r ^
905R84117
FINAL *">?> A
ORGANOCHLORINE CONTAMINANTS OF WINTERING DUCKS
FORAGING ON DETROIT RIVER SEDIMENTS
Running Title: ORGANOCHLORINES IN DETROIT RIVER DUCKS
V. Elliott Smith and John M. Spurr
Cranbrook Institute of Science
P.O. Box 801
Bloomfield Hills, Michigan 48013
John C. Filkins
U.S. Environmental Protection Agency
Large Lakes Research Station
9311 Groh Road
Grosse He, Michigan 48138
Jody J. Jones
University of Maine
Orono, Maine 04469
October 1, 1984
Smith -
-------
INTRODUCTION
The Detroit River, connecting Lakes St. Clair and Erie, is one of the
world's busiest waterways, which passes through a highly industrialized area of
the United States and Canada (Fig. 1). Among the principal contaminants found
in Detroit River sediments are organochlorines (Fallon and Horvath, 1983),
including polychlorinated biphenyls (PCBs). Due to their hydrophobic nature
and stability, PCBs tend to collect and persist in organic sediments (Choi and
Chen, 1976; Haque e£al_., 1974; Crump-Wiesner ejt al_., 1973). To the degree
that PCBs and other contaminants accumulate in sediments and associated
benthos, they become available at higher concentrations to larger bottom-
foraging fauna. Thus polluted sediments in many nearshore areas may act as
reservoirs of "in-place" contaminants, which continue to enter the food chain
long after point sources of pollution are controlled.
Each winter, thousands of migratory waterfowl congregate on the Detroit
River, attracted by areas of shallow, ice-free water. These include diving
ducks, such as scaups and goldeneyes, which forage on plant and animal matter
in the sediments. During 1980-82, Drobney et al_. (1982) studied the food
habits and nutritional status of 169 lesser scaups (Aythya affinis), greater
scaups (_A. marila) and goldeneyes (Bucephala clangula) wintering on the lower
Detroit River. This was part of a study designed to evaluate the effects of
winter navigation on survival of waterfowl. The collections presented us with
an opportunity to study the possible uptake and accumulation by ducks of
selected organochlorines, including PCB congeners, from polluted sediment. In
part, our objective was to identify and compare complex PCB mixtures in these
ducks and in their benthic food supply at a foraging site. We thought that
Detroit River sediment might be an important source of PCBs, even for transient
Smith - 3
-------
waterfowl. If so, then distinctive patterns of certain congeners in sediment
might be found also in local benthos, ducks and other bottom feeders. Another
related goal was to examine the relationship between lipid and PCB
concentrations in the ducks during this period of increased fat metabolism.
Weekly counts of waterfowl along the lower Detroit River near its junction
with Lake Erie (Fig. 1) showed concentrations of up to 24,000 ducks and geese
in late January, 1980, which declined steadily to approximately 5,000 birds in
late March, just prior to the final northward migration (Drobney et^ j»l_., 1982).
It is likely that local populations in March included transient ducks from
wintering grounds farther south.
Of the 13 ducks analyzed in this study, 11 were collected at Mud Island;
the remaining two were collected 8 km southward on the river (Fig. 1). The
study site was in shallow (1-3 m) waters on the downstream side of Mud Island,
where aquatic macrophytes grow abundantly in the soft bottom during the summer.
Sediments there are finely textured with a high organic content. Immediately
after ducks were collected at this site, their gut contents were removed and
preserved in formalin for later identification (Drobney e_t al_., 1982). Their
diets were found to include a wide variety of invertebrate and plant materials.
However, the bulk of their food consisted of oligochaete worms (including
Tubifex sp.) and tubers of Vallisneria sp. and Potamogeton sp., two species of
aquatic flowering plants. The percentages of plant and animal matter found in
gut contents are summarized in Table 1 (from Drobney elt al_., 1982). Their
results show that pollution-tolerant oligochaetes may be an important winter
food for diving ducks in the Detroit River, as they were for oldsquaw ducks
(Clangula hyemalis) in Milwaukee Harbor (Rofritz, 1977). Oligochaetes were
Smith - 4
-------
reported to have a higher caloric content than other benthos studied by
Cummins and Wuycheck (1971).
Since Mud Island was a prominent feeding area for ducks, we also sampled
other components of this system: unfiltered water, filterable seston,
surficial sediment, oligochaetes and adult carp (Cyprinus carpio), the latter
representing a bottom-feeding fish. Sediment samples collected represented
only recently deposited material likely to be available to foraging ducks
and fish. At Mud Island this was defined as the upper 4-cm zone of a
sediment core in which 65* of the total PCB mass occurred at nearly uniform
concentrations. Within the next 5-9 cm interval of sediment depth, the PCB
concentration rapidly declined. Biological mixing of surficial sediments
and associated contaminants has been documented elsewhere in the Great Lakes
(Robbins, 1982) and in laboratory microcosms (Karickhoff and Morris, 1985).
Studies have shown that the composition of PCB mixtures in environmental
samples is highly variable due to differences in the chemical and biological
behavior of congeners (Safe, 1980; Hutzinger e_t al_., 1974). Certain PCBs are
more persistent (i.e., conservative) than others, as indicated by the
similarity of these congener ratios in the environment to those in commercial
PCB mixtures (Ballschmiter and Zell, 1980; Ballschmiter ejb al_., 1978). We
evaluated all of the possible ratios of congeners in Mud Island sediment and
fauna, and selected 6 congeners that were involved most frequently in the
most consistent ratios. The distribution of these "conservative" congeners
was compared to that of PCBs as a whole.
Smith - 5
-------
METHODS
A detailed description of the field and laboratory methods is included in
a more comprehensive report on the Detroit River studies (Smith et_ al_., in
preparation). The following is a brief summary of methods.
Each water sample (12 L), collected in four, 3.8 L brown glass jugs, was
extracted by partitioning twice with dichloromethane in a 15:1 ratio. Combined
extracts were concentrated, transferred into n-hexane, dried through sodium
sulfate columns and evaporated under nitrogen to volumes of 2-10 ml, as
adopted from standard EPA procedures (Thompson, 1974). Excess lipids were
removed by treating 2 ml extracts with concentrated sulfuric acid, and
subsequently freezing the aqueous layer in an acetone-dry ice bath (Murphy,
1972). Cleaned extracts in hexane were stored temporarily in sealed glass
ampules (8°C, dark).
Seston was collected by pressure filtration (<35 kpa) through glass fiber
filters (nominally, 0.6 ym porosity). Filters, stored temporarily in glass
jars with acetone, were exhaustively extracted by Soxhlet with n-hexane-acetone
(1:1), transferred into n-hexane and processed as above.
Oligochaetes were hand-picked from grab samples (Ponar dredge) of sediment,
after the larger particles were collected by washing the mud through a 0.5 mm
mesh bronze wire sieve (U.S. 130). Frozen oligochaetes (5 g) were mixed with
anhydrous sodium sulfate (1:3) and exhaustively extracted by Soxhlet with
n-hexane:dichloromethane (1:1) for 6 hours. Extracts were processed as for
water and seston samples.
Sediment cores, approximately 15 cm long, were collected by pushing a pole-
mounted aluminum tube (5 cm diameter) into the bottom. Three cores were
sectioned at 1 on intervals and equivalent sections were combined. Sediments
Smith - 6
-------
were exhaustively extracted by Soxhlet with n-hexane-acetone (1:1) and
processed as above. The final extracts were treated with reactive copper to
remove sulfur (Environment Canada, 1979). Based on preliminary analysis, the
upper four 1-cm sections were shown to have similar concentrations of total
PCB. Extracts of these sections were combined as representing the mixed zone
of sediment.
Extracts of monthly water and seston samples collected during July-November
1982 were combined to form one sample of each. Eleven oligochaete samples from
the same period were also combined. Three sediment samples, collected on
December 14, 1982, were combined.
Carp were gill-netted on September 17, 1982, and stored frozen, prior to
grinding and homogenizing the whole fish. Ducks collected by shotgun during
January-March 1980, were eviscerated, defeathered and the head and feet were
removed prior to freezing. Homogenized aliquots (20 g) of duck or fish tissue
were mixed with sodium sulfate (3:1), and exhaustively extracted with
n-hexane-dichloromethane (1:1). Extracts were processed as above. Total
lipids in carp and duck tissue were determined gravimetrically after drying at
room temperature a 1 ml aliquot of the 10 ml final extract. All solvents used
were of "pesticide grade" from Burdick and Jackson, Muskegon, Michigan. All
glassware was cleaned rigorously with detergents and solvent rinses, followed
by baking at 580°C. Glassware assemblies were refluxed or rinsed again with
solvents before use.
For organochlorine analysis we used a Varian 3700 capillary column gas
chromatograph fitted with dual Ni electron capture detectors, auto-
samplers and 50 m X 0.2 mm (i.d.) SE-54 thin film, fused silica columns.
Chroma to graphic conditions were as described elsewhere (Smith e£ al_., 1984, in
Smith - 7
-------
preparation). Component peaks were baseline-corrected prior to quantitation.
Individual PCB congeners were analyzed with reference to a standard mixture of
Aroclors 1016, 1254 and 1260, in which 95 congeners had been identified and 72
of these quantitated using synthetic PCB standards (Mullin et.al_., 1983;
Mullin et al_., 1984). Detach!oronaphthalene was used as an internal standard
to verify retention times. Homolog and total PCB values were based on the
appropriate summations of congener values. Other organochlorines were
similarly quantitated.
Glassware and solvent blank values for Mud Island water samples averaged
5.7% and 4.5% of sample values for PCBs and other organochlorines,
respectively. Corresponding blank values for PCBs in sediment, oligochaetes
and ducks averaged less than 3% of those in the samples. Analytical precision
calculated for total PCBs in 9 replicate duck samples was 6.48% RSD. For 11
replicate analyses of an Aroclor 1254 standard, the precision was 10.8% RSD.
The 72 congener values that were summed for total PCBs represented 84% of the
true mass, based on later analysis of the same Aroclor 1254 standard. The
Aroclor contained 15 more congeners that were not accounted for by standards
at the time of analysis.
RESULTS AND DISCUSSION
Characteristics of the 13 duck samples analyzed are given in Table 2
according to Drobney (1983). The content of water, fat and lean dry mass in
each case is given as a percentage of the dressed weight (head, feet, feathers
and viscera removed). Most of the variation in dressed weight was due to
differences in fat and water content; the percentage of lean dry mass, largely
protein, was relatively constant at 21.5 - 28.2% of dressed weight. We also
measured lipid (fat) dry weight for an aliquot of the crude extract used for
Smith - 8
-------
organochlorine analysis. This value was used here to express the concentration
of contaminants per unit lipid. Comparison of these two sets of lipid
measurements per unit of tissue showed agreement within 9t of the lower value,
despite the difference in extraction solvents used (Methods and Table 2).
Table 3 summarizes the concentrations of various organochlorines, including
total PCBs, in ducks and other compartments of the Mud Island system. The
water, seston, sediment and oligochaete values represent composites of samples
collected over a 5-month period. For carp (n=9) and ducks (n=13) the mean and
standard deviation are shown, along with the range and percent occurrence of
each contaminant. The total PCB values represent a summation of congener
concentrations: 72 different congeners were quantitated in carp, and 68 in
ducks. The variability of total PCB concentrations in carp and duck species,
expressed as %RSD, was: carp, 48.5; lesser scaup, 47.2; greater scaup, 17.3;
goldeneye, 23.6.
Mean concentrations of total PCB adjusted for lipid (PCB/% lipid) in ducks
were: lesser scaup, 79 mg/kg; greater scaup, 89 mgAg; goldeneye, 38 mgAg.
Although total PCB values were higher for both scaup species relative to
goldeneye, the survey of gut contents from 169 ducks (Table 1) indicates that a
higher proportion of animal matter, mainly oligochaetes, was consumed by
goldeneyes. However, the gut contents were not available for organochlorine
analysis. The seston PCB concentration in the water column (5.2 mg/kg)
suggests a greater input of PCBs to the bottom than was indicated by the
concentration in sediment (0.63 mgAg).
At lower concentrations alpha- and gamma-BHC (hexachlorocyclohexane) had a
distribution similar to that of PCB in the carp and ducks (Table 3). Hexa-
chlorobenzene concentrations were relatively high in ducks, and particularly in
Smith - 9
-------
goldeneyes (mean, 1.7 mg/kg). C1s- and trans-chlordanes were found in scaup
ducks (means, 0.0025 - 0.0097 mg/kg), but not in goldeneyes. Concentrations of
cis- and trans-nonachlor (means, 0.013 - 0.33 mgAg) were measured 1n most
ducks of all three species. Of the DDT group, 4,4'-DDE levels were highest
(means, 0.48 - 1.3 mgAg}, especially in greater scaups. Other organochlorines
that occurred less consistently were beta- and delta-BHC, heptachlor epoxide,
and alpha- and gamma-chlordane. All standards of organochlorines reported
here were shown to be stable under the acid conditions used to clean extracts
(Murphy, 1972)
High resolution gas chromatography/mass spectroscopy analysis, applied to
extracts of a female greater scaup (no. 50) and female goldeneye (no. 1) by
Kuehl (EPA/ERL-Duluth, 1983, personal communication), confirmed the presence
of trichlorobiphenyls through nonachlorobiphenyls, cis- and trans-chlordane,
cis- and trans-nonachlor, 4,4'-DDE and hexachlorobenzene, other chlorinated
benzenes, and octachlorostyrene. Low levels of chlorinated cyclohexanes
(alpha-, beta-, gamma-, and delta-BHC) and heptachlor epoxide, reported here,
were not confirmed by mass spectroscopy.
Concentrations of the individual congeners comprising PCB mixtures in
sediment, oligochaetes, carp and ducks from Mud Island are shown in Table 4.
Included are the homolog values representing groups of congeners with two to
nine chlorines per molecule. Congeners in Table 4 are identified by their
structures and by peak numbers (Ballschmiter and Zell, 1980). The percent
composition by congener of PCBs in all 13 ducks is summarized in Fig. 2. Only
18 of the 72 congeners reported accounted for 90% of the total PCB mass in
scaups and goldeneyes combined.
Smith - 10
-------
The advantages of congener-specific analysis of PCBs have been widely
recognized (Mullin et al_., 1984; Mullin, et_ aJL, 1983; Duinker et al_., 1980;
Ballschmiter and Zell, 1980). One important benefit is that concentrations of
potentially toxic congeners can be measured individually. Toxicities measured
with respect to the induction of aryl hydrocarbon hydroxylase (AHH) enzymes
and other effects have been reported (Safe £t al_., 1982), and structural
characteristics of these congeners are known (Ballschmiter e£ al_., 1978).
Among the potentially more toxic PCB congeners which occur as major
components in ducks, oligochaetes, carp and sediments from Mud Island are 9
tri-, tetra-, penta-, hexa- and heptachlorobiphenyls (Table 4, asterisks).
These account for averages of 28.3% of the total PCB mass in lesser scaups,
31.0% in great scaups and 36.8% in goldeneyes. In each species, over 84% of
this fraction is represented by only three congeners: 2,3',4,4',5-
pentachlorobiphenyl, 2,2',3,4,4',5'-hexachlorbiphenyl and 2,2',3,3',4,4',5-
heptachlorobiphenyl. These congeners, respectively, caused embryotoxicity in
chickens (Ax and Hansen, 1975), accumulation of rat liver porphyrins (Stonard
and Grieg, 1976) and AHH enzyme induction (Parkinson and Safe, 1981).
Previous work (Haseltine and Prouty, 1980; Custer and Heinz, 1980; Heath
et al_., 1972) has indicated that ducks may be relatively insensitive to
environmental levels of PCBs. However, in any study which relates exposure to
effects it is relevant to know the bioaccumulated levels of specific congeners
in addition to that of total PCBs. The more toxic congeners among hexa- and
heptachlorobiphenyls tend to persist and become enriched in fauna (Safe et al.,
1982; Ballschmiter et al_., 1978), as indicated in Figure 2. At a given level
of total PCBs, concentrations of the more toxic components may vary widely
among different organisms.
Smith - 11
-------
The proportions of congeners were notably different for PCB mixtures in
sediment and various fauna from Mud Island. These differences were most
apparent when the congener concentrations in fauna were grouped by homolog and
expressed as ratios to the corresponding values for sediment. The resulting
distribution coefficients (K .) were plotted for oligochaete/sediment, carp/-
sediment and ducks/sediment (Fig. 3). The K. values for oligochaete/sediment
range near unity, indicating that in oligochaetes there was little, if any,
biomagnification of sediment PCBs as a whole. However, the plot for
oligochaetes shows relatively lower concentrations of tri- through pentachloro-
biphenyls and slightly higher levels of hepta- and octachlorobiphenyls than in
sediment. In contrast, the carp and duck Krf plots (mean values) indicate
much greater elevations of PCB levels over those in sediment. The K. values
for carp homologs ranged from 11 for trichlorobiphenyls to 47 for
octachlorobiphenyls. In ducks the range was much greater, from 2.7 from
dichlorobiphenyls to 43 for octachlorobiphenyls. Apparently in ducks there was
more selective bioaccumulation and/or retention of the heavier, more
chlorinated congeners than in carp or oligochaetes.
Although the source of PCBs in oligochaetes was clearly the Mud Island
sediment they inhabited, it is less certain where carp and ducks obtained the
bulk of their PCB content. Carp may experience varying exposures to PCBs
through feeding within different areas of the Detroit River system. However,
much greater variability of exposures is likely for ducks due to their seasonal
migrations and the greater diversity of food items available to them. Further
complicating the effects of varying exposure on PCB composition are the
biological differences in uptake and accumulation that may occur in different
species.
Smith - 12
-------
In order to evaluate sediment as a direct source of PCBs in fauna, we
sought to identify congeners that appear to be least affected by selective
accumulation processes in different organisms. We expected that the ratios of
such congeners would provide a basis for comparing PCB mixtures that was
independent of concentration differences. As a first step we compared the
variability of all PCB ratios in Mud Island samples. Ballschmiter et al.
(1978) used a similar approach to identify conservative ratios of
"recalcitrant" congeners for purposes of estimating past loadings of PCB
Aroclors 1254 and 1260 to the environment. Similarly, we found that six PCB
congeners (Fig. 2), occurring in all Mud Island samples, were most often
represented in the most consistent ratios, as indicated by their low root mean
square error. Three of the six congeners (peak nos. 170, 180 and 183) were
najor constituents of the samples (Fig. 2). Among 13 ducks studied, the six
congeners comprised 22.1 - 34.5% of the total PCB mass. Their structures are
shown in Table 4. One compound, 2,2l,3,3',4,4',5-heptachlorobiphenyl (peak no.
170), which constitutes 9.5% (mean value) of the PCB mass in 13 ducks, is known
to induce hepatic microsomal monoxygenase enzymes of both the PB- and MC-type
(Parkinson et^al_., 1981; Parkinson and Safe, 1981). The capacity of compounds
to induce the MC-type or aryl hydrocarbon hydroxylase (AHH) enzymes is thought
to be highly correlated with their toxicity (Poland et^ aJL, 1979; Safe ejt al.,
1982). Another of the six congeners, 2,2',3,4,4',5,5'-heptachlorobiphenyl
(peak no. 180), which comprises 12.0% of the duck PCBs, is also likely to be a
AHH inducer based on its structural characteristics (Safe et al_., 1982). Two
additional congeners, 2,2',3,4,4',5-hexachlorobiphenyl (peak no. 138) and
2,3,3',4,4',5-hexachlorobiphenyl (peak no. 156), which are known to be AHH
inducers (Safe £t al_., 1982), also form consistent ratios with the six
Smith - 13
-------
compounds above. The combined toxicity and persistence of these congeners
increases their potential for adverse effects on ducks and other fauna that
bioaccumulate them selectively. Most of the same congeners were prominent in
human subjects exposed to PCBs in the Yusho incident (Bandiera e_t al_., 1984).
We used the six congeners to form three non-redundant ratios identified
here by their peak numbers as 180/170, 183/172 and 194/195. The magnitude and
variability of the ratios in sediment, oligochaetes, carp and ducks were
compared as an indication of similarity within and between sample types. The
results of a multivariate analysis of variance indicated no significant
differences (a = .05) between the 3 ratios in sediment, oligochaetes and 9
carp. However, carp ratios were significantly different (a = .05) from those
in the 13 ducks. Within the 3 duck species, the ratio differences were not
significant (a = .05).
While these results are very limited, they suggest the following
interpretation. We infer that the PCB compositions of both oligochaetes and
carp were strongly influenced by exposure to Mud Island sediment since their
ratios were very similar. The different ratios which occurred in ducks suggest
that their PCBs were derived from other sources as well. Also, the ducks were
collected two years prior to the other samples. However, the striking
consistency of these ratios among the 3 species of migratory ducks suggests a
different explanation: that their PCBs were largely from a common source
(possibly Mud Island sediment), but were proportioned differently as a group by
more selective uptake and/or retention of congeners. Their distinctive pattern
may be seen in the plots of homolog distribution coefficients described earlier
(Fig. 3). The duck PCBs were consistently more enriched in hexa-, hepta- and
Smith - 14
-------
octachlorobiphenyls than were PCBs 1n other system compartments, including
water and seston.
In order to investigate the possible influence of accelerated fat
metabolism on the PCB content of wintering ducks, we examined the relationship
between PCB concentrations in lipid and the percentage of lipid in 13 duck
carcasses during the February-March period. Tissue composition data collected
for all 169 Detroit River scaups and goldeneyes (Drobney, 1983) indicated that
the ratio of fat/lean dry mass declined during the January-March period (Fig.
4). Most of the decline occurred in January, and reached a plateau in mid-
February for the population as a whole. A smaller increase in percent fat than
occurred during late February through early March. The latter trend may have
represented an increase in the number of transient birds with higher fat
content, or improved feeding conditions. Also, it may be explained by a
relative decrease in lean dry mass, as occurs in other wintering duck species
(Reinecke jst al_., 1982; Peterson and Ellarson, 1979). However, this tendency
toward increasing fat/lean mass was not apparent in the subset of 13 ducks that
we analyzed (Fig. 4).
We used a linear regression analysis to relate the concentration of PCBs in
lipid to the percentage of lipid in 13 carcasses (lipid/wet weight). This
analysis was applied to both total PCB mass and the fraction of 6 congeners
that were selected as most conservative. The results (Fig. 5) showed that
concentrations of both were inversely correlated to percent lipid (r = 0.76
and 0.71, respectively). Moreover, the 6-congener fraction (PCB,) as a
percentage of total PCBs (PCBt) appeared to increase as the percent lipid
declined. Based on the regression lines, PCB^/PCBt would be predicted to
increase slightly from 0.25 at 25% lipid to 0.31 at 5% lipid. In other words,
Smith - 15
-------
PCBs as a whole behaved somewhat less conservatively than the 6-congener
fraction during this period.
Stickel (1973) noted that, despite cyclic increases in lipid metabolism,
losses of fat-soluble contaminants in birds are usually slow. White et al.
(1981) attempted a similar regression analysis of DDE and dieldrin with percent
lipid in wintering blue-winged teal, but the results were not significant due
to the low concentrations of pesticides present.
CONCLUSIONS
Wintering scaup and goldeneye ducks which foraged on contaminated sediments
near Mud Island in the lower Detroit River, were exposed to organochlorines,
including sediment PCB concentrations on the order of 0.63 mgAg. Similar
concentrations were found in benthic oligochaetes, confirmed two years earlier
as a major component of duck diets at this site. Body burdens of PCBs in the
ducks, ranging from 2.7 to 20 mg/kg in the carcass and from 38 to 89 mg/kg in
the lipid fraction, were high relative to 1979 means of <1 mgAg for wing pools
of mallards and black ducks from the Atlantic flyway (Cain, 1981). They also
exceeded most 1979-1980 levels of PCB in 17 species of dabbling and diving
ducks, including greater scaups and goldeneyes, from New York state (Kim et^
al., 1984). Levels in Detroit River ducks were more comparable to those in
greater scaup and goldeneyes wintering on the Baltic Sea (Falandysz and Szefer,
1982). Adult carp from Mud Island contained slightly higher PCB levels of 7.6
to 31. mgAg. Total PCB levels in all ducks and carp from Mud Island exceeded
the 2 mgAg guideline for edible fish established by the U.S. Food and Drug
Administration (1984).
The composition of PCB mixtures in Mud Island ducks, and that in sediment,
oligochaetes and carp, followed two distinct patterns. While all of the fauna
Smith - 16
-------
accumulated larger fractions of the more chlorinated PCBs than did sediment,
the highest proportion of hepta- and octachlorobiphenyls was in ducks. This
fraction included 6 congeners which formed consistent ratios in the sediment
and fauna. Multivariate analysis of variance indicated a significant
difference (a = .05) between 3 ratios formed from the 6 congeners in carp and
in ducks. There was no significant difference among the 3 duck species.
The greater enrichment of heavier PCB congeners in ducks may have resulted
from a more selective process of bioaccumulation and/or from dietary exposure
to different PCB sources elsewhere. On the other hand, the similarity of
homolog ratios in all 3 duck species suggests exposure to one dominant source
of PCBs, possibly Detroit River sediments.
For the 13 ducks, lower fat content was correlated (a = .05) with higher
PCB concentrations over a two month period. Since the proportion of 6
conservative congeners in total PCBs increased by 6% as predicted fat levels
decreased from 25% to 5%, fat mobilization seemed to have some effect on PCB
composition. To the extent that differential loss of less persistent PCBs
occurred during fat loss, an increase in the ratio of the 6 congeners to total
PCBs was expected.
Some of the principal organochlorines observed in sediment, oligochaetes,
carp and ducks from Mud Island were compounds of documented toxicity. These
included pesticides as well as 9 PCB congeners that comprised roughly a third
of the total PCB mass in ducks. Among the latter were congeners which induce
aryl hydrocarbon hydroxylase (AHH) enzymes in the manner of other known toxins
(Safe, 1980). Any toxicity of these to scaup and goldeneye ducks remains
unknown. Although we are not aware of any adverse conditions among Detroit
River ducks which night be attributed to contaminant exposures, such effects
Smith - 17
-------
would seem more likely to occur when ducks are stressed by cold weather and
starvation. It is important to recognize that the contaminants reported here
in ducks are likely to represent only part of their total exposure to
potentially toxic substances, both organic and inorganic, which occur in
Detroit River sediments. Due to biological mixing of recent sediments these
substances continue to be available to bottom-feeding ducks and other fauna.
As game species, scaups and goldeneyes contaminated by Detroit River sediments
may also present a hazard to human populations throughout their extensive
flyway.
Smith - 18
-------
ACKNOWLEDGEMENTS
We owe special thanks to R.D. Drobney for use of his unpublished data in
Table 2 and Figure 4 of this paper. We also thank all other contributors to
this work, especially J. Rood, S. Rood, J. Rathbun, S. Mathews and M.
Kasprzyk of Cranbrook Institute of Science, D. Do!an and M. Mull in of the
U.S. Environmental Protection Agency, and K. McGunagle, D. Griesmer, R. Brown
and D. Caudill of Computer Sciences Corporation. The project was supported
by EPA Grant ICR810232-01 to Cranbrook Institute of Science.
Smith - 19
-------
REFERENCES
Ax, R.L. and Hansen, L.G. 1975. Effects of purified PCB analogs on chicken
reproduction. Poultry Sci., 54:895-900.
Ballschmiter, K. and Zell, M. 1980. Analysis of polychlorinated biphenyls
(PCBs) by glass capillary gas chromatography. Composition of technical
Aroclor- and Clophen-PCB mixtures. Fres. Z. Anal. Chem., 302:20-31.
Ballschmiter, K., Zell, M., and Neu, H.J. 1978. Persistence of PCBs in the
ecosphere: Will some PCB-components "never" degrade? Chemosphere, 7:
173-176.
Bandiera, S., Farrell, K., Mason, G., Kelley, M., Romkes, M., Bannister, R.,
and Safe, S. 1984. Comparative toxicities of the polychlorinated
dibenzofuran (PCDF) and biphenyl (PCB) mixtures which persist in Yusho
victims. Chemosphere, 13:507-512.
Cain, B.W. 1981. Nationwide residues of organochlorine compounds in wings of
adult mallard and black ducks, 1979-1980. Pest. Monit. J., 15:128-134.
Choi, W.W. and Chen, K.Y. 1976. Associations of chlorinated hydrocarbons with
fine particulates and humic substances in nearshore surficial sediments.
Environ. Sci. Techno!., 10:782-786.
Crump-Wiesner, H.J., Feltz, H.R., and Yates, M.L. 1973. A study of the
distribution of polychlorinated biphenyls in the aquatic environment. J.
Res. U.S. Geol. Surv., 1:603-607.
Cummins, K.W. and Huycheck, J.C. 1971. Caloric equivalents for investigations
in ecological energetics. Int. Assoc. Theor. Appl. Limnol., Comm. No. 18.
158 p.
Smith - 20
-------
Custer, T.VI. and Heinz, G.H. 1980. Reproductive success and nest
attentiveness of mallard ducks fed Aroclor 1254. Environ. Poll. Ser. A.,
21:313-318.
Drobney, R.D., Jones, J.O., and Noseworthy, S.M. 1982. The effects of winter
navigation on waterfowl and benthic communities. Final Report to the U.S.
Fish and Wildlife Service, Grant DOI-C-14-16-009, Patuxent Wildlife
Research Center, Laurel, Maryland.
Drobney, R.D. 1983. Unpublished data.
Duinker, J.C., Hillebrand, M.T.J., Palmork, K.H., and Wilhelmsen, S. 1980. An
evaluation of existing methods for quantisation of polychlorinated
biphenyls in environmental samples and suggestions for an improved method
based on measurement of individual components. Bull. Environ. Contain.
Toxicol., 25:956-964.
Environment Canada. 1979. Analytical Methods Manual. Inland Waters
Directorate, Water Quality Branch, Ottawa, Canada.
Falandysz, J. and Szefer, P. 1982. Chlorinated hydrocarbons in diving ducks
wintering in Gdansk Bay, Baltic Sea. The Science of the Total Environment.
24:119-127.
Fallen, M.E. and Horvath, F.J. 1983. A preliminary assessment of pollutants
in sediment deposit!onal areas of the Detroit River. Report to the U.S.
Environmental Protection Agency, Grant R-005666-01. Regional Office,
Region V, Chicago, Illinois.
Hague, R., Schmedding, D., and Freed, V. 1974. Aqueous solubility, adsorption
and vapor behavior of polychlorinated biphenyl Aroclor 1254. Environ. Sci.
Techno!., 8:139-142.
Smith - 21
-------
Haseltine, S.D. and Prouty, R.M. 1980. Aroclor 1242 and reproductive success
of adult mallards (Anas platyrhynchos). Environ. Res., 23:29-34.
Heath, R.G., Spann, J.W., Kreitzer, J.F., and Vance, C. 1972. Effects of
polychlorinated biphenyls on birds. Proc. Int. Ornithol. Congress, 1?:
475-485.
Hutzinger, 0., Safe, S., and Zitko, V. 1974. The Chemistry of PCBs. CRC
Press, Inc., Cleveland, Ohio.
Karickhoff, S.W. and Morris, K.R. 1985. Impact of tubificid oligochaetes on
pollutant transport in bottom sediments. Environ. Sci. Techno!., 19:51-
56.
Kim, K.S., Pastel, M.J., Kim, J.S., and Stone, W.B. 1984. Levels of
polychlorinated biphenyls, DDE and mirex in waterfowl collected in New York
state, 1979-1980. Archiv. Environ. Contam. Toxicol., 13:373-381.
Murphy, P.6. 1972. Sulfuric acid for cleanup of animal tissues for analysis
of acid-stable chlorinated hydrocarbon residues. J. Assoc. Offic. Anal.
Chem., 55:1360-1362.
Mullin, M.D., Pochini, C.M., Safe, S.H., and Safe, L.M. 1983. Analysis of
PCBs using specific isomer high resolution capillary gas chromatography.
In: PCBs: Human and Environmental Hazards, ed., F.M. D'ltrie and M.A.
Kamrin, p. 165-176. Boston: Butterworth Publishers.
Mullin, M.D., Pochini, C.M., McCrindle, S., Romkes, M., Safe, S.H., and Safe,
L.M. 1984. High-resolution PCB analysis: synthesis and characterization
of all 209 PCB congeners. Environ. Sci. Techno!., 18:468-476.
Parkinson, A., Robertson, L., Safe, L., and Safe, S. 1981. Polychlorinated
biphenyls as inducers of hepatic microsomal enzymes: effects of di-ortho
substitution. Chem. Biol. Interact., 31:1-12.
Smith - 22
-------
Parkinson, A. and Safe, S. 1981. Aryl hydrocarbon hydroxylase Induction and
its relationship to the toxicity of halogenated aryl hydrocarbons.
Toxicol. Environ. Chem., 4:1-46.
Peterson, S.R. and Ell arson, R.S. 1978. p,p'-DDE, polychlorinated biphenyls
and endrin in Oldsquaws in North America, 1969-73. Pest. Monit. J., 11:
170-181.
Poland, H., Greenlee, W.F., and Kende, A.S. 1979. Studies on the mechanism of
toxicity of the chlorinated dibenzo-p-dioxins and related compounds. Ann.
N.Y. Acad. Sci., 320:214-230.
Reinecke, K.J., Stone, T.L., and Owen, R.B. 1982. Seasonal carcass
composition and energy balance of female black ducks in Maine. Condor 84:
420-426.
Robbins, J.A. 1982. Stratigraphic and dynamic effects of sediment reworking
by Great Lakes zoobenthos. Hydrobiologia, 92:611-622.
Rofritz, D.J. 1977. Oligochaeta as a winter food source for the Oldsquaw. J.
Wild!. Manage., 41:590-591.
Safe, S. 1980. Metabolism, uptake, storage and bioaccumulation. In:
Halogenated Biphenyls, Terphenyls, Naphthalenes, Dibenzodioxins and Related
Products, ed., R. Kimbrough, pp. 81-107. Amsterdam: Elsevier Publishing.
Safe, S., Robertson, L., Sawyer, T., Bandiera, S., Safe, L., Parkinson, A.,
Campbell, M.A., and Mullin, M.D. 1982. PCBs: structure-activity
relationships. In: Advances in Exposure. Health and Environmental Effects
Studies of PCBs, pp. 229-248. Report to the U.S. Environmental Protection
Agency, Contract 68-01-6554. ICAIR Life Systems, Inc., Cleveland, Ohio.
Smith - 23
-------
Smith, V.E., Spurr, J.S., Rathbun, J.E., Filkins, J.C., Rood, J.T. 1984.
Organochlorine contamination of sediments and biota in the nearshore Great
Lakes. EPA Project Report (CR810232-01). In preparation.
Stickel, L.F. 1973. Pesticide residues in birds and mammals. In:
Environmental Pollution By Pesticides, ed., C.A. Edwards, pp. 254-312. New
York: Plenum Press.
Stonard, M. D. and Grieg, J.B. 1976. Different patterns of hepatic microsomal
enzyme activity produced by administration of pure hexachlorobiphenyl
isomers and hexachlorobenzene. Chem.-Biol. Interact., 15:365-379.
Thompson, J.F. 1974. Analysis of pesticide residues in human and
environmental samples. U.S. Environmental Protection Agency, Research
Triangle Park, North Carolina.
U.S. Food and Drug Administration. 1984. Polychlorinated biphenyls (PCBs) in
fish and shellfish; reduction of tolerances; final decision. Federal
Register, 49(100):21514-21520.
White, D.H., King, K.A., Mitchell, C.A., and Krynitsky, A.J. 1981. Body
lipids and pesticide burdens of migrant blue winged teal. J. Field
Ornithol., 52:23-28.
Smith - 24
-------
LIST OF FIGURES
Figure 1. Mud Island study site on the lower Detroit River, Michigan.
Figure 2. Aggregate percent of PCB congeners in 13 scaup and goldeneye
ducks. Asterisks indicate 6 congeners forming conservative
ratios.
Figure 3. Median distribution coefficients (Kj) of PCB homologs for
ducks/sediment, carp/sediment and oligochaetes/sediment.
Figure 4. Fat/lean dry mass versus week collected for 169 ducks
(Drobney, 1983) and the analyzed subset of 13.
Figure 5. Linear regression analysis of total PCBs/% lipid and 6
congeners/% lipid on percent lipid in 13 ducks.
LIST OF TABLES
Table 1. Aggregate percent of animal and plant ( ) matter in gut
contents of 169 ducks (after Drobney et al_., 1982).
Table 2. Characteristics and tissue composition of 13 ducks analyzed
(after Drobney, 1983).
Table 3. Organochlorine concentrations in Mud Island samples.
Table 4. PCB composition by congener and homolog of Mud Island
samples. Compounds of potentially greater toxicity are
indicated (*).
Smith - 25
-------
FIG. 1
KILOMETERS
to
20
MICHIGAN
DETROIT
LAKE ST. CLAIR
MUD ISLAND
LAKE ERIE
-------
FIG. 2
HB
L-.-Jj.'.^-:J 1
lf=FH
|fTlt
I E=
» f" r"i i
i r*w^*
rTtfl
LQZ
902
SOZ
ZOZ
IOZ
661.
861.
S6L*
eei.
161
091
£91
981.
081*
9ZL
CZL
05 L
991.
C9L
ISl
O
frfi Z
9CL
«^
s
-------
FIG. 3
(PX) 'dd300 -amisia
-------
FIG. 4
00
1 I
I
<0
I
o
I I
° *-s
I *- o
00
hO> O)
00
-ID ^
-m O
z
LU
UJ
- CO
SSVW AdQ NV31/lVd
-------
FIG. 5
(0
oc
ILI
0°
H- <0
I
I
I
I
^.
-------
TABLE 1
MALE
FEMALE
BOTH
CQWON SOLDENEYE
Bucephala clangula 27.3(72.6} 44.0(55.9) 35.6(64.8)
LESSER SCAUP
Aythya affinis 24.6 (75.3) 33.1 (66.9) 27.9 (72.1)
GREATER SCAUP
Aythya narila 12.7(87.3) 28.4(71.6) 18.6(81.6)
-------
TABLE 2
SPECIES
LESSER SCAUP
Aythya
af finis
GREATER SCAUP
Aythva
an'1! a
GOLDENEYE
Bucephala
cTangula
COLL.
NO.
8
13
53
59
72
79
82
47
50
66
1
2
29
WK
COLL.*
7
7
10
n
12
12
12
10
10
n
6
7
8
AGE/
SEX
jv. F
jv. F
ad. F
ad. F
ad. F
jv. F
ad. F
ad. F
ad. F
jv. H
ad. F
jv. M
jv. F
TOTAL
WT. (G)
969.9
1029.1
974.5
778.7
710.4
714.9
710.4
910.7
1006.3
969.9
796.9
1293.2
860.6
DRESSED.
VTT. (6)1
710.4
746.8
787.8
487.2
528.2
519.1
482.7
601.1
614.7
642.1
555.5
979.0
592.0
WATER2
54.2
51.0
54.5
65.4
59.9
62.4
51.8
62.9
54.4
57.7
54.8
49.9
62.2
F^
23.3
27.5
18.6
6.7
13.3
12.0
23.5
8.9
22.8
17.2
23.0
27.3
11.9
1 LEAK'
DRY MASS4
22.5
21.5
26.9
28.0
26.8
25.6
24.7
28.2
22.8
25.1
22.2
22.8
25.9
'Wet weight irinus head, feet, feathers
and viscera
2 Recovered by freeze drying.
"Recovered by petroleuir ether extraction.
4 Regaining after water and fat extracted.
*Weeks 1-12 are Jan.-March 1980
-------
TABLE 3
yi
if
D
Z>
3
GOLDENEYE 1
Bucephala clangula !
D. «0
=>
«t -r-
o v-
i/i re
et
UJ 1C
SI-
UJ «->
g£
LESSER SCAUP
Aythya af finis
O
D.
1.
«
a. u
ac.
< in
0 3
c
T
ex
£
/I
n
o
o
Cf>
x r->
^ H
r^
CT>.
X n
-^ n
Cn C
E
en
* r-
^i. 11
en c
en
j* o>
^ «
r.5
i/>
i UJ &>
§»-.*.-
UJ ~^- n
« < en c
d5*~
*
UJ CJV
S<7
o en c
uj e
i/>
ac
o en-
r- j* r-
I/O »^. H
uj cn c
E
DC
UJ c- ,_
r- -» B
< en e
* c
COMPOUND
O 1- CO
O r- r- O CM
r o r- C* O O O CM
^ . *oCf «v o t
oo r- o - o o »-- i .
.. ... .« rOC^O*1 ^O
8r o » o o r~- oc o t o «
r-OO*-> O O O *-> tVJ O
r- p- o n O*->OCTOO*J'
o *J O ^ ro ic i 4-» <»)
CC4J(«J *«l ' 1 1 1 Or O lf> Or <"1
.IOOOO O -Hr^+jQOCh-t-vc -
r- «VJ r > O> O Po f O<*5O-*'fMOOO
,i.+l.O- 0 eC'r-'OO'CC-V
lCi»r~OOO O OOOOxfOOOO
r^' . ' O O O C5 O O C3
tVJ CM
ro in CM o r^
OCNJO--I^O r^ oo -~* »
ooooioo r-i r o Oi~- O0f-«r>
00 t . ^ - . >OO % . . r^OO O» «O O
OP^' O' ^O^O * ^ O **- <*o * ' ' ^l£)O»^^-
~ O O O~. O ITIO r- <£>.. O O r O W O
OrOi OlDOt OCO i OO' OO Or- Cf
Or- o*jo4jo»-1 oo o+^r- MOI inooo
r r^-oo o - rn O4->m f>oO4-> o *t 4^ o *J
Or-»J -«J MD O «*> <> OOO«-> in*JO
O4J O^ffOOD ' ' r OCSJ- +!m OmcsJ 'COf^
o*c 4,0 ^o *oo it +'r- e>i coo oec +IM «* or^ 4ic\j
l\in -fi» mo o
. r~-f « - PJO' . . O O *J O 4-* OOO4->O-*->lDOO*-)^ CMO
r- CMOO O O4->. . r~*->O fOOO4->»~
Or-4J .10 -U1O*- Or Ot^O .^<-14J . «t
c*4-» o«tf ooo 4'<\j i Or 4'cv 41 1/1 »to on oc* r
om 4>o 41 o OO 4i r mo r-*o oin 410 OCfi 41 n *-
«-r^ 4r- *oo coo ir>o «^o CMO eno 4>o coo 4CM no 1/1
,.CO.r-.r--O- r-.O-O-l^'CM.O'O-CJ
PJ no oo oo oo oo oo oo r-o oo coo oo o
oooo' ocJocSesoocJ
t^
o *o -~^r
OCM Or- COO ^PJ VC
Or~Or~ t O no^~O>' r- >OOK> *»
Or-r-O O r- O n O CM O O ~ ^ O4Ji OCSlO*OOrO r O
*4JO4Jr- f «»«Jr-*JVO4-JO+->inO>COO*->
~ OO r- OOO. OVOO O O r 4-> CM 4J
r-^4J .000*0 ^CM 4>CM »m «cp r^O'tf- « oo
OCM 4', n no CM f^o on Ov or- 4'r Or oio 41,
r-«o 40 mo CM >co no tio +> o 4, o mo 4i«Mj 4!, coo
4' . O * < O O-O Q-r- OC U3 rO Op CO C\J
r- r-O OO O OO O OO r-O r-O r-o OO no "*O OO
vo .............
r-OOOOOOOOOOOOO
vo CJN oo r- « h- ch
ror r-rjc\joi/>m
5oo i i i 1008000 i
ooo oooooo
r CM n r- \O ^ n
COOO f r-^-OCMCOCM
nOQQ QOQQp CMr-
IOOOO t 1 lOOOOOOO
oooo ooooooo'
cr in co
r-ODCMco^-i co >n >£> rr>
cM«or-cMoncsi*oo^om
CvJCMOOOOOOOOOr-^O
mooooooooooooo
m M>ont~.vna>
r~ -r-^-oOr-CMCMOinoon
in CM ...........
*Or-Or-OOOOOOOOO
01
c
O) tl U
N £ 0
C 10 «l r- L.
a> -e» c jc o
jO t. us u i
P 41 1- O T3 "O JC
CO L.O W C O O Ol r- £. C O
o o z o xieZ r--t)jc o o 10 ui o 1-
fi. CO X CO-OCD JC~-O r- Z C O O O
f eo c uxijciooao
r- U « «~- VI Z
w m j= « _ i
-------
TABLE 4
U.
>-
u.
X
£
ft
Q.
ZD
«t
<_)
IS!
CL
^J
«t
UJ
cr:
CJ
CL
ID
«l
t_)
to
cc
UJ
to
in
_j
o.
OL
<
t_>
VI
U
ij
<:
3;
j
3
_j
^
*
LU
l/>
)N6ENER
'_>
o
o
3
er
c:
0
u
V
rv
.c:
CD
u
Z3
ra
1-
o
(TJ
>,
x:
4-J
>!
«I
hya affinls
>.
«t
Cypn'nus carpio
(Composite)
E
u
*}
1
en
^ ro
. M
I"5
CTV .
jtf ro
--^ n
cr tr
e -
ov ,
jf r--
--. II
C7> c
E-
_* CT.
-- II
1~
.^
\ tl
-E-5
_*
^ 11
i1^.
STRUCTURE
£
jj «.
a.
n n
ro ro
r-- f--
T--. r-~
O O
0 0
' O ~ O
gtx, O cs.
0 O O
C-o "0
r-v i r-- i
t «t
O (V O CM
1 O 00 O 00
O -0
o o o o
4 4
WO « O
0 O
o o
w «
-CM -CM
r-^ O r~ o
u^ . in .
- O - O
1C 1 1C 1
»» w
(~> vc f~^ vc
O 0 O O
mo ir) o
o o
0 O
cr>
- CM CT>
no vc
n .
o . -o
n
01 I ro I
CD
8 1C VO
VO CTi VC
O CM C
CO - O
* . o
if O 4 O
1C Tf
>C O CSJ
O O O
1 I I
in c* ^-
*c oo
o o o
00 0
O O 0
fl
">»
0.
1 £.
t- Q.
a
J3
C L-
e°
r\j v 3 j:
OJ (NJ
* I--
*^as -oo
8^ 8^
o - o
1 . .Id 1 . 1
ro ro rn m i
ro - i ro ro r--
. rv ^^ CM
i CT' + ' O O, 4' O
W O U"1 c\j CD
O O Tf O «T
O O O O O
O~
O «3
CM ^
ro ^ o * *CTI
. .cr. . o
r~- r- o O O
ic o in
^ - o ' - *
ro i O
i ro 1 ro ul CO r-^ \O
ro * O O *~ ro c\f
. . m -^' O O
«t O 1C O
vc vc +1 o o 4, o
ro o oo t
O O f^ O u~>
o o o o o
cy.
.-^uo ->C
cr. o o CNJ
C\J O O O
~~~ o --v C^o o f
O 1 f^ t CO 1 i
I i O ^ « IOO 1
. ^ r- co o cr «
O O ' CM ' O ro O
-t , O ro O O Co
u-iOPO *>OOOO -"O
§O ro r^
o vc o ^
O O O O C
tc
CSJ O O O ro O
- -ro - *ir> o O ro . ~csj o Csj vo
8.Q .^-O -O
csj O O OO 'O W
i 0
' i ^ i in vn i i csj i o i
^j cri in
ocsj^-inovcocsjo^ovc * CM
VCrOi CM -O ' roOO -ro CM
O -OOOOO -OOO roO
O O 4 . *. . O *i
j,O-'OcsjOmO*OCMO ' O
vc in CM vo vc CT> -f1
ro O O ro O O
O C3 O O O O '
in i vc CM
««j vc in r-. in
BOO CSJ O «»
o o i o o o
OOO OOO
OO <"O
"> vc CM ro w ro CM
O O » CTi
O O O O O O O
O O O O O O O
in
'x
c
1 r^ror^CM oror^ooro
O ICOVOO O VO O
CLo ~" o"~O ~~"o"^ o
S^r vc m ' CM
i in i - ^ i o '
ro I i i i< rooocoooroo
ro ro ocr>ro oroowro OOOCDOOO
ro *-^ CM * ' Csj O ro
OO OOOO OOOOOO
CM cn roooo CM o o cr, «ooocno
f O VCOCMCMOCsjt ro
OO OOOOOOOO
OO OOOOOOOO
1C ^~~- *~ ~^ ro '^~~
. .! OVC' -C^ ovo-^ Om
r-^ o ' -vc O^r-*.^- Oror--OOin
vc- o- 10x00 o vc o
^-o o - o-
O O O O O
en i 01 m ro i o
ro i vc i co i ro i in i
*" - ^~* ^3 OO CTl ^~- C2 C3 C? C? ^J f~>
ro i ro 0*3-1 i rororo ocrtOr«-i oroocr-ovc
ro ro »O Ovcro < -CM 'CM-o-ro
OO .0^- OOOO OOOOOO
+ [. O» 41-4. 41. 4. 4.
*t oo 4iom ooooo oo- c-ao
Ownro rov
0 OO OOO OOO
O OO OOOO OOO
CM
.^-.ro ^^vo' -m^ ~CM ' -00- -CM- -ro
cri r -co cr>cMoaoooo oinicoocr'
CMO OCMCMOOOOO OOOO-OO
^- . o i ^-'O
O i O O' ^O' -O ^-O -O
« eo i
^^ Oi iCMt^-i«e-i miin *ci
i * i O i «^*- rn o csj CM i f or*-CM
O CM w 00 r~- o O O >» O W OroOVOO
- .< i OCM.CM*' ' .r -O'1
OO - -OOOOOOO OOOOOO
OO roO f 4'OroOCnoooO OOO-OicO
O * CM CM ro ^ r-, r ^
o o o ooo ooo
O O C3OOC3O OOC3
i O 00
cr> - . in ' -CM CM
~OOVC' -O -«i-vOO i -i '-CM' -VO CM ~O
en .OCMCr, -ro o in .. « OOO i OOOO -O -O
4 .41 .41 ,O »O "4; »O .4' .O "O «4. .41 .O -O '
i OOOOr O4iO4 OOCM in cnr^
ro r CM- ro* r CM CM CMOO
O OOOOOO 0 OO
IO IOOOOOOOO IOO
O OOOOOOOO OO
CMI o or^ooco room
in r » r-v ro ' ^ro^« m< OOCM
OOO OOOO CMOOOrsj
oooooooooooooo
cooooooooooooo
-- .. .- ... u-)
ro^ininvCvc^-ininmvcmvc »
rorororororo^-^j-^-mm. - w
CMCMCMCSJCMCMCSJCOCMrsjCMrorOro
CMCMCMCMCMCMCMCMCMCMCMCMCMCM
ocsjn*in«or->oocncMrnn«o
^^^^^^>-^«3-w-ininvcior^
a.1
cr-
c
-------
TABLE 4 (CONT.)
LLJ
>-
UJ
z
UJ
o
_J
o
O
Jj
1J
<;
I
<_>
CD
_l
O
E
*
LU
C^>
CC
Z
LU
O
z
(_J
oo
o
«3
c
«tj
u
IT)
(C
.C
Q.
OJ
U
a
,
jr
>,
1
in
c
**_
U-
P
IO
X
.C
>.
«£
0
^
1-
«5
U
l/>
3
C
g
(_>
Oi
*->
c
o
<_J
"?
u
0-
L
M ro
\ II
en c
E
"pi-
.^ ro
cr c
E -
cn^
.* rv
^- II
£s5
.* CT.
^ II
if -5
en-
j*
~x II
_~5
a <
-- n
i~
UJ
cc
rD
u
r>
a:
h-
2
LU*h
°"
8*-
p . .
8>c - O p vo
r\j - « o TJ
. o
O CM »-^O
O I
ao " o a? i
«- o vc *f
o in o OD
. p o O CM
0 > 0
t' O r^ O 4 o
oo
o ro
o o o
- - o r~ *~^
8O 0 0 Oun
ro ' O 1C
r . ^^O
^O 'O
1C 1
t~* l i 1C I
cv o ro vc
O n O CH
. ro o O «
4 o (O O + O
<£> *t r^
CM O f>
O C O
ro
in
-o
.^o o
O to O O O CM
O d i - --'
i rv. \£> i
m ,3-
i O O fl CM
» « O CNJ
+0-0 + o
o *- «
ro ro ^O
COO
^viri
CSJ O TI
vc o o
g. . x r-»
0^-0 O
O TT
^ i og l
*c in o o
O> . r-> rj
O O O -03
--1 O O Jl O
n CM o^
(V O O
O O CM
OO ro
o o o
8 $
§ g 2
o o o
*/t
1 ">-,
0 C
*J jC
ai ex
u^ in jo
- O l.
*^ **. E ^
TS- « 3 .C
CM ro
*
«
r^ CO
^ -O ^~ r^ - - ^< ir>
r^ . o --^o* o f O ' -~-*<
icO OOr^- Oi-^OO' r- o »
- OOC . o o ^r -o o ^
ro . O O
§l OO '3 f^ ^ ' 'O VC? l O
CM ^P i i^ i ^3 ro
o r"' . - i ooio^O O O * ' lOCOf-^t
O~' ro OTOOOOOOOCiw O««C
OOro O^f «O f ' «tT) »OO^ »O Oin
4 O~ OOOOOOOO -CMOO -in
r^« o-*!«4[--*.4.o*^«O-
c^or^ *IO-»OO~OCMOOO + OOO *!O
OVO U^ ^ VD i 1C
O ^O" pcnocM OCM
^ . p o Co* o M o
o orvi - . Q f»- -ptM
CM O ~O O O O O
CM i 'Or~ r~ r O in *-
81 i * . CM i en i r^ i *w^
vo^~ 10 i o ro i O 10 i
roro 1 cv O*~ro O1 O^IO O1 ID
OO"^ OW " ^ -VO . r- r CO U1
-.0- o o oooo-woo -oo
r-^Oro -^'OroOf^- (MOf-*O4O*"~O 4>O
Oir> i^. ocO' C^CM oo
OO O O O O IT) O O">
oo ooooooo o
. .« *~.^ t-^ O VT)
ic ^- - . o ro --^ru ^* p ro ' -CM
COO CT>CT> CMOO- OpldOO U3O
^ - C\JC\J VO' .f-v'O' --^CO OO - -
o o . .-_o^-O' .^-opc^^o po
^- . o - o o o
VI OOOI-v O'CMI p^lr- Or I' CM
O 1 CD 1^ O 1 ** * lf> -
o- 01 o*coov OLD ior- '
ro. . O oor^O«*OioOvoO*3-ld OV »
OO^± <*' CM »P 'O *CM 'OCMID »O IT)O
-*O- OCM «OOOOOOOOO ' OO «CM
ODO f-^ 4OlT)OroOU~!Or-HO-*'OCMO 4O
gOCMOTj- r^i r\j» '
OCMOOOOUOO CT>
oooooooooo o
r- .*} 1 -~C
^-vc vc ~CM pro
80 - p o LO o ooo oo
. ^ .CM -iri o ^ - r-^ .. - f
O O vo O ' -CM i . O O * O O ' ^O
O -OOQ -OO O O ^r-x
i o ocs, n i r- oil o i p-
vc -^- - ' o ' o ^- vn ^^ co o u^
r- 1 -iro OCD IOU-) IOOD-
or-ic rovo ocr>-rooo Oir»v£> om i
O' r-~ CT> r^ CM - OOCMOO -OCMCTi «o
OO ' OLO^OOO-' O 'CMOO * OO ^-«^
o o » o ooOCT>oaoo-"O o»oino r-
roro' 4. LT> O CM < CM" t1
OroCMCMOOixloVO CO
OOO^-OOOOOO CM
vp vo r^ oo en
p VO CM K. * * 1C CO
OOPCMPP^~ O UO
0000000 to i o
ooooooo o o
CM op ro ro in r*v
ro^j-crivo^1 copr^p
§*~ o 5 o p f~~ o » o CM
OOOOOOOOO '
oooooooooo o
in
1 >1
10 c
vvc-icvnvo iniD ex:
-in « .- L/> «. * OJQ.
-- . - ^- - u-) . - - a.-^
"oro*yir> »v *>-^-^ JD
rororororo^- > »^-^- 01.
--.. .... E,
CMCMrsitsjcMCMCMrororo njz
CMCMCMCMCMCMCMCMCMCM
CM^f^-ul^p' »*-Cocn
aocococr^c^oO' >
^O O IT* O «
^co p co o «s o CM ' -
8 OO- -CM O OO
*- p Cn- O ro
O --O O O O
' O vc « ^O
1 CM 1 - U"> 1 CM 1
VO 1 1 If- 1 O O O I
rove ro OCMro ou~>O'*-'^
OOO ro -\r> o CM i o c^
O -~^ OO-*»OOOO-O
O +l O 41 . > O
I'O »» CMO-l'OmOOO-tO
ro IT) 1C r ro CM O-i
CO O t^ O O '
O O O O O O O
~~a P °
- ^ Q CO O CO -
§ -~«5 OOr-OOro
ro t~. »-» i -Ol
. vc f-v^o O
^~O -O 10 ro ^O
i ro i
«t \ *-^ ^ < CM O CO 1
ID l - i ro CM mioroor^r*.
OOO ro ro OOw> .ro»- O«-
ro * .1 OroOOOO -csj
4'O r -MOOirOOCMO^'O
«» f^ CMCO-KVOCMSO
CM < O ro O O ro
O O OO--OOO
. ,ir>
- - - O O r°
- .*» ^m O cn O *C O O O
8ro i O'-'^ O 0 t .- iri
l~^ . r- . Q (0 OO
O **-~ O ' 'O O *~* O O O
-^ » ro l
i o i ro i - vo i co ' '
« 1 CO 1 ^ tn 1 r o r
cr»o OO «» OVTJO o^or^**CM
Ort -ro >»ro.. .0 vf
O OO ^-- OO-CMOOOO-O
-HO CnO VO - O-fOOOCTiO^O
OO CO r O> If, TJ t».
o OOOIOOCM
o o oooooo
--o «oo ^-
. *vc p vc p r^ - * P ^ ^
8 CM O O ~-^CM OOP' O ' VC *00 - ^ - " **
CM«S-« »OrO' voov/^o 'P
> O OCM - -O .^-O- . -O .OOOO
-O 'O Q CM . O
l^-l Cf!> 1 O 1 -1
S' ' 1 ' ID 1 CO 1 1
coo«-oo ocvjf^ oo ID OT- VD
OLD Oo> 'CNjOi *f t^ ' CM' CT11 cr
OOO O O O O 1C ro .O -O
TOCnO-nO O-t'Oroo . O ti O + O' O
rocMOTjcnr^. -fioooro
i OrOO" OOCMCMCM
OOOOOO OOO
i vp Cft«-iD roiciD
CMO Oror^vovcro^
§o p o o CM o o o
O 1 O O O O O O O
oo ooooooo
voch IDOQVO iDtn^f
§O ' OOrsif^r^u^vD
P pOpCMOOP
0 1 OOOOOOO
oo ooooooo
1
*3-lDVOVCVOlDlD- VOID
«*VTflDVO- - - IT)
rororororovv^^v
ro ro ro ro ro ro ro ro ro ro
CMCMCMCMCM CMCMCMCMCM
CMrsJCMfMCM CMCMCMCMCM
coeft' vvcr^oO' «jvc
ruCMrororo rorovv-«cr
c
v
-------
TABLE 4 (CONT.)
u.
u.
z
UJ
c
o
Q.
X
«t
CL
CD
Q.
_
Ct
4.
>
Q.
CC
U
E
0
_J
C
AJ
a:
UJ
z
0
s
3
cr
c
u
JC
a.'
u
1C
L_
10
«?
>
jr
>.
<
c
n-
X-
«
jr
o
a
u
in
3
C
g
03
4~>
0
O
O
IT
L.
f~~
u~-~
J£ PO
"^ n
a- c
en. .
.* PO
cr cr
e
-^ ii
cr,-
it
I^5
-s. II
!~
LU
i-
u
t
uo
xJ **
gco *-^*^ ro
oo CD CT '""'"- c^
8fVJ r- O G O r- O
. ^_- . .in t . vc . ..^j-
O O CD - 'O -O O *
** C? VO cvj CD ^
1 ^ t CO ' ^ ' '
f O x^ | r- ^ 0
rococo o co o co 4->
OOO T r \c ^ in vc
ooovovcoooo com
VD CSJ -* VC r- *i
^~ C3 *&- C^} CD CT^
O O * O O csj
8cc cp
csj O ^- * -
8r^ o o co
co - - o
^ CD CD O*> C) r~~ ' *"** C?^
x'O CO O * -O O >
uo i f ro en i o m
r-x. i O O ro i
SO CO » O CO CO " ' (
VC1 O CO O CSj
- O O m co O o *» to
O O O ' CO O *-. O rv
m c\j -t-. ci m 4i
(xj O in o ^^
O O oj o O ^
ro
ro yD O ^-.rv. VD
\C ' -o *- -r** co *C O
COO Q 'O CD O *-*
*- ^. t ^^ PO x_^ C> ^>.
i & *- ^i IO
5?OU->OOOOO*'O r-O
CT> CX1 -t ' CTt1 ^~" * '
O O> O *~~ CO
O O O O ro
*""* O CO
^-^ Ci *^i
8O -"^-O » CO * CTi
vC ^ *^~» ^--^p ^x. t^ ^-^,
--^ojin.gw^(- -| g
1 1 1 CM W '
p m o ww- i
PO" O!~>OO -O CO PO
OO \£} * \C '"" 4>
O O O O in
f 10 in
O O"* ^- » csj
cv o * 0 O ro
O 0 O O 0
o o o o o o
CSJ tf CTi
O CO UO t\J CSJ
C\J O ^3- O O ^
O O O O O f
o o o o o o
LA
to in in u> ic:
- » « * * TO Q;
tn in - - x. jc.
^ in «* ^- ^ ^
ro no «» - o l-
"* ** m ^ ' <)
ro rvj cv ro ro 5 x:
CM (NV CM CJ CSJ
CT> r ro vO CO
O QO*
.v_^o- ^ O O » O i^- O-
- f cv I t r- t m 1 % I r- co i * f i Q f ^ r^i
- 00 i O r»» ro r-* - ' i t ^- t CO m » cT> »
IF oic ooomocoroo c\» o ID vo o ^ » tc o f^ i
p**! ^^ ^«^ CM iO in C- ** O C\J ro r*- (NJ CO CO in iO ^~ CT^ f*-- C5 ^2 C^ *~ C? "" M
cv CO CNj ^^ ^2 C1 ^D * O* tO C? ro ro C> *~~ CD ^D ^~ ^^ ^D * CD f**-. f\j
CNJOO-nOO »O -O O »O -O -O
-^oooinocc cr co » t 4-1 41 r-^ 41 ^- ou r^. ^i
r~> vc O O^> ro c\j co O r~ O^ »~~ ro ro to
oooocooooooooo
g %O Q* OCOOChp'
ro - -O *^^ O CNJ * . O c\j O 10 O f--
8< o VD *~~ o r*- o ^o ^ -co ooc^*~~Or~o*^o ^^»
t c> ^~ > ' t£j C* C^ fv *"- ^*- C) in * ^ Cj tf" ^^-^ x -^ * * * Cp
p - O "- ' O - - -P -O O O O O O 1C
^I-ICC PO WIO Co 00 CM 1 -1 >O OOIO O O
f C^ rO CM C3 C^i C^ VO W C5 C? PO W CO W C^ in t> 00 W 1
r-^ o ' OOOO CVOO -POOO -WOOOOOO «r^
OC-I^OCM *'Owor~-o+>ooO4(ooo+,oooaoo o 4'co
O'-O'-oocMPowomooo
~oooooo« oooooo
*~ »^ O - »cj **- O C\j O ro - vt£> in
8vOtno 'tf-vOO' O ro iOOO OOOCf -O>
x_^ Q co »tc *Q""~ If} I f O ^~ < O f**-
i i i o ( (*x. 1/1 t ^ t in * r«x in i in i o
-t«tf O^C* O-'S'Cs.iicr^ s -i Oro t O » Csj co ^-i
Of**- » roioO'ctOCOinCTi CsjOOinO O^-OOOro »
m co CD CD * ' C* * vO CD CD CD ^5 CD 's- CO CD CD CD *~ C^ CD CD CD CD CD CT^ ^-*
OO^OroO 4'OOOinO 41 OOO 4>oinO -*1OOOOc\iO' O O
-«-O^OOCsjCOOO-*'^t* f~~ ptntO 4I
ooo- oo ^^i-oinooo co
r-ooOOOOr-OOOOOO ro
O CO QOO- ' * > Qro^-*ro
O^-OO Oroo^tf- CM R1^ OOOCO'^-'f
CO * 1> * CD ^^ * CD ** * CD CD CD CD » CD ^ 'CD 25 * ' CD ** ** CD ^^ro
r^r- i - O Q O fsj r- o P ^f i Q'
-~^ co ' m -^- c>i * o ' ' * ' co ' * co ^ t r-^ t o co
i ro o r^- i O ro ^ i i ro i O r*. csa co *
CT> O ro O *t O O fsj O O *~~ Cti CO O co, CO O \O O Csj O ro - I
«a-o*ooo oooo 'O^o^r » o" o TSJO'OOOCDO om
4'OcsjO' O + OCoOroO 4iOOO -'-Of^O 4'OvoOCOO« O csj-
CT\ co ^~ ^~ co vo ro 41 t*- ^* r*x. *~ *$" i+) 41
COOO^-OOCSJCvjCOOVOOOO VO
ooooooo oooooo
roOO'~~OOOrO' OO O O v~
ooooooooooo too
ooooooooooo oo o
UO- «-to-vo-vO VD ID VO W
^inuominvo * in in - ininmuo *oc
«*^^«e-^'«3-^" - *in- - in Q.JC
POCMCMCMCMCMCMCMCMCMCMPOPOPT' §j=
CMCMCMCMCMCMCVJCMCNjeMCMCVCMCM
l^pT^pC^^gScSSsIsS
vo
r^ --^ ^** o o
ooo
p o o
e o o-
- 1 1 CO
in vo oo
m o in o 01
PO VO O CM O O
O O O O
O -0
4' cr o
> W CM
0 0
ooo
« O CO O W
80 o o
t . . * .
O 0
O in w
o- i in i
CM 1 O O
CM O C^ O OO
O > w rj
4.1 o in o w o
in vc PO
O O
ooo
~ . .ir,
O O vc p O
. -w o o
g. . 0
o o
r 1
S- o
VC O CO
ro o
o o o o o
41 o cr o O
f*~ ^» PO
i O O
ooo
^ "~- P m
Cr? "" CO ""
CO O- 0 O
" 1 00 1 W 1
W CM
W CM O VO O W
o in «CM
o o o o o
*< O VO O O O
O m PO
r^ 0 0
ooo
^- ^v. fx^
ro csj
888
ooo
PO csj ro
SCO f
O 0
ooo
ooo
m vo vo
in in -
- - in
«*« in
#1 »
* * *
ro ro ro
ro ro ro
* * *
CSJ CSJ CNJ
CO (SJ CO
^- in co
Cft O*i CT"!
------- |