United States
Environmental Protection
Agency
Office of Research and
Development
Washington, DC 20460
EPA/600/AP-92/001a
August 1992
Workshop Review Draft
&EPA
Chapter 1.
Disposition and
Pharmacokinetics
Review
Draft
(Do Not
Cite or
Quote)
600AP92001a
Notice
This document is a preliminary draft. It has not been formally released by EPA and should not
at this stage be construed to represent Agency policy. It is being circulated for comment on
its technical accuracy and policy implications.
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DRAFT EPA/600/AP-92/001a
DO NOT QUOTE OR CITE August 1992
Workshop Review Draft
Chapter 1. Disposition and Pharmacokinetics
Health Assessment for
2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD)
and Related Compounds
NOTICE
THIS DOCUMENT IS A PRELIMINARY DRAFT. It has not been formally released by the U.S.
Environmental Protection Agency and should not at this stage be construed to represent Agency
policy. It is being circulated for comment on its technical accuracy and policy implications.
Office of Health and Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Washington, D.C.
Printed on Recycled Paper
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DISCLAIMER
This document is a draft for review purposes only and does not constitute Agency policy.
Mention of trade names or commercial products does not constitute endorsement or recommendation
for use.
Please note that this chapter is a preliminary draft and as such represents work
in progress. The chapter is intended to be the basis for review and discussion at
a peer-review workshop. It will be revised subsequent to the workshop as
suggestions and contributions from the scientific community are incorporated.
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CONTENTS
Tables v
Figures vi
List of Abbreviations vii
Authors and Contributors xii
1. DISPOSITION AND PHARMACOKINETICS 1-1
1.1. ABSORPTION/BIOAVAILABILITY FOLLOWING EXPOSURE 1-1
1.1.1. Oral 1-1
1.1.2. Dermal Absorption 1-8
1.1.3, Transpulmonary Absorption 1-13
1.1.4. Parenteral Absorption 1-13
1.2. DISTRIBUTION 1-14
1.2.1, Distribution in Blood and Lymph 1-14
1.2.2, Tissue Distribution 1-16
1.2.3, Time-Dependent Tissue Distribution 1-22
1.2.4. Dose-Dependent Tissue Distribution 1-34
1.2.5, Potential Mechanisms for the Dose-Dependent
Tissue Distribution 1-39
1.3. METABOLISM AND EXCRETION 1-42
1.3.1, Structure of Metabolites 1-43
1.3.2. Toxicity of Metabolites " 1-51
1.3.3, Autoinduction of Metabolism 1-53
1.3.4, Excretion in Animals 1-57
1.3.5. Excretion in Humans 1-61
1.4. PHYSIOLOGICALLY BASED PHARMACOKINETIC MODELS 1-65
1.5. PHARMACOKINETICS IN SPECIAL POPULATIONS 1-74
1.5.1, Pregnancy and Lactation (Prenatal and
Postnatal Exposure of Offspring) 1-74
1.5.2. Aging 1-81
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1.6. REFERENCES
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LIST OF TABLES
1.1 Gastrointestinal Absorption of 2,3,7,8-TCDD and Related
Compounds Following a Single Oral Exposure by Gavage 1-2
1.2 Percentage of 2,3,7,8-TCDD in the Liver of Rats 24 Hours
After Oral Administration of 0.5 mL of Various Formulations
Containing TCDD 1-7
1.3 Dermal Absorption of 2,3,7,8-TCDD and Related Compounds in
the Rat 1-9
1.4 Tissue Distribution of [14C]-2-3,7,8-TCDD in Female Wistar
Rats 1-18
1.5 Elimination of 2,3,7,8-TCDD and Related Compounds from Major Tissue Depots 1-25
1.6 Elimination Constants and Half-Lives of Various 2,3,7,8-Substituted CDDs and CDFs in
Hepatic and Adipose Tissue of Marmoset Monkeys 1-31
1.7 2,3,7,8-TCDD Concentrations in Liver and Adipose Tissue
Following Different Doses and Calculated Concentration
Ratios (Liver/Adipose Tissue) 1-36
1.8 Metabolism and Excretion of 2,3,7,8-TCDD and Related
Compounds 1-44
1.9 Half-Life Estimates for 2,3,7,8-TCDD and Related Compound
in Humans 1-62
1.10 Pharmacokinetic Parameters for 2,3,7,8-TCDD Used in
PB-Pk Models 1-66
1.11 Pharmacokinetic Parameters for 2,3,7,8-TCDF Used in the
PB-Pk Model Described by King et al. (1983) 1-72
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LIST OF FIGURES
1-1 Time Course of the Concentration of 14C-TCDD in Rat Liver and Adipose
Tissue After a Single Subcutaneous Injection of 300 ng TCDD/kg bw to
Female Rats (M±SD) 1_24
1-2 Dose Dependency of the Percentage of the Administered Dose of 14C-TCDD/g
of Tissue Recovered in Liver and Adipose Tissue After Single Subcutaneous
Doses _
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LIST OF ABBREVIATIONS
ACTH Adrenocorticotrophic hormone
Ah Aryl hydrocarbon
AHH Aryl hydrocarbon hydroxylase
ALT L-alanine aminotransferase
AST L-asparate aminotransferase
BDD Brominated dibenzo-p-dioxin
BDF Brominated dibenzofuran
BCF Bioconcentration factor
BGG Bovine gamma globulin
bw Body weight
cAMP Cyclic 3,5-adenosine monophosphate
CDD Chlorinated dibenzo-p-dioxin
cDNA Complementary DNA
CDF Chlorinated dibenzofuran
CNS Central nervous system
CTL CytOtoxic T lymphocyte
DCDD 2,7-Dichlorodibenzo-p-dioxin
DHT 5oc-Dihydrotestosterone
DMBA Dimethylbenzanthracene
DMSO Dimethyl sulfoxide
DNA Deoxyribonucleic acid
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DRE
DTG
DTK
ECOD
EGF
EGFR
ER
EROD
EOF
FSH
GC-ECD
GC/MS
GOT
GnRH
GST
HVH
HAH
HCDD
HDL
HxCB
HpCDD
HpCDF
Dioxin-responsive enhancers
Delayed type hypersensitivity
Delayed-type hypersensitivity
Dose effective for 50% of recipients
7-Ethoxycoumarin-O-deethylase
Epidermal growth factor
Epidermal growth factor receptor
Estrogen receptor
7-Ethoxyresurofin 0-deethylase
Enzyme altered foci
Follicle-stimulating hormone
Gas chromatograph-electron capture detection
Gas chromatograph/mass spectrometer
Gamma glutamyl transpeptidase
Gonadotropin-releasing hormone
Glutathione-S-transferase
Graft versus host
Halogenated aromatic hydrocarbons
Hexachlorodibenzo-p-dioxin
High density lipoprotein
Hexachlorobiphenyl
Heptachlorinated dibenzo-p-dioxin
Heptachlorinated dibenzofuran
vin
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LIST OF ABBREVIATIONS (cont.)
HPLC
HRGC/HRMS
HxCDD
HxCDF
High performance liquid chromatography
High resolution gas chromatography/high resolution mass spectrometry
Hexachlorinated dibenzo-p-dioxin
Hexachlorinated dibenzofuran
I-TEF
LH
LDL
LPL
LOAEL
LOEL
MCDF
MFO
mRNA
MNNG
NADP
NADPH
NK
NOAEL
NOEL
International TCDD-toxic-equivalency
Dose lethal to 50% of recipients (and all other subscripter dose levels)
Luteinizing hormone
Low density liproprotein
Lipoprotein lipase activity
Lowest-observable-adverse-effect level
Lowest-observed-effect level
6-Methyl- 1 ,3,8-trichlorodibenzofuran
Mixed function oxidase
Messenger RNA
/V-methyl-./V-nitrosoguanidme
Nicotinamide adenine dinucleotide phosphate
Nicotinamide adenine dinucleotide phosphate (reduced form)
Natural killer
No-observable-adverse-effect level
No-observed-effect level
IX
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LIST OF ABBREVIATIONS (cont.)
OCDD
OCDF
PAH
PB-Pk
PCB
OVX
PEL
PCQ
PeCDD
PeCDF
PEPCK
PGT
PHA
PWM
ppm
PPQ
ppt
RNA
SAR
SCOT
SGPT
Octachlorodibenzo-p-dioxin
Octachlorodibenzofuran
Polyaromatic hydrocarbon
Physiologically based pharmacokinetic
Polychlorinated biphenyl
Ovariectomized
Peripheral blood lymphocytes
Quaterphenyl
Pentachlorinated dibenzo-p-dioxin
Pentachlorinated dibenzo-p-dioxin
Phosphopenol pyruvate carboxykinase
Placental glutathione transferase
Phytohemagglutinin
Pokeweed mitogen
Parts per million
Parts per trillion
Ribonucleic acid
Structure-activity relationships
Serum glutamic oxaloacetic transaminase
Serum glutamic pyruvic transaminase
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LIST OF ABBREVIATIONS (cont.)
SRBC
t*
TCAOB
TCB
TCDD
TEF
TGF
tPA
TNF
TNP-LPS
TSH
TTR
UDPGT
URO-D
VLDL
v/v
w/w
Sheep erythrocytes (red blood cells)
Half-time
Tetrachloroazoxybenzene
Tetrachlorobiphenyl
Tetrachlorodibenzo-p-dioxin
Toxic equivalency factors
Thyroid growth factor
Tissue plasminogen activator
Tumor necrosis factor
lipopolysaccharide
Thyroid stimulating hormone
Transthyretrin
UDP-glucuronosyltransferases
Uroporphyrinogen decarboxylase
Very low density lipoprotein
Volume per volume
Weight by weight
XI
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AUTHORS AND CONTRIBUTORS
The Office of Health and Environmental Assessment (OHEA) within the Office of Research
and Development was responsible for the preparation of this chapter. The chapter was prepared
through Syracuse Research Corporation under EPA Contract No. 68-CO-0043, Task 20, with Carol
Haynes, Environmental Criteria and Assessment Office in Cincinnati, OH, serving as Project Officer.
During the preparation of this chapter, EPA staff scientists provided reviews of the drafts as
well as coordinating internal and external reviews.
AUTHORS
James Olson
Department of Pharmacology and Therapeutics
State University of New York - Buffalo
Buffalo, NY
EPA CHAPTER MANAGER
Jerry Blancato
Office of Research and Development
Environmental Monitory Systems Laboratory
Las Vegas, NV
Xli 08/10/92
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1. DISPOSITION AND PHARMACOKINETICS
The disposition and pharmacokinetics of 2,3,7,8-TCDD and related compounds
have been investigated in several species and under various exposure conditions.
There are several reviews on this subject that focus on 2,3,7,8-TCDD and related
halogenated aromatic hydrocarbons (Neal et al., 1982; Gasiewicz et al., 1983a;
Olson et al., 1983; Birnbaum, 1985). During the last 6 years, considerably more
data have been published on this class of compounds that includes 2,3,7,8-
substituted CDDs, BDDs, CDFs, BDFs and the coplaner PCBs and PBBs. This chapter
reviews the disposition and pharmacokinetics of these agents and identifies
congener and species specific factors that may have an impact on the dose-related
biological responses of these compounds.
1.1. ABSORPTION/BIOAVAILABILITY FOLLOWING EXPOSURE
The gastrointestinal, dermal and transpulmonary absorption of these
compounds are discussed herein because they represent potential routes for human
exposure to this class of persistent environmental contaminants. Parenteral
absorption is reviewed since this route of exposure has been used to generate
disposition and pharmacokinetic data on these compounds.
1.1.1. Oral
1.1.1.1. GASTROINTESTINAL ABSORPTION IN ANIMALS — A major source of human
exposure to 2,3,7,8-TCDD and related compounds is thought to be through the diet.
Experimentally, these compounds are commonly administered in the diet or by
gavage in an oil vehicle. Gastrointestinal absorption is usually estimated as
the difference between the administered dose (100%) and the % of the dose that
was not absorbed. The unabsorbed fraction is estimated as the recovery of parent
compound in feces within 24-48 hours of a single oral exposure by gavage. Table
1-1 summarizes gastrointestinal absorption data on 2,3,7,8-TCDD and related
compounds.
In Sprague-Dawley rats given a single oral dose of 1.0 pg [ 14C]-2,3,7,8-
TCDD/kg bw in acetone:corn oil (1:25, v/v), the fraction absorbed ranged from
66-93% with a mean of 84% (Rose et al., 1976). With repeated oral dosing of rats
at 0.1 or 1.0 pg/kg/day (5 days/week for 7 weeks), gastrointestinal absorption
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TABLE 1-1 (cont.)
Chemical
Species (Sex)
Dose
((tmol/kg)
(Kg/kg)
Vehicle
% Administered
Dose Absorbed
[Mean (Range)]
Reference
PCBs
3,3'4,4'-T4CB
C57BL mouse (F)
34.5
10,000
corn oil
77
Uehler et al.,
1989
Absorption is generally estimated as the difference between the administered dose (100%) and the X of the dose that was not absorbed. The
unabsorbed fraction is estimated as the recovery of parent compound in feces within 48 hours of exposure.
NR = Not reported
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of 2,3,7,8-TCDD was observed to be approximately that observed for the single
oral exposure (Rose et al., 1976). Oral exposure of Sprague-Dawley rats to a
larger dose of 2,3,7,8-TCDD in acetone: corn oil (50 pg/kg) resulted in an
average absorption of 70% of the administered dose (Piper et al., 1973).
One study in the guinea pig reported that -50% of a single oral dose of
2,3,7,8-TCDD in acetonercorn oil was absorbed (Nolan et al., 1979). The
gastrointestinal absorption of 2,3,7,8-TCDD was also examined in the hamster, the
species most resistant to the acute toxicity of this compound (Olson et al.,
1980a). Hamsters were given a single, sublethal, oral dose of [1,6-3H]-2,3,7,8-
TCDD in olive oil (650 pig/kg), and an average of 75% of the dose was absorbed.
When 2,3,7,8-TCDD was administered to rats in the diet at 7 or 20 ppb (0.5 or 1.4
A*9/kg/day) for 42 days, 50-60% of the consumed dose was absorbed (Fries and
Marrow, 1975). These findings indicate that oral exposure to 2,3,7,8-TCDD in the
diet or in an oil vehicle results in the absorption of >50% of the administered
dose.
The intestinal absorption of 3H-2,3,7,8-TCDD has also been investigated in
thoracic duct-cannulated rats (Lakshmanan et al., 1986). The investigators
concluded that 2,3,7,8-TCDD was absorbed into chylomicrons and transported
through the lymphatic system prior to entering the systemic circulation.
The absorption of 2,3,7,8-TBDD in male Fischer 344 rats was studied after
oral exposure by gavage at 5 ^g/kg in Emulphor:ethanol:water (1:1:3) (Diliberto
et al., 1990). The percent of the dose absorbed for this study was defined as
100 — (% total oral dose in feces on day 1 and 2 — % total intravenous dose in
feces on day 1 and 2) using the intravenous pharmacokinetic data of Kedderis et
al. (1991).
The relative absorbed dose or bioavailability of 2,3,7,8-TBDD after oral
exposure was estimated at 78, 82, 60 and 47% at dose levels of 0.001, 0.01, 0.1
and 0.5 pmol/kg, respectively. These results suggest nonlinear absorption at the
higher doses, with maximal oral absorption at an exposure of <0.01 ^imol/kg (5
/^g/kg).
The absorption of 2,3,7,8-TCDF has been investigated after oral exposure by
gavage. Approximately 90% of the administered dose (0.1 and 1.0 ^imol/kg) of
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2,3,7,8-TCDF in Emulphor:ethanol (1:1) was absorbed in male Fischer 344 rats
(Birnbaum et al., 1980). [Emulphor EL-620 is a polyoxyethylated vegetable oil
preparation (GAF Corp., New York, NY)]. Similarly, >90% of the administered dose
(0.2 pmol/kg, 6 pg/kg and 1-15 pg/kg) of 2,3,7,8-TCDF in Emulphor: ethanol:water
(1:1:8) was absorbed in male Hartley guinea pigs (Decad et al., 1981a; loannou
et al., 1983). Thus, 2,3,7,8-TCDF appears to be almost completely absorbed from
the gastrointestinal tract. This may be related to the greater relative
solubility of 2,3,7,8-TCDF compared to that of 2,3,7,8-TCDD or 2,3,7,8-TBDD.
The oral bioavailability of 2,3,4,7,8-PeCDF and 3,3',4,4'-TCB in corn oil
were similar to that of 2,3,7,8-TCDD (Brewster and Birnbaum, 1987; Wehler et al.,
1989; Clarke et al., 1984). Furthermore, 2,3,4,7,8-PeCDF absorption was
independent of the dose (0.1, 0.5 or 1.0 pmol/kg). Incomplete and variable
absorption of 1,2,3,7,8-PeCDD was reported in rats, with 19-71% of the dose
absorbed within the first 2 days after oral exposure (Wacker et al., 1986).
Early studies on the pharmacokinetic behavior of OCDD by Williams et al.
(1972) and Norback et al. (1975) demonstrated that OCDD was poorly absorbed after
oral exposure. More recently, Birnbaum and Couture (1988) also found that the
gastrointestinal absorption of OCDD in rats was very limited, ranging from 2-15%
of the administered dose. Lower doses (50 pg/kg) in a o-dichlorobenzene:corn oil
(1:1) vehicle were found to give the best oral bioavailability for this extremely
insoluble compound.
1.1.1.2. GASTROINTESTINAL ABSORPTION IN HUMANS — Poiger and Schlatter
(1986) investigated the absorption of 2,3,7,8-TCDD in a 42-year-old man after
ingestion of 105 ng 3H-2,3,7,8-TCDD (1.14 ng/kg bw) in 6 mL corn oil and found
that >87% of the oral dose was absorbed from the gastrointestinal tract.
Following absorption, the half-life for elimination was estimated to be 2120
days.
The above data indicate that gastrointestinal absorption of 2,3,7,8-TCDD and
related compounds is variable, incomplete and congener specific. More soluble
congeners, such as 2,3,7,8-TCDF, are almost completely absorbed, while the
extremely insoluble OCDD is very poorly absorbed. In some cases, absorption has
been found to be dose dependent, with increased absorption occurring at lower
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doses (2,3,7,8-TBDD, OCDD). The limited data base also suggests that there are
no major interspecies differences in the gastrointestinal absorption of these
compounds.
1.1.1.3. BIOAVAILABILITY FOLLOWING ORAL EXPOSURE — Oral exposure of
humans to 2,3,7,8-TCDD and related compounds usually occurs as a complex mixture
of these contaminants in food, soil, dust, water or other mixtures that would be
expected to alter absorption.
The influence of dose and vehicle or adsorbent on gastrointestinal
absorption has been investigated in rats by Poiger and Schlatter (1980), using
hepatic concentrations 24 hours after dosing as an indicator of the amount
absorbed (Table 1-2). Administration of 2,3,7,8-TCDD in an aqueous suspension
of soil resulted in a decrease in the hepatic levels of 2,3,7,8-TCDD as compared
with hepatic levels resulting from administration of 2,3,7,8-TCDD in 50% ethanol.
The extent of the decrease was directly proportional to the length of time the
2,3,7,8-TCDD had been in contact with the soil. When 2,3,7,8-TCDD was mixed in
an aqueous suspension of activated carbon, absorption was almost totally
eliminated (<0.07% of the dose in hepatic tissues).
Philippi et al. (1981) and Hutter and Philippi (1982) have shown that
radiolabeled 2,3,7,8-TCDD becomes progressively more resistant with time to
extraction from soil. Similarly, the feeding of fly ash, which contains CDDs,
to rats in the diet for 19 days resulted in considerably lower hepatic levels of
CDDs than did the feeding of an extract of the fly ash at comparable dietary
concentrations of CDDs (van den Berg et al., 1983). The CDDs were tentatively
identified as 2,3,7,8-TCDD, 1,2,3,7,8-PeCDD, 1,2,3,6,7,8-HxCDD and 1,2,3,7,8,9-
HxCDD and the difference in hepatic levels noted between fly ash-treated and
extract-treated rats was greater for the more highly chlorinated isomers than it
was for 2,3,7,8-TCDD. These results indicate the importance of the formulation
or vehicle containing the toxin(s) on the relative bioavailability of 2,3,7,8-
TCDD, PeCDD and HxCDDs after oral exposure.
Since 2,3,7,8-TCDD in the environment is likely to be absorbed to soil,
McConnell et al. (1984) and Lucier et al. (1986) compared the oral bioavail-
ability of 2,3,7,8-TCDD from environmentally contaminated soil to that from
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TABLE 1-2
Percentage of 2,3,7,8-TCDD in the Liver of Rats 24 Hours
After Oral Administration of 0.5 mL of Various
Formulations Containing TCDD*
Formulation
50% ethanol
Aqueous suspension of soil
(37%, w/w) that had been in
contact with TCDD for:
10-15 hours
8 days
Aqueous suspension of activated
carbon (25%, w/w)
TCDD Dose
(ng)
14.7
12.7, 22.9
21.2, 22.7
14.7
No. of
Animals
7
17
10
6
Percentage of
Dose in the Liver
36.7±1.2
24.1±4.8
16.0+2.2
<0.07
*Source: Poiger and Schlatter, 1980
w/w = Weight by weight
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2,3,7,8-TCDD administered in corn oil in rats and guinea pigs and rats, respec-
tively. As indicated by biological effects and the amount of 2,3,7,8-TCDD in the
liver, the intestinal absorption from Times Beach and Minker Stout, Missouri,
soil was -50% less than from corn oil. Shu et al. (1988) reported an oral
bioavailability of -43% in the rat dosed with three environmentally contaminated
soil samples from Times Beach, Missouri. This figure did not change signifi-
cantly over a 500-fold dose range of 2-1450 ng 2,3,7,8-TCDD/kg bw for soil
contaminated with -2, 30 or 600 ppb of 2,3,7,8-TCDD. In studies of other soil
types, Umbreit et al. (1986a,b) estimated an oral bioavailability in the rat of
0.5% for soil at a New Jersey manufacturing site and 21% for a Newark salvage
yard. These results indicate that bioavailability of 2,3,7,8-TCDD from soil
varies between sites and that 2,3,7,8-TCDD content alone may not be indicative
of potential human hazard from contaminated environmental materials. Although
these data indicate that substantial absorption may occur from contaminated soil,
soil type and duration of contact, as suggested from the data that demonstrated
decreased extraction efficiency with increasing contact time between soil and
2,3,7,8-TCDD (Philippi et al., 1981; Huetter and Philippi, 1982), may substan-
tially affect the absorption of 2,3,7,8-TCDD from soils obtained from different
contaminated sites.
1.1.2. Dermal Absorption. Brewster et al. (1989) examined the dermal absorp-
tion of 2,3,7,8-TCDD and three CDFs in male Fischer 344 rats (10 weeks old;
200-250 g). The fur was clipped from the intrascapular region of the back of
each animal. A single compound was then applied over a 1.8 cm^ area of skin in
60 pL of acetone and covered with a perforated stainless steel cap. Table 1-3
summarizes data on the absorption of each compound at 3 days after a single
dermal exposure. At an exposure of 0.1 pmol/kg, the absorption of 2,3,7,8-TCDF
(49% of administered dose) was greater than that of 2,3,4,7,8-PeCDF, 1,2,3,7,8-
PeCDF and 2,3,7,8-TCDD. For each compound, the relative absorption (percentage
of administered dose) decreased with increasing dose while the absolute absorp-
tion (jug/kg) increased nonlinearly with dose. Results also suggest that the
majority of the compound remaining at the skin exposure site was associated with
the stratum corneum and did not penetrate through to the dermis. In a subsequent
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TABLE 1-3
Dermal Absorption of 2,3,7,8-TCDD and Related Compounds in the Rata
Chemical
2,3,7,8-TCDD
2,3,7,8-TCDF
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
Dose
(jumol/kg)
0.00015
0.001
0.01
0.1
0.5
1.0
0.1
0.5
1.0
0.1
0.5
1.0
0.1
0.5
1.0
(pg/kg)
0.05
0.32
3.2
32
160
321
31
153
306
34
170
340
34
170
340
% Administered Dose
Skin Siteb
61.73±4.37
59.71±1.90
72.60±0.41
82.21+2.85
80.92±2.74
82.68±3.69
51.18±11.95
82.14±11.22
88.70±5.17
74.7213.58
91.67±2.46
84.23+5.44
65.77+4.80
75.50±1.81
81.84±1.67
Absorbed
38.27±4.37
40.29±1.89
27.40±0.41
17.78+2.85
19.08±2.74
17.30±3.67
48.84±11.95
17.86+11.22
11.32+5.17
25.2713.58
8.3312.46
15.7615.44
34.1914.78
24.5011.80
18.1611.67
aSource: Brewster et al., 1989
'Values are the meaniSD of three to four animals and represent the
amount of administered dose of radiolabeled congener remaining at
the application site 3 days after dermal exposure.
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study, Banks and Birnbaum (1991a) examined the rate of absorption of 2,3,7,8-TCDD
over 120 hours after the dermal application of 200 pmol (1 nmol/kg) to male
Fischer 344 rats. The absorption kinetics appeared to be first-order, with an
absorption rate constant of 0.005 hour . Using a similar exposure protocol, the
dermal absorption of 2,3,7,8-TCDF was found to follow a first-order process with
a rate constant of 0.009 hour"1 (Banks and Birnbaum, 1991b). Together, these
results on dermal absorption indicate that at lower doses (<0.1 /jmol/kg), a
greater percent of this administered dose of 2,3,7,8-TCDD and three CDFs was
absorbed. Nonetheless, the rate of absorption of 2,3,7,8-TCDD is still very slow
(rate constant of 0.005 hour"1) even following a low dose dermal application of
200 pmol (1 nmol/kg). Results from Table 1-3 also suggest that the dermal
absorption of 2,3,7,8-TCDF, 2,3,4,7,8-PeCDF and 1,2,3,7,8-PeCDF occurs at a very
slow rate. Using a similar exposure protocol, the dermal absorption of 2,3,7,8-
TBDD was only 30-40% of that observed for 2,3,7,8-TCDD (Jackson et al., 1991).
Rahman et al. (1992) and Gallo et al. (1992) compared the in vitro
permeation of 2,3,7,8-TCDD through hairless mouse and human skin. In both
species, the amount of 2,3,7,8-TCDD permeated increased with the dose, but the
percent of the dose permeated decreased with increasing dose. The permeability
coefficient of 2,3,7,8-TCDD in human skin was about one order of magnitude lower
than that in mouse skin. The hairless mouse skin does not appear to be a
suitable model for the permeation of 2,3,7,8-TCDD through human skin since the
viable tissues were the major barrier to 2,3,7,8-TCDD permeation in hairless
mouse skin, while the stratum corneum layer provided the greater resistance in
human skin. A significant increase in 2,3,7,8-TCDD permeation through human skin
was observed when the skin was damaged by tape-stripping. Gallo et al. (1992)
suggested that washing and/or tape-stripping of the exposed area might remove
most of the 2,3,7,8-TCDD and reduce the potential for systemic exposure and
toxicity since most of the 2,3,7,8-TCDD remained within the horny layer of human
skin even at 24 hours following exposure.
Weber et al. (1991) also investigated the penetration of 2,3,7,8-TCDD into
fy
human cadaver skin at concentrations of 65-6.5 ng/cm . This study also found
that the stratum corneum acted as a protective barrier, as its removal increased
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the amount of 2,3,7,8-TCDD absorbed into layers of the skin. With intact skin
and acetone as the vehicle, the rate of penetration of 2,3,7,8-TCDD into the
f\
dermis ranged from 6-170 pg/hour/cm , while penetration into the dermis and
o
epidermis ranged from 100-800 pg/hour/cm . With mineral oil as the vehicle,
there was about a 5- to 10-fold reduction in the rate of penetration of 2,3,7,8-
TCDD into the intact skin.
1.1.2.1. BIOAVAILABILITY FOLLOWING DERMAL EXPOSURE — Dermal exposure of
human to 2,3,7,8-TCDD and related compounds usually occurs as a complex mixture
of these contaminants in soil, oils or other mixtures which would be expected to
alter absorption. Poiger and Schlatter (1980) presented evidence that the
presence of soil or lipophilic agents dramatically reduces dermal absorption of
2,3,7,8-TCDD compared to absorption of pure compound dissolved in solvents. In
a control experiment, 26 ng of 2,3,7,8-TCDD in 50 pL methanol was administered
to the skin of rats, and 24 hours later the liver contained 14.8±2.6% of the
dose. By comparing this value to the hepatic levels obtained after oral
administration in 50% ethanol (in the same study), the amount absorbed from a
dermal application can be estimated at -40% of the amount absorbed from an
equivalent oral dose. This comparison assumes that hepatic levels are valid
estimates of the amount absorbed from both oral and dermal routes and that
absorption from methanol is equivalent to absorption from 50% ethanol. The dose-
dependent distribution of 2,3,7,8-TCDD in the liver is another factor that may
limit quantitative conclusions regarding bioavailability which are based solely
on hepatic levels following exposure to 2,3,7,8-TCDD. As compared with dermal
application in methanol, dermal application of 2,3,7,8-TCDD to rats in vaseline
or polyethylene glycol reduced the percentage of the dose in hepatic tissue to
1.4 and 9.3%, respectively, but had no observable effect on the dose of 2,3,7,8-
TCDD required to induce skin lesions (-1 pg/ear) in the rabbit ear assay.
Application of 2,3,7,8-TCDD in a soil/water paste decreased hepatic 2,3,7,8-TCDD
to -2% of the administered dose and increased the amount required to produce skin
lesions to 2-3 pg in rats and rabbits, respectively. Application in an activated
carbon/water paste essentially eliminated absorption, as measured by percent of
dose in the liver, and increased the amount of 2,3,7,8-TCDD required to produce
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skin lesions to -160 pg. These results suggest that the dermal absorption and
acnegenic potency of 2,3,7,8-TCDD depend on the formulation (vehicle or
adsorbent) containing the toxin.
Shu et al. (1988) investigated the dermal absorption of soil-bound 2,3,7,8-
TCDD in rats. Relative dermal bioavailability was estimated by comparing the
level of 2,3,7,8-TCDD in the liver of rats given soil-bound 2,3,7,8-TCDD dermally
to that of rats given oral doses of 2,3,7,8-TCDD dissolved in corn oil. The
level of 2,3,7,8-TCDD in livers of rats dosed orally with 2,3,7,8-TCDD in corn
oil, following correction for unabsorbed 2,3,7,8-TCDD, is assumed to represent
100% bioavailability. The dermal penetration of 2,3,7,8-TCDD after 4 hours of
contact with skin was -60% of that after 24 hours of contact. After 24 hours of
contact with the skin, the degree of dermal uptake from contaminated soil was -1%
of the administered dose. The authors observed that the degree of uptake does
not appear to be influenced significantly by the concentration of 2,3,7,8-TCDD
in soil, the presence of crankcase oil as co-contaminants or by environmentally-
versus laboratory-contaminated soil.
A major limitation of the above studies is the uncertainty regarding the
extrapolation of dermal absorption data on these compounds from the rat to the
human. The in vitro dermal uptake of 2,3,7,8-TCDD has been investigated in
hairless mouse and human skin (Gallo et al., 1992; Rahman et al., 1992). In
vitro dermal uptake of 2,3,7,8-TCDD from laboratory-contaminated soil found that
aging of soils (up to 4 weeks) and the presence of additives (2,4,5-trichloro-
phenol and motor oil) in the soil did not have any significant effect of dermal
uptake (Gallo et al., 1992). Since most of the 2,3,7,8-TCDD remained in the
stratum corneum layer of human skin, the permeation of 2,3,7,8-TCDD was signifi-
cantly lower in human than in hairless mouse skin. Although there are no
published quantitative in vivo data on the dermal absorption of 2,3,7,8-TCDD and
related compounds in the human, there are very limited data on the rhesus monkey.
Brewster et al. (1988) found that 1,2,3,7,8-PeCDF was poorly absorbed in the
monkey after dermal application with <1% of the administered dose being absorbed
in 6 hours. This provides further evidence for the very slow rate of dermal
absorption of 2,3,7,8-TCDD and related compounds.
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1.1.3. Transpulmonary Absorption. The use of incineration as a means of solid
and hazardous waste management results in the emission of contaminated particles
that may contain TCDD and related compounds into the environment. Thus,
significant exposure to TCDD and related compounds may result from inhalation of
contaminated fly ash, dust and soil. In an attempt to address the bioavail-
ability and potential health implications of inhaling contaminated particles,
Nessel et al. (1990) examined the potential for transpulmonary absorption of TCDD
after intratracheal instillation of the compound administered to female Sprague-
Dawley rats either in a corn oil vehicle or as a laboratory-prepared contaminant
of gallium oxide particles. Several biomarkers of systemic absorption were
measured, including the dose-dependent effects of TCDD on hepatic microsomal
cytochrome P-450 content, AHH activity and liver histopathology. Significant
dose-related effects were observed at an exposure of >0.55 fjg TCDD/kg. The
authors found that induction was slightly higher when animals received TCDD in
corn oil than when animals received TCDD-contaminated particles and was compar-
able to induction after oral exposure. The results from Nessel et al. (1990)
indicate that systemic effects occur after pulmonary exposure to TCDD, suggesting
that transpulmonary absorption of TCDD does occur.
The transpulmonary absorption of 2,3,7,8-TCDD was assessed in male Fischer
344 rats following intratracheal instillation of a 1 nmol/kg dose in Emulphor:
ethanol:water (Diliberto et al., 1992). Transpulmonary absorption was -92%,
suggesting that there was almost complete absorption of 2,3,7,8-TCDD by inhala-
tion under these conditions. Similar results were also observed for the
transpulmonary absorption of 2,3,7,8-TBDD under similar exposure conditions
(Diliberto et al., 1991). These results suggest that the transpulmonary
absorption of 2,3,7,8-TCDD and 2,3,7,8-TBDD was similar to that observed
following oral exposure.
1.1.4. Parenteral Absorption. In an effort to obtain more reproducible and
complete absorption of 2,3,7,8-TCDD and related compounds for pharmacokinetic
studies, Abraham et al. (1989) investigated the absorption of 2,3,7,8-TCDD after
parenteral application in rats, using various vehicles. These investigators
observed optimal results with the subcutaneous injection of 2,3,7,8-TCDD using
a mixture of toluene:DMSO (1:2) as vehicle. At 3 and 5 days after treatment, the
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percentages of administered dose remaining at the injection site under the skin
of the back were -10 and 2%, respectively. The vehicle did not cause adverse
effects at an applied volume of 0.2 mL/kg bw. The absorption of a defined
mixture of CDDs and CDFs in the rat was also examined after subcutaneous
injection using toluene:DMSO (1:2) as a vehicle. Of the 97 congeners analyzed,
70 were >95% absorbed 7 days after exposure, 21 were 90-95% absorbed and 1,2,3,9-
TCDD, 1,2,3,6,7,9-/ 1,2,3,6,8,9-HxCDD, 1,2,3,4,6,7,9-HpCDD, OCDD, 1,2,4,6,8,9-
HxCDF and 1,2,3,7,8,9-HxCDF were 84-89% absorbed. Greater than 90% absorption
of CDDs and CDFs was also observed under these conditions in the marmoset monkey,
with the exception of 1,2,3,4,7,8,9-HpCDF, OCDF and OCDD, which had -50-80% of
the administered dose absorbed (Neubert et al., 1990; Abraham et al., 1989).
Although the absorption of CDDs and CDFs after subcutaneous administration in
toluene:DMSO (1:2) is somewhat slow, in rats and monkeys, absorption of most
congeners was >90% within 7 days. Even for highly chlorinated insoluble
congeners, such as OCDD and OCDF, subcutaneous absorption was >84% in the rat and
>50% in the monkey.
Less complete and slower absorption of CDDs and CDFs was observed after
subcutaneous injection of these compounds using an oil-containing vehicle
(Brunner et al., 1989; Abraham et al., 1989). Using a corn oil:acetone vehicle
(24:1, v/v), Lakshmanan et al. (1986) observed that only 7% of the administered
dose of 2,3,7,8-TCDD was absorbed 24 hours after subcutaneous injection and that
only 35% was absorbed after intraperitoneal injection. Also, Brunner et al.
(1989) reported that intraperitoneal administration of CDDs and CDFs revealed a
delayed absorption from the abdominal cavity which varied for the different
congeners. Therefore, concentrations measured in abdominal adipose tissue after
intraperitoneal administration may not represent average values of adipose tissue
in the whole body, particularly at early time points following exposure.
1.2. DISTRIBUTION
1.2.1. Distribution in Blood and Lymph. Once a compound is absorbed, its
distribution is regulated initially by its binding to components in blood and its
ability to diffuse through blood vessels and tissue membranes. Lakshmanan et al.
(1986) investigated the absorption and distribution of 2,3,7,8-TCDD in thoracic
duct-cannulated rats. Their results suggest that following gastrointestinal
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absorption, 2,3,7,8-TCDD is absorbed primarily by the lymphatic route and is
transported predominantly by chylomicrons. Ninety percent of the 2,3,7,8-TCDD
in lymph was associated with the chylomicron fraction. The plasma disappearance
of 2,3,7,8-TCDD-labeled chylomicrons followed first-order decay kinetics, with
67% of the compound leaving the blood compartment very rapidly (t 1^=0.81
minutes), whereas the remainder of the 2,3,7,8-TCDD had a ti^ of 30 minutes.
2,3,7,8-TCDD was then found to distribute primarily to the adipose tissue and the
liver.
In vitro studies have investigated the distribution of 2,3,7,8-TCDD in human
whole blood. Henderson and Patterson (1988) found -80% of the compound associ-
ated with the lipoprotein fraction, 15% associated with protein (primarily human
serum albumin) and 5% associated with cellular components. Theoretical and
limited experimental data also suggest that 2,3,7,8-TCDD and related compounds
may be associated with plasma prealbumin (McKinney et al., 1985; Pedersen et al.,
1986). The distribution of [%]-2,3,7,8-TCDD among lipoprotein fractions from
three fasting, normolipemic donors indicated a greater percentage associated with
LDL (55.3±9.03% SD) than with VLDL (17.4±9.07% SD) or HDL (27.3±10.08% SD). The
distribution of 2,3,7,8-TCDD among the lipoprotein fractions was similar to that
reported earlier by Marinovich et al. (1983). When the binding of 2,3,7,8-TCDD
was calculated per mole of lipoprotein, it was suggested that the maximal binding
capacity was exerted by VLDL, followed by LDL and HDL (Marinovich et al., 1983).
The results also suggest that variations in the amounts of each lipoprotein class
may alter the distribution of 2,3,7,8-TCDD among lipoproteins in a given subject.
Significant species differences also exist; in the case of the rat, which has
markedly lower plasma lipids compared to humans, 2,3,7,8-TCDD was distributed
almost equally among the lipoprotein fractions (Marinovich et al., 1983).
In addition, there is indirect evidence that suggests that the binding of
2,3,7,8-TCDD to lipoproteins may alter the pharmacokinetics and toxic potency of
the compound. Marinovich et al. (1983) found that experimentally induced
hyperlipidemia in rats delayed the development of overt toxicity (lethality).
However, the disposition of 2,3,7,8-TCDD was not investigated under these
conditions. These investigators suggest that the release of lipoprotein-bound
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2,3,7,8-TCDD is related to the metabolic turnover of lipoproteins. In hyperlipi-
demic rats, the turnover of VLDL and LDL is delayed significantly compared to
normolipidemic animals, and this may contribute to the plasma lipoprotein binding
modifying the toxicity of 2,3,7,8-TCDD in hyperlipidemic rats.
The uptake of lipoprotein-associated 2,3,7,8-TCDD by cultured human
fibroblasts found the time- and temperature-dependent cellular uptake was
greatest from LDL, intermediate from HDL and least from serum (Shireman and Wei,
1986). Decreased cellular uptake of LDL and 2,3,7,8-TCDD was observed in mutant
fibroblasts, which lack the normal cell membrane receptor for LDL. This provides
some evidence that specific binding of LDL and the LDL receptor pathway may
account for some of the rapid early uptake of 2,3,7,8-TCDD with LDL entry. The
results suggest that the entry of 2,3,7,8-TCDD into cells may not be solely by
simple diffusion. However, nonspecific binding of the LDL and transfer of
2,3,7,8-TCDD from LDL to the cell membranes are probably also important, since
significant time- and temperature-dependent uptake of 2,3,7,8-TCDD and LDL
occurred in the mutant fibroblasts.
Thus, upon absorption, 2,3,7,8-TCDD and probably related compounds are bound
to chylomicrons, lipoproteins and other serum proteins that assist in distri-
buting these uncharged, lipophilic compounds throughout the vascular system.
These compounds then partition from blood components into cellular membranes and
tissues, probably largely by passive diffusion. In addition, cellular uptake may
be facilitated partly through the cell membrane LDL receptor, the hepatic
receptor for albumin (Weisiger et al., 1981) and/or other systems.
1.2.2. Tissue Distribution. Once absorbed into blood, 2 , 3, 7,8-TCDD and related
compounds readily distribute to all organs. Tissue distribution within the first
hour after exposure parallels blood levels and reflects physiological parameters
such as blood flow to a given tissue and relative tissue size. For example, high
initial concentrations of 2,3,7,8-TCDD, 1,2,3,7,8-PeCDF and 3,3',4,4'-TCB were
observed in highly perfused tissue such as the adrenal glands during the 24-hour
period after a single exposure (Birnbaum et al., 1980; Olson et al., 1980;
Pohjanvirta et al., 1990; Brewster and Birnbaum, 1988; Durham and Brouwer, 1990).
A high percentage of the dose of 2,3,7,8-TCDF and 1,2,3,7,8-PeCDF was also found
in muscle within the first hour after intravenous exposure, due to the large
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volume of this tissue (Birnbaum et al., 1980; Birnbaum, 1985; Brewster and
Birnbaum, 1988). Nevertheless, within several hours the liver, adipose tissue
and skin become the primary sites of disposition, when expressed as percent of
administered dose per g tissue and as percent of dose per organ. Liver, adipose
tissue, skin and thyroid were the only tissue to show an increase in the concen-
tration of 2,3,7,8-TCDD during the initial 4 days after a single intraperitoneal
exposure of rats (Pohjanvirta et al., 1990). In this study, a similar general
pattern of disposition was observed in Han/Wistar and Long-Evans rats which are
respectively most resistant and susceptible to the acute toxicity of 2,3,7,8-TCDD
(Pohjanvirta et al., 1990).
Table 1-4 illustrates the tissue distribution of 2,3,7,8-TCDD in female
Wistar rats 7 days after a single subcutaneous exposure (Abraham et al., 1988).
This general pattern of distribution is similar to that observed in mice, rats,
rhesus monkeys, hamsters and guinea pigs, where liver and adipose tissue
consistently have the highest concentrations of 2,3,7,8-TCDD (Piper et al., 1973;
Fries and Marrow, 1975; Rose et al., 1976; Allen et al., 1975; Van Miller et al.,
1976; Kociba et al., 1978a,b; Gasiewicz et al., 1983; Manara et al., 1982; Olson
et al., 1980; Gasiewicz and Neal, 1979; Birnbaum, 1986; Pohjanvirta et al., 1990;
Abraham et al., 1988). A similar pattern of disposition also was observed for
2,3,7,8-TCDF in the guinea pig, rat, C57BL/6J and DBA/2J mouse and rhesus monkey,
with 2,3,7,8-TCDF concentrations highest in liver and adipose tissue (Decad et
al., 1981b; Birnbaum et al., 1980, 1981). In summary, there do not appear to be
major species or strain differences in the tissue distribution of 2,3,7,8-TCDD
and 2,3,7,8-TCDF, with the liver and adipose tissue being the primary disposition
sites.
The tissue distribution of the coplanar PCBs and PBBs also appears to be
similar to that of 2,3,7,8-TCDD and 2,3,7,8-TCDF. Limited studies in rats and
mice found that 3,3',4,4'-TCB, 3,3•,4,4'-TBB and 3,3'4,4'5, 5'-HxBB distributed
preferentially to adipose tissue and liver (Clarke et al., 1983, 1984; Millis et
al., 1985; Wehler et al., 1989; Clevenger et al., 1989).
While the liver and adipose tissue contain the highest concentrations of
2,3,7,8-TCDD and 2,3,7,8-TCDF, there are some congener-specific differences in
the relative tissue distribution of related compounds. 2,3,7,8-TBDD and
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TABLE 1-4
Tissue Distribution of [14C]-2-3,7,8-TCDD in
Female Wistar Ratsa'b
Tissue
Liver
Adipose tissue
Adrenal glands
Ovaries
Thymus
Skin
Lung
Kidney
Pancreas
Spleen
Serum
Bone (with marrow)
Muscle
Brain
Range of 2,3,7,8-TCDD Concentrations
(ng/g)
29.23-30.99
3.72-4.14
0.89-1.08
0.76-0.96
0.60-1.05
0.64-0.68
0.32-0.33
0.27-0.29
0.21-0.31
0.18-0.23
0.16-0.18
0.16-0.16
0.08-0.12
0.07-0.09
aSource: Abraham et al., 1988
''Distribution was assessed 7 days
single subcutaneous exposure (3 jug/kg bw)
after
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1,2,3,7,8-PeCDD disposition in the rat was very similar to that of 2,3,7,8-TCDD
(Kedderis et al., 1991; Wacker et al., 1986). The hepatic concentration of OCDD
and 2,3,4,7,8-PeCDF in the rat, however, was approximately 10- to 20-fold greater
than that in adipose tissue, which generally contains the second highest levels
of these compounds (Birnbaum and Couture, 1988; Norback et al., 1975; Williams
et al., 1972; Brewster and Birnbaum, 1987). The tissue distribution of a defined
mixture of CDDs and CDFs (28.8 nq CDDs+CDFs/kg bw containing 120 ng 2,3,7,8-
TCDD/kg bw) was measured in marmoset monkeys 7 days after a single subcutaneous
exposure (Abraham et al., 1990). For most of the 2,3,7,8-substituted congeners,
the highest concentrations were detected in hepatic and adipose tissue, with
correspondingly lower values detected in kidney, brain, lung, heart, thymus or
testes. The hepatic and adipose tissue concentrations were similar for 2,3,7,8-
TCDD, 1,2,3,7,8-PeCDD, 2,3,7,8-TCDF and 1,2,3,7,8-/ 1,2,3,4,8-PeCDF. Nonethe-
less, the hepatic concentrations were approximately 10-fold or more greater than
those of adipose tissue for 1,2,3,4,7,8-HxCDD, 1,2,3,6,7,8-HxCDD, 1,2,3,7,8,9-
HxCDD, 1,2,3,4,6,7,8-HpCDD, OCDD, 2,3,4,7,8-PeCDF, 1,2,3,4,7,8-/1,2,3,4,7,9-
HxCDF, 1,2,3,6,7,8-HxCDF, 1,2,3,7,8,9-HxCDF, 2,3,4,6,7,8-HxCDF, 1,2,3,4,6,7,8-
HpCDF, 1,2,3,4,7,8,9-HpCDF and OCDF. The lungs and thymus contained higher
concentrations of all of the above congeners than were detected in kidney, brain,
heart and testes. Unexpectedly, the concentrations of the above HxCDDs, HpCDDs,
OCDD and HxCDFs were similar in the adipose tissue, lungs and thymus. In the
case of HpCDFs and OCDF, the concentrations were greater in the lungs than the
adipose tissue. The enhanced disposition of highly chlorinated congeners to the
lungs and thymus is of interest and deserves further investigation. For example,
it is possible that the high concentration in the lungs could be related to the
insolubility of these compounds.
Whole-body autoradiography of mice and rats after intravenous administration
of [ C]-2,3,7,8-TCDD showed a selective localization of radioactivity in the
liver and nasal olfactory mucosa (Appelgren et al., 1983; Gillner et al., 1987).
The selective localization of 2,3,7,8-TCDD in the nasal olfactory mucosa was
apparently overlooked by other distribution studies that only examined selected
organs. Gillner et al. (1987) found no 2,3,7,8-TCDD-derived radioactivity in the
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olfactory mucosa after solvent extraction of sections, suggesting that 2,3,7,8-
TCDD was not covalently bound in this tissue. In addition, Gillner et al. (1987)
reported induction of mRNA coding for cytochrome P-4501A2 in the absence of
P-4501A1 induction in olfactory mucosa of rats. The selective distribution of
2,3,7,8-TCDD in the liver and olfactory mucosa correlates with the tissue
specific induction of cytochrome P-4501A2, which represents a potential seques-
tration (binding) protein (see Section 1.2.5). Increases in the incidence of
squamous cell carcinoma of nasal turbinates and carcinoma of the liver were
observed in rats after a 2-year exposure to 2,3,7,8-TCDD in rat chow (Kociba et
al., 1978); however, this effect was not observed in nasal tissues of mice or
rats intubated with 2,3,7,8-TCDD. Gillner et al. (1987) suggested that 2,3,7,8-
TCDD may not be an initiator in this tissue and indicated that future studies
should investigate the possibility that 2,3,7,8-TCDD may act as a promoter or
cocarcinogen in nasal tissue.
Evidence has also been reported that suggests that 2,3,7,8-TCDD uptake and
retention by the liver is dependent on the cell type within the liver. Hakansson
et al. (1989) found that at 4 days after exposure of rats to 2,3,7,8-TCDD, 60%
of the dose distributed to hepatocytes and 12% was retained by stellate cells.
Half-lives for 2,3,7,8-TCDD in hepatocytes and stellate cells were also
calculated to be 13 and 50 days, respectively, suggesting that 2,3,7,8-TCDD is
more persistent in nonparenchymal cells. Further studies are needed to
understand the pharmacokinetic and pharmacodynamic significance of the cell-
specific distribution of 2,3,7,8-TCDD and related compounds.
1.2.2.1. TISSUE DISTRIBUTION IN HUMANS — Fachetti et al. (1980) reported
tissue concentrations of 2,3,7,8-TCDD at levels of 1-2 ng/g in adipose tissue and
pancreas, 0.1-0.2 ng/g in the liver and <0.1 ng/g in thyroid, brain, lung, kidney
and blood in a woman who died 7 months after potential exposure to 2,3,7,8-TCDD
from the Seveso accident. This pattern of 2,3,7,8-TCDD distribution, however,
may not be representative for humans since the woman at the time of death had an
adenocarcinoma (which was not considered related to the accident) involving the
pancreas, liver and lung.
1-20 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
Ryan et al. (1985a) examined the distribution of 2,3,7,8-TCDD in two humans
at autopsy. They determined on a weight basis that 2,3,7,8-TCDD distributed in
descending order to fat (-6 ppt) and liver (-2 ppt), with levels in muscle and
kidney below detection; however, 2,3,7,8-TCDD levels compared on a per lipid
basis were similar between tissues. These data should be interpreted with
caution, since only two subjects were examined and one of the subjects was
suffering from fatty liver syndrome; therefore, the data cannot be generalized
to the entire population.
Poiger and Schlatter (1986) estimated that -90% of the body burden of
2,3,7,8-TCDD was sequestered in the fat after a volunteer ingested ^H-2,3,7,8-
TCDD in corn oil at a dose of 1.14 ng/kg. During this 135-day study, elevated
radioactivity was detected in the blood only during the first 2 days after
treatment. The data would be consistent with the high lipid bioconcentration
potential of 2,3,7,8-TCDD in humans, as calculated by Geyer et al. (1986) from
daily intake assumptions, levels in human adipose tissue and pharmacokinetic
models. Geyer et al. (1986) estimated a BCF of between 104 and 206 for 2,3,7,8-
TCDD in human adipose tissue.
In human adipose tissue, levels of 2,3,7,8-TCDD averaging 5-10 ppt have been
reported for background populations in St. Louis, MO, by Graham et al. (1986),
in Atlanta, GA, and Utah by Patterson et al. (1986), and in Canada by Ryan et al.
(1985b). Sielken (1987) evaluated these data and concluded that the levels of
2,3,7,8-TCDD in human adipose are log-normally distributed and positively
correlated with age. Among the observed U.S. background levels of 2,3,7,8-TCDD
in human adipose tissue, more than 10% were >12 ppt.
Patterson et al. (1987) developed a HRGC/HRMS analysis for 2,3,7,8-TCDD in
human serum. The arithmetic mean of the individual human serum samples was 47.9
ppq on a whole-weight basis and 7.6 ppt on a lipid-weight basis. Paired human
serum and adipose tissue levels of 2,3,7,8-TCDD have been compared by Patterson
et al. (1988) and Kahn et al. (1988). Both laboratories reported a high
correlation between adipose tissue and serum 2,3,7,8-TCDD levels when the samples
were adjusted for total lipid content. This correlation indicates that serum
1-21 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
2,3,7,8-TCDD is a valid estimate of the 2,3,7,8-TCDD concentration in adipose
tissue.
In a study of potentially heavily exposed Vietnam veterans, MMMR (1988)
reviewed an Air Force study of Ranch Hand veterans who were either herbicide
loaders or herbicide specialists in Vietnam. The mean serum 2,3,7,8-TCDD levels
of 147 Ranch Hand personnel was 49 ppt in 1987, based on total lipid-weight,
while the mean serum level of the 49 controls was 5 ppt. In addition, 79% of the
Ranch Hand personnel and 2% of the controls had 2,3,7,8-TCDD levels >10 ppt. The
distribution of 2,3,7,8-TCDD levels in this phase of the Air Force health study
indicates that only a small number of Ranch Hand personnel had unusually heavy
2,3,7,8-TCDD exposure. This report also estimated the half-life of 2,3,7,8-TCDD
in humans to be -7 years on the basis of 2,3,7,8-TCDD levels in serum samples
taken in 1982 and 1987 from 36 of the Ranch Hand personnel who had 2,3,7,8-TCDD
levels >10 ppt in 1987. Similar results were obtained by Kahn et al. (1988) who
compared 2,3,7,8-TCDD levels in blood and adipose tissue of Agent Orange-exposed
Vietnam veterans and matched controls (Kahn et al., 1988). This study also
examined moderately exposed Vietnam veterans who handled herbicides regularly
while in Vietnam. Although this study can distinguish moderately exposed men
from others, the data do not address the question of identifying persons whose
exposures are relatively low and who constitute the bulk of the population, both
military and civilian, who may have been exposed to greater than background
levels of 2,3,7,8-TCDD.
1.2.3. Time-Dependent Tissue Distribution. 2,3,7,8-TCDD and related compounds
exhibit congener specific disposition, which depends on tissue, species and time
after a given exposure. In general, these compounds are cleared rapidly from the
blood and distributed to liver, muscle, skin, adipose tissue and other tissues
within the first hour(s) after exposure. This is followed by redistribution to
the liver and adipose tissue, which exhibit increasing tissue concentrations over
several days after exposure. Elimination from tissues then occurs at rates that
are congener-, tissue- and species-specific. Thus, the ratio of the concentra-
tion of 2,3,7,8-TCDD and related compounds in different tissues (i.e., liver/
adipose) may not remain constant over an extended period after a single exposure.
Abraham et al. (1988) examined the concentrations of 2,3,7,8-TCDD in liver and
1-22 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
adipose tissue of female Wistar rats over a 91-day period after a single
subcutaneous exposure at a dose of 300 ng/kg bw (Figure 1-1). The maximum
concentration of 2,3,7,8-TCDD in the liver and adipose tissue was reached at 3
and 7 days after exposure, respectively. The liver/adipose tissue concentration
ratio does not remain constant over time since the concentration of 2,3,7,8-TCDD
decreases more rapidly in the liver than in the adipose tissue. For example, the
liver/adipose tissue concentration ratio (for 2,3,7,8-TCDD) was 10.3 at 1 day
after exposure and 0.5 at 91 days after exposure (Figure 1-1). Results from
other disposition studies also indicate that the ratio of the concentration of
2,3,7,8-TCDD and related compounds in liver, adipose tissue and other tissues
does not remain constant over an extended period after a single exposure
(Pohjanvirta et al., 1990; Birnbaum, 1986; Birnbaum et al., 1980; Decad et al.,
1981a; Birnbaum and Couture, 1988; Olson et al., 1980; Kedderis et al., 1991;
Brewster and Birnbaum, 1987, 1988; Neubert et al., 1990). This relationship is
important in attempting to correlate dose-response data with tissue concentra-
tions of 2,3,7,8-TCDD and related compounds.
In an attempt to maintain constant 2,3,7,8-TCDD levels in tissues to study
long-term effects, Krowke et al. (1989) investigated several loading-dose/
maintenance-dose exposure regimens. They found that similar liver/adipose tissue
concentrations ranging from 5-8 could be maintained in rats over a 22-week period
using a loading dose of 25 /jg/kg followed by weekly maintenance doses of 5 ^g/kg.
A large body of data on the tissue concentrations of 2,3,7,8-TCDD and
related compounds over time after exposure can be evaluated by estimating
congener-specific half-life values for a given tissue and species. Table 1-5
summarizes pharmacokinetic elimination parameters for 2,3,7,8-TCDD and related
compounds from major tissue depots. Data from Abraham et al. (1988) (see Figure
1-1) were used to estimate the half-life for 2,3,7,8-TCDD in the liver and
adipose tissue of rats (Table 1-5). The decrease in the 2,3,7,8-TCDD concentra-
tion in adipose tissue is a linear function in the semi-logarithmic plot in
Figure 1-1 (log concentration versus time), which indicates apparent first-order
elimination kinetics with a half-life of 24.5 days (Table 1-5). Liver tissue
exhibits a biphasic (two-component) exponential decay pattern with a half-life
of 11.5 days for the first component (days 10-49) and a half-life of 16.9 days
1-23 08/11/92
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PRAFT—DO NOT QUOTE OR CITE
A liter hssuf
A odipost I issue
0 10
0.01
0 7 14 21 28 35 42 49 56 63 70 77 84 91
«t»s tiler IrettiKitl
FIGURE 1-1
Time Course of the Concentration of 14C-TCDD in Rat Liver and Adipose Tissue
After a Single Subcutaneous Injection of 300 ng TCDD/kg bw to Female Rats (M±SD).
Source: Abraham et al., 1988
1-24
08/11/92
-------
TABLE 1-5
• Elimination of 2,3,7,8-TCDD and Related Compounds from Major Tissue Depots
Chemical
CDDs
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
Species (Sex)
Dose
T i ssue
Half-Life
(days)
Remarks
Reference
Wistar rat (F)
Wistar rat (M)
Sprague-Dawley rat (H)
Sprague-Dawley rat (F)
C57BL/6J mice (M)
Ahb/Ahr
C57BL/6J mice
-------
ro
o
00
TABLE 1-5 (cont.)
Chemical
OCDD
Species (Sex)
Fischer 344 rat (M)
BDDs
2,3,7,8-TBDD
CDFs
2,3,7,8-TCDF
2,3,7,8-TCDF
2,3,7,8-TCDF
Fischer 344 rat (M)
Fischer 344 rat (M)
C57BL/6J mice (M)
DBA/2J mice (M)
Dose
50 /ig/kg, i.v.
T i ssue
liver
adipose
skin
Half-Life
(days)
84
38
3
69
0.5 fig/kg, i.v.
(0.001 junol/kg)
liver
adipose
skin
muscle
blood
4.5
16.5
57.8
2.5
57.8
1.6
26.7
18.2
30.6 /ig/kg, i.v.
(0.1 /unol/kg)
30.6 jig/kg, i.v.
(0.1 /unol/kg)
30.6 /ig/kg, i.v.
(0.1 /unol/kg)
liver
adipose
skin
muscle
blood
liver
adipose
skin
muscle
liver
adipose
muscle
0.10
1.25
3.75
0.45
11.09
0.02
0.72
0.02
1.14
1.9
1.6
0.15
4.0
0.015
1.1
1.8
7.0
0.02
4.0
Remarks
Pool size (% of total dose):
72.7
7.1
9.0, 1st component
0.3, 2nd component
Reference
Birnbaum and
Couture, 1988
1st component
2nd component
1st component
2nd component
1st component
2nd component
Pool size (X of total dose)
29.09 1st component
31.39 2nd component
17.85
6.84 1st component
1.22 2nd component
24.85 1st component
6.73 2nd component
1.31 1st component
0.89 2nd component
1st component
2nd component
1st component
2nd component
1st component
2nd component
Kedderis et
al., 1991
Birnbaum et
al., 1980
Decad et a I.,
1981b
Brewster and
Birnbaum,
1988
8
•8
o
H
VO
-------
TABLE 1-5 (cont.)
Chemi ca I
1,2,3,7.8-PeCDF
1,2,3,7,8-PeCDF
2,3,4,7,8-PeCDF
2,3,4,7,8-PeCDF
1,2,3,6,7,8-HxCDF
Species (Sex)
Fischer 344 rat (M)
Sprague-Dawley rat (F)
Fischer 344 rat (M)
Sprague-Dawley rat (F)
Sprague-Dawley rat (F)
PCBs
S.S'A.A'-TCB
Sprague-Dawley rat (F)
Dose
34 jig/ kg, i ,v.
(0.1 pmol/kg)
4.0 /ig/kg, p.o.
34 /tg/kg, i.v.
(0.1 junol/kg)
5.6 jig/kg, p.o.
6.0 fig/kg, p.o.
T i ssue
liver
adipose
skin
muscle
adrenal
blood
liver
liver
adipose
skin
muscle
blood
liver
liver
Half-Life
(days)
1.36
25.72
12.91
1.32
14.53
0.03
6.96
0.14
2.36
0.07
12.42
3.3
193
69
0.62
1.23
0.04
0.51
9.84
0.04
1.32
55
108
73
Remarks
Pool size (X of total dose):
42.59 1st component
1.27 2nd component
10.19
7.14 1st component
1.49 2nd component
34.81 1st component
7.42 2nd component
0.26 1st component
0.02 2nd component
5.33 1st component
1.29 2nd component
69.8% of total dose
Pool size (X of total dose):
67.71
10.53
3.54 1st component
1.37 2nd component
29.40 1st component
2.01 2nd component
0.78 3rd component
3.18 1st component
0.37 2nd component
0.008 3rd component
78.3% of total dose
63.4% of total dose
5 mg/kg/day, p.o.,
for 21 days,
liver
adipose
0.8
2.5
21 -Day exposure produced steady
state with 300 ng/g in liver and 8
jig/9 in adipose tissue.
Elimination was assessed over a 22-
day post-exposure period.
Reference
Brewster and
Birnbaum,
1988
Van den Berg
et al., 1989
Brewster and
Birnbaum,
1987
Van den Berg
et al., 1989
Van den Berg
et al., 1989
CLarke et
al., 1984
D
O
•8
o
H
M
8
O
H
H
M
VO
10
-------
Chemical
3,3'4,«'-TCB
TABLE 1-5 (cont.)
Species (Sex)
ICR mice (M)
Dose
8 mg/kg, p.o., every
other day for 10
doses
T i ssue
liver
adipose
serum
Half-Life
(days)
2.15
2.60
1.07
Remarks
Steady state tissue concentrations:
1.5 itg/g
19.2 itg/g
0.04 /19/mL
Reference
Clevenger et
al., 1989
i.v. = Intravenous; s.c. = subcutaneous; i.p. = intraperitoneal; p.o. * per os
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DRAFT—DO NOT QUOTE OR CITE
for the second component (days 49-91) (see Figure 1-1 and Table 1-5). Results
of Abraham et al. (1988) and Lakshmanan et al. (1986) indicate that in the rats,
2,3,7,8-TCDD is more persistent in the adipose tissue than in the liver. This
is in contrast to the mouse, where liver and adipose tissue have similar half-
lives (Birnbaum, 1986). 2,3,7,8-TCDD is exceptionally persistent in the adipose
tissue of the rhesus monkey, with a half-life approximately 10- to 40-fold
greater than that observed in the rat and mouse (Bowman et al., 1989). Thus, the
relative persistence of 2,3,7,8-TCDD is tissue specific and exhibits marked
interspecies variability.
Most of the pharmacokinetic data on the relative persistence of other
congeners in Table 1-5 has been reported in rat studies, which limits inter-
species comparisons. Results in the rat suggest that the distribution and
elimination of 2,3,7,8-TBDD from tissue are similar to that of 2,3,7,8-TCDD. The
most persistent congeners are OCDD, 2,3,4,7,8-PeCDF and 1,2,3,6,7,8-HxCDF, which
distribute almost entirely to the liver. OCDD and 2,3,4,7,8-PeCDF also exhibit
similar elimination kinetics, with a relative half-life in the liver more than
2-fold greater than that in adipose tissue. The least persistent congeners are
2,3,7,8-TCDF, 1,2,3,7,8-PeCDF and 3,3'4,4'-TCB. These congeners exhibit similar
elimination kinetics in the rat with half-lives in the adipose tissue greater
than those in liver. The relative tissue distribution of these congeners varies,
however, with 2,3,7,8-TCDF and 1,2,3,7,8-PeCDF distributing primarily to the
liver, while 3,3',4,4'-TCB distributes predominantly to the adipose tissue.
The experimental tissue distribution and elimination data in Table 1-5 were
obtained after exposure to a single congener, while real world exposure to
2,3,7,8-TCDD and related compounds occurs as a complex mixture of congeners.
Recently, Neubert et al. (1990) examined the persistence of various CDDs and CDFs
in hepatic and adipose tissue of male and female marmoset monkeys. Animals
received a single subcutaneous exposure to a defined CDD/CDF mixture (total dose
of 27.8 /jg/kg bw), which contained 0.12 jjg 2,3,7,8-TCDD/kg bw. Using the I-TE
factors (NATO, 1988; U.S. EPA, 1989), the total administered dose corresponded
to 0.464 ng I-TE/kg bw. The concentrations of specific congeners in liver and
adipose tissue were measured at 1, 6, 16 or 28 weeks after exposure, and
elimination constants and half-lives were estimated assuming first-order kinetics
1-29 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
(Table 1-6). Data in Table 1-6 were determined from pregnant and nonpregnant
female and male marmosets (total of 12 animals) since no obvious differences in
tissue concentrations were observed among these groups. All 2,3,7,8-substituted
CDDs and CDFs were consistently more persistent in the adipose tissue than in the
liver of marmoset monkeys. In general, the persistence in adipose tissue was
from -1.3- to 2.0-fold greater than that in liver, with the exception of
l,2,3,4,7,8-/l,2,3,4,7,9-HxCDF, HpCDFs and OCDF, which were even more persistent
in adipose tissue. For the latter congeners and OCDD, there was marked variance
in half-life values, which may be due to delayed and incomplete absorption of the
exceptionally persistent congeners and the relatively short (28 weeks) period of
investigation. A significant species difference exists for OCDD and 2,3,4,7,8-
PeCDF, which, in contrast to the marmoset monkey, was found to be more persistent
in the liver of the rat, with half-lives more than 2-fold greater than that in
adipose tissue (Birnbaum and Couture, 1988; Brewster and Birnbaum, 1987) (see
Table 1-5). Further comparison of tissue elimination data in the rat (Table 1-5)
and monkey (Table 1-6) indicates that 2,3,7,8-TCDD, OCDD, 2,3,7,8-TCDF,
1,2,3,6,7,8-HxCDF and 2,3,4,7,8-PeCDF (adipose tissue only) are more persistent
in the marmoset monkey than in the rat. The exception to this relationship is
2,3,4,7,8-PeCDF, which is more persistent in rat liver, compared to the monkey.
The exposure of marmoset monkeys to a complex mixture of CDDs and CDFs
included exposure to both 2,3,7,8- and non-2,3,7,8-substituted congeners (Neubert
et al., 1990). One week after exposure to this complex mixture, the non-2,3,7,8-
substituted CDDs and CDFs were present in liver and adipose tissue in relatively
minor quantities when compared with 2,3,7,8-substituted congeners; however, non-
2,3, 7, 8-substituted compounds represented a considerable percent of the exposure
mixture. In this study, none of the non-2,3,7,8-substituted TCDDs, PeCDDs, TCDFs
or PeCDFs could be detected in the liver. Some of the hexa and hepta congeners
were detected in adipose tissue and liver, but after 1 week, the total amount in
the liver was >5% of the administered dose only in the case of 1,2,4,6,8,9-HxCDF.
Similar results were obtained in rats after exposure to a defined, complex
mixture of CDDs and CDFs (Abraham et al., 1989). Additional short-term studies
in rats provide evidence that the low tissue concentration of non-2,3,7,8-
substituted congeners, measured 1 week after exposure, were the result of rapid
1-30 08/11/92
-------
DRAFT—DO NOT QUOTE OR CITE
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1-31
08/11/92
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Congener
1,2,3,4,7,8.9-HpCDF
OCDF
TABLE 1-6 (cont.)
Hepatic Tissue
Ke 1
(weeks )
0.0088±0.0127
0.004010.0096
Half-Life
(weeks)
79
174
95% Conf . Interval
(weeks)
20-»d
30-»d
Adipose Tissue
K« 1
(weeks'1)
0.0011±0.0112
-0.0042±0.0148
Half-Life
(weeks)
660
«d
95% Conf. Interval
(weeks)
30-»d
28-d
9Source: Neubert et al., 1990
I
ui
NJ
Animals were treated subcutaneously with a single dose of a defined COO\CDF mixture, and the tissues were analyzed at different times following
treatment. Half-lives were calculated from tissue concentrations of the 2,3,7,8-substituted congeners in hepatic and adipose tissue. Values
are given as elimination rate constant K including estimated SD and half-life including 95% confidence intervals.
Calculated from the time period: >6 weeks after injection.
Calculated half-life is apparently infinite. Data for OCDD and OCDF are unreliable due to delayed absorption.
8Not detected in hepatic tissue 6 weeks after treatment; limits of detection used for calculation
f,
Due to interference
O
O
1
1
3
O
CO
NA - Not applicable
O
H
M
VO
ro
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DRAFT—DO NOT QUOTE OR CITE
elimination, since these congeners were detected at higher levels in the liver
13-14 hours after exposure (Abraham et al., 1989). These results in monkeys and
rats are compatible with data from analysis of human tissue samples and milk in
which the non-2,3,7,8-substituted congeners have also not been shown to be
present in significant concentrations, when compared with the 2,3,7,8-substituted
congeners (Schecter et al., 1985, 1986; Ryan, 1986; Rappe et al., 1986; Beck et
al., 1987, 1988; Thoma et al., 1989).
A potential problem of tissue distribution and elimination studies after
exposure to a complex mixture of CDDs and CDFs is the possible interaction of the
mixture during the uptake and elimination of specific congeners from tissues.
A similar hepatic distribution (-25% of dose) and liver/adipose tissue concentra-
tions ratio (~2) for 2,3,7,8-TCDD were observed in rats 7 days after exposure to
2,3,7,8-TCDD (100 ng/kg bw) when the compound was administered alone or in
combination with a large amount of other CDDs/CDFs (total 23,222 ng/kg bw)
(Abraham et al., 1988, 1989). This suggests that under these experimental
conditions, the tissue distribution of 2,3,7,8-TCDD was not altered when the
exposure included a complex mixture of CDDs/CDFs. Van den Berg et al. (1989)
studied the hepatic disposition and elimination of CDFs when administered
individually (see Table 1-5) and as mixtures. Co-administration of 1,2,3,7,8-
and 2,3,4,7,8-PeCDF resulted in 46% of the dose of 1,2,3,7,8-PeCDF distributing
to the liver, while 70% was distributed to the liver after administration of the
single compound (see Table 1-5). Nevertheless, this combined exposure did not
alter the rate of elimination of 1,2,3,7,8-PeCDF from the liver. Co-administra-
tion of 2,3,4,7,8-PeCDF and 1,2,3,6,7,8-HxCDF did not alter the hepatic uptake
of either congener or the hepatic elimination of 2,3,4,7,8-PeCDF but increased
the hepatic half-life of 1,2,3,6,7,8-HxCDF to 156 days from the single compound
exposure half-life of 73 days (see Table 1-5). However, these values must be
considered rough estimates since the experimental period of 42 days was too short
to accurately calculate half-lives. Although there are few investigations of
potential interactions of mixtures of CDDs and CDFs on the uptake and elimination
of individual congeners, the limited available data suggests that exposure to
complex mixtures (see Table 1-6) may alter the tissue disposition of individual
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congeners. There is clearly a need for more understanding of possible pharmaco-
kinetic interactions of complex mixtures of these and other compounds.
1.2.4. Dose-Dependent Tissue Distribution. Recent evidence suggests that the
tissue distribution of 2,3,7,8-TCDD and related compounds is dose dependent.
Abraham et al. (1988) investigated the distribution of 2,3,7,8-TCDD in liver and
adipose tissue of rats 7 days after a single subcutaneous exposure to 2,3,7,8-
TCDD at doses of 1-3000 ng/kg bw. Greater than 97% of the administered 2,3,7,8-
TCDD was absorbed at all doses, with the exception of the 3000 ng/kg group where
84% of the dose was absorbed. Figure 1-2 illustrates the dose-dependent
disposition of 2,3,7,8-TCDD in liver and adipose tissue (% dose/g) 7 days after
exposure. A sharp increase in 2,3,7,8-TCDD concentration in liver was observed
at exposure levels >10 ng/kg bw. Disposition in the liver increased from -11%
of the administered dose at an exposure level of 1-10 ng/kg bw to -37% of the
dose at an exposure level of 300 ng/kg bw. The increase in distribution to the
liver was accompanied by a dose-related decrease in the concentration of 2,3,7,8-
TCDD in the adipose tissue. As a result, the liver/adipose tissue concentration
ratio for 2,3,7,8-TCDD at 7 days after exposure increased with increasing doses,
starting at an exposure level of 30 ng/kg bw (Table 1-7). Thus, the tissue-
specific disposition of 2,3,7,8-TCDD is regulated by a complex relationship,
which includes species, time after a given exposure and dose (see Figures 1-1 and
1-2; Tables 1-5 and 1-6).
Other studies on the tissue disposition of 2,3,7,8-TCDD and related
compounds report similar dose-dependent behavior with disproportionally greater
concentrations in the liver at high doses compared with low doses. Poiger et al.
(1989) observed a dose-related increase in distribution to the liver (% of
dose/liver) and/or an increase in the liver/adipose tissue concentration ratio
for 2,3,7,8-TCDD, 2,3,4,7,8-PeCDF, 1,2,3,7,8-PeCDF and 1,2,3,6, 7,8-HxCDF in the
rat. Kedderis et al. (1991a) also observed a dose-related increase in hepatic
disposition (1.27 versus 10.05 % of dose/liver) and an increase in the liver/
adipose tissue concentration ratio (0.16 versus 2.59) for 2,3,7,8-TBDD at 56 days
after exposure at doses of 0.001 and 0.1 /jmol/kg bw, respectively. In a related
study, pretreatment of mice with 2,3,7,8-TCDD (5 or 15 /jg/kg) produced a dose-
related, enhanced hepatic accumulation of a subsequent oral dose of 2,3,7,8-TCDD
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FIGURE 1-2
Dose Dependency of the Percentage of the Administered Dose of 14C-TCDD/g of
Tissue Recovered in Liver and Adipose Tissue After Single Subcutaneous Doses
(values from animals treated with 3000 ng TCDD/kg bw were corrected for 84%
absorption). Concentrations were measured 7 days after the injection.
Source: Abraham et al., 1988
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TABLE 1-7
2,3,7,8-TCDD Concentrations in Liver and Adipose Tissue Following Different Doses and Calculated
Concentration Ratios (Liver/Adipose Tissue)8'
Dose
(ng/kg)
1
3
10
30
100
300
1000
3000
Number
6
6
12
6
6
6
6
5
TCDD Concentration
Liver
(ng/g)
0.0031*0.0009
0.0102±0.0020
0.0406±0.0121
0.162±0.032
0.699*0.130
3.38±0.22
10.7*2.2
27.9±2.4
TCDD Concentration
Adipose Tissue
(ng/g)
ND
0.0139±0.0015
0.0494*0.0084
0.139*0.021
0.335*0.065
0.819*0.075
2.02*0.17
3.66*0.31
Concentration Ratio:
Liver/Adipose Tissue
NA
0.74*0.15
0.82*0.20
1.16*0.07
2.10*0.27
4.14*0.31
5.27*0.96
7.65*0.64
Source: Abraham et al., 1988
Concentrations were measured 7 days after injection
ND = Not detectable; NA = not applicable
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(Curtis et al., 1990). Similarly, a dose-related increase in hepatic uptake of
[12^I]-2-iodo-3,7,8-trichlorodibenzo-p-dioxin was observed after pretreatment of
mice with 2,3,7,8-TCDD (Poland et al., 1989; Leung et al., 1990). Shen and Olson
(1987) also observed an increase in the uptake of 2,3,7,8-TCDD by isolated
hepatocytes from 2,3,7,8-TCDD pretreated mice.
Chronic studies also support dose-dependent alterations in the tissue
distribution of these compounds. Kociba et al. (1978a,b) found that female rats
maintained on a daily dietary 2,3,7,8-TCDD intake of 100 ng for 2 years had an
average 2,3,7,8-TCDD content of 8100 ppt in fat and 24,000 ppt in the liver.
Rats given 10 ng/kg/day had an average of 1700 ppt 2,3,7,8-TCDD in the fat and
5100 ppt in the liver. For both of these exposures the liver/ adipose tissue
concentration ratio of 2,3,7,8-TCDD was -3. At the lowest dose level of 1
ng/kg/day, both fat and liver contained an average of 540 ppt 2,3,7,8-TCDD.
Kociba et al. (1976) presented evidence that steady state had been reached by <13
weeks of feeding of 2,3,7,8-TCDD.
Other studies do not support the dose-dependent tissue distribution of
2,3,7,8-TCDD and related compounds described above. Rose et al. (1976) reported
a lack of a dose-dependent accumulation of ^C-TCDD in male and female rat liver
and adipose tissue following 7, 21 and 49 days of exposure at 0.01, 0.1 or 1.0
pg/kg/day, Monday through Friday. The rates of accumulation of TCDD-derived
radioactivity were similar in fat, liver and whole body; however, the concentra-
tion in the liver was -5-fold greater than that in fat. Recently, Clark et al.
(1991) and Tritscher et al. (1992) also reported a lack of a dose-dependent
hepatic disposition of TCDD in female Sprague-Dawley rats exposed biweekly to
TCDD for 30 weeks at doses equivalent to 3.5, 10.7, 35.7 and 125 ng/kg/day. A
linear relationship between administered dose and the concentration in the liver
was observed over the dose range used in this chronic exposure study. Brewster
and Birnbaum (1987) also observed similar concentrations (% dose/g) of 2,3,4,7,8-
PeCDF in liver, adipose tissue and other tissues at 3 days after oral exposure
at doses of 0.1, 0.5 or 1.0 pmol/kg bw. These results conflict with the above
studies which support the dose-dependent tissue distribution of these compounds.
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While it is not possible at this time to explain these differences, most of the
available data support a dose-dependent relationship.
The dose-dependent tissue distribution of 2,3,7,8-TCDD and related compounds
is a critical factor that must be considered in estimating the concentration of
these compounds in human tissues after chronic low-level exposure. This is
particularly important since the general human population is exposed to much
smaller daily doses (possibly 0.3 pg 2,3,7,8-TCDD/kg/day) than those used in
experimental disposition studies. Due at least partly to the long half-life of
2,3,7,8-TCDD in humans, however, this exposure results in concentrations of 3-18
pg/g in human adipose tissue (Leung et al., 1990). Similar levels of 2,3,7,8-
TCDD in adipose tissue (14 pg/g) were observed in rats 7 days after subcutaneous
exposure to 3 ng/kg bw (see Table 1-7) (Abraham et al., 1988). Under these
experimental conditions, the liver/adipose tissue 2,3,7,8-TCDD concentration was
0.74. Nonetheless, steady state was definitely not reached under these
conditions, and, with increasing time after exposure, this ratio may decrease,
based on the observation that 2,3,7,8-TCDD was more persistent in adipose tissue
than in liver in rats exposed to 300 ng/kg bw (see Figure 1-1 and Table 1-5)
(Abraham et al., 1988). Human data on the liver/adipose tissue concentration
ratio of 2,3,7,8-TCDD and related compounds are limited but suggest that the
ratio may vary by at least an order of magnitude between individuals. Leung et
al. (1990) observed a geometric mean adipose tissue 2,3,7,8-TCDD concentration
of 7.78 ppt in 26 individuals and a concentration in liver at about one-tenth of
that in adipose tissue on a whole weight basis. When measured on a total lipid
basis, the concentrations of 2,3,7,8-TCDD in both tissues were approximately the
same. More variability between individuals was observed in the CDD and CDF
concentrations in liver and adipose tissue from 25 subjects from the Munich area
(Thoma et al., 1989). For example, the liver/adipose tissue concentration ratio
for 2,3,7,8-TCDD was >1.0 for 5 of the 25 individuals (Thoma et al., 1989).
While the majority of individuals had liver/adipose tissue concentration ratios
<1.0 for CDDs and CDFs, ratios >1.0 were observed for HpCDD (5 of 25), OCDD (2
of 25), PeCDF (2 of 25), HxCDF (1 of 25), HpCDF (7 of 25) and OCDF (3 of 25).
Considerable variability in CDD and CDF concentrations in liver and adipose
tissues was also observed between individual marmoset monkeys (Neubert et al.,
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DRAFT—DO NOT QUOTE OR CITE
1990), suggesting that individual variability may also contribute to the
difficulty in assigning a constant liver/adipose tissue ratio for CDDs and CDFs
in humans and nonhuman primates.
1.2.5. Potential Mechanisms for the Dose-Dependent Tissue Distribution. The
observation that exposure to higher doses of 2,3,7,8-TCDD and related compounds
results in a disproportionally greater hepatic concentration of these compounds
may be explained by a hepatic binding species that is induced by 2,3,7,8-TCDD and
other agonists for the Ah receptor. The studies of Voorman and Aust (1987, 1989)
and Poland et al. (1989a,b) provide evidence that this binding species is
cytochrome P-4501A2.
Poland et al. (1989) reported that TCDD and other Ah agonists (2, 3,7,8-TCDF,
B-naphthoflavone, 3,3',4,4',5,5'-hexabromobiphenyl) act through the Ah receptor
to increase a liver binding species that increases the hepatic uptake of
[12^I]-2-iodo-3,7,8-trichlorodibenzo-p-dioxin (a radiolabeled isosteric analogue
of TCDD) in vivo and binding of this radioligand to liver homogenate in vitro.
Twenty-four hours after the administration of a non-AHH-inducing dose (1x10"
mol/kg) of [125I]-2-iodo-3,7,8-trichlorodibenzo-p-dioxin to C576BL/6J mice, the
hepatic concentration of radioactivity was 1-2% of the administered dose, whereas
in mice pretreated 48 hours earlier with an AHH inducing dose of TCDD (1x10"'
mol/kg), the hepatic accumulation of radiolabel was 25-30% of that administered.
A similar, though less dramatic effect, was observed in vitro, with liver
19S
homogenate from TCDD-treated mice binding about four times more [* I]-2-iodo-
3,7,8-trichlorodibenzo-p-dioxin than homogenate from control mice. The adminis-
tration of TCDD to C57BL/6J mice produced a dose-related stimulation of in vivo
hepatic uptake of [125I]-2-iodo-3,7,8-trichlorodibenzo-p-dioxin, increased
binding of radioligand to liver homogenate and induction of hepatic activity,
with an ED5Q ranging from 1.5 to 4.0xlO"9 mol/kg. In congenic C57BL/6J (Ahd/Ahd)
mice, which express the lower affinity Ah receptor, the £050 values for all three
responses were shifted to doses that were about 10-fold higher. The observed
effects on hepatic disposition were tissue specific, with no remarkable disposi-
tional changes being observed in kidney, lung, spleen, small intestine or muscle.
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This is significant in that TCDD and other agonists for the Ah receptor induce
cytochrome P-4501A1 in liver and other tissues, whereas cytochrome P-4501A2 is
apparently inducible only in liver and nasal olfactory mucosa (Tuteja et al.,
1985; Gillner et al. 1987). Furthermore, the changes in hepatic disposition were
not species specific; similar responses were observed in guinea pigs, rats, mice
and hamsters (Poland et al., 1989).
The following evidence reported by Poland et al. (1989) supports the
hypothesis that the TCDD-inducible hepatic binding protein is cytochrome
P-4501A2: the TCDD-induced hepatic binding species was found predominantly in
the microsomal fraction and was inactivated by heating at 60°C, trypsin and
mercurials; the TCDD-induced hepatic binding species was specific for the liver,
with a large pool size (Bmax of 22±5 nmol/g liver); and the major microsomal
binding species covalently labeled with the photo-affinity ligand [^I]-2-iodo-
3-azido-7,8-dibromodibenzo-p-dioxin migrates with that immunochemically stained
with polyclonal antiserum that binds to cytochrome P-4501A2.
One observation of Poland et al. (1989) does not support the hypothesis that
the TCDD-inducible hepatic protein is cytochrome P-4501A2. These investigators
found that dietary administration of isosafrole did not stimulate hepatic uptake
1 ye
of [1'"l]-2-iodo-3,7,8-trichlorodibenzo-p-dioxin or the in vitro binding of this
ligand to liver homogenate. Isosafrole is not an agonist for the Ah receptor,
but it selectively induces cytochrome P-4501A2 (Ryan et al., 1980). Poland et
al. (1989) suggest that this may be attributable to the high affinity binding of
an isosafrole metabolite to the protein, which might inhibit the binding of
19S
[1'"I]-iodo-3,7,8-trichlorodibenzo-p-dioxin to cytochrome P-4501A2 at or near the
active site of the enzyme. This does not explain why TCDD, which also has high
affinity for cytochrome P-4501A2, cannot displace some of the isosafrole
metabolite from the protein, which should produce enhanced hepatic disposition
of TCDD.
Voorman and Aust (1987, 1989) support further the hypothesis that cytochrome
P-4501A2 is the TCDD-inducible hepatic binding species. These investigators
found that 3,3'4,4'5, B'-HxBB, an agonist for the Ah receptor, was associated only
with cytochrome P-4501A2 through the immunoprecipitation of cytochromes P-4501A1
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and 1A2, which were induced in 3,3'4,4'5,5'-HxBB treated rats. In addition, they
found that 3,3'4,4'5,5'-HxBB inhibited estradiol 2-hydroxylase activity of
purified cytochrome P-4501A2. A similar association of PAHs with immunoprecipi-
tated cytochrome P-4501A2 was observed for other agonists for the Ah receptor,
including 2,3,7,8-TCDD, 3,3' ,4,4'-TCB, 3,3 ' ,4,4 ' , 5-PeCB and 3,3 ' ,4,4', 5,5 '-HxCB.
The association of 2,3,7,8-TCDD with cytochrome P-4501A2 occurred within 2
minutes, with maximum inhibition of estradiol 2-hydroxylase occurring at a
concentration comparable to the concentration of the enzyme (50 nm). Cytochrome
P-4501A2 was inhibited (complexed) by 2,3,7,8-TCDD with nearly 1:1 stoichiometry
and the Kj for 2,3,7,8-TCDD was calculated to be 8 nM. Therefore, 2,3,7,8-TCDD
can be considered a higher binding inhibitor of cytochrome P-4501A2.
The TCDD-induced binding species was found to have an apparent equilibrium
19S
dissociation constant, KJ-J, [ 1-"I]-2-iodo-3,7,8-trichlorodibenzo-p-dioxin of 56±16
nM and a pool size, Bmax, of 22±5 nmol/g of liver in C57BL/6J mice (Poland et
al., 1989). The induced microsomal binding species has an affinity about 10
times less than the Ah receptor but a pool size that is -2x10^ greater. Thus,
agonists for the Ah receptor may significantly affect their disposition through
a dose-related enhancement of hepatic uptake which should correlate with
induction of cytochrome P-4501A2.
The disposition and pharmacokinetics of 2,2' ,4,4',5,5'-HxCB and -HxBB have
been investigated in several species (Tuey and Matthews, 1980; Lutz et al.,
1984). These lipophilic compounds are similar to 2,3,7,8-TCDD in that they are
slowly metabolized and that metabolism is required for urinary and biliary
elimination. 2,2',4,4',5,5'-HxCB and -HxBB distribute primarily to adipose
tissue with partition coefficients (tissue/blood ratio) ranging from 300-500 in
the mouse, rat, monkey, dog and human. The liver is not a major site for the
disposition of 2,2',4,4',5,5'-HxCB and -HxBB, in contrast to 2,3,7,8-TCDD and
related compounds. Partition coefficients in the liver range from 10-30 in these
species. 2,2' ,4,4' ,5,5'-HxCB and -HxBB do not induce hepatic cytochrome P-4501A1
or 1A2 and do not exhibit dioxin-like activity. The lack of induction of hepatic
cytochrome P-4501A2 may explain the lack of a dose-dependent hepatic disposition
of these compounds.
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Kedderis et al. (1991b) assessed the dose-response relationship for the
induction of hepatic cytochrome P-4501A1 and P-4501A2 in male Fischer 344 rats
exposed to 2,3,7,8-TBDD at doses as low as 0.1 nmol/kg. They reported that
induction of P-4501A2 by 2,3,7,8-TBDD appeared to be a more sensitive response
than P-4501A1 induction over the dose-range studied. In addition, comparison of
hepatic P-4501A2 levels and liver:adipose tissue concentration ratios suggested
that induction of P-4501A2 alone would not directly account for the preferential
hepatic accumulation of 2,3,7,8-TBDD, and additional factors must be involved.
One explanation may be that at low 2,3,7,8-TBDD concentrations, endogenous
substrates bind to CYP1A2, not allowing 2,3,7,8-TBDD to be sequestered by the
protein (Birnbaum, 1992). At higher 2,3,7,8-TBDD concentrations, new protein is
formed and 2,3,7,8-TBDD can compete for binding to CYP1A2, resulting in the
increased hepatic deposition observed at higher exposures of 2,3,7,8-TBDD
(Kedderis et al., 1991a).
Other factors may also regulate the intracellular distribution of 2,3,7,8-
TCDD and related compounds. The possible role of hepatic lipoproteins as
intracellular carriers in the transport of 2,3,7,8-TCDD has been assessed by in
vitro and in vivo studies (Soues et al., 1989a,b). 2,3,7,8-TCDD and 2,3,7,8-TCDF
were bound to lipoproteins in mouse and rat liver, which subsequently underwent
rapid and pronounced degradative processing, possibly catalyzed by lipoprotein
lipase, to heavier entities. The in vitro incubation of 2,3,7,8-TCDD-lipoprotein
complex with separated Ah receptor demonstrated that a passive transfer occurred.
The authors suggest a carrier-role for lipoproteins in the intracellular
transport of 2,3,7,8-TCDD and related compounds.
1.3. METABOLISM AND EXCRETION
Although early in vivo and in vitro investigations were unable to detect the
metabolism of 2,3,78-TCDD (Vinopal and Casida, 1973; Van Miller et al., 1976),
there is evidence that a wide range of mammalian and aquatic species are capable
of biotransforming 2,3,7,8-TCDD to polar metabolites (Ramsey et al., 1979, 1982;
Poiger and Schlatter, 1979; Olson et al., 1980; Olson, 1986; Gasiewicz et al.,
1983; Poiger et al., 1982; Sijm et al., 1990; Kleeman et al., 1986a,b, 1988).
Although metabolites of 2,3,7,8-TCDD have not been directly identified in humans,
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recent data regarding feces samples from humans in a self-dosing experiment
suggests that humans can metabolize 2,3,7,8-TCDD (Wendling et al., 1990).
Table 1-8 summarizes data on the metabolism and excretion of 2,3,7,8-TCDD
and related compounds after exposure to a single radiolabeled congener.
Investigations of 2,3,7,8-TCDD in rats, mice, guinea pigs and hamsters found that
>90% of the radiolabeled material excreted in urine and bile represented polar
metabolites. Similar results were also observed for other congeners (see Table
1-8), with the exception of OCDD; however, although studies were often limited
to the rat. OCDD is apparently not metabolized by the rat or metabolized to a
very minimal extent (Birnbaum and Couture, 1988). For all of the congeners in
Table 1-8, essentially all of the CDD, BDD, CDF or PCB-derived radioactivity in
liver, adipose tissue and other tissues represented parent compound, suggesting
that the metabolites of these compounds were readily excreted. Thus, with the
exception of OCDD, the metabolism of 2,3,7,8-TCDD and related compounds is
required for urinary and biliary elimination and therefore plays a major role in
regulating the rate of excretion of these compounds. In addition, direct
intestinal excretion of parent compound is another route for excretion of
2,3,7,8-TCDD and related compoundst that is not regulated by metabolism.
1.3.1. Structure of Metabolites. Several metabolites of 2,3,7,8-TCDD have
recently been identified, Sawahata et al. (1982) investigated the in vitro
metabolism of 2,3,7,8-TCDD in isolated rat hepatocytes. The major product was
deconjugated with B-glucurpnidase, derivatized with diazomethane and separated
into two compounds by HPLC. These metabolites were subsequently identified as
l-hydroxy-2,3,7,8-TCDD and 8-hydroxy-2,3,7-trichlorodibenzo-p-dioxin. Poiger et
al. (1982b) identified six metabolites in the bile of dogs that were given a
lethal dose of [%]-2,3,7,8-TCDD. The major metabolite was 1,3,7,8-tetrachloro-
2-hydroxydibenzo-p-dioxin; however, 3,7,8-trichloro-3-hydroxydibenzo-p-dioxin and
l,2-dichloro-4,5-hydroxybenzene were identified as minor metabolites. The
structures of the three remaining metabolites were not determined; however, two
appeared to be trichlorohydroxydibenzo-p-dioxins and the third was apparently a
chlorinated 2-hydroxydiphenyl ether. Poiger and Buser (1984) reported differ-
ences in the relative amounts of various 2,3,7,8-TCDD metabolites in dog and rat
1-43 OB/11/92
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TABLE 1-8
Metabolism and Excretion of 2,3,7,8-TCDD and Related Compounds8
Chemical
Spec i es
Dose
CDDs
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8, -TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCOD
Sprague-Dawley
rat (M)
Sprague-Dawley
rat
Sprague-Dawley
rat (F)
Sprague-Dawley
rat (M, F)
Sprague-Dawley
rat (M, F)
Han/Wistar rat
(M)
Long-Evans rat
(M)
Sprague-Dawley
(M)
C57BL/6J mice
(M)
DBA/2 J mice
(M)
B6D2F1J mice
50 fig/kg, P.O.
7 or 72 ppb
in diet for
42 days
7 or 72 ppb
in diet for
42 days
1.0 jig/kg,
p.o
0.1 and 1.0
/ig/kg/day,
5 days/week
for 7 weeks
5 /ig/kg, i.p.
5 M9/kg, i.p.
500 /ig/kg, i.p.
10 /tg/kg, i.p.
10 /ig/kg, i.p.
10 /ig/kg, i.p.
Chemical Nature of
Excretion Products
(% Metabolites)
Urine
NA
NA
NA
NA
NA
>90
>90
100
100
100
100
Bile
NA
NA
NA
NA
NA
NA
NA
100
100
100
100
Feces
Ratio of % of
Dose Excreted
(Feces/Urine)
NA
NA
NA
NA
NA
"70-90
"20-90
NA
85
82
86
4.0
NA
NA
9.9
8.5
14.1
12.0
NA
2.7
1.2
2.5
Half-Life6
(days)
17.4±5.6C
12
15
31±6d
23.7
21.9
20.8
NA
11.0t1.2d
24.4±1.0d
12.6±0.8d
Comment
NC
NC
NC
NC
NC
NC
NC
NC
NC
NC
NC
Reference
Piper et al.,
1973
Fries and
Morrow, 1975
Fries and
Morrow, 1975
Rose et al.,
1976
Rose et a I.,
1976
Pohjanvirta
et al., 1990
Pohjanvirta
et al., 1990
Neal et al.,
1982
Gasiewicz et
al., 1983
Gasiewicz et
al., 1983
Gasiewicz et
al., 1983
O
O
§
H
3
M
O
50
O
H
VO
10
-------
I
•t»
in
O
00
TABLE 1-8 (cont.)
Chemical
2,3,7,8-TCDO
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCDO
2-Iodo-3,7,8-TCDD
2-Iodo-3,7,8-TCDD
2,3,7,8-TCDO
2,3,7,8-TCDO
2,3,7,8-TCDD
2,3,7,8-TCDD
Species
C57BL/6J mice
AhD/Aha (M)
C57BL/6J mice
Aha/Aha (M)
DBA/2J Ahb/Ahd
(F)
DBA/2J Ahd/Ahd
(F)
C57BL/6J mice
(F)
C57BL/6J mice
-------
I
*»
O\
O
CO
TABLE 1-8 (cont.)
Chemical
2,3,7,8-TCOD
2,3,7,8-TCDD
2,3,7,8-TCDD
2,3,7,8-TCOD
1,2,3,7,8-PeCOD
OCDD
OCDO
BDDs
2,3,7,8-TBDD
Species
Golden Syrian
hamster (M)
human (M)
rainbow trout
yellow perch
Sprague-Dawley
rat (M, F)
Fischer 344 rat
Fischer 344 rat
(M)
Fischer 344 rat
(M)
Dose
[3H] 650
/ig/kg, p.o.
1.14 ng/kg,
p.o.
494 ppt in diet
for 13 weeks
494 ppt in diet
for 13 weeks
8.42-10.06
Jig/kg, p.o.
50 pg/kg, iv
50 /ig/kg/day,
p.o., for 10
days
Chemical Nature of
Excretion Products
(X Metabolites)
Urine
NA
NA
NA
NA
NA
<33
NA
Bile
NA
NA
"75
'90
100
0
NA
Feces
NA
"50
NA
NA
NA
0
NA
0.001 junol/kg,
iv
NA
100
80-90
Ratio of X of
Dose Excreted
(Feces/Urine)
NA
>3.1
NA
NA
12
>65
NA
Half-Life6
(days)
14.96±2.53
2120e
105
126
29.5±2.7
'70
"173
11.1
0.7
2.9
17.8
Comment
NC
NC
elimination followed
for 13 weeks following
exposure
elimination followed
for 13 weeks following
exposure
NC
whole body t^
estimated from body
burden in liver, skin
and adipose tissue over
56-day period
whole body t^
estimated from body
burden in liver, skin
and adipose tissue over
112-day period
Pool size (X of dose):
11.63 1st component
2.78 2nd component
1 .45 3rd component
Reference
Olson et at.,
1980; Neal et
al., 1982
Poiger and
Sch latter,
1986;
Wend 1 ing et
al., 1990
K Iceman et
al., 1986
Kleeman et
al., 1986
Wacker et
al., 1986
Birnbaum and
Couture, 1988
Birnbaum and
Couture, 1988
Kedderis et
al., 1991
D
O
I
M
8
O
H
VO
10
-------
DRAFT—DO NOT QUOTE OR CITE
XX
4-1
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01
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4-1
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01
5 «
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O X *x.
UJ CA
O 01
**~ 01 O
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CO O u.
ae o xx
£
u
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O *•* xx U-
«> 3 g
3 O.t!
CO CX "Q 01
5 Z *
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.C X 01
U UJ C
=
01
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1-47
08/11/92
-------
Chemical
2,3,4,7,8-PeCDF
CBs
3,3'4,4'-TCB
3,3'4,4'-TCB
TABLE 1-8 (cont.)
Species
rhesus monkey
(M)
CD rat (M, F)
rhesus monkey
(F)
Dose
0.1 pmol/kg, iv
Chemical Nature of
Excretion Products
(X Metabolites)
Urine
NA
Bile
NA
Feces
63-70
Ratio of % of
Dose Excreted
(Feces/Urine)
"34
0.6 mg/kg, iv
0.6 mg/kg, iv
>90
97
NA
NA
>90
97
42
7.2
Half-Life13
(days)
38-49
"1.3-1.5
"8-10
Comment
t^/2 represents minimum
value; all animals lost
body weight and
exhibited other signs
of toxicity
Reference
Brewster et
al., 1988
NC
NC
Abdel-Hamid
et al., 1981
Abdel-Hamid
et al., 1981
aAll studies measure the excretion of radiolabeled parent compound and metabolites following exposure to a single congener labeled with H, C or
1 •
'•'Half-life for excretion estimates assume first-order elimination kinetics
c(mean±SE)
d(mean±SD)
§
s
M
O
»
O
M
M
en=1
i.p. = Intraperitoneal; i.v. = intravenous; NA = not available; NC = no comment; p.o. = per os
O
GO
VO
IO
-------
DRAFT—DO NOT QUOTE OR CITE
bile. Trichlorodihydroxydibenzo-p-dioxin and tetrachlorodihydroxydiphenyl ether
appear to be major metabolites in rat bile. Furthermore, conjugates, presumably
glucuronides, were formed in the rat but not in the dog. The investigators also
observed a generally higher metabolism rate of 2,3,7,8-TCDD in the dog. This is
in good agreement with the unique ability of the dog to readily metabolize
persistent PCBs such as 2,4,5,2'415'-HxCB (Sipes et al., 1982).
Biliary metabolites of 2,3,7,8-TCDF have been investigated by Poiger et al.
(1984); however, unequivocal structure assignment of the metabolites could not
be made using GC/MS. With the use of synthetic standards and GC/MS, Burka et al.
(1990) identified 4-hydroxy-2,3,7,8-TCDF and 3-hydroxy-2,7,8-TCDF as major
biliary metabolites of 2,3,7,8-TCDF in rats. Small amounts of 3-hydroxy-2,4,7,8-
TCDF and 2,2 '-dihydroxy-4,41, 5,5 '-TCB were also detected. This suggests that the
preferred site of metabolism of 2,3,7,8-TCDF is near the furan oxygen with
oxygenation at C4 predominating over C3. The authors speculate that epoxidation
of the C4-C4a bond or the C3-C4 bond could lead to formation of the above
metabolites. The results of Burka et al. (1990) and Sawahata et al. (1982)
suggest that oxygenation of the unsubstituted carbon nearest the bridging oxygen
in both 2,3,7,8-TCDF and 2,3,7,8-TCDD is the major route of metabolism of these
compounds in the rat. Furthermore, data on the rate of elimination of these
compounds summarized in Tables 1-5 and 1-8 indicate that this reaction occurs at
a faster rate for the furan, since the rate of urinary and biliary elimination
and resulting persistence of these compounds depends on metabolism.
Data summarized in Tables 1-5 and 1-8 indicate that 1,2,3,7,8-PeCDF is
metabolized and eliminated at a greater rate than 2,3,4,7,8-PeCDF. The
preference for oxygenation at C4 in 2,3,7,8-TCDF offers an explanation for the
observation that 2,3,4,7,8-PeCDF is metabolized at a much slower rate than
1,2,3,7,8-PeCDF, because one of the preferred sites for metabolism is blocked in
the 2,3,4,7,8-substituted compound. The rate of metabolism of these compounds
and their resulting relative persistence in rodents correlate with analysis of
human tissues from the Yusho cohort where the relative concentrations were
2,3,4,7,8-PeCDF > 1,2,3,7,8-PeCDF > 2,3,7,8-TCDF (Masuda et al., 1985).
Pluess et al. (1987) investigated the structure of 1,2,3,7,8-PeCDF metabo-
lites in rat bile. A dihydroxy-tetra-CDF was identified as the major metabolite.
1-49 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
The authors propose that this compound could be formed either via further
oxidation of the hydroxy-tetra-CDF or possibly via hydrolytic dechlorination of
a hydroxy-penta-CDF. Minor metabolites include a dihydroxy-tri-CDF, hydroxy-
tetra-CDF and hydroxy-penta-CPF.
Pluess et al. (1987) also investigated the metabolites of 2,3,4,7,8-PeCDF
in rat bile. A total of 10 metabolites were detected with a dihydroxy-penta-CB
and a hydroxy-penta-CDF representing the major metabolites. The biphenyl
metabolite indicates that cleavage of the ether bridge of the furan is an
important pathway for metabolism of this congener. Other less abundant metabo-
lites of 2,3,4,7,8-PeCDF include a hydroxy-tetra-CDF, dihydroxy-tri-CDF,
dihydroxy-tetra-CDF and a thio-tetra-CDF. Sulfur-containing metabolites were
also identified as minor metabolites of 2,3,7,8-TCDF and 1,2,3,7,8-PeCDF in rats
(Kuroki et al., 1990). These sulfur-containing metabolites probably arise from
CDF-glutathione conjugates.
In another study, a dihydroxy-PeCDF was identified as the only detectable
biliary metabolite of 1,2,3,6,7,8-HxCDF, while no metabolites of 1,2,3,4,6,7,8-
HpCDF were detected in the bile of rats that were treated with this congener
(Poiger et al., 1989).
Several in vivo and in vitro studies have investigated the metabolism of
3,3',4,4'-TCB. Rat feces were found to contain 5-hydroxy-3,3',4,4'-TCB and 4-
hydroxy-3,3'4',5-TCB as major metabolites (Yoshimura et al., 1987) and
2,5-dihydroxy-3,3',4,4'-TCB, 4,4'-dihydroxy-3,3'5,5'-TCB, 5,6-dihydroxy-
3,3',4,4'-TCB, 4-hydroxy-3,3',4-TCB and 4-hydroxy-4',5'-epoxy-3,3'4',5-TCB as
minor metabolites (Koga et al., 1989).
Mouse feces were found to contain 5-hydroxy- and 6-hydroxy-3,3' ,4,4'-TCB and
4-hydroxy-3,3 ' ,4'5-TCB while urine contained 2-hydroxy-3,3 ' ,4,4'-TCB in addition
to these metabolites (Wehler et al., 1989). 3,3'4,4'-TCB was the major compound
present in mouse liver, while a minor portion was due to 4-hydroxy-3,3'4,4'-TCB
(Wehler et al., 1989). Darnerud et al. (1986) found 2-hydroxy-3,3'4,4'-TCB and
a methylsulphonyl-TCB as major metabolites in the mouse fetus. Sulphur-
containing metabolites of noncoplanar PCBs have also been reported to accumulate
in the bronchial mucosa and uterine luminal fluid of mice (Bergman et al., 1979;
Brandt et al., 1982) and in human lung, liver and adipose tissue (Haraguchi et
1-50 OB/11/92
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DRAFT—DO NOT QUOTE OR CITE
al., 1986, 1989). PCB methyl sulphones are stable lipophilic metabolites formed
by the mercapturic acid pathway. The toxicological significance of these
metabolites remains generally unknown.
1.3.2. Toxicity of Metabolites. The above discussion indicates that the
metabolism of 2,3,7,8-TCDD and related compounds is required for urinary and
biliary elimination and thus plays a major role in regulating the rate of
excretion of these compounds. At present, metabolism is also generally
considered a detoxification process.
Data on the metabolism of 2,3,7,8-TCDD suggests that reactive epoxide
intermediates may be formed. Poland and Glover (1979) have investigated the in
vivo binding of [l,2-3H]-2,3,7,8-TCDD derived radioactivity to rat hepatic
macromolecules, and found maximum levels equivalent to 60 pmol of 2,3,7,8-
TCDD/mol of nucleotide in RNA, and 6 pmol of 2,3,7,8-TCDD/mol of nucleotide in
DNA. This corresponds to one 2,3,7,8-TCDD-DNA adduct/35 cells. These investi-
gators suggest that it is unlikely that 2,3,7,8-TCDD-induced oncogenesis is
through a mechanism gf covalent binding to DNA and somatic mutation. Further
studies of 2,3,7,8-TCDD and related compounds are needed to confirm these results
and assess the relationship between covalent binding and the short and long-term
toxicity of these compounds.
Weber et al. (1982) investigated the toxicity of 2,3,7,8-TCDD metabolites
by administering extracts of bile from 2,3,7,8-TCDD-treated dogs to male guinea
pigs in single oral doses equivalent to 0.6, 6.0 and 60 pg/kg of parent compound.
Other groups of guinea pigs were given bile extract form untreated dogs or
2,3,7,8-TCDD itself. A comparison of the mortality data at 5 weeks after dosing
indicated that the acute toxicity of 2,3,7,8-TCDD to guinea pigs was at least 100
times higher than wa» the acute toxicity of its metabolites.
Mason and Safe (1986) synthesized 2-hydroxy-3,7,8-TCDD and 2-hydroxy-
1,3,7,8-TCDD, which are metabolites of 2,3,7,8-TCDD, and assessed the toxicity
of these compounds in male Wistar rats. The compounds produced no significant
effect on body weight gain, thymus, liver or spleen weights after exposure to a
dose of <5000 ^g/kg bw. 2-Hydroxy-3,7,8-TCDD induced hepatic microsomal AHH,
EROD and 4-chlorobiphenylhydroxylase activity at an exposure of 1000 and
1-51 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
5000 pg/kg bw, while 2-hydroxy-l,3,7,8-TCDD was inactive as an inducer. Thus,
while 2-hydroxy-3,7,8-TCDD has dioxin-like activity as an inducer of the hepatic
monooxygenase system, the potency of the metabolite is more than three orders of
magnitude less than that of 2,3,7,8-TCDD. Furthermore, results are consistent
with the expected rapid conjugation and excretion of these 2,3,7,8-TCDD metabo-
lites (Weber et al., 1982).
Metabolism of coplanar PCBs and PBBs also appears to be a detoxification
process. 5-Hydroxy-3,3•,4,4'-TCB and 4-hydroxy-3,3•,4',5-TCB did not produce
liver hypertrophy, induction of hepatic AHH or DT-diaphorase activities or thymus
atrophy (Yoshimura et al., 1987). Thus, monohydroxy metabolites of 3,3',4,4'-TCB
are much less toxic than the parent compound. Further evidence for metabolism
as a detoxification process comes from comparison of the metabolism and toxicity
of two coplanar PBBs. Millis et al. (1985) found that 3,3•,4,4',5,5'-HxBB
exhibited greater toxic potency in rats than 3,3',4,4'-TBB, even though
3,3',4,4'-TBB had about a 10-fold greater affinity for the Ah receptor. Although
receptor binding affinities imply that 3,3'4,4'-TBB should be more toxic than
3,3',4,4'5,5'-HxBB, it was less toxic than the HxBB because 3,3',4,4'-TBB was
metabolized at a much greater rate than 3,3•,4,4',5,5'-HxBB. In addition to
supporting metabolism as a detoxification process, the results of Millis et al.
(1985) also suggest that receptor binding and in vitro AHH induction do not
accurately reflect toxicity for PAHs, which are more readily metabolized,
presumably because continued occupation of the receptor is required for toxicity.
Structure-activity studies of 2,3,7,8-TCDD and related compounds support the
widely accepted principle that this parent compound is the active species. The
relative lack of activity of readily excreted monohydroxylated metabolites of
2,3,7,8-TCDD and 3,3'4,4'-TCB suggests that metabolism is a detoxification
process necessary for the biliary and urinary excretion of these compounds. This
concept has also been generally applied to 2,3,7,8-TCDD related compounds,
although data are lacking on the structure and toxicity of metabolites of other
CDDs, BDDs, CDFs, BDFs, PCBs and PBBs.
It is possible that low levels of unextractable and/or unidentified
metabolites may contribute to one or more of the toxic responses of 2,3,7,8-TCDD
1-52 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
and related compounds. Further studies on the nature of the biotransformation
products of these compounds will help to address this uncertainty.
. 1.3.3. Autoinduction of Metabolism. Accurate rate constants for metabolism are
important in developing pharmacokinetic models that describe the disposition of
2,3,7,8-TCDD and related compounds. Metabolism plays a major role in regulating
the excretion and relative persistence of these compounds, since metabolism is
required for urinary and biliary excretion. Although the relative rate of
metabolism of 2,3,7,8-TCDD and related compounds can be estimated from tissue and
excretion half-life data (see Tables 1-5 and 1-8), other factors such as relative
body composition, hepatic and extrahepatic binding proteins and direct intestinal
elimination of the parent compound can also regulate the excretion of 2,3,7,8-
TCDD and related compounds. Therefore, in vivo disposition data (see Tables 1-5
and 1-8) provide only a limited approximation of the relative rate of metabolism
of a specific congener in a given species. In vivo disposition data were also
obtained at exposures that were associated with induction of cytochromes P-4501A1
and 1A2 and other potentially adverse responses that could alter metabolism and
disposition. Therefore, it may not be appropriate to directly extrapolate these
data to predict the pharmacokinetics at low levels of exposure. Low dose
extrapolations can be assisted by assessments of the potential for autoinduction
of metabolism which may occur at exposures which are associated with enzyme
induction. Characterization of the dose dependent disposition of 2,3,7,8-TCDD
and related compounds is particularly important in the extrapolation of high
exposure animal data to low exposure human data.
The excretion of metabolites of 2,3,7,8-TCDD and related compounds into bile
represents a direct means for estimating the rate of metabolism, since biliary
elimination depends on metabolism and is the major route for excretion of these
compounds. The rate of metabolism of CDFs was estimated from the relative
abundance of metabolites in rat bile (Poiger et al., 1989). The rate of
biotransformation of 2,3,7,8-TCDF, 1,2,3,7,8-PeCDF, 2,3,4,7,8-PeCDF and
1,2,3,6,7,8-HxCDF were characterized as fairly high, moderate, low and very low,
respectively. Kedderis et al. (1991b) observed 10% of the dose of [3H]-2,3,7,8-
TBDD excreted in bile 5 hours after intravenous administration of 1 nmol/kg to
1-53 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
male Fischer 344 rats. All biliary radioactivity was attributable to metabo-
lites. This rate of elimination is similar to the fecal excretion (-8% of the
dose) 24 hours after intravenous administration of 1 nmol/kg [3H]-2,3,7,8-TBDD
(Kedderis et al., 1991a) and reflects the effect of intravenous bolus versus oral
administration on distribution and elimination. The large % dose excreted within
the first few days may also be due to a rapidly excreted impurity in the
radiolabeled 2,3,7,8-TBDD (Kedderis et al., 1992a). To assess the ability of
2,3,7,8-TCDD and 2,3,7,8-TBDD to induce their own metabolism (biliary elimina-
tion), rats were pretreated with 100 nmol/kg, per os, of each compound 3 days
prior to intravenous injection of 1 nmol/kg of the respective [3H] congeners.
Biliary excretion of the radiolabeled dose was quantitatively and qualitatively
unaffected by pretreatment, despite a 2-fold increase in hepatic levels of [3H]
in the pretreated animals and significant induction of cytochrome P-4501A1 and
1A2 (Kedderis et al., 1991b). Therefore, under these conditions, autoinduction
of 2,3,7,8-TCDD and 2,3,7,8-TBDD metabolism did not occur in the rat in vivo at
doses that elicited enhanced hepatic uptake. Similarly, Curtis et al. (1990)
observed no change, or even an apparent decrease, in gastrointestinal contents
and fecal elimination of TCDD equivalents in pretreated versus naive mice 24
hours after oral administration of [14C]-2,3,7,8-TCDD, despite significantly
enhanced levels of 2,3,7,8-TCDD in the livers of pretreated mice.
While the above studies suggest that autoinduction of 2,3,7,8-TCDD metabo-
lism does not occur, other results indicate that metabolism may be induced under
certain conditions. Poiger and Buser (1984) observed a small yet significant
increase in biliary excretion over a 72-hour period, with pretreated rats (10
jug/kg, intraperitoneal) excreting 9.7±1.9% of the radiolabeled dose of 2,3,7,8-
TCDD (200-300 pg/kg, per os) compared to 7.0±0.9% excreted by naive animals. In
addition to being small changes, these results were obtained using a dose of
2,3,7,8-TCDD in excess of the LD50 in the rat. Poiger and Schlatter (1985)
examined the influence of pretreatment with phenobarbital and 2,3,7,8-TCDD on the
biliary excretion of [3H]-2,3,7,8-TCDD metabolites in a dog given a single oral
dose of the [3H] congener (31 or 33.8 ng/kg). Without pretreatment, 24.5% of the
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DRAFT—DO NOT QUOTE OR CITE
absorbed dose was excreted in the bile within 110 hours. Phenobarbital did not
alter this rate, whereas pretreatment with 2,3,7,8-TCDD (10 /jg/kg) 9 days earlier
resulted in a doubling of the amount of metabolites excreted in bile (47.4%).
Although this observation is limited to one dog and requires further investiga-
tion, the results suggest that significant autoinduction of 2,3,7,8-TCDD
metabolism and biliary excretion may occur in the dog. Nonetheless, the small
increase in metabolism and biliary excretion of 2,3,7,8-TCDD in the rat observed
by Poiger and Buser (1984) and the negative results of Redderis et al. (1991b)
and Curtis et al. (1990) suggest that autoinduction of 2,3,7,8-TCDD metabolism
and biliary excretion in the rat may not occur, or occurs to an extent that is
not biologically relevant.
Limited data suggest that autoinduction of metabolism and biliary excretion
does occur for CDFs in contrast to CDDs and BDDs. Pretreatment of rats with
2,3,7,8-TCDF (1.0 /jmol/kg, 3 days earlier) significantly increased the biliary
excretion of a subsequent dose of [ 14C]-2,3,7,8-TCDF (McKinley et al., 1991).
The naive rats excreted 5.69±2.35% of the dose over the initial 8 hours, while
the pretreated rats excreted 13.18±3.15% of the [14C]-2,3,7,8-TCDF. Similarly,
pretreatment of rats with 2,3,4,7,8-PeCDF (500 /jg/kg, per os, 3 days earlier)
resulted in a 2-fold increase in the biliary elimination of a subsequent dose of
[14C]-2,3,4,7,8-PeCDF (Brewster and Birnbaum, 1987). These results suggest that
pretreatment with 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF induces the metabolism of
these congeners.
3,3',4,4'-TCB and 3,3',4,4'-TBB appear to be metabolized by a 3-methylchol-
anthrene-inducible form of hepatic cytochrome P-450 (1A1 and/or 1A2), which is
also induced by 3,3',4,4'-TCB (Shimada and Sawabe, 1983; Mills et al., 1985).
This suggests that these compounds can induce their own rate of metabolism and
subsequent excretion.
Isolated hepatocytes in suspension culture have been demonstrated to provide
a useful in vitro system for studying the hepatic metabolism of 2,3,7,8-TCDD
under the same conditions in species that have a wide range of sensitivity to the
compound (Olson et al., 1981). The in vitro rate of metabolism of 2,3,7,8-TCDD
in guinea pig, rat, C57BL/6J mouse, DBA/2J mouse and hamster hepatocytes was
1-55 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
estimated to be 0.2, 1.2, 1.1, 0.9 and 1.2 pmol/mg protein/hour, respectively
(Wroblewski and Olson, 1985, 1988; Shen and Olson, 1987). These results indicate
that 2,3,7,8-TCDD is metabolized by the guinea pig liver at a rate -5-fold less
than that observed for the rat, mouse and hamster. The limited ability of the
guinea pig to metabolize 2,3,7,8-TCDD can explain the limited excretion of
2,3,7,8-TCDD metabolites in feces, which represents the major route for 2,3,7,8-
TCDD excretion (Olson, 1986). In addition, the limited metabolism in the guinea
pig may partly explain the relatively long excretion half-life for 2,3,7,8-TCDD
in the guinea pig and may contribute to the remarkable sensitivity of the guinea
pig to the acute toxicity of this agent (Olson, 1986).
Isolated hepatocytes in suspension culture have been used as an in vitro
system for studying the autoinduction of metabolism of 2,3,7,8-TCDD and related
compounds. Wroblewski and Olson (1988) investigated the metabolism of
[ C]-2,3,7,8-TCDD (2.2 /^M) in hepatocytes isolated from untreated 2,3,7,8-TCDD-,
3-MC-, isosafrole- and phenobarbital-pretreated rats and hamsters. In both
species, 2,3,7,8-TCDD and 3-MC pretreatments elevated the rate of 2,3,7,8-TCDD
metabolism by 5- to 6-fold, while phenobarbital pretreatment had no effect.
Isosafrole produced a 1.8- to 2.5-fold increase in metabolism. These in vitro
results at a high substrate concentration (2.2 pM) indicate that 2,3,7,8-TCDD can
induce its own rate of metabolism in the rat and hamster. In contrast, 2,3,7,8-
TCDD was not able to induce its own rate of metabolism in guinea pig and mouse
hepatocytes (Wroblewski and Olson, 1985; Shen and Olson, 1987). Together, these
results indicate that 2,3,7,8-TCDD is metabolized in the liver by a 2,3,7,8-TCDD
inducible enzyme, which is expressed in the rat and hamster, but not in the
guinea pig and mouse. More recently, the kinetics of 2,3,7,8-TCDD metabolism was
investigated in isolated rat hepatocytes incubated with [3H]-2,3,7,8-TCDD at
concentrations of 0.01, 0.1 and 1.0 pM (Olson et al., 1991). Lower 2,3,7,8-TCDD
concentrations in the media result in concentrations in hepatocytes which are
more similar to the levels in the liver after in vivo exposure. For example, the
concentration of 2,3,7,8-TCDD in hepatocytes incubated at 0.01 pM are similar to
hepatic levels after in vivo exposure of rats at a dose of -10 /jg/kg. At 0.01
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DRAFT—DO NOT QUOTE OR CITE
•a
and 0.1 ^iM, the rate of metabolism of [JH]-2,3,7,8-TCDD was similar in hepato-
cytes isolated from control and 2,3,7,8-TCDD pretreated rats, while at 1.0 pH,
2,3,7,8-TCDD metabolism was greater in hepatocytes isolated from 2,3,7,8-TCDD
pretreated rats. The results indicate that 2,3,7,8-TCDD can induce its own rate
of metabolism in the rat, but only at high hepatic concentrations, which are
generally not attained after in vivo exposure. A dose-dependent autoinduction
of 2,3,7,8-TCDD metabolism is consistent with the lack of autoinduction of
2,3,7,8-TCDD metabolism and biliary excretion in the rat (Kedderis et al., 1991b;
Curtis et al., 1990).
The metabolism of [%]-2,3,7,8-TCDF was also investigated in isolated rat
hepatocytes incubated at concentrations of 0.01, 0.1 and 1.0 pM (Olson et al.,
1991). At all concentrations, hepatocytes from 2,3,7,8-TCDD pretreated rats
metabolized 2,3,7,8-TCDF at a rate from 4- to 25-fold greater than that observed
in hepatocytes from control rats. The results indicate that 2,3,7,8-TCDF is
metabolized in rat liver by a 2,3,7,8-TCDD inducible enzyme, possibly cytochrome
P-4501A1 or 1A2. These in vitro results support the in vivo autoinduction of
2,3,7,8-TCDF metabolism and biliary elimination observed in the rat (McKinley et
al., 1991).
There is in vivo and in vitro data suggesting that autoinduction of 2,3,7,8-
TCDD and 2,3,7,8-TBDD metabolism does not occur in the rat after exposure to
sublethal doses of these agents. This is in contrast to 2,3,7,8-TCDF and
2,3,4,7,8-PeCDF where in vivo and in vitro results support the autoinduction of
metabolism and biliary elimination of these compounds in the rat.
1.3.4. Excretion in Animals. Data regarding the excretion of 2,3,4,7-TCDD and
related compounds after exposure to a single radiolabeled congener (see Table
1-8) support the assumption of a first-order elimination process consisting of
one or more components. These studies show that 2,3,7,8-TCDD was excreted slowly
from all species tested, with half-lives ranging from 11 days in the hamster to
2120 days in humans. 2,3,7,8-TCDD is exceptionally persistent in humans relative
to other animal models. Elimination data in tissues (see Tables 1-5 and 1-6)
also indicate that 2,3,7,8-TCDD and related compounds are exceptionally persis-
tent in nonhuman primates (Bowman et al., 1989; Neubert et al., 1990). These
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differences may also be in part related to the dose-dependency of the excretion
of these compounds. In general, the congener and species specific rate of
elimination of 2,3,7,8-TCDD and related compounds from major tissue depots (see
Table 1-5) is similar to the excretion data summarized in Table 1-8.
In the Syrian Golden hamster, the mammalian species least sensitive to the
acute toxicity of 2,3,7,8-TCDD, excretion occurred readily through both the urine
(35% of administered dose, 41% of total excreted radioactivity) and feces (50%
of the administered dose, 59% of total excreted radioactivity) (Olson et al.,
1980b). A similar excretion pattern was observed in mice, although there was
significant strain variability (Gasiewicz et al., 1983; Birnbaum, 1986). In all
the other species, excretion occurred mainly through the feces, with relatively
minor amounts of 2,3,7,8-TCDD metabolites found in the urine (Piper et al., 1973;
Allen et al., 1975; Olson, 1986; Rose et al., 1976; Gasiewicz and Neal, 1979;
Pohjanvirta et al., 1990). Results in Table 1-8 also indicate that fecal
elimination was the primary route for the excretion of 1,2,3,7,8-PeCDD, OCDD,
2,3,7,8-TBDD, 2,3,7,8-TCDF, 1,2,3,7,8-PeCDF, 2,3,4,7,8-PeCDF, and 3,3',4,4'-TCB.
Only Piper et al. (1973) reported the excretion of metabolites in the expired
air. During 21 days following administration of a single oral dose of [ C]-
2,3,4,7-TCDD to rats, 3.2% of the administered radioactivity (4.6% of the
excreted radioactivity) was recovered in the expired air.
Studies in the rat, guinea pig, hamster and mouse have found that
essentially all of the 2,3,7,8-TCDD-derived radioactivity excreted in the urine
and bile corresponds to metabolites of 2,3,7,8-TCDD (see Table 1-8). The
apparent absence of 2,3,7,8-TCDD metabolites in liver and fat suggests that once
formed, the metabolites of 2,3,7,8-TCDD are excreted readily. Thus, urinary and
biliary elimination of 2,3,7,8-TCDD depends on metabolism of the toxin. The more
limited data for other compounds also suggest that this relationship may be true
for 1,2,3,7,8-PeCDD, 2,3,7,8-TBDD, 2,3,7,8-TCDF, 1,2,3,7,8-PeCDF, 2,3,4,7,8-
PeCDF, and 3,3',4,4'-TCB (see Table 1-8).
Although urine and bile appear to be free of unmetabolized 2,3,7,8-TCDD,
data indicate that 2,3,7,8-TCDD and its metabolites are excreted in the feces of
guinea pigs, rats, mice and hamsters treated with [ H]- and/or [ C]-2,3,7,8-TCDD
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(see Table 1-8). While 15-35% of the 2 , 3 , 7 , 8-TCDD-derived radioactivity in rat,
mouse and hamster feces represents unchanged 2,3,7,8-TCDD, 81% of the radioa-
ctivity in guinea pig feces represents unmetabolized 2,3,7,8-TCDD (Olson, 1986;
Neal et al., 1982; Gasiewicz et al., 1983; Olson et al., 1980). The daily
presence of unchanged 2,3,7,8-TCDD in feces and its absence in bile suggests that
direct intestinal elimination may be the source for the fecal excretion of
2,3,7,8-TCDD. Data also suggest that direct intestinal elimination of parent
compound contributes to the fecal excretion for 2,3,7,8-TBDD (Kedderis et al.,
1991). While the direct intestinal elimination of parent compound may occur for
other congeners (see Table 1-8), this conclusion cannot be made at this time due
to the lack of experimental data. Nonetheless, the species-specific fecal
excretion of 2,3,7,8-TCDF is very similar to that observed for 2,3,7,8-TCDD, with
>90% of the 2, 3, 7, 8-TCDF-derived radioactivity excreted in guinea pig feces
representing parent compound (Decad et al., 1981a) . In addition, the excretion
of unchanged CDDs and CDFs was detected in rat feces after subcutaneous exposure
to a defined mixture of congeners (Abraham et al., 1989). Studies in lactating
rats have also found that unchanged 2,3,7,8-TCDD may be excreted in the milk of
lactating animals (Moore et al., 1976; Lucier et al., 1975; Nau et al., 1986).
Lactation, direct intestinal elimination, and perhaps sebum may serve as routes
for excretion of 2,3,7,8-TCDD, which do not depend on metabolism of the toxin.
These data suggest that the in vivo half -life for elimination of 2,3,7,8-TCDD and
related compounds only provides an approximation of the rate of metabolism of
these compounds in a given animal. The results in Table 1-8 do suggest that
2,3,7,8-TCDF, 1, 2 , 3 , 7, 8-PeCDF, and 3,3 ' ,4,4'-TCB are metabolized and excreted
more rapidly that 2,3,7,8-TCDD, 2,3,7,8-TBDD, 1,2, 3, 7, 8-PeCDD, 2, 3, 4, 7, 8-PeCDF
and OCDD.
The rate of excretion of 2,3,7,8-TCDD and related compounds is species- and
congener-specific (see Table 1-8). 2,3,7,8-TCDD is most persistent in human and
nonhuman primates. In the hamster, the least sensitive species to the acute
toxicity of 2,3,7,8-TCDD, the mean ti was 10.8 days (Olson et al., 1980a,b) , and
in the guinea pig, the most sensitive species to the acute toxicity of 2,3,7,8-
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DRAFT—DO NOT QUOTE OR CITE
TCDD, the mean ti^ was 94 days (Olson, 1986). 2,3,7,8-TCDF was also most persis-
tent in the guinea pig, with a t!/6 of 20-40 days (Decad et al., 1981a; loannou
et al., 1983). Furthermore, results indicate that the relatively limited ability
of the guinea pig to metabolize 2,3,7,8-TCDD and -TCDF may contribute to the
greater persistence and greater acute toxicity of these congeners in the guinea
pig.
The time distribution, metabolism and excretion of 2,3,7,8-TCDD were also
investigated in Han/Wistar and Long-Evans rats, which were, respectively, more
resistent (LD50>3000 /jg/kg) versus more susceptible (LD50 -10 /Lig/kg) to the acute
toxicity of 2,3,7,8-TCDD (Pohjanvirta et al., 1990). The results suggest that
the metabolism and disposition of 2,3,7,8-TCDD do not have a major role in
explaining the strain differences in toxicity.
The intraspecies differences in the ti^ of 2,3,7,8-TCDD in three mouse
strains may be due to the finding that the DBA/2J strain possesses ~2-fold
greater adipose tissue stores than the C57BL/6J and B6D2FJ/J strains (Gasiewicz
et al., 1983b). The sequestering of the lipophilic toxin in adipose tissue
stores of the DBA/2J mouse may contribute to the greater persistence of 2,3,7,8-
TCDD in this strain. Birnbaum (1986) examined the effect of genetic background
on the distribution and excretion of 2,3,7,8-TCDD in two sets of congenic mouse
strains in which the congenic pairs differed only at the Ah locus. The Ah locus
had no effect on the tissue distribution or excretion of 2,3,7,8-TCDD. Thus, the
distribution and excretion of 2,3,7,8-TCDD were primarily governed by the total
genetic background rather than the allele present at the Ah locus. These
findings are consistent with the in vitro results of Shen and Olson (1987), who
found that the hepatic uptake and metabolism of 2,3,7,8-TCDD do not correlate
with genetic differences at the murine Ah locus. However, it is important to
note that all of these are relatively high-dose studies, which may not allow for
detection of Ah receptor-mediated effects on disposition.
Although the dose-related tissue distribution of 2,3,7,8-TCDD and related
compounds has been described recently, very limited data are available on the
dose-related excretion of these compounds. Rose et al. (1976) investigated the
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DRAFT—DO NOT QUOTE OR CITE
elimination of [14C]-2,3,7,8-TCDD in rats given repeated oral doses of 0.01, 0.1
or 1.0 pg/kg/day Monday through Friday for 7 weeks or a single dose of 1.0 ^g/kg.
In the single-dose study, no ^C was excreted in the urine or expired air; in the
repeated-dose study, however, 3-18% of the cumulative dose was excreted in the
urine by 7 weeks. This study indicated that steady-state concentrations will be
reached in the bodies of rats in -13 weeks. The rate constant defining the
approach to steady-state concentrations was independent of the dose of 2,3,7,8-
TCDD over the range studied. Relatively small changes in the excretion of
2,3,7,8-TBDD were also observed after exposures at 1 and 100 nmol/kg (Kedderis
et al., 1991). These results are consistent with the in vivo and in vitro
evidence suggesting that autoinduction of 2,3,7,8-TCDD and 2,3,7,8-TBDD metabo-
lism does not occur in the rat after exposure to sublethal doses of these
compounds (Kedderis et al., 1991b; Curtis et al., 1990; Olson et al., 1991). In
contrast to these compounds, 2,3,7,8-TCDF and 2,3,4,7,8-PeCDF can induce their
own rate of metabolism and biliary excretion (Brewster and Birnbaum, 1987;
McKinley et al., 1991; Olson et al., 1991). Autoinduction of metabolism would
suggest that these compounds may exhibit dose-related excretion, with longer
half-lives for elimination at lower doses, which are not associated with enzyme
induction. Further data are needed to test this hypothesis.
1.3.5. Excretion in Humans. Poiger and Schlatter (1986) investigated the
excretion of 2,3,7,8-TCDD in a 42-year-old man (92 kg) after ingestion of 105 ng
(1.14 ng/kg) [3H]-2,3,7,8-TCDD in 6 mL corn oil (see Table 1-8). The half-life
for elimination was estimated to be 2120 days. Table 1-9 summarizes additional
half-life estimates for 2,3,7,8-TCDD and related compounds in humans, based on
serum and/or adipose tissue concentrations at two or more time points. In
another study, the half-life of 2,3,7,8-TCDD in humans was estimated to be -7
years on the basis of 2,3,7,8-TCDD levels in serum samples taken in 1982 and 1987
from 36 of the Ranch Hand personnel who had 2,3,7,8-TCDD levels >10 ppt in 1987
(Pirkle et al., 1989). These studies indicate that 2,3,7,8-TCDD is exceedingly
persistent in humans. Estimated half-lives for other congeners in Table 1-9
range from 0.8-10 years. The half-life values in Table 1-9 are rough estimates
based on a small number of individuals and based on analysis at as few as two
1-61 08/11/92
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TABLE 1-9
Half-Life Estimates for 2,3,7,8-TCOD and Related Compound in Humans
Chemical
Exposure Incident
Number of
Individuals
Sample
Time Period
Between First and
Last Analysis
CDDs
2,3,7,8-TCDD
1,2,3,6,7,8-HxCDO
1,2,3,4,6,7,8-HpCDD
OCDD
Ranch Hand
Vietnam veterans
technical pentachlorophenol in
wood of home
technical pentachlorophenol in
wood of home
technical pentachlorophenol in
wood of home
36
1
1
1
serum
adipose tissue
adipose tissue
adipose tissue
CDFs
2,3,4,7.8-PeCDF
1,2,3,4,7,8-HxCDF
1,2,3,6,7,8-HxCDF
1,2,3,4,6,7,8-HpCDF
2,3,4,7,8-PeCDF
1, 2,3,4, 7,8-HxCDF
1,2,3,4,6,7,8-HpCDF
Binghamton, New York, state
office building
Binghamton, New York, state
office building
Binghamton, New York, state
office building
Binghamton, New York, state
office building
Yu-Cheng
Yu-Cheng
Yu-Cheng
1
1
1
1
4
3
2
4
3
2
4
3
2
adipose tissue
blood
combined
adipose tissue
blood
combined
adipose tissue
blood
combined
adipose tissue
blood
combined
blood
blood
blood
5 years
28 months
28 months
28 months
Number of
Time Points
2
2
2
2
initial 43 months
final 29 months
total 6 years
initial 43 months
final 29 months
total 6 years
initial 43 months
final 29 months
total 6 years
initial 43 months
final 29 months
total 6 years
initial 2.9 years
final 2.7 years
total 5.6 years
initial 2.9 years
final 2.7 years
total 5.6 years
initial 2.9 years
final 2.7 years
total 5.6 years
4
4
7
4
4
7
4
4
7
4
4
7
2
2
3
2
2
3
2
2
3
Half-Life
(years)
Reference
7.1a
3.5
3.2
5.7
4.7
7.2
4.5
2.9
4.4
4.0
3.5
4.3
4.9
6.5
4.1
6.8
1.3
2.9
1.7
2.1
5.1
2.4
1.6
6.1
2.4
Pirkle et
al., 1989
Gorski et
al., 1984
Gorski et
al., 1984
Gorski et
al., 1984
Schecter et
at., 1990
Schecter et
al., 1990
Schecter et
al., 1990
Schecter et
al., 1990
Ryan, 1989
Ryan, 1989
Ryan, 1989
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DRAFT—DO NOT QUOTE OR CITE
time points. Phillips (1989) discusses this issue. Estimates also assume a
simple, single compartment, first-order elimination process. Recent data suggest
a biphasic elimination of 2,3,7,8-TCDD, with a half-life >7 years (Birnbaum,
1992).
Ryan and Masuda (1991) reported on their continuing investigation into the
elimination of CDFs in humans from the Yusho and Yu-Cheng rice oil poisonings.
Yu-Cheng individuals had CDF blood levels on a lipid basis of 1-50 pg/kg, while
Yusho patients had levels of 0.1-5 pg/kg. In the Yu-Cheng individuals, half-
lives for three CDFs were 2-3 years, while elimination from Yusho individuals was
more variable and slower, with half-lives >5 years (see Table 1-9) and, in
several cases, no measurable elimination during the 7 years in which samples were
available. The limited results suggest that clearance of these CDFs in the human
is biphasic, with faster elimination at higher exposure. Schecter et al. (1990)
and Ryan (1989) also reported longer half-life values for CDFs in humans at later
time points after exposure, when concentrations are closer to the background
levels of individuals with no unusual exposure.
Due to the lipophilic nature of milk, milk secretion can provide a
relatively efficient mechanism for decreasing the body burden of 2,3,7,8-TCDD in
females. As discussed by Graham et al. (1986), this elimination of 2,3,7,8-TCDD
through mother's milk can result in high exposure levels in the infant. Since
both milk and the fatty tissues of fish are essentially providing an oily
vehicle, it would be likely that these sources would provide 2,3,7,8-TCDD in a
form that is readily bioavailable.
Several investigators have guantified the levels of 2,3,7,8-TCDD in human
milk samples. Many of the milk samples were pooled (Jensen, 1987). Rappe et al.
(1984) reported levels of 1-3 ppt 2,3,7,8-TCDD in milk fat (lipid adjusted) from
five volunteers in West Germany, and in a later report, Rappe et al. (1985)
reported an average level of 0.6 ppt 2,3,7,8-TCDD in milk fat from four
volunteers in northern Sweden. Furst et al. (1986) reported an average level of
9.7 ppt 2,3,7,8-TCDD in milk fat from three individuals in the Netherlands and
<1.0 ppt 2,3,7,8-TCDD in milk fat from two individuals in Yugoslavia. Nygren et
al. (1986) reported average levels of 2,3,7,8-TCDD in human milk samples from
four subjects in Sweden to be 0.6 ppt in milk fat, in five subjects from West
1-64 08/11/92
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DRAFT—DO NOT QUOTE OR CITE
Germany to be 1.9 ppt in milk fat, and in four subjects from Vietnam to be <0.5
ppt in milk fat.
High levels of 2,3,7,8-TCDD have been detected in the milk of mothers
exposed to high levels of 2,3,7,8-TCDD in the environment. Reggiani et al.
(1980) reported levels between 2.3 and 28.0 ppt 2,3,7,8-TCDD in whole milk from
mothers in Seveso. Baughman (1975) reported levels between 40.0 and 50.0 ppt
2,3,7,8-TCDD in whole milk from mothers in South Vietnam. Schecter et al. (1987)
also found high ppt levels of 2,3,7,8-TCDD in human milk samples from South
Vietnam. These authors found that samples taken in 1985 from South Vietnamese
mothers were comparable to the level of 2,3,7,8-TCDD presently found in North
American human milk samples (5 ppt).
1.4. PHYSIOLOGICALLY BASED PHARMACOKINETIC MODELS
PB-PK models have been developed for 2,3,7,8-TCDD in C57BL/6J and DBA/2J
mice (Leung et al., 1988), rats (Leung et al., 1990) and humans (Kissel and
Robarge, 1988). PB-Pk models incorporate known or estimated anatomical,
physiological and physicochemical parameters to describe quantitatively the
disposition of a chemical in a given species. PB-Pk models can assist in the
extrapolation of high-to-low dose kinetics within a species, estimating exposures
by different routes of administration, calculating effective doses and extrapo-
lating these values across species (Scheuplein et al., 1990). Table 1-10
summarizes the pharmacokinetic parameters for 2,3,7,8-TCDD that were used in
developing these PB-Pk models. In many cases, these parameters were estimated
from in vivo experimental data.
A five-compartment (blood, liver, fat, muscle/skin, viscera), flow-limited
PB-Pk model for 2,3,7,8-TCDD was developed for the Ah-responsive C57BL/6J mouse
and the Ah-nonresponsive DBA/2J mouse (Leung et al., 1988). The model also
included binding in the hepatic cytosol and hepatic microsomes and first-order
hepatic metabolism. There was general agreement between the simulated descrip-
tion generated by the model and the experimental disposition data of Gasiewicz
et al. (1983). The greater accumulation of 2,3,7,8-TCDD in the liver of the
C57BL/6J mouse, compared to the DBA/2J mouse, was not attributable to the 2-fold
greater total fat content in the DBA/2J strain. The authors suggested that
strain difference in hepatic disposition was due to differing affinity of
1-65 08/11/92
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TABLE 1-10
Pharmacokinetic Parameters for 2,3,7,8-TCOD Used in PB-Pk Models
PARTITION COEFFICIENT (TISSUE/BLOOD)
Liver
Fat
Richly perfused (kidney)
Slowly perfused (skin)
Slowly perfused (muscle)
BIOCHEMICAL CONSTANTS
Binding capacity to hepatic cytosolic protein (nmol/liver)
Binding affinity to hepatic cytosolic protein (nM)
Binding capacity to hepatic microsomal protein (nmol/liver)
Noninduced binding capacity to hepatic microsomal protein (nmol/liver)
Induced binding capacity to hepatic microsomal protein (/unol/liver)
Binding affinity to hepatic microsomal protein (nM)
First-order metabolic rate constant (per hour per kg liver)
Absorption constant from gastrointestinal tract into liver (per hour)
Binding to blood
C57BL/6J
Mouse3
DBA/2J
Mouse9
Sprague-Dawley
RatD
Human0
20
350
20
250
250
20
350
20
250
250
20
350
20
40
40
25
300
7-10
30
4
Female C57BL/6J Miced
Naive
10
300
10
200
3
Pretreated
10
300
10
200
3
0.0042
0.29
20
-
-
20
3.25
0.02
2.5
0.0042
2
20
-
-
75
1.75
0.02
2.5
0.054
0.015
-
25
175
7
2.0
0.2
2.5
-
-
-
-
-
-
-
-
-
0.0042
0.29
1.75
-
-
20
1.0
0.04
1.0
0
0.29
20
-
-
20
3.0
0.15
3.0
O
O
1
I
w
3
(-3
M
\eung et al. (1988)
bLeung et al. (1990)
GKissel and Robarge (1988)
i 1 OC
Leung et al. (1990) modeled the disposition of [ "l]-2-iodo-3,7,8-TCDD, an analog of 2,3,7,8-TCOD, following a single exposure (0.1 nmol/kg)
in naive female C57BL/6J mice and in mice pretreated 3 days earlier with an inducing dose of 2,3,7,8-TCDD (0.1 /unol/kg).
o
oo
vo
N)
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2,3,7,8-TCDD for the microsomal binding protein in the two strains, and proposed
using a different microsomal dissociation constant for each strain. Alterna-
tively, the strain difference in hepatic disposition may be due to the different
doses of 2,3,7,8-TCDD needed to induce cytochrome P-4501A2 in the two strains.
In contrast to the high capacity/low affinity hepatic microsomal binding
proteins, the low capacity/high affinity hepatic cytosolic binding protein (Ah
receptor) did not play a major role in determining the overall tissue distribu-
tion pattern of 2,3,7,8-TCDD in this model.
A similar five-compartment PB-Pk model was developed to describe the tissue
disposition of 2,3,7 8-TCDD in the Sprague-Dawley rat (Leung et al., 1990). This
description included blood, liver (cytosolic receptor and microsomal binding
protein), fat, muscle/skin and visceral tissue compartments. The authors found
generally good agreement between the PB-Pk model simulated data and the experi-
mental data for the single-dose study of Rose et al. (1976) and the 7- and
13-week multiple-dose studies of Kociba et al. (1978). The model was not
satisfactory for the 2-year feeding study of Kociba et al. (1978), underpre-
dicting the 2,3,7,8-1000 concentration in the fat at the low dose (0.001
pg/kg/day) and overestimating the concentration achieved at the high dose (0.1
pg/kg/day). The model found the hepatic disposition of 2,3,7,8-TCDD to be
dependent on the high capacity/low affinity hepatic microsomal binding protein
with a dissociation constant of 7 nM and a basal and induced concentration in the
liver of 25 and 175 nmol/liver, respectively. As discussed earlier, Voorman and
Aust (1987, 1989) and Poland et al. (1989a,b) provided evidence that this binding
species is cytochromg P-4501A2. Induction of the microsomal binding protein was
necessary in order to account for the differences in hepatic disposition at low
(0.01 /jg/kg) and high (1.0 ^g/kg) daily doses of 2,3,7,8-TCDD. The dose-
dependent tissue distribution of 2,3,7,8-TCDD was also discussed earlier. As in
the mouse PB-Pk model (Leung et al., 1988), the low capacity/high affinity
hepatic cytosolic binding protein (Ah receptor) was not a major factor in
directly influencing the hepatic disposition of 2,3,7,8-TCDD. The dissociation
constant of the cytosolic Ah receptor in vivo was estimated to be 15 pM by
fitting enzyme induction data from McConnell et al. (1984).
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A PB-Pk model was also developed for female C57BL/6J mice for [125I]-2-iodo-
3,7,8-TCDD, an analog of 2,3,7,8-TCDD (Leung et al., 1990). Mice were pretreated
with 0.1 fjmol/kg of 2,3,7,8-TCDD or the vehicle only, followed by 2-iodo-3,7,8-
TCDD (0.1 nmol/kg) 3 days later. Naive mice had liver/fat 2-iodo-3,7,8-TCDD
concentration ratios of 0.17-0.38, while the 2,3,7,8-TCDD pretreated mice had
ratios of 2.0-6.1. This is in agreement with the dose-dependent tissue distribu-
tion of 2,3,7,8-TCDD described earlier (Abraham et al., 1988; Poiger et al.,
1989). As with 2,3,7,8-TCDD, the model found that the 2-iodo-3,7,8-TCDD
concentration in the liver was most sensitive to the binding capacity of the
hepatic microsomal protein. Whole-body elimination of 2-iodo-3,7,8-TCDD
approximated first-order kinetics, and induction by pretreatment with an inducing
dose of 2,3,7,8-TCDD almost doubled the rate of excretion (ti^ of 14.2 days in
naive versus 8.0 days in induced mice) (see Table 1-8). The distribution in
naive and pretreated mice was described by a PB-Pk model in which induction
(2,3,7,8-TCDD pretreatment) increased the amount of hepatic microsomal binding
protein from 1.75-20 nmol/liver and increased the rate constant for metabolism
of free 2-iodo-3,7,8-TCDD from 1-3 hours/kg liver. Although the more rapid
elimination of 2-iodo-3,7,8-TCDD in 2,3,7,8-TCDD pretreated mice suggests that
the rate of metabolism of 2-iodo-3,7,8-TCDD was induced by pretreatment, no data
were provided on the effect of this pretreatment of body weight and composition,
which may in turn alter the rate of elimination. 2,3,7,8-TCDD pretreatment may
also alter deiodinase activity. Furthermore, in vivo and in vitro studies
suggest that the autoinduction of 2,3,7,8-TCDD metabolism may not occur under
these conditions. Kedderis et al. (1991b) and Curtis et al. (1990) found no
autoinduction of 2,3,7,8-TCDD metabolism and biliary excretion in the rat. In
addition, Shen and Olson (1987) found that while 2,3,7,8-TCDD pretreatment of
C57BL/6J mice increased the uptake of 2,3,7,8-TCDD by hepatocytes in suspension
culture, pretreatment did not increase the rate of metabolism of 2,3,7,8-TCDD by
hepatocytes. Therefore, this PB-Pk model may not accurately describe the
metabolism of 2,3,7,8-TCDD for exposures which result in varying degrees of
induction of the hepatic monooxygenase system. While the dose-dependent
pharmacokinetics of 2,3,7,8-TCDD may not include autoinduction of 2,3,7,8-TCDD
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metabolism, this may not be the case for other CDDs, HDDs, CDFs, BDFs, PCBs and
PBBs. For example, the autoinduction of metabolism has been reported for CDFs
(Brewster and Birnbaum, 1987; McKinley et al., 1991; Olson et al., 1991).
Andersen et al. (1992) recently derived a receptor-mediated PB-PK model for
the tissue distribution and enzyme inducing properties of 2,3,7,8-TCDD. The data
used for this analysis were from two previously published studies with Wistar
rats (Abraham et al., 1988; Krowke et al., 1989). The model was used to examine
the tissue disposition of 2,3,7,8-TCDD and the induction of both a dioxin-binding
protein (presumably cytochrome P-4501A2) and cytochrome P-4501A1.
Kohn et al. (1992) recently developed a mechanistic model of the effects of
dioxin on glue expression in the rat liver (referred to as the NIEHS model). The
model includes the tissue distribution of 2,3,7,8-TCDD in the rat and its effect
on the concentrations of CYP1A1 and CYP1A2, and the effects of 2,3,7,8-TCDD on
the Ah, estrogen and EGF receptors over a wide 2,3,7,8-TCDD dose range.
Experimental data from Tritscher et al. (1992) and Sewall et al. (1992) were
incorporated into the NIEHS model. Female Sprague-Dawley rats were injected with
an initiating dose of diethylnitrosamine, and after 20 days, the rats were
exposed biweekly to 2,3,7,8-TCDD in corn oil by gavage at doses equivalent to
3.5-125 ng/kg/day for 30 weeks. The NIEHS model predicts a linear relationship
between administered dose and the concentration in the liver over this dose
range, which is in agreement with the data of Tritscher et al. (1992). The
biochemical response curves for all these proteins were hyperbolic, indicating
a proportional relationship between target tissue dose and protein concentration
at low administered doses of 2,3,7,8-TDCDD.
A fugacity-based PB-Pk model for the elimination of 2,3,7,8-TCDD from humans
was developed by Kissel and Robarge (1988). Transport within the body was
assumed to be perfusion-limited (flow-limited). 2,3,7,8-TCDD was assumed to be
uniformly distributed within each tissue or fluid phase, and tissue levels were
considered to be in equilibrium with exiting fluids (blood, bile, urine).
2,3,7,8-TCDD appears to be poorly metabolized in humans, thus reducing the
necessity of modeling the fate of metabolites. 2,3,7,8-TCDD also seems to
exhibit fugacity-based partitioning behavior in humans as evidenced by relatively
constant lipid-based tissue distribution (Leung et al., 1990; Ryan et al., 1987),
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although this is not the case in rodents (Leung et al., 1988, 1990). With a
daily human background intake of 2,3,7,8-TCDD in North America of -50 pg/day
(Travis and Hattemer-Frey, 1987), the steady state adipose tissue concentration
predicted by the model, assuming no metabolism, was 7.7 ppt. This is similar to
the lipid-based blood tissue levels reported in the general population with no
known unusual exposure. The model was also used to predict the elimination of
2,3,7,8-TCDD from Ranch Hand Vietnam veterans. The model simulation assumed a
background exposure of 50 pg/day and no metabolism. Under these conditions,
apparent half-lives of 4.4, 5.2, 5.9, 7.2, 9.1 and 20 years were estimated for
individuals with adipose tissue concentrations of 100, 50, 30, 20, 15 and 10 ppt,
respectively. The model predicted half-lives are similar to the experimental
value of 7.1 years, based on analysis of 2,3,7,8-TCDD in blood lipids of veterans
with adipose burdens greater than 10 ppt (Pirkle et al., 1989) (see Table 1-9).
The apparent half-lives derived from the model increased as the adipose tissue
concentrations approached the steady-state level associated with background
exposure. Ryan and Masuda (1991) also reported a similar relationship for CDFs,
with experimentally derived half-lives increasing in individuals with lower body
burdens of the compounds. Finally, the model was also found to approximate the
elimination of 2,3,7,8-TCDD from one volunteer as reported by Poiger and
Schlatter (1986). Taken together, the comparisons described above suggest that
a fugacity-based PB-Pk model for 2,3,7,8-TCDD in humans can provide one method
for describing the elimination of 2,3,7,8-TCDD from humans.
Kedderis et al. (1992a,b) recently developed a PB-PK model for 2,3,7,8-TBDD
in the rat. The model is based on previously developed physiologically-based
models for 2,3,7,8-TCDD (Leung et al., 1990; Poland et al., 1989) and utilizes
published data on the disposition of a single exposure to 2,3,7,8-TBDD at a dose
of 1 or 100 nmol/kg, intravenous (Kedderis et al., 1991a,b) and dermal
disposition data (Jackson et al., 1991). In the model, the dose- and time-
dependent accumulation in the liver was attributed to specific binding of
2,3,7,8-TBDD with the inducible protein, CYP1A2. The model also includes
diffusion-limited tissue uptake of 2,3,7,8-TBDD, transluminal excretion of parent
compound via the gut into the feces, growth of tissue compartments and a separate
skin compartment. This model provides further validation of the model structure
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originally developed to describe important dispositional determinants for
2,3,7,8-TCDD.
A five-compartment (blood, liver, fat, skin, muscle) flow-limited physiolog-
ical model was developed to describe the tissue distribution and excretion of
2,3,7,8-TCDF-derived material in rats, mice and monkeys (King et al., 1983),
based on experimental data reported earlier (Birnbaum et al., 1980, 1981; Decad
et al., 1981b). Partition coefficients (tissue/blood distribution ratios) and
metabolic clearances were estimated from in vivo experimental data and are
summarized in Table 1-11. All pharmacokinetic parameters for 2,3,7,8-TCDF were
based on in vivo data after a single intravenous exposure at a dose of 0.1
pmol/kg (30.6 pg/kg). Therefore, the model is limited in not considering the
potential dose-related distribution and excretion of 2,3,7,8-TCDF. Recent
studies indicate that 2,3,7,8-TCDF is able to induce its own rate of metabolism
and biliary excretion at higher doses (McKinley et al., 1991; Olson et al.,
1991). This model will need to be revised as additional data on the dose-related
distribution and excretion of 2,3,7,8-TCDF become available.
PB-Pk models are primarily limited by the availability of congener and
species-specific date that accurately describe the dose- and time-dependent
disposition of 2,3,7,8-TCDD and related compounds. The pharmacokinetic
parameters summarized in Tables 1-10 and 1-11 were derived from available in vivo
and in vitro experimental data. As additional data become available, partic-
ularly on the dose-dependent disposition of these compounds, more accurate models
can be developed. Jn developing a suitable model in the human, it is also
important to consider that the half-life estimate of 7.1 years for 2,3,7,8-TCDD
was based on two se?um values taken 5 years apart, with the assumption of a
single compartment, first-order elimination process (Pirkle et al., 1989). It
is likely that the excretion of 2,3,7,8-TCDD in humans is more complex, involving
several compartment, tissue-specific bonding proteins and a continuous daily
background exposure. Furthermore, changes in body weight and body composition
should also be considered in developing PB-Pk models for 2,3,7,8-TCDD and related
compounds in humans.
An empirical model of dioxin (toxic equivalents) disposition in animals and
humans has also been recently developed by Carrier and Brodeur (1991). The
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TABLE 1-11
Pharmacokinetic Parameters for 2,3,7,8-TCDF Used in the PB-Pk Model
Described by King et al. (1983)
C57BL/6J
Mouse
DBA/2J
Mouse
Fischer 344
Rat
Rhesus
Monkey
PARTITION COEFFICIENTS
Liver
Fat
Skin
Muscle
130
25
8
2
100
40
12
4
100
35
4
2
30
30
7
2
CLEARANCES
Metabolism
Km (mL/minute/kg)
Metabolism excretion ratio K^/K^*
0.07
2.8
0.14
0.06
2.4
O.27
1.0
4.0
0.03
2.25
0.45
O.19
aUrinary clearance/biliary clearance
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kinetic analysis begins with the observation that the tissue distribution of
dioxin-like HPAH in humans and in animals is dose-dependent (or, alternately,
body-burden dependent). As total body burden of TCDD equivalents increases, the
proportion of the body burden associated with the liver increases toward a
maximum value. The data were then analyzed with an empirical, saturable binding
isotherm equation:
liver fraction (fH) =
If the body burden— Cj^y (pg/kg)— is considered a surrogate for liver concentra-
tion, this equation can be loosely interpreted as the induction of binding
species in the liver as dose increases. In the analyses, Kd was found to be very
similar for people and experimental animals, indicating similar protein induction
dynamics in various animal species. This model, however, is not physiologically-
based and the terms, Cj^,, and f^x, are difficult to interpret in biological
terms. In working with different isomers, fmax and K^ values vary somewhat,
presumably due to binding affinities in the liver.
This empirical model is successful in providing a description that "fits"
the observed data in various species. It still is largely a fitting exercise to
a particular equation, not an examination of biology by computer modeling. In
addition, there are at least two assertions that seem incorrect. First, the
assumption that the limit of the hepatic fraction at very low doses is zero. It
seems more likely that the limit is some finite value, determined by liver
partitioning of dioxin and the binding parameters of the Ah receptor, and the
dioxin binding species in the liver in the linear, low-dose region. Secondly,
that metabolism of dioxin becomes saturated with the maximum induction of liver
sequestration of dioxin. There is no justification for this at present.
Nevertheless, the model indicates clearly that with respect to dosimetry and
induction of hepatic binding species for dioxin, people and rodents are very
similar. Furthermore, the empirical model of Carrier and Brodeur (1991) is
generally consistent with the PB-PK models.
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1.5. PHARMACOKINETICS IN SPECIAL POPULATIONS
1.5.1. Pregnancy and Lactation (Prenatal and Postnatal Exposure of Offspring).
The distribution and excretion of [14C]-2,3,7,8-TCDD (30 pg/kg) and [14C]-
2,3,7,8-TCDF (800 /t/g/kg) were studied in pregnant C57BL/6N mice after oral
exposure on gestation day 11 (Weber and Birnbaum, 1985). The distribution and
excretion of 2,3,7,8-TCDD and 2,3,7,8-TCDF in pregnant mice were similar to that
of males of the same strain (Gasiewicz et al., 1983; Decad et al., 1981b) (see
Tables 1-5 and 1-8), although elimination rates were higher in the pregnant mice
for both congeners. For 2,3,7,8-TCDD, liver, urinary and fecal elimination were
3.0, 3.4 and 14.4 times faster than that reported for males. For 2,3,7,8-TCDF,
liver, urinary and fecal elimination were 1.3, 1.8 and 1.8 times faster than
observed for males. Elimination data from pregnant mice was based on only three
time points (gestation days 12, 13 and 14) and thus represents only rough
estimates. In addition, the greater fecal excretion could have been due to
incomplete absorption of 2,3,7,8-TCDD after oral exposure. Although these
results need further substantiation, it is conceivable that the sex of the
animal, pregnancy and/or the route of exposure could have a significant impact
on the pharmacokinetics of these compounds.
In a related study, Krowke (1986) compared the 2,3,7,8-TCDD concentrations
in the liver of pregnant and nonpregnant NMRI mice exposed subcutaneously to 12.5
or 25 nmol/kg/day on gestation days 9-11. At 7 days after exposure to the lower
dose, the hepatic 2,3,7,8-TCDD concentrations were 7 and 32 ng/g in pregnant and
nonpregnant mice, respectively. At the higher exposure, 5.5 times lower
concentrations of 2,3,7,8-TCDD were found in the livers of pregnant animals on
gestation day 18. A similar effect on hepatic 2,3,7,8-TCDD levels was observed
also in combined exposure, which contained 1,2,3,7,8-PeCDD, 1,2,3,4,7,8-HxCDD or
2,3,7,8-TCDF. The decreased hepatic levels of 2,3,7,8-TCDD in pregnant mice are
consistent with the Weber and Birnbaum (1985) observation of more rapid elimina-
tion of 2,3,7,8-TCDD in pregnant mice. Further investigations are necessary to
better characterize the apparently significant effects of pregnancy on the
disposition of 2,3,7,8-TCDD and related compounds.
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Weber and Birnbaum (1985) also investigated the distribution of [14C]-
2,3,7,8-TCDD (30 ^g/kg) and [14C]-2,3,7,8-TCDF (800 ^g/kg) to the embryos of
pregnant C57BL/6N mice after oral exposure on gestation day 11. On gestation
days 12, 13 and 14, the percent of the maternal dose in the embryo remained
constant at 0.032-0.037%/ embryo, while the concentrations in the embryo were
0.34, 0.17 and 0.15% of the dose/g embryo, respectively. Embryos had approxi-
mately 11-fold higher concentrations of 2,3,7,8-TCDD than 2,3,7,8-TCDF when
exposed on a percent of total dose/g tissue basis. This may be due to the more
rapid metabolism and excretion of 2,3,7,8-TCDF compared to 2,3,7,8-TCDD.
Assuming that all radioactive material found in embryos was parent compound, at
most, 2.6 ng (8 pmol) of 2,3,7,8-TCDD and 6.4 ng (21 pmol) of 2,3,7,8-TCDF/g
tissue were detected under these conditions.
The transfer of [14C]-2,3,7,8-TCDD to the embryo during early gestation was
assessed in NMRI mice given a dose of 25 pg/kg by intraperitoneal injection on
either days 7, 8, 9, 10, 11 or 13 of gestation (Nau and Bass, 1981). The mice
were sacrificed after 48 hours, and 2,3,7,8-TCDD concentrations were determined
by liquid scintillation counting of solubilized tissue and by GC-ECD and GC/MS.
Similar results were given by these methods, suggesting that 2,3,7,8-TCDD derived
[ C] in maternal and embryonic tissue was the parent compound. The maternal
liver contained from 4-8% of the dose/g or 40-80 ng/g. 2,3,7,8-TCDD in embryonic
tissue from gestation days 11-15 ranged from 0.04-0.1% of the dose/g or 0.4-1.0
ng/g. In contrast, higher levels were found earlier in gestation, with 10 ng/g
embryo on gestation day 9 and 2 ng/g on day 10. The higher levels may be related
to placentation, which occurs at approximately gestation days 10-11 in this mouse
strain. The affinity of fetal liver for 2,3,7,8-TCDD was relatively low, as
compared to maternal liver; however, 2,3,7,8-TCDD levels in fetal livers were 2-4
times higher than levels in other fetal organs. Nau and Bass (1981) also
attempted to correlate 2,3,7,8-TCDD levels in the fetuses with the observed
incidence of cleft palate. Three groups of mice were given either a single
intraperitoneal exposure to 25 /jg/kg 2,3,7,8-TCDD on gestation day 7 or 10 or 5
pg/kg/day, intraperitoneally, on gestation days 7-11. On gestation day 13,
2,3,7,8-TCDD concentrations in maternal tissues were very similar in the three
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exposure groups. At day 13, however, the embryo contained 0.038±0.011% (0.36
ng/g), 0.096±0.027% (0.92 ng/g) and 0.12±0.05% (1.1 ng/g) of the dose (mean±SD)
in the 7-, 10- and 7- to 11-day exposure groups, respectively. Cleft palate
incidence on gestation day 18 was 16, 84 and 65% for the 7-, 10- and 7- to 11-day
exposure groups, respectively. Although further studies are needed, these
results suggest that cleft palate incidence is generally related to the 2,3,7,8-
TCDD concentration in the embryo. In a related study, Couture et al. (1990)
found that gestation day 12 was the peak period of sensitivity for 2,3,7,8-TCDD-
induced cleft palate in C57BL/6N mice; however, tissue levels were not investi-
gated .
In the same laboratory, Abbott et al. (1989) investigated the distribution
of 2,3,7,8-TCDD in the C57BL/6N mouse fetus following maternal exposure on
gestation day 11 to 30 pg/kg. 2,3,7,7-TCDD was detected in the gestation day-11
embryo at 3 hours post-exposure and was equally distributed between the embryonic
head and body. At 72 hours post-exposure, 0.035% of the total dose was in fetal
tissues, and 1% of the 2,3,7,8-TCDD in the fetus (1.4-3.5 pg was found in the
palatal shelf.
Krowke (1986) also measured the concentration of 2,3,7,8-TCDD in the
placenta, amniotic fluid and fetus of NMRI mice exposed to 2.5 nmol/kg by
subcutaneous injection on days 9-11 of gestation. Similar concentrations of
2,3,7,8-TCDD were observed in the placenta, amniotic fluid and fetus (-0.5 ng/g)
on day 16 of gestation. Fetal liver 2,3,7,8-TCDD concentrations were at least
five times greater than other fetal tissue. Krowke (1986) reported slightly
lower 2,3,7,8-TCDD levels in the fetal head relative to other extrahepatic fetal
tissue, while Weber and Birnbaum (1985) found a slightly higher 2,3,7,8-TCDD
concentration in the head relative to other extrahepatic fetal tissue.
Nau et al. (1986) investigated the transfer of 2,3,7,8-TCDD via the placenta
and milk in NMRI mice exposed to 25 /^g/kg on day 16 of gestation. The authors
confirmed the relatively low fetal tissue levels with prenatal exposure to
2,3,7,8-TCDD (Nau and Bass, 1981) and found that postnatally, 2,3,7,8-TCDD was
transferred efficiently to mouse neonates and offspring by lactating mothers.
During the first 2 postnatal weeks, the pups were given doses of 2,3,7,8-TCDD via
the milk that were, on a body weight basis, similar to those that had been
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administered prenatally to their mothers. 2,3,7,8-TCDD levels in the tissue of
lactating mothers decreased within the first 3 postnatal weeks by two to three
orders of magnitude to reach levels that were only -2% of the corresponding
levels in the pups that these mothers had nursed. Thus in mice, excretion into
milk represents a major pathway for maternal elimination of 2,3,7,8-TCDD and for
the subsequent exposure of pups.
The disposition of 2,3,7,8-TCDD in rat pups was assessed after the prenatal
(via placental transfer) and/or postnatal (via milk) exposure from pregnant
Wistar rats given a single dose of 3, 30 or 300 ng/kg, subcutaneously, on day 19
of gestation (Korte et al., 1990). Lactation resulted in the rapid elimination
of 2,3,7,8-TCDD from maternal tissues, with the half-life of 2,3,7,8-TCDD in the
liver of lactating rats estimated to be -7 days. This compares to a half-life
of 13.6 days in the liver of nonlactating rats (Abraham et al., 1988). At
postnatal day 7, exposure via the milk resulted in pup liver 2,3,7,8-TCDD
concentrations that were greater than the corresponding levels in maternal liver.
In cross-fostering experiments, the concentrations of 2,3,7,8-TCDD in the liver
of offspring at postnatal day 7 were 0.47, 2.59 and 4.16 ng/g in the 300 ng/kg
groups exposed through the placenta only, via the milk only and through the
placenta and via the milk, respectively. These results support the earlier
observations that the placental transfer of 2,3,7,8-TCDD in rats and mice is
relatively limited compared with the efficient transfer via maternal milk.
Van den Berg et al. (1987) investigated the transfer of CDDs and CDFs to
fetal and neonatal rats. Prenatal exposure of the fetus was assessed in pregnant
Wistar rats fed a diet containing a fly ash extract from a municipal incinerator
on days 10-17 of gestation. Postnatal exposure of 10-day-old pups was assessed
through feeding lactating mothers the same contaminated diet for the first 10
days after delivery. Although the fly ash extract contained almost all of the
136 tetra- to octa-CDDs and -CDFs, only 17 CDD and CDF congeners were detected
as major compounds in the tissue of fetuses, pups and dams. All of the congeners
were 2,3,7,8-substituted with the exception of 2,3,4,6, 7-PeCDF. 2,3,7,8-TCDD had
the highest retention (0.0092% of the dose/g) in the fetus, while 2,3,7,8-TCDF,
1,2,3,7,8-PeCDF and hepta- and octa-CDDs and -CDFs were not detected in the
fetus. In the liver of offspring, the highest retention was found for 2,3,7,8-
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TCDD, 1,2,3,7,8-PeCDD and the three 2,3,7,8-substituted HxCDDs (0.74-1.13%
dose/g). The 2,3,7,8-substituted penta- and hexachlorinated congeners showed the
highest retention in the livers of dams (2.05-5.17% of dose/g liver), while
2,3,7,8-TCDF, 1,2,3,7,8-PeCDF and 2,3,4,6,7-PeCDF had the lowest retention. A
linear relationship was found between the retention of CDDs and CDFs in the
livers of pregnant and lactating rats. Furthermore, a linear relationship was
found between the retention of CDDs and CDFs in the livers of the lactating rats
and livers of the offspring.
In a related study, Hagenmaier et al. (1990) investigated the transfer of
CDDs and CDFs through the placenta and via milk in a marmoset monkey. A defined
mixture of CDDs and CDFs was given as a single subcutaneous injection to a
pregnant marmoset monkey at the end of the organogenesis period (week 10 of
gestation, 11 weeks prior to delivery). Transfer of CDDs and CDFs through the
placenta was investigated in a newborn 1 day after birth, and transfer through
the placenta and via milk was assessed in an infant of the same litter after a
lactation period of 33 days. Tissue concentrations of the offspring were
compared with those of the mother at the end of the lactation period and with
data from other adult marmosets obtained at this time of maximum absorption (1
week after injection) and 6 weeks after injection. Deposition of CDDs and CDFs
into the newborn liver was very low, suggesting very little transplacental
transport and hepatic accumulation of these compounds. 2,3,7,8-TCDD and
1,2,3,7,8-PeCDD were found at the highest concentration in the liver of the
newborn (-0.15% of dose/g). For all other congeners, the concentrations in the
liver of the newborn were <10% of the corresponding concentrations in adults.
In contrast to liver, concentrations of 2,3,7,8-substituted congeners in the
adipose tissue of the newborn were at least 33% of the levels in adults, and in
the case of OCDD and OCDF, levels were 3-fold higher in the newborn than in the
adult. The adipose tissue/liver concentration ratios for 2,3,7,8-substituted
congeners in the newborn ranged from 2.2 for 1,2,3,4,6,7,8-HpCDF to 10.9 for
2,3,7,8-TCDF. Furthermore, the concentration of these congeners in the newborn
was highest in the adipose tissue, followed by the skin and liver. This is in
contrast to the relative distribution in the adult where the liver generally
contains the highest levels of these congeners. The results indicate that
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hepatic concentrations in the fetus may not be representative of the rate of
placental transfer of CDDs and CDFs. In the marmoset monkey, substantial
placental transfer into fetal adipose tissue can be observed for most of the
2,3,7,8-substituted congeners during the fetal period. As expected from rodent
studies, the transfer of CDDs and CDFs via mothers' milk was considerable,
resulting in hepatic concentrations of 2,3,7,8-TCDD, 1,2,3,7,8-PeCDD and
1,2,3,6,7,8-HxCDD in the suckled infant (postnatal day 33) higher than those in
the dam. The hepatic concentration of 2,3,7,8-TCDD in the 33-day-old infant was
-0.9% of the dose/g tissue. Transfer of hepta- and octa-CDDs and CDFs to the
suckled infant was rather low, only -10% of the levels in the dam. When total
exposure of the mother and offspring at the end of the 33-day nursing period was
assessed in terms of I-TE factors (U.S. EPA, 1989), the liver of the mother
contained 2494 pg I-TE/g, while the offspring liver contained 2022 pg I-TE/g.
This approach is necessary to assess total exposure due to the congener-specific
transfer via lactation.
The pre- and postnatal transfer of 2,3,7,8-TCDD to the offspring of rhesus
monkeys was investigated by Bowman et al. (1989). Animals were fed a diet
containing 2,3,7,8-TCDD at concentrations of 5 or 25 ppt for -4 years and were
on a 2,3,7,8-TCDD-free diet for -18 months prior to parturition. Maternal
2,3,7,8-TCDD levels (mean±SE) in adipose tissue were 49±11 (n=7) and 173±81 (n=3)
ppt in the 5 and 25 ppt groups, respectively. Corresponding levels in the
adipose tissue of offspring at weaning (4 months) were 187±58 an 847±298 ppt in
the 5 and 25 ppt groups, respectively. From these data, a 2,3,7,8-TCDD BCF of
4.29 was estimated from mother to nursing infant. This value is similar to that
observed for 2,3,7,8-TCDD in the marmoset monkey (Hagenmaier et al., 1990). The
milk of the rhesus monkeys in the 25 ppt group contained from 4-14 ppt of
2,3,7,8-TCDD, which corresponds to 150-500 ppt on a lipid basis. The authors
calculated that the three mothers in the 25 ppt group excreted from 17-44% of
their 2,3,7,8-TCDD body burden by lactation. They also concluded that the
results are generally consistent with overall triglyceride movement as mediating
the excretion of 2,3,7,8-TCDD in milk.
In a subsequent study, Bowman et al. (1990) reported the relative persis-
tence of 2,3,7,8-TCDD in the offspring of rhesus monkeys that were exposed
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earlier to 5 or 25 ppt of 2,3,7,8-TCDD in the diet. The concentration of
2,3,7,8-TCDD in adipose tissue was measured in offspring at -4-5, 12 and 24
months of age. The decrease of 2,3,7,8-TCDD levels in adipose tissue of seven
young monkeys departed somewhat from first-order, single-compartment kinetics,
but with the limited data and an assumption of first-order kinetics, a half-life
of 121 days was estimated. When the data were adjusted within each animal for
body weight gain and for average fat content at each age, the adjusted data
apparently followed first-order, single-compartment kinetics, with a half-life
of -181 days. Thus, young monkeys apparently eliminate 2,3,7,8-TCDD from adipose
tissue at a faster rate than adult rhesus monkeys, which had individual half-
lives ranging from 180-550 days (Bowman et al., 1989).
Furst et al. (1989) examined the levels of CDDs and CDFs in human milk and
the dependence of these levels on the period of lactation. The mean concentra-
tions of CDDs in human milk (on a fat basis) ranged from 195 ppt for OCDD to 2.9
ppt for 2,3,7,8-TCDD, with the levels of the other congeners decreasing with
decreasing chlorination. This is in contrast to the generally lower levels of
CDFs in human milk, which range from 25.1 ppt for 2,3,4,7,8-PeCDF to 0.7 ppt for
1,2,3,7,8-PeCDF. An evaluation of the CDD and CDF levels in relation to the
number of breast-fed children found that the concentrations in milk generally
decreased with the greater number of children. The CDD and CDF levels in milk
from mothers nursing their second child are on average 20-30% lower than those
for mothers breast-feeding their first child. CDD and CDF levels were also
analyzed in one mother over a period of 1 year after delivery of her second baby
to assess the effect of duration of lactation. After breast-feeding for 1 year,
the mother had CDD and CDF levels that were 30-50% of the starting concentration.
Levels in milk fat (ppt) at 1, 5 and 52 weeks after delivery were 251, 132 and
119 for OCDD, 7.9, 5.9 and 1.4 for 2,3,7,8-TCDD and 33.1, 24.5 and 10 for
2,3,4,7,8-PeCDF, respectively. The results suggest a more rapid mobilization of
CDDs and CDFs and excretion into human milk during the first few weeks post-
partum. Although further studies are necessary, the limited data suggest that
there are time-dependent, isomer-specific differences in the excretion of CDDs
and CDFs in human milk.
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Although data are more limited for the coplanar PCBs, 3,3',4,4'-TCB,
3,3',4,4',5-PeCB and 3,3 ' ,4,4',5,5'-HxCB have been detected in human milk from
Swedish mothers, at concentrations of 16-32, 72-184 and 46-129 ppt on a fat
basis, respectively (Noren et al., 1990). Therefore, lactation appears to be an
effective means for the excretion of coplanar PCBs from mothers and a major
source of postnatal exposure of nursing infants. Since 3,3',4,4',5-PeCB and
other coplanar PCBs are present in human milk at concentrations up to 60-fold
higher than 2,3,7,8-TCDD, it is important to consider the relative toxic potency
of these dioxin-like compounds and their potential health impact on nursing
infants.
1.5.2. Aging. The influence of aging on the intestinal absorption of 2,3,7,8-
TCDD was studied in 13-week-, 13-month- and 26-month-old (senescent) male Fischer
344 rats (Hebert and Birnbaum, 1987). Absorption was measured by an in situ
intestinal recirculation perfusion procedure. When absorption was calculated in
terms of ng 2,3,7,8-TCDD absorbed/g mucosal dry weight/hour, the decrease between
the senescent rats and the two younger age groups, from 544 ng/g/hour (young) to
351 ng/g/hour (senescent), was not statistically significant (p<0.05). The
results indicate that, as with other molecules that depend on diffusion for their
absorption, aging does not affect the intestinal absorption of 2,3,7,8-TCDD.
Banks et al. (1990) studied the effect of age on the dermal absorption and
disposition of 2,3,7,8-TCDD and 2,3,4,7,8-PeCDF in male Fischer 344 rats. When
rats were administered the same dose per body weight, dermal absorption of
2,3,7,8-TCDD, at 3 days after exposure, decreased from 17.7±2.7% (mean±SD) to
5.6±2.5% of the administered dose in 10- and 36-week-old rats, respectively.
Dermal absorption in the 96-week-old rats was similar to that of the 36-week-old
rats. Dermal absorption of 2,3,4,7,8-PeCDF also decreased from 22.2±0.2 to
14.7±3.8% of the administered dose in 10- and 36-week-old rats, respectively.
Dermal absorption of both compounds was also decreased in older rats given the
same total dose per surface area. Older animals may have decreased blood flow
in the upper dermis, which will decrease the clearance of these compounds from
the application site. Potential age-related changes in the intercellular stratum
corneum lipids may also play a role in the decreased dermal absorption observed
in older animals. Changes in the percentage of the administered dose detected
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in various depots reflected age-related changes in dermal absorption, while age-
related changes in the tissue distribution of the absorbed dose reflected changes
in the total mass of these tissues at various ages. Overall elimination of the
absorbed dose was not affected by age. Although this investigation was conducted
using a lipophilic solvent system and an animal model with skin that is more
permeable than human skin, the results suggest that systemic bioavailability
after dermal exposure to 2,3,7,8-TCDD or 2,3,4,7,8-PeCDF may be reduced in older
age groups.
In a similar study, absorption, tissue distribution and elimination were
examined 72 hours after dermal application of a lower dose of 200 pmol (111
2
pmol/cnr) 2,3,7,8-TCDD to weanling (3-week-old), juvenile (5-week-old), pubescent
(8-week-old), young adult (10-week-old) and middle-aged (36-week-old) rats
(Anderson et al., 1992). Dermal absorption using acetone as vehicle was greatest
in 3-week-old rats (129 pmol; 64% of the administered dose), decreasing to -80
pmol (40%) in 5-, 8- and 10-week-old rats and to 45 pmol (22%) in 36-week-old
rats. The results indicate that 2,3,7,8-TCDD is absorbed to a greater degree
through skin of very young animals and that a significant decrease in potential
for systemic exposure may occur during maturation and again during aging.
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