67, 17–31 (2002)
Copyright © 2002 by the Society of Toxicology
TOXICOLOGICAL SCIENCES
PBPK Modeling of the Percutaneous Absorption of Perchloroethylene
from a Soil Matrix in Rats and Humans
Torka S. Poet,* ,1 Karl K. Weitz,* Richard A. Gies,* Jeffrey A. Edwards,* Karla D. Thrall,* Richard A. Corley,*
Hanafi Tanojo,† Xiaoying Hui,† Howard I. Maibach,† and Ronald C. Wester†
*Battelle, Pacific Northwest Division, P.O. Box 999, Richland, Washington 99352; and †Department of Dermatology,
P.O. Box 0989, University of California, San Francisco, California 94143
Received August 17, 2001; accepted October 30, 2001
degreaser. Chronic inhalation exposures to PCE result in liver
tumors in male mice and renal tumors in male rats (National
Cancer Institute, 1986). PCE is poorly metabolized, primarily
through CYP450-mediated oxidation to trichloracetic acid and
trichloroethanol, and most of the absorbed dose of PCE is
excreted unchanged in the exhaled air (Monster and Houtkooper, 1979). PCE is poorly water soluble and has a vapor
pressure of 14 mmHg. Since PCE is volatile, it is available for
absorption by inhalation and by percutaneous absorption.
Environmental contaminants, such as PCE, have the potential
to collect in soil; therefore, the assessment of dermal absorption
from soil exposures is needed for a complete toxicokinetic exposure assessment (Sedman, 1989). PCE leaks into soil from storage
tanks and can be deposited from contaminated air during rain.
PCE is more soluble in soil than water and volatilization rates
from soil are lower than from water. The EPA National Priorities
List of hazardous waste sites identified PCE in at least 771 of 1430
sites (HAZDAT, 1996). PCE is 31st on the 1999 CERCLA list of
priority hazardous substances based on frequency of detection
(detection limit of 1 to 5 ␮g/kg) in the environment and potential
for human exposure (ATSDR, 2000).
A noninvasive real-time mass spectrometric (MS/MS)
breath analysis technique was employed to determine the circulating levels of PCE following dermal absorption. The percutaneous absorption of PCE from soil in rats and human
volunteers and a physiologically based pharmacokinetic
(PBPK) model that includes dermal absorption parameters to
quantify the bioavailability of PCE were developed. In dermal
absorption studies, bioavailability is often expressed either as
an absolute amount or percentage of the dose absorbed. The
percentage of the applied dose that is absorbed is dependent
upon exposure conditions, such as length of exposure and
exposure concentration, and is not useful in extrapolating beyond the experimental conditions. Of greater utility is the
calculation of the percutaneous permeability coefficient (K P),
which can be used in validated kinetic models to extrapolate
beyond the experimental conditions.
A PBPK model was used to estimate the K P for dermal
absorption of PCE in rats and humans. Equations for dermal
Perchloroethylene (PCE) is a widely used volatile organic chemical. Exposures to PCE are primarily through inhalation and dermal
contact. The dermal absorption of PCE from a soil matrix was
compared in rats and humans using real-time MS/MS exhaled breath
technology and physiologically based pharmacokinetic (PBPK) modeling. Studies with rats were performed to compare the effects of
loading volume, concentration, and occlusion. In rats, the percutaneous permeability coefficient (K P) for PCE was 0.102 ⴞ 0.017, and was
independent of loading volume, concentration, or occlusion. Exhaled
breath concentrations peaked within 1 h in nonoccluded exposures,
but were maintained over the 5 h exposure period when the system
was occluded. Three human volunteers submerged a hand in a container of PCE-laden soil for 2 h and their exhaled breath was continually monitored during and for 2.5 h following exposure. The
absorption and elimination kinetics of PCE were slower in these
subjects than initially predicted based upon the PBPK model developed from rat dermal kinetic data. The resulting K P for humans was
over 100-fold lower than for the rat utilizing a single, well-stirred
dermal compartment. Therefore, two additional PBPK skin compartment models were evaluated: a parallel model to simulate follicular
uptake and a layered model to portray a stratum corneum barrier.
The parallel dual dermal compartment model was not capable of
describing the exhaled breath kinetics, whereas the layered model
substantially improved the fit of the model to the complex kinetics of
dermal absorption through the hand. In real-world situations, percutaneous absorption of PCE is likely to be minimal.
Key Words: human; PBPK modeling; perchloroethylene; rat;
soil matrix.
Perchloroethylene (tetrachloroethylene: PCE) is widely used
as a solvent in the dry cleaning industry and as a metal
This research was supported in full under Cooperative Agreement DEFG07-97ER62509, Environmental Management Science Program, Office of
Science and Technology, Office of Environmental Management, United States
Department of Energy (DOE). However, any opinions, findings, conclusions,
or recommendations expressed herein are those of the authors and do not
necessarily reflect the views of the DOE.
1
To whom correspondence should be addressed at Chemical Dosimetry,
Battelle, Pacific Northwest Division, P.O. Box 999 MSIN P7-59, Richland,
WA 99352. Fax: (509) 376-9064. E-mail: [email protected].
17
18
POET ET AL.
TABLE 1
Experimental Design
Species
n
Exposure type
Target soil
weight
Target PCE
conc. (g/kg)
Exposure
duration (h)
Media
analyzed (h)
Charcoal
trap
Rat
Rat
Rat
Rat
Rat
Rat
Rat
Human
3
3
3
3
3
5
5
3
Nonoccluded
Nonoccluded
Nonoccluded
Nonoccluded
Nonoccluded
Occluded
Occluded
Hand immersion
0.5 g
0.5 g
0.5 g
5.0 g
5.0 g
5.0 g
5.0 g
4 kg
5
15
50
15
50
15
50
30
5
5
5
5
5
5
5
2
0 and 5
0 and 5
0 and 5
0 and 5
0 and 5
0 and 5
0 and 5
0, 0.5, 1, 1.5, 2
Yes
Yes
Yes
Yes
Yes
No
No
No
Note. Actual soil weight and concentrations were measured and are given in Table 5.
absorption in the PBPK model were based upon Fick’s first law
of diffusion, as described by Jepson and McDougal (1997).
Three different models that varied only in the description of the
skin compartment were compared. The simplest model included a single homogenous dermal compartment. The other
two models included two dermal compartments. The first,
parallel model employed a compartment that was designed to
mimic a follicular route of absorption and the remaining composite dermal compartment; the second, multilayered compartment divided the skin into a stratum corneum (sc) barrier and
the remaining viable cutaneous tissue.
A number of different researchers have incorporated Fick’s
law equation into models to determine the transdermal flux
under nonsteady state absorption conditions (Corley et al.,
2000; Jepson and McDougal, 1997; McDougal et al., 1986;
Poet et al., 2000b). Integrating Fick’s law into a PBPK model
allows for an instantaneous estimation of K P and exposure
concentration without requiring that measurements be taken at
steady state or from an infinite source (Poet et al., 2000a). The
K P should be consistent regardless of exposure concentration
and surface area for any given exposure site and chemical, but
it can vary between exposure sites. Unless a mathematical
model is used to account for changing exposure concentration,
the calculation of flux or permeability coefficient must be
assessed at steady state (Poet et al., 2000a).
The intent of the research described in this article was to
describe the absorption of PCE in rats and humans in a manner
that will be applicable to risk assessment. The validity of
different model descriptions of the dermal compartment are
also compared. Like any biokinetic model, these models must
be descriptive enough to obtain reasonable parameters while
still being simple enough to have utility. The development of a
model that adequately describes the data is integral to exposure
assessment.
MATERIALS AND METHODS
Animals and chemicals. Adult male F344 rats (200 –260 g) were obtained
from Charles River Inc. (Raleigh, NC). Prior to use, animals were housed in
solid-bottom cages with hardwood chips, and were acclimated in a humidityand temperature-controlled room with a 12-h light/dark cycle. Rodent feed
(Purina rodent chow) and water were provided ad libitum.
HPLC grade (99.9% pure) PCE (CAS #127-18-4) and all other chemicals
(reagent grade or better) were obtained from Sigma Chemical Co. (St. Louis,
MO).
Dermal Exposures in Rats
Application techniques. Dermal exposures to PCE were conducted in a
soil matrix. The soil sample was collected in Yolo County, California and was
prepared by passing it through a 40-mesh sieve and retaining it on an 80-mesh
sieve. The soil consisted of 30% sand, 18% clay, 52% silt, and it had an
organic content of 1.3% and a pH of 6.8. This soil sample has been used in two
previous studies involving the dermal absorption of methyl chloroform and
trichloroethylene (Poet et al., 2000a,b). Male F344 rats were anesthetized
using a ketamine/xylazine mixture and the hair on the lower back clipper was
shaved the day prior to exposure. Any rats that showed skin irritation or nicks
were excluded from the study. Two soil loading volumes were compared for
nonoccluded exposures. The low volume consisted of a target of 0.5 g of soil
over 8 cm 2 of skin (0.0625 g/cm 2), and the high volume consisted of 5 g of soil
over 5 cm 2 (1 g/cm 2). Occluded exposures employed 1 g/cm 2. PCE was mixed
with the soil the evening before exposures, sealed tightly with minimal
headspace, and mixed overnight in a rotating mixer. The different exposure
conditions are outlined in Table 1.
A system that has been previously described was used for the low volume
exposures (Poet et al., 2000a,b). Target concentrations of 5, 15, or 50 g/kg soil
were placed inside a ring of DuoDerm威 (Bristol-Myers Squibb, Princeton, NJ)
that was attached to the clipped area on the back of the rats, and covered with
a transparent dressing (Bioclusive威, Johnson & Johnson, Arlington, TX) that
allowed free passage of vapor. This was covered with an aerated weighboat
and a muslin patch containing activated charcoal. The entire patch system was
secured with self-adherent wrap. Preliminary studies were conducted to verify
that all volatilizing PCE was trapped by the charcoal. The patch system was
initially set up on the surface of a glass plate and placed in the exposure
chamber. No PCE was detected using the MS/MS system when the charcoal
patch was in place. After removing the charcoal patch, chamber PCE concentrations rose quickly. In additional studies, rats were exposed to the highest
concentration (50 g/kg), and after 30 min rats were removed from the chamber,
humanely sacrificed, and replaced in the chamber from which PCE had been
flushed out. Peak chamber concentrations of ⬃3,500 ppb were reduced to
nondetectable after the rats were sacrificed.
A glass cell was used for the high volume exposures. Target concentrations
of 15 or 50 mg/kg were placed inside a 2.5-cm diameter hand-blown glass cell
(O.Z. Glass Co., Pinole, CA). The 2.5 cm deep cell was partitioned into two
compartments by a vapor-permeable ceramic frit. The lower compartment
19
DERMAL ABSORPTION OF PERCHLOROETHYLENE
TABLE 2
Human Subjects
Surface area (cm 2) a
ID #
Gender
Age
Race
BW (kg)
Height (cm)
% Body fat b
Total body
Exposed hand
1
2
3
F
F
M
30
47
66
Black
Cauc.
Cauc.
63.5
70.3
79.4
162.3
162.3
177.5
34.8
32.0
27.4
17065
17987
19881
443.7
467.7
516.9
Note. The study participants were African American (Black) or Caucasian (Cauc).
a
The total body surface area was estimated by the following equation from Gehan and George (1970) as reported in U.S. EPA, 1996: SA (cm2) ⫽ Weight
(kg) 0.517 ⫻ Height (cm) 0.417 ⫻ 239. The exposed surface area of the hand for each volunteer was calculated as 2.6% of total surface area.
b
Body fat percentages were measured for each volunteer using a hand-held near-infrared body fat analyzer (Futrex威, Gaithersburg, MD).
contained the PCE-laden soil and the upper compartment contained activated
charcoal to trap volatilized chemical. The glass cell only had space for the
addition of 2 g of charcoal above the ceramic frit. Preliminary studies (as
outlined above for the DuoDerm system) revealed that the 2 g of charcoal was
not sufficient to block the escape of PCE into the chamber, so an additional
charcoal patch was placed above the glass exposure cell. Following the
addition of the charcoal patch, no PCE escaped into the chamber.
For occluded soil samples, target concentrations of 15 and 50 g/kg PCE in
5 g of soil were placed in a hand-blown glass cell (O.Z. Glass Co., Pinole, CA).
The occluded cell had the same dimensions as the lower compartment of the
nonoccluded cell and was completely enclosed. The glass cells were attached
to the clipped area using a cyanoacrylate adhesive.
Dermal exposures. To quantify total absorbed dose, the exact weight of
administered soil was recorded and samples of exposure soil, the amount
remaining at the end of the exposure, and the amount volatilized to the charcoal
were analyzed using gas chromatography. Soil samples were analyzed using a
Headspace Autosampler (Perkin-Elmer 40XL) linked to a Hewlett-Packard
5890 Series II GC (Hewlett-Packard, Avondale, PA). A Restek Rt x-Volatiles
column (30 m ⫻ 0.32 mm ⫻ 1.5 ␮m) cross-bonded with phenylmethyl
polysiloxane was used. The oven temperature was set at 90°C and the injection
and FID detector temperatures were set at 120°C. Helium was the carrier gas
at 8 psi. Charcoal from the nonoccluded patch system was extracted using
toluene and PCE concentrations measured using similar GC conditions, except
2 ␮l of toluene was injected (splitless) onto the GC.
Immediately following dermal application, rats were individually placed in
small off-gassing chambers as described by Gargas (1990) and a Teledyne
Discovery II MS/MS equipped with an atmospheric sampling glow discharge
ionization source sampled from the off-gassing chamber (representing exhalation from the animal) approximately every 5 s as described previously (Poet
et al., 2000b). Breathing air was continually supplied to the rat through the lid
of the off-gassing chamber at a measured rate (200 ml/min). Airflow rates were
measured using flow meters from Sierra Instruments (Carmel Valley, CA). The
ASGDI source derived reagent ions directly from the volatile chemicals in the
sampled air. An electric potential was established by applying 400 V between
two plates. Ions were then focused onto the MS/MS trap. Helium was used as
a buffer and collision gas. The MS intensity data was converted to concentration (ppb) through the use of external standards prepared in Tedlar威 bags. A
standard curve was generated each day of experimentation. PCE was quantified
by selective ion monitoring of the most abundant product m/z ratios 160 –164.
Dermal Exposures in Human Subjects
Study participants. Two healthy female volunteers and one healthy male
volunteer participated in the study; demographic data including gender, race,
age, body weight, and height are given in Table 2. The studies were conducted
under approval from both the University of California at San Francisco
(Committee on Human Research) Institutional Review Board (IRB) and the
Pacific Northwest National Laboratory IRB in compliance with multiple
project assurance number DOE.MPA.PNNL96-2000. Written consent was
obtained from each subject prior to participation. Subjects reported no chronic
conditions; no significant cardiovascular, hepatic, central nervous system,
renal, hematological, or gastrointestinal diseases; and no dermatological problems. Since PCE is highly lipophilic and PCE pharmacokinetics are sensitive
to the amount of body fat, the percent body fat for each subject was determined
using a handheld near-infrared body fat analyzer (Futrex威, Gaithersburg, MD).
Dermal exposure conditions. Human volunteers were dermally exposed
to PCE by immersing a hand in 4 kg of soil within a covered container. The
surface area for human exposures was approximately 2.6% of the total body
surface area. For comparison, the rats were exposed over approximately 2 and
2.7% of the total body surface area for high and low volume exposures,
respectively. Surface areas for each exposure are given in Table 2. The target
PCE concentration was 30 g/kg of soil. PCE-laden soil was prepared in a
separate room and covered with plastic wrap prior to use. Soil samples were
collected from the container at different locations near the hand (n ⫽ 3 per time
point) before and during the exposure at 0, 0.5, 1, 1.5, and 2 h for GC analysis
of PCE concentrations in the exposure media over time.
Subjects were provided clean breathing air via a facemask with a two-way
nonrebreathing valve to prevent potential inhalation exposures. The exhaled
breath was passed through a heated mixing chamber (1.3 l volume) from which
the MS/MS drew a sample for analysis approximately every 5 s, as described
previously (Poet et al., 2000b). Excess exhaled air was vented from the mixing
chamber to a hood with negligible flow restriction via a large borehole exit
tube. Background exhaled breath measurements were taken for 2 min before
each exposure.
PBPK Model
The overall model structure was based on the flow-limited model for volatile
chemicals initially described by Ramsey and Andersen (1984), as it was
developed specifically for PCE by Reitz et al. (1996). The initial model
structure was comprised of five compartments: fat, liver, rapidly perfused
tissues, slowly perfused tissues, and skin linked by the systemic circulation. In
a standard blood flow-limited model, the blood is assumed to equilibrate with
exhaled breath. For rat exposures, an equation was also added to calculate the
amount of chemical in the off-gassing chamber in terms of input from the
exhaled breath and removal from the chamber either by rebreathing or as it was
drawn into the MS/MS, as described previously (Poet et al., 2000a).
Chemical-specific partition coefficients and metabolism rates were obtained
from a previously established PBPK model for PCE (Reitz et al., 1996). In this
study, human blood/air partition coefficients were determined experimentally,
and human tissue/blood partition coefficients were estimated by dividing the
rat tissue/air by the human blood/air partition coefficient (Reitz et al., 1996).
Reitz et al. (1996) also conducted a sensitivity analysis that showed that the
blood/air partition coefficient had the most impact on the model estimation of
20
POET ET AL.
TABLE 3
Parameters Used in the PBPK Model for Rats and Humans
Body wt. (kg)
Tissue volume (% body weight)
Liver
Rapidly perfused
Slowly perfused
Fat
Skin a
Flows (l/h)
Alveolar ventilation
Cardiac output
Percentage of cardiac output
Liver
Richly perfused
Poorly perfused
Fat
Skin a
Biochemical constants
Vmax (mg/h)
Km (mg/l)
Rat
Human
0.198–0.258
63.5–79.4
4.0
5.0
75.0
7.0
10.0
3.1
3.7
61.1
27.4–34.8
3.5
4.5–5.8
4.5–5.8
300–350
300–350
24.0
53.0
19.0
5.0
5.8
0.325
5.62
25.0
49.0
19.0
9.0
5.8
32.9
4.66
Note. Parameters except those relating to the skin compartment were obtained from Reitz et al. (1996). Parameters relating to the skin compartment
were obtained from Jepson and McDougal (1997) and Brown et al. (1997).
a
Values are for total skin. The skin compartment in the PBPK model was
comprised solely of the area of the skin under the exposure cell. The remainder
of the skin was included in the slowly perfused compartment.
amount of PCE metabolized. However, this was for inhalation exposures, and
blood/air partitioning would affect the transfer of PCE from inhaled air. Soil
partition coefficients were determined using the method of Gargas et al.
(1989), with modifications. Quadruplicate samples of 1 g soil were placed in
sealed vials and incubated for 2 h with PCE at 32°C (the approximate
temperature at the skin surface) with vigorous shaking in a heating block. PCE
was added as vapor from Tedlar威 bags containing approximately 10,000 ppm
PCE. The amount of PCE in the headspace of vials containing soil was
compared to empty reference vials containing PCE only. The soil:skin partition
coefficient was calculated by dividing the soil:air partition coefficient by the
skin:air partition coefficient. The skin:air partition coefficient was determined
by Mattie et al. (1994) in male F344 rats. Parameters for the rat and human
PBPK models are given in Tables 3 and 4.
Dermal compartments. The uptake of PCE through the skin was initially
described using a single well-stirred compartment that incorporated Fick’s law
of diffusion into the PBPK model. Equations to describe the rate of change of
chemical concentration across the skin (Jepson and McDougal, 1997) and
evaporation to patch components were:
dA sk
⫽
dt
冉
K p ⫻ SA
1000
冊 冉
⫻
C sk
C surf ⫺
P skm
冊
⫹ Qsk ⫻ 共C art ⫺ C ven 兲
dA coal
⫽ K loss 共C surf ⫻ V s 兲
dt
(1)
(2)
where A sk is the amount of PCE in the skin (mg), C sk is the PCE concentration
in the skin (mg/l), SA is the surface area exposed (cm 2), C surf is the concentration of PCE at the skin surface (mg/l), P skm is the skin:soil partition
coefficient (unitless), C art is the arterial blood PCE concentration (mg/l), Q sk is
the blood flow to the skin (l/h), C ven is the venous blood PCE concentration
(mg/l), A coal is the concentration in the charcoal from evaporative loss (mg/l),
V s represents the volume of the dosing solution (kg), and K loss is the rate of
evaporation loss from the media (/h).
K loss and K p were estimated for each individual animal or human subject by
a least-squares fit of the model to the exhaled breath, charcoal, and soil data
(where appropriate) using the Simusolv™ optimization subroutine (Dow
Chemical Co.). For human subjects, the observed delay before the appearance
of PCE in exhaled breath was estimated by visually extrapolating the appearance of PCE in the exhaled breath back to the x-axis. Thus, each parameter was
not only optimized to the exhaled breath concentrations, but also to the
chemical concentrations in exposure media and escaping into the charcoal
patches.
When it became apparent that the single homogenous dermal compartment
model would not adequately describe the human exhaled breath data, two
additional model structures with dual dermal compartments were established.
These two dermal models consisted of a parallel dermal compartment, and a
dual layered compartment structure, as described previously by Bookout et al.
(1996, 1997). The parallel model is predicated on the theory that the follicles
in the skin serve as a shunt for chemical absorption (Kao et al., 1988).
Therefore, the skin is divided into a follicle compartment that contains the
follicles and sweat glands and a composite dermal compartment that contains
the remaining layers of the skin. The area of the follicular compartment was
assumed to be 1/100th of the rest of the skin and the depth was assumed to be
388/560th that of the entire skin (Grabau et al., 1995).
The permeability coefficients through the two parallel compartments are
related to the K P through the skin as a whole such that the combined permeability coefficients must match the single compartment coefficient following an
area correction, as described in Bookout et al. (1997). Therefore, the composite
K P was first optimized for the one compartment model, then fixed in the two
compartment model, and the two new K Ps were optimized to the data but
constrained by their relationship to the composite K P. The equations used in the
PBPK model to describe the dual parallel compartments and the relationship
between the composite K P and the K Ps for the dual compartments are given in
the Appendix.
The second alternative dermal model described by Bookout et al. (1996)
separates the stratum corneum from the underlying viable cutaneous tissue.
The barrier function of the skin primarily resides in the stratum corneum
(Scheuplein, 1978). In this dual layered model, the dermis and viable epidermis
are combined into the viable cutaneous compartment. Blood exchange occurs
only in the viable cutaneous compartment, and the stratum corneum imparts a
barrier between the surface exposure and the viable cutaneous tissue. The
stratum corneum compartment was assumed to be 11/560th of the total skin
volume (Bookout et al., 1996).
As for the parallel model, the total permeability coefficient for both compartments is related to the permeability coefficient for a single well-stirred
TABLE 4
Tissue Partition Coefficients for Rats and Humans
Tissue
Rat
Human
Blood/air
Liver/air
Richly perfused/air
Poorly perfused/air
Fat/air
Skin/air a
Soil/air b
18.85
70.3
70.3
20.0
1638
41.5
108.5
10.3
60.6
60.6
31.9
1227
41.5
108.5
Note. All partition coefficients except those relating to the skin compartment
were obtained from Reitz et al. (1996).
a
The skin/air partition coefficient was obtained from Mattie et al. (1994).
b
The soil partition coefficient was determined as described in Materials and
Methods.
21
DERMAL ABSORPTION OF PERCHLOROETHYLENE
Data Analysis
Data are presented as the mean ⫾ SD of n ⫽ 3–5 exposures. A Student’s
t-test or an ANOVA followed by the Student-Newman-Keuls test was used for
statistical comparisons of parameters for different exposure conditions, where
appropriate. Statistical significance was considered at p ⬍ 0.01.
Dermal compartment evaluation. As the model becomes progressively
complicated with the increase in the number of dermal compartments, there is
a possibility that more parameters than necessary to explain the data have been
added. In order to discriminate the goodness of fit of the dual compartment
models versus the one-compartment model, likelihood ratio tests were performed. Models are considered to be nested when the basic model structures
are identical except for the addition of complexity, such as the added dermal
compartments. In this case, the model with the single well-stirred dermal
compartment was nested within the dual dermal compartment models. Under
these conditions, the likelihood ratio can be used to statistically compare the
relative ability of the models to describe the same data, as described in the
“Reference Guide for Simusolv” (Steiner et al., 1990). The hypothesis that one
model is better than another is calculated using the likelihood functions
evaluated at the maximum likelihood estimates. Since the parameters are
optimized in the model using the maximum log likelihood function (LLF), the
resultant LLF is used for the statistical comparison of the models. The equation
states that two times the log of the likelihood ratio follows a chi-square (␹ 2)
distribution with r degrees of freedom:
⫺ 2log␥ 共x兲 ⬵ ␹ r2
FIG. 1. Schematics of the single homogenous and dual dermal compartments used to describe the uptake of PCE in rats and humans.
(3)
The likelihood ratio test states that if the difference between the maximum
log likelihood functions of the two models is greater than the chi-square
distribution then the model fit has been improved (Devore, 1995; Steiner et al.,
1990). Example calculations of the likelihood ratio analysis are given in the
Results.
RESULTS
compartment. For a layered model, the K Ps for each new compartment are
inversely related to the composite K P for the single compartment (Bookout et
al., 1996). Therefore, the K P first optimized for the one compartment model
was also used as the composite K P for the dual-layered model. A diagrammatic
representation of the two alternative compartment models is given in Figure 1,
and the equations used in the PBPK model to describe each model are given
in the Appendix. Since only the sc compartment is in contact with the media
and only the viable cutaneous compartment is involved with blood exchange,
the partition coefficients for the sc compartment:media (PSC/media) and the
viable cutaneous compartment:blood (PVC/blood) were calculated using the
media/air and blood/air partition coefficients, respectively. The permeability
coefficient for the viable cutaneous compartment (K PVC) was not optimized
directly, but was based on the inverse relationship between the overall K P and
K P for the sc compartment (K P SC), as described by Bookout et al. (1996). The
two new partition coefficients (PSC, PVC) were calculated in the same fashion
as for the parallel model. PSC and K PSC were optimized using the log
likelihood function (LLF) of Simusolv™, and PVC and K PVC were optimized
indirectly using the equation in the Appendix as described by Bookout et al.
(1996).
The inclusion of additional skin compartments increased the number of
variables. The skin:air PCE partition coefficient was measured in whole skin of
male F344 rats (Mattie et al., 1994), but the partition coefficients for the dual
parallel and layered compartments were not measured directly and had to be
optimized in the PBPK model. The partition coefficients for the dual compartments were equal to the partition coefficient measured in whole skin with a
volume correction. Equations used to optimize the partition coefficients for the
dual compartments are given in the Appendix.
Rat Exposures
Target PCE concentrations in soil were 5, 15, and 50 g/kg
soil. The weight of the applied soil was measured and precise
concentrations were determined using GC headspace analysis.
Actual exposure concentrations are given in Table 5. The PCE
mass balance was tracked by determining soil and charcoal
patch PCE concentrations after exposures using GC analysis.
In nonoccluded systems, PCE volatilized from the exposure
cells and 39 to 90% of the initial PCE amount was recovered
in the charcoal patch, depending upon the exposure conditions.
Due to a combination of uptake into the body and volatilization
to the charcoal patch, the amount of PCE remaining in the soil
postexposure (5 h) was less than 1%. Conversely, in occluded
exposures, where volatilization was prevented, 45% of the
original concentration of PCE was recovered in the soil 5 h
after the initiation of exposure, with the remainder being absorbed (Table 6).
The impact of various PBPK model parameters on predicted
exhaled breath concentrations was assessed. Variables pertaining to the skin had the most effect, particularly dermal blood
flow and K P. The rate of loss (K loss) also had an effect on the
peak exhaled breath concentration and rate of decline. The
model fit to literature data was verified, and all parameters
22
POET ET AL.
TABLE 5
Exposure Conditions for Dermal Absorption in Rats
PBPK estimates
Nonoccluded
Low vol., low conc.
Low vol., mid conc.
Low vol., high conc.
Overall avg.
High vol., mid conc.
High vol., high conc.
Overall avg.
Occluded
High vol., mid conc.
High vol., high conc.
Overall avg.
PCE conc. (g/kg)
Soil applied (g)
4.56 ⫾ 0.70 (2.4 mg)
14.8 ⫾ 1.70 (7.6 mg)
51.0 ⫾ 5.09 (26 mg)
0.522 ⫾ 0.125
0.514 ⫾ 0.030
0.509 ⫾ 0.058
15.5 ⫾ 2.02 (76 mg)
52.9 ⫾ 6.13 (267 mg)
4.93 ⫾ 0.58
5.04 ⫾ 0.179
15.5 ⫾ 4.84 (80 mg)
49.9 ⫾ 1.27 (267 mg)
5.17 ⫾ 0.388
5.35 ⫾ 1.34
Skin permeability
coefficient (K P, cm/h)
Evaporative loss rate
(K loss/h)
0.090 ⫾ 0.016
0.088 ⫾ 0.019
0.086 ⫾ 0.014
0.087 ⫾ 0.013
0.112 ⫾ 0.014
0.111 ⫾ 0.012
0.112 ⫾ 0.013
0.818 ⫾ 0.034
0.794 ⫾ 0.052
0.764 ⫾ 0.106
0.791 ⫾ 0.065
0.944 ⫾ 0.083
0.970 ⫾ 0.074
0.953 ⫾ 0.076
0.110 ⫾ 0.018
0.111 ⫾ 0.011
0.111 ⫾ 0.013
NA
NA
NA
Note. PCE exposure concentrations were determined using headspace gas chromatography. The number in parenthesis is the average amount of PCE in the
exposure. K loss is significantly different between low and high volume exposures (p ⬍ 0.05), which was attributed to the use of different exposure systems. NA,
not applicable.
(such as metabolism rate constants and partition coefficients)
other than those relating to dermal absorption were held constant.
PCE exhaled breath levels in rats exposed to PCE in 0.5 g of
soil (low volume exposures) under nonoccluded conditions
peaked in less than 1 h (Fig. 2). The rate of loss (K loss) was
concurrently estimated by fitting to the exhaled breath data, the
concentration remaining in the soil, and the amount volatilizing
to the patch. The parameters K P and K loss were optimized in an
iterative manner until all three endpoints (soil concentration,
charcoal concentration, and exhaled breath profile) were de-
scribed by the model using the LLF function of Simusolv™.
An example of rat chamber concentration data and PBPK
model predictions is given in Figure 2. Model predictions of
charcoal and soil concentrations over time are compared with
actual soil and charcoal concentrations as determined using GC
analysis in Figure 3. The K P and K loss values that resulted in the
best fit to the data for the nonoccluded low volume exposures
were 0.087 ⫾ 0.013 cm/h and 0.791 ⫾ 0.065/h, respectively
(Table 5). Exposures to PCE in 5 g of soil resulted in similar
exhaled breath profiles (Fig. 4). As for the low volume exposures, the peak exhaled breath concentration was reached under
TABLE 6
Recovery of PCE in Rat Exposures
Nonoccluded
Low vol., low dose
Low vol., mid dose
Low vol., high dose
Overall avg.
High vol., mid dose
High vol., high dose
Overall avg.
Occluded
High vol., mid dose
High vol., high dose
Overall avg.
% Absorbed
% Volatilized
% Remaining in soil
Total % recovered
57.0 ⫾ 6.28 (1.4 mg)
54.9 ⫾ 6.13 (4.2 mg)
51.2 ⫾ 5.20 (13.3 mg)
54.4 ⫾ 5.90
11.1 ⫾ 2.15 (8.4 mg)
8.71 ⫾ 0.63 (23 mg)
10.3 ⫾ 2.09
43.9 ⫾ 6.44
39.3 ⫾ 3.41
42.7 ⫾ 5.65
42.0 ⫾ 4.32
87.5 ⫾ 2.77
90.2 ⫾ 0.74
88.4 ⫾ 2.60
0.58 ⫾ 0.064
0.69 ⫾ 0.13
0.44 ⫾ 0.24
0.57 ⫾ 0.18
0.51 ⫾ 0.098
0.54 ⫾ 0.049
0.52 ⫾ 0.082
101.5 ⫾ 13.1
95.0 ⫾ 5.13
94.3 ⫾ 8.11
96.9 ⫾ 8.82
99.1 ⫾ 0.72
99.4 ⫾ 0.16
99.2 ⫾ 0.60
49.2 ⫾ 6.07 (39 mg)
52.9 ⫾ 2.01 (141 mg)
50.9 ⫾ 4.47
NA
NA
NA
45.2 ⫾ 5.75
45.7 ⫾ 8.73
45.4 ⫾ 6.62
94.4 ⫾ 5.27
98.3 ⫾ 8.91
96.4 ⫾ 6.89
Note. The total % of PCE recovered was assessed using a combination of PBPK estimations and GC determinations. The % absorbed was estimated using
the PBPK model. The number in parentheses is the average amount of PCE absorbed. The % volatilized was determined from the amount remaining in the
charcoal patch using GC analysis as described in Materials and Methods. The % remaining in the soil was measured using GC headspace analysis as described
in Materials and Methods. The total % recovered was calculated from % absorbed, % volatilized, and % remaining in soil.
DERMAL ABSORPTION OF PERCHLOROETHYLENE
FIG. 2. Concentration-time plots for PCE exposures of rats to low volume,
nonoccluded soil. Points are exhaled breath data (averaged every 1 min
interval), lines are PBPK simulations of representative rats exposed to target
concentrations of 5 (filled box), 15 (open diamond), and 50 g/kg (filled circle)
in 0.5 g of soil over a 8 cm 2 area of the back.
an hour and steadily declined. The K P was not significantly
different between any of the exposure conditions (Table 5).
When the patch system was occluded, PCE volatilization
was prevented and K loss was therefore set to 0. As a result of
occlusion, the concentration of PCE in the exhaled breath was
maintained over the 5 h exposure period (Fig. 5), rather than
declining after an initial peak as observed for the nonoccluded
exposures where PCE was allowed to evaporate during the
exposures. By factoring in the rate of loss of PCE from the
nonoccluded patch system into the analysis, PBPK estimations
of K P were consistent between occluded and nonoccluded
exposures (Table 5).
FIG. 3. Percentage of PCE in the exposure media (soil) and charcoal
(nonoccluded exposures only) from rat studies. Lines are PBPK estimations,
points are concentrations measured by headspace analysis for percentage in
occluded soil (filled box; n ⫽ 10), low and high volume, nonoccluded soil
(filled diamond; n ⫽ 15), charcoal from high volume exposures (open diamond; n⫽6), and charcoal from low volume exposures (open circle; n ⫽ 9).
Values are percentage calculated for all exposure concentrations ⫾ SD.
23
FIG. 4. Concentration-time plots for PCE exposures in high volume,
nonoccluded soil. Points are exhaled breath data (averaged every 1 min
interval), lines are PBPK simulations of representative rats exposed to target
concentrations of 15 (open diamond) and 50 g/kg (filled circle) in 5 g of soil
over 5 cm 2 of the back.
The percent of each applied dose of PCE absorbed was also
estimated using the PBPK model. At 5 h, the percentage
absorbed was highly dependent on the exposure conditions.
The nonoccluded, low volume and occluded, high volume
exposures resulted in 50% or more of the original dose being
absorbed. The percentage absorbed was not dependent upon
exposure concentration and no significant differences in percent absorbed were noted between exposure doses within an
exposure scenario. The amount absorbed, however, was highly
dependent on exposure volume (Table 6).
The percentage (and amount) absorbed was considerably
higher for the high volume occluded exposure over the nonoccluded exposure. For the occluded exposures, the exhaled
breath concentrations reached a plateau and remained steady
between 1 and 5 h whereas in the nonoccluded exposures, the
FIG. 5. Concentration-time plots for PCE exposures in occluded soil.
Points are exhaled breath data (averaged every 1 min interval), lines are
PBPK simulations of representative rats exposed to target concentrations of
15 (open diamond), and 50 g/kg (filled circle) in 5 g of soil over a 5 cm 2
area of the back.
24
POET ET AL.
Human Exposures
FIG. 6. Soil concentration of PCE over time from human studies. The line
is the PBPK estimation, points are concentrations measured by headspace
analysis of triplicate samples taken from the soil around the hand at the times
indicated (mean ⫾ SD; n ⫽ 3).
exhaled breath concentrations began to decline in less than 1 h
(Figs. 2, 4, and 5). The PBPK model predictions indicated that
the exposure concentration declines to less than 10% of the
original concentration in the low volume exposures and less
than 30% in the high volume exposures by 1 h, about the time
that exhaled breath concentrations peaked (Fig. 3). In the
occluded exposures, the percentage absorbed would have continued to increase for exposures longer than 5 h since 45% of
the original concentration remained in the soil. In contrast,
there was less than 1% of the original concentration of PCE
remaining in either of the nonoccluded exposure systems to be
absorbed at 5 h.
A cyclical pattern in the exhaled breath profile was noted for
most rat exposures. This same pattern was observed previously
for similar exposures to trichloroethylene (Poet et al., 2000a).
These waves in the exhaled breath levels were associated with
patterns of activity where the rats moved around in the chamber. In the model, the pulmonary rate was held constant over
the exposure period, whereas breathing rate would be expected
to change with activity level and cause the variations in exhaled breath levels that were observed.
To track PCE soil concentrations over time, aliquots of soil
were taken from the exposure container every 30 min. The
large exposure volume (4 kg) and surface area of the container
resulted in a slow decline of PCE from the soil as it was
volatilized and absorbed into the body (Fig. 6). The rate of loss
(K loss) was concurrently estimated by fitting to the exhaled
breath data and the concentration remaining in the soil over
time. The parameters (K P and K loss) were optimized in an
iterative manner using the LLF function of Simusolv™ until
both endpoints (soil concentration and exhaled breath profile)
were described by the model. The K loss (rate of volatilization
from the container) that resulted in the best fit of the data was
0.851 ⫾ 0.054/h (Table 7). This Kloss is remarkably similar to
that determined for the rat exposures (0.953 ⫾ 0.076/h: Tables
5 and 7).
The exhaled breath profiles for the three volunteers had
similar lag phases, slow increases, and slow declines of PCE
levels over the 4.5 h that the exhaled breath was monitored. A
lag phase was added to the model to account for the delay in
the appearance of PCE in the exhaled breath. The delay was
estimated by visually extrapolating back to the x-axis from the
initial appearance of PCE in the exhaled breath. A single
well-stirred dermal compartment model prediction of the exhaled breath profile predicted a more rapid initial uptake and
decline than shown by the exhaled breath profile (Fig. 7).
Dual Compartment Models
The slow rise and decline of PCE in exhaled breath indicated
a more complex dermal absorption than the single well-stirred
compartment was able to describe. Therefore, two additional
dermal description models that differed only in the type of dual
dermal compartment were employed. The first, parallel model
included a follicular compartment that represented a small area
of the total skin but could potentially account for enhanced
dermal uptake through the skin appendages. The second, layered model included a stratum corneum layer that would provide a rate-limiting absorption of chemical through the outermost covering of the skin and an underlying viable cutaneous
compartment in equilibrium with the blood supply.
Four new parameters were added to each of the dual com-
TABLE 7
Human PCE Absorption Parameters
Subject
Delay (h)
K P (cm/h)
K loss (/h)
Amount absorbed (mg)
1
2
3
Avg.
0.30
0.40
0.45
0.38 ⫾ 0.08
0.0011
0.0006
0.0010
0.0009 ⫾ 0.0003
0.818
0.913
0.820
0.851 ⫾ 0.054
25.7
11.3
23.6
20.2 ⫾ 7.77
Note. The delay in appearance of PCE in exhaled breath, permeability coefficient (Kp), and % absorbed were predicted using the PBPK model described in
Materials and Methods. K loss, rate of evaporation loss from the media.
DERMAL ABSORPTION OF PERCHLOROETHYLENE
FIG. 7. PBPK model predictions (lines) and exhaled breath data (points
averaged every 1 min) for human subjects exposed to PCE for 2 h by
immersing a hand in 4 kg of soil. The concentration to which the subjects were
exposed is given after the subject number on the figure. The different lines
represent model predictions using the three different dermal compartment
descriptions; single well-stirred, dual parallel, and dual layered.
partment models. To fit the parallel compartment model to the
data, the overall K P was set equal to the one compartment K P
and two new K P parameters were introduced, K PFO for the
follicular compartment and K PCD for the remaining composite
dermal compartment. K PFO and K PCD were related to K P as
described by Bookout et al. (1997) using the equation given in
the Appendix. Likewise, the two new partition coefficients for
the follicular subcompartment (PFO) and the composite dermal
subcompartment (PCD) were related to the partition coefficient
of the homogenous skin, which was measured using a vial
equilibrium technique by Mattie et al. (1994) following a
25
volume correction. PFO and K PFO were optimized using the
LLF of Simusolv, and PCD and K PCD were optimized indirectly using the equation in the Appendix as described by
Bookout et al. (1997). Additional partition coefficients for
follicle/media follicle/blood, and composite/media, composite/
blood were calculated by dividing by the media and blood
partition coefficients, respectively. Since the new permeability
parameters and partition coefficients for the two dual dermal
compartment models were related to the one compartment K P,
the volatilization rate and total amount absorbed were held
constant, equal to the results from the single compartment
model. As expected, the parallel model resulted in a slightly
steeper slope for the initial rise in PCE breath concentrations,
and a concomitantly faster decline. Visually, the apparent fit to
the data was not improved using the parallel model (Fig. 7).
The four new parameters for the layered dermal model were
K PSC and PSC for the sc compartment and K PVC and PVC for
the viable cutaneous compartment. The sc barrier imparted a
slower absorption and elimination from the skin, and a substantially improved fit to the data, by visual inspection (Fig. 7).
The rate-limiting K PSC was much lower than the K PVC (Table
8). K PVC was very similar to the K P estimated for the absorption of PCE in the rats.
Although the fit of the one homogenous skin compartment
model to the F344 rat absorption data was adequate, the fits of
the dual skin compartment models were also evaluated for the
purpose of comparison to the human results. The behavior of
the models was the same in rats as for humans, where the
parallel model predicted a slightly faster absorption and elimination and the layered model resulted in slightly slower kinetics. K PCD (from the parallel model) and K PSC (from the
layered model) were not significantly different than the aggregate K P from the one compartment model (Table 9). A visual
comparison of the fit of the models indicated no improvement
for the dual compartment models over the one compartment
model for any of the exposure scenarios (Fig. 8).
To compare the results of the two dual dermal compartment
models to the single dermal compartment model, a statistical
evaluation was conducted by comparing the log likelihood
ratios. The single dermal compartment model structure is
nested within the dual compartment models. The degrees of
freedom represent the additional parameters in the more complex models. Although the dual parallel and dual layered
compartment models have six and four new permeability coefficients, respectively, the permeability coefficient/blood and
permeability coefficient/media are calculated from the single
optimized coefficient for each new compartment by dividing
by the media/air or blood/air partition coefficients. Therefore,
for either of the dual compartment models, the additional
parameters (and additional degrees of freedom) are four—two
new partition coefficients and two new permeability coefficients.
The chi-square (␹ 2) value for 4 degrees of freedom at a
confidence level of 0.99 is 13.28 and the hypothesis is that the
26
POET ET AL.
TABLE 8
Human Dual Compartment PCE Absorption Parameters
Parallel
Layered
Subject
K PFO
K PCD
PFO
PCD
K PSC
K PVC
PSC
PVC
1
2
3
Avg.
0.0061
0.0091
0.0118
0.009 ⫾ 0.003
0.0010
0.00052
0.00089
0.0008 ⫾ 0.0003
0.114
0.094
0.095
0.01 ⫾ 0.001
41.8
41.8
41.8
41.8 ⫾ 0.0
0.0011
0.00061
0.0010
0.0009 ⫾ 0.0003
0.087
0.077
0.073
0.077 ⫾ 0.010
302
212
190
235 ⫾ 59.4
37.0
36.1
38.5
37.2 ⫾ 1.21
Note. Parallel and layered subcompartment model permeability coefficients and partition coefficients were estimated using the LLF function of Simusolv™.
FO, follicular compartment; CD, composite dermal; SC, stratum corneum; and VC, viable cutaneous.
more complicated models result in better fits to the data. The
comparison of the LLF for the rat single versus parallel model
using Equation 3 gives:
–
2共 – 1850 – – 1991兲 ⫽
–
281
Since –281 ⬍ 9.488 the dual parallel model does not give a
statistically better fit than the single dermal compartment
model. The remaining statistical model comparisons are given
in Table 10. Using the log likelihood ratio, only the dual
layered model for human dermal absorption of PCE fits the null
hypothesis and gives a statistically improved fit to the data; this
verifies the visual assessment of the data.
DISCUSSION
Real time breath analysis, MS monitoring, and similar exposure systems have been used previously in this laboratory to
assess the dermal absorption of methyl chloroform and trichloroethylene (Poet et al., 2000a,b). The PBPK model from Reitz
et al. (1996) was used to estimate the permeability coefficient
of PCE in rats and humans.
The exhaled breath profile for PCE was similar to previous
results with trichloroethylene in rats, but PCE showed a slower
absorption and elimination than trichloroethylene in humans.
PCE is the least volatile and the most lipophilic of the three
chemicals. Also, at 108.5, the soil/air partition coefficient is
13-fold greater than for methyl chloroform and 100-fold
greater than for trichloroethylene. The permeability coefficient
for PCE in rats (0.111 cm/h) was between methyl chloroform
(K P ⫽ 0.15) and TCE (K P ⫽ 0.088), although in practical
terms, the K P for these three chlorinated solvents were very
similar.
Rat dermal exposures to PCE in soil were used to assess the
effect of exposure concentrations and soil loading volume.
Rats were exposed to 15 and 50 mg PCE/kg soil at loading
volumes of either 0.0625 or 1 g/cm 2. The fraction absorbed
decreased with increasing load (Table 6); this is in agreement
with model predictions of McKone (1990). The increased
fractional absorption is related to the lower amount of PCE
applied and the increased surface area/soil volume that allowed
for increased absorption on a percent basis. However, the
amount absorbed and K P were essentially the same for the low
and high volume exposures. Trapping PCE at the skin surface
in the occluded exposure system also resulted in an increased
percentage absorbed, but did not alter the K P (Table 6). By
definition, K P should be independent of concentration, loading
volume, volatilization, and time of exposure, although each of
these variables can alter the percentage absorbed (Poet et al.,
2000a; U.S. EPA, 1992).
Absorption parameters (K P, fraction absorbed) and loss rate
(K loss) did not vary across doses (Table 6). The high volume
exposure systems (5 g of soil) demonstrate the importance of
loss of chemical, when PCE is permitted to volatilize away
from the exposure system; only ⬃10% is absorbed, much less
than the ⬃50% absorbed from the occluded system. In a real
world exposure scenario, it seems likely that PCE concentra-
TABLE 9
Rat Dual Compartment PCE Absorption Parameters
Parallel
Avg.
Layered
K PFO
K PCD
PFO
PCD
K PSC
K PVC
PSC
PVC
0.0002 ⫾ 0.00001
0.100 ⫾ 0.027
2.32 ⫾ 0.231
2.20 ⫾ 0.0015
0.102 ⫾ 0.030
22.0 ⫾ 21.9
37.3 ⫾ 1.39
249 ⫾ 39.4
Note. Parallel and layered subcompartment model permeability coefficients and partition coefficients from rat exposures. FO, follicular compartment; CD,
composite dermal; SC, subcutaneous; and VC, viable cutaneous; n ⫽ 7 rats representing different exposure scenarios.
27
DERMAL ABSORPTION OF PERCHLOROETHYLENE
FIG. 8. Example PBPK model predictions (lines) for rat exposures using
the three different dermal compartment descriptions; single homogenous, dual
parallel, and dual layered compared to the exhaled breath data (points averaged
every 1 min) for (A) PCE exposures in high volume, nonoccluded soil exposed
to target concentrations of 15 (open diamond), and 50 g/kg (filled circle), and
(B) PCE exposures in high volume, occluded soil to a target concentration of
50 g/kg.
tion in the soil will be low due to evaporation and movement
through the soil. In addition, chemical soil sorption follows a
process of aging by which the chemical contact with the soil
shifts from a reversible interaction on the surface of the soil
particle to deeper sites within the soil particles (Pignatello and
Xing, 1996). In such aged soils, dermal bioavailability may
decrease even further than observed in the exposures conducted in this study. The process of aging takes varying length
of time depending upon the soil type, temperature, microbial
content, etc. Thus, this study focused on the impact of reversible binding of PCE in a specific soil matrix and volatility. The
solubility of PCE in water is 0.01, and at concentrations below
this, no PCE was detected in exhaled breath of either rats
exposed to 5 ml of water or a single human subject exposed to
4 l of water using the hand immersion method (data not
shown).
The low K P and the expected volatilization of PCE from soil
would indicate that dermal exposures would not result in
significant uptake of PCE. In a PBPK model for PCE developed by Rao and Brown (1993), the octanol/water partition
coefficient was used to calculate a maximal dermal permeability coefficient of 0.125 cm/h (U.S. EPA, 1992). Applying this
coefficient to their model, they estimated that approximately
7– 8% of the total dose of PCE would be contributed by the
dermal absorption from the combined inhalation and dermal
uptake while showering. Bogen et al. (1992) calculated a K P of
0.37 cm/h in hairless guinea pigs from a dilute aqueous exposure, and Nakai et al. (1999) determined a PCE permeability
coefficient of 0.018 cm/h in human skin in vitro. The permeability of PCE determined here for rats (0.111 cm/h) was
similar to the K P of 0.125 cm/h estimated from the octanol/
water partition coefficient (Rao and Brown, 1993; U.S. EPA,
1992). However, the human K P for PCE absorption through the
hand was considerably lower than that estimated for human
abdominal and breast skin in vitro reported by Nakai et al.
(1999). Absorption through human abdominal and breast skin
is expected to be less than through the much thicker hand (U.S.
EPA, 1992), and the difference between rat and human absorption is consistent with previous studies that have shown that
percutaneous absorption can range from 1–20-fold higher in
rats compared to humans (Bronaugh, 1998; U.S. EPA, 1992).
In the previous studies with trichloroethylene and methyl chloroform, the K P in humans were approximately 20- and 40-fold
TABLE 10
Comparison of Log Likelihood Functions (LLF) for Single and Dual Compartment Models
Model
Parallel
Rat
Human
Layered
Rat
Human
␹ r2
LLF for single
LLF for dual
Likelihood function
Accept or reject
13.28
13.28
–1850
–1178
–1991
–1174
–281
7.73
Reject
Reject
13.28
13.28
–1850
–1178
–2287
–1017
–874
321
Reject
Accept
Note. The models were optimized for all of the human data sets and a subset of rat data sets (n ⫽ 7, representing each exposure dose and scenario). The values
given here are averages for all optimizations. The LLF optimization algorithms are for the rate limiting permeability coefficient, KP for the single compartment,
KPCD for the parallel, and KPSC for the layered model. Significance level for chi-square (␹ 2) distribution ⫽ 0.99 at r ⫽ 4 degrees of freedom.
28
POET ET AL.
FIG. 9. PBPK model predictions (lines) for rat exposures to methyl chloroform in soil (from Poet et al., 2000a) using a single, well-stirred dermal compartment with a delay added before the end of exposures (dashed lines) and a dual
layered dermal compartment compared (solid lines) to the exhaled breath data
(points averaged every 1 min) for the three human volunteers from that study. The
K PVC and KPSC for methyl chloroform dermal absorption from hand immersion
soil exposures were 0.004 ⫾ 0.002 and 0.003 ⫾ 0.001, respectively.
higher, respectively. The difference in K P for PCE between the
rats and humans was over 100-fold. Some of these differences
are likely due to the different exposure sites. The thickness of
the epidermis on the human hand ranges from ⬃85 ␮m on the
back of the hands to ⬃370 ␮m on the fingertips (International
Commission on Radiological Protection, 1992). The epidermis
of the F344 rat is in the range of 18 ␮m (Grabau et al., 1995).
Even more important is the difference between the stratum
corneum layer between the hands and feet and the rest of the
body. Over most of the body, the stratum corneum is uniform
and ranges from 13–16 ␮m, but on the palms of the hands and
the soles of the feet, the stratum corneum can be more than 600
␮m thick (International Commission on Radiological Protection, 1992). The stratum corneum rats has been estimated at
6 –13 ␮m in frozen sections from different sites on SpragueDawley rats (Monteiro-Riviere et al., 1990) and 11.2 ⫾ 5.1 ␮m
in the back of F344 rats (Grabau et al., 1995).
The fit of the PBPK model to the data using a single,
well-stirred skin compartment was poor for the human exposures, with less than 75% of the variability of the data explained by optimizing K P using the LLF using Simusolv™.
The slower than predicted elimination might be thought to
involve a lower rate of metabolism than predicted. Estimated
human metabolic rate constants for PCE have varied from
6.17–32.9 mg/h and 2.66 – 4.66 mg/l for V max and K M, respectively (Hattis et al., 1993; Reitz et al., 1996). However, as is
typical for flow limited models, simulations of the low exposures were insensitive to changes in the metabolic rate constants.
Since the concentrations of PCE in blood and liver were well
below the K M, the most likely cause of the poor fit to the human
data was the use of the overly simplistic well-stirred dermal
compartment. Because the stratum corneum is so prominent on
the hand, and it has been proposed to function as a barrier to
percutaneous absorption, a model that separated out the stratum corneum and the viable epidermis into two compartments
was used. The stratum corneum compartment represented a
barrier to absorption with a rate-limiting K p and the viable
epidermis compartment contained the rest of the skin and the
blood exchange. The optimized K p for the stratum corneum
compartment (K pSC) was not different from the K p estimate
for the one compartment model. However, the optimized K p
for the transfer of PCE from the stratum corneum to the viable
cutaneous compartment (K pVC) was very similar to the K p
estimated for the dermal absorption of PCE in rats (Tables 5
and 8).
The second prevalent theory for the different rate of absorption between laboratory animals and humans is the existence of
skin structures, such as follicles, that act as shunts (Kao et al.,
1988; McKone, 1990; Scheuplein and Blank, 1971). Therefore,
an additional two-compartment model that might explain the
increased rat absorption over the human was compared. For the
parallel model, both subcompartments were in contact with the
exposure dose and the blood exchange. However, the utilization of this parallel model form did not improve the fit of the
data (Fig. 7).
Since the depth of the stratum corneum and the number of
follicles can vary greatly between species (Grabau et al.,
1995), the two different dual skin compartment models were
also tested against the rat exhaled breath data (Fig. 8). No
improvement in the fit of the model was observed using either
of the dual dermal compartment models (Table 10). The permeability coefficients for the composite dermal (K PFO) in the
parallel model and the subcutaneous barrier (K PSC) in the
layered model were not different form the aggregate K P for the
one, homogenous compartment, indicating that, in these mod-
29
DERMAL ABSORPTION OF PERCHLOROETHYLENE
TABLE 11
Comparison of Log Likelihood Functions (LLF) for Methyl Chloroform
Layered model
vs. scenario
␹ r2
Avg. exposure end
(h) a
LLF for
single
LLF for
dual
Likelihood
function
Accept or
reject
1
2
13.28
13.28
2.8
2.0
–1075
–1134
–1052
–1052
45.3
164
Accept
Accept
Note. The dual layered model developed for PCE exposures was modified to describe methyl chloroform exposures in humans to contaminated water from
Poet et al. (2000b). The values given here are averages for all optimizations. The LLF optimization algorithms are for the rate limiting permeability coefficient,
K P for the single compartment, and K PSC for the layered model. Significance level for chi-square (␹ 2) distribution ⫽ 0.99 at r ⫽ 4 degrees of freedom.
a
To get acceptable data fits to the original methyl chloroform exhaled breath data, both the lag time before the appearance of methyl chloroform in the exhaled
breath and the time of exposure end were modified (scenario 1) as described in the text. The second scenario did not include modifying the end of exposure
(exposure stop ⫽ 2 h).
els, the sc barrier drives the absorption rate, not the existence
of skin appendages such as follicles.
Researchers in this laboratory group have previously conducted dermal absorption studies with other volatile organic
compounds. For temperature-dependent absorption of chloroform from whole-body bathwater exposures, hand or forearm
exposures to trichloroethylene, or arm-only exposures to 2-butoxyethanol vapor a single, well-stirred skin compartment provided a good fit to the data (Corley et al., 1997, 2000; Poet et
al., 2000a). For methyl chloroform, however, a reasonable fit
to the data was only obtained after an initial delay was added
and the length of the exposure was artificially adjusted to
account for the delay at the end of exposure. In this way,
exposures that were continued for 2 h were modeled as if they
were extended to up to 3.3 h (Poet et al., 2000b). This assumes
that the skin is acting as a depot that continues to release
chemical after the cessation of the actual exposure. Since the
dual-layered dermal structure seems to describe the dermal
absorption of PCE in these very similar exposures, the same
model structure was used to describe the methyl chloroform
exposures in three human subjects exposed to methyl chloroform by immersing a hand in a bucket of water containing
1.2 ⫾ 0.276 g/l or soil containing 7.1 ⫾ 0.896 g/kg (Fig. 9), as
reported previously. The updated model structure employed a
more empirical model scenario with two differences. First, the
lag time in the model was described by extrapolating the delay
in the appearance of methyl chloroform in the exhaled breath
back to the x-axis, instead of optimizing to the fit of the data.
Second, no time was added to the model describing the exposure beyond the actual exposure time, thus, the end of the
exposure described in the model was 2 h, the time the hand was
removed from the water. The fit using the dual layered model
was significantly improved over the simple well-stirred compartment, using the likelihood ratio test as described in the
Materials and Methods. A similar improvement of fit was
demonstrated for the exposures to methyl chloroform in water
(Table 11).
Data from the whole-body exposures to chloroform and
hand exposures to trichloroethylene were all adequately fit
using a single, well-stirred skin compartment (Corley et al.,
1997, 2000; Poet et al., 2000a). Whole-body submersion in
water and hand-only exposures are very different and comparisons are difficult, particularly when the differences in dermal
composition of the hand discussed earlier are taken into account. The reasoning behind the need for a two compartment
model for PCE and methyl chloroform, but not for trichloroethylene are not as clear. Methyl chloroform and PCE are both
less water soluble (0.078 and 0.015%, respectively) than trichloroethylene (0.1%). The biggest difference between these
two chemicals and trichloroethylene is in their partition coefficients for skin/soil and skin/blood. Methyl chloroform and
PCE both have higher skin/blood partitioning (4.3 and 4.0,
respectively) than trichloroethylene (1.5). In addition the skin/
media partitioning is much lower for methyl chloroform and
PCE (1.3 and 0.4, respectively) compared to trichloroethylene
(7.1). Therefore, based on partitioning alone, methyl chloroform and PCE would be expected to partition less readily into
the blood than trichloroethylene. Additionally, PCE and
methyl chloroform are poorly metabolized and the higher rate
of metabolism of trichloroethylene may mask the effects of
dermal absorption through the stratum corneum.
As models become more complex, there is a risk of increasing the number of parameters beyond that which are needed.
When it becomes necessary to add complexity to the model,
additional unknown parameters are also added and it is essential to justify their use, both biologically and practically. The
use of the layered dermal model to describe human percutaneous absorption is justified by a visual assessment, statistically,
and physiologically. The ability of the dual-layered model to
drastically improve the fit of the human PCE exposure data and
the inability of the parallel model to improve the fit for either
the rat or human exposures indicates that the stratum corneum
plays an important role in the barrier function of the skin
toward absorption of PCE, particularly when exposure is to the
hands. That the estimation of a K p through the viable cutaneous
tissue so closely resembles the rat K p may suggest that the role
of the stratum corneum is very important in species differences
in dermal absorption, but the role of skin appendages such as
hair follicles may be minimal.
The impact of the two-compartment dermal model on risk
30
POET ET AL.
assessment for PCE will be most evident in species comparisons. Currently, it is difficult to extrapolate from rodent to
human exposure assessments. For PCE, it appears that taking
into account skin morphology into species-specific models may
facilitate such extrapolations in the future. The thick stratum
corneum on the hand may have influenced the human K P
determined in these experiments, as indicated by the effect of
separating out the skin in the PBPK model to a dual layered
compartment, and exposures to other body sites may alter the
assessment of K P for PCE.
APPENDIX
Equations and abbreviations for the alternative parallel or layered subcompartments used in the PBPK model were adapted from Bookout et al. (1996,
1997) and are discussed below.
Permeability constants. With the layered model, the K Ps have an inverse
relationship to each other. Therefore, 1/K P equals the sum of the recripocals of
the subcompartment K Ps. The permeability for the viable cutaneous (VC)
compartment was determined by the relationship between the optimized K P for
the subcutaneous (SC) compartment and the overall K P for the single wellstirred compartment.
KPCD ⫽
共KP ⫻ KPSC兲
共KP ⫺ KPSC兲
(8)
Partition coefficients. The partition coefficients measured experimentally for
the single skin compartment is equal to the sum of the partition coefficients for
each of the subcompartments following a volume correction. The partition
coefficient for the VC compartment was determined based on the optimized
partition coefficient for the SC compartment and the partition coefficient
measured experimentally.
PVC ⫽
共PSK ⫻ VSK兲 ⫺ 共PSC ⫻ VSC兲
VVc
(9)
Parallel Subcompartment Model
The parallel subcompartment model partitioned the skin compartment into a
follicular (FO) and the remaining composite dermal (CD). Both compartments
were in contact with the PCE-laden soil and were involved in blood exchange
(Fig. 1).
Permeability constants. The permeability coefficient (K P) for the single
well-stirred dermal compartment times the total surface area (SA) equals the
sum of the K P times the SA for each subcomartment. The permeability for the
CD compartment was determined by the relationship between the optimized
K P for the FO compartment and the overall K P for the single well-stirred
compartment once the equation was rearranged to the following:
共KP ⫻ SA兲 ⫺ 共KPFO ⫻ SAFO兲
KPCD ⫽
SACD
PCD ⫽
共PSK ⫻ VSK兲 ⫺ 共PFO ⫻ VFO兲
VCD
(4)
(5)
Partition coefficients. According to Bookout et al. (1996), the partition
coefficients measured experimentally for the single skin compartment is equal
to the sum of the partition coefficients for each of the subcompartments
following a volume correction. The partition coefficient for the CD compartment was determined based on the optimized partition coefficient for the FO
compartment and the partition coefficient measured experimentally for whole
skin by Mattie et al. (1994).
Mass-balance equations for dermal absorption. Equations for the transfer of
PCE through the FO and CD compartments replace the single equation
(Equation 1) for the well-stirred dermal compartment model.
冉
KPFO ⫻ SAFO
dAskFO
⫹
dt
1000
冊 冉
⫻
冉
CFO
csurf ⫺
PMFO
冉
dAskCD
KPCD ⫻ SACD
⫽
dt
1000
冊 冉
⫻
csurf ⫺
冉
CCD
PMCD
(6)
冊冊
⫹ QCD ⫻ 共Cart ⫺ Cven兲
冉
KPSC ⫻ SASC
dAskSC
⫽
dt
1000
冊 冉
⫻
csurf ⫺
⫹
冉
KPVC ⫻ SAVC
dAskVC
⫽
dt
1000
冉
CSC
PMSC
冊
(10)
⫹ QVC ⫻ 共Cart ⫺ Cven兲
(11)
KPVC ⫻ SAVC
1000
冊 冉冉 冊
⫻
冊冊
CSC
PSC
⫺ CVC
冉
CVC ⫺
CSC
PSC
冊
where: art, arterial; C, concentrations (mg/l); CD, composite dermal; FO,
follicular; KP, permeability (cm/h); P, partition coefficient (unitless); Q, blood
flow (l/h); SA, surface area (cm 2); SC, subcutaneous; SK, skin; t, time (h); V,
volume (l); VC, viable cutaneous; ven, venous.
ACKNOWLEDGMENT
The authors thank Amber Alford for her assistance in conducting the rat
exposures.
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