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EPA #540-F-00-006
OSWER #9285.7-32
October 1999
SHORT SHEET:
IEUBK MODEL BIOAVAILABILITY VARIABLE
Office of Solid Waste and Emergency Response
U.S. Environmental Protection Agency
Washington, DC 20460
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NOTICE
This document provides guidance to EPA staff. It also provides guidance to the public and to the
regulated community on how EPA intends to exercise its discretion in implementing the National
Contingency Plan. The guidance is designed to implement national policy on these issues. The document
does not, however, substitute for EPA's statutes or regulations, nor is it a regulation itself. Thus, it
cannot impose legally-binding requirements on EPA, States, or the regulated community, and may not
apply to a particular situation based upon the circumstances. EPA may change this guidance in the
future, as appropriate.
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U.S. ENVmONMENTAL PROTECTION AGENCY
TECHNICAL REVIEW WORKGROUP FOR LEAD
The Technical Review Workgroup for Lead (TRW) is an interoffice workgroup convened by the U.S.
EPA Office of Solid Waste and Emergency Response/Office of Emergency and Remedial Response
(OSWER/OERR).
Region 8
Jim Luey
Denver, CO
CO-CHAIRPERSONS
NCEA/Washington
Paul White
MEMBERS
Region 1
Mary Ballew
Boston, MA
Region 2
Mark Maddaloni
New York, NY
Region 4
Kevin Koporec
Atlanta, GA
Region 5
Patricia VanLeeuwen
Chicago, IL
Region 6
Ghassan Khoury
Dallas, TX
Region 7
Michael Beringer
Kansas City, KS
Region 10
Marc Stifelman
Seattle, WA
NCEAAVashington
Karen Hogan
NCEA/Cincinnati
Harlal Choudhuiy
NCEA/Research Triangle Park
Robert Elias
OERR Mentor
Larry Zaragoza
Office of Emergency and Remedial Response
Washington, DC
Executive Secretary
Richard Troast
Office of Emergency and Remedial Response
Washington, DC
Associate
Scott Everett
Department of Environmental Quality
Salt Lake City, UT
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IEUBK Model Bioauailabilfty Variable
Introduction
Definitions
Performance of the Integrated Exposure Uptake Biokinetic Model
for Lead in Children (IEUBK) is a function of site-specific param-
eter input values. A site-specific determination of soil-borne
lead bioavailability is, therefore, advantageous for improving
predictiveness of the model. This short sheet discusses issues
to consider and applicable methods for determining a site-
specific bioavailability value for soil-borne lead.
The current default estimate for the bioavailability of soil/dust in
the IEUBK model is 30 percent as an absolute value. This ab-
sorption fraction is partitioned into a non-saturable component
(6 percent) and a saturable component (24 percent). Investiga-
tors (Casteel; Henningsen et al, 1998) have observed variable
bioavailability across different soil/lead matrices, although the
majority of samples are generally consistent with the IEUBK
default value. Soil particle size (for soils sieved to <250 um or
60-mesh), mineralogy, and lead speciation are among the fac-
tors that influence bioavailability (Steele et al., 1990).
In Vitro techniques, such as the physiologically-based extrac-
tion test (PBET - Ruby et al., \ 996), have been developed as a
means of capturing the impact of the soil/lead matrix on
bioavailability. However, physico-chemical characteristics of
the soil/lead matrix are not the sole determinants of the highly
complex biological process of gastrointestinal absorption. In
effect, solubility and bioavailability are not interchangeable
terms. Until such time that fully validated in vitro techniques
become generally accepted, the recommended approach to
demonstrating site-specific bioavailability will need to be sup-
ported by an appropriate animal bioassay.
This short sheet reaffirms the provisions of the 1995 Adminis-
trative Reform for Lead that requires review of data that may
set a precedent. Bioavailability data (other than from pub-
lished studies using the juvenile swine model) that are intended
for use in an EPA risk assessment using the IEUBK should be
sent for review by the Office of Emergency and Remedial Re-
sponse. This review not only promotes better science but also
promotes sharing of information so that all EPA Regions can
benefit from new information/analyses.
As indicated in the Guidance Manual for the IEUBK Model,
bioavailabilitv refers to "the fraction of the total amount of
material in contact with a body portal-of-entry (lung, gut, skin)
that enters the blood." Bioavailability is also described as
absolute or relative (USEPA, 1994). Absolute bioavailabilirv is
the amount of a substance entering the blood via a particular
route of exposure (e.g., gastrointestinal) divided by the total
amount administered (e.g., soil lead ingested). Relative
bioavailabilitv is indexed by measuring the bioavailability of a
particular substance relative to the bioavailability of a stan-
dardized reference material, such as soluble lead acetate.
It should be noted that the bioavailability input parameter in
the IEUBK model is an absolute value, but it may be experi-
mentally determined by relative means, provided that the ab-
solute bioavailability of the "standardized reference material"
is known. For the IEUBK model, soluble lead in water and food
is estimated to have 50 percent absolute bioavailability. The
model presumes that the relative bioavailability of lead in soil
is 60 percent, thus producing an absolute bioavailability for
soil lead of 30 percent (i.e., 60% x 50% = 30%). It is acknowl-
edged that this value has significant variability and uncer-
tainty, but it is the estimate under which the IEUBK model was
validated with comprehensive blood lead study results.
"Bloaccessabilitv" is a term used in describing an event that
relates to the absorption process. It generally refers to the
fraction of administered substance that becomes solubilized in
the gastrointestinal fluid. For the most part, solubility is a
prerequisite of absorption, although small amounts of lead in
particulate or suspended/emulsified form may be absorbed by
pinocytosis. Moreover, it is not simply the fraction dissolved
that determines bioavailability, but also the rats of dissolu-
tion, which has physiological and geochemical influences. In
and of itself, bioaccessability is not a direct measure of the
movement of a substance across a biological membrane (i.e.,
absorption or bioavailability). The relationship of
bioaccessability to bioavailability is ancillary and the former
need not be known in order to measure the latter.
Produced by the Technical Review Workgroup for Lead (TRW) /
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However, bioaccessability (i.e., solubility) may serve as a sur-
rogate for bioavailability if certain conditions are met (see
Methods and Issues to Consider when Determining Site-Spe-
cific Bioavailability of Soil-Borne Lead).
As previously mentioned, lead absorption is believed to occur
by both active and passive mechanisms. Although the precise
subcellular processes involved in lead absorption are not en-
tirely known, active/passive absorption processes (depend-
ing on dose) can impart a curvilinear shape to a graph of dose
v.? blood lead concentration. The potential impact of active
and passive absorption processes on the determination of rela-
tive bioavailability is discussed in a latter section (Methods
and Issues to Consider when Determining Site-Specific
Bioavailabilily of Soil-Borne Lead).
When to Consider Adjustments in Bioavailability
As stated in the Introduction, the bioavailability of soil-borne
lead is influenced by numerous characteristics of the soil-lead
matrix. Particle size has been demonstrated to effect soil-lead
bioavailability (Steele et al., 1990). Although a strong quan-
titative relationship between particle size and bioavailability has
not been established, an understanding of particle size distri-
bution in a soil-lead source may provide qualitative information
on the potential bioavailability of the source material. Perhaps
more importantly, available data (Henningsen el a!., 1998)
indicate that lead speciation can have a significant effect on
bioavailability.
Currently, in vivo bioassays are the only way to quantitatively
measure and adjust default bioavailability to fit site soils.
However, validation studies are in progress which show prom-
ise for in vitro tests which may be correlated to the in vivo
results. Such a test would have obvious and much needed
advantages of speed, affordability, simplicity, and higher
throughput. Until such tests are sufficiently validated with in
vivo data, the use of in vitro bioaccessibility results are deemed
by EPA to represent insufficient evidence for quantitative ad-
justment of bioavailability. The reason for this position is that
small changes in in vitro assays, such as pH, time, tempera-
ture, volume, other solutes, and agitation regimes, can have
relatively large impacts on results of lead solubility. Until vali-
dation is confirmed, the use of a simpler, faster, and cheaper
lab benchtop test will not, in and of itself, be judged an ad-
equate surrogate for measuring bioavailability.
Results of tests by EPA using animal models have shown a
general pattern of relative bioavailability for certain lead salts.
While lead speciation is not the sole factor influencing
bioavailability, these patterns can, nonetheless, be used to
compare a site's form of soil lead to explore differences in
bioavailability relative to the defaults. If the lead speciation pro-
file suggests a bioavailability estimate substantially different
from the IEUBK mode! default, then the costs and benefits of
performing supporting animal tests for now, and possibly of in
vitro tests after validation, can be considered for quantitative
measures of bioavailability, and adjustments for a specific site.
Furthermore, qualitative estimates of relative bioavailability
can be made in the uncertainty section of a risk assessment.
General patterns of relative bioavailability determined by EPA
Region 8 studies of 20 soil lead samples (Henningsen et al.,
1998), compared to the default soil relative bioavailability of 60
percent, are shown as groups in the following table:
Potentially
Lower
Bioavailability
(RBA<25%)
Galena (PbS)
Angfcsite (PbSO4)
Pb (M) Oxides
Pb Fe (M)
Sulfates
Native Pb
Intermediate
Bioavailability
(RB A =25% to 75%)
Pb Oxide
PbFe(M) Oxides
Pb Phosphate
Slags
Potentially
Higher
Bioavailability
(RBA>75%)
Cerrusile
(PbCO,)
PbMn(M)
Oxides
Pb = lead, S = sulfur, M = metals, Fe = iron, Mn = manganese
Results of well-conducted blood lead studies can infer relatively
low bioavailability of lead in soil. Such findings would not
support a quantitative adjustment of bioavailability, but could
assist in identifying soils for further study and/or support a
qualitative adjustment in the risk characterization section of a
risk assessment.
Methods and Issues to Consider When Determining
Site-Specific Bioavailability of Soil-Borne Lead
Ethics aside, in a hypothetical setting the ideal method for
making a bioavailability adjustment for soil-borne lead in the
IEUBK model would be to dose a large group of young children
with soil-borne lead and compare the area-under-the-concen-
tration/time curve (AUC) with the AUC of the same or similar
group which received an equal lead dose by intravenous ad-
ministration. This is the conventional pharmacological and
toxicological method for measuring absolute bioavailability.
Realistically, issues of ethics, cost, and implementation are im-
portant determinants of study design. Consequently, an alter-
nate approach is to measure soil-lead bioavailability relative to
a "standardized reference material" (see Definitions section).
Determination of relative bioavailability needs to consider the
experimental evidence suggesting that gastrointestinal lead ab-
sorption follows first-order saturation kinetics. An example is
presented to illustrate that relative bioavailability, as estimated
from experimental studies, can depend strongly on the response
levels at which comparisons are made. The approach used to
estimate relative bioavailability is to compare doses of lead (in
different forms) that, upon ingestion by an experimental animal,
produce equal levels of biological response (in this example,
blood lead concentrations). The curves in the Figure illustrate
relationships that may be fit to experimental data on the relation-
ship between the ingested dose of lead and resulting blood lead
measures. The two curves are of the Michaelis-Menten form
Produced by the Technical Review Workgroup for Lead (TRW)
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(Equation 1} with vmax =30, km = 1 in the soluble lead relation-
ship and vmax = 10, km = 0.4 for the soil lead relationship.
Absorption rale
vmax * dose
km + dose
Equation 1
where
vmax - maximum rate at which an enzyme can
function.
km - concentration of substrate that produces
50% maximum velocity of the enzyme.
This example is hypothetical, in that the curves shown are for
purposes of illustration and are not intended to represent a
specific data set. However, similar models, using Michaelis-
Menten form equations, have been presented to EPA as models
of bioavailability data from rodent studies conducted with soils
from Superfund sites.
To estimate relative bioavailability in this example, a reference
blood lead concentration is selected (5 ug/dL). The dose lev-
els of soluble lead and lead in soil, respectively, at which this
blood concentration is produced are then estimated. As illus-
trated in the Figure, a dose of 0.2 mg/kg/d of soluble lead is
associated with a blood lead level of 5 ug/dL, while a dose of
lead in soil of 0.4 mg/kg/d is required to achieve this same
level. The relative bioavailability is estimated to be 0.5 or 50
percent based on the ratio of these doses (0.2/0.4).
However, in this example, where the soluble lead graph and the
soil lead graph show different curvatures (specifically result-
ing from the different km values in the example), the estimated
relative bioavailability depends on blood lead level at which
the comparisons are made (see figure at right).
Hypothetical response curve for lead uptake study
Example showing different stages of response for soil lead and soluble lead
t, if-
Lead In soil
T [ I I I
0.0 0.5 1.0 1.5 2.0
Ingested lead, mg/kg/d
Note that at low doses the relative bioavailability of the two
materials is similar, while at high doses the relative bioavailability
of lead in soil is estimated to be low compared with soluble
lead. A variety of different mechanistic factors may affect the
bioavailability of lead administered at high doses. In experi-
mental studies of bioavailability, substantial amounts of soil
may be administered to the experimental animals, and the pres-
ence of these high quantities of soil in the diet may affect the
bioavailability of lead. Such effects may be due to alterations
to the chemical environment of the GI tract.
Blood bad
concentration for
calculating relative
bioavailablity foig/dL)
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8.00
Dose of soluble lead
to achieve this
concentration
(ma/kg/day)
0.03
0.07
0.11
0.15
0.20
0.25
0.30
0.36
Dose of soil lead
to achieve this
concentration
(rng/kg/day)
0.04
0.10
0.17
0.27
0.40
0.60
0.93
1.60
Relative
bioavailability
0.78
0.71
0.65
0.58
0.50
0.42
0.33
0.23
Produced by the Technical Review Workgroup for Lead (TRW)
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For example, the presence of substantial quantities of soil may
provide additional binding sites for lead, reducing the likeli-
hood that any lead which becomes solubilized will remain in
solution and be absorbed. Due to the potential for high doses
of either soil or lead itself to affect (reduce) absorption, experi-
mental bioavailabiiity studies need to be performed at low
enough doses to provide a reasonable comparison with the
quantities of soil and lead that humans are likely to ingest.
Where experimental limitations necessitate that the quantities
of soil or lead administered substantially exceed the expected
human doses (on a body weight basis), it should be recog-
nized that an extrapolation to lower doses may be appropriate.
This extrapolation step may take the form of an explicit math-
ematical treatment of the data (and as such would need to
address the uncertainty in the predictions at low dose) or it
may involve a more qualitative demonstration that under the
particular experimental conditions utilized, the estimated
bioavailabiiity is not highly sensitive to the lead dosage used
for comparison.
Site Soil Homogeneneity for Sample Collection and
Preparation
Soil samples that are tested for in vivo bioavailabiiity or in
vitro bioaccessibilhy should be obtained from areas that are
reasonably similar (i.e., similar geophysical and chemical prop-
erties of lead in soil). The top 2 inches of surface soil from
residential yards should be representatively sampled and
composited for testing. It is critical to sieve soil samples to
<250 um (60 mesh) to more closely represent the size of soil
particles that would be expected to adhere to children's hands.
An extremely useful tool for geophysical-chemical character-
ization of lead in soil is the electron microprobe (Medlin, 1997).
Soil samples which are characterized or tested for bioavailabiiity
must retain their integrity, including chain of custody docu-
mentation, and proper mixing that provides a uniform subsample
without physically degrading the soil particles.
Appropriate Animal Model
Because of the difficulties in gathering data on oral absorption
of lead in children, there is no validated absolute model for
experimental uses in measuring bioavailabiiity. Each candi-
date animal model is expected to respond uniquely to absolute
lead absorption (i.e., oral uptake vs. intravenous dosing), com-
pared to children, because of differences in physiology, diet,
behavior, and development. However, it is possible to use a
similar mammalian gastrointestinal system to measure rela-
tive absorption in comparison to the uptake of a soluble lead
reference material (e.g., lead acetate). This is the concept
underlying the juvenile swine model (Weis et al, 1994) which
has further advantages of permitting sequential blood sam-
pling and responding to doses similar to those experienced
by children. Further details on the appropriate design as-
pects of such studies can be obtained from Weis et al, 1994;
Casteel
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Henningsen, G., Weis, C., Casteel, S., Brown, L., Hoffman, E.,
Brattin, W,, Drexler, J., Christensen, S. 1998. Differential
Bioavailability of Lead Mixtures from twenty different soil
matrices at Superfund mining sites. Abstract. Toxicological
Sciences. 42(l-s).
Ruby M., et at. 1996. Estimation of Lead and Arsenic
Bioavailability using a Physiologically-Based Extraction
Test. EnvironSci TecAno/30: 422-430.
Steele, M., Beck, B., Murphy, B. 1990. Assessing the contribu-
tion from lead in mining wastes to blood lead. Reg. Toxicol,
Pharm. 11:158-190.
U.S.EPA. 1994. Guidance Manual for the Integrated Exposure
Uptake Biokinetic Model for Lead in Children. U.S. Environ-
mental Protection Agency, EPA/540/R-93/081, PB93-963510.
Weis, C., Poppenga, R., Henningsen, G., Thanker, B., Harpstead,
T., Jolly, R. 1994. Lead Absorption from Mine Waste in an
Immature-Swine Model. Presented at the 33^ Annual Meet-
ing of the Society of Toxicology, Dallas, TX, March 1994.
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