteca in two rivers affected by metal mining. Environ
Pollut 145: 753-765. http://dx.doi.Org/10.1016/i.envpol.2006.05.020
November 2012 7-254 Draft - Do Not Cite or Quote
-------
Borgmann. U; Kramar. O; Loveridge. C. (1978). Rates of mortality, growth, and biomass production of
Lymnaea palustris during chronic exposure to lead. J Fish Res Board Can 35: 1109-1115.
http://dx.doi.org/10.1139/f78-175
Boughriet. A; Proix. N; Billon. G: Recourt. P; Ouddane. B. (2007). Environmental impacts of heavy
metal discharges from a smelter in Deule-canal sediments (Northern France): Concentration levels and
chemical fractionation. Water Air Soil Pollut 180: 83-95. http://dx.doi.org/10.1007/sll270-006-9252-5
Boyle. D; Amlund. H; Lundebve. AK; Hogstrand. C: Bury. NR. (2010). Bioavailability of a natural lead-
contaminated invertebrate diet to zebrafish. Environ Toxicol Chem 29: 708-714.
http://dx.doi.org/10.1002/etc.61
Bradham. KD; Dayton. EA; Basta. NT; Schroder. J; Pavton. M; Lanno. RP. (2006). Effect of soil
properties on lead bioavailability and toxicity to earthworms. Environ Toxicol Chem 25: 769-775.
http://dx.doi.Org/10.1897/04-552R.l
Brix. KV; DeForest. DK; Burger. M; Adams. WJ. (2005). Assessing the relative sensitivity of aquatic
organisms to divalent metals and their representation in toxicity datasets compared to natural aquatic
communities. Hum Ecol Risk Assess 11: 1139-1156. http://dx.doi.org/10.1080/10807030500278628
Brown. CS: Luebbert J: Mulcahy. D; Schamber. J: Rosenberg. PH. (2006). Blood lead levels of wild
Steller's eiders (Polysticta stelleri) and black scoters (Melanitta nigra) in Alaska using a portable blood
lead analyzer. J Zoo Wildl Med 37: 361-365. http://dx.doi.Org/10.1638/05-092.l
Brumbaugh. WG: Schmitt. CJ: May. TW. (2005). Concentrations of cadmium, lead, and zinc in fish from
mining-influenced waters of northeastern Oklahoma: Sampling of blood, carcass, and liver for aquatic
biomonitoring. Arch Environ Contam Toxicol 49: 76-88. http://dx.doi.org/10.1007/s00244-004-0172-3
Bryan. CE; Christopher. SJ: Balmer. BC: Wells. RS. (2007). Establishing baseline levels of trace elements
in blood and skin of bottlenose dolphins in Sarasota Bay, Florida: Implications for non-invasive
monitoring. Sci Total Environ 388: 325-342. http://dx.doi.Org/10.1016/i.scitotenv.2007.07.046
Buekers. J: Redeker. ES: Smolders. E. (2009). Lead toxicity to wildlife: Derivation of a critical blood
concentration for wildlife monitoring based on literature data [Review]. Sci Total Environ 407:
3431-3438. http://dx.doi.Org/10.1016/i.scitotenv.2009.01.044
Burger. J: Campbell. KR; Murray. S: Campbell. TS: Gaines. KF; Jeitner. C: Shukla. T; Burke. S:
Gochfeld. M. (2007). Metal levels in blood, muscle and liver of water snakes (Nerodia spp.) from
New Jersey, Tennessee and South Carolina. Sci Total Environ 373: 556-563.
http://dx.doi.0rg/10.1016/i.scitotenv.2006.06.018
Burger. J: Gochfeld. M. (2005). Effects of lead on learning in herring gulls: An avian wildlife model for
neurobehavioral deficits. Neurotoxicology 26: 615-624. http://dx.doi.Org/10.1016/i.neuro.2005.01.005
Burger. J: Murray. S: Gaines. KF; Novak. JM; Punshon. T; Dixon. C: Gochfeld. M. (2006). Element
levels in snakes in South Carolina: Differences between a control site and exposed site on the
savannah river site. Environ Monit Assess 112: 35-52. http://dx.doi.org/10.1007/sl0661-006-0695-3
Businelli. D: Onofri. A; Massaccesi. L. (2011). Factors involved in uptake of lead by some edible crops
grown in agricultural soils of central Italy. Soil Sci 176: 472-478.
http://dx.doi.org/10.1097/SS.Ob013e31822769fa
Bustamante. P; Bertrand. M; Boucaud-Camou. E; Miramand. P. (2006). Subcellular distribution of Ag,
Cd, Co, Cu, Fe, Mn, Pb, and Zn in the digestive gland of the common cuttlefish Sepia officinalis.
Journal of Shellfish Research 25: 987-993.
Bustamante. P: Miramand. P. (2005). Subcellular and body distributions of 17 trace elements in the
variegated scallop Chlamys variafrom the French coast of the Bay of Biscay. Sci Total Environ 337:
59-73. http://dx.doi.0rg/10.1016/j.scitotenv.2004.07.004
Cade. TJ. (2007). Exposure of California condors to lead from spent ammunition. J Wildl Manag 71:
2125-2133. http://dx.doi.org/10.2193/2007-084
November 2012 7-255 Draft - Do Not Cite or Quote
-------
Caetano. M; Fonseca. N; Carlos Vale. RC. (2007). Mobility of Pb in salt marshes recorded by total
content and stable isotopic signature. Sci Total Environ 380: 84-92.
http://dx.doi.0rg/10.1016/i.scitotenv.2006.ll.026
Camara Pellisso. S: Mufioz. MJ; Carballo. M; Sanchez-Vizcaino. JM. (2008). Determination of the
immunotoxic potential of heavy metals on the functional activity of bottlenose dolphin leukocytes in
vitro. Vet Immunol Immunopathol 121: 189-198. http://dx.doi.Org/10.1016/i.vetimm.2007.09.009
Campos. JA; Tejera. NA. (2011). Bioconcentration factors and trace elements bioaccumulation in
sporocarps of fungi collected from quartzite acidic soils. Biol Trace Elem Res 143: 540-554.
http://dx.doi.org/10.1007/sl2011-010-8853-4
Carine. F; Enrique. AG: Steven. C. (2009). Metal effects on phenol oxidase activities of soils. Ecotoxicol
Environ Saf 72: 108-114. http://dx.doi.Org/10.1016/i.ecoenv.2008.03.008
Carreras. HA; Pignata. ML. (2007). Effects of the heavy metals Cu2+, Ni2+, Pb2+, and Zn2+ on some
physiological parameters of the lichen Usnea amblyoclada. Ecotoxicol Environ Saf 67: 59-66.
http://dx.doi.0rg/10.1016/i.ecoenv.2006.05.005
Carvalho. F; Oliveira. J: Alberto. G. (2011). Factors affecting 210Po and 210Pb activity concentrations in
mussels and implications for environmental bio-monitoring programmes. J Environ Radioact 102:
128-137. http://dx.doi.0rg/10.1016/i.jenvrad.2010.ll.003
Casas. S: Gonzalez. JL; Andral. B; Cossa. D. (2008). Relation between metal concentration in water and
metal content of marine mussels (Mytilus galloprovincialis): Impact of physiology. Environ Toxicol
Chem 27: 1543-1552. http://dx.doi.Org/10.1897/07-418.l
Casteel. SW: Weis. CP; Henningsen. GM; Brattin. WL. (2006). Estimation of relative bioavailability of
lead in soil and soil-like materials using young swine. Environ Health Perspect 114: 1162-1171.
http://dx.doi.org/10.1289/ehp.8852
Cenkci. S: Cioerci. IH; Yildiz. M; Oezav. C: Bozdao. A; Terzi. H. (2010). Lead contamination reduces
chlorophyll biosynthesis and genomic template stability in Brassica rapa L. Environ Exp Bot 67:
467-473. http://dx.doi.0rg/10.1016/i.envexpbot.2009.10.001
Chadwick. EA; Simpson. VR; Nicholls. AE; Slater. FM. (2011). Lead levels in Eurasian otters decline
with time and reveal interactions between sources, prevailing weather, and stream chemistry. Environ
SciTechnol45: 1911-1916. http://dx.doi.org/10.1021/esl034602
Chapin. FS. Ill; Sala. OE; Burke. 1C: Grime. JP; Hooper. DU; Lauenroth. WK; Lombard. A; Moonev.
HA; Mosier. AR; Naeem. S: Pacala. SW: Roy. J: Steffen. WL; Tilman. D. (1998). Ecosystem
consequences of changing biodiversity. Bioscience 48: 45-52.
Chapman. E.. E.; Dave. G..: Murimboh. J.. D. (2010). Ecotoxicological risk assessment of undisturbed
metal contaminated soil at two remote lighthouse sites. Ecotoxicol Environ Saf 73: 961-969.
http://dx.doi.0rg/10.1016/i.ecoenv.2010.02.014
Chaube. R; Mishra. S: Singh. RK. (2010). In vitro effects of lead nitrate on steroid profiles in the post-
vitellogenic ovary of the catfish Heteropneustes fossilis. Toxicol In Vitro 24: 1899-1904.
http://dx.doi.0rg/10.1016/i.tiv.2010.07.021
Chen. TH; Gross. JA; Karasov. WH. (2006b). Sublethal effects of lead on northern leopard frog (Rana
pipiens) tadpoles. Environ Toxicol Chem 25: 1383-1389. http://dx.doi.Org/10.1897/05-356R.l
Chen. WP: Li. LQ; Chang. AC: Wu. LS: Chanev. RL; Smith. R; Aiwa. H. (2009b). Characterizing the
solid-solution partitioning coefficient and plant uptake factor of As, Cd, and Pb in California
croplands. Agric Ecosyst Environ 129: 212-220. http://dx.doi.Org/10.1016/i.agee.2008.09.001
Chen. ZZ; Zhu. L; Wilkinson. KJ. (2010c). Validation of the biotic ligand model in metal mixtures:
Bioaccumulation of lead and copper. Environ Sci Technol 44: 3580-3586.
http://dx.doi.org/10.1021/esl003457
November 2012 7-256 Draft - Do Not Cite or Quote
-------
Chiesa. ME; Rosenberg. CE; Fink. NE; Salibian. A. (2006). Serum protein profile and blood cell counts
in adult toads Bufo arenarum (Amphibia: Anura: Bufonidae): Effects of sublethal lead acetate. Arch
Environ Contam Toxicol 50: 384-391. http://dx.doi.org/10.1007/s00244-004-0252-4
Cho. YS: Bolick. JA; Butcher. DJ. (2009). Phytoremediation of lead with green onions (Allium
fistulosum) and uptake of arsenic compounds by moonlight ferns (Pteris cretica cv Mayii). Microchem
J 91: 6-8. http://dx.doi.0rg/10.1016/i.microc.2008.05.008
Chrastny. V: Komarek. M; Haiek. T. (2010). Lead contamination of an agricultural soil in the vicinity of a
shooting range. Environ Monit Assess 162: 37-46. http://dx.doi.org/10.1007/sl0661-009-0774-3
Church. ME; Gwiazda. R; Risebrough. RW: Sorenson. K; Chamberlain. CP; Parry. S: Heinrich. W:
Rideout. BA; Smith. DR. (2006). Ammunition is the principal source of lead accumulated by
California Condors re-introduced to the wild. Environ Sci Technol 40: 6143-6150.
http://dx.doi.org/10.1021/es060765s
Ciardullo. S: Aureli. F; Coni. E; Guandalini. E; losi. F; Raggi. A; Rufo. G: Cubadda. F. (2008).
Bioaccumulation potential of dietary arsenic, cadmium, lead, mercury, and selenium in organs and
tissues of rainbow trout (Oncorhyncus mykiss) as a function offish growth. J Agric Food Chem 56:
2442-2451. http://dx.doi.org/10.1021/if703572t
Cid. N; Ibanez. C: Palanques. A; Prat. N. (2010). Patterns of metal bioaccumulation in two filter-feeding
macroinvertebrates: Exposure distribution, inter-species differences and variability across
developmental stages. Sci Total Environ 408: 2795-2806.
http://dx.doi.0rg/10.1016/i.scitotenv.2010.03.030
Clark. HF; Brabander. DJ: Erdil. RM. (2006). Sources, sinks, and exposure pathways of lead in urban
garden soil. J Environ Qual 35: 2066-2074. http://dx.doi.org/10.2134/ieq2005.0464
Coeurdassier. M; Scheifler. R; de Vaufleury. A; Crini. N; Saccomani. C: Du Mont. LS: Badot. PM.
(2007). Earthworms influence metal transfer from soil to snails. Appl Soil Ecol 35: 302-310.
http://dx.doi.0rg/10.1016/i.apsoil.2006.08.004
Comino. E; Menegatti. S: Fiorucci. A; Schwitzguebel. JP (2011). Accumulation and translocation
capacity of As, Co, Cr and Pb by forage plants. Agrochimica 55: 105-115.
Company. R; Serafim. A; Lopes. B; Cravo. A; Kalman. J: Riba. I; DelValls. TA; Blasco. J: Delgado. J:
Sarmiento. AM; Nieto. JM; Shepherd. TJ; Nowell. G: Bebianno. MJ. (2011). Source and impact of
lead contamination on delta-aminolevulinic acid dehydratase activity in several marine bivalve species
along the Gulf of Cadiz. Aquat Toxicol 101: 146-154. http://dx.doi.Org/10.1016/i.aquatox.2010.09.012
Conkova. M; Kubiznakova. J. (2008). Lead isotope ratios in tree bark pockets: An indicator of past air
pollution in the Czech Republic. Sci Total Environ 404: 440-445.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.04.025
Costanza. R: d'Arge. R: De Groot. R: Farber. S: Grasso. M; Hannon. B; Limburg. K; Naeem. S: O'Neill.
RV; Paruelo. J: Raskin. RG: Sutton. P; Van Den Belt. M. (1997). The value of the world's ecosystem
services and natural capital. Nature 387: 253-259. http://dx.doi.org/10.1038/387253aO
Couture. RM; J-F. C: Auger. D; Claisse. D; Gobeil. C: Cossa. D. (2010). Seasonal and decadal variations
in lead sources to eastern North Atlantic mussels. Environ Sci Technol 44: 1211-1216.
http://dx.doi.org/10.1021/es902352z
Cravo. A; Lopes. B; Serafim. A; Company. R; Barreira. L; Gomes. T; Bebianno. MJ. (2009). A
multibiomarker approach in Mytilus galloprovincialis to assess environmental quality. J Environ
Monit 11: 1673-1686. http://dx.doi.org/10.1039/b909846a
Croisetiere. L; Hare. L; Tessier. A. (2006). Afield experiment to determine the relative importance of prey
and water as sources of As, Cd, Co, Cu, Pb, and Zn for the aquatic invertebrate Sialis velata. Environ
Sci Technol 40: 873-879. http://dx.doi.org/10.1021/es0516209
November 2012 7-257 Draft - Do Not Cite or Quote
-------
Cui. S: Zhou. QX; Chao. L. (2007). Potential hyperaccumulation of Pb, Zn, Cu and Cd in endurant plants
distributed in an old smeltery, northeast China. Environ Geol 51: 1043-1048.
http://dx.doi.org/10.1007/s00254-006-0373-3
Currie. M; Hodson. ME; Arnold. RE; Langdon. CJ. (2005). Single versus multiple occupancy: Effects on
toxicity parameters measured on Eisenia fetida in lead nitrate-treated soil. Environ Toxicol Chem 24:
110-116. http://dx.doi.0rg/10.1897/03-686.l
Cutter. BE; Guvette. RP (1993). Anatomical, chemical, and ecological factors affecting tree species
choice in dendrochemistry studies. J Environ Qual 22: 611-619.
httrj://dx.doi.org/10.2134/ieal993.00472425002200030028x
Dai. W: Du. H; Fu. L; Jin. C: Xu. Z; Liu. H. (2009a). Effects of dietary Pb on accumulation,
histopathology, and digestive enzyme activities in the digestive system of tilapia (Oreochromis
niloticus). Biol Trace Elem Res 127: 124-131. http://dx.doi.org/10.1007/sl2011-008-8227-3
Dai. W: Fu. L; Du. H; Jin. C: Xu. Z. (2009b). Changes in growth performance, metabolic enzyme
activities, and content of Fe, Cu, and Zn in liver and kidney of tilapia (Oreochromis niloticus) exposed
to dietary Pb. Biol Trace Elem Res 128: 176-183. http://dx.doi.org/10.1007/sl2011-008-8259-8
Daily. GC. (1997). Introduction: What are ecosystem services? In GC Daily (Ed.), Nature's services:
Societal dependence on natural ecosystems (pp. 1-10). Washington, DC: Island Press.
Daily. GC: Ehrlich. PR. (1999). Managing earth's ecosystems: An interdisciplinary challenge. Ecosystems
2: 277-280.
Das. M; Maiti. SK. (2007). Metal accumulation in A. baccifera growing naturally on abandoned copper
tailings pond. Environ Monit Assess 127: 119-125. http://dx.doi.org/10.1007/sl0661-006-9265-y
Dauwe. T; Snoeijs. T; Bervoets. L; Blust. R; Eens. M. (2006). Calcium availability influences lead
accumulation in a passerine bird. Animal Biology 56: 289-298.
http://dx.doi.org/10.1163/157075606778441822
Dayton. EA; Basta. NT; Pavton. ME; Bradham. KD; Schroder. JL; Lanno. RP. (2006). Evaluating the
contribution of soil properties to modifying lead phytoavailability and phytotoxicity. Environ Toxicol
Chem 25: 719-725. http://dx.doi.Org/10.1897/05-307R.l
Dazy. M; Masfaraud. JF; Ferard. JF. (2009). Induction of oxidative stress biomarkers associated with
heavy metal stress in Fontinalis antipyretica Hedw. Chemosphere 75: 297-302.
http://dx.doi.0rg/10.1016/i.chemosphere.2008.12.045
De Boeck. G: Eyckmans. M; Lardon. I; Bobbaers. R; Sinha. AK; Blust. R. (2010). Metal accumulation
and metallothionein induction in the spotted dogfish Scyliorhinus canicula. Cornp Biochem Physiol A
Mol Integr Physiol 155: 503-508. http://dx.doi.Org/10.1016/i.cbpa.2009.12.014
de Groot. AC: Peijnenburg. WJG. M; van den Hoop. MAG. T; Ritsema. R: van Veen. RPM. (1998).
Zware metalen in Nederlandse veldbodems: een experimentele en theoretische studie over
evenwichtspartitie. (607220001). Bilthoven, The Netherlands: National Institute of Public Health and
the Environment. http://www.rivm.n^ibliotheek/rapporten/607220001.html
De Jonge. M..; Eyckmans. M..; Blust. R..: Bervoets. L.. (2011). Are accumulated sulfide-bound metals
metabolically available in the benthic oligochaete Tubifex tubifex. Environ Sci Technol 45:
3131-3137. http://dx.doi.org/10.1021/esl037595
De Jonge. M; Blust. R: Bervoets. L. (2010). The relation between acid volatile sulfides (AVS) and metal
accumulation in aquatic invertebrates: Implications of feeding behavior and ecology. Environ Pollut
158: 1381-1391. http://dx.doi.0rg/10.1016/i.envpol.2010.01.001
De Jonge. M; Dreesen. F; De Paepe. J: Blust. R: Bervoets. L. (2009). Do acid volatile sulfides (AVS)
influence the accumulation of sediment-bound metals to benthic invertebrates under natural field
conditions? Environ Sci Technol 43: 4510-4516. http://dx.doi.org/10.1021/es8034945
November 2012 7-258 Draft - Do Not Cite or Quote
-------
de Vaufleury. A; Coeurdassier. M; Pandard. P; Scheifler. R; Low. C: Crini. N; Badot PM. (2006). How
terrestrial snails can be used in risk assessment of soils. Environ Toxicol Chem 25: 797-806.
http://dx.doi.Org/10.1897/04-560R.l
de Vries. W: Groenenberg. JE. (2009). Evaluation of approaches to calculate critical metal loads for forest
ecosystems. Environ Pollut 157: 3422-3432. http://dx.doi.Org/10.1016/j.envpol.2009.06.021
de Vries. W: Lofts. S: Tipping. E; Meili. M; Groenenberg. JE; Schiitze. G. (2007). Impact of soil
properties on critical concentrations of cadmium, lead, copper, zinc, and mercury in soil and soil
solution in view of ecotoxicological effects [Review]. Rev Environ Contam Toxicol 191: 47-89.
http://dx.doi.org/10.1007/978-0-387-69163-3 3
DeAngelis. PL; Bartell. SM; Brenkert. AL. (1989). Effects of nutrient recycling and food-chain length on
resilience. Am Nat 134: 778-805.
Debelius. B; Forja. JM; DelValls. A; Lubian. LM. (2009). Toxicity and bioaccumulation of copper and
lead in five marine microalgae. Ecotoxicol Environ Saf 72: 1503-1513.
http://dx.doi.0rg/10.1016/i.ecoenv.2009.04.006
DeForest. DK; Brix. KV; Adams. WJ. (2007). Assessing metal bioaccumulation in aquatic environments:
The inverse relationship between bioaccumulation factors, trophic transfer factors and exposure
concentration. Aquat Toxicol 84: 236-246. http://dx.doi.Org/10.1016/i.aquatox.2007.02.022
Del Toro. I; Floyd. K; Gardea-Torresdev. J: Borrok. D. (2010). Heavy metal distribution and
bioaccumulation in Chihuahuan Desert Rough Harvester ant (Pogonomyrmex rugosus) populations.
Environ Pollut 158: 1281-1287. http://dx.doi.Org/10.1016/i.envrjol.2010.01.024
Delistratv. D; Yokel. J. (In Press) Ecotoxicological study of arsenic and lead contaminated soils in former
orchards at the Hanford site, USA. Environ Toxicol. http://dx.doi.org/10.1002/tox.20768
Deng. H; Ye. ZH; Wong. MH. (2009). Lead, zinc and iron (Fe2+) tolerances in wetland plants and relation
to root anatomy and spatial pattern of ROL. Environ Exp Bot 65: 353-362.
http://dx.doi.0rg/10.1016/i.envexpbot.2008.10.005
Desrosiers. M; Gagnon. C: Masson. S: Martel. L; Babut. MP (2008). Relationships among total
recoverable and reactive metals and metalloid in St. Lawrence River sediment: Bioaccumulation by
chironomids and implications for ecological risk assessment. Sci Total Environ 389: 101-114.
http://dx.doi.0rg/10.1016/i.scitotenv.2007.08.019
Devall. MS: Thien. LB; Ellgaard. E; Flowers. G. (2006). Lead transport into Bayou Trepagnier wetlands
in Louisiana, USA. J Environ Qua! 35: 758-765. http://dx.doi.org/10.2134/jeq2005.0217
Pi Toro. DM; Allen. HE; Bergman. HL; Meyer. JS: Paquin. PR; Santore. RC. (2001). Biotic ligand model
of the acute toxicity of metals. 1. Technical basis. Environ Toxicol Chem 20: 2383-2396.
http://dx.doi.org/10.1002/etc.5620201034
Pi Toro. DM; Mahonev. JD; Hansen. DJ; Scott. KJ; Carlson. AR. (1992). Acid volatile sulfide predicts
the acute toxicity of cadmium and nickel in sediments. Environ Sci Technol 26: 96-101.
http://dx.doi.org/10.1021/es00025a009
Pi Toro. DM; McGrath. JA; Hansen. DJ: Berry. WJ: Paquin. PR; Mathew. R; Wu. KB; Santore. RC.
(2005). Predicting sediment metal toxicity using a sediment biotic ligand model: Methodology and
initial application. Environ Toxicol Chem 24: 2410-2427. http://dx.doi.Org/10.1897/04-413r.l
Dickinson. NM; Baker. AJM; Doronila. A; Laidlaw. S: Reeves. RD. (2009). Phytoremediation of
inorganics: Realism and synergies. Int J Phytoremediation 11: 97-114.
http://dx.doi.org/10.1080/15226510802378368
Dirilgen. N.. (2011). Mercury and lead: Assessing the toxic effects on growth and metal accumulation by
Lemnaminor. Ecotoxicol Environ Saf 74: 48-54. http://dx.doi.Org/10.1016/i.ecoenv.2010.09.014
Dogan. M; Savgideger. SD; Colak. U. (2009). Effect of lead toxicity on aquatic macrophyte Elodea
canadensis Michx. Bull Environ Contam Toxicol 83: 249-254.
http://dx.doi.org/10.1007/s00128-009-9733-5
November 2012 7-259 Draft - Do Not Cite or Quote
-------
Douav. F; Pruvot. C: Waterlot C: Fritsch. C: Fourrier. H; Loriette. A; Bidar. G: Grand. C: de Vaufleury.
A; Scheifler. R. (2009). Contamination of woody habitat soils around a former lead smelter in the
North of France. Sci Total Environ 407: 5564-5577. http://dx.doi.Org/10.1016/j.scitotenv.2009.06.015
Douglas-Stroebel. E; Hoffman. DJ; Brewer. GL; Sileo. L. (2004). Effects of lead-contaminated sediment
and nutrition on mallard duckling brain growth and biochemistry. Environ Pollut 131: 215-222.
http://dx.doi.0rg/10.1016/i.envpol.2004.02.002
Du. M; Wang. D. (2009). The neurotoxic effects of heavy metal exposure on GABAergic nervous system
in nematode Caenorhabditis elegans. Environ Toxicol Pharmacol 27: 314-320.
httrj://dx.doi.org/10.1016/i.etap.2008.11.011
Duman. F; Cicek. M; Sezen. G. (2007). Seasonal changes of metal accumulation and distribution in
common club rush (Schoenoplectus lacustris) and common reed (Phragmites australis). Ecotoxicology
16: 457-463. http://dx.doi.org/10.1007/sl0646-007-0150-4
Duman. F; Obali. O: Demirezen. D. (2006). Seasonal changes of metal accumulation and distribution in
shining pondweed (Potamogeton lucens). Chemosphere 65: 2145-2151.
http://dx.doi.0rg/10.1016/i.chemosphere.2006.06.036
Dwivedi. S: Srivastava. S: Mishra. S: Dixit B; Kumar. A; Tripathi. RD. (2008). Screening of native plants
and algae growing on fly-ash affected areas near National Thermal Power Corporation, Tanda, Uttar
Pradesh, India for accumulation of toxic heavy metals. J Hazard Mater 158: 359-365.
http://dx.doi.0rg/10.1016/i.jhazmat.2008.01.081
Dzubai. A; Backor. M; Tomko. J; Peli. E; Tuba. Z. (2008). Tolerance of the lichen Xanthoria parietina (L.)
Th. Fr. to metal stress. Ecotoxicol Environ Saf 70: 319-326.
http://dx.doi.0rg/10.1016/i.ecoenv.2007.04.002
Ebenso. IE; Ologhobo. AD. (2009a). Effects of lead pollution against juvenile Achatina achatina fed on
contaminated artificial diet. Bull Environ Contam Toxicol 82: 583-585.
httrj://dx.doi.org/10.1007/s00128-008-9625-0
Ebenso. IE; Ologhobo. AD. (2009b). Effects of lead pollution at industrial contaminated sites on sentinel
juvenile Achatina achatina. Bull Environ Contam Toxicol 82: 106-110.
http://dx.doi.org/10.1007/s00128-008-9525-3
Ebrahimpour. M; Mushrifah. I. (2009). Variation and correlations of selected heavy metals in sediment
and aquatic plants in Tasik Chini, Malaysia. Environ Geol 57: 823-831.
http://dx.doi.org/10.1007/s00254-008-1362-5
Einsporn. S: Bressling. J: Koehler. A. (2009). Cellular localization of lead using an antibody-based
detection system and enzyme activity changes in the gills and digestive gland of the blue mussel
Mytilus edulis. Environ Toxicol Chem 28: 402-408. http://dx.doi.Org/10.1897/08-174.l
Einsporn. S: Koehler. A. (2008). Immune-localisations (GSSP) of subcellular accumulation sites of
phenanthrene, aroclor 1254 and lead (Pb) in relation to cytopathologies in the gills and digestive gland
of the mussel Mytilus edulis. Mar Environ Res 66: 185-186.
http://dx.doi.0rg/10.1016/i.marenvres.2008.02.053
Eisler. R. (2000). Handbook of chemical risk assessment: Health hazards to humans, plants, and animals,
Volume 1: Metals. Boca Raton, FL: Lewis Publishers.
El-Gendv. K.. S.: Radwan. M.. A.; Gad. A.. F. (2011). Feeding and growth responses of the snail Theba
pisana to dietary metal exposure. Arch Environ Contam Toxicol 60: 272-280.
http://dx.doi.org/10.1007/s00244-010-9542-l
El-Rjoob. AWO: Massadeh. AM; Omari. MN. (2008). Evaluation of Pb, Cu, Zn, Cd, Ni and Fe levels in
Rosmarinus officinalis labaiatae (Rosemary) medicinal plant and soils in selected zones in Jordan.
Environ Monit Assess 140: 61-68. http://dx.doi.org/10.1007/sl0661-007-9847-3
November 2012 7-260 Draft - Do Not Cite or Quote
-------
Epelde. L; Becerril. JM; Barrutia. O: Gonzalez-Oreja. JA; Garbisu. C. (2010). Interactions between plant
and rhizosphere microbial communities in a metalliferous soil. Environ Pollut 158: 1576-1583.
http://dx.doi.0rg/10.1016/i.envpol.2009.12.013
ERG (Eastern Research Group Inc.). (2004). Issue paper on the bioavailability and bioaccumulation of
metals. Lexington, MA.
Erickson. RJ; Mount. PR; Highland. TL; Hockett. JR; Leonard. EN; Mattson. VR; Dawson. TD; Lott.
KG. (2010). Effects of copper, cadmium, lead, and arsenic in a live diet on juvenile fish growth. Can J
Fish Aquat Sci 67: 1816-1826. http://dx.doi.org/10.1139/F10-098
Ernst. G: Zimmermann. S: Christie. P; Frev. B. (2008). Mercury, cadmium and lead concentrations in
different ecophysiological groups of earthworms in forest soils. Environ Pollut 156: 1304-1313.
http://dx.doi.0rg/10.1016/i.envpol.2008.03.002
Esbaugh. A.. J.: Brix. K.. V.; Mager. E.. M.; Grosell. M.. (2011). Multi-linear regression models predict
the effects of water chemistry on acute lead toxicity to Ceriodaphnia dubia and Pimephales promelas.
Comp Biochem Physiol C Toxicol Pharmacol 154: 137-145.
http://dx.doi.0rg/10.1016/i.cbpc.2011.04.006
Ettler. V; Vanek. A; Mihaljevic. M; Bezdicka. P. (2005). Contrasting lead speciation in forest and tilled
soils heavily polluted by lead metallurgy. Chemosphere 58: 1449-1459.
http://dx.doi.0rg/10.1016/i.chemosphere.2004.09.084
Evans. RD; Balch. GC: Evans. HE; Welbourn. PM. (2006). Uptake and elimination of lead, zinc, and
copper by caddisfly larvae (Trichoptera: Hydropsychidae) using stable isotope tracers. Arch Environ
Contain Toxicol 51: 35-42. http://dx.doi.org/10.1007/s00244-005-2080-6
Fairbrother. A; Wenstel. R; Sappington. K; Wood. W. (2007). Framework for metals risk assessment.
Ecotoxicol Environ Saf 68: 145-227. http://dx.doi.Org/10.1016/j.ecoenv.2007.03.015
Farag. AM; Nimick. DA; Kimball. BA; Church. SE; Harper. DP; Brumbaugh. WG. (2007).
Concentrations of metals in water, sediment, biofilm, benthic macroinvertebrates, and fish in the
Boulder River watershed, Montana, and the role of colloids in metal uptake. Arch Environ Contam
Toxicol 52: 397-409. http://dx.doi.org/10.1007/s00244-005-0021-z
Farley. KJ: Carbonaro. RF: Fanelli. CJ: Costanzo. R: Rader. KJ: Pi Toro. DM. (2011). TICKET-UWM: A
coupled kinetic, equilibrium, and transport screening model for metals in lakes. Environ Toxicol Chem
30: 1278-1287. http://dx.doi.org/10.1002/etc.518
Fernandez-Leborans. G: Antonio-Garcia. MT. (1988). Effects of lead and cadmium in a community of
protozoans. Acta Protozool 27: 141-159.
Fernandez Severini. MD; Botte. SE; Hoffmever. MS: Marcovecchio. JE. (2011). Lead concentrations in
zooplankton, water, and paniculate matter of a southwestern Atlantic temperate estuary (Argentina).
Arch Environ Contam Toxicol 61: 243-260. http://dx.doi.org/10.1007/s00244-010-9613-3
Field. PM; Sherrell. RM. (2003). Direct determination of ultra-trace levels of metals in fresh water using
desolvating micronebulization and HR-ICP-MS: Application to Lake Superior waters. J Anal At
Spectrom 18: 254-259. http://dx.doi.org/10.1039/b210628k
Firat. O..: Cogun. H.. Y; Yuzereroghu T.. A.; Gok. G..: Firat. O..: Kargin. F..; Kotemen. Y.. (2011). A
comparative study on the effects of a pesticide (cypermethrin) and two metals (copper, lead) to serum
biochemistry of Nile tilapia, Oreochromis niloticus. Fish Physiol Biochem 37: 657-666.
http://dx.doi.org/10.1007/sl0695-011-9466-3
Fritioff. A; Greger. M. (2006). Uptake and distribution of Zn, Cu, Cd, and Pb in an aquatic plant
Potamogeton natans. Chemosphere 63: 220-227. http://dx.doi.Org/10.1016/i.chemosphere.2005.08.018
Fritsch. C: Giraudoux. P; Coeurdassier. M; Douav. F; Raoul. F; Pruvot. C: Waterlot. C: de Vaufleury. A;
Scheifler. R. (2010). Spatial distribution of metals in smelter-impacted soils of woody habitats:
Influence of landscape and soil properties, and risk for wildlife. Chemosphere 81: 141-155.
http://dx.doi.0rg/10.1016/i.chemosphere.2010.06.075
November 2012 7-261 Draft - Do Not Cite or Quote
-------
Fujiwara. FG: Gomez. PR; Dawidowski. L; Perelman. P; Faggi. A. (2011). Metals associated with
airborne paniculate matter in road dust and tree bark collected in a megacity (Buenos Aires,
Argentina). Ecol Indicat 11: 240-247. http://dx.doi.Org/10.1016/i.ecolind.2010.04.007
Fukunaga. A. .; Anderson. M.. J. (2011). Bioaccumulation of copper, lead and zinc by the bivalves
Macomona liliana and Austrovenus stutchburyi. Exp Mar Bio Ecol 396: 244-252.
http://dx.doi.0rg/10.1016/i.jembe.2010.10.029
Fullmer. CS. (1997). Lead-calcium interactions: Involvement of 1,25-dihydroxyvitamin D. Environ Res
72: 45-55. http://dx.doi.org/10.1006/enrs.1996.3689
Furman. O; Strawn. DG: Heinz. GH; Williams. B. (2006). Risk assessment test for lead bioaccessibility to
waterfowl in mine-impacted soils. J Environ Qua! 35: 450-458.
http://dx.doi.org/10.2134/ieq2005.0316
Gagnon. C: Fisher. NS. (1997). Bioavailability of sediment-bound methyl and inorganic mercury to
marine bivalve. Environ Sci Technol 31: 993-998. http://dx.doi.org/10.1021/es960364k
Gai. N; Yang. Y; Li. T; Yao. J: Wang. F; Chen. H. (2011). Effect of lead contamination on soil microbial
activity measured by microcalorimetry. Chinese Journal of Chemistry 29: 1541-1547.
http://dx.doi.org/10.1002/cjoc.201180277
Gal. J: Markiewicz-Patkowska. J: Hursthouse. A; Tamer. P. (2008). Metal uptake by woodlice in urban
soils. Ecotoxicol Environ Saf 69: 139-149. http://dx.doi.Org/10.1016/i.ecoenv.2007.01.002
Gallagher. FJ; Pechmann. I; Bogden. JD; Grabosky. J: Weis. P. (2008). Soil metal concentrations and
productivity of Betula populifolia (gray birch) as measured by field spectrometry and incremental
annual growth in an abandoned urban brownfield in New Jersey. Environ Pollut 156: 699-706.
htrp://dx.doi.org/10.1016/i.envpol.20Q8.06.013
Gambrell. RP. (1994). Trace and toxic metals in wetlands: Areview. J Environ Qual 23: 883-891.
http://dx.doi.org/10.2134/ieql994.00472425002300050005x
Gandhi, N,.; Bhavsar, S,. R; Diamond, M,. L. (2011). Critical load analysis in hazard assessment of
metals using a Unit World Model. Environ Toxicol Chem 30: 2157-2166.
http://dx.doi.org/10.1002/etc.606
Gandois. L; Probst. A; Dumat C. (2010). Modelling trace metal extractability and solubility in French
forest soils by using soil properties. Eur J Soil Sci 61: 271-286.
http://dx.doi.0rg/10.llll/i.1365-2389.2009.01215.x
Gao. Y; Zhou. P; Mao. L; Zhi. YE; Shi. WJ. (201 Ob). Assessment of effects of heavy metals combined
pollution on soil enzyme activities and microbial community structure: modified ecological
doseresponse model and PCR-RAPD. Environmental Earth Sciences 60: 603-612.
http://dx.doi.org/10.1007/sl2665-009-0200-8
Garcia-Garcia. G: Nandini. S: Sarma. SSS. (2006). Turbidity mitigates lead toxicity to cladocerans
(Cladocera). Ecotoxicology 15: 425-436. http://dx.doi.org/10.1007/sl0646-006-0064-6
Garcia-Garcia. G: Picazo-Paez. EA; Nandini. S: Sarma. SSS. (2007). Combined effects of sediment and
lead (PbC12) on the demography of Brachionus patulus (Rotifera: Brachionidae). Hydrobiologia 593:
209-218. http://dx.doi.org/10.1007/sl0750-007-9039-8
Garcia-Rico. L..; Tejeda-Valenzuela. L..; Burgos-Hernandez. A.. (2010). Seasonal variations in the
concentrations of metals in Crassostrea corteziensis from Sonora, Mexico. Bull Environ Contam
Toxicol 85: 209-213. http://dx.doi.org/10.1007/s00128-010-0055-4
Garg. P; Satva. S: Sharma. S. (2009). Effect of heavy metal supplementation on local (Allolobophora
parva) and exotic (Eisenia fetida) earthworm species: A comparative study. J Environ Sci Health A
Tox Hazard Subst Environ Eng 44: 1025-1032. http://dx.doi.org/10.1080/10934520902996997
Gaur. JP; Noraho. N; Chauhan. YS. (1994). Relationship between heavy metal accumulation and toxicity
in Spirodela polyrhiza (L) Schleid and Azolla pinnata R Br. Aquat Bot 49: 183-192.
November 2012 7-262 Draft - Do Not Cite or Quote
-------
Gebologlu. N: Cetin. SC: Ece. A; Yilmaz. E; Elmastas. M. (2005). Assessment of lead and cadmium
contents of tomatoes and beans grown in the vicinity of highway of Tokat, Turkey. Asian J Chem 17:
730-736.
Gedan. K; Silliman. BR; Bertness. MD. (2009). Centuries of human-driven change in salt marsh
ecosystems [Review]. Ann Rev Mar Sci 1: 117-141.
http://dx.doi.org/10.1146/annurev.marine.010908.163930
Gitav. H; Brown. S: Easterling. W: Jallow. B. (2001). Ecosystems and their goods and services. In
Climate change 2001: Impacts, adaptation and vulnerability: Contribution of Working Group II to the
third assessment report of the Intergovernmental Panel on Climate Change. Cambridge, United
Kingdom: Cambridge University Press.
Giusi. G: Alo. R; Crudo. M; Facciolo. RM; Canonaco. M. (2008). Specific cerebral heat shock proteins
and histamine receptor cross-talking mechanisms promote distinct lead-dependent neurotoxic
responses in teleosts. Toxicol Appl Pharmacol 227: 248-256.
http://dx.doi.0rg/10.1016/i.taap.2007.10.018
Gnandi. K; Tchangbedji. G: Killi. K; Baba. G: Abbe. K. (2006). The impact of phosphate mine tailings on
the bioaccumulation of heavy metals in marine fish and crustaceans from the coastal zone of Togo.
Mine Water Environ 25: 56-62. http://dx.doi.org/10.1007/sl0230-006-0108-4
Gomez-Ramirez. P..; Martinez-Lopez. E..; Maria-Mojica. P..: Leon-Ortega. M..; Garcia-Fernandez. A..
1 (2011). Blood lead levels and 5-ALAD inhibition in nestlings of Eurasian Eagle Owl (Bubo bubo)
to assess lead exposure associated to an abandoned mining area. Ecotoxicology 20: 131-138.
http://dx.doi.org/10.1007/sl0646-010-0563-3
Gopal. R; Rizvi. AH. (2008). Excess lead alters growth, metabolism and translocation of certain nutrients
in radish. Chemosphere 70: 1539-1544. http://dx.doi.Org/10.1016/i.cheniosphere.2007.08.043
Gopalakrishnan. S: Thilagam. H; Raja. PV. (2007). Toxicity of heavy metals on embryogenesis and larvae
of the marine sedentary polychaete Hydroides elegans. Arch Environ Contam Toxicol 52: 171-178.
http://dx.doi.org/10.1007/s00244-006-0038-v
Gopalakrishnan. S: Thilagam. H; Raja. PV. (2008). Comparison of heavy metal toxicity in life stages
(spermiotoxicity, egg toxicity, embryotoxicity and larval toxicity) of Hydroides elegans. Chemosphere
71: 515-528. http://dx.doi.Org/10.1016/i.chemosphere.2007.09.062
Gorski. J; Nugegoda. D. (2006). Sublethal toxicity of trace metals to larvae of the blacklip abalone,
Haliotis rubra. Environ Toxicol Chem 25: 1360-1367. http://dx.doi.Org/10.1897/05-060R.l
Goriir. G. (2007). Effects of host plant contaminated with heavy metals on the life history traits of aphids
(Brevicoryne brassicae L.). Polish Journal of Ecology 55: 113-120.
Goto. D; Wallace. WG. (2010). Metal intracellular partitioning as a detoxification mechanism for
mummichogs (Fundulus heteroclitus) living in metal-polluted salt marshes. Mar Environ Res 69:
163-171. http://dx.doi.0rg/10.1016/i.marenvres.2009.09.008
Goulder. LH; Kennedy. D. (1997). Valuing ecosystem services: Philosophical bases and empirical
methods. In GC Daily (Ed.), Nature's Services: Societal Dependence on Natural Ecosystems (pp.
23-47). Washington, DC: Island Press.
Groenenberg. JE; Koopmans. GF; Comans. RNJ. (2010). Uncertainty analysis of the nonideal competitive
adsorption-Donnan model: Effects of dissolved organic matter variability on predicted metal
speciation in soil solution. Environ Sci Technol 44: 1340-1346. http://dx.doi.org/10.1021/es902615w
Grosell. M; Brix. KV (2009). High net calcium uptake explains the hypersensitivity of the freshwater
pulmonate snail, Lymnaea stagnalis, to chronic lead exposure. Aquat Toxicol 91: 302-311.
http://dx.doi.0rg/10.1016/i.aquatox.2008.10.012
Grosell. M; Gerdes. R: Brix. KV. (2006a). Influence of Ca, humic acid and pH on lead accumulation and
toxicity in the fathead minnow during prolonged water-borne lead exposure. Comp Biochem Physiol
C Toxicol Pharmacol 143: 473-483. http://dx.doi.Org/10.1016/i.cbpc.2006.04.014
November 2012 7-263 Draft - Do Not Cite or Quote
-------
Grosell. M; Gerdes. RM; Brix. KV. (2006b). Chronic toxicity of lead to three freshwater invertebrates -
Brachionus calyciflorus, Chironomus tentans, and Lymnaea stagnalis. Environ Toxicol Chem 25:
97-104. http://dx.doi.Org/10.1897/04-654R.l
Guillen. J; Baeza. A; Ontalba. MA; Miguez. MR (2009). Pb-210 and stable lead content in fungi: Its
transfer from soil. Sci Total Environ 407: 4320-4326.
http://dx.doi.0rg/10.1016/i.scitotenv.2009.03.025
Quo. XY; Zhang. SZ; Shan. XQ; Luo. L; Pel ZG: Zhu. YG: Liu. T; Xie. YN; Gault. A. (2006b).
Characterization of Pb, Cu, and Cd adsorption on paniculate organic matter in soil. Environ Toxicol
Chem 25: 2366-2373. http://dx.doi.Org/10.1897/05-636R.l
Guo. YL; Yang. YC: Wang. DY. (2009). Induction of reproductive deficits in nematode Caenorhabditis
elegans exposed to metals at different developmental stages. Reprod Toxicol 28: 90-95.
http://dx.doi.0rg/10.1016/i.reprotox.2009.03.007
Gupta. DK; Huang. HG: Yang. XE; Razafindrabe. BHN; Inouhe. M. (2010). The detoxification of lead in
Sedum alfredii H. is not related to phytochelatins but the glutathione. J Hazard Mater 177: 437-444.
http://dx.doi.0rg/10.1016/i.jhazmat.2009.12.052
Guvette. RP; Cutter. BE; Henderson. GS. (1991). Long-term correlations between mining activity and
levels of lead and cadmium in tree-rings of eastern red-cedar. J Environ Qua! 20: 146-150.
http://dx.doi.org/10.2134/ieql991.00472425002000010022x
Gvedu-Ababio. TK; Baird. D. (2006). Response of meiofauna and nematode communities to increased
levels of contaminants in a laboratory microcosm experiment. Ecotoxicol Environ Saf 63: 443-450.
http://dx.doi.0rg/10.1016/i.ecoenv.2005.01.010
Ha. MH; Choi. J. (2009). Effects of environmental contaminants on hemoglobin gene expression in
Daphnia magna: A potential biomarker for freshwater quality monitoring. Arch Environ Contain
Toxicol 57: 330-337. http://dx.doi.org/10.1007/s00244-007-9079-0
Ha. NTH: Sakakibara. M; Sano. S: Hori. RS: Sera. K. (2009). The potential of Eleocharis acicularis for
phytoremediation: Case study at an abandoned mine site. CLEAN - Soil, Air, Water 37: 203-208.
http://dx.doi.org/10.1002/clen.200900009
Hall. JR; Ashmore. M; Fawehinmi. J: Jordan. C: Lofts. S: Shotbolt. L; Spurgeon. DJ; Svendsen. C:
Tipping. E. (2006). Developing a critical load approach for national risk assessments of atmospheric
metal deposition. Environ Toxicol Chem 25: 883-890. http://dx.doi.Org/10.1897/04-571R.l
Han. YL; Huang. SZ; Gu. JG: Qiu. S: Chen. JM. (2008). Tolerance and accumulation of lead by species
of Iris L. Ecotoxicology 17: 853-859. http://dx.doi.org/10.1007/sl0646-008-0248-3
Hansen. JA; Audet. D; Spears. BL; Healy. KA; Brazzle. RE; Hoffman. DJ: Dailev. A; Beyer. WN.
(2011 a). Lead exposure and poisoning of songbirds using the Coeur d'Alene River Basin, Idaho, USA.
Integr Environ Assess Manag 7: 587-595. http://dx.doi.org/10.1002/ieam.201
Hassan. R; Scholes. R; Ash. N. (2005). Ecosystems and human well-being: Current state and trends.
Washington, DC: Island Press.
Heier. LS: Lien. IB; Stromseng. AE; Ljones. M; Rosseland. BO; Tollefsen. KE; Salbu. B. (2009).
Speciation of lead, copper, zinc and antimony in water draining a shooting range: Time dependant
metal accumulation and biomarker responses in brown trout (Salmo trutta L.). Sci Total Environ 407:
4047-4055. http://dx.doi.Org/10.1016/i.scitotenv.2009.03.002
Henczova. M; Deer. AK; Filla. A; Komlosi. V: Mink. J. (2008). Effects of Cu(2+) and Pb(2+) on different
fish species: Liver cytochrome P450-dependent monooxygenase activities and FTIR spectra. Comp
Biochem Physiol C Toxicol Pharmacol 148: 53-60. http://dx.doi.Org/10.1016/j.cbpc.2008.03.010
Hirsch. HVB; Possidente. D: Possidente. B. (2010). Pb2+: An endocrine disrupter in Drosophila?
[Review]. Physiol Behav 99: 254-259. http://dx.doi.Org/10.1016/i.physbeh.2009.09.014
November 2012 7-264 Draft - Do Not Cite or Quote
-------
Hong. FS: Wu. C: Liu. C: Wang. L; Gao. FQ; Yang. F; Xu. JH; Liu. T; Xie. YN; Li. XZ. (2007a). Direct
evidence for interaction between lead ions and kidney DNAfrom silver crucian carp. Chemosphere
68: 1442-1446. http://dx.doi.Org/10.1016/i.chemosphere.2007.04.020
Hooper. DU; Vitousek. PM. (1997). The effects of plant composition and diversity on ecosystem
processes. Science 277: 1302-1305. http://dx.doi.org/10.1126/science.277.5330.1302
Horng. CY; Wang. SL; Cheng. IJ. (2009). Effects of sediment-bound Cd, Pb, and Ni on the growth,
feeding, and survival of Capitella sp I. Exp Mar Bio Ecol 371: 68-76.
http://dx.doi.0rg/10.1016/i.jembe.2009.01.008
Horvat. T; Vidakovic-Cifrek. Z; Orescanin. V; Tkalec. M; Pevalek-Kozlina. B. (2007). Toxicity
assessment of heavy metal mixtures by Lemna minor L. Sci Total Environ 384: 229-238.
http://dx.doi.0rg/10.1016/i.scitotenv.2007.06.007
Hou. JL; Zhuang. P; Zhang. LZ; Feng. L; Zhang. T; Liu. JY; Feng. GP (2011). Morphological
deformities and recovery, accumulation and elimination of lead in body tissues of Chinese sturgeon,
Acipenser sinensis, early life stages: A laboratory study. Journal of Applied Ichthyology (Print) 27:
514-519. http://dx.doi.0rg/10.llll/i.1439-0426.2011.01703.x
Hovmand. MF; Nielsen. SP: Johnsen. I. (2009). Root uptake of lead by Norway spruce grown on 210Pb
spiked soils. Environ Pollut 157: 404-409. http://dx.doi.Org/10.1016/i.envpol.2008.09.038
Hu. Q; Qi. HY; Zeng. JH; Zhang. HX. (2007b). Bacterial diversity in soils around a lead and zinc mine. J
Environ Sci 19: 74-79. http://dx.doi.org/10.1016/S1001-0742(07)60012-6
Hu. X; Ding. Z. (2009). Lead/cadmium contamination and lead isotopic ratios in vegetables grown in
peri-urban and mining/smelting contaminated sites in Nanjing, China. Bull Environ Contam Toxicol
82: 80-84. http://dx.doi.org/10.1007/s00128-008-9562-y
Hu. X; Hu. C: Sun. X; Lu. M; Su. B; Cao. A. (2009b). Effects of simulated acid rain on soil acidification,
availabilities and temporal and spatial variations of Cu and Pb in a vegetable field under natural
conditions. J Food Agr Environ 7: 92-96.
Huang. CC: Xu. Y; Briggler. JT; McKee. M; Nam. P; Huang. YW. (2010). Heavy metals, hematology,
plasma chemistry, and parasites in adult hellbenders (Cryptobranchus alleganiensis). Environ Toxicol
Chem 29: 1132-1137. http://dx.doi.org/10.1002/etc.148
Huang. JH; Kalbitz. K; Matzner. E. (2008). Mobility of trimethyllead and total lead in the forest floor.
Soil Sci Soc Am J 72: 978-984. http://dx.doi.org/10.2136/sssai2006.0118
Hui. N; Selonen. S: Hanzel. J: Tuomela. M; Rainio. A; Kontio. H; Hakala. K; Lankinen. P; Steffen. K;
Fingerroos. T; Strommer. R; Setala. H; Hatakka. A; Romantschuk. M. (2009). Influence of lead on
organisms within the detritus food web of a contaminated pine forest soil. Boreal Environ Res 14:
70-85.
Hurd. NA; Sternberg. SPK. (2008). Bioremoval of aqueous lead using Lemna minor. Int J
Phytoremediation 10: 278-288. http://dx.doi.org/10.1080/15226510802096036
Inouve. LS: Yoo. LJ; Talent. LG: Clarke. JU: Jones. RP; Steevens. JA; Boyd. RE. (2007). Assessment of
lead uptake in reptilian prey species. Chemosphere 68: 1591-1596.
http://dx.doi.0rg/10.1016/i.chemosphere.2006.12.045
IPCC (Intergovernmental Panel on Climate Change). (2007). Climate change 2007: Impacts, adaptation
and vulnerability. Cambridge, U.K.: Cambridge University Press.
Isidori. M..; Cangiano. M..; Palermo. F. .A.; Parrella. A.. (2010). E-screen and vitellogenin assay for the
detection of the estrogenic activity of alkylphenols and trace elements. Comp Biochem Physiol C
Toxicol Pharmacol 152: 51-56. http://dx.doi.Org/10.1016/i.cbpc.2010.02.011
Islam. E; Liu. D; Li. T; Yang. X; Jin. X; Mahmood. Q: Tian. S: Li. J. (2008). Effect of Pb toxicity on leaf
growth, physiology and ultrastructure in the two ecotypes of Elsholtzia argyi. J Hazard Mater 154:
1-3. http://dx.doi.0rg/10.1016/i.ihazmat.2007.10.121
November 2012 7-265 Draft - Do Not Cite or Quote
-------
Jakl. M; Dytrtova. JJ: Miholova. D; Kolihova. D; Szakova. J: Tlustos. P. (2009). Passive diffusion
assessment of cadmium and lead accumulation by plants in hydroponic systems. Chem Speciation
Bioavailability 21: 111-120. http://dx.doi.org/10.3184/095422909x456870
Jara-Marini. ME; Soto-Jimenez. MF; Paez-Osuna. F. (2009). Trophic relationships and transference of
cadmium, copper, lead and zinc in a subtropical coastal lagoon food web from SE Gulf of California.
Chemosphere 77: 1366-1373. http://dx.doi.Org/10.1016/i.chemosphere.2009.09.025
Jeffree. RA; Oberhansli. F; Tevssie. JL. (2008). The accumulation of lead and mercury from seawater and
their depuration by eggs of the spotted dogfish Scyliorhinus canicula (Chondrichthys). Arch Environ
Contam Toxicol 55: 451-461. http://dx.doi.org/l0.1007/s00244-007-9103-4
Jezierska. B; Lugowska. K; Witeska. M. (2009). The effects of heavy metals on embryonic development
offish (a review) [Review]. Fish Physiol Biochem 35: 625-640.
http://dx.doi.org/10.1007/sl0695-008-9284-4
Jing. G: Li. Y; Xie. L; Zhang. RQ. (2007). Different effects of Pb2+ and Cu2+ on immune and
antioxidant enzyme activities in the mantle of Pinctada fucata. Environ Toxicol Pharmacol 24:
122-128. http://dx.doi.0rg/10.1016/i.etap.2007.04.002
John. R; Ahmad. P; Gadgil. K; Sharma. S. (2009). Heavy metal toxicity: Effect on plant growth,
biochemical parameters and metal accumulation by Brassica juncea L. International Journal of Plant
Production 3: 65-75.
Jones. JC. (1939). On the occurrence of lead shot in stomachs of North American gruiformes. J Wildl
Manag 3: 353-357.
Jones. RP; Bednar. AJ; Inouye. LS. (2009b). Subcellular compartmentalization of lead in the earthworm,
Eisenia fetida: Relationship to survival and reproduction. Ecotoxicol Environ Saf 72: 1045-1052.
http://dx.doi.0rg/10.1016/i.ecoenv.2008.12.011
Jordaens. K; De Wolf. H; Vandecasteele. B; Blust. R; Backeljau. T. (2006). Associations between shell
strength, shell morphology and heavy metals in the land snail Cepaea nemoralis (Gastropoda,
Helicidae). Sci Total Environ 363: 285-293. http://dx.doi.Org/10.1016/j.scitotenv.2005.12.002
Joshi. SR. (2008). Influence of roadside pollution on the phylloplane microbial community of Alnus
nepalensis (Betulaceae). Rev Biol Trop 56: 1521-1529.
Jothinavagi. N; Anbazhagan. C. (2009). Heavy metal monitoring of Rameswaram coast by some
Sargassum species. Am-Eurasian J Sci Res 4: 73-80.
Kalis. EJJ; Temminghoff. EJM; Visser. A; van Riemsdijk. WH. (2007). Metal uptake by Lolium perenne
in contaminated soils using a four-step approach. Environ Toxicol Chem 26: 335-345.
http://dx.doi.Org/10.1897/06-173R.l
Kalis. EJJ: Temminghoff. EJM: Weng. LP; van Riemsdijk. WH. (2006). Effects of humic acid and
competing cations on metal uptake by Lolium perenne. Environ Toxicol Chem 25: 702-711.
http://dx.doi.Org/10.1897/04-576R.l
Kalman. J: Riba. I; Blasco. J: Delvalls. TA. (2008). Is delta-aminolevulinic acid dehydratase activity in
bivalves from south-west Iberian Peninsula a good biomarker of lead exposure? Mar Environ Res 66:
38-40. http://dx.doi.0rg/10.1016/j.marenvres.2008.02.016
Kamaruzzaman. BY; Akbar. B; Jalal. K; Shahbudin. S. (2010). Accumulation of metals in the gills of
tilapia fingerlings (Oreochromis niloticus) from in vitro toxicology study. J Fish Aquat Sci 5: 503-509.
http://dx.doi.org/10.3923/ifas.2010.503.509
Kannan. K; Agusa. T; Perrotta. E; Thomas. NJ; Tanabe. S. (2006). Comparison of trace element
concentrations in livers of diseased, emaciated and non-diseased southern sea otters from the
California coast. Chemosphere 65: 2160-2167. http://dx.doi.Org/10.1016/i.chemosphere.2006.06.003
Kaufman. CA; Bennett. JR; Koch. I; Reimer. KJ. (2007). Lead bioaccessibility in food web intermediates
and the influence on ecological risk characterization. Environ Sci Technol 41: 5902-5907.
http://dx.doi.org/10.1021/es062443u
November 2012 7-266 Draft - Do Not Cite or Quote
-------
Kawakami. SK; Gledhill. M; Achterberg. EP. (2006). Production of phytochelatins and glutathione by
marine phytoplankton in response to metal stress. J Phycol 42: 975-989.
http://dx.doi.0rg/10.llll/j.1529-8817.2006.00265.x
Kay. SH; Haller. WT. (1986). Heavy metal bioaccumulation and effects on waterhyacinth weevils,
Neochetina eichhorniae, feeding on waterhyacinth, Eichhornia crassipes. Bull Environ Contam
Toxicol 37: 239-245. http://dx.doi.org/10.1007/BF01607756
Kempter. H; Krachler. M; Shotyk. W. (2010). Atmospheric Pb and Ti accumulation rates from Sphagnum
moss: Dependence upon plant productivity. Environ Sci Technol 44: 5509-5515.
http://dx.doi.org/10.1021/esl00366d
Kertesz. V; Bakonyi. G: Farkas. B. (2006). Water pollution by Cu and Pb can adversely affect mallard
embryonic development. Ecotoxicol Environ Saf 65: 67-73.
http://dx.doi.0rg/10.1016/i.ecoenv.2005.05.016
Kertesz. V; Fancsi. T. (2003). Adverse effects of (surface water pollutants) Cd, Cr and Pb on the
embryogenesis of the mallard. Aquat Toxicol 65: 425-433.
http://dx.doi.org/10.1016/S0166-445X(03)00155-3
Khan. MAQ; Ahmed. SA; Catalin. B; Khodadoust. A; Ajayi. O: Vaughn. M. (2006). Effect of temperature
on heavy metal toxicity to juvenile crayfish, Orconectes immunis (Hagen). Environ Toxicol 21:
513-520. http://dx.doi.org/10.1002/tox.20213
Khan. S: Hesham. AEL; Qiao. M; Rehman. S: He. JZ. (2010c). Effects of Cd and Pb on soil microbial
community structure and activities. Environ Sci Pollut Res Int 17: 288-296.
http://dx.doi.org/10.1007/sll356-009-0134-4
Khangarot. BS. (1991). Toxicity of metals to a freshwater tubificid worm, Tubifex tubifex (Muller). Bull
Environ Contam Toxicol 46: 906-912.
Khozhina. El; Sherriff. BL. (2008). The accumulation of Cu, Zn, Cd, and Pb in the aquatic biomass of
sulphide tailing ponds. Geochem Int 46: 897-911. http://dx.doi.org/10.1134/s0016702908090048
Kibblewhite. MG: Ritz. K; Swift. MJ. (2008). Soil health in agricultural systems. Philos Trans R Soc
Lond B Biol Sci 363: 685-701. http://dx.doi.org/10.1098/rstb.2007.2178
Kimbrough. KL; Lauenstein. GG: Christensen. JD; Apeti. DA. (2008). An assessment of two decades of
contaminant monitoring in the nation's coastal zone. Silver Spring, MD: National Centers for Coastal
Ocean Science, http://aquaticcommons.org/2232/
King. CK; Dowse. MC: Simpson. SL. (2010). Toxicity of metals to the Bivalve tellina deltoidalis and
relationships between metal bioaccumulation and metal partitioning between seawater and marine
sediments. Arch Environ Contam Toxicol 58: 657-665. http://dx.doi.org/10.1007/s00244-009-9413-9
King. CK; Gale. SA; Stauber. JL. (2006). Acute toxicity and bioaccumulation of aqueous and sediment-
bound metals in the estuarine amphipod Melita plumulosa. Environ Toxicol 21: 489-504.
http://dx.doi.org/10.1002/tox.20211
Kitvatanachai. S: Apiwathnasorn. C: Leemingsawat. S: Wongwit. W: Tornee. S. (2005). Determination of
lead toxicity in Culex quinquefasciatus mosquitoes in the laboratory. Southeast Asian J Trop Med
Public Health 36: 862-874.
Klaminder. J: Bindler. R; Emteryd. O: Renberg. I. (2005). Uptake and recycling of lead by boreal forest
plants: Quantitative estimates from a site in northern Sweden. Geochim Cosmo Act 69: 2485-2496.
http://dx.doi.0rg/10.1016/i.gca.2004.ll.013
Klok. C: Van der Hout. A; Bodt. J. (2006). Population growth and development of the earthworm
Lumbricus rubellus in a polluted field soil: Possible consequences for the godwit (Limosa limosa).
Environ Toxicol Chem 25: 213-219. http://dx.doi.Org/10.1897/05-286R.l
Knowlton. MF; Boyle. TP; Jones. JR. (1983). Uptake of lead from aquatic sediment by submersed
macrophytes and crayfish. Arch Environ Contam Toxicol 12: 535-541.
http://dx.doi.org/10.1007/BF01056549
November 2012 7-267 Draft - Do Not Cite or Quote
-------
Kobler. J; Fitz. WJ: Dirnbock. T; Mirtl. M. (2010). Soil type affects migration pattern of airborne Pb and
Cd under a spruce-beech forest of the UN-ECE integrated monitoring site Zobelboden, Austria.
Environ Pollut 158: 849-854. http://dx.doi.Org/10.1016/i.envpol.2009.09.026
Kohler. HR; Alberti. G: Seniczak. S: Seniczak. A. (2005). Lead-induced hsp70 and hsp60 pattern
transformation and leg malformation during postembryonic development in the oribatid mite,
Archegozetes longisetosus Aoki. Comp Biochem Physiol C Toxicol Pharmacol 141: 398-405.
http://dx.doi.0rg/10.1016/i.cbpc.2005.09.003
Koivula. MJ; Eeva. T. (2010). Metal-related oxidative stress in birds. Environ Pollut 158: 2359-2370.
http://dx.doi.0rg/10.1016/i.envpol.2010.03.013
Komarek. M; Ettler. V; Szakova. J: Sebek. O; Tlustos. P. (2009). Bioavailability of lead and cadmium in
soils artificially contaminated with smelter fly ash. Bull Environ Contam Toxicol 83: 286-290.
http://dx.doi.org/10.1007/s00128-009-9742-4
Kominkova. D; Nabelkova. J. (2005). The risk assessment of heavy metals in the ecosystem of urban
creeks. Water Sci Technol 53: 10. http://dx.doi.org/10.2166/wst.2006.298
Komjarova. I; Blust. R. (2008). Multi-metal interactions between Cd, Cu, Ni, Pb and Zn in water flea
Daphnia magna, a stable isotope experiment. Aquat Toxicol 90: 138-144.
http://dx.doi.0rg/10.1016/i.aquatox.2008.08.007
Komjarova. I; Blust. R. (2009a). Application of a stable isotope technique to determine the simultaneous
uptake of cadmium, copper, nickel, lead, and zinc by the water flea Daphnia magna from water and
the green algae Pseudokirchneriella subcapitata. Environ Toxicol Chem 28: 1739-1748.
http://dx.doi.0rg/10.1897/08-437.l
Komjarova. I; Blust. R. (2009b). Effect of Na, Ca and pH on simultaneous uptake of Cd, Cu, Ni, Pb, and
Zn in the water flea Daphnia magna measured using stable isotopes. Aquat Toxicol 94: 81-86.
http://dx.doi.0rg/10.1016/i.aquatox.2009.05.018
Komjarova. I; Blust. R. (2009c). Effects of Na, Ca, and pH on the simultaneous uptake of Cd, Cu, Ni, Pb,
and Zn in the zebrafish Danio rerio: A stable isotope experiment. Environ Sci Technol 43: 7958-7963.
http://dx.doi.org/10.1021/es9016987
Kools. SAE: Boivin. MEY: Van Per Wurff. AWG: Berg. MP: Van Gestel. CAM: Van Straalen. NM.
(2009). Assessment of structure and function in metal polluted grasslands using Terrestrial Model
Ecosystems. Ecotoxicol Environ Saf 72: 51-59. http://dx.doi.Org/10.1016/j.ecoenv.2008.03.016
Korean. SE; Cigerci. IH; Konuk. M. (2007). Screening of delta-aminolevulinic acid dehydratase from
Pseudomonas strains as biosensor for lead and some other metals contamination. Environ Monit
Assess 134: 263-269. http://dx.doi.org/10.1007/sl0661-007-9615-4
Kramer. JL; Redig. PT. (1997). Sixteen years of lead poisoning in eagles, 1980-95: An epizootiologic
view. Journal of Raptor Research 31: 327-332.
Kuang. YW: Zhou. GY; Wen. DZ; Liu. SZ. (2007). Heavy metals in bark of Pinus massoniana (Lamb.) as
an indicator of atmospheric deposition near a smeltery at Qujiang, China. Environ Sci Pollut Res Int
14: 270-275. http://dx.doi.org/10.1065/espr2006.09.344
Kudirat. AO. (2008). Bioconcentration of lead in the tissues of feral and laboratory exposed Clarias
gariepinus. Journal of Medical Sciences 8: 281-286.
Kuklova. M..; Kukla. J..: Hnilicka. F.. (2010). The soil-to-herbs transfer of heavy metals in spruce
ecosystems. Pol J Environ Stud 19: 1263-1268.
Kumar. G: Tripathi. R. (2008). Lead-induced cytotoxicity and mutagenicity in grass pea. Turkish Journal
of Biology 32: 73-78.
Kurilenko. VV; Osmolovskava. NG. (2006). Ecological-biogeochemical role of macrophytes in aquatic
ecosystems of urbanized territories (an example of small water bodies of St. Petersburg). Russian
Journal of Ecology 37: 147-151. http://dx.doi.org/10.1134/sl067413606030015
November 2012 7-268 Draft - Do Not Cite or Quote
-------
Kwartirnikov. MA; Lavchieva-Nacheva. GV: Lavchiev. VI. (1999). The effects of lead intoxication on the
survival and behavior of Leptinotarsa decemlineata Say (Coloeptera: Chrysomelidae). Acta Zoolog
BulgSl: 103-108.
Lacoue-Labarthe. T; Warnau. M; Metian. M; Oberhansli. F; Rouleau. C: Bustamante. P. (2009).
Biokinetics of Hg and Pb accumulation in the encapsulated egg of the common cuttlefish Sepia
officinalis: Radiotracer experiments. Sci Total Environ 407: 6188-6195.
http://dx.doi.0rg/10.1016/i.scitotenv.2009.09.003
Lagisz. M; Laskowski. R. (2008). Evidence for between-generation effects in carabids exposed to heavy
metals pollution. Ecotoxicology 17: 59-66. http://dx.doi.org/10.1007/sl0646-007-0176-7
Lamelas. C: Pinheiro. JP; Slaveykova. VI. (2009). Effect of humic acid on Cd(ll), Cu(ll), and Pb(ll)
uptake by freshwater algae: Kinetic and cell wall speciation considerations. Environ Sci Technol 43:
730-735. http://dx.doi.org/10.1021/es802557r
Lamelas. C: Slaveykova. VI. (2007). Comparison of Cd(II), Cu(II), and Pb(II) biouptake by green algae
in the presence of humic acid. Environ Sci Technol 41: 4172-4178.
http://dx.doi.org/10.1021/es063102i
Lamelas. C: Slaveykova. VI. (2008). Pb uptake by the freshwater alga Chlorella kesslerii in the presence
of dissolved organic matter of variable composition. Environ Chem 5: 366-372.
http://dx.doi.org/10.1071/en08043
Lamelas. C: Wilkinson. KJ; Slaveykova. VI. (2005). Influence of the composition of natural organic
matter on Pb bioavailability to microalgae. Environ Sci Technol 39: 6109-6116.
http://dx.doi.org/10.1021/es050445t
Landers. DH; Simonich. SL; Jaffe. DA; Geiser. LH; Campbell. DH; Schwindt. AR; Schreck. CB; Kent.
ML; Hafner. WD; Taylor. HE; Hageman. KJ: Usenko. S: Ackerman. LK; Schrlau. JE; Rose. NL; Blett.
TF; Erwav. MM. (2008). The fate, transport, and ecological impacts of airborne contaminants in
western national parks (USA). (EPA/600/R-07/138). Corvallis, Oregon: U.S. Environmental
Protection Agency, NHEERL, Western Ecology Division.
http://www.nature.nps.gov/air/studies/air toxics/WACAPreport.cfm
Landre. AL; Watmough. SA; Dillon. PJ. (2010). Metal pools, fluxes, and budgets in an acidified forested
catchment on the Precambrian Shield, Central Ontario, Canada. Water Air Soil Pollut 209: 209-228.
http://dx.doi.org/10.1007/sll270-009-0193-7
Langdon. CJ: Hodson. ME; Arnold. RE; Black. S. (2005). Survival, Pb-uptake and behaviour of three
species of earthworm in Pb treated soils determined using an OECD-style toxicity test and a soil
avoidance test. Environ Pollut 138: 368-375. http://dx.doi.Org/10.1016/i.envpol.2005.03.002
Larsson. P; Holmqvist. N; Stenroth. P; Berglund. O: Nystrom. P; Graneli. W (2007). Heavy metals and
stable isotopes in a benthic omnivore in a trophic gradient of lakes. Environ Sci Technol 41:
5973-5979. http://dx.doi.org/10.1021/es0704838
Lazzaro. A; Schulin. R; Widmer. F; Frev. B. (2006). Changes in lead availability affect bacterial
community structure but not basal respiration in a microcosm study with forest soils. Sci Total
Environ 371: 110-124. http://dx.doi.Org/10.1016/i.scitotenv.2006.08.033
Le Faucheur. S: Schildknecht. F; Behra. R: Sigg. L. (2006). Thiols in Scenedesmus vacuolatus upon
exposure to metals and metalloids. Aquat Toxicol 80: 355-361.
http://dx.doi.0rg/10.1016/i.aquatox.2006.10.002
Leal. RB; Ribeiro. SJ: Posser. T; Cordova. FM; Rigon. AP; Filho. EZ; Bainy. ACD. (2006). Modulation
of ERK1/2 and p38(MAPK) by lead in the cerebellum of Brazilian catfish Rhamdia quelen. Aquat
Toxicol 77: 98-104. http://dx.doi.Org/10.1016/i.aquatox.2005.ll.002
Leland. HV; Kuwabara. JS. (1985). Trace metals. In GM Rand; SR Petrocelli (Eds.), Fundamentals of
aquatic toxicology: Methods and applications (pp. 374-415). Washington, DC: Hemisphere
Publishing.
November 2012 7-269 Draft - Do Not Cite or Quote
-------
Levin. SA. (1998). Ecosystems and the biosphere as complex adaptive systems. Ecosystems 1: 431-436.
http://dx.doi.org/10.1007/sl00219900037
Li. LZ; Zhou. DM; Wang. P; Luo. XS. (2008b). Subcellular distribution of Cd and Pb in earthworm
Eisenia fetida as affected by Ca2+ ions and Cd-Pb interaction. Ecotoxicol Environ Saf 71: 632-637.
http://dx.doi.0rg/10.1016/i.ecoenv.2008.04.001
Li. M; Liu. ZT; Xu. Y; Cui. YB; Li. PS: Kong. ZM. (2009b). Comparative effects of Cd and Pb on
biochemical response and DNA damage in the earthworm Eisenia fetida (Annelida, Oligochaeta).
Chemosphere 74: 621-625. http://dx.doi.Org/10.1016/i.chemosphere.2008.10.048
Li. Y; Zu. Y; Fang. Q; Gao. Z; Schvartz. C. (2009d). Relationship between heavy metal concentrations of
herbaceous plants and soils at four Pb-Zn mining sites in Yunnan, China. Front Environ Sci Eng China
3: 325-333. http://dx.doi.org/10.1007/sll783-009-0024-5
Ling. Q; Hong. F. (2009). Effects of Pb2+ on the structure and function of photosystem II of Spirodela
polyrrhiza. Biol Trace Elem Res 129: 251-260. http://dx.doi.org/10.1007/sl2011-008-8283-8
Ling. Q; Hong. F. (2010). Antioxidative role of cerium against the toxicity of lead in the liver of silver
crucian carp. Fish Physiol Biochem 36: 367-376. http://dx.doi.org/10.1007/sl0695-008-9301-7
Liu. DW: Liu. XM; Chen. ZW: Xu. HZ; Ding. XF (2010a). Bioaccumulation of lead and the effects of
lead on catalase activity, glutathione levels, and chlorophyll content in the leaves of wheat. Commun
Soil Sci Plant Anal 41: 935-944. http://dx.doi.org/10.1080/00103621003646022
Lock. K; Waegeneers. N; Smolders. E; Criel. P; Van Eeckhout H; Janssen. CR. (2006). Effect of leaching
and aging on the bioavailability of lead to the springtail Folsomia Candida. Environ Toxicol Chem 25:
2006-2010. http://dx.doi.Org/10.1897/05-612R.l
Locke. LN; Thomas. NJ. (1996). Lead poisoning of waterfowl and raptors. In A Fairbrother; LN Locke;
GL Hoff (Eds.), Noninfectious diseases of wildlife (2nd ed., pp. 108-117). Ames, IA: Iowa State Press.
Lofts. S: Chapan. PM; Dwver. R; Mclaughlin. MJ; Choeters. I; Sheppard. SC: Adams. WJ: Allowav. BJ;
Antunes. PM; Campbell. PG: Davies. B; Degryse. F; De Vries. W: Farley. K; Garrett. RG: Green. A;
Jan Groenenberg. B; Hale. B; Harrass. M; Hendershot. WH; Keller. A; Lanno. R; Liang. T; Liu. WX;
Ma. Y; Menzie. C: Moolenaar. SW: Piatkiewicz. W: Reimann. C: Rieuwerts. JS: Santore. RC: Sauve.
S: Schuetze. G: Schlekat. C: Skeaff. J: Smolders. E; Tao. S: Wilkins. J: Zhao. FJ. (2007). Critical loads
of metals and other trace elements to terrestrial environments. Environ Sci Technol 41: 6326-6331.
http://dx.doi.org/10.1021/es0726106
Luoma. SN; Rainbow. PS. (2005). Why is metal bioaccumulation so variable? Biodynamics as a unifying
concept [Review]. Environ Sci Technol 39: 1921-1931. http://dx.doi.org/10.1021/es048947e
Ma. L; Xu. R; Jiang. J. (2010). Adsorption and desorption of Cu(II) and Pb(II) in paddy soils cultivated
for various years in the subtropical China. J Environ Sci 22: 689-695.
http://dx.doi.org/10.1016/S1001-0742(09)60164-9
Ma. Y; Dickinson. NM; Wong. MH. (2006). Beneficial effects of earthworms and arbuscular mycorrhizal
fungi on establishment of leguminous trees on Pb/Zn mine tailings. Soil Biol Biochem 38: 1403-1412.
http://dx.doi.0rg/10.1016/i.soilbio.2005.10.016
MacDonald. DP; Ingersoll. CG: Berger. TA. (2000). Development and evaluation of consensus-based
sediment quality guidelines for freshwater ecosystems. Arch Environ Contam Toxicol 39: 20-31.
http://dx.doi.org/10.1007/s002440010075
MacDonald. DP; Smorong. DE; Ingersoll. CG: Besser. JM; Brumbaugh. WG: Kemble. NE; May. TE;
Ivev. CD; Irving. S: O'Hare. M. (2009). Development and evaluation of sediment and pore-water
toxicity thresholds to support sediment quality assessments in the Tri-state Mining District (TSMD),
Missouri, Oklahoma and Kansas. Nanaimo, Canada: MacDonald Environmental Sciences.
http://www.fws.gov/soutfawest/es/Oklahoma/Documents/Contaminants/01TSMD%20Biological%20D
ata%20Evaluation%202008 Volume%201 Vers4 Feb09.pdf
November 2012 7-270 Draft - Do Not Cite or Quote
-------
MacFarlane. GR: Koller. CE; Blomberg. SP. (2007). Accumulation and partitioning of heavy metals in
mangroves: A synthesis of field-based studies. Chemosphere 69: 1454-1464.
http://dx.doi.0rg/10.1016/i.chemosphere.2007.04.059
MacFarlane. GR: Schreider. M; McLennan. B. (2006). Biomarkers of heavy metal contamination in the
red fingered marsh crab, Parasesarma erythodactyla. Arch Environ Contam Toxicol 51: 584-593.
http://dx.doi.org/10.1007/s00244-005-5067-4
Mackie. GL. (1989). Tolerances of five benthic invertebrates to hydrogen ions and metals (Cd, Pb, Al).
Arch Environ Contam Toxicol 18: 215-223. http://dx.doi.org/10.1007/BF01056206
Mager. EM. (2012). Lead. In Homeostasis and toxicology of non-essential metals. U.K.: Academic Press.
http://www.elsevierdirect.com/ISBN/9780123786340/Fish-Phvsiologv-Homeostasis-and-Toxicologv-
of-NonEssential-Metals
Mager. EM; Brix. KV; Gerdes. RM; Ryan. AC: Grosell. M. (2011 a). Effects of water chemistry on the
chronic toxicity of lead to the cladoceran, Ceriodaphnia dubia. Ecotoxicol Environ Saf 74: 238-243.
http://dx.doi.0rg/10.1016/i.ecoenv.2010.ll.005
Mager. EM; Brix. KV: Grosell. M. (2010). Influence of bicarbonate and humic acid on effects of chronic
waterborne lead exposure to the fathead minnow (Pimephales promelas). Aquat Toxicol 96: 135-144.
http://dx.doi.0rg/10.1016/i.aquatox.2009.10.012
Mager. EM; Esbaugh. AJ; Brix. KV: Ryan. AC: Grosell. M. (20 lib). Influences of water chemistry on the
acute toxicity of lead to Pimephales promelas and Ceriodaphnia dubia. Comp Biochem Physiol C
Toxicol Pharmacol 153: 82-90. http://dx.doi.Org/10.1016/i.cbpc.2010.09.004
Mager. EM; Grosell. M. (2011). Effects of acute and chronic waterborne lead exposure on the swimming
performance and aerobic scope of fathead minnows (Pimephales promelas). Comp Biochem Physiol C
Toxicol Pharmacol 154: 7-13. http://dx.doi.Org/10.1016/i.cbpc.2011.03.002
Mager. EM; Wintz. H; Vulpe. CD; Brix. KV: Grosell. M. (2008). Toxicogenomics of water chemistry
influence on chronic lead exposure to the fathead minnow (Pimephales promelas). Aquat Toxicol 87:
200-209. http://dx.doi.0rg/10.1016/i.aquatox.2008.02.001
Magrisso. S: Belkin. S: Erel. Y. (2009). Lead bioavailability in soil and soil components. Water Air Soil
Pollut 202: 315-323. http://dx.doi.org/10.1007/sll270-009-9978-v
Mahler. BJ; Van Metre. PC: Callender. E. (2006). Trends in metals in urban and reference lake sediments
across the United States, 1970 to 2001. Environ Toxicol Chem 25: 1698-1709.
Mahmoudi. E; Essid. N; Beyrem. H; Hedfi. A; Boufahja. F; Vitiello. P: Aissa. P. (2007). Individual and
combined effects of lead and zinc on a free-living marine nematode community: Results from
microcosm experiments. Exp Mar Bio Ecol 343: 217-226.
http://dx.doi.0rg/10.1016/i.jembe.2006.12.017
Maitv. S: Roy. S: Bhattacharya. S: Chaudhury. S. (2011). Metallothionein responses in the earthworm
Lampito mauritii (Kinberg) following lead and zinc exposure: A promising tool for monitoring metal
contamination. European Journal of Soil Biology 47: 69-71.
http://dx.doi.0rg/10.1016/i.eisobi.2010.10.001
Maitv. S: Roy. S: Chaudhury. S: Bhattacharya. S. (2008). Antioxidant responses of the earthworm
Lampito mauritii exposed to Pb and Zn contaminated soil. Environ Pollut 151: 1-7.
http://dx.doi.0rg/10.1016/i.envpol.2007.03.005
Malik. N; Biswas. AK; Qureshi. TA; Borana. K; Virha. R. (2010). Bioaccumulation of heavy metals in
fish tissues of a freshwater lake of Bhopal. Environ Monit Assess 160: 267-276.
http://dx.doi.org/10.1007/sl0661-008-0693-8
Mander. U; Mitsch. WJ. (2009). Pollution control by wetlands. Ecol Eng 35: 153-158.
http://dx.doi.0rg/10.1016/i.ecoleng.2008.10.005
November 2012 7-271 Draft - Do Not Cite or Quote
-------
Manzo. S: Buono. S: Cremisini. C. (2010). Cadmium, lead and their mixtures with copper: Paracentrotus
lividus embryotoxicity assessment, prediction, and offspring quality evaluation. Ecotoxicology 19:
1209-1223. http://dx.doi.org/10.1007/sl0646-010-0506-z
Marchand. L; Mench. M; Jacob. PL; Otte. ML. (2010). Metal and metalloid removal in constructed
wetlands, with emphasis on the importance of plants and standardized measurements: A review
[Review]. Environ Pollut 158: 3447-3467. http://dx.doi.Org/10.1016/i.envpol.2010.08.018
Maret. TR; Cain. DJ; MacCov. DE; Short. TM. (2003). Response of benthic invertebrate assemblages to
metal exposure and bioaccumulation associated with hard-rock mining in northwestern streams, USA.
J North Am Benthol Soc 22: 598-620.
Marin-Guirao. L; Atucha. AM; Barba. JL; Lopez. EM; Fernandez. AJG. (2005). Effects of mining wastes
on a seagrass ecosystem: Metal accumulation and bioavailability, seagrass dynamics and associated
community structure. Mar Environ Res 60: 317-337.
http://dx.doi.0rg/10.1016/i.marenvres.2004.ll.002
Martin. M; Osborn. KE; Billig. P; Glickstein. N. (1981). Toxicities often metals to Crassostrea gigas and
Mytilus edulis embryos and Cancer magister larvae. Mar Pollut Bull 12: 305-308.
http://dx.doi.org/10.1016/0025-326X(81)90081-3
Martinez-Haro. M; Green. AJ; Mateo. R. (2011). Effects of lead exposure on oxidative stress biomarkers
and plasma biochemistry in waterbirds in the field. Environ Res 111: 530-538.
http://dx.doi.0rg/10.1016/i.envres.2011.02.012
McGlothan. JL; Karcz-Kubicha. M; Guilarte. TR. (2008). Developmental lead exposure impairs
extinction of conditioned fear in young adult rats. Neurotoxicology 29: 1127-1130.
http://dx.doi.0rg/10.1016/i.neuro.2008.06.010
McLaughlin. MJ; Lofts. S: Warne. M; Amorim. MJB; Fairbrother. A; Lanno. R; Hendershot. W: Schlekat.
CE; Ma. Y; Paton. GI. (2010). Derivation of ecologically-based soil standards for trace elements. In G
Merrington; I Schoeters (Eds.), Soil quality standards for trace elements: Derivation, implementation,
and interpretation (pp. 7-80). Boca Raton, FL: CRC Press, http://dx.doi.org/10.1201/bl0293-3
Mebane. CA; Hennessv. DP; Dillon. FS. (2008). Developing acute-to-chronic toxicity ratios for lead,
cadmium, and zinc using rainbow trout, a mayfly, and a midge. Water Air Soil Pollut 188: 41-66.
http://dx.doi.org/10.1007/sll270-007-9524-8
Mehrotra. V: Saxena. VL; Saxena. AK. (2008). Impact of different doses of lead on internal organs of
quails. J Environ Biol 29: 147-149.
Mellem. JJ; Baijnath. H; Odhav. B. (2009). Translocation and accumulation of Cr, Hg, As, Pb, Cu and Ni
by Amaranthus dubius (Amaranthaceae) from contaminated sites. J Environ Sci Health A Tox Hazard
Subst Environ Eng 44: 568-575. http://dx.doi.org/10.1080/10934520902784583
Menta. C: Maggiani. A; Vattuone. Z. (2006). Effects of Cd and Pb on the survival and juvenile production
of Sinella coeca and Folsomia Candida. European Journal of Soil Biology 42: 181-189.
http://dx.doi.0rg/10.1016/i.ejsobi.2006.01.001
Metian. M; Warnau. M; Oberhansli. F; Bustamante. P. (2009). Delineation of Pb contamination pathways
in two Pectinidae: The variegated scallop Chlamys varia and the king scallop Pecten maximus. Sci
Total Environ 407: 3503-3509. http://dx.doi.Org/10.1016/i.scitotenv.2009.02.010
Meyer. C: Gilbert. D; Gaudry. A; Franchi. M; Nguyen-Viet. H; Fabure. J: Bernard. N. (2010).
Relationship of atmospheric pollution characterized by gas (NO2) and particles (PM10) to microbial
communities living in bryophytes at three differently polluted sites (rural, urban, and industrial).
Microb Ecol 59: 324-334. http://dx.doi.org/10.1007/s00248-009-9580-2
Meyer. JS: Adams. WJ: Brix. KV; Luoma. SN; Mount. PR; Stubblefield. WA; Wood. CM. (2005).
Toxicity of dietborne metals to aquatic organisms. In JS Meyer; WJ Adams; KV Brix; SN Luoma; DR
Mount; WA Stubblefield; CM Wood (Eds.). Pensacola, FL: Society of Environmental Toxicology and
Chemistry.
November 2012 7-272 Draft - Do Not Cite or Quote
-------
Migula. P; Binkowska. K. (1993). Feeding strategies of grasshoppers (Chorthippus spp) on heavy metal
contaminated plants. Sci Total Environ 134: 1071-1083.
http://dx.doi.org/10.1016/S0048-9697(05)80112-3
Miller. JR; Anderson. JB; Lechler. PJ; Kondrad. SL; Galbreath. PF; Salter. EB. (2005). Influence of
temporal variations in water chemistry on the Pb isotopic composition of rainbow trout
(Oncorhynchus mykiss). Sci Total Environ 350: 204-224.
http://dx.doi.0rg/10.1016/i.scitotenv.2005.01.030
Miramand. P; Bustamante. P; Bentlev. D; Koueta. N. (2006). Variation of heavy metal concentrations
(Ag, Cd, Co, Cu, Fe, Pb, V, and Zn) during the life cycle of the common cuttlefish Sepia officinalis.
Sci Total Environ 361: 132-143. http://dx.doi.Org/10.1016/j.scitotenv.2005.10.018
Miretzky. P; Munoz. C: Carrillo-Chavez. A. (2007). A sandy loam soil as a natural control for Pb
contamination. Environ Chem Lett 5: 131-136. http://dx.doi.org/10.1007/sl0311-007-0093-2
Mishra. S: Srivastava. S: Tripathi. RD; Kumar. R; Seth. CS: Gupta. DK. (2006b). Lead detoxification by
coontail (Ceratophyllum demersum L.) involves induction of phytochelatins and antioxidant system in
response to its accumulation. Chemosphere 65: 1027-1039.
http://dx.doi.0rg/10.1016/i.chemosphere.2006.03.033
Mishra. VK; Upadhyav. AR; Pandev. SK; Tripathi. BD. (2008). Concentrations of heavy metals and
aquatic macrophytes of Govind Ballabh Pant Sagar an anthropogenic lake affected by coal mining
effluent. Environ Monit Assess 141: 49-58. http://dx.doi.org/10.1007/sl0661-007-9877-x
Mo. MH; Chen. WM; Yang. HR; Zhang. KQ. (2008). Diversity and metal tolerance of nematode-trapping
fungi in Pb-polluted soils. J Microbiol 46: 16-22. http://dx.doi.org/10.1007/sl2275-007-0174-8
Mogren. CL; Trumble. JT. (2010). The impacts of metals and metalloids on insect behavior. Entomol Exp
Appl 135: 1-17. http://dx.doi.0rg/10.llll/i.1570-7458.2010.00967.x
Mohapatra. A; Rautrav. TR; Patra. AK; Vijavan. V; Mohantv. RK. (2009). Trace element-based food
value evaluation in soft and hard shelled mud crabs. Food Chem Toxicol 47: 2730-2734.
http://dx.doi.0rg/10.1016/i.fct.2009.07.037
Monteiro. V; Cavalcante. DG: Vilela. MB; Sofia. SH; Martinez. CB. (2011). In vivo and in vitro
exposures for the evaluation of the genotoxic effects of lead on the Neotropical freshwater fish
Prochilodus lineatus. Aquat Toxicol 104: 291-298. http://dx.doi.Org/10.1016/j.aquatox.2011.05.002
Montufar-Melendez. AI; Sanchez-Ortiz. JR; Sarma. SSS: Nandini. S. (2007). Combined effects of
temperature and lead concentration on the competition between the rotifers Brachionus havanaensis
and Brachionus rubens (Rotifera: Brachionidae). J Environ Sci Health A Tox Hazard Subst Environ
Eng 42: 393-398. http://dx.doi.org/10.1080/10934520601144717
Morris. S: van Aardt. WJ: Ahern. MD. (2005). The effect of lead on the metabolic and energetic status of
the Yabby, Cherax destructor, during environmental hypoxia. Aquat Toxicol 75: 16-31.
http://dx.doi.0rg/10.1016/i.aquatox.2005.07.001
Morselli. L; Bernardi. E; Passarini. F; Tesini. E. (2006). Critical loads for Cd and Pb in the province of
Bologna. Ann Chim 96: 697-705. http://dx.doi.org/10.1002/adic.200690072
Mosher. S: Cope. WG: Weber. FX; Shea. D; Kwak. TJ. (2012). Effects of lead on Na(+), K(+)ATPase and
hemolymph ion concentrations in the freshwater mussel Elliptic complanata. Environ Toxicol 27:
268-276. http://dx.doi.org/10.1002/tox.20639
Mouchet. F; Cren. S: Cunienq. C: Devdier. E; Guilet. R; Gauthier. L. (2007). Assessment of lead
ecotoxicity in water using the amphibian larvae (Xenopus laevis) and preliminary study of its
immobilization in meat and bone meal combustion residues. Biometals 20: 113-127.
http://dx.doi.org/10.1007/sl0534-006-9019-x
Mulhern. BM; Reichel. WL; Locke. LN; Lamont. TG: Belisle. A; Cromartie. E; Baglev. GE; Proutv. RM.
(1970). Organochlorine residues and autopsy data from bald eagles 1966-68. Pestic Monit J 4:
141-144.
November 2012 7-273 Draft - Do Not Cite or Quote
-------
Murray. H; Thompson. K; Macfie. SM. (2009). Site- and species-specific patterns of metal bioavailability
in edible plants. Botany 87: 702-711. http://dx.doi.org/10.1139/b09-019
Nahmani. J; Hodson. ME; Black. S. (2007). Effects of metals on life cycle parameters of the earthworm
Eisenia fetida exposed to field-contaminated, metal-polluted soils. Environ Pollut 149: 44-58.
http://dx.doi.0rg/10.1016/i.envpol.2006.12.018
Nahmani. J; Hodson. ME; Devin. S: Vijver. MG. (2009). Uptake kinetics of metals by the earthworm
Eisenia fetida exposed to field-contaminated soils. Environ Pollut 157: 2622-2628.
http://dx.doi.0rg/10.1016/i.envpol.2009.05.002
Nain. S: Smits. JE. (2011). Subchronic lead exposure, immunotoxicology and increased disease resistance
in Japanese quail (Corturnix coturnix japonica). Ecotoxicol Environ Saf 74: 787-792.
http://dx.doi.0rg/10.1016/i.ecoenv.2010.10.045
Nakavama. SM; Ikenaka. Y; Hamada. K; Muzandu. K; Choongo. K; Teraoka. H; Mizuno. N; Ishizuka. M.
(2011). Metal and metalloid contamination in roadside soil and wild rats around a Pb-Zn mine in
Kabwe, Zambia. Environ Pollut 159: 175-181. http://dx.doi.Org/10.1016/i.envpol.2010.09.007
Nakhle. KF; Cossa. D; Khalaf. G: Beliaeff. B. (2006). Brachidontes variabilis and Patella sp as
quantitative biological indicators for cadmium, lead and mercury in the Lebanese coastal waters.
Environ Pollut 142: 73-82. http://dx.doi.Org/10.1016/i.envpol.2005.09.016
Nam. DH; Lee. DP. (2006). Monitoring for Pb and Cd pollution using feral pigeons in rural, urban, and
industrial environments of Korea. Sci Total Environ 357: 288-295.
http://dx.doi.0rg/10.1016/i.scitotenv.2005.08.017
Nandi. S: Gupta. PS: Selvaraiu. S: Roy. SC: Ravindra. JP (2010). Effects of exposure to heavy metals on
viability, maturation, fertilization, and embryonic development of buffalo (Bubalus bubalis) oocytes in
vitro. Arch Environ Contam Toxicol 58: 194-204. http://dx.doi.org/10.1007/s00244-009-9342-7
Nguyen-Viet. H; Bernard. N; Mitchell. BAD; Badot. PM; Gilbert. D. (2008). Effect of lead pollution on
testate amoebae communities living in Sphagnum fallax: An experimental study. Ecotoxicol Environ
Saf 69: 130-138. http://dx.doi.Org/10.1016/i.ecoenv.2007.02.007
Nilsson. J; Grennfelt. P. (1988). Critical loads for sulphur and nitrogen: Report from a workshop held at
Skokloster, Sweden, 19-24 March 1988. Skokloster, Sweden: Nordic Council of Ministers:
Copenhagen.
Nimptsch. J: Wunderlin. DA; Dollan. A; Pflugmacher. S. (2005). Antioxidant and biotransformation
enzymes in Myriophyllum quitense as biomarkers of heavy metal exposure and eutrophication in
Suquia River basin (Cordoba, Argentina). Chemosphere 61: 147-157.
http://dx.doi.0rg/10.1016/i.chemosphere.2005.02.079
Niyogi. S: Wood. CM. (2004). Biotic ligand model, a flexible tool for developing site-specific water
quality guidelines for metals [Review]. Environ Sci Technol 38: 6177-6192.
http://dx.doi.org/10.1021/es0496524
NOAA (National Oceanic and Atmospheric Administration). (1999). Sediment quality guidelines
developed for the national status and trends program. Silver Spring, MD: National Ocean Service.
Notten. MJM; Oosthoek. AJP; Rozema. J: Aerts. R. (2005). Heavy metal concentrations in a soil-plant-
snail food chain along a terrestrial soil pollution gradient. Environ Pollut 138: 178-190.
http://dx.doi.0rg/10.1016/i.envpol.2005.01.011
Notten. MJM: Oosthoek. AJP: Rozema. J: Aerts. R. (2006). Heavy metal pollution affects consumption
and reproduction of the landsnail Cepaea nemoralis fed on naturally polluted Urtica dioica leaves.
Ecotoxicology 15: 295-304. http://dx.doi.org/10.1007/sl0646-006-0059-3
Notten. MJM: Walraven. N; Beets. CJ: Vroon. P; Rozema. J: Aerts. R. (2008). Investigating the origin of
Pb pollution in a terrestrial soil-plant-snail food chain by means of Pb isotope ratios. Appl Geochem
23: 1581-1593. http://dx.doi.0rg/10.1016/i.apgeochem.2008.01.010
November 2012 7-274 Draft - Do Not Cite or Quote
-------
NFS (U.S. National Park Service). (2011). WACAP database.
http://www.nature.nps.gov/air/studies/air toxics/WACAPreport.cfm
Nwachukwu. OI; Pulford. ID. (2011). Microbial respiration as an indication of metal toxicity in
contaminated organic materials and soil. J Hazard Mater 185: 1140-1147.
http://dx.doi.0rg/10.1016/i.jhazmat.2010.10.024
Odjegba. VJ: Fasidi. IP. (2006). Effects of heavy metals on some proximate composition of Eichhornia
crassipes. J Appl Sci Environ Manag 10: 83-87.
Odum. HT. (2000). Heavy metals in the environment: Using wetlands for their removal. Boca Raton, FL:
Lewis Publishers.
Ojo. AA; Wood. CM. (2007). In vitro analysis of the bioavailability of six metals via the gastro-intestinal
tract of the rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 83: 10-23.
http://dx.doi.0rg/10.1016/i.aquatox.2007.03.006
Oliveira Ribeiro. CA; Filipak Neto. F; Mela. M; Silva. PH; Randi. MAP; Rabitto. IS: Alves Costa. JRM;
Pelletier. E. (2006). Hematological findings in neotropical fish Hoplias malabaricus exposed to
subchronic and dietary doses of methylmercury, inorganic lead, and tributyltin chloride. Environ Res
101: 74-80. http://dx.doi.0rg/10.1016/j.envres.2005.ll.005
Osman. AGM; McKkawv. IA; Verreth. J: Kirschbaum. F. (2007a). Effects of lead nitrate on the activity of
metabolic enzymes during early developmental stages of the African catfish, Clarias gariepinus
(Burchell, 1822). Fish Physiol Biochem 33: 1-13. http://dx.doi.org/10.1007/sl0695-006-9111-8
Osman. AGM: Wuertz. S: Mekkawv. IA; Exner. HJ; Kirschbaum. F. (2007b). Lead induced
malformations in embryos of the African catfish Clarias gariepinus (Burchell, 1822). Environ Toxicol
22: 375-389. http://dx.doi.org/10.1002/tox.20272
Owojori. OJ; Healev. J: Princz. J: Siciliano. SD. (2011). Can avoidance behavior of the mite Oppia nitens
be used as a rapid toxicity test for soils contaminated with metals or organic chemicals. Environ
Toxicol Chem 30: 2594-2601. http://dx.doi.org/10.1002/etc.658
Paczkowska. M; Kozlowska. M; Golinski. P. (2007). Oxidative stress enzyme activity in Lemna minor
L. exposed to cadmium and lead. Acta Biol Cracov Bot 49: 33-37.
Palaniappan. PR; Krishnakumar. N; Vadivelu. M. (2009). Bioaccumulation of lead and the influence of
chelating agents in Catla catla fmgerlings. Environ Chem Lett 7: 51-54.
http://dx.doi.org/10.1007/sl0311-008-0134-5
Paauin. PR: Gorsuch. JW: Apte. S: Batlev. GE: Bowles. KG: Campbell. PGC: Delos. CG: Pi Toro. DM:
Dwver. RL; Galvez. F; Gensemer. RW: Goss. GG: Hogstrand. C: Janssen. CR; McGeer. JC: Naddv.
RB; Plavle. RC: Santore. RC: Schneider. U; Stubblefield. WA; Wood. CM; Wu. KB. (2002). The
biotic ligand model: A historical overview [Review]. Comp Biochem Physiol C Toxicol Pharmacol
133: 3-35. http://dx.doi.org/10.1016/S1532-0456(02)00112-6
Patel. M; Rogers. JT; Pane. EF; Wood. CM. (2006). Renal responses to acute lead waterborne exposure in
the freshwater rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 80: 362-371.
http://dx.doi.0rg/10.1016/i.aquatox.2006.09.012
Patfaak. SP; Gopal. K. (2009). Bacterial density and antibiotic resistance of Aeromonas sp. in organs of
metal-stressed freshwater fish Channa punctatus. Toxicol Environ Chem 91: 331-337.
http://dx.doi.org/10.1080/02772240802098222
Patrick. GJ: Farmer. JG. (2006). A stable lead isotopic investigation of the use of sycamore tree rings as a
historical biomonitor of environmental lead contamination. Sci Total Environ 362: 278-291.
http://dx.doi.0rg/10.1016/i.scitotenv.2005.12.004
Patrick. L. (2006). Lead toxicity, a review of the literature. Part I: Exposure, evaluation, and treatment
[Review]. Altern Med Rev 11: 2-22.
November 2012 7-275 Draft - Do Not Cite or Quote
-------
Pauget B; Gimbert. F; Coeurdassier. M; Scheifler. R; de Vaufleury. A. (2011). Use of chemical methods
to assess Cd and Pb bioavailability to the snail Cantareus aspersus: A first attempt taking into account
soil characteristics. J Hazard Mater 192: 1804-1811. http://dx.doi.Org/10.1016/i.jhazmat.2011.07.016
Pearce. NJG: Mann. VL. (2006). Trace metal variations in the shells of Ensis siliqua record pollution and
environmental conditions in the sea to the west of mainland Britain. Mar Pollut Bull 52: 739-755.
http://dx.doi.0rg/10.1016/i.marpolbul.2005.ll.003
Penttinen. OP; Kilpi-Koski. J; Jokela. M; Toivainen. K; Vaisanen. A. (2008). Importance of dose metrics
for lethal and sublethal sediment metal toxicity in the oligochaete worm Lumbriculus variegates.
Journal of Soils and Sediments 8: 59-66. http://dx.doi.org/10.1065/iss2007.12.267
Peralta-Videa. JR; Lopez. ML; Naravan. M; Saupe. G: Gardea-Torresdev. J. (2009). The biochemistry of
environmental heavy metal uptake by plants: Implications for the food chain. Int J Biochem Cell Biol
41: 1665-1677. http://dx.doi.Org/10.1016/i.biocel.2009.03.005
Pereira. AA; van Hattum. B; de Boer. J: van Bodegom. PM; Rezende. CE; Salomons. W. (2010). Trace
elements and carbon and nitrogen stable isotopes in organisms from a tropical coastal lagoon. Arch
Environ Contam Toxicol 59: 464-477. http://dx.doi.org/10.1007/s00244-010-9489-2
Perez-Legaspi. IA; Rico-Martinez. R. (2001). Acute toxicity tests on three species of the genus Lecane
(Rotifera: Monogononta). Hydrobiologia 446/447: 375-381.
http://dx.doi.Org/10.1023/A:1017531712808
Pernet-Coudrier. B; Companys. E; Galceran. J: Morev. M; Mouchel. JM; Puy. J: Ruiz. N; Varrault. G.
(2011). Pb-binding to various dissolved organic matter in urban aquatic systems: Key role of the most
hydrophilic fraction. Geochim Cosmo Act 75: 4005-4019. http://dx.doi.Org/10.1016/j.gca.2011.04.030
Perugini. M; Manera. M; Grotta. L; Abete. MC: Tarasco. R: Amorena. M. (2011). Heavy metal (Hg, Cr,
Cd, and Pb) contamination in urban areas and wildlife reserves: Honeybees as bioindicators. Biol
Trace Elem Res 140: 170-176. http://dx.doi.org/10.1007/sl2011-010-8688-z
Peterson. G: Allen. CR: Holling. CS. (1998). Ecological resilience, biodiversity, and scale. Ecosystems 1:
6-18. http://dx.doi.org/10.1007/sl00219900002
Petraitis. PS: Latham. RE. (1999). The importance of scale in testing the origins of alternative community
states. Ecology 80: 429-442. http://dx.doi.org/10.1890/0012-9658(1999)080r0429:TIOSIT12.0.CO:2
Pimentel. D; Wilson. C: McCullum. C: Huang. R; Dwen. P; Flack. J: Tran. Q; Saltman. T; Cliff. B.
(1997). Economic and environmental benefits of biodiversity. Bioscience 47: 747-757.
Pimm. SL. (1984). The complexity and stability of ecosystems [Review]. Nature 307: 321-326.
http://dx.doi.org/10.1038/307321aO
Piotrowska. A; Bajguz. A; Godlewska-Zylkiewicz . B; Zambrzycka. E. (2010). Changes in growth,
biochemical components, and antioxidant activity in aquatic plant Wolffia arrhiza (Lemnaceae)
exposed to cadmium and lead. Arch Environ Contam Toxicol 58: 594-604.
http://dx.doi.org/10.1007/s00244-009-9408-6
Poulton. BC: Allert. AL; Besser. JM; Schmitt. CJ: Brumbaugh. WG: Fairchild. JF. (2010). A
macroinvertebrate assessment of Ozark streams located in lead-zinc mining areas of the Viburnum
Trend in southeastern Missouri, USA. Environ Monit Assess 163: 619-641.
http://dx.doi.org/10.1007/sl0661-009-0864-2
Probst. A; Liu. H; Fanjul. M; Liao. B; Hollande. E. (2009). Response of Vicia faba L. to metal toxicity on
mine tailing substrate: Geochemical and morphological changes in leaf and root. Environ Exp Bot 66:
297-308. http://dx.doi.0rg/10.1016/i.envexpbot.2009.02.003
Quan. W: Zhu. J: Ni. Y; Shi. L; Chen. Y. (2009). Fauna! utilization of constructed intertidal oyster
(Crassostrea rivularis) reef in the Yangtze River estuary, China. Ecol Eng 35: 1466-1475.
http://dx.doi.0rg/10.1016/i.ecoleng.2009.06.001
November 2012 7-276 Draft - Do Not Cite or Quote
-------
Rabitto. IS: Alves Costa. JRM; Silva de Assis. HC: Pelletier. E; Akaishi. FM; Anjos. A; Randi. MAP;
Oliveira Ribeiro. CA. (2005). Effects of dietary Pb(II) and tributyltin on neotropical fish, Hoplias
malabaricus: Histopathological and biochemical findings. Ecotoxicol Environ Saf 60: 147-156.
http://dx.doi.0rg/10.1016/i.ecoenv.2004.03.002
Radwan. MA; El-Gendv. KS: Gad. AF. (2010). Oxidative stress biomarkers in the digestive gland of
Theba pisana exposed to heavy metals. Arch Environ Contam Toxicol 58: 828-835.
http://dx.doi.org/10.1007/s00244-009-9380-l
Rai. PK. (2008). Heavy metal pollution in aquatic ecosystems and its phytoremediation using wetland
plants: An ecosustainable approach [Review]. Int J Phytoremediation 10: 133-160.
http://dx.doi.org/10.1080/15226510801913918
Raimundo. J: Vale. C: Duarte. R; Moura. I. (2008). Sub-cellular partitioning of Zn, Cu, Cd and Pb in the
digestive gland of native Octopus vulgaris exposed to different metal concentrations (Portugal). Sci
Total Environ 390: 410-416. http://dx.doi.Org/10.1016/i.scitotenv.2007.10.029
Rainbow. PS. (1996). Heavy metals in aquatic invertebrates. In WN Beyer; GH Heinz; AW Redmon-
Norwood (Eds.), Environmental contaminants in wildlife: Interpreting tissue concentrations (pp.
405-425). Boca Raton, FL: CRC Press.
Rainbow. PS; Poirier. L; Smith. BD; Brix. KV; Luoma. SN. (2006). Trophic transfer of trace metals from
the polychaete worm Nereis diversicolor to the polychaete N-virens and the decapod crustacean
Palaemonetes varians. Mar Ecol Prog Ser 321: 167-181. http://dx.doi.org/10.3354/meps321167
Ramesh. M; Saravanan. M; Kavitha. C. (2009). Hormonal responses of the fish, Cyprinus carpio, to
environmental lead exposure. Afr J Biotechnol 8: 4154-4158.
Rau. S; Miersch. J; Neumann. D; Weber. E; Krauss. GJ. (2007). Biochemical responses of the aquatic
moss Fontinalis antipyretica to Cd, Cu, Pb and Zn determined by chlorophyll fluorescence and protein
levels. Environ Exp Bot 59: 299-306. http://dx.doi.Org/10.1016/j.envexpbot.2006.03.001
Reddv. AM; Kumar. SG; Jvothsnakumari. G; Thimmanaik. S; Sudhakar. C. (2005). Lead induced changes
in antioxidant metabolism of horsegram (Macrotyloma uniflorum (Lam.) Verde.) and bengalgram
(Cicer arietinum L.). Chemosphere 60: 97-104. http://dx.doi.Org/10.1016/i.chemosphere.2004.ll.092
Reglero. MM; Monsalve-Gonzalez. L; Taggart. MA; Mateo. R. (2008). Transfer of metals to plants and
red deer in an old lead mining area in Spain. Sci Total Environ 406: 287-297.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.06.001
Reglero. MM; Taggart. MA; Castellanos. P; Mateo. R. (2009a). Reduced sperm quality in relation to
oxidative stress in red deer from a lead mining area. Environ Pollut 157: 2209-2215.
http://dx.doi.0rg/10.1016/i.envpol.2009.04.017
Reichmuth. JM; Weis. P; Weis. JS. (2010). Bioaccumulation and depuration of metals in blue crabs
(Callinectes sapidus Rathbun) from a contaminated and clean estuary. Environ Pollut 158: 361-368.
http://dx.doi.0rg/10.1016/i.envpol.2009.09.009
Reinfelder. JR; Fisher. NS; Luoma. SN; Nichols. JW; Wang. WX. (1998). Trace element trophic transfer
in aquatic organisms: A critique of the kinetic model approach. Sci Total Environ 219: 117-135.
http://dx.doi.org/10.1016/S0048-9697(98)00225-3
Reynolds. KD; Schwarz. MS; Mcfarland. CA; Mcbride. T; Adair. B. (2006). Northern pocket gophers
(Thomomys talpoides) as biomonitors of environmental metal contamination. Environ Toxicol Chem
25: 458-469. http://dx.doi.Org/10.1897/05-130Rl.l
Rhea. DT; Harper. DP; Farag. AM; Brumbaugh. WG. (2006). Biomonitoring in the Boulder River
watershed, Montana, USA: Metal concentrations in biofilm and macroinvertebrates, and relations with
macroinvertebrate assemblage. Environ Monit Assess 115: 381-393.
http://dx.doi.org/10.1007/sl0661-006-7086-7
November 2012 7-277 Draft - Do Not Cite or Quote
-------
Rice. C: Ghorai. JK; Zalewski. K; Weber. DN. (2011). Developmental lead exposure causes startle
response deficits in zebrafish. Aquat Toxicol 105: 600-608.
http://dx.doi.0rg/10.1016/i.aquatox.2011.08.014
Ringenary. MJ; Molof. AH; Tanacredi. JT; Schreibman. MP; Kostarelos. K. (2007). Long-term sediment
bioassay of lead toxicity in two generations of the marine amphipod Elasmopus laevis, SI Smith,
1873. Environ Toxicol Chem 26: 1700-1710. http://dx.doi.Org/10.1897/06-303Rl.l
Robinson. GR; Sibrell. PL; Boughton. CJ: Yang. LH. (2007). Influence of soil chemistry on metal and
bioessential element concentrations in nymphal and adult periodical cicadas (Magicicada spp.). Sci
Total Environ 374: 367-378. http://dx.doi.Org/10.1016/i.scitotenv.2006.12.031
Rodriguez-Estival. J: Martinez-Haro. M; Monsalve-Gonzalez. L; Mateo. R. (2011). Interactions between
endogenous and dietary antioxidants against Pb-induced oxidative stress in wild ungulates from a Pb
polluted mining area. Sci Total Environ 409: 2725-2733.
http://dx.doi.0rg/10.1016/i.scitotenv.2011.04.010
Rogers. JT; Patel. M; Gilmour. KM; Wood. CM. (2005). Mechanisms behind Pb-induced disruption of
Na+ and Cl- balance in rainbow trout (Oncorhynchus mykiss). Am J Physiol Regul Integr Comp
Physiol 58: R463-R472. http://dx.doi.org/10.1152/aipregu.00362.2004
Rogers. JT: Wood. CM. (2004). Characterization of branchial lead-calcium interaction in the freshwater
rainbow trout (Oncorhynchus mykiss). J Exp Biol 207: 813-825. http://dx.doi.org/10.1242/jeb.00826
Rogers. JT: Wood. JG. (2003). lonoregulatory disruption as the acute toxic mechanism for lead in the
rainbow trout (Oncorhynchus mykiss). Aquat Toxicol 64: 215-234.
http://dx.doi.org/10.1016/S0166-445X(03)00053-5
Rogival. D; Scheirs. J: Blust. R. (2007). Transfer and accumulation of metals in a soil-diet-wood mouse
food chain along a metal pollution gradient. Environ Pollut 145: 516-528.
http://dx.doi.0rg/10.1016/i.envpol.2006.04.019
Rokytova. L; Kula. E; Kodarova. L; Peslova. A. (2004). Feeding of the willow leaf beetle Lochmaea
capreae L. (Coleoptera, Chrysomelidae) on leaves of birch (Betula pendula Roth) contaminated by
heavy metals. J Forest Sci 50: 109-117.
Roodbergen. M; Klok. C: van der Hout. A. (2008). Transfer of heavy metals in the food chain earthworm
Black-tailed godwit (Limosa limosa): Comparison of a polluted and a reference site in The
Netherlands. Sci Total Environ 406: 407-412. http://dx.doi.Org/10.1016/i.scitotenv.2008.06.051
Roulier. JL; Tusseau-Nuillemin. MH; Coquerv. M; Geffard. O: Garric. J. (2008a). Measurement of
dynamic mobilization of trace metals in sediments using DOT and comparison with bioaccumulation
in Chironomus riparius: First results of an experimental study. Chemosphere 70: 925-932.
http://dx.doi.0rg/10.1016/i.chemosphere.2007.06.061
Rozon-Ramilo. LD; Dube. MG: Squires. AJ; Niyogi. S. (2011). Examining waterborne and dietborne
routes of exposure and their contribution to biological response patterns in fathead minnow
(Pimephales promelas). Aquat Toxicol 105: 466-481. http://dx.doi.Org/10.1016/i.aquatox.2011.07.006
Rubio-Franchini. I; Saavedra. JM; Rico-Martinez. R. (2008). Determination of lead in samples of
zooplankton, water, and sediments in a Mexican reservoir: Evidence for lead biomagnification in
lower/intermediate trophic levels? Environ Toxicol 23: 459-465. http://dx.doi.org/10.1002/tox.20357
Ruelas-Inzunza. J: Paez-Osuna. F. (2008). Trophic distribution of Cd, Pb, and Zn in a food web from
Altata-Ensenada del Pabellon subtropical lagoon, SE Gulf of California. Arch Environ Contam
Toxicol 54: 584-596. http://dx.doi.org/10.1007/s00244-007-9075-4
Ruiz. E; Alonso-Azcarate. J: Rodriguez. L. (2011). Lumbricus terrestris L. activity increases the
availability of metals and their accumulation in maize and barley. Environ Pollut 159: 722-728.
http://dx.doi.0rg/10.1016/i.envpol.2010.ll.032
Sadiq. M. (1992). Lead in marine environments. In Toxic metal chemistry in marine environments. New
York, NY: Marcel Dekker, Inc.
November 2012 7-278 Draft - Do Not Cite or Quote
-------
Salice. CJ: Suski. JG: Bazar. MA; Talent. LG. (2009). Effects of inorganic lead on Western fence lizards
(Sceloporus occidentalis). Environ Pollut 157: 3457-3464.
http://dx.doi.0rg/10.1016/i.envpol.2009.06.013
Samecka-Cymerman. A; Kempers. AJ. (2007). Heavy metals in aquatic macrophytes from two small
rivers polluted by urban, agricultural and textile industry sewages SW Poland. Arch Environ Contam
Toxicol 53: 198-206. http://dx.doi.org/10.1007/s00244-006-0059-6
Sanchez-Marin. P..; Bellas. J..: Mubiana. V.. K.; Lorenzo. J.. I.; Blust. R..: Beiras. R.. (2011). Pb
uptake by the marine mussel Mytilus sp. Interactions with dissolved organic matter. Aquat Toxicol
102: 48-57. http://dx.doi.Org/10.1016/j.aquatox.2010.12.012
Sanchez-Marin. P; Beiras. R. (2011). Adsorption of different types of dissolved organic matter to marine
phytoplankton and implications for phytoplankton growth and Pb bioavailability. Journal of Plankton
Research 33: 1396-1409. http://dx.doi.org/10.1093/plankt/fbr039
Sanchez-Marin. P; Lorenzo. JI; Blust. R: Beiras. R. (2007). Humic acids increase dissolved lead
bioavailability for marine invertebrates. Environ Sci Technol 41: 5679-5684.
http://dx.doi.org/10.1021/es070088h
Sanchez-Marin. P; Santos-Echeandia. J: Nieto-Cid. M; Alvarez-Salgado. XA; Beiras. R. (2010a). Effect
of dissolved organic matter (DOM) of contrasting origins on Cu and Pb speciation and toxicity to
Paracentrotus lividus larvae. Aquat Toxicol 96: 90-102.
http://dx.doi.0rg/10.1016/i.aquatox.2009.10.005
Sanchez-Marin. P; Slaveykova. VI; Beiras. R. (2010b). Cu and Pb accumulation by the marine diatom
Thalassiosira weissflogii in the presence of humic acids. Environ Chem 7: 309-317.
http://dx.doi.org/10.1071/enl0015
Savard. MM; Begin. C: Parent. M; Marion. J: Smirnoff. A. (2006). Dendrogeochemical distinction
between geogenic and anthropogenic emissions of metals and gases near a copper smelter.
Geochemistry: Exploration, Environment, Analysis 6: 237-247.
http://dx.doi.org/10.1144/1467-7873/05-096
Sawasdee. B; Kohler. HR. (2010). Metal sensitivity of the embryonic development of the ramshorn snail
Marisa cornuarietis (Prosobranchia). Ecotoxicology 19: 1487-1495.
http://dx.doi.org/10.1007/sl0646-010-0534-8
Scheidegger. C: Behra. R: Sigg. L. (2011). Phytochelatin formation kinetics and toxic effects in the
freshwater alga Chlamydomonas reinhardtii upon short- and long-term exposure to lead(II). Aquat
Toxicol 101: 423-429. http://dx.doi.Org/10.1016/i.aauatox.2010.ll.016
Scheifler. R; Coeurdassier. M; Morilhat. C: Bernard. N; Faivre. B; Flicoteaux. P; Giraudoux. P; Noel. M;
Piotte. P: Rieffel. D; de Vaufleury. A; Badot. PM. (2006a). Lead concentrations in feathers and blood
of common blackbirds (Turdus merula) and in earthworms inhabiting unpolluted and moderately
polluted urban areas. Sci Total Environ 371: 197-205.
http://dx.doi.0rg/10.1016/i.scitotenv.2006.09.011
Scheifler. R; De Vaufleury. A; Coeurdassier. M; Crini. N; Badot. PM. (2006b). Transfer of Cd, Cu, Ni, Pb,
and Zn in a soil-plant-invertebrate food chain: A microcosm study. Environ Toxicol Chem 25:
815-822. http://dx.doi.Org/10.1897/04-675R.l
Schipper. AM; Wiinhoven. S: Leuven. R; Ragas. AMJ; Hendriks. AJ. (2008). Spatial distribution and
internal metal concentrations of terrestrial arthropods in a moderately contaminated lowland
floodplain along the Rhine River. Environ Pollut 151: 17-26.
http://dx.doi.0rg/10.1016/i.envpol.2007.03.007
Schmitt. CJ: Whyte. JJ: Brumbaugh. WG: Tillitt. DE. (2005). Biochemical effects of lead, zinc, and
cadmium from mining on fish in the Tri-States District of northeastern Oklahoma, USA. Environ
Toxicol Chem 24: 1483-1495. http://dx.doi.Org/10.1897/04-332R.l
November 2012 7-279 Draft - Do Not Cite or Quote
-------
Schmitt. CJ: Whyte. JJ: Roberts. AP; Annis. ML; May. TW: Tilitt. DE. (2007b). Biomarkers of metals
exposure in fish from lead-zinc mining areas of southeastern Missouri, USA. Ecotoxicol Environ Saf
67: 31-47. http://dx.doi.Org/10.1016/i.ecoenv.2006.12.011
Schroth. AW: Bostick. BC: Kaste. JM; Friedland. AJ. (2008). Lead sequestration and species
redistribution during soil organic matter decomposition. Environ Sci Technol 42: 3627-3633.
http://dx.doi.org/10.1021/es703002b
Semreen. MH; Aboul-Enein. HY. (2011). Determination of heavy metal content in wild-edible mushroom
from Jordan. Anal Lett 44: 932-941. http://dx.doi.org/10.1080/00032711003790072
Serafim. A; Lopes. B; Company. R; Cravo. A; Gomes. T; Sousa. V: Bebianno. MJ. (2011). Amulti-
biomarker approach in cross-transplanted mussels Mytilus galloprovincialis. Ecotoxicology 20:
1959-1974. http://dx.doi.org/10.1007/sl0646-011-0737-7
Shacklette. HT; Boerngen. JG. (1984). Element concentrations in soils and other surficial materials of the
conterminous United States. (1270). Washington D.C.: Government Printing Office.
http://pubs.usgs.gov/pp/1270/
Shaheen. SM; Tsadilas. CD. (2009). Concentration of lead in soils and some vegetable plants in north
Nile Delta as affected by soil type and irrigation water. Commun Soil Sci Plant Anal 40: 327-344.
http://dx.doi.org/10.1080/00103620802649237
Shakya. K; Chettri. MK; Sawidis. T. (2008). Impact of heavy metals (copper, zinc, and lead) on the
chlorophyll content of some mosses. Arch Environ Contam Toxicol 54: 412-421.
http://dx.doi.org/10.1007/s00244-007-9060-y
Shore. RF; Rattner. BA. (2001). Ecotoxicology of wild mammals. In RF Shore; BA Rattner (Eds.).
Hoboken, NJ: John Wiley & Sons.
Shu. YH; Zhou. JL; Tang. WC: Lu. K; Zhou. Q; Zhang. GR. (2009). Molecular characterization and
expression pattern of Spodoptera litura (Lepidoptera: Noctuidae) vitellogenin, and its response to lead
stress. J Insect Physiol 55: 608-616. http://dx.doi.Org/10.1016/i.jinsphys.2009.03.005
Shuhaimi-Othman. M; Yakub. N; Ramie. NA; Abas. A. (2011 a). Sensitivity of the freshwater prawn,
Macrobrachium lanchesteri (Crustacea: Decapoda), to heavy metals. Toxicol Ind Health 27: 523-530.
http://dx.doi.org/10.1177/0748233710391993
Shuhaimi-Othman. M; Yakub. N; Ramie. NA; Abas. A. (20 lib). Toxicity of metals to a freshwater
ostracod: Stenocypris major. Journal of Toxicology 2011: 136104.
http://dx.doi.org/10.1155/2011/136104
Shuhaimi-Othman. M; Yakub. N; Umirah. NS: Abas. A. (201 Ic). Toxicity of eight metals to Malaysian
freshwater midge larvae Chironomus javanus (Diptera, Chironomidae). Toxicol Ind Health 27:
879-886. http://dx.doi.org/10.1177/0748233711399318
Simkiss. K; Jenkins. KGA; Mclellan. J; Wheeler. E. (1982). Methods of metal incorporation into
intracellular granules. Experientia 38: 333-335. http://dx.doi.org/10.1007/BF01949375
Simpson. SL; Batlev. GE. (2007). Predicting metal toxicity in sediments: A critique of current approaches
[Review]. Integr Environ Assess Manag 3: 18-31. http://dx.doi.org/10.1002/ieam.5630030103
Singer. C: Zimmermann. S: Sures. B. (2005). Induction of heat shock proteins (hsp70) in the zebra mussel
(Dreissena polymorpha) following exposure to platinum group metals (platinum, palladium and
rhodium): Comparison with lead and cadmium exposures. Aquat Toxicol 75: 65-75.
http://dx.doi.0rg/10.1016/i.aquatox.2005.07.004
Singh. R; Tripathi. RD; Dwivedi. S: Kumar. A; Trivedi. PK; Chakrabartv. D. (2010). Lead
bioaccumulation potential of an aquatic macrophyte Najas indica are related to antioxidant system.
Bioresour Technol 101: 3025-3032. http://dx.doi.Org/10.1016/i.biortech.2009.12.031
Sizmur. T; Hodson. ME. (2009). Do earthworms impact metal mobility and availability in soil? A review
[Review]. Environ Pollut 157: 1981-1989. http://dx.doi.Org/10.1016/i.envpol.2009.02.029
November 2012 7-280 Draft - Do Not Cite or Quote
-------
Sizmur. T; Hodson. MF. (2008). The impact of Eisenia veneta on As, Cu, Pb and Zn uptake by ryegrass
(Lolium perenne L.). Mineral Mag 72: 495-499. http://dx.doi.Org/10.1180/minmag.2008.072.l.495
Sizmur. T; Tilston. EL; Charnock. J; Palumbo-Roe. B; Watts. MJ; Hodson. ME. (2011). Impacts of
epigeic, anecic and endogeic earthworms on metal and metalloid mobility and availability. J Environ
Monit 13: 266-273. http://dx.doi.org/10.1039/cOem00519c
Skorbiowicz. E. (2006). Cadmium and lead in bottom sediments and reed sweet grass (Glycerii aquatica)
roots in selected rivers of upper Narew river basins. Pol J Environ Stud 15: 482-489.
Slaveykova. VI; Wilkinson. KJ. (2005). Predicting the bioavailability of metals and metal complexes:
Critical review of the biotic ligand model [Review]. Environ Chem 2: 9-24.
http://dx.doi.org/10.1071/EN04076
Sloman. KA; Lepage. O: Rogers. JT; Wood. CM; Winberg. S. (2005). Socially-mediated differences in
brain monoamines in rainbow trout: Effects of trace metal contaminants. Aquat Toxicol 71: 237-247.
http://dx.doi.0rg/10.1016/i.aquatox.2004.ll.008
Smith. DM; Mielke. HW: Heneghan. JB. (2009a). Subchronic lead feeding study in male rats and
micropigs. Environ Toxicol 24: 453-461. http://dx.doi.org/10.1002/tox.20448
Smith. JT; Walker. LA; Shore. RF; Durell. S: Howe. PD; Taylor. M. (2009b). Do estuaries pose a toxic
contamination risk for wading birds? Ecotoxicology 18: 906-917.
http://dx.doi.org/10.1007/sl0646-009-0352-z
Smolders. E; McGrath. S: Fairbrother. A; Hale. B; Lombi. E; McLaughlin. M; Rutgers. M; Van der Vliet.
L_. (2007). Hazard assessment of inorganic metals and metal substances in terrestrial systems. In
Assessing the hazard of metals and inorganic metal substances in aquatic and terrestrial systems. Boca
Raton, FL: CRC Press, http://dx.doi.org/10.1201/9781420044416.ch6
Smolders. E; Ports. K; van Sprang. P; Schoeters. I; Janssen. CR; McGrath. SP; McLaughlin. MJ. (2009).
Toxicity of trace metals in soil as affected by soil type and aging after contamination: Using calibrated
bioavailability models to set ecological soil standards. Environ Toxicol Chem 28: 1633-1642.
http://dx.doi.0rg/10.1897/08-592.l
Snoeijs. T; Dauwe. T; Pinxten. R; Darras. VM; Arckens. L; Eens. M. (2005). The combined effect of lead
exposure and high or low dietary calcium on health and immunocompetence in the zebra finch
(Taeniopygia guttata). Environ Pollut 134: 123-132. http://dx.doi.Org/10.1016/i.envpol.2004.07.009
Sobrino-Figueroa. A; Caceres-Martinez. C. (2009). Alterations of valve closing behavior in juvenile
Catarina scallops (Argopecten ventricosus Sowerby, 1842) exposed to toxic metals. Ecotoxicology 18:
983-987. http://dx.doi.org/10.1007/sl0646-009-0358-6
Sobrino-Figueroa. AS: Caceres-Martinez. C: Botello. AV; Nunez-Nogueira. G. (2007). Effect of
cadmium, chromium, lead and metal mixtures on survival and growth of juveniles of the scallop
Argopecten ventricosus (Sowerby II, 1842). J Environ Sci Health A Tox Hazard Subst Environ Eng
42: 1443-1447. http://dx.doi.org/10.1080/10934520701480821
Sobrino. AS: Miranda. MG: Alvarez. C: Quiroz. A. (2010). Bio-accumulation and toxicity of lead (Pb) in
Lemnagibba L (duckweed). J Environ Sci Health A Tox Hazard Subst Environ Eng 45: 107-110.
http://dx.doi.org/10.1080/10934520903389267
Son. J; Ryoo. MI; Jung. J: Cho. K. (2007). Effects of cadmium, mercury and lead on the survival and
instantaneous rate of increase of Paronychiurus kimi (Lee) (Collembola). Appl Soil Ecol 35: 404-411.
http://dx.doi.0rg/10.1016/i.apsoil.2006.07.002
Sonmez. O; Bukun. B; Kava. C: Aydemir. S. (2008). The assessment of tolerance to heavy metals (Cd,Pb
and Zn) and their accumulation in three weed species. Pakistan J Bot 40: 747-754.
Soto-Jimenez. MF; Arellano-Fiore. C: Rocha-Velarde. R; Jara-Marini. ME; Ruelas-Inzunza. J: Paez-
Osuna. F (2011 a). Trophic transfer of lead through a model marine four-level food chain: Tetraselmis
suecica, Artemia franciscana, Litopenaeus vannamei, and Haemulon scudderi. Arch Environ Contam
Toxicol 61: 280-291. http://dx.doi.org/10.1007/s00244-010-9620-4
November 2012 7-281 Draft - Do Not Cite or Quote
-------
Soto-Jimenez. MF; Arellano-Fiore. C: Rocha-Velarde. R; Jara-Marini. ME; Ruelas-Inzunza. J: Voltolina.
D; Frias-Espericueta. MG: Quintero-Alvarez. JM; Paez-Osuna. F. (201 Ib). Biological responses of a
simulated marine food chain to lead addition. Environ Toxicol Chem 30: 1611-1617.
http://dx.doi.org/10.1002/etc.537
Southward Hogan. L; Marschall. E; Folt. C: Stein. RA. (2007). How non-native species in Lake Erie
influence trophic transfer of mercury and lead to top predators. J Great Lakes Res 33: 46-61.
http://dx.doi.org/10.3394/0380-1330(2007)33r46:HNSrLE12.0.CO:2
Sparling. DW: Krest S: Ortiz-Santaliestra. M. (2006). Effects of lead-contaminated sediment on Rana
sphenocephala tadpoles. Arch Environ Contam Toxicol 51: 458-466.
http://dx.doi.org/10.1007/s00244-005-0243-0
Spears. BL; Hansen. JA; Audet. DJ. (2007). Blood lead concentrations in waterfowl utilizing Lake Coeur
d'Alene, Idaho. Arch Environ Contam Toxicol 52: 121-128.
http://dx.doi.org/10.1007/s00244-006-0061-z
Spokas. EG: Spur. BW: Smith. H; Kemp. FW: Bogden. JD. (2006). Tissue lead concentration during
chronic exposure of Pimephales promelas (fathead minnow) to lead nitrate in aquarium water. Environ
Sci Technol 40: 6852-6858. http://dx.doi.org/10.1021/es060811o
Spry. DJ; Wiener. JG. (1991). Metal bioavailability and toxicity to fish in low-alkalinity lakes: a critical
review [Review]. Environ Pollut 71: 243-304. http://dx.doi.org/10.1016/0269-7491(91)90034-T
Spurgeon. DJ: Hopkin. SP; Jones. DT. (1994). Effects of cadmium, copper, lead and zinc on growth,
reproduction and survival of the earthworm Eisenia fetida (Savigny): Assessing the environmental
impact of point-source metal contamination in terrestrial ecosystems. Environ Pollut 84: 123-130.
http://dx.doi.org/10.1016/0269-7491(94)90094-9
Stavros. HCW: Bossart. GD; Hulsev. TC: Fair. PA. (2007). Trace element concentrations in skin of free-
ranging bottlenose dolphins (Tursiops truncatus) from the southeast Atlantic coast. Sci Total Environ
388: 300-315. http://dx.doi.Org/10.1016/i.scitotenv.2007.07.030
Stefanowicz. AM; Niklinska. M; Laskowski. R. (2008). Metals affect soil bacterial and fungal functional
diversity differently. Environ Toxicol Chem 27: 591-598. http://dx.doi.Org/10.1897/07-288.l
Stobrawa. K; Lorenc-Plucinska. G. (2008). Thresholds of heavy-metal toxicity in cuttings of European
black poplar (Populus nigra L.) determined according to antioxidant status of fine roots and
morphometrical disorders. Sci Total Environ 390: 86-96.
http://dx.doi.0rg/10.1016/i.scitotenv.2007.09.024
Strom. SM; Patnode. KA; Langenberg. JA; Bodenstein. BL; Scheuhammer. AM. (2005). Lead
contamination in American woodcock (Scolopax minor) from Wisconsin. Arch Environ Contam
Toxicol 49: 396-402. http://dx.doi.org/10.1007/s00244-005-7063-0
Sudova. R: Vosatka. M. (2007). Differences in the effects of three arbuscular mycorrhizal fungal strains
on P and Pb accumulation by maize plants. Plant Soil 296: 77-83.
http://dx.doi.org/10.1007/slll04-007-9291-8
Suicmez. M; Kayim. M; Koseoglu. D; Hasdemir. E. (2006). Toxic effects of lead on the liver and gills of
Oncorhynchus mykiss Walbaum 1792. Bull Environ Contam Toxicol 77: 551-558.
http://dx.doi.org/10.1007/s00128-006-1099-3
Sun. SQ: Wang. DY; He. M; Zhang. C. (2009). Monitoring of atmospheric heavy metal deposition in
Chongqing, China: Based on moss bag technique. Environ Monit Assess 148: 1-9.
http://dx.doi.org/10.1007/sl0661-007-0133-l
Sun. SQ: Wang. GX; He. M; Cao. T. (2011). Effects of Pb and Ni stress on oxidative stress parameters in
three moss species. Ecotoxicol Environ Saf 74: 1630-1635.
http://dx.doi.0rg/10.1016/i.ecoenv.2011.04.002
November 2012 7-282 Draft - Do Not Cite or Quote
-------
Sundbv. B; Caetano. M; Vale. C: Gobeil. C: Luther. GW. Ill; Nuzzio. DB. (2005). Root-induced cycling
of lead in salt marsh sediments. Environ Sci Technol 39: 2080-2086.
http://dx.doi.org/10.1021/es048749n
Suter. GW: Norton. SB; Fairbrother. A. (2005). Individuals versus organisms versus populations in the
definition of ecological assessment endpoints. Integr Environ Assess Manag 1: 397-400.
http://dx.doi.org/10.1002/ieam.5630010409
Suter. GW: Rodier. DJ; Schwenk. S: Trover. ME; Tyler. PL; Urban. DJ; Wellman. MC: Wharton. S.
(2004). The U.S. Environmental Protection Agency's Generic Ecological Assessment Endpoints. Hum
Ecol Risk Assess 10: 967-981. http://dx.doi.org/10.1080/10807030490887104
Suthar. S: Singh. S: Dhawan. S. (2008). Earthworms as bioindicator of metals (Zn, Fe, Mn, Cu, Pb and
Cd) in soils: Is metal bioaccumulation affected by their ecological category? Ecol Eng 32: 99-107.
http://dx.doi.0rg/10.1016/i.ecoleng.2007.10.003
Svendsen. C: Hankard. PK; Lister. LJ; Fishwick. SK; Jonker. MJ; Spurgeon. DJ. (2007). Effect of
temperature and season on reproduction, neutral red retention and metallothionein responses of
earthworms exposed to metals in field soils. Environ Pollut 147: 83-93.
http://dx.doi.0rg/10.1016/i.envpol.2006.08.012
Swarup. D: Naresh. R; Varshnev. VP; Balagangatharathilagar. M; Kumar. P; Nandi. D; Patra. RC. (2007).
Changes in plasma hormones profile and liver function in cows naturally exposed to lead and
cadmium around different industrial areas. Res Vet Sci 82: 16-21.
http://dx.doi.0rg/10.1016/i.rvsc.2006.05.002
Szivak. I; Behra. R; Sigg. L. (2009). Metal-induced reactive oxygen species production in
Chlamydomonas reinhardtii (chlorophyceae). J Phycol 45: 427-435.
http://dx.doi.0rg/10.llll/i.1529-8817.2009.00663.x
Tamura. H; Honda. M; Sato. T; Kamachi. H. (2005). Pb hyperaccumulation and tolerance in common
buckwheat (Fagopyrum esculentum Moench). J Plant Res 118: 355-359.
http://dx.doi.org/10.1007/sl0265-005-0229-z
Theegala. CS: Suleiman. AA; Carriere. PA. (2007). Toxicity and biouptake of lead and arsenic by
Daphnia pulex. J Environ Sci Health A Tox Hazard Subst Environ Eng 42: 27-31.
http://dx.doi.org/10.1080/10934520601015404
Tilman. D. (1996). Biodiversity: Population versus ecosystem stability. Ecology 77: 350-363.
http://dx.doi.org/10.2307/2265614
Tilman. D. (2000). Causes, consequences and ethics of biodiversity. Nature 405: 208-211.
http://dx.doi.org/10.1038/35012217
Tilman. D; Downing. JA. (1994). Biodiversity and stability in grasslands. Nature 367: 363-365.
http://dx.doi.org/10.1038/367363aO
Tollett. VD: Benvenutti. EL; Deer. LA; Rice. TM. (2009). Differential toxicity to Cd, Pb, and Cu in
dragonfly larvae (Insecta: Odonata). Arch Environ Contam Toxicol 56: 77-84.
http://dx.doi.org/10.1007/s00244-008-9170-l
U.S. EPA (U.S. Environmental Protection Agency). (1977). Air quality criteria for lead [EPA Report].
(EPA-600/8-77-017). Washington, D.C.
http://nepis.epa.gov/Exe/ZvPURL.cgi?Dockev=20013GWR.txt
U.S. EPA (U.S. Environmental Protection Agency). (1985). Ambient water quality criteria for lead - 1984
[EPA Report]. (EPA 440/5-84-027). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (1986a). Air quality criteria for lead [EPA Report].
(EPA/600/8-83/028aF-dF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
November 2012 7-283 Draft - Do Not Cite or Quote
-------
U.S. EPA (U.S. Environmental Protection Agency). (1986b). Air quality criteria for lead: Volume I of IV
[EPA Report]. (EPA-600/8-83/028aF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=32647
U.S. EPA (U.S. Environmental Protection Agency). (1998). Guidelines for ecological risk assessment
[EPA Report]. (EPA/630/R-95/002F). Washington, D.C.
http://www.epa.gov/raf/publications/guidelines-ecological-risk-assessment.htm
U.S. EPA (U.S. Environmental Protection Agency). (2002b). National recommended water quality
criteria: 2002 [EPA Report]. (EPA-822-R-02-047). Washington, D.C.
http://nepis.epa.gov/Exe/ZvPURL.cgi?Dockev=P1005EYQ.txt
U.S. EPA (U.S. Environmental Protection Agency). (2003a). Generic ecological assessment endpoints
(GEAEs) for ecological risk assessment [EPA Report]. (EPA-630/P-02/004F). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2003b). Guidance for developing ecological soil
screening levels (Eco-SSLs): Review of background concentration for metals - Attachment 1-4 [EPA
Report]. (OSWER Directive 92857-55). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2004). Air quality criteria for paniculate matter [EPA
Report]. (EPA/600/P-99/002aF-bF). Research Triangle Park, NC.
http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=87903
U.S. EPA (U.S. Environmental Protection Agency). (2005b). Ecological soil screening levels for lead:
Interim final [EPA Report]. (OSWER Directive 9285.7-70). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2005d). Procedures for the derivation of equilibrium
partitioning sediment benchmarks (ESBs) for the protection of benthic organisms: Metal mixtures
(cadmium, copper, lead, nickel, silver and zinc) [EPA Report]. (EPA-600-R-02-011).
Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2006b). Air quality criteria for lead: Volume I of II
[EPA Report]. (EPA/600/R-05/144aF). Research Triangle Park, NC.
http ://cfpub. epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
U.S. EPA (U.S. Environmental Protection Agency). (2006c). Air quality criteria for lead: Volume II of II
[EPA Report]. (EPA/600/R-05/144bF). Research Triangle Park, NC.
http ://cfpub .epa. gov/ncea/CFM/recordisplav. cfm?deid= 158823
U.S. EPA (U.S. Environmental Protection Agency). (2007a). Aquatic life ambient freshwater quality
criteria - Copper: 2007 Revision [EPA Report]. (EPA-822/R-07-001). Washington, D.C.
http://water.epa.gov/scitech/swguidance/standards/criteria/aqlife/pollutants/copper/2007 index.cfm
U.S. EPA (U.S. Environmental Protection Agency). (2007c). Framework for metals risk assessment [EPA
Report]. (EPA 120/R-07/001). Washington, D.C. http://www.epa.gov/raf/metalsframework/index.htm
U.S. EPA (U.S. Environmental Protection Agency). (2007d). Guidance for developing ecological soil
screening levels [EPA Report]. (OSWER Directive 92857-55). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2008b). Ambient aquatic life water quality: Lead
(draft) [EPA Report]. (EPA Contract No. 68-C-04-006). Washington, D.C.
U.S. EPA (U.S. Environmental Protection Agency). (2008e). Integrated science assessment for oxides of
nitrogen and sulfur: Ecological criteria [EPA Report]. (EPA/600/R-08/082F). Research Triangle Park,
NC. http://cfpub.epa.gov/ncea/cfm/recordisplav.cfm?deid=201485
U.S. EPA (U.S. Environmental Protection Agency). (2009b). National recommended water quality criteria
[EPA Report]. Washington, D.C. http://www.epa.gov/waterscience/criteria/wqctable/nrwqc-2009.pdf
UNEP (United Nations Environment Programme). (2003). Ecosystems and human well-being: A
framework for assessment. Washington, DC: Island Press.
November 2012 7-284 Draft - Do Not Cite or Quote
-------
Uvsal. Y; Taner. F. (2009). Effect of pH, temperature, and lead concentration on the bioremoval of lead
from water using Lemna minor. Int J Phytoremediation 11: 591-608.
http://dx.doi.org/10.1080/15226510902717648
Vaisvalavicius. R; Motuzas. A; Prosvcevas. I; Levinskaite. L; Zakarauskaite. D; Grigaliuniene. K; Butkus.
V (2006). Effect of heavy metals on microbial communities and enzymatic activity in soil column
experiment. Arch Agron Soil Sci 52: 161-169. http://dx.doi.org/10.1080/03650340600566431
van Capelleveen. HE; van Straalen. NM; van den Berg. M; van Wachem. E. (1986). Avoidance as a
mechanism of tolerance for lead in terrestrial arthropods. In HWH Veldthuis (Ed.), Proceedings of the
3rd European Congress of Entomology (pp. 251-254). Amsterdam, The Netherlands: Nederlandse
Entomologische Vereniging.
Vandenhove. H; Olyslaegers. G: Sanzharova. N; Shubina. O: Reed. E; Shang. Z; Velasco. H. (2009).
Proposal for new best estimates of the soil-to-plant transfer factor of U, Th, Ra, Pb and Po. J Environ
Radioact 100: 721-732. http://dx.doi.Org/10.1016/i.jenvrad.2008.10.014
Vedamanikam. VJ; Shazilli. NAM. (2008a). Comparative toxicity of nine metals to two Malaysian
aquatic dipterian larvae with reference to temperature variation. Bull Environ Contam Toxicol 80:
516-520. http://dx.doi.org/10.1007/s00128-008-9413-x
Vedamanikam. VJ: Shazilli. NAM. (2008b). The effect of multi-generational exposure to metals and
resultant change in median lethal toxicity tests values over subsequent generations. Bull Environ
Contam Toxicol 80: 63-67. http://dx.doi.org/10.1007/s00128-007-9317-l
Veltman. K; Huijbregts. MAJ; Hendriks. AJ. (2010). Integration of biotic ligand models (BLM) and
bioaccumulation kinetics into a mechanistic framework for metal uptake in aquatic organisms.
Environ Sci Technol 44: 5022-5028. http://dx.doi.org/10.1021/es903697c
Vermeulen. F; Van Den Brink. NW: D'Have. H; Mubiana. VK; Blust. R; Bervoets. L; De Coen. W.
(2009). Habitat type-based bioaccumulation and risk assessment of metal and As contamination in
earthworms, beetles and woodlice. Environ Pollut 157: 3098-3105.
http://dx.doi.0rg/10.1016/i.envpol.2009.05.017
Vigneshkumar. B; Pandian. SK; Balamurugan. K. (In Press) Catalase activity and innate immune
response of Caenorhabditis elegans against the heavy metal toxin lead. Environ Toxicol.
http://dx.doi.org/10.1002/tox.20722
Viiver. MG: Van gestel. CAM; Lanno. RP; Van straalen. NM; Peijnenburg. WJG. M. (2004). Internal
Metal Sequestration and Its Ecotoxicological Relevance: A Review. Environ Sci Technol 38:
4705-4712. http://dx.doi.org/10.1021/es040354g
Viiver. MG: van Gestel. CAM; van Straalen. NM; Lanno. RP; Peijnenburg. W. (2006). Biological
significance of metals partitioned to subcellular fractions within earthworms (Aporrectodea
caliginosa). Environ Toxicol Chem 25: 807-814. http://dx.doi.Org/10.1897/05-128R.l
Vink. JPM. (2009). The origin of speciation: Trace metal kinetics over natural water/sediment interfaces
and the consequences for bioaccumulation. Environ Pollut 157: 519-527.
http://dx.doi.0rg/10.1016/i.envpol.2008.09.037
Vitousek. PM; Aber. JD; Howarth. RW: Likens. GE; Matson. PA; Schindler. DW: Schlesinger. WH;
Tilman. DG. (1997). Human alteration of the global nitrogen cycle: Sources and consequences. Ecol
Appl 7: 737-750. http://dx.doi.org/10.1890/1051-0761(1997)007r0737:HAOTGN12.0.CO:2
Vlahogianni. TH; Valavanidis. A. (2007). Heavy-metal effects on lipid peroxidation and antioxidant
defence enzymes in mussels Mytilus galloprovincialis. Chem Ecol 23: 361-371.
http://dx.doi.org/10.1080/02757540701653285
Wall. DH. (1999). Biodiversity and ecosystem functioning. Bioscience 49: 107-108.
Wang. C: Gu. X; Wang. X; Quo. H; Geng. J: Yu. H; Sun. J. (2011 a). Stress response and potential
biomarkers in spinach (Spinacia oleracea L.) seedlings exposed to soil lead. Ecotoxicol Environ Saf
74: 41-47. http://dx.doi.Org/10.1016/i.ecoenv.2009.02.009
November 2012 7-285 Draft - Do Not Cite or Quote
-------
Wang. C: Tian. Y; Wang. X; Geng. J; Jiang. J; Yu. H; Wang. C. (2010a). Lead-contaminated soil induced
oxidative stress, defense response and its indicative biomarkers in roots of Vicia faba seedlings.
Ecotoxicology 19: 1130-1139. http://dx.doi.org/l0.1007/s 10646-010-0496-x
Wang. C: Yan. X; Wang. P; Hou. J: Hui. Z. (2010b). Effects of salinity and pH on the uptake and
distribution of Pb and Cd in the aquatic plant Artemisia selengensis in different seasons. Agrochimica
54: 65-78.
Wang. CR: Tian. Y; X-R. W: Yu. HX; Lu. XW: Wang. C: Wang. H. (2010c). Hormesis effects and
implicative application in assessment of lead-contaminated soils in roots of Vicia faba seedlings.
Chemosphere 80: 965-971. http://dx.doi.Org/10.1016/i.chemosphere.2010.05.049
Wang. CR: Wang. XR: Tian. Y: Xue. YG: Xu. XH: Sui. YX: Yu. HX. (2008a). Oxidative stress and
potential biomarkers in tomato seedlings subjected to soil lead contamination. Ecotoxicol Environ Saf
71: 685-691. http://dx.doi.Org/10.1016/i.ecoenv.2008.01.002
Wang. CR: Wang. XR: Tian. Y: Yu. HX: Gu. XY: Du. WC: Zhou. H. (2008b). Oxidative stress, defense
response, and early biomarkers for lead-contaminated soil in Vicia faba seedlings. Environ Toxicol
Chem 27: 970-977. http://dx.doi.Org/10.1897/07-344.l
Wang. DY; Peng. Y. (2007). Multi-biological defects caused by lead exposure exhibit transferable
properties from exposed parents to their progeny in Caenorhabditis elegans. J Environ Sci 19:
1367-1372. http://dx.doi.org/10.1016/S1001-0742(07)60223-X
Wang. DY; Xing. XJ. (2008). Assessment of locomotion behavioral defects induced by acute toxicity
from heavy metal exposure in nematode Caenorhabditis elegans. J Environ Sci 20: 1132-1137.
http://dx.doi.org/10.1016/S1001-0742(08)62160-9
Wang. G: Su. MY; Chen. YH; Lin. FF; Luo. D; Gao. SF. (2006b). Transfer characteristics of cadmium
and lead from soil to the edible parts of six vegetable species in southeastern China. Environ Pollut
144: 127-135. http://dx.doi.0rg/10.1016/i.envpol.2005.12.023
Wang. MZ: Jia. XY. (2009). Low levels of lead exposure induce oxidative damage and DNA damage in
the testes of the frog Rana nigromaculata. Ecotoxicology 18: 94-99.
http://dx.doi.org/10.1007/sl0646-008-0262-5
Wang. N; Ingersoll. CG: Ivev. CD; Hardestv. DK; May. TW: Augspurger. T; Roberts. AD; van Genderen.
E; Barnhart. MC. (2010e). Sensitivity of early life stages of freshwater mussels (Unionidae) to acute
and chronic toxicity of lead, cadmium, and zinc in water. Environ Toxicol Chem 29: 2053-2063.
http://dx.doi.org/10.1002/etc.250
Wang. Q; Liu. B; Yang. H; Wang. X; Lin. Z. (2009d). Toxicity of lead, cadmium and mercury on
embryogenesis, survival, growth and metamorphosis of Meretrix meretrix larvae. Ecotoxicology 18:
829-837. http://dx.doi.org/10.1007/sl0646-009-0326-l
Wang. WX; Rainbow. PS. (2008). Comparative approaches to understand metal bioaccumulation in
aquatic animals [Review]. Comp Biochem Physiol C Toxicol Pharmacol 148: 315-323.
http://dx.doi.0rg/10.1016/i.cbpc.2008.04.003
Wang. ZW: Nan. ZR; Wang. SL; Zhao. ZJ. (20 lie). Accumulation and distribution of cadmium and lead
in wheat (Triticum aestivum L.) grown in contaminated soils from the oasis, north-west China. J Sci
Food Agric 91: 377-384. http://dx.doi.org/10.1002/isfa.4196
Ward. DM; Nislow. KH; Chen. CY; Folt. CL. (2010). Reduced trace element concentrations in fast-
growing juvenile atlantic salmon in natural streams. Environ Sci Technol 44: 3245-3251.
http://dx.doi.org/10.1021/es902639a
Watanabe. K; Monaghan. MT; Takemon. Y; Omura. T. (2008). Biodilution of heavy metals in a stream
macroinvertebrate food web: Evidence from stable isotope analysis. Sci Total Environ 394: 57-67.
http://dx.doi.0rg/10.1016/i.scitotenv.2008.01.006
November 2012 7-286 Draft - Do Not Cite or Quote
-------
Watmough. SA. (1999). Monitoring historical changes in soil and atmospheric trace metal levels by
dendrochemical analysis. Environ Pollut 106: 391-403.
http://dx.doi.org/10.1016/S0269-7491(99)00102-5
Watmough. SA; Dillon. PJ. (2007). Lead biogeochemistry in a central Ontario forested watershed.
Biogeochemistry 84: 143-159. http://dx.doi.org/10.1007/sl0533-007-9110-6
Watson. AP; Van Hook. RI; Jackson. PR; Reichle. DE. (1976). Impact of a lead mining smelting complex
on the forest- floor litter arthropod fauna in the new lead belt region of southeast Missouri.
(ORNL/NSF/EATC-30). Oak Pudge, Tennessee: Oak Pudge National Laboratory.
Weber. DN. (1993). Exposure to sublethal levels of waterborne lead alters reproductive behavior patterns
in fathead minnows (Pimephales promelas). Neurotoxicology 14: 347-358.
Weber. DN. (1996). Lead-induced metabolic imbalances and feeding alterations in juvenile fathead
minnows (Pimephales promelas). Environ Toxicol 11: 45-51.
Weir. PA; Hine. CH. (1970). Effects of various metals on behavior of conditioned goldfish. Arch Environ
Health 20: 45-51.
Weis. JS: Weis. P. (1998). Effects of exposure to lead on behavior of mummichog (Fundulus heteroclitus
L) larvae. J Exp Zool Part A Ecol Genet Physiol 222: 1-10.
Westman. WE. (1977). How much are nature's services worth? Measuring the social benefits of
ecosystem functioning is both controversial and illuminating. Science 197: 960-964.
http://dx.doi.org/10.1126/science.197.4307.960
Wetmore. A. (1919). Lead poisoning in waterfowl. US Dep Agric Bull 793: 1-12.
Witte. KM; Wantv. RB; Ridley. WI. (2004). Engelmann Spruce (Picea engelmannii) as a biological
monitor of changes in soil metal loading related to past mining activity. Appl Geochem 19:
1367-1376. http://dx.doi.0rg/10.1016/i.apgeochem.2004.01.022
Wong. CC: Wu. SC: Kuek. C: Khan. AG: Wong. MH. (2007). The role of mycorrhizae associated with
vetiver grown in Pb-/Zn-contaminated soils: Greenhouse study. Restor Ecol 15: 60-67.
http://dx.doi.Org/10.llll/i.1526-100X.2006.00190.x
Wong. LC: Kwok. KWH; Leung. KMY; Wong. CK. (2009). Relative sensitivity distribution of freshwater
planktonic crustaceans to trace metals. Hum Ecol Risk Assess 15: 1335-1345.
http://dx.doi.org/10.1080/10807030903307115
Worms. I; Simon. DF; Hassler. CS: Wilkinson. KJ. (2006). Bioavailability of trace metals to aquatic
microorganisms: Importance of chemical, biological and physical processes on biouptake [Review].
Biochimie 88: 1721-1731. http://dx.doi.Org/10.1016/i.biochi.2006.09.008
WRI (World Resources Institute). (2000). World resources 2000-2001: People and ecosystems: The
fraying web of life. Washington, D.C.
Wu. X; Hong. F: Liu. C: Su. MY; Zheng. L; Gao. FQ; Yang. F. (2008c). Effects of Pb2+ on energy
distribution and photochemical activity of spinach chloroplast. Spectrochim Acta A Mol Biomol
Spectrosc 69: 738-742. http://dx.doi.Org/10.1016/i.saa.2007.05.047
Xie. W: Zhou. J; Wang. H; Liu. Q; Xia. J: Lv. X. (2011). Cu and Pb accumulation in maize (Zea mays L.)
and soybean (Glycine max L.) as affected by N, P and K application. African Journal of Agricultural
Research 6: 1469-1476.
Xing. X; Du. M; Zhang. Y; Wang. D. (2009a). Adverse effects of metal exposure on chemotaxis towards
water-soluble attractants regulated mainly by ASE sensory neuron in nematode Caenorhabditis
elegans. J Environ Sci 21: 1684-1694. http://dx.doi.org/10.1016/sl001-0742(08)62474-2
Xing. X; Guo. Y; Wang. D. (2009b). Using the larvae nematode Caenorhabditis elegans to evaluate
neurobehavioral toxicity to metallic salts. Ecotoxicol Environ Saf 72: 1819-1823.
http://dx.doi.0rg/10.1016/i.ecoenv.2009.06.006
November 2012 7-287 Draft - Do Not Cite or Quote
-------
Xing. X; Rui. Q; Du. M; Wang. D. (2009c). Exposure to lead and mercury in young larvae induces more
severe deficits in neuronal survival and synaptic function than in adult nematodes. Arch Environ
Contam Toxicol 56: 732-741. http://dx.doi.org/10.1007/s00244-009-9307-x
Xiong. ZT; Zhao. F; Li. MJ. (2006). Lead toxicity in Brassica pekinensis Rupr.: Effect on nitrate
assimilation and growth. Environ Toxicol 21: 147-153. http://dx.doi.org/10.1002/tox.20167
Xu. J; Ke. X; Krogh. PH; Wang. Y; Luo. YM; Song. J. (2009a). Evaluation of growth and reproduction as
indicators of soil metal toxicity to the Collembolan, Sinella curviseta. Insect Sci 16: 57-63.
http://dx.doi.0rg/10.llll/i.1744-7917.2009.00254.x
Xu. J; Wang. Y; Luo. YM; Song. J: Ke. X. (2009b). Effects of copper, lead and zinc in soil on egg
development and hatching of Folsomia Candida. Insect Sci 16: 51-55.
http://dx.doi.0rg/10.llll/i.1744-7917.2009.00253.x
Yang. H; Shen. Z; Zhu. S: Wang. W. (2008a). Heavy metals in wetland plants and soil of Lake Taihu,
China. Environ Toxicol Chem 27: 38-42. http://dx.doi.Org/10.1897/07-089.l
Yang. R; Tang. J: Chen. X; Hu. S. (2007). Effects of coexisting plant species on soil microbes and soil
enzymes in metal lead contaminated soils. Appl Soil Ecol 37: 240-246.
http://dx.doi.0rg/10.1016/i.apsoil.2007.07.004
Yang. R; Yu. G: Tang. J: Chen. X. (2008b). Effects of metal lead on growth and mycorrhizae of an
invasive plant species (Solidago canadensis L.). J Environ Sci 20: 739-744.
http://dx.doi.org/10.1016/S1001-0742(08)62121-X
Yang. Y; Campbell. CD; Clark. L; Cameron. CM; Paterson. E. (2006). Microbial indicators of heavy
metal contamination in urban and rural soils. Chemosphere 63: 1942-1952.
http://dx.doi.0rg/10.1016/i.chemosphere.2005.10.009
Yu. DY; Xu. ZR; Yang. XG. (2005). Effects of lead and particulate montmorillonite on growth
performance, hormone and organ weight in pigs. Asian - Australasian Journal of Animal Sciences 18:
1775-1779.
Zaprjanova. P; Dospatliev. L; Angelova. V: Ivanov. K. (2010). Correlation between soil characteristics
and lead and cadmium content in the aboveground biomass of Virginia tobacco. Environ Monit Assess
163: 253-261. http://dx.doi.org/10.1007/sl0661-009-0831-v
Zeng. LS: Liao. M; Chen. CL; Huang. CY. (2007). Effects of lead contamination on soil enzymatic
activities, microbial biomass, and rice physiological indices in soil-lead-rice (Oryza sativa L.) system.
Ecotoxicol Environ Saf 67: 67-74. http://dx.doi.Org/10.1016/i.ecoenv.2006.05.001
Zhang. CB; Huang. LN; Luan. TG: Jin. J: Lan. CY. (2006). Structure and function of microbial
communities during the early stages of revegetation of barren soils in the vicinity of a Pb/Zn smelter.
Geoderma 136: 555-565. http://dx.doi.Org/10.1016/i.geoderma.2006.04.011
Zhang. H; Luo. Y; Song. J: Zhang. H; Xia. J: Zhao. Q. (20lib). Predicting As, Cd and Pb uptake by rice
and vegetables using field data from China. J Environ Sci 23: 70-78.
http://dx.doi.org/10.1016/sl001-0742(10)60375-0
Zhang. Y; Song. JM; Yuan. HM; Xu. YY; He. ZP; Duan. LQ. (201 Ob). Biomarker responses in the
bivalve (Chlamys farreri) to exposure of the environmentally relevant concentrations of lead, mercury,
copper. Environ Toxicol Pharmacol 30: 19-25. http://dx.doi.Org/10.1016/i.etap.2010.03.008
Zhang. YM; Huang. DJ; Zhao. DQ: Long. J: Song. G: Li. A. (2007b). Long-term toxicity effects of
cadmium and lead on Bufo raddei tadpoles. Bull Environ Contam Toxicol 79: 178-183.
http://dx.doi.org/10.1007/s00128-007-9152-4
Zhang. Z; Song. X; Wang. Q: Lu. X. (2012). Cd and Pb contents in soil, plants, and grasshoppers along a
pollution gradient in Huludao City, northeast China. Biol Trace Elem Res 145: 403-410.
http://dx.doi.org/10.1007/sl2011-011-9199-2
November 2012 7-288 Draft - Do Not Cite or Quote
-------
Zheng. RQ; Li. CY. (2009). Effect of lead on survival, locomotion and sperm morphology of Asian
earthworm, Pheretima guillelmi. J Environ Sci 21: 691-695.
http://dx.doi.org/10.1016/sl001-0742(08)62325-6
Zhuang. P; Zou. HL; Shu. WS. (2009). Biotransfer of heavy metals along a soil-plant-insect-chicken food
chain: Field study. J Environ Sci 21: 849-853. http://dx.doi.org/10.1016/sl001-0742(08)62351-7
November 2012 7-289 Draft - Do Not Cite or Quote
-------