vvEPA
United States
Environmental Protection
Agency
EPA/690/R-18/002 | July 31, 2018 | FINAL
Provisional Peer-Reviewed Toxicity Values for
Technical Toxaphene
(CASRN 8001-35-2)
Weathered Toxaphene, and Toxaphene Congeners
swerfumd
U.S. EPA Office of Research and Development
National Center for Environmental Assessment, Superfund Health Risk Technical Support Center (Cincinnati, OH)
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FINAL
07-31-2018
Provisional Peer-Reviewed Toxicity Values for
Technical Toxaphene
(CASRN 8001-35-2),
Weathered Toxaphene, and Toxaphene Congeners
Superfund Health Risk Technical Support Center
National Center for Environmental Assessment
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, OH 45268
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AUTHORS, CONTRIBUTORS, AND REVIEWERS
CHEMICAL MANAGER
Scott C. Wesselkamper, PhD
National Center for Environmental Assessment, Cincinnati, OH
CONTRIBUTORS
J. Phillip Kaiser, PhD, DABT
National Center for Environmental Assessment, Cincinnati, OH
Q. Jay Zhao, PhD, MPH, DABT
National Center for Environmental Assessment, Cincinnati, OH
DRAFT DOCUMENT PREPARED BY
SRC, Inc.
7502 Round Pond Road
North Syracuse, NY 13212
PRIMARY INTERNAL REVIEWERS
Elizabeth Owens, PhD
National Center for Environmental Assessment, Cincinnati, OH
Q. Jay Zhao, PhD, MPH, DABT
National Center for Environmental Assessment, Cincinnati, OH
This document was externally peer reviewed under contract to:
Eastern Research Group, Inc.
110 Hartwell Avenue
Lexington, MA 02421-3136
Questions regarding the content of this PPRTV assessment should be directed to the U.S. EPA
Office of Research and Development's National Center for Environmental Assessment,
Superfund Health Risk Technical Support Center (513-569-7300).
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TABLE OF CONTENTS
COMMONLY USED ABBREVIATIONS AND ACRONYMS iv
BACKGROUND 1
DISCLAIMERS 1
QUESTIONS REGARDING PPRTVs 1
INTRODUCTION 2
TECHNICAL TOXAPHENE 2
WEATHERED TOXAPHENE 5
REVIEW OF POTENTIALLY RELEVANT DATA (NONCANCER AND CANCER) 11
HUMAN STUDIES 23
ANIMAL STUDIES 25
Oral Exposures 25
Inhalation Exposures 52
OTHER DATA (SHORT-TERM TESTS, OTHER EXAMINATIONS) 52
Older Studies Identifying the Liver and Kidney as Toxicity Targets of Technical Toxaphene
52
Genotoxicity and Mutagenicity 53
Metabolism/Toxicokinetic Studies for Technical Toxaphene 65
DERIVATION 01 PROVISIONAL VALUES 65
DERIVATION OF ORAL REFERENCE DOSES 66
Derivation of a Subchronic Provisional Reference Dose 66
Derivation of a Chronic Provisional Reference Dose 76
DERIVATION OF INHALATION REFERENCE CONCENTRATIONS 78
Derivation of a Subchronic Provisional Reference Concentration 78
Derivation of a Chronic Provisional Reference Concentration 78
CANCER WEIGHT-OF-EVIDENCE DESCRIPTOR AND POTENCY VALUES 78
APPENDIX A. SCREENING PROVISIONAL VALUES 79
APPENDIX B. DATA TABLES 89
APPENDIX C. BENCHMARK DOSE MODELING RESULTS 108
APPENDIX D. REFERENCES 208
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COMMONLY USED ABBREVIATIONS AND ACRONYMS1
a2u-g
alpha 2u-globulin
MN
micronuclei
ACGIH
American Conference of Governmental
MNPCE
micronucleated polychromatic
Industrial Hygienists
erythrocyte
AIC
Akaike's information criterion
MOA
mode of action
ALD
approximate lethal dosage
MTD
maximum tolerated dose
ALT
alanine aminotransferase
NAG
7V-acetyl-P-D-glucosaminidase
AR
androgen receptor
NCEA
National Center for Environmental
AST
aspartate aminotransferase
Assessment
atm
atmosphere
NCI
National Cancer Institute
ATSDR
Agency for Toxic Substances and
NOAEL
no-observed-adverse-effect level
Disease Registry
NTP
National Toxicology Program
BMD
benchmark dose
NZW
New Zealand White (rabbit breed)
BMDL
benchmark dose lower confidence limit
OCT
ornithine carbamoyl transferase
BMDS
Benchmark Dose Software
ORD
Office of Research and Development
BMR
benchmark response
PBPK
physiologically based pharmacokinetic
BUN
blood urea nitrogen
PCNA
proliferating cell nuclear antigen
BW
body weight
PND
postnatal day
CA
chromosomal aberration
POD
point of departure
CAS
Chemical Abstracts Service
PODadj
duration-adjusted POD
CASRN
Chemical Abstracts Service registry
QSAR
quantitative structure-activity
number
relationship
CBI
covalent binding index
RBC
red blood cell
CHO
Chinese hamster ovary (cell line cells)
RDS
replicative DNA synthesis
CL
confidence limit
RfC
inhalation reference concentration
CNS
central nervous system
RfD
oral reference dose
CPN
chronic progressive nephropathy
RGDR
regional gas dose ratio
CYP450
cytochrome P450
RNA
ribonucleic acid
DAF
dosimetric adjustment factor
SAR
structure activity relationship
DEN
diethylnitrosamine
SCE
sister chromatid exchange
DMSO
dimethylsulfoxide
SD
standard deviation
DNA
deoxyribonucleic acid
SDH
sorbitol dehydrogenase
EPA
Environmental Protection Agency
SE
standard error
ER
estrogen receptor
SGOT
serum glutamic oxaloacetic
FDA
Food and Drug Administration
transaminase, also known as AST
FEVi
forced expiratory volume of 1 second
SGPT
serum glutamic pyruvic transaminase,
GD
gestation day
also known as ALT
GDH
glutamate dehydrogenase
SSD
systemic scleroderma
GGT
y-glutamyl transferase
TCA
trichloroacetic acid
GSH
glutathione
TCE
trichloroethylene
GST
glutathione-S-transferase
TWA
time-weighted average
Hb/g-A
animal blood-gas partition coefficient
UF
uncertainty factor
Hb/g-H
human blood-gas partition coefficient
UFa
interspecies uncertainty factor
HEC
human equivalent concentration
UFc
composite uncertainty factor
HED
human equivalent dose
UFd
database uncertainty factor
i.p.
intraperitoneal
UFh
intraspecies uncertainty factor
IRIS
Integrated Risk Information System
UFl
LOAEL-to-NOAEL uncertainty factor
IVF
in vitro fertilization
UFs
subchronic-to-chronic uncertainty factor
LC50
median lethal concentration
U.S.
United States of America
LD50
median lethal dose
WBC
white blood cell
LOAEL
lowest-observed-adverse-effect level
Abbreviations and acronyms not listed on this page are defined upon first use in the PPRTV document.
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PROVISIONAL PEER-REVIEWED TOXICITY VALUES FOR
TECHNICAL TOXAPHENE (CASRN 8001-35-2),
WEATHERED TOXAPHENE, AND TOXAPHENE CONGENERS
BACKGROUND
A Provisional Peer-Reviewed Toxicity Value (PPRTV) is defined as a toxicity value
derived for use in the Superfund Program. PPRTVs are derived after a review of the relevant
scientific literature using established Agency guidance on human health toxicity value
derivations. All PPRTV assessments receive internal review by at least two National Center for
Environmental Assessment (NCEA) scientists and an independent external peer review by at
least three scientific experts.
The purpose of this document is to provide support for the hazard and dose-response
assessment pertaining to chronic and subchronic exposures to substances of concern, to present
the major conclusions reached in the hazard identification and derivation of the PPRTVs, and to
characterize the overall confidence in these conclusions and toxicity values. It is not intended to
be a comprehensive treatise on the chemical or toxicological nature of this substance.
PPRTV assessments are eligible to be updated on a 5-year cycle to incorporate new data
or methodologies that might impact the toxicity values or characterization of potential for
adverse human-health effects and are revised as appropriate. Questions regarding nomination of
chemicals for update can be sent to the appropriate U.S. Environmental Protection Agency
(EPA) Superfund and Technology Liaison (https://www.epa.gov/research/fact-sheets-regional-
science).
DISCLAIMERS
The PPRTV document provides toxicity values and information about the adverse effects
of the chemical and the evidence on which the value is based, including the strengths and
limitations of the data. All users are advised to review the information provided in this
document to ensure that the PPRTV used is appropriate for the types of exposures and
circumstances at the site in question and the risk management decision that would be supported
by the risk assessment.
Other U.S. EPA programs or external parties who may choose to use PPRTVs are
advised that Superfund resources will not generally be used to respond to challenges, if any, of
PPRTVs used in a context outside of the Superfund program.
This document has been reviewed in accordance with U.S. EPA policy and approved for
publication. Mention of trade names or commercial products does not constitute endorsement or
recommendation for use.
QUESTIONS REGARDING PPRTVS
Questions regarding the content of this PPRTV assessment should be directed to the
U.S. EPA Office of Research and Development's (ORD's) NCEA, Superfund Health Risk
Technical Support Center (513-569-7300).
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INTRODUCTION
This PPRTV assessment details the hazard and dose-response assessment pertaining to
chronic and subchronic exposures to technical toxaphene (CASRN 8001-35-2), weathered
toxaphene, and toxaphene congeners.
TECHNICAL TOXAPHENE
Technical toxaphene, CASRN 8001-35-2, is a yellow, waxy solid with a turpentine odor.
It is a manufactured pesticide consisting of a complex mixture of hundreds of chlorinated
terpenes produced by the chlorination of camphene under ultraviolet (UV) light (ATSI)R. 2014;
de Geus et ai, 1999; Sal eh. 1991). This manufactured mixture, referred to as technical
toxaphene in this document, was used extensively as an insecticide, piscicide, and acaricide
beginning in the mid-1940s, but the U.S. EPA canceled the registration for most uses as a
pesticide or pesticide ingredient in 1982 (ATSDR. 2014; de Geus et ai. 1999; Vetter and Qehme.
1993; Sal eh. 1991). All registered uses of toxaphene mixtures were subsequently canceled in the
United States in 1990 (ATSI)R. 2014; l.acavo et ai. 2004; de Geus et ai. 1999; Sal eh. 1991).
The use of technical toxaphene was also banned in most of Europe during the 1980s (Barbini et
ai. 2007; Alder and Vieth. 1996). Table 1 summarizes the physicochemical properties of
technical toxaphene.
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Table 1. Physicochemical Properties of Technical Toxaphene (CASRN 8001-35-2)
Property (unit)
Value
Physical state
Solid
Boiling point (°C)
NA (dechlorinates at 155°C)fl
Melting point (°C)
65-903
Density (g/cm3 at 25°C)
1.65a
Vapor pressure (mm Hg at 20°C)
6.69 x 10-6 3
pH (unitless)
NV
pKa (unitless)
NV
Solubility in water (mg/L at 20°C)
0.55a
Octanol-water partition coefficient (log Kow)
5.9b
Henry's law constant (atm-m3/mol at 20°C)
6.0 x 10 6 a
Soil adsorption coefficient (log Koc)
5b
Atmospheric OH rate constant (cm3/molecule-sec at 25°C)
2.3 x 10 12 (estimated)13
Atmospheric half-life (d)
4.7 (estimated)13
Relative vapor density (air = 1)
14.3°
Molecular weight (g/mol)
431.8a
Flash point (°C)
135 (closed cup, 60% solution)3;
115 (tag closed cup, 90% solution)3
a AT SDR (2014).
bU.S. EPA (2012c).
°NTP (2014).
NA = not applicable; NV = not available.
Congeners (components) in technical toxaphene include chlorinated bornanes, bornenes,
bornadienes, camphenes, and dihydrocamphenes, each containing 6-10 chlorine atoms (ATSDR,
2014; de Geus et al.. 1999). Figure 1 shows carbon skeleton structures for these congeners. The
approximate relative composition of technical toxaphene has been reported to be
76% chlorobornanes, 18% chlorobornenes, 2% chlorobornadienes, 1% other chlorinated
hydrocarbons, and 3% nonchlorinated hydrocarbons. The actual composition, however, is likely
to have varied depending on manufacturing conditions (ATSDR. 2014; de Geus et al. 1999;
Sal eh. 1991). Table 2 contains names and CASRNs of selected toxaphene congeners reported in
technical toxaphene (or weathered toxaphene residues; see "Weathered Toxaphene" section), as
well as alternative names in the Andrews and Vetter (1995) and Parlar (Burhenne et al. 1993)
identification systems.
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Bornane Bornene Bornadiene
Camphene Dihydrocamphene
Figure 1. Carbon Skeleton Structures for Congeners (with numbered carbon atoms) in
Technical Toxaphene2
2
Congeners typically contain 6-10 chlorine atoms. Sources: ATSDR (2014): de Geus et al. (1999).
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Table 2. Names of Selected Congeners in Technical Toxaphene or Weathered Toxaphene3
Chemical Name {other ID)
CASRN
Parlar No.
Andrews-
Vetter Code
2,2,3-exo,8,9,10(E)-Hexachlorocamphene
NV
p-11
NV
2-exo,3-endo,8,8,9,10(E)-Hexachlorocamphene
NV
p-12
NV
2,2,5,5,9,10,10-Heptachlorobornane
165820-13-3
p-21
B7-499
2-endo,3-exo,5-endo,6-exo,8,8,10,10-Octachlorobornane (T2, Tox8)
142534-71-2
p-26
B8-1413
2,2,5-endo,6-exo,8,9,10-Heptachlorobornane (Toxicant B)
51775-36-1
p-32
B7-515
2,2,5,5,9,9,10,10-Octachlorobornane
165820-15-5
p-38
B8-789
2,2,3-exo,5-endo,6-exo,8,9,10-Octachlorobornane
64618-67-3
p-39
B8-531
2-endo,3-exo,5-endo,6-exo,8,9,10,10-Octachlorobornane
166021-27-8
p-40
B8-1414
2-exo,3-endo,5-exo,8,9,9,10,10-Octachlorobornane (T(Y>)
165820-16-6
p-41
B8-1945
2,2,5-endo,6-exo,8,8,9,10-Octachlorobornane (TC8, Toxicant Al)
58002-19-0
p-42a
B8-806
2,2,5-endo,6-exo,8,9,9,10-Octachlorobornane (TC8, ToxicantA2)
177695-50-0
p-42b
B8-809
2-exo,5,5,8,9,9,10,10-Octachlorobornane (TC7)
165820-17-7
p-44
B8-2229
2-endo,3-exo,5-endo,6-exo,8,8,9,10,10-Nonachlorobornane
{Toxicant Ac, T12, Tox9)
6680-80-8
p-50
B9-1679
2,2,5,5,8,9,10,10-Octachlorobornane
165820-18-8
p-51
B8-786
2,2,5-endo,6-exo,8,8,9,10,10-Nonachlorohornane
64618-71-9
p-56
B9-1046
2,2,5-endo,6-exo,8,9,9,10,10-Nonachlorobornane
155750-49-5
p-59
B9-1049
2,2,5,5,8,9,9,10,10-Nonachlorobornane
154159-06-5
p-62
B9-1025
2-exo,3-endo,5-exo,6-exo,8,8,9,10,10-Nonachlorobornane
182266-92-8
p-63
B9-2206
2,2,5,5,6-exo,8,9,9,10,10-Decachlorobornane
151183-19-6
p-69
B10-1110
2-exo,3-endo,6-exo,8,9,10-Hexachlorobornane (Hx-Sed)
57981-29-0
NV
B6-923
2-endo,3-exo,5-endo,6-exo,8,9,10-Heptachlorobornane (Hp-Sed)
70649-42-2
NA
B7-1001
2-exo,3 -endo,5-exo,8,9,10,10-Heptachlorobornane (TMX-1)
NV
NA
B7-1450
2 -exo, 3 -endo ,5-exo,9,9,10,10 -Heptachlorobornane
NV
NA
B7-1453
2-endo,3-exo,5-endo,6-exo,8,8,9,10-Octachlorobornane
NV
NA
B8-1412
"ATSDR (2014): Bntekevelt et al. (2001): de Gens et al. (1999): Andrews and Vetter (1995): Burhenne et al.
(1993).
NA = not applicable; NV = not available.
WEATHERED TOXAPHENE
Following release of technical toxaphene into the environment, the congeners are
expected to undergo differential transformation, and degradation via abiotic and biotic processes,
resulting in different mixtures of persistent toxaphene congeners, commonly termed weathered
toxaphene [for reviews, see ATS PR (2014); Braekevelt et al. (2001); de Geus et al. (1999);
Geygr et al. (1999); Sal eh (1991)1. Transformation and degradation processes are expected to
include dechlorination and dehydrochlorination. The composition of weathered toxaphene is
expected to vary depending on the environmental media and conditions (e.g., soil, sediment, or
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air; anaerobic vs. aerobic), distance from source, length of time since release, and the varying
biological matrices in which toxaphene congeners may be found [for reviews, see AT SDR
(2014); deGeusetal. (1999); U.S. EPA (2010); U.S. EPA (2005a); U.S. EPA (2005b); Saleh
(1991)1. Biological processes that can produce different weathered toxaphene profiles include
the ways toxaphene is metabolized and excreted, which vary across congeners and species of
organisms. For example, studies of tissue samples of aquatic mammals and fish found that the
relative amounts of certain octachlorinated and nonachlorinated congeners (p-26 [B8-1413], p-50
[B9-1679], p-62 [B9-1025], and to a lesser extent p-40/41 [B8-1414/B8-1945] and p-44
[B8-2229]), were higher than the relative amounts in technical toxaphene PEkici et al. (2008);
Ruppe et al. (2004); Ruppe et al. (2003); Vetter et al. (2001); Angerhofer et al. (1999); de Geus
et al. (1999); Vetter et al. (1999); Saleh (1991); Vetter and Scherer (1999) as cited in Bernardo et
al. (2005)1. In addition, Simon and Manning (2006) showed that the average percentage of three
persistent congeners (PCs) (p-26, p-50, and p-62, dubbed £3PC) jn tissue samples from more
than 10 northern-latitude aquatic species (22.45%, based on ng/g wet weight) was greater than
the average percentage of these three congeners in tissue from 11 aquatic species sampled from a
former U.S. manufacturing plant in Georgia in 1997 (4.47%).
Further support that weathered toxaphene composition varies in different environmental
settings comes from observed differences in the compositional distribution of toxaphene
congeners in toxaphene-contaminated environmental media (e.g., sediments or air), as well as
biological tissues and fluids sampled from different species and regions of the world. Table 3
presents an example of reported compositional differences between a technical toxaphene
standard and weathered toxaphene in environmental media and biological tissues, showing a
shift to hexachlorinated and heptachlorinated congeners in sediments from a Canadian lake,
compared with the proportions of these congeners in technical toxaphene (Braekevelt et al..
2001). The same study found lesser relative amounts of heptachlorinated congeners and greater
amounts of nonachlorinated congeners, compared with those in technical toxaphene, in pelagic
fish and mammals (i.e., lake trout and beluga whales) and a similar, although less pronounced,
pattern in the demersal amphipod Cyclocaris guilelmi (see Table 3). In contrast, tissue samples
from bottom-dwelling fish species from a tidal creek near a former U.S. toxaphene
manufacturing plant showed a clear shift toward hexachlorinated and heptachlorinated congeners
that reflected the composition of sediments, with a predominance of Hx-Sed and Hp-Sed among
the detected congeners (Maruva et al.. 2001b; Maruva et al.. 2001a). Because Hx-Sed and
Hp-Sed appear to be eliminated more rapidly from fish than octachlorinated and nonachlorinated
congeners (Smalling et al.. 2000; Fisk et al.. 1998). Maruva et al. (2001b) postulated that the
predominance of lower chlorinated congeners in bottom-dwelling fish species was due to
continued exposure to weathered toxaphene in sediments enriched in these lower chlorinated
congeners.
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Table 3. Percent Distribution of Chlorine Homologue Groups in Technical Toxaphene,
Environmental Media, and Biological Samples"
Hexachloro
Heptachloro
Octachloro
Nonachloro
Technical toxaphene
1.4
26.6
63.1
8.9
Air (Northwest Territory, Canada)
3.8
41.2
48.3
6.7
Sediments (Lake Winnipeg, Canada)
31.2
39.8
25.6
3.5
Demersal amphipod Cyclocaris guilelmi
(Beaufort Sea, Canada)
1.2
19.2
68.0
11.6
Lake trout (Lake Laberge, Canada)
0.4
11.8
69.3
18.5
Beluga blubber (Nunavut, Canada)
0.4
15.4
67.5
16.7
aBraekevelt et al. (2001).
Information about actual exposure levels and toxicokinetic properties of toxaphene
congeners in mammalian species is inadequate to explain the congener profile observed in
human biological fluids. However, analysis of human fluids shows a congener profile dominated
by octachlorinated congeners (p-26, and to a lesser extent p-40/41 and p-44) and nonachlorinated
congeners (p-50 and p-62). In an analysis of human serum and breast milk samples collected in
five studies, the major detected congeners and their average percentages of total toxaphene
congeners detected were as follows: p-26 (32.8%), p-50 (54.7%), p-62 (6.3%), p-44 (3.5%), and
p-40/41 (2.7%) (Simon and Manning. 2006). More than 90% of the total congeners detected in
these samples was the three major PCs, p-26, p-50, and p-62 (dubbed £3PC).
A summary of available toxicity values for technical toxaphene from U.S. EPA and other
agencies/organizations is provided in Table 4. No toxicity values are available for weathered
toxaphene or individual toxaphene congeners.
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Table 4. Summary of Available Toxicity Values for Technical Toxaphene
(CASRN 8001-35-2)
Source (parameter)3'b
Value (applicability)
Notes
Reference
Noncancer
IRIS
NV
NA
U.S. EPA (2018)
HEAST
NV
NA
U.S. EPA (201 la)
DWSHA (RfD)
4 / 10 4 mg/kg-d
NA
U.S. EPA (2012a)
ATSDR (acute-duration
oral MRL)
0.05 mg/kg-d
Derived from a NOAEL of 5 mg/kg-d
for neurological effects in Beagle dogs
exposed for 13 wk (Cki et al.. 1986).
ATSDR (2014)
ATSDR
(intermediate-duration
oral MRL)
0.002 mg/kg-d
Derived from data on depressed
humoral immunity in female
cynomolgus monkeys.
ATSDR (2014)
IPCS
NV
NA
IPCS (2018):
WHO (2018)
WHO (ADI)
Not established; however,
reservations remain about
the safety of toxaphene in
food.
Could not establish an ADI for a
material that varied in composition
according to the method of
manufacture.
WHO (1969):
WHO (1974):
WHO (1984):
WHO (1990)
CalEPA (PHG)
10 ppb in drinking water
Based on a NOAEL of 0.35 mg/kg-d
in rats exposed for 90 d (Chu et al..
1986s).
CalEPA (2003)
CalEPA (REL)
NV
NA
CalEPA (2016)
CalEPA (Prop 65 List)
NV
Not listed for causing developmental
or reproductive toxicity.
CalEPA (2018a)
OSHA (PEL)
0.5 mg/m3
Skin designation; 8-hr TWA for
general industry, construction, and
shipyard employment.
OSHA (2017b):
OSHA (2017c):
OSHA (2017a)
ACGIH (TLV-TWA)
0.5 mg/m3
Potential for acute CNS effects that
include salivation, nausea, vomiting,
and muscle spasms that may lead to
convulsions; potential for liver
damage. Skin notation based on
absorption of technical toxaphene
through skin of treated rabbits leading
to systemic CNS effects and lethality.
ACGIH (2001):
ACGIH (2017)
ACGIH (TLV-STEL)
1 mg/m3
Potential for acute CNS effects that
include salivation, nausea, vomiting,
and muscle spasms that may lead to
convulsions; potential for liver
damage. Skin notation based on
absorption of technical toxaphene
through skin of treated rabbits leading
to systemic CNS effects and lethality.
ACGIH (2001):
ACGIH (2017)
DOE (PAC)
PAC-1: 1 mg/m3;
PAC-2: 20 mg/m3;
PAC-3: 200 mg/m3
Based on TEELs.
DOE (2015)
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Table 4. Summary of Available Toxicity Values for Technical Toxaphene
(CASRN 8001-35-2)
Source (parameter)3'b
Value (applicability)
Notes
Reference
USAPHC (air-MEG)
1-hr critical: 200 mg/m3;
1-hr marginal: 20 mg/m3;
1-hr negligible: 1 mg/m3;
8-hr negligible: 0.5 mg/m3;
14-d negligible: 0.12 mg/m3;
1-yr negligible: 0.015 mg/m3
1-hr values based on TEELs for
hepatocellular carcinomas and
neoplastic nodules, 8-hr and 14-d
values based on TLVs for CNS
convulsions and liver damage, 1-yr
value based on IRIS.
U.S. APHC (2013)
USAPHC (water-MEG)
1-yr negligible: 0.014 mg/L
Based on MRL for mild anisokaryosis.
U.S. APHC (2013)
USAPHC (soil-MEG)
1-yr negligible: 212 mg/kg
Basis: noncancer.
U.S. APHC (2013)
Cancer
IRIS (WOE)
B2, probable human
carcinogen;
OSF: 1.1 (nig/kg-d)1;
IUR: 3.2 x 10-4 (ng/m3)-1
Both values based on increased
incidence of hepatocellular tumors in
mice and thyroid tumors in rats
following oral exposure; states values
supported by mutagenicity in
Salmonella.
U.S. EPA (1988a)
HEAST
NV
NA
U.S. EPA (2011a)
DWSHA (WOE)
B2, probable human
carcinogen
NA
U.S. EPA (2012a)
NIOSH (REL)
Potential occupational Ca
Skin designation.
NIOSH (2016)
NIOSH (IDLH)
200 mg/m3; potential
occupational Ca
No toxic responses were noted in
25 volunteers exposed to 500 mg/m3
for 30 min/d for 10 consecutive days
IShclanskv CI947) as cited in NIOSH
(2014)1. However, the orieinal IDLH
of 200 mg/m3 is not being revised at
this time.
NIOSH (2014);
NIOSH (2016)
NTP (WOE)
Reasonably anticipated to be
a human carcinogen
Based on sufficient evidence of
carcinogenicity from studies in mice
and rats.
NTP (2014)
I ARC (WOE)
Group 2B, possibly
carcinogenic to humans
Based on sufficient evidence of
carcinogenicity in mice and rats and
inadequate evidence in humans.
I ARC (2001)
CalEPA (OSF)
1.2 (mg/kg-d) 1
PHG of 0.03 ppb in drinking water is
based on the OSF.
CalEPA (2003);
CalEPA (2018b)
CalEPA (ISF)
1.2 (mg/kg-d)"1
NA
CalEPA (2003):
CalEPA (2018b)
CalEPA (IUR)
3.4 x 10-4 (iig/m3)-1
NA
CalEPA (2003):
CalEPA (2018b)
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Table 4. Summary of Available Toxicity Values for Technical Toxaphene
(CASRN 8001-35-2)
Source (parameter)3'b
Value (applicability)
Notes
Reference
CalEPA (Prop 65 List)
NV
Listed as cancer causing.
CalEPA (2018a)
ACGIH (WOE)
A3, confirmed animal
carcinogen with unknown
relevance to humans
Based on induction of hepatocellular
carcinomas in mice and an increased
incidence of thyroid tumors in rats.
ACGIH (2001);
ACGIH (2015)
aSources: ACGIH = American Conference of Governmental Industrial Hygienists; ATSDR = Agency for Toxic
Substances and Disease Registry; CalEPA = California Environmental Protection Agency; DOE = U.S.
Department of Energy; DWSHA = Drinking Water Standards and Health Advisories; HEAST = Health Effects
Assessment Summary Tables; IARC = International Agency for Research on Cancer; IPCS = International
Programme on Chemical Safety; IRIS = Integrated Risk Information System; NIOSH = National Institute for
Occupational Safety and Health; NTP = National Toxicology Program; OSHA = Occupational Safety and Health
Administration; USAPHC = U.S. Army Public Health Command; WHO = World Health Organization.
Parameters: ADI = acceptable daily intake; IDLH = immediately dangerous to life or health concentrations;
ISF = inhalation slope factor; IUR = inhalation unit risk; MEG = military exposure guideline; MRL = minimal risk
level; OSF = oral slope factor; PAC = protective action criteria; PEL = permissible exposure limit; PHG = public
health goal; REL = recommended exposure limit; RfD = reference dose; STEL = short-term exposure limit;
TEEL = temporary emergency exposure limit; TLV = threshold limit value; TWA = time-weighted average;
WOE = weight of evidence.
Ca = carcinogen; CNS = central nervous system; NA = not applicable; NOAEL = no-observed-adverse-effect
level; NV = not available.
Non-date-limited literature searches were conducted in March 2016 and updated in
July 2018 for studies relevant to the derivation of provisional toxicity values for technical
toxaphene (CASRN 8001-35-2), weathered toxaphene, and toxaphene congeners. Searches were
conducted using the U.S. EPA's Health and Environmental Research Online (HERO) database of
scientific literature. HERO searches the following databases: PubMed, ToxLine (including
TSCATS1), and Web of Science. The following databases were searched outside of HERO for
health-related data: American Conference of Governmental Industrial Hygienists (ACGIH),
Agency for Toxic Substances and Disease Registry (ATSDR), California Environmental
Protection Agency (CalEPA), U.S. EPA Integrated Risk Information System (IRIS), U.S. EPA
Health Effects Assessment Summary Tables (HEAST), U.S. EPA Office of Water (OW),
U.S. EPA TSCATS2/TSCATS8e, U.S. EPA High Production Volume (HPV), European Centre
for Ecotoxicology and Toxicology of Chemicals (ECETOC), Japan Existing Chemical Database
(JECDB), European Chemicals Agency (ECHA), Organisation for Economic Co-operation and
Development (OECD) Screening Information Data Sets (SIDS), OECD International Uniform
Chemical Information Database (IUCLID), OECD HPV, National Institute for Occupational
Safety and Health (NIOSH), National Toxicology Program (NTP), Occupational Safety and
Health Administration (OSHA), and Defense Technical Information Center (DTIC).
10
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REVIEW OF POTENTIALLY RELEVANT DATA
(NONCANCER AND CANCER)
Tables 5 A and 5B provide overviews of the relevant noncancer and cancer databases,
respectively, for technical toxaphene (CASRN 8001-35-2). The tables include all potentially
relevant repeated-dose short-term-, subchronic-, and chronic-duration studies, as well as
reproductive and developmental toxicity studies. Principal studies are identified in bold. The
phrase "statistical significance," used throughout the document, indicates ap-value of < 0.05
unless otherwise specified.
11
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Human
Convulsions (presumably from effects on the nervous system) are the most common effect reported in acute poisoning case reports. A limited number of
epidemiological studies have examined possible associations between occupational exposure to TT and noncancer diseases. In U.S. male pesticide applicators,
self-reDorted hypothyroidism was associated with "ever-u.se" of 50 specific insecticides (including TT) (Goldner et al.. 20131 but no statistically sienificantlv elevated
ORs were found for amyotrophic lateral sclerosis and "ever-u.se" of any of the subject pesticides, including TT (kamcl et al.. 2012). Additionally, a statistically
significant exposure-response trend in association with rheumatoid arthritis was observed for lifetime davs of toxaphene use (Mever et al.. 2017).
Animal
1. Oral (mg/kg-d)
Short term
40 M/0 F, S-D, rat, TT in
corn oil by gavage; 0, 100
(then 75) mg/kg-d for 28 d
0, 78
Increased TSH serum
levels; increased
incidences for thyroid
follicular hypertrophy and
diffuse intrafollicular
hyperplasia
NDr
78
Waritz et al. (1996)
(Due to dose adjustments after the
first 3 d of exposure, a TWA dose
of 78 mg/kg-d has been calculated
for this assessment.)
PR
Short term
5 M/5 F, S-D, rat, TT in corn
oil by gavage; 0, 6 mg/kg-d
for 21 d, starting at
5-6 wk-of-age
0,6
No difference in maze
learning and learning
transfer testing
6
NDr
Crowder et al. (1980)
PR
Short term
5 M/0 F, B6C3Fi, mouse,
TT in diet; 0, 10, 40, 80,
160, 320 ppmfor 14 d
0, 1.8, 7.3, 15,
29.6, 60.1
Increased absolute and
relative liver weight
(>10%)
1.8
7.3
Wane et al. (2015)
PR
Short term
36 M/0 F, B6C3Fi, mouse,
TT in diet; 0, 3, 32, 320 ppm
for up to 28 d (sacrifices at
7, 14, and 28 d)
0,0.6, 5.9, 60.3
Increased absolute and
relative liver weight
(>10%), serum ALT
activities, hepatic cell
proliferation rates (BrdU
labeling index), and MDA
concentrations in liver
5.9
60.3
Wane et al. (2015)
PR
12
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Short term
5 M/0 F, C57BL/6, mouse,
TT in diet; 0 or 320 ppm for
14 d
0, 64.5
Decreased body weight;
increased absolute and
relative liver weight
(>10%); increased hepatic
cell proliferation rates
(BrdU labeling index);
liver enzyme activity
changes
NDr
64.5
Wane et al. (2017)
PR
Subchronic
2 M/2 F, cynomolgus,
monkey, TT (corn oil) in
gelatin capsules; 0,
1 mg/kg-d for 52 wk
0, 1
Increased relative spleen
and thymus weights;
decreased mean IgG and
IgM responses to SRBC,
mean percentages of
CD2+CD4+ lymphocytes,
and mean CD4:CD8 ratios
NDr
1
Arnold et al. (2001); Brvce et al.
(2001); Trvohonas et al. (2000)
(The small number of animals
limits confidence in the LOAEL)
PR
Subchronic
5 M/10 F, cynomolgus,
monkey, TT (corn oil) in
gelatin capsules; 0, 0.1, 0.4,
0.8 mg/kg-d for up to 75 wk
M: 0, 0.8
F: 0,0.1,0.4,
0.8
Decreased IgM responses
to SRBC
NDr (M)
0.1 (F)
0.8 (M)
0.4 (F)
Arnold et al. (2001); Trvohonas et
al. (2001)
PR
Subchronic
6 M/6 F, Beagle, dog, TT
(corn oil); 0, 0.2, 2.0,
5.0 mg/kg-d in gelatin
capsules for 13 wk
0, 0.2, 2.0, 4.5
Increased relative liver
weight (>10%)
2.0 (M)
NDr (F)
4.5 (M)
0.2 (F)
Chi et al. (1986)
(Due to dose adjustments
throughout the exposure duration,
an approximate TWA dose of
4.5 mg/kg-d was calculated for the
high-dose group for this
assessment.)
PR
Subchronic
10 M/10 F, S-D, rat, TT in
diet; 0, 4, 20, 100, 500 ppm
for 13 wk
M: 0,0.35, 1.8,
8.6, 45.9
F: 0. 0.50,2.6,
12.6, 63
Increased incidences of
lesions in the thyroid,
kidney, and liver in both
sexes
0.35 (M)
NDr (F)
1.8 (M)
0.50 (F)
Chu et al. (1986)
PR
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Subchronic
12 M/0 F, S-D, rat, TT in
diet; 0, 30, 300 ppm for 9 wk
0,2.6,25.8
Transient
immunosuppression
(decreased KLH IgG titers
at 15 d, but not at 21 d)
and increased relative liver
weight
NDr
2.6
Roller et al. (1983)
PR
Subchronic
0 M/23-26 F,
Swiss-Webster, mouse, TT
in diet; 0, 10, 100, 200 ppm
for 8 wk
0, 1.9, 19.1,39.2
Immune suppression
(decreased BSA IgG
titers). Increased absolute
and relative liver weight
(>10%) and variation in
hepatocyte size
1.9
19.1
Allen etal. (1983)
PR
Chronic
50 M/50 F per exposed
group; 10 M/10 F controls,
Osborne-Mendel, rat, TT in
diet; 0, 556,
1,112 TWA ppm (M);0,
540, 1,080 TWA ppm (F) for
80 wk followed by
28-30 wk on control diet
M: 0, 38.9,
77.88
F: 0,41.6, 83.29
Statistically and
biologically significant
(>10%) decreased body
weight in females and
clinical signs in both sexes
38.9 (M)
41.6 (F)
77.88 (M)
83.29 (F)
NCI (1979)
PR
Chronic
50 M/50 F per exposed
group; 10 M/10 F controls,
B6C3Fi, mouse, TT in diet;
0, 99, 198 TWA ppm for
80 wk followed by
10-11 wk on control diet
M: 0, 17, 34.0
F: 0, 17, 34.2
No effects at the low dose
in either sex.
17
NDr
NCI (1979)
(Increased late mortality in
high-dose males and females may
have been secondary to high
incidence of hepatocellular
carcinomas in these groups.)
PR
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Reproductive
3 M/6 F, S-D, rat, TT in
diet; 0,4,20,100,500 ppm
for up to 29 wk; F0 dams
and sires exposed for 13 wk
prior to mating and
throughout mating,
gestation, and lactation
(~29 wk). Fla pups were
exposed to same diet as
parents at weaning, culled
to 15 M and 30 F at 5 wk,
and maintained on same
diet for an additional
26 wk. F0 rats were
maintained on diets after
Fla weaning and
production of Fib litters
F0 M: 0,0.36,
1.8,9.2,45
F0 F: 0,0.36,
1.9, 8.5,46
Fla M: 0,0.29,
1.4, 7.5,37
Fla F: 0,0.38,
1.9,9.4,49
Reproductive: No effects
on F0 fertility and
gestation indices, litter
sizes, or pup survival.
Systemic: Increased
absolute liver weight
(>10%) in F0 females;
increases in lesions in the
thyroid, kidney, and
liver, in the F0 and
Fla adults and Fib pups.
45 (F0 M)
46 (F0 F)
NDr (Fla M)
NDr (F0 F)
NDr (F0 M)
NDr (F0 F)
0.29 (Fla M)
0.36 (F0 F)
CM et al. (1988)
PR, PS
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Reproductive
8 M/16 F, S-D, rat, TT in
diet; 0, 25, 100 ppm for up to
42 wk; F0 dams and sires
exposed from
28-100 d-of-age, when
mating occurred, continuing
for the dams through
gestation, lactation, and
production of two litters.
This procedure was
continued through three
successive two-litter
generations until F3b pups
were born. F0 dams were
sacrificed after 42 wk of
exposure; Fib and
F2b parents were sacrificed
after 39 wk of exposure
M: 0, 1.7, 6.88
F: 0,2.0,7.99
Reproductive: No effects
on F0 fertility and
pregnancy indices, mean
litter size, live birth index,
number of pups at birth
through lactation, or
growth of pups through
lactation
Systemic: Increased
relative liver weight
(>10%)
6.88 (F0 M)
7.99 (F0 F)
NDr(M)
NDr(F)
NDr (F0 M)
NDr (F0 F)
1.7 (F1 M)
2.0 (F0 F)
Keimedv et al. (1973)
(NOAEL/LOAEL determinations
are of low confidence due to
reporting deficiencies in the
available report and uncertainties
regarding the reliability of studies
conducted by Industrial Bio-Test
Laboratories. Reporting of results
is inadequate to conduct BMD
analysis on endpoints evaluated.)
PR
16
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Reproductive
4 M/14 F, Swiss-white,
mouse, TT in diet; 0, 25 ppm
through 5 generations;
exposure to parental dams
and sires before mating and
continuing through
production of two litters
until F5b litters were
delivered. The report did not
clearly specify F0 exposure
duration before mating, but
specified that parental
animals in all generations
were sacrificed at
120 d-of-age
M: 0, 4.7
F: 0, 5.1
Parental reproductive: No
effects on fertility, pup
viability, or pup survival
indices
4.7 (M)
5.1 (F)
NDr
NDr
Keolmeer et al. (1970)
(Although histology, serum
chemistry, and hematology
endpoints were reported to be
evaluated, reporting of results was
inadequate for NOAEL/LOAEL
determination for parental systemic
effects from TT.)
PR
Developmental
16-39 pregnant F, CD, rat,
TT in corn oil by gavage; 0,
15, 25, 35 mg/kg-d on
GDs 7-16; dams sacrificed
on GD 21 and fetuses
examined for ossification
centers
0, 15,25,35
Maternal: Decreased
weight gain
Developmental: Decreased
number of sternal, but not
caudal, ossification centers
in fetuses; biologically
significant (>5%)
decreased fetal body
weight
NDr
NDr
15
15
Oiemoff and Carver (1976)
PR
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Developmental
26-90 pregnant F, CD-I,
mouse, TT in corn oil by
gavage; 0, 15, 25,
35 mg/kg-d on GDs 7-16;
dams were sacrificed on
GD 18 and fetuses examined
for ossification centers
0, 15,25,35
Maternal: Increased
relative liver weight
Developmental: No
statistically significant
changes observed
NDr
35
15
NDr
Chemoff and Carver (1976)
PR
Developmental
25 pregnant F, CD-I, mouse,
TT in corn oil by gavage; 0,
75 mg/kg-d on GDs 8-12;
dams gave birth and litters
were counted and weighed
on PNDs 1 and 3
0, 75
Maternal: Decreased
body-weight gain
Developmental: Decreased
pup body weight on
PND 1, but not PND 3
NDr
NDr
75
75
CJiemoff and Kavlock (1983)
(Comprehensive examination of
fetuses for visceral or skeletal
variations or anomalies was not
conducted.)
PR
Developmental
~25 exposed pregnant F,
~50 control pregnant F, S-D,
rat, TT in corn oil by gavage;
0, 32 mg/kg-d on GDs 6-15;
sacrifice on GD 20
0, 32
Maternal: Increased
mortality and decreased
body-weight gain
Developmental: Increased
mean proportion of fetuses
with supernumerary ribs
NDr
NDr
32 (FEL)
32
Chemoff etal. (1990)
PR
Developmental
3 pregnant F, S-D, rat, TT in
corn oil by gavage; 0,
6 mg/kg-d on GDs 7-21;
righting, grasp-hold, and
startle reflexes assessed in
all offspring, starting at
PND 7; 5 M/5 F offspring
evaluated in maze tests,
PNWs 14-16
0,6
Developmental: Delayed
attainment of righting
reflex ability in offspring
NDr
6
Crowder et al. (1980)
(LOAEL is considered potentially
minimal; righting reflex was
delayed, but not grasp-hold or
startle reflexes.)
PR
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category"
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
Developmental
3 pregnant F, Holtzman
albino, rat; TT, Toxicant A
(p-42), or Toxicant B (p-32)
in diet; 0, 0.050 (TT), 0.002
(p-42), 0.002 (p-32) mg/kg-d
from GD 5 through weaning
and PND 30; offspring
received same diet as dams
through PND 90; due to
insufficient p-32 test
substance, p-32-exposed
offspring received p-42 from
PNDs 40-90
0, 0.050 (TT)
0.002 (p-42)
0.002 (p-32)
Developmental: No clear
effects in offspring
neurobehavioral tests
(righting reflex and
swimming ability at
PNDs 7-17 [all offspring];
maze testing of
motivational behavior,
learning, and learning
retention between
PNDs 70-90
[n = 15-16 per group])
NDr (TT)
0.002 (p-42)
0.002 (p-32)
NDr (TT)
NDr (p-42)
NDr (p-32)
Olson etal. (1980)
(Delays in righting reflex were
noted in TT-exposed rats [but not
in p-42 or p-32 groups]; however,
the magnitude of effect was not
reported, precluding the
determination of an effect level.)
PR
Developmental
0 M/12 F, Swiss-Webster,
mouse, TT in diet; 0, 10,
100, 200 ppm for 3 wk
before mating and
throughout gestation and
lactation (~9 wk)
0, 1.9, 19.1,39.2
Immune suppression
(decreased ability of
macrophages to engulf
SRBCs) in offspring
NDr
1.9
Allen etal. (1983)
(Offspring received control diet at
weaning and were aged 8 wk prior
to immune testing.)
PR
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Table 5A. Summary of Potentially Relevant Noncancer Data for Technical Toxaphene (CASRN 8001-35-2)
Number of Male/Female,
Strain, Species, Study
Type, Reported Doses,
Category"
Study Duration
Dosimetryb
Critical Effects
NOAELb
LOAELb
Reference (comments)
Notes0
2. Inhalation (mg/m3)
ND
aDuration categories are defined as follows: Acute = exposure for <24 hours; short term = repeated exposure for 24 hours to <30 days; long term (subchronic) = repeated
exposure for >30 days <10% lifespan for humans (>30 days up to approximately 90 days in typically used laboratory animal species); and chronic = repeated exposure
for >10% lifespan for humans (>~90 days-2 years in typically used laboratory animal species) (U.S. EPA. 20021.
bDosimetry: Values are presented as ADDs (mg/kg-day) for oral noncancer effects. In contrast to other repeated exposure studies, values from animal gestational
exposure studies are not adjusted for exposure duration in calculation of the ADD or HEC.
°Notes: PR = peer reviewed; PS = principal study.
ADD = adjusted daily dose; ALT = alanine aminotransferase; BMD = benchmark dose; BrdU = bromodeoxyuridine; BSA = bovine serum albumin; F = female(s);
FEL = frank effect level; GD = gestation day; HEC = human equivalent concentration; IgG = immunoglobulin G; IgM = immunoglobulin M; KLH = keyhole limpet
hemocyanin; LOAEL = lowest-observed-adverse-effect level; M = male(s); MDA = malondialdehyde; ND = no data; NDr = not determined;
NOAEL = no-observed-adverse-effect level; OR = odds ratio; PND = postnatal day; PNW = postnatal week; S-D = Sprague-Dawley; SRBC = sheep red blood cell;
TSH = thyroid stimulating hormone; TT = technical toxaphene; TWA = time-weighted average.
20
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Table 5B. Summary of Potentially Relevant Cancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category
Number of Male/Female, Strain, Species,
Study Type, Reported Doses, Study Duration
Dosimetry"
Critical Effects
Reference (comments)
Notesb
Human
Several epidemiological studies of agricultural workers have exhibited some statistically significant associations between self-reported past occupational exposure to TT
or regional indicators of agricultural use of toxaphene and increased risk for several tvpcs of cancer as follows: rectal cancer (Lee et al.. 2007; Purdue et al.. 20071
melanoma (Purdue et al.. 20071 leukemia or non-Hodgkin's lvmphoma (Mills et al.. 2005; Schroeder et al.. 2001). and breast cancer (Mills and Yang. 2006). In soouses
of the male pesticide applicators in the Lee et al. (2007) and Purdue et al. (2007) studies, glioma was significantly associated with "ever-use" of anv of the subject
insecticides (including TT) (Louis et al.. 2017). Additionally, based on the observation that telomere length in surrogate tissues is associated with some cancer tvpcs in
other epidemiological studies, mean relative telomere length in buccal DNA decreased significantly with toxaphene use in male pesticide applicators (Hou et al.. 2013).
No significant association between exposure to toxaphene and the occurrence of non-Hodgkin's lvmphoma was reported in several studies (Louis et al.. 2017; Purdue et
al.. 2007; Mills et al.. 2005; De Roos et al.. 2003; Cantor et al.. 1992). In contrast to the Mills et al. (2005) study, the studv bv Purdue and colleagues did not see a
significant association between exposure to toxaphene and leukemia (Purdue et al.. 2007).
Animal
1. Oral (mg/kg-d)
Carcinogenicity
54 M/54 F, B6C3Fi, mouse, TT in diet; 0, 7, 20,
or 50 ppm for 18 mo followed by a 6-mo
observation period
0,0.91,2.6,6.5
Statistically significantly increased
incidence of combined hepatocellular
adenomas and carcinomas in male mice at
6.5 mg/kg-d. No significantly elevated
tumor incidences in exposed-female
groups
Litton Bionetics (1978)
(The IRIS OSF of
1.1 [mg/kg-d]"1 is based
on the liver tumor data in
male mice.)
U.S. EPA
(1988a);
NPR
Carcinogenicity
50 M/50 F, B6C3Fi, mouse, TT in diet; 99,
198 ppm (TWA) for 80 wk (10 M/10 F in
matched 0-ppm control group)
M: 0, 17, 34.0
F: 0, 17,34.2
Statistically significantly increased
incidences of combined hepatocellular
carcinomas and adenomas in
exposed-male and female groups
NCI (1979)
PR
Carcinogenicity
50 M/50 F, Osborne-Mendel, rat, TT in diet;
556, 1,112 ppm (TWA) (M); 540, 1,080 ppm
(TWA) (F) for 80 wk (10 M/10 F in matched
0-ppm control group)
M: 0, 38.9, 77.88
F: 0,41.6, 83.29
Statistically significantly increased
incidences of thyroid tumors in high-dose
males and females
NCI (1979)
PR
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Table 5B. Summary of Potentially Relevant Cancer Data for Technical Toxaphene (CASRN 8001-35-2)
Category
Number of Male/Female, Strain, Species,
Study Type, Reported Doses, Study Duration
Dosimetry"
Critical Effects
Reference (comments)
Notesb
2. Inhalation (mg/m3)
Human
ND
Animal
ND
aDosimetry: The units for oral exposures are expressed as ADDs (mg/kg-day).
bNotes: PR = peer reviewed; NPR = not peer reviewed.
ADD = adjusted daily dose; DNA = deoxyribonucleic acid; F = female(s); IRIS = Integrated Risk Information System; M = male(s); ND = no data; OSF = oral slope
factor; TT = technical toxaphene; TWA = time-weighted average.
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HUMAN STUDIES
Technical Toxaphene
Case reports of humans intentionally or accidentally ingesting products containing
technical toxaphene reported convulsions as the principal and common effect in both fatal and
nonfatal poisoning events (Wells and Milhorn. 1983; McGee et aL 1952). For some of these
cases, other reported effects included changes in liver and kidney function, cardiac dilatation,
swelled kidneys, and temporary memory loss.
Two reports of acute or short-term exposure of human subjects to airborne technical
toxaphene were identified in the literature search. No changes in blood cell profiles, urinalysis,
or gross appearance of the skin were found in a group of 25 human subjects (15 males,
10 females) following 10 daily 30-minute exposures to an aerosol containing a maximum of
500 mg technical toxaphene/m3 or following three additional 30-minute exposures after a 3-week
nonexposure period (Keplinger. 1963). Acute pulmonary insufficiency and changes in chest
x-rays were noted in two Egyptian agricultural workers who applied a spray containing
60% technical toxaphene, 35% kerosene, 3% xylol, and 2% emulsifier during a 2-month period
in 1958 (Warraki. 1963). Treatment with cortisone, streptomycin, and isoniazid produced rapid
recovery in both subjects.
Several epidemiological studies of pesticide applicators or agricultural workers have
reported some statistically significant associations between self-reported past occupational
exposure to technical toxaphene, or regional indicators of agricultural use of toxaphene, with
increased risk for several types of cancer. These positive associations included rectal cancer
(Lee et aL 2007; Purdue et aL 2007). melanoma (Purdue et aL 2007). leukemia or
non-Hodgkin's lymphoma (Mills et aL 2005; Schroeder et aL 2001). and breast cancer (Mills
and Yang. 2006). Louis et al. (2017) evaluated possible associations between various types of
cancer and the use of seven specific organochlorine insecticides (including technical toxaphene)
in female spouses of male pesticide applicators from the Lee et al. (2007) and Purdue et al.
(2007) studies. A statistically significant association was observed between "ever-use" of any of
the subject organochlorine insecticides and increased glioma risk. Although statistically
significant associations were observed between specific organochlorine insecticide (e.g., lindane)
use and increased risk of cancer, no association was observed specifically for toxaphene use.
No significant association between exposure to toxaphene and the occurrence of
non-Hodgkin's lymphoma was reported in several studies (Louis et al.. 2017; Purdue et al.. 2007;
Mills et al.. 2005; De Rous et al.. 2003; Cantor et al.. 1992). In contrast to the Mills et al. (2005)
study, the study by Purdue and colleagues did not see a significant association between exposure
to toxaphene and leukemia (Purdue et al.. 2007).
Goldner et al. (2013) evaluated possible associations between self-reported thyroid
disease and use of 50 specific insecticides (including technical toxaphene), herbicides, and
fungicides within a cohort of U.S. male pesticide applicators. Hypothyroidism was associated
with "ever-use" of eight insecticides, including technical toxaphene. Additionally, results from
exposure-response analyses using the intensity-weighted exposure measure showed technical
toxaphene had an association at low exposure, but not high exposure.
Hon et al. (2013) evaluated possible associations between telomere length in buccal cell
deoxyribonucleic acid (DNA) samples and the use of 48 specific pesticides (including technical
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toxaphene) within the Agricultural Health Study cohort of U.S. male pesticide applicators. Mean
relative telomere length (measured with real-time polymerase chain reaction) decreased
significantly with two metrics of pesticide use (lifetime days of use, and lifetime
intensity-weighted days of use) for technical toxaphene and six other pesticides. No significant
associations were found for relative telomere lengthening and use of any of the pesticides. The
rationale for examining telomere length in DNA from buccal cells was based on the observation
from other case-control studies that telomere length in surrogate tissues (e.g., blood or buccal
cells) is associated with some, but not all, cancer types [see Hou et al. (2013) for a list of
references]. Some studies found associations between cancer and shorter telomeres, and others
found associations with longer telomeres (Hou et al.. 2013).
In another case-control analysis of data collected within the U.S. Agricultural Health
Study of male pesticide applicators, no statistically significantly elevated odds ratios (ORs) were
found for amyotrophic lateral sclerosis and "ever-use" of any of the subject pesticides, including
toxaphene (Kamel et al.. 2012). The OR for toxaphene (adjusted for age and sex) was elevated,
but not to a statistically significant degree (OR = 2.0, 95% confidence interval [CI] = 0.8-4.9;
7 cases, 6,937 controls).
In a recent case-control analysis of data collected within the U.S. Agricultural Health
Study of male pesticide applicators, a statistically significant exposure-response trend in
association with rheumatoid arthritis was observed for the top tertile of lifetime days of
toxaphene use (Mever et al.. 2017). The OR for rheumatoid arthritis and "ever-use" of
toxaphene (adjusted for age, state of enrollment, pack-years smoking, and education) was
elevated, but not statistically significantly (OR = 1.44, 95% CI = 0.90-2.14; 23 cases,
1,487 controls).
A hospital-based prospective cohort study was designed to evaluate possible associations
between levels of 29 persistent organochlorine pesticides in blood samples of pregnant women
and health outcomes of offspring during infancy and early childhood. In a report of preliminary
findings, concentrations of a considerable number of these chemicals (including the toxaphene
congeners p-26 and p-50) showed significant increases associated with increasing age of the
subject. Additionally, decreases in congener concentrations were associated with with increasing
number of pregnancies experienced (gravidity) or carried to a viable gestation age (parity)
(Kanazawa et al.. 2012). Health outcome data for offspring from this study has not been
identified.
Weathered Toxaphene and Toxaphene Congeners
Studies evaluating possible associations between health effects in humans and exposure
to weathered toxaphene or individual toxaphene congeners have not been identified.
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ANIMAL STUDIES
Oral Exposures
Short-Term-Duration Studies
Technical Toxaphene
Waritz et al. (1996)
Groups of 40 male Sprague-Dawley (S-D) rats (8 weeks old) were given 100 mg/kg-day
technical toxaphene in corn oil by gavage for 3 days, followed by a lower dose, 75 mg/kg-day,
for 25 days, due to deaths of two rats on Day 4. A time-weighted average (TWA) dose of
78 mg/kg-day has been calculated for this assessment. A group of 40 control rats received the
corn oil vehicle for 28 days. Technical toxaphene (purity unknown) was mixed with corn oil and
administered once daily by intragastric gavage. Feed consumption and body weight were
measured at study initiation (Day 0) and weekly thereafter. Ten animals per group were
sacrificed on exposure Days 1, 8, and 15, and 1 day following the last administered dose
(Day 29). Blood was immediately collected from the aorta. Levels of serum thyroid stimulating
hormone (TSH), triiodothyronine (T3), thyroxine (T4), reverse triiodothyronine (rT3), and
corrected triiodothyronine (CrT3) were determined by radioimmunoassay (RIA). At necropsy,
the thyroids, parathyroid glands, pituitary glands, and brains were grossly examined, fixed in
formalin, and weighed. Slices of thyroids including the parathyroids and slices of pituitary
glands were prepared for sectioning and stained with hematoxylin/eosin for microscopic
examination. Colloid levels in thyroid follicles were determined by follicle cell sizes as follows:
(1) small, nondistended follicles were taken as indicators of no or little colloid storage, typically
found in young rats; and (2) large, >60% distended follicles were taken as indicators of large
stores of colloid, typically found in older rats.
No differences in mean thyroid or brain weights, or in thyroid:brain-weight ratios
between exposed and control groups were observed. Other organ-weight results were not
described. TSH levels significantly increased with exposure time (39, 141, and 192% increases
over controls after 7, 14, and 28 days of exposure, respectively). Mean differences between
exposed and control values for levels of T3, T4, rT3, and CrT3 were reported as not statistically
significant at any exposure interval. Histological features of pituitary sections were reported to
be similar between control and exposed groups. Thyroid follicular hypertrophy (indicated by
columnar thyroid follicular epithelial cells) was not found in control rats or exposed rats
sacrificed on Day 1, but occurred in 70% of exposed rats sacrificed on Days 8 and 15, and 100%
of exposed rats on Day 29. Diffuse intrafollicular hyperplasia also was found in thyroids of 10,
90, and 80% of exposed rats on Days 8, 15, and 21, but was not found in control rats. The
percentage of exposed rats with small follicle sizes (indicative of low stores of colloid) increased
with exposure duration: 0% on Day 1; 20% on Day 8; 30% on Day 15; and 50% on Day 29.
The results of this study indicate that 78 mg/kg-day, the only dose tested, is a
lowest-observed-adverse-effect level (LOAEL) for increased TSH levels in serum and increased
incidences of thyroid follicular hypertrophy and diffuse intrafollicular hyperplasia in male S-D
rats given gavage doses of technical toxaphene for 28 days. Waritz et al. (1996) postulated that
the study results, along with other observations showing that technical toxaphene induces
cytochrome P450 (CYP450) enzymes in the liver (including UDP-glucuronyl transferases), are
consistent with the hypothesis that technical toxaphene perturbs thyroid homeostasis in rats
(i.e., induces thyroid hyperactivity) via stimulation of the synthesis of UDP-glucuronyl
transferases.
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Crowder et al. (1980)
Groups of 5- to 6-week-old S-D rats (five/sex/group) were administered technical
toxaphene (Boots-Hercules, Inc.; purity 99.9%) at doses of 0 or 6 mg/kg-day, once daily by
gavage (in 0.1 mL corn oil) for 21 days. Animals were given free access to food and water.
Body-weight measurements, feed consumption, or clinical observations were not reported by the
study authors. Approximately 6-8 weeks after the exposure ended and between
14-16 weeks-of-age, five male and five female rats/group were chosen randomly for evaluation
of learning and learning transfer abilities using maze testing. Each rat received 10 maze
trials/day.
No statistically significant differences in maze learning or learning transfer between the
exposed and control groups were described. The study authors concluded that exposure to
6 mg/kg-day technical toxaphene did not interfere with the learning ability of adult rats.
The study results indicate a no-observed-adverse-effect level (NOAEL) of 6 mg/kg-day
for the lack of prolonged effects on learning and learning transfer abilities in maze testing of
adult male and female S-D rats exposed to technical toxaphene via gavage for 21 days, starting at
5-6 weeks-of-age. No LOAEL could be determined.
Wang et al. (2015)
In an initial 14-day range-finding study, B6C3Fi mice (five males/group) were exposed
to technical toxaphene in the diet at concentrations of 0, 10, 40, 80, 160, and 320 ppm for
14 days. Based on group averages between reported means for initial and final body weights and
and an allometric equation for food consumption (food consumption in kg/day = 0.056 x [body
weight in kg06611]) described by U.S. EPA (1988b). estimated daily intakes of 1.8, 7.3, 15, 29.6,
and 60.1 mg/kg-day were calculated. In a follow-up 28-day mechanistic study, B6C3Fi mice
(36 males/exposure group) were exposed to 0, 3, 32, and 320 ppm for up to 28 days (sacrifices
occurred at 7, 14, and 28 days). Based on group averages between reported means for initial and
final body weights at 28 days and an allometric equation for food consumption (food
consumption in kg/day = 0.056 x [body weight in kg0 6611]) described in U.S. EPA (1988b),
estimated daily intakes of 0.6, 5.9, and 60.3 mg/kg-day were calculated. In a final knockout
study, a total of 30 male mice (15 wild-type C57BL/6 and 15 constitutive androstane receptor
knockout [CAR] mice) were exposed to 0 or 320 ppm for 14 days. Because body-weight or
food-consumption data for the wild-type C57BL/6 or CARknockout mice were not reported,
an estimated daily intake of 60.4 mg/kg-day was calculated for the exposed groups based on the
mean reference body weights and food-consumption rates for BAFi and B6C3Fi mice in a
subchronic-duration study.3 Groups of male mice exposed to 750 ppm phenobarbital (PB) in
food were included as a positive control. Food and water were available ad libitum. Seven days
prior to sacrifice, all of the range-finding and knockout study animals and five randomly selected
mice/group from the mechanistic study were given bromodeoxyuridine (BrdU, 0.8 mg/mL in
drinking water with 1% glucose) to measure hepatic DNA synthesis to provide an indicator of
cell proliferation. Body weights of all animals were recorded weekly, although only initial and
terminal weights were provided in the report. For the range-finding and mechanistic studies,
liver weight and weights of other major organs were recorded. Blood was collected to determine
3Dose estimates were calculated using the mean reference body weight and food-consumption rate values for male
BAFi and B6C3Fi mice in a subchronic-duration study (U.S. EPA. 1988b). Mean reference body
weight = 0.0270 kg, and mean reference food-consumption rate = 0.0051 kg/day.
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serum activities of alanine aminotransferase (ALT) and aspartate aminotransferase (AST).
Survival data were obtained and necropsies were performed on all animals from each study.
Portions of the liver and gut (in the range-finding and mechanistic studies) were fixed for
histopathological analysis and for immunohistochemistry staining for anti-BrdU. Liver tissues
from sacrificed animals in the mechanistic study and the constitutive androstane receptor (CAR)
knockout study were also analyzed for concentrations of total protein, acyl-CoA oxidase (ACO)
activity, 8-isoprostane, and malondialdehyde (MDA) (to test for lipid peroxidation), as well as
8-hydroxydeoxyguanasine (8-OHdG) (to test for oxidative DNA damage). Quantitative
real-time polymerase chain reaction (PCR) was used for gene expression analysis of CAR, aryl
hydrocarbon receptor (AhR), peroxisome proliferator-activated receptor alpha (PPAR-alpha)
target genes, cell growth genes, and oxidative stress-related genes in the liver.
No mortalities or clinical signs of toxicity were recorded in any of the control or exposed
groups in any of the studies. In the 14-day range-finding and 28-day mechanistic studies, mean
terminal body weights were statistically significantly decreased only in the high-dose groups by
about 8 and 6% respectively, compared with control groups. In the 14-day study, the mean
initial body weight within the 60.1-mg/kg-day group was higher than the terminal body weight
(28.79 g vs. 27.57 g), indicating a significant decrease in body-weight gain during exposure. In
the 28-day study, control mean body-weight values increased by about 8% during the study
period, but the increase during exposure was only about 0.5% in the 60.3-mg/kg-day group. In
the CAR knockout 14-day study, body weights and body-weight gain were reported to be not
significantly different between exposed (60.4 mg/kg-day) groups and untreated controls. In the
14-day range-finding study, absolute and relative liver weights were increased by >10%
(compared with controls) at concentrations >7.3 mg/kg-day (see Table B-l). In the 28-day
mechanistic study, absolute and relative liver weights were increased by >10% (compared with
controls) in the 60.3-mg/kg-day group only (see Table B-2), with similar magnitudes of increase
observed across all exposure durations. After a 14-day exposure of wild-type (WT) C57BL/6
mice to 60.4 mg/kg-day, increased absolute and relative liver weights were seen (27 and
57%) increase compared with WT controls; see Table B-l), but the liver weight responses were
diminished in CAR knockout mice (-5 and +7%, respectively, compared with CAR
controls). In the 14- and 28-day studies of B6C3Fi mice, serum ALT activities were statistically
significantly increased only in the highest dose groups (the increases ranged from 68-98%),
see Tables B-l and B-2). Histopathological examinations were reported to be similar between
the exposed and control groups in the 28-day study; no necrosis or compensatory hyperplasia
was reportedly observed (no incidence data were reported).
Statistically significant increases in hepatic DNA synthesis (BrdU labeling index),
compared with controls, were observed at concentrations of 29.6 mg/kg-day (~8-fold) and
60.1 mg/kg-day (~14-fold) in the 14-day study (see Table B-l), and at 60.3 mg/kg-day (~3-fold)
in the 28-day study (see Table B-2). Statistically significant increases in hepatic DNA synthesis
also were observed in 60.4-mg/kg-day C57BL/6 mice (>10-fold increase compared with control)
exposed for 14 days, but no increase was observed in exposed CAR 1 knockout mice. Liver
levels of PPAR-alpha and 8-OHdG were comparable to controls in B6C3Fi mice at 7, 14, and
28 days, but MDA concentrations were elevated in 60.3-mg/kg-day animals at 7, 14, and 28 days
(27-35%o increase compared with controls) (see Table B-2). In 14-day exposed wild-type
C57BL/6 mice, hepatic MDA concentrations were elevated by 22%; no MDA changes were
observed in exposed CAR 7 knockout mice.
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Results for several endpoints in the positive control (PB) B6C3Fi group were not
significantly different from the negative control (i.e., body weight, serum ALT activities, and
hepatic MDA concentrations) at any time point evaluated, but statistically significant increased
relative liver-weight values were found at 7, 14, and 28 days, and increased hepatic BrdU
labeling index values were found at Days 7 and 14, but not at 28 days.
In B6C3Fi mice, statistically significant changes in expression of genes in liver tissue
(>2-fold change) were primarily limited to the highest-dose group at 7, 14, and 28 days and
included increased expression of the CAR responsive genes Cyp3all and Cyp2bl0 (up to a
1,973-fold increase), a decrease in the PPAR-alpha related geneAcotl (occurring at 7 and
14 days at >5.9 mg/kg-day, with no other PPAR-alpha related gene changes), increased
expression of the cell growth-related gene, c-myc, suppression of p21 expression, and increased
expression of the oxidative-stress related gene, Aoxl. In 60.4-mg/kg-day wild-type C57BL/6
mice exposed for 14 days, increased hepatic expression of the following genes was found:
Cyp3all, Cyp2b9, CyplblO, c-myc, and Aoxl. These gene expression changes were not
observed in the exposed CAR 1 mice.
Results from the 14-day range-finding study indicate a LOAEL of 7.3 mg/kg-day and a
NOAEL of 1.8 mg/kg-day for increased absolute and relative liver weight in male B6C3Fi mice.
At 29.6 mg/kg-day, significantly increased hepatic cell proliferation rates (BrdU labeling
indices) were observed, and at 60.1 mg/kg-day, significantly increased serum ALT activity was
also observed. The 28-day study identified a LOAEL of 60.3 mg/kg-day and a NOAEL of
5.9 mg/kg-day for increases in absolute and relative liver weight, serum ALT activities, hepatic
cell proliferation rates, and hepatic MDA concentrations in male B6C3Fi mice. Histological
examination of livers from the exposed groups reportedly revealed no evidence for necrosis or
compensatory hyperplasia. Changed expression of examined genes was only observed in livers
of high-dose B6C3Fi mice (increased expression for Cyp3all, Cyp2bl0, c-myc, and Aoxl and
decreased expression of p2T). In the 14-day CAR knockout study, exposure of technical
toxaphene to wild-type C57BL/6 male mice to 60.4 mg/kg-day increased relative liver weight,
hepatic cell proliferation rates, and hepatic MDA concentrations, but these changes were absent
in CAR 1 knockout mice exposed to 60.4 mg/kg-day. The study authors submit that taken
together, the results implicate a nongenotoxic, CAR-mediated mode of action (MOA). However,
the data do not unequivocally prove this to be the primary MOA.
Wang ft al. (2017)
Jfi mimmmmmmmmSm
In a follow-up knockout study by Wang and colleagues, a total of 50 male mice
(15 wild-type C57BL/6 and CAR 7 , and 10 pregnane X receptor knockout mice [PXR ] and
PXR/CAR double knockout mice [PXR /CAR ]; 5 mice/group) were exposed to 0 or
320 ppm technical toxaphene in the diet for 14 days. Due to increased mortality (3/5 mice)
observed in PXR /CARmice in initial studies, the dose was lowered to 160 ppm for
subsequent studies in the double knockout mice. Based on group averages between reported
means for initial and final body weights and an allometric equation for food consumption (food
consumption in kg/day = 0.056 x [body weight in kg0 6611]) described by U.S. EPA (1988b).
estimated daily intakes were calculated as follows: 64.5 mg/kg-day for C57BL/6 mice,
63.7 mg/kg-day for CAR mice, 64.6 mg/kg-day for PXR mice, and 32.6 mg/kg-day for
PXR /CAR~h mice. Groups of male mice exposed to 750 ppm PB in food were included as a
positive control. Food and water were available ad libitum. Seven days prior to sacrifice, mice
were given BrdU (0.8 mg/mL in drinking water with 1% glucose) to measure hepatic DNA
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synthesis to provide an indicator of cell proliferation. Body weights of all animals were recorded
weekly, although only initial and terminal weights were provided in the report. Survival data
were obtained and necropsies were performed on all animals from each study. The liver was
removed and weighed, portions of the liver were fixed for histopathological analysis and for
immunohistochemistry staining for anti-BrdU, and the remainder was snap frozen for enzyme
activity and gene expression analysis. Hepatic microsomal ethoxyresorufin-O-deethylase
(EROD; as a measure of AhR activation), pentoxyresorufin-O-dealkylase (PROD; as a measure
of CAR activation), and 7-benzyloxyquinoline (BQ; as a measure of PXR activation) activities
were determined. Quantitative real-time PCR was used for gene expression analysis of CAR,
PXR, and AhR target genes in the liver, as well as cell growth-related genes.
After the dose change in the PXR /CARmice, no mortalities or clinical signs of
toxicity were recorded in the control or exposed groups tested, of any strain tested. Mean
terminal body weights were statistically significantly decreased (13-18%) in all exposed groups
compared with controls. The mean initial body weight for PXR 1 mice was higher than the
terminal body weight (24.0 g vs. 21.3 g), indicating a significant decrease in body-weight gain
during exposure. This decrease was also observed in PXR /CARbut to a much lesser extent.
Control mean body-weight values increased by 5-14% during the study period. Absolute and
relative liver weights were increased by >10% (compared with controls) in exposed C57BL/6
and PXR ' mice, but the liver weight responses were greatly diminished in CAR knockout
mice (-13 and +7%, respectively, compared with CAR controls). Relative liver weight
increased by 21% in PXR /CAR double knockout mice, which the study authors contributed
to the significantly lower body weight compared to controls.
Statistically significant increases in hepatic DNA synthesis (BrdU labeling index),
compared with controls, were observed in C57BL/6 and PXR ' mice, but no increase was
observed in exposed CAR 1 or PXR /CAR mice. Statistically significant liver enzyme
activity changes were as follows: increased PROD in PXR 1 mice, increased BQ in C57BL/6
and PXR 1 mice, decreased BQ in PXR /CAR 1 mice, and increased EROD in C57BL/6,
CAR 1 and PXR mice. Statistically significant changes in expression of genes in liver tissue
induction of CAR, PXR, and AhR target genes generally followed the same pattern seen with the
induction of enzyme activities. The expression levels of CAR/PXR target genes (Cyp3all,
Cyp2b9, and Cyp2bl0) were significantly increased (30- to 570-fold) in exposed C57BL/6 and
PXR 1 mice compared to controls. Cyp3all and Cyp2b9 expression levels were not
significantly altered in exposed CAR 7 mice compared to controls. Cyp2bl0, however, was
significantly increased in CAR mice. Cyp3all, Cyp2b9, and Cyp2bl0 expression levels were
unchanged in exposed PXR /CARmice compared to untreated controls. Among the AhR
target genes examined, expression of Cyplal and Cypla2 was significantly increased in exposed
C57BL/6 and PXR ' mice compared to untreated controls. Ponl was significantly increased in
C57BL/6 mice but not in PXR ' mice, and Cyplbl was unchanged in C57BL/6 and significantly
decreased in PXR 1 mice. No change in expression of Cypla2, Cyplbl, or Ponl was observed
in CAR 1 or PXR /CAR mice compared to controls, whereas Cyplal was significantly
decreased in CAR 1 mice. Of the cell growth-related genes examined (C-myc, Ccndl, Cdc25a,
and P21), the only significant increase in expression observed was that of C-myc in exposed
PXR mice. As with the Wang et al. (2015) study, the results of this study implicate a
nongenotoxic, CAR-mediated MO A, but do not unequivocally prove this to be the primary
MOA.
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Results from this 14-day study indicate a LOAEL of 64.5 mg/kg-day for increased
absolute and relative liver weight in male C57BL/6 mice (wild-type). No NOAEL could be
determined.
Weathered Toxaphene
No short-term-duration toxicity studies of weathered toxaphene in laboratory animals by
oral exposure have been identified.
Toxaphene Congeners
No short-term-duration toxicity studies of individual toxaphene congeners in laboratory
animals by oral exposure have been identified.
Subchronic-Duration Studies
Technical Toxaphene
Arnold et al. (2001); Brvce et al. (2001); Trypfaonas et al. (2000) (52 weeks)
Cynomolgus monkeys (Macacafascicularis) (two males and two females/group; -3.5 to
-3.9 years old) were administered technical toxaphene (Hercules Powder Company Inc.,
Wilmington, DE, Lot #71-132-b8-17-20) in gelatin capsules (glycerol/corn oil vehicle) at doses
of 0 or 1 mg/kg-day, 7 days/week, for 52 weeks. Animals were allotted -240 g of monkey chow
daily along with small amounts of fruit and vegetables (quantities not reported). Water was
available ad libitum and supplemented once/week with a vitamin preparation. Monkeys were
provided with environmental enrichment activities and allowed time for paired exercise runs
(within groups only).
On a daily basis, feed and water consumption were measured, visual inspections for
general health and behavior status were conducted, and menstrual status of female monkeys was
determined by vaginal swabbing. Body weight and detailed clinical evaluations were performed
weekly, including examination of the animals' skin/coat; lymph glands; eyelids; hydration status;
ears; mouth; teeth; nares; heart and respiration rate; body temperature; cardiac, pulmonary and
abdominal sounds; and abdominal, uterine, or prostate status. Monthly blood samples were
collected for 26 comprehensive standard serum biochemistry endpoints including cholesterol,
chloride, potassium, sodium, ALT, alkaline phosphatase (ALP), AST, thyroxine (T4), and
thyroxine unbound (TU). Additionally, comprehensive hematology endpoints were measured
bimonthly (Arnold et al.. 2001).
Immune system endpoints were evaluated between Weeks 36-50 (Trvphonas et al,
2000). Flow cytometry analysis and measurement of phagocytic and respiratory burst activities
in peripheral blood of the control and exposed monkeys were performed during Week 35.
Natural killer cell (NKC) activity and the response of fractionated peripheral blood leukocytes to
several mitogens (phytohemagglutinin, Concanavalin A, pokeweed mitogen) were measured
during exposure Week 36. Monkeys in exposed and control groups were immunized
intravenously with sheep red blood cells (SRBCs) (1 x io9 per kg body weight) on Weeks 36 and
42. Blood was collected just prior to the initial immunization (Day 0) and then weekly (to
Day 63 postimmunization), and titers to SRBC immunoglobulin M (IgM) and
immunoglobulin G (IgG) were determined. On treatment Week 46, monkeys were immunized
intramuscularly with 0.5 mL pneumococcus antigens (pneumovax-23; Merk, Shar and Dohme,
Canada, Kirkland, Quebec) and 0.5 mL of tetanus toxoid. Blood was collected just prior to
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immunizations (Day 0) and then weekly (to Day 35 postimmunization). IgG titers to pneumovax
and tetanus toxoid were determined using enzyme-linked immunosorbent assays (ELISAs).
All exposed monkeys and two control (one/sex) animals were necropsied after 1 year
(52 weeks) of exposure. Organ weights were obtained for liver, spleen, kidneys, adrenals,
thyroid, thymus, ovaries/testes, brain, and pituitary. Sections of these and 21 other internal
organs were processed for light microscopy; however, no histopathological incidence data were
provided in the available reports. Hearts were subsectioned to include atrium, septum,
ventricles, and conducting system. Microsomal fractions were prepared immediately from liver
samples to test for CYP450 oxygenase activities (aminopyrine, methoxyresorufin, and
ethoxyresorufin metabolism). Toxaphene contents were determined in blood samples, nuchal fat
pad samples biopsied at the time of blood sampling, portions of one kidney, and liver samples
from necropsied monkeys, as well as 24-hour urine and fecal samples 1 day prior to fat pad
biopsy.
Food consumption, body weights, and all hematology and serum biochemistry endpoints
(including T4 and TU) were plotted versus time on test for each control and exposed monkey, as
well as mean and standard deviation (SD) values for the control and exposed groups for visual
examination of time trends, or differences in exposed and control animals. Serum data for
cholesterol, sodium, potassium, and chloride levels were fit to polynomial models and assessed
for fit to the models by analysis of variance (ANOVA) techniques. Statistical analyses for
phagocytosis and respiratory burst activities used ANOVA, and a paired Mest was used for IgG
titer data for SRBC, pneumococcus, and tetanus toxoid.
No exposure-related changes in body weight, feed consumption, or water intake were
reported, but these data were not shown. Relative spleen and thymus weights were increased in
each exposed monkey of each sex, compared with values for one control monkey of each sex.
The magnitudes of increase following exposure ranged from about 22-100% for spleens and
from 66-216% for thymuses, but were reported to be "not explained" by the histological
examinations. Absolute and relative liver weights were increased by 30 and 32%, respectively,
in one exposed female monkey. Absolute and relative liver weights in the other three exposed
monkeys were generally within about 10% of reported control values (in one male, relative liver
weight was decreased by 11% compared with control). Clinical monitoring identified increased
inflammation and/or enlargement of tarsal glands (3/4 treated animals vs. 0/4 control) during
treatment Weeks 8-13, and impacted diverticulae in the upper and lower eyelids
(4/4 vs. 0/4 control) during Weeks 10-41. One exposed female showed additional clinical signs
including edema-like swelling on the eyelids and excessive dry skin, as well as increased serum
potassium and sodium levels and a steady significant decrease in cholesterol during the study.
Serum chemistry in the remaining animals was comparable to controls, with the exception of a
slight increase in chloride levels in exposed animals. Based on graphically presented data, there
was an increase in liver microsomal metabolic activities, with average increases of -35%
(methoxyresorufin), -55% (ethoxyresorufin), and -50% (aminopyrine) in exposed animals
(average of two males and one female) compared with controls, excluding the female with
swollen eyelids and dry skin. No differences in T4 or TU serum levels, or hematologic
endpoints were reported in exposed monkeys, compared with control monkeys. No statistically
significant differences in immune endpoints were observed between the exposed and control
groups, but there were consistent (across most time points) decreases in mean percentages of
CD2+CD4+ lymphocytes (16% lower compared with control), mean CD4:CD8 ratios
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(-25% reduction), and mean IgG and IgM responses to SRBCs (-10-30% lower than control
values), all suggesting a possible immune suppression effect. The study authors noted that no
remarkable histological changes in examined tissues due to toxaphene treatment were found.
The single exposure level in this study, 1 mg/kg-day, is a LOAEL for increased relative
spleen and thymus weights, clinical signs of toxicity (e.g., inflammation and enlargement of
tarsal glands), and small (but not statistically significant) decreases in three endpoints indicative
of possible immune suppression. Confidence in this determination is low because of the limited
number of animals in the study and the small magnitude of changes in the immune suppression
indicators.
Arnold et al. (2001) and Trypfaonas et al. (2001) (up to 75 weeks)
Groups of cynomolgus monkeys (Macaca fascicularis) (10 females and 5 males/group)
were administered technical toxaphene (Hercules Powder Company Inc., Wilmington, DE,
Lot #71-132-b8-17-20) in gelatin capsules (glycerol/corn oil vehicle) at doses of 0, 0.1, 0.4, or
0.8 mg/kg-day (females) and 0 or 0.8 mg/kg-day (males) for up to 75 weeks. The monkeys were
obtained from the Animal Breeding Colony, Health Canada, Ottawa, Ontario and housed
individually in stainless-steel cages in rooms maintained at 22 ± 3°C with a relative humidity of
50 ± 10%) where they acclimatized for at least 5 months. Age-matched females were randomly
distributed into four test groups, which were then assigned to one of two test rooms
(20 females/room). Older males that were proven sires were assigned to the control group.
Younger males, similar in age to females, formed the exposed groups, and males were randomly
assigned to either test room (6 or 4 males/room). Male and female animals were daily allotted
-200 and -165 g of Laboratory Monkey Chow, respectively, along with small amounts of fruits
and vegetables; water was available ad libitum. For enrichment, female monkeys were paired
(within groups) and moved to exercise cages at least 1 day/week; males were exercised
individually.
General health status, feed and water consumption, and menstrual status in females were
recorded daily, and body weight was measured weekly. Comprehensive hematology endpoints
were measured 4 months prior to dosing and during Weeks 5, 11, 19, 23, 41, and 64; samples for
analysis of 26 standard serum biochemistry endpoints (including ALT, ALP, AST, T4, and T4
binding capacity) were taken 4 months prior to dosing and during Weeks 9, 18, 36, and 63 after
the start of dosing. Because of the age difference between the control and exposed males, serum
biochemistry endpoints were measured in females only. In females, estrogen and progesterone
were monitored daily throughout a complete menstrual cycle during Weeks 22-31. Serum
hydrocortisone (i.e., Cortisol) and toxaphene concentrations in blood and adipose tissue collected
from the nuchal fat pad were determined at least monthly. Available reports for this study,
however, did not present the blood and tissue toxaphene concentration results. Immunological
testing was initiated on study Week 33 and performed in blocks until completion at Week 70.
Results from a pilot study indicated that, by Week 20, steady-state concentrations of toxaphene
were attained in blood and fat tissues ( Andrews et al.. 1996). Immune testing occurred between:
exposure Weeks 33-46 for immune components in blood, Weeks 44-53 for SRBC responses,
and Weeks 53-63 for tetanus toxoid responses and pneumococcus responses. Blood was
collected immediately prior to immunizations and at weekly intervals (for 3-5 weeks). Titers to
SRBC (IgM and IgG), pneumococcus (IgG), and tetanus toxoid (IgG) were determined using
ELISAs. Delayed-type dermal hypersensitivity (DTH) to dinitrochlorobenzene (DNCB) was
measured during Weeks 66-70. Challenged sites were evaluated by measuring skin-fold
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thickness prior to application and at 24 and 48 hours postapplication of DNCB or acetone. The
sites were scored for macroscopic appearance (erythema, contour, induration, texture, and
appearance of superficial layers compared to adjacent skin). Endpoint scores were combined to
generate total clinical scores. The monkeys were not sacrificed after the exposure period, and no
histological analyses of tissues were performed in this study.
No statistically significant exposure-related changes in feed or water consumption were
observed. Although some statistically significant differences in weight gain during
Weeks 24-42 were discussed, the study authors concluded that no toxaphene-related effects on
body weight were observed (body weight and body-weight gain data were not provided).
Weight changes in the control and exposed males were not compared because the groups differed
in age. Females in the 0.8-mg/kg-day exposure group were reported to have a higher incidence
of a slight nail bed prominence, edema of the eyelids, as well as a lower incidence of dry skin,
but the numbers of females affected were not provided. No other clinical signs were reported.
Statistical analysis found no consistent effect of exposure on hematological endpoints. The only
consistent exposure-related change in serum biochemistry endpoints was a trend with time for
decreasing cholesterol in exposed female monkeys. Menstrual status endpoints (e.g., menses
duration) were comparable between exposed and control female monkeys.
Mean anti-SRBC (IgM) responses were significantly decreased (by >27-53% compared
with control values) at several postimmunization periods in females administered 0.4 and
0.8 mg/kg-day and in males given 0.8 mg/kg-day (see Table B-3). High-dose females also had
significantly decreased anti-SRBC (IgG) and antitetanus toxoid (IgG) responses at several
postimmunization periods (see Table B-3). No significant differences in response to
pneumococcus were detected between exposed and control females (males were not tested).
Peripheral blood leukocyte populations were unaffected by exposure, with the exception of a
significant decrease (-37%) in absolute CD20+ B lymphocytes in high-dose females only,
compared with control values. The DTH response, mean percent NKC activity, and
lymphoproliferative responses of peripheral blood leukocytes to mitogens (phytohemagglutinin
and pokeweed mitogen) were comparable to control values. No exposure-related effects on
Cortisol levels were seen.
The results of this study indicate a LOAEL of 0.4 mg/kg-day for impaired immune
responses to SRBC in female cynomolgus monkeys exposed to encapsulated technical toxaphene
for up to 75 weeks. The NOAEL from this study is 0.1 mg/kg-day. At the high dose of
0.8 mg/kg-day, female monkeys also showed suppressed immune response to tetanus toxoid, and
male monkeys showed decreased anti-SRBC IgM (which was the only dose level tested in
males).
Cfau et al. (1986) (dog)
Groups of Beagle dogs (six/sex/group) were administered technical toxaphene (FBC
Chemicals, Scarborough, Ontario) via gelatin capsules (in corn oil vehicle) at nominal doses of
0, 0.2, 2.0, or 5.0 mg/kg-day for 13 weeks. An initial high dose of 10 mg/kg-day was
administered during the first 2 days, but brief convulsions, salivation, and vomiting were
observed in one male and two females, and some animals consumed little to no diet at this dose.
Subsequently, the high dose was reduced to 5 mg/kg-day for 4 weeks, followed by erroneous
administration of 2.5 mg/kg-day in Weeks 4-8. Doses of 5.0 mg/kg-day were then administered
during Weeks 9-13. For this assessment, an approximate TWA dose of 4.5 mg/kg-day was
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calculated for the high-dose group. Beagles, 7-8 months-of-age, were acclimated for 2 weeks
prior to treatment. Capsules were prepared weekly using corn oil (vehicle) and administered
daily at 8:00 AM. Animals were offered 400 g/day of standard dog food during a 1-hour feeding
period in the afternoon. Fresh water was provided ad libitum. Clinical signs were monitored
twice daily, food consumption was measured daily, and body weights were recorded weekly.
Comprehensive serum biochemistry, comprehensive hematology, and nonfasting urinalytic
(glucose, creatinine, urea nitrogen, protein, and albumin) endpoints were measured in samples
collected 2 weeks before study initiation and at Weeks 5, 10, and 13 of treatment. Gross
examinations were performed at necropsy, and all major tissues and organs were collected for
histological examination. Tissue samples were also collected for hepatic mixed function oxidase
activities (liver) and tissue residue analysis (liver and fat). Analyses were carried out in a similar
manner for the rat experiment by the same researchers (described below).
No dose-related differences in body weight, gross pathological changes, or other clinical
signs were reported (other than those noted when high-dose dogs received 10 mg/kg doses for
2 days). Body-weight and feed-consumption data were not provided. Increased mean relative
liver weights (compared with controls; see Table B-4) were observed in high-dose males (19%)
and in all treated females (13, 20, and 30% increases in low-, medium-, and high-dose groups,
respectively; statistically significant at medium and high doses). Neither absolute liver weights
nor other organ-weight differences were reported. Statistically significant increases in ALP
(see Table B-4) were observed in high-dose males and females at Weeks 5 (99 and
107%) increase, respectively) and 13 (140 and 150%> increase, respectively), compared with
controls. No other hematological, serum biochemistry or urinalytic endpoints were affected in
the exposed groups.
Incidences for hepatic periportal eosinophilia and increased cytoplasmic density were
100%) (6/6) and significantly increased in high-dose male dogs, compared with controls (1/6);
eosinophilia was also 6/6 in high-dose females, but due to higher incidence in controls (2/6), the
difference was not statistically significant (p = 0.06) (see Table B-5). Incidences for
non-neoplastic lesions in the kidney were not significantly increased in exposed groups of either
sex, compared with controls (see Table B-5). In exposed females, significantly increased
incidences for thyroidal reduced colloid density occurred at 0.2 and 2.0 mg/kg-day, but not in the
high-dose group; significantly increased incidence of reduced follicle size/follicular collapse was
only found in the 2.0-mg/kg-day female group (see Table B-5). The relationship of the thyroid
lesions to toxaphene exposure is uncertain due to the lack of a clear dose-response. No
exposure-related increased incidences of non-neoplastic lesions were reported in other tissues.
Concentrations of toxaphene residues in liver and fat samples from both sexes increased in a
dose-dependent manner, and higher concentrations were found in fat than in liver samples.
The results of this study indicate a LOAEL of 0.2 mg/kg-day for a biologically
significant relative liver-weight increase (>10%) in the absence of body-weight changes in
female Beagle dogs administered encapsulated technical toxaphene for 13 weeks. No NOAEL
could be determined. Additional changes observed in both sexes at the high dose of
4.5 mg/kg-day were increased mean serum activity of ALP and histological liver changes
(periportal eosinophilia and increased cytoplasmic density, but not cytoplasmic vacuolation).
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C.'hu et al. (1986) (rat)
Groups of Sprague-Dawley (S-D) rats (10/sex/group) were administered technical
toxaphene (FBC Chemicals, Scarborough, Ontario) via diet at concentrations of 0, 4, 20, 100, or
500 ppm for 13 weeks. The commercial test material (90% w/w solution in xylene) was vacuum
distilled to remove 80% of the xylene, and then dissolved in corn oil and incorporated into food.
The frequency of food preparation was not reported. Rats were acclimatized for 1 week before
initiation of the study with free access to food and water. The animals were randomly divided
into exposure groups. Based on body weights and food consumption, average technical
toxaphene intakes were reported by the study authors to be 0, 0.35, 1.8, 8.6, and 45.9 mg/kg-day
(males) and 0, 0.50, 2.6, 12.6, and 63 mg/kg-day (females). Clinical observations were made
daily. Food consumption and body-weight gain were recorded weekly. At necropsy, all animals
were grossly examined. Organ weights for the brain, heart, liver, spleen, and kidney were
recorded, and -32 internal organs/tissues were processed for histological examination. Blood
samples were collected and analyzed for standard hematology endpoints and numerous serum
chemistry endpoints (total protein, glucose, sodium, potassium, calcium, cholesterol, uric acid,
inorganic phosphate, total bilirubin, ALP, AST, and lactate dehydrogenase [LDH]). Enzymatic
activities of several CYP450 oxygenases were determined in liver samples (aniline hydroxylase
[AH], aminopyrine demethylase [APDM], and ethoxyresorufin-O-deethylase [EROD]). Aspirate
from femoral bone marrow was stained for cytological evaluation. Toxaphene residue
concentrations in samples of liver and perirenal fat were determined.
No clinical signs of toxicity or mortality were observed. There were slight, but
nonsignificant, increases in weight gain (816%) and food consumption (59%) in exposed male
rats, compared with controls; however, no exposure-related patterns were seen in female body
weight or food-consumption data. Mean concentrations of toxaphene in liver and fat samples
were similar in the control and low-dose groups, but increased in the higher dose groups (1.8,
8.6, and 45.9 mg/kg-day [males] and 2.6, 12.6, and 63 mg/kg-day [females]). In the highest dose
group, concentrations were 10.6 ± 4.9 ppm in liver and 103 ± 28 ppm in fat for females, and
7.4 ±3.9 ppm and 57 ± 28 ppm for males. Gross examination at necropsy revealed fatty liver in
single animals in the control and several exposed groups, as well as enlarged kidneys in
two 45.9-mg/kg-day males, one 0.35-mg/kg-day male, and one 0.50-mg/kg-day female.
Statistically significant organ-weight changes (compared with controls) were increased relative
liver weights (see Table B-6) in 45.9-mg/kg-day males (18%) and 63-mg/kg-day females (32%),
and increased relative kidney weights in 45.9-mg/kg-day males (23%). Liver activities for
CYP450 APDM and AH were also statistically significantly increased in 45.9-mg/kg-day males
(96 and 99% increased) and 63-mg/kg-day females (98 and 72% increased). Values for
hematology and serum chemistry endpoints were reported to be similar between all exposed
groups and controls.
Histochemical evaluations found statistically significant increased incidences of
non-neoplastic changes in the liver, kidneys, and thyroid of exposed groups of both sexes,
compared with controls (see Table B-7). In both sexes, liver lesions with significantly increased
incidences included anisokaryosis (at >1.8 mg/kg-day in males and >0.50 mg/kg-day in females;
moderate to severe in both sexes at the highest dose tested and minimal to mild in both sexes at
lower doses), architectural changes described as accented zonation (at >8.6 mg/kg-day in males
and >2.6 mg/kg-day in females; moderate to severe in females at the highest dose tested and
minimal to mild in both sexes at lower doses), and nuclear necrosis (at 1.8 and 45.9 mg/kg-day in
males and 63 mg/kg-day in females; minimal to mild in both sexes) (see Table B-7). The study
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authors considered the liver changes observed at doses >1.8 mg/kg-day in males and
>2.6 mg/kg-day in females to be biologically significant. Kidney lesions with significantly
increased incidences included primary tubular injury described as large eosinophilic inclusions,
which protruded into the tubular lumen (at >1.8 mg/kg-day in males and >12.6 mg/kg-day in
females; moderate to severe in males at 45.9 mg/kg-day and minimal to mild in females at
63 mg/kg-day and in both sexes at lower doses) and focal tubular necrosis (at >8.6 mg/kg-day in
males and >0.5 mg/kg-day in females; minimal to mild in both sexes) (see Table B-7). The
study authors considered the renal injury in the 0.35- and 1.8-mg/kg-day males and 0.50- and
2.6-mg/kg-day females to be minimal and focal and not biologically significant. Thyroid lesions
with significantly increased incidences in females included reduced follicular size, increased
epithelial height, cytoplasmic vacuolation, and reduced colloid density at doses >0.50 mg/kg-day
(see Table B-7). In males, thyroid lesions consisted of moderate to severe increased epithelial
height and cytoplasmic vacuolation (all severity) at >8.6 mg/kg-day, and moderate to severe
reduced colloid density at >1.8 mg/kg-day (see Table B-7). The study authors considered the
moderate to severe changes in the thyroid at >1.8 mg/kg-day in males and 63 mg/kg-day in
females to be biologically significant. No exposure-related histological changes in other organs
were found. Because the liver, kidney, and thyroid were distinct target organs within this study
and in other studies in which a dose-response was seen for the incidence of various pathologies,
all severity grades of lesions observed in these organs were considered biologically relevant for
the purpose of this PPRTV assessment.
The results of this study indicate a LOAEL of 0.50 mg/kg-day for histopathological
changes in the thyroid, kidney, and liver of female S-D rats exposed to toxaphene in food for
13 weeks. No NOAEL could be determined.
Roller ft al. (1983)
Groups of male S-D rats (12/group) were administered technical toxaphene at
concentrations of 0, 30, or 300 ppm in their diet for 9 weeks. Based on reference body weights
and food-con sum pti on rates for male S-D rats in a subchroni c-durati on study (U.S. EPA. 1988b).
estimated daily intakes of 2.6 and 25.8 mg/kg-day were calculated for this assessment.4
Cesarean-derived rats (Charles River) were obtained at 2 weeks-of-age, acclimated for 1 week,
and placed on exposure regimens postweaning. Specific details of animal facilities or access to
feed and water were not provided. After 6 weeks of exposure, the animals were antigenically
challenged with 1 mg KLH in 0.2 mL of sterile water via subcutaneous (s.c.) injection at the base
of the tail. After 15 days, the animals received a second KLH challenge. Blood samples were
collected via cardiac puncture on Days 8, 15 (primary response), and 21 (secondary response)
following the initial KLH antigen challenge and used for measuring IgG titers by ELISAs. A
separate positive control group (six rats) was given intraperitoneal (i.p.) doses of 75 mg/kg of
cyclophosphamide on Days 2, 13, and 15 after antigen challenge. The study also included other
groups of rats exposed to 50 or 500 ppm Aroclor 1254. At necropsy, the liver, spleen, and
thymus were weighed and samples were fixed for microscopic examination along with samples
of kidney, heart, lung, and the gastrointestinal (GI) tract.
4Dose estimates were calculated using reference values for body weight and food-consumption rate (U.S. EPA.
1988b)- Reference body weight for male S-D rats in a subchronic-duration study = 0.267 kg. Reference
food-consumption rate for male S-D rats in a subchronic-duration study = 0.023 kg/day.
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Mean terminal body weights in exposed groups were lower than negative control values
by <5%. Mean relative liver weight was increased in both exposure groups (11 and
24% increased compared with the control mean at the low and high doses, respectively)
(see Table B-8). The mean relative weights of thymus and spleen in exposed animals were
comparable to control values. Technical toxaphene was qualitatively described to produce less
severe liver lesions than a 9-week exposure to Aroclor 1254 at the highest dose tested (which
induced liver lesions described as megalocytosis of hepatocytes, accompanied by enlarged
hepatocytes showing various degrees of vacuolar degeneration). Incidence data were not
reported for histological findings. The histology of other organs in the exposed groups "did not
appear to differ significantly from the controls." Decreased IgG titers in response to KLH
challenge reached significance at the 15-day collection point in the 2.6- and 25.8-mg/kg-day
groups (46 and 50% decreased, respectively, compared with negative controls). However,
6 days after a secondary KLH challenge on Day 15, KLH IgG titers were comparable between
exposed groups and negative controls. Immunosuppressive effects from technical toxaphene
were not as marked or prolonged as those observed with the positive control, cyclophosphamide.
The results of this study indicate that 2.6 mg/kg-day, the lowest tested exposure level,
was a LOAEL for transient immunosuppressive effects and increased relative liver weight in
male S-D rats exposed to technical toxaphene in the diet for 9 weeks. No NOAEL could be
determined. Other liver effects (qualitatively described megalocytosis of hepatocytes) were
observed at 25.8 mg/kg-day, but not at 2.6 mg/kg-day.
Allen et al. (1983)
In two immunological studies, groups of 23-26 female Swiss-Webster mice were
administered technical toxaphene in their diet at concentrations of 0, 10, 100, or 200 ppm for up
to 8 weeks. Toxaphene (Hercules Lot X-l88825-6) dissolved in acetone was mixed into ground
rat pellets. The acetone was evaporated and the dried food fed to the animals in powder form. In
the first study, 3-week-old female mice (Washington State University Laboratory Animal
Resources) were housed in groups of five animals/cage. Estimated daily intakes of 1.9, 19.1, and
39.2 mg/kg-day were calculated based on group averages between reported means for initial and
final (8 weeks) body weights and an allometric equation for food consumption (food
consumption in kg/day = 0.056 x [body weight in kg0 6611]) described by U.S. EPA (1988b). In a
second study, 36 females (3 animals/cage; presumably 12 dams/group) were exposed beginning
3 weeks prior to mating, and throughout gestation, and lactation (~9 weeks). The offspring
(exposed ~6 weeks in duration from Gestation Day [GD] 0 through lactation) were placed on
standard control diets at weaning and aged to 8 weeks prior to immunological testing. The
numbers of offspring in each group differed between immune tests, but were clearly specified in
the report. All animals were provided with free access to food and water throughout the duration
of the study.
All mice were weighed weekly (but only data from the first study were reported). In both
studies, DTH antibody response and phagocytosis assays were performed on subsets of
8-week-old mice from each exposure group. For the DTH assays, the mice were sensitized to
mycobacteria plus 0.1 mL Freund's complete adjuvant (FCA) 29 and 20 days before testing.
Testing began with i.p. injection of tritiated thymidine, followed 24 hours later by s.c. injection
of purified protein derivative into the left rear footpad and saline into the right rear footpad. The
animals were sacrificed 24 hours later, and radioactivity levels in the foot pads were measured.
To test IgG antibody response, the mice were immunized with BSA mixed with FCA and saline
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at 29 and 8 days prior to determining IgG titers (by ELISA) in serum from blood collected from
the hearts of sacrificed mice. For phagocytosis assays conducted in the second study, the same
animals also received i.p. injections of mineral oil 5 days before sacrifice, when peritoneal fluid
was collected for macrophage analysis. Isolated macrophages were incubated in tubes
containing SRBCs for 1 hour and examined microscopically. If a macrophage had engulfed two
or more SRBCs, it was reported as phagocytic. Positive control animals for the DTH and
antibody assays in the first study were i.p. injected with 100 mg/kg cyclophosphamide on 13 and
6 days prior to testing and on the day of testing. At necropsy, liver weights were recorded and
samples of liver, kidney, spleen, heart, lungs, thymus, and GI tract were collected for
microscopic examination. Statistical analysis was performed using a Student's Mest and x2
contingency methods where appropriate.
No mortalities or exposure-related clinical signs were reported in either study. In the first
study, mean body weights in the exposed and control groups were comparable. Mean absolute
and relative liver weights in the 19.1- and 39.2-mg/kg-day groups were increased by >10%
compared with control means (14 and 27% higher absolute weights and 16 and 44% higher
relative weights, respectively) (see Table B-9). Histological changes in the liver were described
as moderate to severe, with variation in hepatocyte size (enlarged up to 2-3 times) in the
19.1- and 39.2-mg/kg-day dose groups, and some fatty infiltration in the 19.1-mg/kg-day dose
group, but incidence data were not provided. No statistically significant differences in delayed
hypersensitivity were found between exposed groups and negative controls; only the positive
control group showed DTH suppression. IgG titers to BSA were statistically significantly lower
in the 19.1- and 39.2-mg/kg-day groups than titers in the control and 1.9-mg/kg-day group (as
assessed with a %2 contingency table). Median IgG BSA titers (x 103) were 512 in the control
and 1.9-mg/kg-day groups, 128 for the 19.1-mg/kg-day group, 64 for the 39.2-mg/kg-day group,
and 32 for the positive control group.
For the second study, body-weight, organ-weight, and histology data were not provided.
The DTH and BSA IgG antibody responses were statistically significantly decreased only in the
19.1-mg/kg-day group, compared with control values: a 17.2% decrease for the DTH test, and a
median BSA IgG titer of 256 x 103 versus 512 x 103 for controls. All exposed groups had
statistically significantly decreased phagocytosis ability, compared with controls. For control,
1.9-, 19.1-, and 39.2-mg/kg-day groups, respective mean percentage (± SD) phagocytosis values
were: 75 ±28 (n = 34 mice); 51 ± 37 (n = 28); 16 ±13 (n = 24); and 28 ±12 (n = 40).
The results of the first study indicate a LOAEL of 19.1 mg/kg-day and a NOAEL of
1.9 mg/kg-day for immune suppression (reduced BSA IgG titers), increased absolute and relative
liver weights, and possible histologic changes in the liver (variation in hepatocyte size) in female
Swiss-Webster mice fed technical toxaphene in food for 8 weeks, starting at weaning. In the
second study, for similarly aged mice exposed through their dams in utero and during lactation
for about 6 weeks, the lowest exposure level, 1.9 mg/kg-day, is a LOAEL for immune
suppression assayed as decreased ability of macrophages to engulf SRBCs. No NOAEL could
be determined from the second study.
Weathered Toxaphene
No sub chronic-duration toxicity studies of weathered toxaphene in laboratory animals by
oral exposure have been identified.
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Toxaphene Congeners
No sub chronic-duration toxicity studies of individual toxaphene congeners in laboratory
animals by oral exposure have been identified.
Chronic-Duration and Carcinogenicity Studies
Technical Toxaphene
Litton Bionetics (1978)
Groups of B6C3Fi mice (54/sex/group) were fed technical toxaphene in food at
concentrations of 0, 7, 20, or 50 ppm for 18 months, followed by a 6-month observation period
(see Table 5B). The numbers of early deaths in treated groups did not differ from controls.
Available accounts of this study only report cancer findings from the histological examination of
major organs of sacrificed mice (U.S. EPA, 1988a, 1985, 1980). Other data (e.g., incidence data
for non-neoplastic histologic lesions) that may have been collected are not available (U.S. EPA,
1988a, 1985, 1980). Only incidence data for liver tumors were reported in available sources.
Based on an assumed body weight of 0.030 kg and an animal lifetime of 735 days, U.S. EPA
(U.S. EPA. 1988a, 1985, 1980) calculated the estimated doses to be 0, 0.91, 2.6, and
6.5 mg/kg-day. Liver tumor incidence data are reported in Table B-10. A statistically significant
increase in the incidence of combined hepatocellular adenomas and carcinomas was found in
male mice at 6.5 mg/kg-day, but no significantly elevated tumor incidences occurred in exposed
female groups.
NCI (1979)
Groups of Osborne-Mendel rats (50/sex/dose group) and B6C3Fi mice (50/sex/dose
group) were fed technical toxaphene in the diet for 80 weeks at TWA concentrations of 556 and
1,112 ppm for male rats, 540 and 1,080 ppm for female rats, and 99 and 198 ppm for mice (both
sexes). Due to clinical signs of toxicity, high and low dietary concentrations were lowered from
initial concentrations in both species (twice for male rats and once for female rats and male and
female mice). TWA toxaphene concentrations were calculated by NCI (1979). Estimated daily
intakes of 38.9 and 77.88 mg/kg-day for male rats, 41.6 and 83.29 mg/kg-day for female rats,
17 and 34.0 mg/kg-day for male mice, and 17 and 34.2 mg/kg-day for female mice were
calculated for this assessment based on reference body weights and food-consumption rates for
these strains (U.S. EPA, 1988b).5 Groups of 10 matched rats or mice/sex served as nonexposed
concurrent controls. The pooled control groups contained up to 52 rats/sex or 48 mice/sex from
other studies started within a 5-month period before or after the start of the technical toxaphene
study. Toxaphene was dissolved in acetone and mixed into feed with 2% corn oil. The animals
were acclimatized for 7 days (rats) or 13 days (mice), and at 5 weeks-of-age, were assigned to
exposure groups. After 80 weeks of exposure, the exposed animals were placed on control diets
and observed for an additional 28-30 weeks (rats) or 10-11 weeks (mice) prior to sacrifice.
The animals were observed twice daily for clinical signs of toxicity. The pathological
evaluation consisted of gross and microscopic examination of major tissues, major organs, and
all gross lesions from moribund animals and animals that survived to the end of the postexposure
5Dose estimates were calculated using reference values for body weight and food-consumption rate (U.S. EPA.
1988b)- Reference body weights for Osborne-Mendel rats in a chronic-duration study: 0.514 kg (male) and 0.389 kg
(female). Reference food-consumption rates for Osborne-Mendel rats in a chronic-duration study = 0.036 kg/day
(male) and 0.030 kg/day (female). Reference body weights for B6C3Fi mice in a chronic-duration study: 0.0373 kg
(male) and 0.0353 kg (female). Reference food-consumption rates for B6C3Fi mice in a chronic-duration study:
0.0064 kg/day (males) and 0.0061 kg/day (females).
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period. The following tissues were examined microscopically (following preservation in
10% buffered formalin, embedding in paraffin, and staining with hematoxylin and eosin): skin,
lungs and bronchi, trachea, bone and bone marrow, spleen, lymph nodes, heart, salivary gland,
liver, gallbladder (mice), pancreas, stomach, small intestine, large intestine, kidney, urinary
bladder, pituitary, adrenal, thyroid, parathyroid, mammary gland, prostate or uterus, testis or
ovary, and brain. Data were analyzed using the one-tailed Fisher's exact test for pairwise
comparison of tumor incidence data, the Cochran-Armitage test for detecting tumor incidence
dose-response trends, and Kaplan-Meier procedures for mortality data.
In rats, clinical signs of toxicity (hyperactivity) were observed in high-dose males
2 weeks after study initiation; consequently, the high and low dietary concentrations were
lowered. At Week 53, the majority of male and female high-dose rats developed body tremors,
and concentrations were again lowered. From Weeks 52-80, other clinical signs included
alopecia, diarrhea, dyspnea, pale mucous membranes, rough hair coats, dermatitis, ataxia, leg
paralysis, epistaxis, hematuria, abdominal distention, and vaginal bleeding, which were noted
primarily in the exposed groups (incidence data were not provided). Impaired equilibrium in
one low-dose and one high-dose female was also reported. Kaplan-Meier survival curves
indicated decreased survival probabilities in high-dose males after approximately 60 weeks, but
the Tarone test for dose-related trend in mortality was not significant in either sex. Mean body
weights of low- and high-dose females, but not males, were lower than matched controls
throughout most of the study; TWA mean body weights were lower by 8 and 13%, respectively.
A few non-neoplastic lesions in the liver and thyroid were observed in exposed groups; however,
incidences were not statistically significantly different from controls (see Table B-l 1 for
incidence data for non-neoplastic lesions found in liver, kidney, or thyroid of exposed rat
groups), and it is unknown if these lesions are related to the observed carcinogenicity.
Statistically significantly increased incidences (compared with pooled controls) were found for
liver neoplastic nodules in low-dose males; thyroid follicular-cell carcinoma or adenoma in
high-dose males; thyroid follicular cell adenomas in high-dose females; and pituitary
chromophobe adenoma, adenoma (not otherwise specified [NOS]), or carcinoma in high-dose
female rats (see Table B-12).
The study results indicate LOAELs of 77.88 (males) and 83.29 mg/kg-day (females), and
NOAELs of 38.9 (males) and 41.6 mg/kg-day (females) for technical toxaphene in
Osborne-Mendel rats exposed in their diet for 80 weeks based on statistically and toxicologically
(>10%) significant decreased body weight in females and clinical signs of toxicity in both sexes.
In mice, several died before exposure Week 19 in both dose groups (causes of death were
not described), and the dietary concentrations were lowered. By the second year of the study
abdominal distention was observed predominantly in high-dose males. Other clinical signs
included alopecia, diarrhea, rough hair coats, and dyspnea (incidences were not provided). From
Weeks 60-76, low-dose males were described as appearing hyperexcitable. Tarone tests for
dose-related trends in mortality were statistically significant, with Kaplan-Meier survival curves
showing decreased survival after about 75 weeks in high-dose males and females. The increase
in late mortality may have been related to high incidence of hepatocellular carcinomas in these
groups. Mean body weights were lower than controls in high-dose males throughout the study,
but comparable to control values in low-dose males and low- and high-dose female mice. TWA
mean body weights for high-dose males were about 6% lower than the control value. No
statistically significant increased incidences of any non-neoplastic lesions were found in any of
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the dose groups, compared with controls (see Table B-l 1 for liver, kidney, or thyroid lesions that
were reported at very low incidences in exposed groups). Significantly increased incidences of
liver tumors (carcinomas alone, or combined carcinomas and adenomas) were found in the
low- and high-dose groups of males and females (see Table B-12).
The low dose of 17 mg/kg-day is a NOAEL in both male and female B6C3Fi mice
exposed to technical toxaphene in the diet for 80 weeks. Increased late mortality at the high dose
of 34.0 and 34.2 mg/kg-day in male and female mice, respectively, may have been secondary to
carcinogenic effects in these groups. Incidences for non-neoplastic histological lesions were not
elevated in exposed groups, compared with controls, thus no LOAEL was identified.
Weathered Toxaphene
No chronic-duration toxicity or carcinogenicity oral-exposure animal studies of
weathered toxaphene have been identified.
Toxaphene Congeners
No chronic-duration toxicity or carcinogenicity oral-exposure animal studies of
individual toxaphene congeners have been identified.
Reproductive Toxicity Studies
Technical Toxaphene
Cfau et al. (1988)
S-D rats (aged 5-6 weeks; six females and three males/group) were administered
technical toxaphene at concentrations of 0, 4, 20, 100, or 500 ppm in their diet for up to
29 weeks in a one-generation reproductive toxicity study. The commercial test material
(90% w/w solution in xylene) was vacuum distilled to remove 8% of the xylene, and then
dissolved in corn oil and incorporated into food. The animals had free access to food and water.
After 13 weeks of exposure, two F0 generation females and one F0 male rat from each dose
group were cohabitated until mating occurred, allowing up to 3 weeks (time to mate was not
provided). To confirm copulation, females were examined daily for the presence of sperm in
vaginal smears or for vaginal plugs. Toxaphene exposure in F0 rats was maintained throughout
mating, gestation, and lactation. At weaning (21 days), Fla pups were switched to the same
exposure diets as their parents. At 5 weeks-of-age, the litters were randomly reduced to 15 males
and 30 females/exposure group, and exposure regimes were maintained for an additional
26 weeks (Fla adults). Following a week of rest postweaning, F0 dams still on their exposure
diets, were remated to a separate exposed male from the same dose group to generate the
Fib litter. Fib pups, along with F0 dams and sires were sacrificed 21 days postpartum.
Exposure periods in F0 dams and sires (depending upon time of mating) ranged from
-25-29 weeks. The Fla adult groups were exposed for -34 weeks total (3 weeks in utero,
3 weeks via lactation, and 28 weeks via diet); Fib pups were exposed for -6 weeks (3 weeks in
utero, 3 weeks lactation). Based on body-weight and feed-consumption data, average daily
intakes of technical toxaphene were reported to be 0, 0.36, 1.8, 9.2, and 45 mg/kg-day
(F0 males); 0, 0.36, 1.9, 8.5, and 46 mg/kg-day (F0 females); 0, 0.29, 1.4, 7.5, and 37 mg/kg-day
(Fla males); and 0, 0.38, 1.9, 9.4, and 49 mg/kg-day (Fla females).
Clinical observations of all animals were made daily. Body weights and feed
consumption were determined weekly for F0 and Fla adults. Body weights and survival of Fla
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and Fib pups were recorded at birth and at Days 4, 7, 14, and 21. Gross examinations were done
at necropsy. Organ weights for F0 and Fla adult liver, brain, heart, kidney, and spleen were
recorded, and -32 internal organs/tissues were processed for histological examination. Blood
samples were collected and analyzed for standard hematology serum chemistry endpoints
including ALP, AST, LDH, and serum sorbitol dehydrogenase (SDH) activity. Enzymatic
activities of several CYP450 oxygenases (AH, APDM, and ER) were determined in liver
samples of all adult rats and in groups of five randomly selected Fib pups/dose/sex. Differential
leukocyte counts and cytological evaluation of femoral bone marrow were performed on the
adult control and highest dose groups. Toxaphene residue concentrations in samples of liver and
perirenal fat from Fla adults, and of liver and total body from Fib pups, were determined.
No exposure-related effects were observed with respect to clinical signs, mortality,
survival indices, fertility and gestation indices, litter sizes, or postnatal F1 growth. Average
weight gain over the exposure period was significantly reduced in Fla males (-10%) and
females (-15%) in the high-dose group, compared with controls. Body-weight gain was not
affected in F0 animals. Slight fatty infiltration of the liver was reported in 9.2- and
45-mg/kg-day F0 males, 8.5- and 46-mg/kg-day F0 females, 7.5- and 37-mg/kg-day Fla males,
and 9.4- and 49-mg/kg-day Fla females. No other gross changes at necropsy were described.
Absolute and relative liver weight means were increased by >10% compared with control
means at the highest dose in F0 males and at concentrations >1.9 mg/kg-day in F0 females
(see Table B-13). In addition, absolute, but not relative, liver weight was increased by >10%
(11%) at the lowest dose of 0.36 mg/kg-day in F0 females (see Table B-13). In Fla males and
females, absolute and relative liver weights were clearly increased by >10% compared with
controls at the highest dose (see Table B-13). The mean relative liver weight reported in the
study for control Fla males was abnormally low (1.7% of body weight), compared with control
means for Fla females (3.9%), F0 males (3.6%), and F0 females (3.7%). It is likely that this
value reflects a typographical error, as the mean and SD are identical to those for absolute kidney
weight reported in a neighboring column in the study table. Based on an expectation that the
Fla male control relative liver weight should be in the range of-3.6, it appears that the only
relevant increase (>10%) is in the highest dose group, consistent with the effect seen in
Fla females.
Absolute and relative kidney weights were increased by >10% in 45-mg/kg-day F0 males
and in 7.5- and 37-mg/kg-day Fla males, but not in any exposed F0 or Fla female groups
(see Table B-13). The report by Chu et al. (1988) did not mention any other exposure-related
changes in other organ-weight data (brain, heart, and spleen).
Toxaphene residue levels in the fat and liver of adult rats, and in the liver and the whole
body of rat pups, increased in a dose-dependent manner. Levels in fat were generally higher than
liver levels (7-17 and 7106% higher in Fla adult males and females, respectively), and also
higher in Fla females than Fla males (58-269% higher). Concentrations in the liver were
comparable between sexes. In the high-dose groups, toxaphene residue levels in the livers of
male and female Fib pups were 54 and 108% higher, respectively, than in the livers of F0 dams.
Changes in biochemical endpoints (compared with controls) included increased (28%)
serum cholesterol in the 46-mg/kg-day F0 females and decreased (up to 21%) serum glucose in
the F0 males exposed to >1.8 mg/kg-day. The study authors were uncertain about the biological
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significance of these changes. Compared with controls, significant increases in hepatic APDM
activity were observed in the 45-mg/kg-day F0 males (59%) and in the Fla adult females at
>9.4 mg/kg-day (up to 45%). In Fib female pups from the 0.36- and 1.9-mg/kg-day F0 dams,
APDM activity decreased significantly (-45 and 26%, respectively). The activities of other
liver oxygenases were not reported. Liver protein (mg/mL homogenate) was reported for
Fib pups only and, compared with controls, was elevated (62%) in Fib male pups from the
46-mg/kg-day F0 dams, and in Fib female pups from the 8.5- and 46-mg/kg-day F0 dams
(40 and 88%, respectively). No other exposure-related changes in biochemical endpoints were
described.
Histochemical evaluations of tissues from F0 and Fla adult rats found statistically
significantly increased incidences of non-neoplastic changes in the liver, kidneys, and thyroid of
exposed groups of both sexes, compared with controls (see Tables B-14 and B-15). Liver lesions
with significantly increased incidences included cytoplasmic density, anisokaryosis, increased
vacuolation, and increased cytoplasmic homogeneity. These lesions were found most
prominently in high-dose F0 and Fla adults of both sexes, but elevated incidences of minimal to
mild gradations occurred in Fla adults at doses >0.29 (males) and 0.38 mg/kg-day (females)
(see Tables B-14 and B-15). The study authors suggested that many of these observed liver
changes could be largely adaptive in nature. Similarly, in the kidney, significant increases in
lesions (primary renal tubular injury in F0 males and females and Fla adult males, and
anisokaryosis, pyknosis, interstitial sclerosis in F0 males) occurred primarily in the highest-dose
group, with increased incidences of minimal to mild gradations of some lesions occurring in
some of the lower dose groups (see Tables B-14 and B-15). Thyroid changes, graded moderate
to severe, were limited to reduced colloid density and colloid inspissation in 45-mg/kg-day
F0 males. Statistically significant increases in incidences of other thyroid lesions (when all
severity grades are considered) were observed in many exposure groups as follows: reduced
colloid density (at >1.8 mg/kg-day in F0 males, 1.4 mg/kg-day in Fla males, and
>0.38 mg/kg-day in Fla females); colloid inspissation (at 0.36, 1.8, and 45 mg/kg-day in
F0 males and >0.29 mg/kg-day in Fla males); increased epithelial height (at 46 mg/kg-day in
F0 females, 0.29, 1.4, and 7.5 mg/kg-day in Fla males, and >0.38 mg/kg-day in Fla females);
follicular collapse/angularity (at 45 mg/kg-day in F0 males, 1.9 mg/kg-day in F0 females,
1.4 and 37 mg/kg-day in Fla males, and 0.38 and 1.9 mg/kg-day in Fla females); and reduced
follicle size (at >9.4 mg/kg-day in Fla males) (see Tables B-14 and B-15). The study authors
expressed uncertainty regarding the biological significance of the changes in the thyroid.
Statistically significant incidences of non-neoplastic changes in thyroid, liver, and kidney were
also present in 3-week-old Fib pups exposed to the highest exposure level (see Table B-15). As
with the Chu et al. (1986) rat study, all severity grades of lesions observed in this study are
considered biologically relevant for the purpose of this PPRTV assessment because the liver,
kidney, and thyroid are distinct target organs that exhibited a dose-response for the incidence of
various pathologies.
A LOAEL of 0.29 mg/kg-day is identified for this study based on increases in
histopathological lesions in the thyroid and liver of Fla adult males. No NOAEL could be
determined.
The results also indicate that the highest dietary exposure level of 500 ppm
(45-46 mg/kg-day in the F0 adults), was a NOAEL for combined reproductive function
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endpoints measured in exposed F0 male and F0 females (fertility and gestation indices, litter
sizes, and pup survival).
Kennedy et al. (1973)6
Groups of S-D albino rats (8 males and 16 females/group) were exposed daily to
technical toxaphene in food at concentrations of 0, 25, and 100 ppm for up to 42 weeks in a
multigeneration reproductive toxicity study. Technical toxaphene was mixed with 2% corn oil
and added to standard rat food. Choice of exposure levels was guided by an early report of
hepatic fatty degeneration and cytoplasmic vacuolation in rats fed 100 ppm technical toxaphene,
but not 25 ppm, in food (Lehman. 1952). Based on reference body weights and
food-con sum pti on rates for S-D rats in a chronic-duration study (U.S. EPA. 1988b). estimated
daily intakes of 0, 1.7, and 6.88 mg/kg-day (males) and 0, 2.0, and 7.99 mg/kg-day (females)
were calculated for this assessment.7 Diets were prepared fresh weekly, and the animals were
allowed free access to feed. F0 animals were received as weanlings and allowed to acclimate for
1 week prior to initiation of the study at 28 days-of-age. The F0 rats were mated at
100 days-of-age (two females to one male). Males were removed upon confirmation of
copulation (sperm-positive vaginal examination). The first litters (Fla) were reduced to
10 pups/litter on Day 5 and retained for 21 days. F0 dams were given 10 days of rest
postweaning and were remated to generate Fib litters. At weaning, 8 males and 16 female
Fib pups were selected from each group at random as parental animals for the F2 generation.
This procedure was continued through three successive two-litter generations until F3b pups
were born. F0 dams were sacrificed after a total of 42 weeks of exposure to technical toxaphene,
whereas Fib and F2b parents were sacrificed after 39 weeks.
All parental rats were necropsied at sacrifice, and absolute and relative organ weights
were determined (for liver, kidneys, spleen, gonads, heart, brain, adrenals, and thyroid).
Thirty-four tissues and organs from parental rats were collected, preserved, and examined
histologically. Weight gains for parental animals (F0, Fib, F2b) were determined prior to the
first mating and up to time of sacrifice. Total and free serum cholesterol were measured in
eight parental animals/sex/group after 39 weeks of exposure. Complete blood and platelet
counts, cell indices (mean corpuscular volume [MCV], mean corpuscular hemoglobin [Hb]
concentration, as well as color, saturation, and volume indices), prothrombin time, activities of
serum glutamic-pyruvic transaminase (GPT = ALT) and glutamic-oxaloacetic transaminase
(GOT = AST), serum ALP, fasting blood sugar, blood urea nitrogen (BUN), icterus indices, and
urinalysis (glucose, albumin, and microscopic exam) were determined on three parental
rats/sex/generation (control and 100-ppm dietary dose group). Reproductive endpoints were
6This study was conducted by Industrial Bio-Test Laboratories (IBT) and published in a peer-reviewed journal prior
to the development of Good Laboratory Practices (GLP) in 1979. In its 2007 Manual for Investigation ofHPV
Chemicals, OECD (2007) noted that 618 of 867 nonacute toxicity studies conducted by IBT prior to its closing in
1978 (including subacute, subchronic-duration, carcinogenicity, reproductive toxicity/teratogenicity, genotoxicity,
and neurotoxicity studies) were found to be invalid during a post hoc audit program conducted by U.S. EPA and the
Canadian Health and Welfare Department. There is no information on whether the study by Kennedy et al. (1973)
was audited.
Dose estimates were calculated using reference values for body weight and food-consumption rate (U.S. EPA.
1988b). Reference body weights for S-D rats in a chronic-duration study: 0.523 kg (male) and 0.338 kg (female).
Reference food-consumption rates for S-D rats in a chronic-duration study = 0.036 kg/day (male) and 0.027 kg/day
(female).
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determined for each litter, including mating index, fertility index, pregnancy index, parturition
index, mean litter size, live birth index, 5-day survival index, and lactation index.
All viable and stillborn progeny were counted, examined for physical abnormalities, and
records of survival/mortalities were maintained. Male and female weanling body weights were
measured in all litters. Gross and microscopic pathologic studies (except for organ weights)
were conducted on F3b weanlings only.
All data were reportedly analyzed by ANOVA, and Student's /-test was used for
intergroup comparisons. However, adequate summary data for independent analysis of most
endpoints (e.g., incidence data for discontinuous variables or mean, SD, and n for continuous
variables) were not provided in the published report.
No treatment-related mortalities, clinical signs of toxicity, or effects on growth of
parental rats were reportedly observed (summary data were not provided). No differences in
cholesterol concentrations, hematologic endpoints, urinalytic endpoints, or other clinical
chemistry findings (e.g., ALT, AST, ALP serum activities) were reportedly observed between
F2 parental exposed and control groups; results for these endpoints in other parental generations
(F0, Fl) were not mentioned. Mean relative liver weight in exposed male groups was increased
compared with controls by >10% in Fl males at 1.7 mg/kg-day and 2-8% in F0 and F2 males.
In females, mean relative liver weights were increased compared with controls at doses of
2.0 and 7.99 mg/kg-day, respectively, as follows: 12 and 26% in the F0 groups, 9 and 0.3% in
the Fl groups, and 9 and 10% in the F2 groups. Gross and microscopic examination reportedly
revealed no exposure-related effects on tissues and organs, except for hepatic cytoplasmic
vacuolization in 63% of all high-dose parental rats (compared with none in controls); incidence
data specific to generation, sex, and exposure groups were not reported. No differences were
reportedly observed between the production of Fl, F2, or F3 litters compared with controls in
mating index, fertility index, pregnancy index, parturition index, mean litter size, live birth
index, 5-day offspring survival index, lactation index, and weanling body weights of offspring.
Mean values were reported for these endpoints by exposure group and generation, but values for
n and SD or standard error (SE) were not provided. No significant differences were reported
between exposed and control groups for the number of pups delivered, number of stillborn pups,
number of viable pups at birth or through lactation, or growth of offspring through lactation
(summary data were not provided). The findings from gross and microscopic examination of
tissues from F3b weanlings were not provided, except for a general statement that no evidence
for exposure-related teratogenicity was found.
The study results indicate LOAELs of 1.7 and 2.0 mg/kg-day for Fl males and
F0 females, respectively, for increased (>10%) relative liver weight in rats fed technical
toxaphene in food for 39-42 weeks. The highest exposure level, 6.88 and 7.99 mg/kg-day in
males and females, respectively, is a reproductive NOAEL for the lack of effects on reproductive
function in three exposed parental male generations, absence of gross abnormalities in offspring,
and lack of effects on offspring survival and growth. Confidence in these determinations is
compromised by reporting deficiencies in the available published report, as well as by
uncertainties regarding the reliability of studies conducted by IBT.
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Keplinger et al. (1970)
tut mBbbbbbbbb8 mma
Groups of Swiss white mice (4 males and 14 females/group) were exposed daily to
technical toxaphene in food at concentrations of 0 or 25 ppm for 120 days, in a five-generation
reproductive toxicity study. Technical toxaphene (source and purity were not provided) was
mixed into standard Purina laboratory chow. This study also tested several other substances (the
pesticides aldrin, dieldrin, chlordane, and dichlorodiphenyltrichloroethane [DDT]), but only the
toxaphene results are reported here. Mice were raised from a laboratory stock. Based on the
mean reference body weights and food-consumption rates for BAFi and B6C3Fi mice in a
subchronic-duration study (U.S. EPA, 1988b), estimated daily intakes of 4.7 mg/kg-day (males)
and 5.1 mg/kg-day (females) technical toxaphene have been calculated for this assessment.8
Study details such as animal care, time of study initiation, or whether time had elapsed between
the start of exposure and mating were not provided. F0 mice (120 days old) were divided into
two cages for mating (two males and seven females/cage). Pregnant females were removed,
housed individually, and continued on their exposure diets. After weaning, F1 litter 1 (Fla) pups
were fed the same exposure diet as the mother until Postnatal Day (PND) 120, when they were
sacrificed for chemical or histological analysis. F0 dams were given 7 days of rest postweaning
then remated to generate a second litter (Fib). At weaning, 4 males and 14 Fib females were
selected as breeders for the next-generation litters (F2a and F2b). Remaining pups and
F0 parents were sacrificed for chemical or histological analysis. This breeding scheme was
continued through five successive, two-litter generations until F5b pups were born. Mice from
all litters were exposed in utero, through lactation, and then in feed until sacrifice at PND 120
(nonbreeders).
At sacrifice, the liver, lungs, kidneys, brain, small intestine, thymus, adrenals, spleen,
stomach, and heart were fixed for microscopic examination. Organ weights were measured for
brain and liver from 10 males and 10 females/group. For exposed and control animals, the
number of females that delivered, total number of pups born, average litter size, survival at
three time points (birth, 4 days, and 4 months), total number of animals weaned, and pup weight
at weaning and at sacrifice were recorded. These data were used to determine fertility, gestation,
pup viability, lactation, and pup survival indices. No specifics regarding the chemical analyses
performed were provided. Statistically significant differences in indices were evaluated using
two unspecified methods.
Compared with controls, no changes in fertility, gestation, lactation, pup viability, or pup
survival indices, resulting from exposure to technical toxaphene, were observed. No significant
changes in brain or liver weight were noted, although data were not provided. The study authors
stated that all compounds tested caused changes in the liver that included fatty metamorphosis,
increased basophilic activity, or hepatic cell necrosis. The study authors also noted that these
liver changes were "usually more marked in the second litter;" however, no substance-specific
incidence data were reported. Damage to the kidney, lungs, and brains was also reported for
most of the test substances (except for the lack of toxaphene-induced brain changes), but
incidence data were not reported. Results from chemical analyses were not presented.
8Dose estimates were calculated using the mean reference body weight and food-consumption rate values for BAFi
and B6C3Fi mice in a subchronic-duration study (U.S. EPA. 1988b"). Mean reference body weights: 0.0270 kg
(male) and 0.0225 kg (female). Mean reference food-consumption rates: 0.0051 kg/day (male) and 0.0046 kg/day
(female).
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The study results indicate a NOAEL of 4.7 mg/kg-day (males) for the lack of effects on
reproductive function endpoints in the four exposed parental generations, and lack of effects on
offspring survival or growth in the animals exposed to technical toxaphene in utero, through
lactation, and in feed to PND 120. The reporting of the histology findings in this study is
inadequate to assess whether the single exposure level is a LOAEL for histological changes in
the liver or other organs.
Weathered Toxaphene
No oral-exposure reproductive toxicity studies of weathered toxaphene in laboratory
animals have been identified.
Toxaphene Congeners
No oral-exposure reproductive toxicity studies of individual toxaphene congeners in
laboratory animals have been identified.
Developmental Toxicity Studies
Technical Toxaphene
Chernoff and Carver (1976)
Groups of pregnant mice (26-90/group) and rats (16-39/group) were administered
technical toxaphene (Hercules, Inc.) at doses of 0, 15, 25, or 35 mg/kg-day in corn oil (vehicle)
by gastric intubation, on GDs 7-16. The CD-I mice and CD rats were obtained from the Charles
River Breeding Laboratories (Wilmington, MA) and were maintained in constant temperature
rooms (22-26°C) on a 12-hour light/dark cycle. Feed and water were available ad libitum.
Pregnancies were confirmed by presence of sperm in the vaginal smear. Body weights measured
on GD 6 were used for dose and body-weight gain calculations. Mortality and the number of
pregnancies that went to term were recorded. Mice and rats were sacrificed on GDs 18 and 21,
respectively. Maternal-weight gain (weight of intact animal - [weight of the removed
uterus + weight measured at GD 6]), liver weight, weights of live fetuses, number of implants,
and fetal mortality were recorded. At sacrifice, fetuses were grossly examined then placed in
fixatives for 3-5 days for either necropsy or skeletal examination (sternal and caudal ossification
centers).
In exposed rats, statistically significantly decreased incidences of pregnancies that went
to term were observed at 25 and 35 mg/kg-day, and increased dam mortality occurred at
35 mg/kg-day (see Table B-16). All exposed groups showed statistically significantly decreased
maternal-weight gain, ranging from 22-41% decrease below the control average
(see Table B-16). Fetal body weights were decreased 5-12% across all dose groups compared
with controls, but decreases were only statistically significant at the 25-mg/kg-day dose
(see Table B-16). No significant differences were observed between the control and exposed
groups in maternal liver to body-weight ratios, the average number of implants, or fetal mortality
(see Table B-16). Exposed groups had statistically significantly lower average numbers of fetal
sternal ossification centers than the control group, but the numbers of caudal ossification centers
were not significantly different in exposed versus control groups, except for a lower number in
the 25-mg/kg-day group. Dose-related changes were not found in the number of fetal anomalies
revealed by necropsy.
In rats, the lowest dose, 15 mg/kg-day, is a maternal LOAEL for decreased
average-weight gain. More severe maternal effects were noted at 25 and 35 mg/kg-day,
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including decreases in the number of pregnancies brought to term and increased maternal
mortality. The 15-mg/kg-day dose is also a developmental LOAEL for decreased numbers of
sternal ossification centers in fetuses and biologically significant (>5%) decreased fetal body
weight. No exposure-related effects were observed on the average number of implants, fetal
mortality, or the occurrence of fetal anomalies. No NOAELs can be identified.
In mice, pregnancies brought to term were not significantly different in the exposed and
control groups (see Table B-16). There was a marginal increase in dam mortality at
35 mg/kg-day (p = 0.07). Maternal-weight gain was significantly reduced in the 25- and
35-mg/kg-day dams, and average liver weights were increased >23% (relative to controls) in all
exposed groups (see Table B-16). Average fetal body weights and numbers of ossification
centers were not significantly different between exposed and control groups. Necropsy of
fetuses found encephaloceles (portions of the brain protruding from the skull) in five high-dose
litters and 11 high-dose fetuses, but none in the control or other exposed groups
(see Table B-16). The elevated incidence of high-dose litters with this anomaly, compared with
the control incidence, did not reach statistical significance (p = 0.07).
In mice, the lowest dose, 15 mg/kg-day, is a maternal LOAEL for increased relative liver
weight. Decreased maternal average-weight gain occurred in the 25- and 35-mg/kg-day groups,
and there was a marginal increase in maternal mortality at 35 mg/kg-day. No maternal NOAEL
can be identified. Due to the lack of statistical significance observed for any of the reproductive
and developmental parameters, the highest dose tested (35 mg/kg-day) is identified as a
reproductive and developmental NOAEL, and no LOAEL can be confidently identified. Note,
however, that the incidence of litters with encephaloceles, although not statistically significant,
was 5/61 (11%) at 35 mg/kg-day. Additionally, the average fetal mortality was twice that of the
control value at 25 mg/kg-day, albeit with a substantial associated variance (i.e., SD) for both
groups.
Chernoff and Kavlock (1983)
Groups of pregnant female CD-I mice (25/group) were exposed daily to technical
toxaphene by gavage (in 0.5 mL corn oil vehicle) at doses of 0 or 75 mg/kg-day on GDs 8-12.
The source of technical toxaphene was not provided. Feed and water were available ad libitum.
Maternal-weight gain and pup survival at birth were assessed. Litters were counted and
weighed on PNDs 1 and 3. Dead pups were necropsied and gross abnormalities were noted.
Dams that did not give birth by GD 22 were sacrificed, and uteri were examined for the presence
of implantation sites. Comprehensive examination of fetuses for visceral and skeletal variations
or anomalies was not conducted.
No significant differences were found between the exposed and control groups in clinical
signs, maternal mortality, or pup survival on PNDs 1 and 3. Statistically significant decreases in
maternal-weight gain (mean of 4.2 g in exposed group vs. mean of 7.4 g in the control group, or
a 43% decrease) and the mean weight of pups on PND 1 (mean of 1.54 g in exposed
group vs. mean of 1.68 g in the control group, or an 8% decrease) were observed. No difference
in pup weight between the exposed and control groups was observed on PND 3.
The results indicate that the only dose tested, 75 mg/kg-day, is a maternal LOAEL for
decreased maternal-weight gain in CD-I pregnant mice exposed by gavage to technical
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toxaphene on GDs 8-12, and also a developmental LOAEL for decreased PND 1 pup body
weight. No NOAEL could be determined.
Chernoff et al. (1990)
Groups of pregnant female S-D rats (~25/exposed group and ~50/control group) were
administered technical toxaphene via gavage at doses 0 or 32 mg/kg-day on GDs 6-15.
Technical toxaphene (purity 100%) was obtained from the U.S. EPA Chemical Repository
(Research Triangle Park, NC) and mixed with 1.0 mL corn oil (vehicle). The animals were
allowed free access to feed and water. Pregnancies were confirmed upon presence of sperm in a
vaginal smear. The animals were monitored for weight loss, mortality, or for other overt signs of
toxicity. Dams were weighed on GD 6 prior to administering the first dose and then every
2 days throughout the dosing period. A subset of animals from each group (4-6 exposed animals
and 6-30 control animals) was sacrificed on GDs 8, 12, 16, and 20. Terminal body weights (on
GD 20) were measured and corrected for gravid uteri weights. At each time point, thymus,
spleen, and adrenal glands were removed and weighed. Developmental endpoints assessed on
GD 20 included the number of pregnancies, number of litters and fetuses, and fetal survival and
body weight. Half of each litter was fixed in formalin and examined for soft tissue anomalies.
The remaining fetuses were cleared in potassium hydroxide (KOH) and stained for skeletal
examination. The lateral and fourth ventricles as well as the renal pelvis lumina were scored on
a scale of 1 (no visible space) to 4 (apparent hydrocephaly or hydronephorsis).
Decreased maternal survival was observed beginning after two exposures on GD 8
(86% survival compared with 100% in controls). By GDs 16 and 20, maternal survival dropped
to -50%), whereas 100%> of control animals survived for the duration of the study. The causes of
death were not explored. Body-weight gain data were presented as the difference between
average body-weight gain of exposed groups and controls. At each time point examined within
the exposure period (GDs 8, 12, and 16), exposed animals gained statistically significantly less
weight (from study initiation to each time point) than the controls (-23.7, -48.5, and -55.2 g,
respectively). No significant differences in maternal-weight gain (from study initiation) were
observed at GD 20. Thymus, spleen, and adrenal organ weights in exposed dams were less than
control values at scattered collection time points, but statistical significance was not consistent
between time points and the reduced organ weights did not appear to be linked to exposure
duration. No statistically significant differences were observed between the exposed and control
groups in fetal mortality or body weight. No statistically significant differences between the
control and exposed groups were found in the mean proportion of fetuses with fourth or lateral
ventricle scores or left/right kidney scores >1.0. A statistically significant increase in the mean
proportion of fetuses with supernumerary ribs, compared with controls, was reported. No other
visceral or skeletal variations or anomalies in fetuses were reported to be induced from exposure.
Litter or fetal incidence data for variations or anomalies were not specified in the available
report.
The study results indicate a LOAEL of 32 mg/kg-day for increased mortality and
decreased maternal-weight gain in pregnant S-D rats exposed to technical toxaphene via gavage
during GDs 6-15. The single exposure level also was a developmental LOAEL for increased
proportion of fetuses with supernumerary ribs, with no effects on fetal mortality and body weight
or evidence for other visceral or skeletal variations or anomalies. No NOAEL could be
determined.
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Crowder et al. (1980)
Groups of S-D rats (three, presumably pregnant, females/group) were administered
technical toxaphene (Boots-Hercules, Inc.; purity 99.9%) at doses of 0 or 6 mg/kg-day by gavage
(in 0.1 mL corn oil) between GDs 7-21. F0 dams were gavaged once daily from Day 7 after
introduction to males (presumed GD 7) through delivery of pups. The rats were caged in pairs
until 2-3 days prior to giving birth, and then caged individually. The animals were given free
access to food and water. Additional housing details were not provided. Body weight and
mortalities of F1 pups (controls or exposed in utero) were recorded daily. Pups were evaluated
for ontogeny of reflexes (grasp-hold reflex, righting reflex, and startle reflex) three times daily,
beginning on PND 7. The ages at which the first responses occurred, and ages when 90% of the
trials by the litter were positive, were recorded. Between 14-16 weeks-of-age, five male and
five female F1 rats/group were chosen randomly for evaluation of learning and learning transfer
abilities using maze testing. Each rat received 10 maze trials/day.
Offspring body weights and survival were reportedly similar between the exposed and
control groups (data were not provided). No statistically significant differences in grasp-hold,
startle reflexes, or in the mean number of days for observation of first correct righting reflex
responses were observed in exposed pups, compared with controls. However, a statistically
significant increase in the mean number of days for >90% of the trials to be positive in litters for
the righting reflex was observed in the exposed group, compared with controls (18.5 days,
exposed; 15.5 days, control). No statistically significant effects were found on maze learning or
learning transfer, assessed between 14-16 weeks-of-age. The study authors concluded that there
was no exposure-related impact on learning, and interpreted the additional time for exposed
litters to master the righting reflex to be due to slight changes in motor function and behavior.
The study results indicate a developmental LOAEL of 6 mg/kg-day for delayed
attainment of the righting reflex ability in offspring of female S-D rats exposed to technical
toxaphene between GDs 7-21 (birth). This exposure level produced no effects on early postnatal
development of grasp-hold, startle reflexes, maze learning, or learning transfer, as assessed
between 14-16 weeks-of-age. No NOAEL could be determined.
Olson et al. (1980)
Groups of pregnant Holtzman albino rats (three/group) were fed diets delivering reported
daily doses of 0 or 0.050 mg technical toxaphene/kg-day, 0.002 mg Toxicant A (p-42)/kg-day, or
0.002 mg Toxicant B (p-32)/kg-day from GD 5 through weaning and PND 30, in a study of
neurobehavioral endpoints in the offspring that were exposed through PND 90. The animals
were housed in stainless steel cages and allowed to acclimate for 5 days prior to initiation of the
study. Food and water were available ad libitum. Test substances (sources and purities were not
reported) were mixed into standard Purina Rat Chow; diets were prepared fresh every
1-2 weeks. Toxicant A and Toxicant B were presumably purified from technical toxaphene in
the laboratory of one of the study authors (F. Matsumura), who published reports on separation
techniques [Nelson and Matsumura (1975) and Matsumura et al. (1975), both cited in Olson et
al. (1980)1. Sufficient quantities of Toxicant A were not available to sustain exposure past
PND 40; the animals in this group were provided Toxicant B through the remainder of the study.
Offspring were kept with their dams until PND 30, then housed in pairs until PND 45, at which
time, the offspring were moved to individual cages for the remainder of the experiment.
Offspring were kept in the same treatment groups as their mothers through PND 90, when
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neurobehavioral testing was completed. All pups were tested for early development of
swimming ability, and 15-16 offspring per group were administered maze tests.
Pup weights were recorded on PNDs 7, 10, 13, and 16. Between PNDs 7-17, tests of
swimming ability and righting reflex were administered to all pups. A subjective scoring system
from "0" through "4" was used for the swimming test: "0" indicated lack of swimming ability
and "4" indicated mature swimming ability. For righting reflex, a score of "1" was assigned if
the rat landed on all fours during the righting test, otherwise a score of "0" was given. Following
these initial tests, 15 control animals (8 females and 7 males) and 16 exposed animals (8/sex),
were randomly selected for symmetrical maze testing to evaluate motivational behavior,
learning, and retention. Selected rats were first aged to PND 70; body weights were reduced to
80-85% of their recorded weight at PND 66, and weight reduction was maintained for the
duration of the study. Behaviors analyzed for motivational testing included: the number of errors
made before mastering a problem; the number of retraces or re-entries made into the "start"
endbox; the sum of the trials necessary for the task to be mastered; and the cumulative time
required to solve the problem. The animals were given 2 days of rest after completing the
motivational test, then challenged with five more trials of the same problem to measure learning
and retention. When behavioral tests were complete (PND 90), the animals were sacrificed.
Liver, kidney, heart, lung, and brain were weighed and fixed for histological examination, along
with tissues from thyroid, adrenal, stomach, small and large intestines, and bladder.
No exposure-related clinical signs or differences in offspring body weight between
exposed and controlled animals were reported for technical toxaphene-, Toxicant A-, or
Toxicant B-exposed groups. Histological examination revealed no significant changes in tissues
for exposed groups, compared with controls. Exposed-group relative liver and kidney weights
were reported comparable to controls (data were not provided). Delays in attaining the ability to
swim were noted in all exposed groups on PNDs 10, 11, and 12, compared with controls, but
swimming abilities in exposed offspring were comparable to controls by PNDs 13-16. Righting
reflex ability between PNDs 7-17 was reported to be statistically significantly inferior to
controls in technical toxaphene-exposed offspring, but not in Toxicant A- or Toxicant B-exposed
offspring. However, these data were not provided and the magnitude of the apparent effect could
not be evaluated. Thus, due to this incomplete reporting, the biological significance of
exposure-related delays in swimming ability and righting reflex is unclear. Results from maze
testing conducted between PNDs 70-90 indicated: (1) no significant differences between
exposed and control groups in motivational endpoints (data not shown in the report), (2) no
consistent differences between exposed and control groups in most measures of learning
(see Table 1 of the study report), and (3) no consistent differences between exposed and control
groups in most measures of retention (see Table 3 of the study report). The report discusses
some of the possible implications of the few measures of maze learning or learning retention
showing statistically significant differences among the groups, but most measures of
performance were similar between exposed and control groups.
The results indicate NOAELs of 0.002 mg Toxicant A (p-42)/kg-day and 0.002 mg
Toxicant B (p-32)/kg-day for the absence of clear biologically significant neurobehavioral
effects, body-weight effects, or effects on tissue histology in Holtzman albino rat offspring
exposed in utero, during lactation, and up to 90 days-of-age. No LOAELs can be identified. No
NOAEL or LOAEL can be identified for technical toxaphene in this particular study due to
incomplete reporting as described above.
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Weathered Toxaphene
No oral-exposure developmental toxicity studies of weathered toxaphene in laboratory
animals have been identified.
Toxaphene Congeners
The only oral developmental toxicity study of toxaphene congeners in laboratory animals
identified was the study reported by Olson et al. (19801 which included tests of two individual
toxaphene congeners (Toxicants A and B [p-42 and p-32]), in addition to technical toxaphene.
Study methods and results for Toxicants A and B are included along with the methods and
results for technical toxaphene in the study summary provided above. The results indicate
NOAELs of 0.002 mg Toxicant A (p-42)/kg-day and 0.002 mg Toxicant B (p-32)/kg-day for the
absence of clear biologically significant neurobehavioral effects, body-weight effects, or effects
on tissue histology in Holtzman albino rat offspring exposed in utero, during lactation, and up to
90 days-of-age. No LOAELs can be identified.
Inhalation Exposures
No toxicity studies of any type have been identified for animals exposed by inhalation to
technical toxaphene, weathered toxaphene, or individual toxaphene congeners.
OTHER DATA (SHORT-TERM TESTS, OTHER EXAMINATIONS)
Older Studies Identifying the Liver and Kidney as Toxicity Targets of Technical
Toxaphene
A number of early animal toxicity studies reviewed by U.S. EPA (1985) and U.S. EPA
(1980) reported finding liver or kidney effects following subacute or longer term repeated
exposure to technical toxaphene in the diet, by gavage, or via capsules with food. However,
primary reports of some of these older studies were not available for this assessment, and
primary reports for others indicate inadequate designs or reporting. Effects reported in the
shorter term studies included "questionable liver pathology" in male and female rats fed 50 or
200 ppm in the diet for 2-9 months (Ortega et al.. 1957). "questionable liver pathology and renal
tubular degeneration" in dogs given gavage doses of 4 mg/kg-day (in corn oil) for 44 or 106 days
[Lackey (1949a) as cited in U.S. EPA (1980)1. and no apparent adverse effects in rats fed
189 ppm in the diet for 12 weeks [Clapp et al. (1971) as cited in U.S. EPA (1980)1. Effects
reported in longer duration studies included: "liver pathology" in rats fed 100 ppm in the diet,
but not 25 ppm, for life [Lehman (1952a) as cited in U.S. EPA (1980)1; "liver pathology" in rats
fed 25 ppm in the diet for life (Fitzhueh and Nelson. 1951); central nervous system (CNS)
stimulation at concentrations of 1,000-1,600 ppm, "slight liver damage" at 100 ppm, and no
effect at 25 ppm in rats fed toxaphene-containing diets for life [Hercules, Inc. updated as cited in
U.S. EPA (1980)1; "moderate liver damage" at 200 ppm, "slight liver damage" at 40 ppm, and no
effects at 5-20 ppm in dogs fed toxaphene-containing diets for life [Hercules, Inc. updated as
cited in U.S. EPA (1980)1; "liver necrosis" in dogs fed encapsulated doses (in corn oil) of
5 mg/kg-day for 1,360 days [Hercules, Inc. updated as cited in U.S. EPA (1980)1; and no clinical
or histological effects in monkeys fed 10-15 ppm in the diet for 2 years [Hercules, Inc. updated
as cited in U.S. EPA (1980)1.
Other reports of liver responses of unclear biological significance following oral
exposure to technical toxaphene include (1) increased serum and liver activities of
gamma-glutamyl transpeptidase (GGTP) in rats given single gavage doses of 110 mg/kg
technical toxaphene (in mineral oil) or 16.5 mg/kg-day for 120 days (Garcia and Nlourelle. 1984)
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and (2) increased (with time of exposure) mean relative liver weights and hepatic enzyme
activities for androgen hydroxylases in rats of unspecified strain or sex given an oral dose of
0.24 mg/kg-day for up to 180 days (Peakall, 1976).
Genotoxicity and Mutagenicity
Results from short-term-duration tests of genotoxic, mutagenic, and clastogenic
endpoints for technical toxaphene, weathered toxaphene, and several toxaphene congeners are
summarized in Table 6.
Technical Toxaphene
In short-term-duration tests of technical toxaphene's mutagenicity in prokaryotes or
nonmammalian eukaryotes, positive results were obtained with some species or strains
(e.g., Salmonella typhimurium TA98 and TA100), but not in others (e.g., S. typhimurium
TA1535 and TA1537), and mutagenic activity could be negatively affected by metabolic
activation in some test strains or species (e.g., Vibrio fischeri M169). Technical toxaphene was
mutagenic in several studies of S. typhimurium strains TA98 and TA100 with and without
metabolic activation (see Table 6 for references), in Escherichia coli B/r strains WP2s and
SR712 with and without activation (Houk and Demarini. 1987). in V fischeri M 169 without
activation (but not mutagenic with activation) (Boon et al.. 1998). and in Neurospora crassa
strain H-59 (uvs-2) without metabolic activation (Brockman et al. 1983). Fractions of technical
toxaphene (e.g., methanol fraction, hexane fraction, and a KOH-treatment of technical
toxaphene) were also mutagenic in S. typhimurium TA100 without metabolic activation (Hooper
et al.. 1979). However, technical toxaphene was (1) not mutagenic with or without metabolic
activation in S. typhimurium strains TA 1535 and TA 1537 (Nlortelmans et al.. 1986);
(2) mutagenic without activation, but not mutagenic or equivocally mutagenic with activation, in
strains TA97 and TA 104 (Schrader et al.. 1998); and (3) equivocally mutagenic without
activation and not mutagenic with activation in strain TA 102 (Schrader et al.. 1998). Technical
toxaphene-induced DNA damage in one assay (SOS chromotest) with E. coli PQ37, but not in
another assay (umuC test) with S. typhimurium TA1535/pSK1002 (Bartos et al.. 2005).
Incubation of isolated DNA with technical toxaphene did not increase DNA breakage rates over
control rates (Griffin and Hill. 1978).
Consistent evidence for genotoxic potential has not been demonstrated across a variety of
in vitro and in vivo tests in mammalian species. In Chinese hamster V79 cells, technical
toxaphene was mutagenic at the hypoxanthine-guanine phosphoribosyltransferase (HGPRT)
locus, induced DNA strand breaks only in the presence of UV-B light, and did not markedly
increase sister chromatid exchanges (SCEs) with or without metabolic activation from human
HepG2 cells or induce unscheduled DN A synthesis above that induced by UV-B light (Schrader
et al.. 1998). Technical toxaphene increased SCEs in human lymphoid LAZ-007 with or without
metabolic activation (Sobti et al.. 1983) and in Chinese hamster lung (CHL) (Don) cells without
metabolic activation (Steinel et al.. 1990). but the responses were mostly less than twofold above
control values. Technical toxaphene also induced micronuclei (MN) in primary beluga whale
skin fibroblasts with or without metabolic activation (Gauthier et al.. 1999) and in human HepG2
cells without metabolic activation (Wu et al.. 2003). Technical toxaphene did not induce
dominant lethal mutations in mice after single i.p. doses as high as 180 mg/kg or 5 days of
gavage doses as high as 80 mg/kg-day (Epstein et al.. 1972). Other short-term in vivo tests
found no evidence for DNA damage (alkaline elution assay) in livers of rats after gavage
administration of technical toxaphene doses of 12 or 36 mg/kg (Kitchin and Brown, 1994. 1989)
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or increased DNA adducts in livers of mice administered seven daily gavage doses of technical
toxaphene as high as 100 mg/kg-day (Hedli et aL 1998).
In summary, mutagenic activity and DNA damage from technical toxaphene has been
demonstrated in some species and strains of nonmammalian prokaryotic and eukaryotic cells, but
not in others. In mammalian test systems, technical toxaphene only induced HGPRT locus
mutations and DNA strand breaks in cultured hamster cells also exposed to UV-B light, but did
not induce SCEs in the absence of UV-B light. In other studies, technical toxaphene induced
SCEs to a moderate degree in human lymphoid LAZ-007 cells and CHL cells, and induced MN
in human HepG2 cells and beluga whale skin fibroblasts. In vivo tests with laboratory animals
found no evidence for technical toxaphene induction of dominant lethal mutations, DNA
damage, or DNA adducts. In addition to the lack of consistent evidence of the genotoxicity of
technical toxaphene in various test systems, there is an absence of increased DNA breakage rates
when technical toxaphene is incubated with isolated DNA samples (Griffin and Hill. 1978).
Weathered Toxaphene
Possible genotoxicity, mutagenicity, and clastogenicity of weathered toxaphene has been
examined in two studies in bacteria. Weathered toxaphene samples (toxaphene residues
extracted from fish, from a contaminated site, and toxaphene residues extracted from soil aged
with technical toxaphene for 104 weeks) were mutagenic in S. typhimurium strain TA100 with
and without metabolic activation, similar to unweathered samples of technical toxaphene (Young
et aL 2009). In the other genotoxicity study with "weathered" toxaphene samples, DNA damage
detected by the umuC assay in S. typhimurium TA1535/pSK1002 was found after exposure to
technical toxaphene treated with UV light for up to 9 hours, but was not detected after exposure
to nontreated technical toxaphene (Bartos et al.. 2005).
Toxaphene Congeners
A few studies in bacteria have found no consistent evidence for mutagenicity of
toxaphene congeners. No definitive increased mutagenic activities (compared with controls)
were detected with or without metabolic activation in S. typhimurium TA100 exposed to p-26,
p-32, p-62, or p-32 (Steinberg et al.. 1998); in S. typhimurium TA1535, TA 1537, TA1538,
TA98, or TA 100 exposed to heptachlorobornane-1 (p-32) (Hooper et aL. 1979); or in V. fischeri
M 169 exposed to p-26, p-50, p-62, or a synthetic mixture of p-32, p-26, p-50, and p-62 (Boon et
al.. 1998). However, in the Mutatox® assay with V. fischeri Ml69, p-32 alone induced mutations
(Boon et al.. 1998). Other genotoxicity studies using toxaphene congeners were not available.
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Studies in prokaryotic organisms
Technical toxaphene
Mutation
Salmonella typhimurium
strains TA98, TA100, TA1535,
and TA1537
0, 100, 333, 1,000,
3,333, 6,666,
10,000 ng/plate
+
TA98, TA100
TA1535, TA1537
+
TA100, TA98
TA1535, TA1537
Plate incorporation assay (S9
activation): Precipitate was noted at
>1,000 |ig/platc. Reproducible
dose-related increases were considered
a positive response even if the increase
was 500 |ig/plate. The number
of revertants was reduced by >50% in
TA98 and TA100 with the addition of
S9 metabolic activation.
Schrader et al.
(1998)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Mutation
S. typhimurium strain TA100
0, 100, 200, 500,
1,000 ng/plate
+
+
Ames test (S9 activation): Data
expressed as number of revertants per
plate (after correction for the solvent
blank). Number of revertants
increased with increasing TT dose.
Young et al.
(2009)
Mutation
(/.-prophage
induction)
Escherichia coli B/r strains
WP2S (X) and SR712
(trpE, uvrDs)
0,0.10,0.19, 0.38,
0.76, 1.52,3.05,6.10,
12.2 mM
+
+
Precipitate was observed at the highest
dose. The minimum TT
concentrations required to produce a
significant positive response at the
99% CI were 1.26 and 0.13 mM, in the
presence and absence of S9 activation,
respectively. Prophage induction
increased with increasing dose.
Honk and
Demarirti
(1987)
Mutatox® assay
Vibrio fischeri M169
0.0083-4.23 mg/L
+
Light levels were measured at 1-hr
intervals from 10-24 hr after
incubation; the concentration that gave
the CMR determined from the
dose-response curve.
LOEC = 3.15 mg/L;
CMR >4.23 mg/L°
Metabolic activation with S9.
Boon et al.
(1998)
Mutation
S. typhimurium strain TA100
(ethanolic KOH treatment of
toxaphene [1:1 or 1:10 molar
ratio; 24 hr at 25°C])
NR
+
ND
Reversion data not reported.
Hooper et al.
(1979)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Mutation
S. typhimurium strain TA100
(fractionated toxaphene:
methanol, hexane, and
recrystallized from
isopropanol)
100-3,000 ng/plate
+
Mother liquor
fraction
+
Methanol fraction
+
Hexane fraction
±
Recrystallized
ND
Revertants:
Mother liquor = 500 rev/mg
Methanol = 2,640 rev/mg
Hexane = 200 rev/mg
Recrystallized =116 rev/mg
Hooper et al.
(1979)
DNA damage
(SOS
chromotest)
E. coli PQ37
0, 2.5, 5.0, 10.0, 20,
40.0 mg/L
+
ND
SOS induction factor was >1.5 at TT
concentrations >10 mg/L.
Bartos et al.
(2005)
DNA damage
(¦umuC test)
S. typhimurium strain
TA1535/pSK1002
0, 2.5, 5.0, 10.0,
40.0 mg/L
ND
A nonsignificant, dose-dependent
increase in/f-galactosidase activity was
observed at 2.5-40.0 mg/L.
Bartos et al.
(2005)
Weathered toxaphene
Mutation
S. typhimurium strain TA100
exposed to TT or a
soil-weathered TT sample
0, 100, 200, 500,
1,000 ng
toxaphene/plate (aged
soil extract)
+
+
Revertant colonies were slightly
increased with the addition of S9. The
number of rcvcrtants/|ig toxaphene
was also plotted. Weathered and TT
were similarly mutagenic.
Hx-Sd and Hp-Sd represented 8.7 and
5.7%, respectively, of the total residual
toxaphene in soil aged for 104 wk.
Youti" et al.
(2009)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Mutation
S. typhimurium strain TA100
exposed to toxaphene residues
extracted from fish collected at
a contaminated site
Approximate doses
from fish tissue extract
of 0, 150, 320, 380,
800, and 900 ng
toxaphene/plate
+
+
The data were expressed as
rcvcrtants/|ig residual toxaphene.
Hx-Sd and Hp-Sd represented 19.5 and
14.6% of residual toxaphene found in
fish collected from a contaminated site.
Weathered and TT displayed similar
mutagenicity. An initial increase
followed by a decrease in the number
of revertants was observed with
increasing concentration in the
presence of S9. The number of
revertants increased with increasing
dose in the absence of S9.
Young et al.
(2009)
DNA damage
(¦umuC test)
S. typhimurium strain
TA1535/pSK1002 exposed to
irradiated TT or TT
0, 7.5, 15, 30, 60 mg/L
+
ND
Toxaphene irradiated with UV for 6
and 9 hr inhibited bacterial growth at
>60 mg/L (-78.8%); after 9-hr growth
inhibition was observed at 30 mg/L
(-85.2%) and at 60 mg/L (-100%).
/;-Galactosidasc activity was increased
at 15 mg/L (9-hr irradiation) and
30 mg/L (6-hr irradiation) by 3.2- and
2.4-fold, respectively. The abundance
of toxaphene congeners decreased with
UV radiation time. TT without
irradiation did not induce
/;-galactosidasc activities beyond
1.5-fold of control values.
Bartos et al.
(2005)
Toxaphene congeners
Mutation
S. typhimurium strains TA98
and TA100 (p-26, p-50, p-62,
and p-32)
0,312.5,625, 1,250,
2,500 ng/plate
Microsuspension modified Ames test
(S9 activation).
Steinberg et
al. (1998)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Mutation
S. typhimurium strain TA100
(p-26, p-50, p-62, and p-32)
0, 156.25,312.5,625,
1,250, 2,500, 5,000,
10,000 ng/plate
p-26 induced an increase in the number
of revertants only at 10,000 |ig/plate
(20%).
p-50 induced increases in the number
of revertants only at 156.25 and
5,000 ng/plate (29 and 15-22%,
respectively).
Precipitation of p-26, p-50, and p-32
was observed at 10,000 ng/plate.
Steinberg et
al. (1998)
Mutation
S. typhimurium strains
TA1535, TA1537, TA1538,
TA98, and TA100
(heptachlorobornane-I = p-32)
100-2,000 ng/plate
NA
Hooper et al.
(1979)
Mutatox® assay
V fischeri M169 (p-32)
9.87-1,500 ng/L
+
Light levels were measured at 1-hr
intervals from 10-24 hr after
incubation; the concentration that gave
the CMR determined from the
dose-response curve.
LOEC = 580 ng/L;
CMR = 770 |ig/Lc
Metabolic activation with S9.
Boon et al.
(1998)
Mutatox® assay
V fischeri M169 (p-26, p-50,
p-62)
9.87-1,500 (ig/L
p-26, p-50, and p-62 were assayed
individually with negative results
without and with S9 activation.
Boon et al.
(1998)
Mutatox® assay
V. fischeri Ml69 (T-4 mix)
9.87-1,500 ng/L
p-32, p-26, p-50, and p-62 were
assayed as a mixture with negative
results without and with S9 activation.
Boon et al.
(1998)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Studies in nonmammalian eukaryotic organisms
Technical toxaphene
Mutation (ad-3
forward mutation
test)
Neurospora crassa
H-12(uvs-2+) and H-59(uvs-2)
Five concentrations
(NR)
+
H-59(uvs-2)
ND
H-59(uvs-2)
Toxaphene induced multilocus
deletions (-20%) and intracistronic
mutations (-40%) in H-59(uvs-2).
Brock man et
al. (1983)
Studies in mammalian cellsin vitro
Technical toxaphene
Mutation
(HGPRT)
Chinese hamster V79 cells
0, 1,2,3,4,5,6,7.5,
10 ng/mL
-
-
Metabolic activation with y-irradiated
human HepG2 cells.
Schrader et al.
(1998)
Mutation
(HGPRT)
Chinese hamster V79 cells
exposed to TT and UV-B light
0.001-1.0 ng/mL
+
+
V79 cells were preincubated with TT
and exposed to 5 J/m2 UV-B light,
resulting in a dose-dependent increase
in HGPRT mutations, up to a twofold
maximum (1 |ig/mL). over cells
exposed to UV-B radiation alone.
Schrader and
Langlois
(1999)
SCE
Chinese hamster V79 cells
0, 1,2,3,4,5,6,7.5,
10 iig/mL
Metabolic activation with y-irradiated
human HepG2 cells. Although small
increases in SCE were observed in
some exposed groups, no consistent
and marked TT-induced increases in
SCE were observed. Positive control
groups exposed to
ethylmethane-sulfonate without
activation or
dimethylbenz[a]anthracene with
activation showed three- to fourfold
increases in SCE frequencies.
Schrader et al.
(1998)
Unscheduled
DNA synthesis
Chinese hamster V79 cells and
human HepG2 cells
NR; 0.001-1.0 ng/mL
(presumably)
TT had no effect on excision repair of
UV-B damage.
Schrader and
Langlois
(1999)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
DNA damage
and repair
Chinese hamster V79 cells
0.001-1.0 ng/mL
+
+
DNA strand breakage was increased in
UV-B exposed cells when
preincubated with TT in a
dose-dependent manner.
Schrader and
Langlois
(1999)
SCE
Human lymphoid LAZ-007
cells
10~4, 105, 10~6 M
+
+
TT induced increased SCE frequency
by 59 and 81% (-S9) and 44 and 54%
(+S9) at low and high doses,
respectively.
Sobti et al.
(1983)
SCE
CHL (Don) cells
0, 5, 10, 15, 20 ng/mL
+
ND
Cells were incubated for 18, 22, or
26 hr in media containing TT and
10 |iIVI BrdU, and then for an
additional 2.5 hr in fresh media
containing colcemid for SCE and
metaphase analysis.
SCEs increased significantly in both a
dose- and time-dependent manner.
Steinel et al.
(1990)
Mammalian
cytokinesis-block
MN assay
Primary beluga whale skin
fibroblasts
0, 0.05, 0.5, 5,
10 iig/mL
+
+
(at 0.5 ng/mL)
(0.05, 5, and
10 ng/mL)
In the absence of S9, mean MN cell
frequency significantly increased at all
doses, ranging from a 1.7- to a 3.6-fold
increase at 10 |ig/mL. In the presence
of S9, a statistically significant
increase in MN cells was observed at
0.5 ng/mL, but then decreased as dose
increased from 0.5-10 |ig/mL.
Gauthier et al.
(1999)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
MN assay
Human HepG2 cells
0, 5, 10, 20, 40 nM
+
ND
MN frequencies increased with
increasing concentration. At 20 and
40 |iIVI increases of 64 and 105%
(compared with controls) were
observed.
When HepG2 cells were pretreated
with 5, 10, 20, or 40 ^M TT then
exposed to 50 |iIVI BaP, the
genotoxicity of BaP alone was
increased by 34 and 56% at 10 and
20 |iIVI; results at 40 |iIVI were
considered questionable due to
significant cytotoxicity.
Wu et al.
(2003)
Studies in mammalsin vivo
Technical toxaphene
Dominant lethal
mutagenicity
Male ICR/Ha Swiss mice
(7 and 9/group in the low- and
high-dose groups, respectively)
were administered a single i.p.
injection of toxaphene.
Exposed males were then
mated to unexposed females
for 8 wk following treatment.
Females were sacrificed 13 d
after the middle of their
breeding period, and uteri were
examined
0, 36, 180 mg/kg
2 mortalities (2 males out of 9 total)
were observed at 180 mg/kg.
TT did not significantly increase
incidences of early fetal
deaths/pregnancy or preimplantation
losses over that observed in controls.
Corpora lutea counts were not
conducted: decreases in total implants
were determined by contrasting total
implants of females mated with control
and treated males.
Epstein et al.
(1972)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
Dominant lethal
mutagenicity
Male ICR/Ha Swiss mice
(7-9/group) were administered
toxaphene via i.p. or gavage
for 5 d. Exposed males were
then mated to unexposed
females for 8 wk. Females
were sacrificed 13 d after the
middle of their breeding
period, and uteri were
examined
0, 40, 80 mg/kg
TT did not significantly increase
incidences of early fetal
deaths/pregnancy or preimplantation
losses over that observed in controls.
Corpora lutea counts were not
conducted: decreases in total implants
were determined by contrasting total
implants of females mated with control
and treated males.
Epstein et al.
(1972)
DNA damage
Female S-D rats (8/group)
were administered 2 doses of
toxaphene (in corn oil) via
gavage at 21 and 4 hr before
sacrifice. Livers were
collected and prepared for the
following assays: DNA
alkaline elution, ODC,
glutathione activity, CYP450,
and serum ALT
0, 12 mg/kg
TT did not induce hepatic DNA
damage (although ODC activity and
CYP450 were increased).
Kitchin and
Brown (1989)
DNA damage
(alkaline elution)
90-d-old female S-D rats
(8-9/group) were administered
2 doses of toxaphene (in
1:1:1 mixture of Tween 80,
Triton X-100, and DMSO) via
gavage at 21 and 4 hr before
sacrifice
0, 12, 36 mg/kg
Endpoints assessed included ODC
activity, ALT activity, and hepatic
DNA damage by alkaline elution.
Hepatic CYP450 content and Ames
test results from other studies were
included in the study report's data
table.
Kitchin and
Brown
(1994);
Kitchin et al.
(1992)
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Table 6. Summary of Genotoxicity and Mutagenicity Studies for Technical Toxaphene, Weathered Toxaphene, and
Toxaphene Congeners
Endpoint
Test System
Doses/Concentrations
Tested3
Results without
Activationb
Results with
Activationb
Comments
References
DNA adducts
(32P-post
labeling)
Male CD-I mice (3-4/group)
were exposed to toxaphene in
corn oil by gavage for
7 consecutive d. Mice were
sacrificed 24 hr after final
dosing and DNA was extracted
from livers
0, 10, 25, 50,
100 mg/kg
DNA adducts in liver DNA were
measured using nuclease-Pl orbutanol
enhancements of the 32P-postlabeling
method.
No evidence of DNA adduct formation
was found in treated or control mice.
Hedli et al.
(1998)
Studies in subcellular systems
Technical toxaphene
DNA damage
ColEl plasmid DNA isolated
from E. coli K-12 strain CR34
thy" B1" thr" leu"
0.1 mg/mL
No increase in DNA breakage rates
compared with controls was observed.
Griffin and
Hill (1978)
aLowest effective dose for positive results, highest dose tested for negative results.
b+ = positive; ± = weakly positive; - = negative.
cBoonetal. (1998) did not clearly define a positive result, but a fourfold increase in bioluminescence is often taken as a positive response with the Mutatox® assay.
ALT = alanine aminotransferase; BaP = benzo[a]pyrene; BrdU = bromodeoxyuridine; CHL = Chinese hamster lung; CI = confidence interval; CMR = calculated
maximum response; CYP450 = cytochrome P450; DMSO = dimethylsulfoxide; DNA = deoxyribonucleic acid; HGPRT = hypoxanthine-guanine
phosphoribosyltransferase; Hp-Sd = heptachlorobornane; Hx-Sd = hexachlorobornane; i.p. = intraperitoneal; KOH = potassium hydroxide;
LOEC = lowest-observed-effect concentration; MN = micronuclei; NA = not applicable; ND = no data; NR = not reported; ODC = ornithine decarboxylase; SCE = sister
chromatid exchange; S-D = Sprague-Dawley; TT = technical toxaphene; UV = ultraviolet.
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Metabolism/Toxicokinetic Studies for Technical Toxaphene
Results from laboratory animal studies indicate rapid and extensive absorption of
technical toxaphene by the GI tract, metabolic dechlorination of absorbed toxaphene congeners,
and extensive and fairly rapid elimination of metabolites in the urine and feces [see AT SDR
(2014) for a more comprehensive review of toxicokinetic data]. For example, oral
administration of single doses of 20 mg/kg 36Cl-labeled technical toxaphene in rats resulted in
excretion of about 52.6% of the administered radioactivity within 9 days in feces (37.3%) and
urine (15.3%) (Crowder and Dindal. 1974). In another study that administered single doses of
14.2 mg/kg 36Cl-labeled technical toxaphene to rats, 14-day urine and feces contained 49.1 and
26.9% of the administered 36Cl-radioactivity, respectively (Ohsawa et al.. 1975). Considerable
amounts of radioactivity in the collected excreta in these studies was in the form of chloride ions,
providing evidence for extensive metabolic dechlorination of congeners in technical toxaphene.
A comparison of 36Cl-radioactivity eliminated in urine following Na36Cl or 36Cl-toxaphene
administration indicated similar elimination half-lives of 2-3 days (Ohsawa et al.. 1975).
Following oral administration of single doses of 14C-labeled technical toxaphene to rats, 14-day
urine and feces contained 31.8 and 27.1% of the administered radioactivity, respectively
(Ohsawa et al.. 1975). Solvent fractionation analysis indicated that about 57 and 39% of
urine- and feces-14C radioactivity, respectively, were in the form of partially dechlorinated
metabolites (Ohsawa et al.. 1975).
DERIVATION OF PROVISIONAL VALUES
Tables 7 and 8 present summaries of noncancer and cancer reference values, respectively,
for technical toxaphene derived in this document. For weathered toxaphene or individual
toxaphene congeners, data were inadequate to derive noncancer provisional reference values, for
either oral or inhalation exposure. Appendix A discusses some options for deriving screening
provisional values for weathered toxaphene.
No effort was made to derive provisional cancer values for technical toxaphene because a
cancer oral slope factor (OSF), inhalation unit risk (IUR), and an associated weight-of-evidence
(WOE) determination for technical toxaphene is currently on the U.S. EPA IRIS website (U.S.
EPA, 1988a). Data for weathered toxaphene or individual toxaphene congeners are inadequate
for assessing their possible carcinogenicity.
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Table 7. Summary of Noncancer Reference Values for
Technical Toxaphene (CASRN 8001-35-2)
Toxicity Type
(units)
Species/Sex
Critical Effect
p-Reference
Value
POD
Method
POD
(HED)
UFc
Principal Study
Subchronic
p-RfD (mg/kg-d)
Rat/M
Cytoplasmic vacuolation
in the thyroid
3 x 1CT4
BMDLio
0.0092
30
Chu et al. (1988)
Chronic p-RfD
(mg/kg-d)
Rat/M
Cytoplasmic vacuolation
in the thyroid
9 x 1CT5
BMDLio
0.0092
100
Cfau et al. (1988)
Subchronic
p-RfC (mg/m3)
NDr
Chronic p-RfC
(mg/m3)
NDr
BMDLio = 10% benchmark dose lower confidence limit; HED = human equivalent dose (in mg/kg-day);
M = male(s); NDr = not determined; p-RfC = provisional reference concentration; p-RfD = provisional reference
dose; POD = point of departure; UFC = composite uncertainty factor.
Table 8. Summary of Cancer Reference Values for
Technical Toxaphene (CASRN 8001-35-2)
Toxicity Type (units)
Species/Sex
Tumor Type
Cancer Value Principal Study
p-OSF (mg/kg-d) 1
An OSF value is available on IRIS (U.S. EPA. 1988a)a
p-IUR (mg/m3)
An IUR value is available on IRIS (U.S. EPA. 1988a)a
"Value based on increased incidence of hepatocellular tumors in mice and thyroid tumors in rats following oral
exposure.
IRIS = Integrated Risk Information System; p-IUR = provisional inhalation unit risk; p-OSF = provisional oral
slope factor.
DERIVATION OF ORAL REFERENCE DOSES
Derivation of a Subchronic Provisional Reference Dose
The rat study designed to evaluate reproductive toxicity by Cfau et al. (1988) is selected
as the principal study for deriving the subchronic provisional reference dose (p-RfD) for
technical toxaphene, with rationale provided below. The critical effect is cytoplasmic
vacuolation (all severity grades) in the thyroid of F0 male S-D rats following 25-29 weeks of
dietary exposure. The study report was published in a peer-reviewed journal. The study is
adequate with regard to design (e.g., inclusion of controls and several exposure levels) and
performance pertaining to examination of potential toxicity endpoints, and presentation of
materials, methods, and results. Details of the study are provided in the "Review of Potentially
Relevant Data" section. No pertinent toxicity data were identified for laboratory animals
repeatedly exposed orally to weathered toxaphene or toxaphene congeners.
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Convulsions (presumably from effects on the nervous system) are the most common
effect reported in acute poisoning case reports. A limited number of epidemiological studies
have examined possible associations between occupational exposure to technical toxaphene and
noncancer diseases. In U.S. male pesticide applicators, self-reported hypothyroidism was
associated with "ever-use" of 50 specific insecticides (including technical toxaphene) (Goldner
et aL 2013). but no statistically significantly elevated ORs were found for amyotrophic lateral
sclerosis and "ever-use" of any of the subject pesticides, including technical toxaphene (Kamel et
al., 2012). Additionally, a statistically significant exposure-response trend in association with
rheumatoid arthritis was observed for lifetime days of toxaphene use (Mever et aL, 2017).
Results from several short-term- and sub chronic-duration oral toxicity studies, as well as
two oral reproductive toxicity studies and a number of oral developmental toxicity studies,
identify the liver, kidney, thyroid, and immune system (including spleen and thymus) as sensitive
noncancer toxicity targets of repeated exposure to technical toxaphene. LOAELs in Table 5A
for systemic effects induced by oral exposure to technical toxaphene range from
0.2-4.5 mg/kg-day for increased liver weight and liver, kidney, and thyroid lesions in 13-week
dietary studies in rats and dogs (Chu et al.. 1986) and a single-generation reproductive toxicity
study in rats (Chu et al.. 1988) to 7.3 mg/kg-day for increased absolute and relative liver weight
(>10%) (14-day dietary study), or 60.3 mg/kg-day for increased absolute and relative liver
weight (>10%), serum ALT activities, hepatic cell proliferation rates, and liver MDA
concentrations in mice (28-day dietary study) (Wane et al.. 2015).
Immune system noncancer effects from oral exposure to technical toxaphene are
examined in multiple species, and several LOAELs were identified (see Table 5A). In
cynomolgus monkeys, a LOAEL of 1 mg/kg-day was identified for decreased IgG and IgM
responses to SRBCs, as well as changed proportions of lymphocytes in females administered
technical toxaphene in gelatin capsules for 52 weeks (Arnold et al.. 2001; Bryce et al.. 2001;
Tryphonas et al.. 2000). as well as LOAELs of 0.4 mg/kg-day in females and 0.8 mg/kg-day in
males for decreased IgM responses to SRBCs in a 75-week study of cynomolgus monkeys
administered encapsulated technical toxaphene (Arnold et al.. 2001; Tryphonas et al.. 2001).
Additionally, a LOAEL of 2.6 mg/kg-day was identified for transient decreased IgG responses to
keyhole limpet hemocyanin (KLH) in male rats fed dietary technical toxaphene for 9 weeks
(Koller et al.. 1983), and a LOAEL of 19.1 mg/kg-day was identified for decreased IgG response
to bovine serum albumin (BSA) in mice fed dietary technical toxaphene for 8 weeks (Allen et al,
1983).
NOAELs in Table 5 A for reproductive toxicity from oral exposures of laboratory animals
to technical toxaphene are higher than the lowest LOAELs for liver, kidney, thyroid, or immune
system effects. The reproductive NOAELs are: 45 (males) and 46 mg/kg-day (females) in a
one-generation rat study (Chu et al, 1988); 6.88 (males) and 7.99 mg/kg-day (females) in a
multipie-generation rat study (Kennedy et al.. 1973); and 4.7 (males) and 5.1 mg/kg-day
(females) in a m ul ti pi e-generati on mouse study (Keplinger et al, 1970).
LOAELs in Table 5A for developmental effects from oral gestational exposure of
laboratory animals to technical toxaphene also are higher than the lowest LOAELs for liver,
kidney, thyroid, or immune system effects. Developmental LOAELs for technical toxaphene
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(given by gavage, unless otherwise noted) in rats are as follows: 15 mg/kg-day for decreased
number of sternal, but not caudal, ossification centers in fetuses exposed on GDs 7-16 (Chernoff
and Carver, 1976); 32 mg/kg-day for increased proportion of fetuses with supernumerary ribs in
fetuses exposed on GDs 6-15 (Chernoff et aL 1990); and 6 mg/kg-day for delayed attainment of
righting reflex in offspring exposed on GDs 7-21 (Crowder et aL 1980). Developmental
LOAELs identified in mice included: 75 mg/kg-day for decreased body weight on PND 1, but
not on PND 3, in pups of mice exposed on GDs 8-12 (Chernoff and Kavlock. 1983), and
1.9 mg/kg-day for suppression of SRBC phagocytosis by macrophages (one of three evaluated
immune responses) in offspring of mice exposed to dietary technical toxaphene for 3 weeks
before mating and continuing throughout gestation and lactation ( Allen et aL. 1983).
Justification
Looking at the laboratory animal oral toxicity database as a whole, degenerative changes
to the liver and kidney are relatively consistent findings across a range of technical toxaphene
exposure levels tested (see Table 5A). In the 28-day rat study by Wane et al. (2015). the effects
on the liver endpoints evaluated (e.g., relative liver weight, hepatocyte cell proliferation rates,
serum ALT) were mostly confined to the highest exposure group evaluated, 60.3 mg/kg-day
(human equivalent dose [HED] = 8.51 mg/kg-day), which reportedly produced no histological
evidence for liver regenerative hyperplasia or hepatocyte necrosis. No changes in any of these
endpoints occurred in rats exposed to 5.9 mg/kg-day (HED = 0.85 mg/kg-day) (Wang et al..
2015). Increased relative liver weight was biologically significant (>10% increase), but not
statistically significant in female Beagle dogs at 0.2 mg/kg-day (HED = 0.1 mg/kg-day), the
lowest dose tested. With respect to technical toxaphene-induced kidney toxicity, although the
Chu et al. (1986) and Chu et al. (1988) studies identified kidney lesions in rats that were
primarily minimal to mild in severity at the lowest doses tested, no histological changes in the
kidney were observed following subchronic exposure in other studies of rats (Koller et al.. 1983)
and mice (Allen et al.. 1983).
Identification of immune suppression as a health hazard from repeated oral exposure to
technical toxaphene comes from several reports on experiments conducted in cynomolgus
monkeys that exhibited decreased immunoglobulin responses to SRBC, as well as spleen and
thymus effects, at low doses (0.4 mg/kg-day; HED = 0.2 mg/kg-day) (Arnold et al.. 2001; Brvce
et al.. 2001; Tryphonas et al.. 2000). This is supported by observations of decreased ability of
macrophages to engulf SRBCs in 8-week postweaning offspring of Swiss-Webster mice exposed
to dietary technical toxaphene (1.6 mg/kg-day) 3 weeks before mating, and throughout gestation
and lactation and decreased IgG response to BSA in adult male Swiss-Webster mice exposed to
19.1 mg/kg-day dietary toxaphene for 8 weeks ( Allen et al.. 1983). and transiently decreased IgG
responses to KLH in male S-D rats exposed to technical toxaphene dietary doses of 2.6 and
25.8 mg/kg-day (Koller et al.. 1983).
There is ample evidence suggesting that thyroid toxicity is a relevant and sensitive
response following exposure to technical toxaphene. In a study of U.S. male pesticide
applicators and "ever-use" of 50 specific insecticides, technical toxaphene was associated with
increased ORs of self-reported hypothyroid disease, showing an association at low exposure
(based on intensity-weighted cumulative days of use), but not high exposure (Goldner et al..
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2013). When 1 1 moderately correlated pairs of pesticides were analyzed by logistic regression
models in this study, the association between technical toxaphene and hypothyroidism remained
statistically significant, and associations with the other pesticides and hypothyroid disease were
reduced. Evidence from animal studies is coherent with the human study indicating effects in the
thyroid. In animal studies, increased incidences of a variety of histopathologic lesions were
observed in the thyroid of both male and female rats across different life stages (i.e., in adult F0
and F1 animals) following subchronic and chronic dietary exposure to low doses
(HEDs = 0.10-0.47 mg/kg-day) of technical toxaphene (Choi et aL 1988; Cfau et aL 1986).
Additionally, increased incidences of thyroid histopathology as well as increased TSH serum
levels were observed in a short-term-duration (28-day) gavage study in rats, albeit at a
significantly higher dose (HED = ~19 mg/kg-day) (Waritz et aL 1996). Finally, the NCI (1979)
chronic-duration study found elevated incidences of thyroid tumors in technical toxaphene
exposed rats, although no elevated incidences of non-neoplastic lesions were observed in any
tissue in the exposed groups (NCI, 1979).
To provide a common basis for comparing potential points of departure (PODs) and
critical effects for a subchronic p-RfD for technical toxaphene (i.e., comparing benchmark dose
[BMD] and benchmark dose lower confidence limits [BMDLs] among the most sensitive
endpoints), data sets from studies in Table 5A with multiple exposure levels for liver, kidney,
thyroid, and immune effects were selected for BMD analysis. In addition, three liver-endpoint
data sets were selected from the most recent repeated-dose oral toxicity study in mice for
comparison purposes [AVanu et al. (2015); relative liver weight, serum ALT, and liver cell
proliferation rates]. Details regarding modeling procedures and detailed BMD modeling results
are presented in Appendix C. For BMD modeling of several of the selected data sets for liver,
kidney, and thyroid lesions observed in rat studies by Chu et al. (1988) and Cfau et al. (1986). the
sum of incidences of minimal-to-mild and moderate-to-severe severity grades were statistically
significantly increased at the lowest dose tested. However, many of these data sets were difficult
to model primarily because the responses at the lowest exposure levels for each of these data sets
were considered far in excess of the benchmark response (BMR), or had near maximal
responses, leaving no data to inform the shape of the dose-response curve in the low-dose region
and requiring extrapolation far below the observable range in order to estimate the BMD. Thus,
the resultant BMDs and BMDLs for several endpoints were considered unreliable, and LOAELs
or NOAELs were selected as PODs for many of the Cfau et al. (1986) and Cfau et al. (1988) rat
data sets to compare with potential PODs from the other selected data sets.
Table 9 summarizes the BMD modeling results for endpoints associated with each of the
aforementioned sensitive toxicity targets following exposure to technical toxaphene to identify
potential PODs (expressed as HEDs) for the derivation of the subchronic p-RfD.
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Table 9. Candidate Principal Studies and PODs for the Derivation of the Subchronic p-RfD
Endpoint/Reference
NOAEL (HED)a
(mg/kg-d)
LOAEL (HED)a
(mg/kg-d)
BMDL (HED)a
(mg/kg-d)
Selected POD (HED)
(mg/kg-d)
Comments
Immune Effects
Decreased mean primary anti-SRBC IgM
response in female cynomolgus monkeys (1-wk
Dostimmunization). 75 wk (Trvohonas et al..
200 nb
0.05
0.2
0.02
0.02 (BMDLisd)
NA
Kidney Effects
Primary tubular injury (all severity grades) in the
kidnev of male S-D rats. 13 wk (Chi et al.. 1986)c
0.090
0.46
0.020
0.020 (BMDLio)
NA
Tubular necrosis (all severity grades) in the
kidnev of male S-D rats. 13 wk (Chi et al.. 1986)°
0.090
0.46
0.045
0.045 (BMDLio)
NA
Tubular necrosis (only minimal to mild observed)
in the kidnev of female S-D rats. 13 wk (Chu et
al.. 1986)°
NDr
0.11
0.0012
0.11 (LOAEL)
Model fit obtained after
dropping 2 high doses, but
not considered reliable due
to lack of data point near the
BMR (see Appendix C)
Primary tubular injury (all severity grades) in the
kidnev of FO male S-D rats. 25-29 wk (Chu et al..
1988)d
NDr
0.10
DUB
0.10 (LOAEL)
No models provided
adequate fit to data; lacks
dose-response
Primary tubular injury (all severity grades) in the
kidnev of FO female S-D rats. 25-29 wk (Chu et
al.. 1988V1
0.083
0.44
0.088
0.088 (BMDLio)
NA
Thyroid Effects
Reduced colloid density (moderate to severe) in
the thvroid of male S-D rats. 13 wk (Chu et al..
1986)°
0.090
0.46
0.013
0.013 (BMDLio)
NA
Reduced colloid density (all severity grades) in
the thvroid of female S-D rats. 13 wk (Chu et al..
1986)°
NDr
0.11
0.00051
0.11 (LOAEL)
Model fit obtained, but not
considered reliable
(see Appendix C)
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Table 9. Candidate Principal Studies and PODs for the Derivation of the Subchronic p-RfD
Endpoint/Reference
NOAEL (HED)a
(mg/kg-d)
LOAEL (HED)a
(mg/kg-d)
BMDL (HED)a
(mg/kg-d)
Selected POD (HED)
(mg/kg-d)
Comments
Reduced colloid density (all severity grades) in
the thvroid of Fla female S-D rats. 34 wk (Chu et
aL 1988V1
NDr
0.089
0.0075
0.089 (LOAEL)
Model fit obtained after
dropping high dose, but not
considered reliable
(see Appendix C)
Colloid inspissation (all severity grades) in the
thvroid of FO male S-D rats. 25-29 wk (Chu et
al.. 1988)d
NDr
0.10
0.0050
0.10 (LOAEL)
Model fit obtained after
dropping 2 high doses, but
not considered reliable
(see Appendix C)
Colloid inspissation (all severity grades) in the
thvroid of Fla male S-D rats. 34 wk (Chu et al..
1988)d
NDr
0.078
0.0089
0.078 (LOAEL)
Model fit obtained after
dropping high dose, but not
considered reliable
(see Appendix C)
Reduced follicular size (all severity grades) in the
thvroid of female S-D rats. 13 wk (Chu et al..
1986)°
NDr
0.11
0.0052
0.11 (LOAEL)
Model fit obtained, but not
considered reliable
(see Appendix C)
Follicle collapse/angularity (minimal to mild
observed) in the thyroid of Fla male S-D rats,
34 wk (Chu et al.. 1988Y1
0.078
0.37
0.014
0.014 (BMDLio)
Model fit obtained after
dropping 2 high doses
Increased epithelial height (all severity grades) in
the thvroid of female S-D rats. 13 wk (Chu et al..
1986)°
NDr
0.11
0.00014
0.11 (LOAEL)
Model fit obtained, but not
considered reliable
(see Appendix C)
Cytoplasmic vacuolation (all severity grades) in
the thvroid of female S-D rats. 13 wk (Chu et al..
1986)°
NDr
0.11
DUB
0.11 (LOAEL)
No models provided
adequate fit to data
Cytoplasmic vacuolation (all severity grades) in
the thyroid of FO male S-D rats, 25-29 wk
(Chu et al.. 1988)d
0.10
0.47
0.0092
0.0092 (BMDLio)
Model fit obtained after
dropping 2 high doses
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Table 9. Candidate Principal Studies and PODs for the Derivation of the Subchronic p-RfD
Endpoint/Reference
NOAEL (HED)a
(mg/kg-d)
LOAEL (HED)a
(mg/kg-d)
BMDL (HED)a
(mg/kg-d)
Selected POD (HED)
(mg/kg-d)
Comments
Cytoplasmic vacuolation (all severity grades) in
the thyroid of FO female S-D rats. 25-29 wk (Chu
etaL 1988V1
0.44
1.9
0.037
0.037 (BMDLio)
Model fit obtained after
dropping high dose, but
considered a borderline case
for passing visual inspection
of the model fit
Cytoplasmic vacuolation (only minimal to mild
observed) in the thyroid of Fla female S-D rats,
34 wk (Chu et al.. 1988)d
NDr
0.089
0.0059
0.089 (LOAEL)
Model fit obtained after
dropping high dose, but not
considered reliable
(see Appendix C)
Liver Effects
Increased relative liver weight (>10%) in female
Beagle doss. 13 wk (Chu et al.. 1986)°
NDr
0.1
DUB
0.1 (LOAEL)
No models provided
adequate fit to data
Increased absolute liver weight (>10%) in
FO female S-D rats. 25-29 wk (Chu et al.. 1988)d
NDr
0.083
0.028
0.028 (BMDLio)
Model fit obtained after
dropping high dose
Increased relative liver weight (>10%) in male
B6C3Fi mice: 28 d (Wang et al.. 2015)
0.1
0.85
DUB
0.1 (NOAEL)
The fit of the nonconstant
variance model provided
only a marginal fit (variance
p = 0.09)
Increased BrdU labeling index in liver of male
B6C3Fi mice: 28 d (Wang et al.. 2015)
0.1
0.85
DUB
0.1 (NOAEL)
No models provided
adequate fit to data
Accented zonation (all severity grades) in the liver
of female S-D rats. 13 wk (Chu et al.. 1986)°
0.11
0.58
0.024
0.11 (NOAEL)
Model fit obtained, but not
considered reliable
(see Appendix C)
Anisokaryosis (all severity grades) in the liver of
male S-D rats. 13 wk (Chu et al.. 1986)°
0.09
0.46
0.0090
0.09 (NOAEL)
Model fit obtained, but not
considered reliable
(see Appendix C)
Anisokaryosis (all severity grades) in the liver of
female S-D rats. 13 wk (Chu et al.. 1986)°
NDr
0.11
0.0030
0.11 (LOAEL)
Model fit obtained, but not
considered reliable
(see Appendix C)
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Table 9. Candidate Principal Studies and PODs for the Derivation of the Subchronic p-RfD
Endpoint/Reference
NOAEL (HED)a
(mg/kg-d)
LOAEL (HED)a
(mg/kg-d)
BMDL (HED)a
(mg/kg-d)
Selected POD (HED)
(mg/kg-d)
Comments
Anisokaryosis (all severity grades) in the liver of
F0 female S-D rats. 25-29 wk (Chi et al.. 1988)d
NDr
0.083
0.0072
0.083 (LOAEL)
Model fit obtained after
dropping 2 high doses, but
not considered reliable
(see Appendix C)
Cytoplasmic homogeneity (all severity grades) in
the liver of Fla male S-D rats. 34 wk (Chu et al..
1988)d
NDr
0.078
0.0039
0.078 (LOAEL)
Model fit obtained after
dropping 2 high doses, but
not considered reliable
(see Appendix C)
'Following U.S. EPA (2011b) guidance, animal doses from candidate principal studies were converted to HEDs through the application of a DAF. DAFs for
each dose are calculated as follows: DAF = (BWa1/4 BWt1'4), where BWa = animal body weight and BWh = human body weight. For all DAF calculations, a
reference human body weight (BWh) of 70 kg (U.S. EPA. 1988b) was used.
bDAFs were calculated using study-specific body weight (B\V) data for female cynomolgus monkeys at the initiation of treatment with technical toxaphene.
Tor the rat portion of the study, DAFs were calculated using body weights (B Wa) estimated from study-specific initial weight and weight-gain data for female
S-D rats. For the dog portion of the study, DAFs were calculated using reference body weights (B\V) for female Beagle dogs following subchronic exposure
(U.S. EPA. 1988b).
dDAFs were calculated using body weights (B Wa) estimated from study-specific initial weight and weight-gain data for male and female S-D rats.
eDAFs were calculated using the mean of study-specific initial and terminal body weights (BWa) of male B6C3Fi mice.
BMD = benchmark dose; BMDLio = 10% benchmark dose lower confidence limit; BMR = benchmark response; BrdU = bromodeoxyuridine; BW = body
weight; DAF = dosimetric adjustment factor; DUB = data unsuitable for BMD modeling (see Appendix C for details); HED = human equivalent dose;
IgM = immunoglobulin M; LOAEL = lowest-observed-adverse-effect level; NA = not applicable; NDr = not determined; NOAEL = no-observed-adverse-effect
level; POD = point of departure; p-RfD = provisional reference dose; S-D = Sprague-Dawley; SD = standard deviation; SRBC = sheep red blood cell.
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In Recommended Use of Body Weight4 as the Default Method in Derivation of the Oral
Reference Dose (U.S. EPA. 2011b). the Agency endorses a hierarchy of approaches to derive
human equivalent oral exposures from data from laboratory animal species, with the preferred
approach being physiologically based toxicokinetic modeling. Other approaches may include
using some chemical-specific information, without a complete physiologically based
toxicokinetic model. In the absence of chemical-specific models or data to inform the derivation
of human equivalent oral exposures, U.S. EPA endorses BW3/4 as a default to extrapolate
toxicologically equivalent doses of orally administered agents from all laboratory animals to
humans for the purpose of deriving an oral reference dose (RfD) under certain exposure
conditions. More specifically, the use of BW3 4 scaling for deriving an RfD is recommended
when the observed effects are associated with the parent compound or a stable metabolite, but
not for portal-of-entry effects. A validated human physiologically based pharmacokinetic
(PBPK) model for technical toxaphene is not available for use in extrapolating doses from
animals to humans. In addition, effects in the liver, kidney, thyroid, and immune system are not
portal-of-entry effects. Therefore, scaling by BW3/4 is relevant for deriving HED for this effect.
From the identified sensitive targets of technical toxaphene toxicity, the thyroid exhibits
consistency and coherence in the evidence across species (including humans), sexes, life stages,
and endpoints. The three lowest potential PODs (HEDs) providing reliable, adequate model fits
are all for thyroid endpoints from subchronic- and chronic-duration studies in S-D rats and
include: (1) a 10% BMDL (BMDLio) of 0.0092 mg/kg-day for cytoplasmic vacuolation (all
severity grades) in the thyroid of F0 males following 25-29 weeks of dietary exposure (Chu et
ai. 1988). (2) a BMDLio of 0.013 mg/kg-day for reduced colloid density (moderate to severe) in
the thyroid of males following 13 weeks of dietary exposure (Chu et ai, 1986), and (3) a
BMDLio of 0.014 mg/kg-day for follicle collapse/angularity (only minimal to mild observed) in
the thyroid of F1 a males following 34 weeks of dietary exposure (Chu et ai, 1988). In the
subchronic-duration (13-week) rat study by (Chu et ai. 1986), moderate to severe reduced
colloid density appears to be the most sensitive effect. When all severity grades are considered,
however, a high control incidence in males precluded BMD modeling and subsequent
identification of a reliable POD. With respect to thyroid effects following chronic exposure,
cytoplasmic vacuolation and reduced colloid density are comparably sensitive in F0 males l"Cfau
et ai (1988); see Table B-14], Therefore, when considering all the BMDL estimates presented in
Table 9, 0.0092 mg/kg-day for cytoplasmic vacuolation (all severity grades) in the thyroid of
F0 males following 25-29 weeks of dietary exposure is considered the most reliable and the
most sensitive (see Appendix C).
Chu et ai (1988) is, therefore, chosen as the principal study, and cytoplasmic vacuolation
(all severity grades) in the thyroid of F0 male S-D rats following 25-29 weeks of dietary
exposure is the critical effect, with a BMDLio (HED) of 0.0092 mg/kg-day as the POD. As
noted in Table 9, this BMDLio was estimated after dropping the two highest doses, which
resulted in a significant improvement of the model fit to the low-dose range. Although this
BMDLio is approximately 10-fold lower than the identified NOAEL of 0.10 mg/kg-day for this
endpoint, there is a 30% (3/10) response at the NOAEL. Thus, due to the animal sample size
examined at 0.10 mg/kg-day, the estimated BMDLio is considered an appropriate POD for this
endpoint. Based on all of the laboratory animal oral toxicity data, this POD is expected to be
protective against all thyroid effects (regardless of severity), as well as any potential liver,
kidney, and immune effects observed in other studies and across other species (including those
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effects for which only LOAELs could be identified) following subchronic or chronic exposure to
technical toxaphene.
The subchronic p-RfD for technical toxaphene is derived as follows:
Subchronic p-RfD = BMDLio (HED) UFc
for Technical Toxaphene = 0.0092 mg/kg-day 30
= 3 x 10"4 mg/kg-day
Table 10 summarizes the uncertainty factors for the subchronic p-RfD for technical
toxaphene.
Table 10. Uncertainty Factors for the Subchronic p-RfD for Technical Toxaphene
UF
Value
Justification
UFa
3
A UFa of 3 (10°5) is applied to account for uncertainty in characterizing the toxicokinetic or
toxicodynamic differences between rats and humans following technical toxaphene exposure. The
toxicokinetic uncertainty has been accounted for by calculating an HED through application of a
DAF as outlined in the U.S. EPA's Recommended Use of Body Weight4 as the Default Method in
Derivation of the Oral Reference Dose ('U.S. EPA. 20 lib).
UFd
1
A UFd of 1 is applied because the oral exposure database contains numerous short-term- and
subchronic-duration toxicity studies of laboratory animals, a one-generation reproductive toxicity
studv of rats (Chi et al.. 19881 multiple-eeneration reproductive toxicity studies in rats (Kennedy
et al.. 1973) and mice (Kcnlinecr et al.. 19701 and several developmental toxicity studies in rats
(Chemoff et al.. 1990; Crowder et al. 1980; Olson et al.. 1980; Chemoff and Carver. 1976) and
mice (Allen et al.. 1983; Chemoff and Kavlock. 1983; Chemoff and Carver. 1976). Analysis of the
database indicates that the effects on reproductive function and early development in rats and mice
occurred at higher exposure levels than the lowest exposure levels resulting in thyroid, liver,
kidney, or immune effects (see Table 5A).
UFh
10
A UFh of 10 is applied for intraspecies variability to account for human-to-human variability in
susceptibility in the absence of quantitative information to assess the toxicokinetics and
toxicodynamics of technical toxaphene in humans.
UFl
1
A UFl of 1 is applied for LOAEL-to-NOAEL extrapolation because the POD is a BMDL.
UFS
1
A UFS of 1 is applied because the principal study is greater than subchronic duration (25-29 wk).
UFC
30
Composite UF = UFA x UFD x UFH x UFL x UFS.
BMDL = benchmark dose lower confidence limit; DAF = dosimetric adjustment factor; HED = human equivalent
dose; LOAEL = lowest-observed-adverse-effect level; NOAEL = no-observed-adverse-effect level; POD = point of
departure; p-RfD = provisional reference dose; UF = uncertainty factor; UFa = interspecies uncertainty factor;
UFc = composite uncertainty factor; UFD = database uncertainty factor; UFH = intraspecies uncertainty factor;
UFl = LOAEL-to-NOAEL uncertainty factor; UFS = subchronic-to-chronic uncertainty factor.
Confidence in the subchronic p-RfD for technical toxaphene is high as explained in
Table 11.
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Table 11. Confidence Descriptors for the Subchronic p-RfD for Technical Toxaphene
Confidence Categories
Designation
Discussion
Confidence in study
H
Confidence in the principal study is high because the study was
published in a peer-reviewed journal and is of quality study
design and performance with regard to examining potential
toxicity endpoints and in presenting materials, methods, and
results. The number of dose groups (4 plus control for males and
females) and group sizes (10-13 males, 10-17 females examined
histologically) were adequate, thus allowing reliable evaluation of
the endpoints investigated and identification of treatment-related
changes in relation to dose. Details of the study are provided in
the "Review of Potentially Relevant Data" section.
Confidence in database
H
There is high confidence in the oral toxicity database because
there are numerous short-term and subchronic-duration toxicity
studies, including studies in monkeys, dogs, and rodents, and
several reproductive and developmental toxicity studies.
Confidence in subchronic p-RfD:i
H
Overall confidence in the subchronic p-RfD is high.
aThe overall confidence cannot be greater than the lowest entry in the table (high).
H = high; p-RfD = provisional reference dose.
Derivation of a Chronic Provisional Reference Dose
The subchronic p-RfD based on cytoplasmic vacuolation (all severity grades) in the
thyroid of F0 male S-D rats following 25-29 weeks of dietary exposure (Chu et aL 1988) is
selected as the basis of the chronic p-RfD for technical toxaphene.
Justification
In the only available chronic-duration animal toxicity bioassay that exposed animals for a
longer duration than the Chu et al. (1988) study and also provided comprehensive examinations
of tissues for non-neoplastic lesions, NCI (1979) did not find significantly elevated incidences of
non-neoplastic lesions (including identified sensitive targets of thyroid, liver, and kidney) in rats
or mice exposed for 80 weeks to average daily doses of technical toxaphene as high as 83.29 and
34.2 mg/kg-day, respectively. However, with respect to thyroid toxicity, it is important to note
that the NCI (1979) study tested a different rat strain (Osborne-Mendel) than did either the
subchronic- or chronic-duration studies by Chu and colleagues (which tested S-D rats).
Nevertheless, there was a 3-12% response (albeit not statistically significant) in noncancer
thyroid lesions in males and females, and there was a significant increase in thyroid tumors in
both sexes, further establishing the thyroid as a target of technical toxaphene following chronic
exposure. Furthermore, the Chu et al. (1988) and NCI (1979) chronic-duration studies did not
examine endpoints of immune suppression, which is also a sensitive noncancer effect identified
in the oral database for technical toxaphene. Thus, in the absence of chronic-duration data on
immune effects, application of an additional database uncertainty factor (UFd) is warranted to
account for uncertainty in identifying potentially more sensitive immune effects following
chronic exposure.
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The chronic p-RfD for technical toxaphene, based on the BMDLio (HED) of
0.0092 mg/kg-day for cytoplasmic vacuolation (all severity grades) in the thyroid of F0 male
S-D rats following 25-29 weeks of dietary exposure (Chu et ai, 1988), is derived as follows:
Chronic p-RfD = BMDLio (HED) UFc
for Technical Toxaphene = 0.0092 mg/kg-day 100
= 9 x 10"5 mg/kg-day
Table 12 summarizes the uncertainty factors for the chronic p-RfD for technical
toxaphene.
Table 12. Uncertainty Factors for the Chronic p-RfD for Technical Toxaphene
UF
Value
Justification
UFa
3
A UFa of 3 (10°5) is applied to account for uncertainty in characterizing the toxicokinetic or
toxicodynamic differences between rats and humans following TT exposure. The toxicokinetic
uncertainty has been accounted for by calculating an HED through application of a DAF as outlined
in the U.S. EPA's Recommended Use of Body Weight3''4 as the Default Method in Derivation of the
Oral Reference Dose (U.S. EPA, 2011b).
UFd
3
A UFd of 3 is applied because the oral exposure database contains numerous short-term- and
subchronic-duration toxicity studies of laboratory animals, 2 chronic-duration toxicity studies of
rats and mice that included comprehensive histological examination of most tissues, a
one-eeneration reproductive toxicitv studv of rats (Chu et al.. 1988s). multiple-eeneration
reproductive toxicitv studies in rats (kennedv et al.. 1973) and mice (Keolineer et al.. 1970). and
several developmental toxicitv studies in rats (Oiernoff et al.. 1990; Crowder et al. 1980; Olson et
al.. 1980; Chemoff and Carver. 1976) and mice (Allen et al.. 1983; Chemoff and Kavlock. 1983;
Oiernoff and Carver. 1976). Analvsis of the database indicates that the effects on reproductive
function and early development in rats and mice occurred at higher exposure levels than the lowest
exposure levels resulting in thyroid, liver, kidney, or immune effects (see Table 5A). However, an
additional uncertainty factor is warranted to account for uncertainty in identifying potentially more
sensitive immune effects following chronic exposure.
UFh
10
A UFh of 10 is applied for intraspecies variability to account for human-to-human variability in
susceptibility in the absence of quantitative information to assess the toxicokinetics and
toxicodynamics of TT in humans.
UFl
1
A UFl of 1 is applied for LOAEL-to-NOAEL extrapolation because the POD is a BMDL.
UFS
1
A UFS of 1 is applied because the principal study is of chronic duration (25-29 wk).
UFC
100
Composite UF = UFA x UFD x UFH x UFL x UFS.
BMDL = benchmark dose lower confidence limit; DAF = dosimetric adjustment factor; HED = human equivalent
dose; LOAEL = lowest-observed-adverse-effect level; NOAEL = no-observed-adverse-effect level; POD = point of
departure; p-RfD = provisional reference dose; TT = technical toxaphene; UF = uncertainty factor;
UFa = interspecies uncertainty factor; UFC = composite uncertainty factor; UFD = database uncertainty factor;
UFh = intraspecies uncertainty factor; UFL = LOAEL-to-NOAEL uncertainty factor; UFS = subchronic-to-chronic
uncertainty factor.
Confidence in the chronic p-RfD for technical toxaphene is medium as explained in
Table 13.
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Table 13. Confidence Descriptors for the Chronic p-RfD for Technical Toxaphene
Confidence Categories
Designation
Discussion
Confidence in study
H
Confidence in the principal study is high because the study was
published in a peer-reviewed journal, and is of quality study design
and performance with regard to examining potential toxicity
endpoints and in presenting materials, methods, and results. The
number of dose groups (4 plus control for males and females) and
group sizes (10-13 males, 10-17 females examined histologically)
were adequate, thus allowing reliable evaluation of the endpoints
investigated and identification of treatment related changes in
relation to dose. Details of the study are provided in the "Review of
Potentially Relevant Data" section.
Confidence in database
M
Confidence in the oral toxicity database is medium because there are
numerous short-term- and subchronic-duration toxicity studies, as
well as chronic-duration toxicity studies, and several reproductive
and developmental toxicity studies, but no dose-response data for
immune suppression endpoints (a sensitive target identified in
subchronic-duration studies) following chronic exposure.
Confidence in chronic p-RfDa
M
Overall confidence in the chronic p-RfD is medium.
aThe overall confidence cannot be greater than the lowest entry in the table (medium).
H = high; M = medium; p-RfD = provisional reference dose.
DERIVATION OF INHALATION REFERENCE CONCENTRATIONS
Derivation of a Subchronic Provisional Reference Concentration
Appropriate data to derive a subchronic provisional reference concentration (p-RfC) for
technical toxaphene, weathered toxaphene, or individual toxaphene congeners has not been
identified.
Derivation of a Chronic Provisional Reference Concentration
Appropriate data to derive a chronic p-RfC for technical toxaphene, weathered
toxaphene, or individual toxaphene congeners has not been identified.
CANCER WEIGHT-OF-EVIDENCE DESCRIPTOR AND POTENCY VALUES
A cancer OSF of 1.1 (mg/kg-day) 1 and IUR of 3.2 x 10 4 (|ig/m3) 1 for technical
toxaphene is currently listed on IRIS (U.S. EPA. 1988a). along with a cancer WOE classification
of Group B2 based on sufficient evidence of carcinogenicity in laboratory animals
(hepatocellular tumors in mice and thyroid tumors in rats) and no studies in humans at the time
of the cancer assessment. The slope factor was based on incidence data for hepatocellular
carcinomas and neoplastic nodules in male B6C3Fi mice in the bioassay conducted by Litton
Bionetics (1978), using a linearized multistage modeling procedure.
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APPENDIX A. SCREENING PROVISIONAL VALUES
For the reasons noted in the main Provisional Peer-Reviewed Toxicity Value (PPRTV)
document, toxicity data for weathered toxaphene or individual toxaphene congeners are
inadequate to derive noncancer provisional toxicity values or assess the carcinogenicity of these
substances. However, information is available for this chemical, which although insufficient to
support deriving a provisional toxicity value under current guidelines, may be of limited use to
risk assessors. In such cases, the Superfund Health Risk Technical Support Center summarizes
available information in an appendix and develops a "screening value." Appendices receive the
same level of internal and external scientific peer review as the main documents to ensure their
appropriateness within the limitations detailed in the document. Users of screening toxicity
values in an appendix to a PPRTV assessment should understand that there is considerably more
uncertainty associated with deriving an appendix screening toxicity value than for a value
presented in the body of the assessment. Questions or concerns about the appropriate use of
screening values should be directed to the Superfund Heath Risk Technical Support Center.
OVERVIEW OF AVAILABLE TOXICITY DATA ON WEATHERED TOXAPHENE
AND TOXAPHENE CONGENERS
The chemical composition of weathered toxaphene mixtures can vary widely depending
on time in the environment and environmental conditions. Known differences in composition
between technical toxaphene and various types of weathered toxaphene could potentially lead to
qualitative or quantitative differences in toxicity. Comparative toxicity studies of technical
toxaphene and weathered toxaphene samples are rare, so the degree to which weathered
toxaphene samples may differ in toxicity from technical toxaphene is uncertain. Toxaphene
residues purified from lake trout samples were reportedly similar to technical toxaphene in
toxicity to mosquito larvae (Gooch and Matsumura. 1987). However, a weathered toxaphene
sample extracted from the livers of cod (cod liver extract [CLE]) exposed to technical toxaphene
was eightfold more potent than ultraviolet (UV)-weathered technical toxaphene or technical
toxaphene in an in vitro assay of gap junctional intercellular communication (GJIC) in mouse
Hepa 1 c 1 c7 cells (Besselink et al., 2008). In a recent in vitro study examining GJIC inhibition in
B6C3Fi mouse hepatocyte cultures, toxaphene congeners and congener mixtures exhibited
comparable or lower potency as that observed for technical toxaphene Kerger et al. (2018). A
synthetic mixture of chlorinated camphenes with very low amounts of congeners in the hepta- to
nona-chlorinated range was 1.3- and 2.3-fold more potent in producing lethal and nonlethal
malformations, respectively, in zebrafish embryos than a synthetic mixture with a compositional
profile similar to technical toxaphene, which had high amounts of congeners in the hepta- to
nona-chlorinated range (Kapp et al.. 2006). A more detailed overview of the limited toxicity
data on samples of weathered toxaphene and toxaphene congeners, as well as considerations for
deriving screening provisional toxicity values for oral exposure to these compounds, follows.
Only one in vivo study, described in several reports (Besselink et al.. 2008; Besselink et
al.. 2000; EU. 2000). has been identified that has any relevance to deriving a screening
provisional reference dose (p-RfD) for weathered toxaphene. In a two-stage liver tumor
initiation/promotion study involving subcutaneous (s.c.) injection, the effects of technical
toxaphene, UV-weathered toxaphene (i.e., UV-irradiated technical toxaphene), and biologically
weathered toxaphene (a liver extract from cod fish fed technical toxaphene) were examined.
Groups of partially hepatectomized female Sprague-Dawley (S-D) rats obtained from
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M0llegaard Breeding Centre in Copenhagen, Denmark were administered the technical and
weathered toxaphene samples once weekly by s.c. injection for 20 weeks after a single
intraperitoneal (i.p.) injection of a tumor-initiating dose of iV-nitrosodiethylamine (NDEA)
(Besselink et aL 2008). To generate the CLE, cod were given toxaphene-enriched feed
(containing 30 ppm technical toxaphene) in their diet for 63 days, followed by a 14-day
postdosing period with untreated feed to permit appropriate metabolism. Approximately 30 kg
of liver enriched in toxaphene residues was harvested from 300 cod for preparing a biologically
weathered toxaphene extract. The rat treatment groups were tested in two experimental
scenarios with rats receiving four different doses. Doses administered to rats via s.c. injection in
Experiment 1 (a comparison of technical toxaphene with UV-weathered toxaphene) were: 0
(corn oil control; n = 14), 0.62, 2, 6, or 18 mg technical toxaphene/kg body weight per week
(n = 10), and 0.3, 0.89, 2.67, or 8 mg UV-weathered toxaphene/kg body weight per week
(n = 10). Doses in Experiment 2 (a comparison of technical toxaphene with CLE) were: 0 (corn
oil control; n = 14), 18 mg technical toxaphene/kg body weight per week (n = 10), and 0.46,
1.39, 4.17, or 12.5 mg CLE/kg body weight per week (n = 10). Both experiments also included a
positive control group that was administered 2,3,7,8-tetrachlorodibenzodioxin (TCDD) (1 |ig/kg
body-weight injection per week; n= 10). One week after the last dosing, the animals were
weighed, blood was collected, and the livers removed and weighed after sacrifice. For each
treatment group, three endpoints evaluating the occurrence of altered liver foci positive for
glutathione-S-transferase-P (number of foci per cm2 of liver, mean foci area [mm2], and area
fraction of liver with foci) were used as measures of liver tumor promotion activity. Serum
alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity were also
measured.
The CLE sample had a markedly different compositional profile of toxaphene congeners
than the technical toxaphene fed to the cod. Compared with technical toxaphene, the CLE
sample had decreased relative percentages of octa- and nona-chlorinated congeners like p-26,
p-32, p-50, and p-62, and increased relative percentages of unidentified lower chlorinated
congeners. The summed concentration of p-26, p-50, and p-62 (dubbed £3PC, approximate
fractional composition of 1:2:2) accounted for <1% of total toxaphene in CLE, which is
significantly lower than the mean £3PC percentages of 22.45% in other examples of biologically
measured toxaphene, specifically 29 tissue biotic samples from several northern latitude species
of fish (e.g., trout, salmon, and burbot) and aquatic mammals (narwhal, beluga whale, walrus),
and 4.47% in >50 aquatic tissue samples from a U.S. Superfund site in Georgia (including blue
crab, red and black drum, croaker, mullet, sea trout, shrimp, spot, yellowtail, and whiting)
[see Simon and Manning (2006) for references of these monitoring studies]. The approximate
fractional composition of p-26, p-50, and p-62 in the UV-weathered toxaphene test substance
was 1:1:2.
Mean values for relative liver weight were statistically significantly increased, compared
with controls, in TCDD-treated positive control groups in both experiments. Means for
body-weight gain, relative liver weight, and relative thymus weight were not significantly
different between vehicle control groups and groups exposed to technical toxaphene,
UV-weathered toxaphene, or CLE. No changes in ALT or AST activity were observed in groups
exposed to technical toxaphene, UV-weathered toxaphene, or CLE. TCDD, however, caused an
increase in AST activity. Mean values for tumor-promotion endpoints in the positive control
groups were statistically significantly increased (about two- to fourfold depending on the
endpoint), compared with vehicle controls, but no statistically significant increases in
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tumor-promotion endpoints were found in any groups exposed to the toxaphene mixtures,
compared with controls. A statistically significantly lower value for the number of foci per cm2
liver was found in the highest CLE-treated dose group, compared with controls. The other
measures of altered foci were also lower in the highest CLE-treated dose group than in the
control group, but the differences were not statistically significant. Besselink et al. (2008)
discussed the hypothesis that this may have been caused by an apoptotic effect leading to
enhanced liver cell death rates at the highest dose. The highest doses used in this study for
technical toxaphene, UV-weathered toxaphene, and CLE are s.c., sub chronic
no-observed-adverse-effect levels (NOAELs) for liver tumor promotion activity.
Besselink et al. (2008) also collected data on in vitro inhibition of GJIC in cultured
mouse Hepalclc7 cells. Twenty percent effective concentration (EC20) values in the in vitro test
of inhibition of intercellular communication indicated the following potency order: CLE
(0.24 |ig/mL) > UV-weathered = technical toxaphene (1.9 (ag/m L) (albeit CLE was 77,000 times
less potent than the positive control, TCDD). The GJIC results indicate that CLE may be about
eightfold more potent than technical toxaphene or UV-weathered toxaphene, at least for this
particular in vitro endpoint. In a recent study that also examined the effects of toxaphene on
GJIC in vitro, Kerger et al. (2018) treated hepatocytes isolated and cultured from male and
female B6C3Fi mice with increasing concentrations of technical toxaphene, individual
toxaphene congeners (p-26, p-50, and p-62), Hx-Sed, and two congener mixtures
(Mixture 1 = p-26, p-50, and p-62 [2:1:1 fractional composition], and Mixture 2 = p-26, p-50,
p-62, and Hx-Sed [2:1:1:1 fractional composition]) for 3 or 24 hours. Phenobarbital was used as
a positive control. The study authors state that compared to untreated control cells, the lowest
concentration of technical toxaphene that produced a significant change in GJIC inhibition
(dubbed the lowest significant effect level [LSEL]) was generally comparable to that observed
for the congeners and mixtures. The LSEL for Hp-Sed was three- to fivefold higher than for
technical toxaphene, while all other treatment groups were within twofold. The male hepatocyte
responses for p-26, Hx-Sed, and Mixture 1 were not significantly different than untreated
controls at any concentration tested. After modeling selected in vitro dose-response data, the
predicted EC20 values for technical toxaphene, toxaphene congeners, and toxaphene mixtures
exhibited the following potency order: p-50 (2.74 (ag/m L) > technical toxaphene
(3.50 |ig/mL) > Hx-Sed (4.06 |ig/mL) > p-62 (4.15 |ig/mL) > p-26 (7.93 |ig/mL) > Hp-Sed
(25.1 |ig/mL) > Mixture 2 (31.9 |ig/mL) > Mixture 1 (32.1 |ig/mL). The EC20 value for technical
toxaphene was within twofold of the EC20 values for technical toxaphene and UV-weathered
toxaphene observed by Besselink et al. (2008) in Hepa 1 c 1 c7 cells. Additionally, Kerger et al.
(2018) suggest that an interaction index of <1, 1, or >1 indicates possible antagonism, additivity,
or synergy, respectively, of components in the congener mixtures measured at the EC20, and the
interaction index of the two congener mixtures was approximately half that expected based on
pure additive responses at the EC20, in contrast to the apparent synergistic response suggested by
the CLE in vitro GJIC data (Besselink et al.. 2008). Taken together, the results of the Besselink
et al. (2008) and Kerger et al. (2018) in vitro studies indicate that relative toxic properties of
technical toxaphene, toxaphene congeners, congener mixtures, or weathered toxaphene can be
dependent on endpoint or test systems, as well as specific substances tested.
The literature search also identified studies of cultured cells (human breast cancer cell
model) (Stel/.er and Chan. 1999). cultured rat embryos (Calciu et al.. 2002: Calciu et al.. 1997).
zebrafish embryos (Kapp et al.. 2006). mosquito larvae (Gooch and Matsumura. 1987). juvenile
rats (Olson et al.. 1980). and mice (Turner et al.. 1977) comparatively exposed to individual
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toxaphene congeners, toxaphene mixtures, or "weathered" toxaphene mixtures (see Table A-l).
Overall, the available information on the toxicity of individual toxaphene congeners is too sparse
to propose a component-based approach (such as the toxic equivalency factor approach for
dioxins and dioxin-like compounds, or the relative potency factor approach for nonsubstituted
polycyclic aromatic hydrocarbons [PAHs]) for assessing health risks from mixtures of toxaphene
congeners, but the available information indicates that individual congeners may differ in toxic
properties. In addition, the results from the most recent of these studies (Kapp et aL 2006)
indicate that mixtures enriched in low-chlorinated congeners (penta- and hexa-chlorinated
congeners) may be more toxic to zebrafish embryos than toxaphene mixtures, like technical
toxaphene, that are more enriched in congeners with >7 chlorines.
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Table A-l. Studies Featuring Comparative Testing of Toxaphene Congeners and Mixtures
Study Description
Reference
Intraperitoneal LD5o values for TT were equivalent in mice with or without a PB pretreatment (42 and 47 mg/kg), whereas LD5o values for
"Toxicant B" (2,2,5-endo,6-exo,8,9,10-heptachlorobornane, or p-32) were higher in mice without pretreatment (75 mg/kg, less toxic) and lower
with pretreatment (9.5 mg/kg, more toxic). Among 5 other tested octachloro-derivatives of Toxicant B, PB pretreatment did not influence LD5o
values to the same degree as Toxicant B values, but 2 compounds were less toxic than TT ("3-exo-B": >100 mg/kg ± PB; 10-C1-B:
>100 mg/kg - PB, 48 mg/kg + PB) and the other 3 were more toxic ("5-exo-B": 24 and 28 mg/kg; 8-C1-B [57%] + 9-C1-B [43%]: 3.3 and
3.1 mg/kg; 10-C1-B: 2.5 and 1.9 mg/kg).
Turner et al. (1977)
Neurobehavioral tests were conducted in offspring of groups of pregnant Holtzman albino rats exposed to control diets, a diet delivering 50 |ig/kg-d
TT, a diet delivering 2 |ig/kg-d "Toxicant A" (p-42a or p-42b), or a diet delivering 2 |ig/kg-d "Toxicant B" (p-32). Dietary exposure started on
GD 5 and lasted until the offspring were 90 d old. The results indicate apparent NOAELs of 0.002 mg Toxicant A (p-42)/kg-d and 0.002 mg
Toxicant B (p-32)/kg-d for the absence of clear biologically significant neurobehavioral effects, body-weight effects, or effects on tissue histology
in Holtzman albino rat offspring. No LOAELs were identified. Neither a NOAEL nor a LOAEL could be identified for TT due to incomplete
reporting.
Olson etal. r1980V
A 48-hr zebrafish (Danio rerio) embryo test for lethal malformations and nonlethal malformations was conducted with 2 synthetic mixtures of
chlorinated camphenes: 1 enriched in higher-chlorinated congeners (hepta- to nona-chlorinated components) and showing a GC/electron capture
negative ion mass spectrometry multiple ion chromatogram similar to TT, and 1 enriched in lower-chlorinated congeners with very small amounts
of chlorinated components in the hepta- to nona-chlorination range. The respective 48-hr EC50 values for lethal and nonlethal malformations were
15.3 and 5.6 mg/L for the lower chlorination mixture and 20.6 and 13.4 mg/L for the higher chlorination mixture. The results suggest that mixtures
containing enhanced amounts of lower chlorinated congeners resulting from environmental weathering (i.e., dechlorination) may be more potent in
this test system than TT.
Kaon et al. (2006)
Numerous morphological endpoints were assessed in explanted rat embryos following a 48-hr exposure to TT, p-26, or p-50, each at 0, 0.1, 1, or
5 uu/mL (Calciu et al.. 1997). Yolk sac diameters, head leneth. and crown rumo leneth were measured after exposure, and 16 moroholoeical
features were scored on a numerical ranking from 0-5. Total morphological score, crown-rump length, and head length were decreased by
exposure to TT, p-26, and p-50 to similar degrees at most of the tested concentrations, indicating equivalent potencies of the test materials on these
endpoints. All 3 test substances also increased the incidence of embryos with neural tube malformations at most tested concentrations. However,
p-26 and p-50 decreased scores for otic development to a greater degree than TT. Based on results from exposure to a 50:50 mixture of p-26 and
d-50. Calciu et al. (1997) oroDosed synergistic effects on certain endooints. These claims, however, cannot be substantiated bv the studv desien and
analytical procedure employed.
The effects of TT, p-26, and p-50 on morphological endpoints were also examined in explanted rat embryos under normal and hyperglycemic
conditions (Calciu et al.. 2002). In the absence of hvoerulvcemia. results were similar to those reported in the earlier studv. Tested hvoerelvcemic
conditions alone caused decreased crown-rump length, head length, and total morphological score, but did not produce malformations. Exposure to
TT or p-26 under the highest hyperglycemic conditions increased the incidence of neural tube malformations, compared with responses in the
absence of hyperglycemia, whereas hyperglycemia appeared to decrease the effects of p-50 on this endpoint.
Calciu et al. (2002);
Calciu et al. (1997)
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Table A-l. Studies Featuring Comparative Testing of Toxaphene Congeners and Mixtures
Study Description
Reference
TT, congener T2 (p-26), and congener T12 (p-50) each displayed estrogenic activities (proliferative effects in MCF7-E3 human breast cancer cells)
at a similar concentration (10 |iIVI). but the response to TT was greater than responses to either of the congeners.
Stelzer and Chan
(1999)
Static 24-hr acute bioassays with mosquito larvae indicated that toxaphene mixtures purified from lake trout samples from 2 U.S. lakes were similar
in toxicity to TT. The fish residue test materials showed a gas chromatogram profile of similar complexity to TT, but a slightly lower combined
concentration of "Toxicant A" (p-42a, p-42b) and "Toxicant B" (p-32) than TT.
Gooch and
Matsumura (1987)
aA more complete description of this study was presented in the main body of this report.
EC50 = median effective concentration; GC = gas chromatograph; GD = gestation day; LD50 = median lethal dose; LOAEL = lowest-observed-adverse-effect level;
NOAEL = no-observed-adverse-effect level; PB = phenobarbital; TT = technical toxaphene.
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Applicability of Computational Toxicology Methods to Fill Data Gaps for Weathered
Toxaphene and Toxaphene Congeners
In light of the limited data available for weathered toxaphene mixtures and individual
toxaphene congeners, U.S. EPA considered that computational toxicology methods, specifically
quantitative structure activity relationships (QSARs), might be used to fill gaps in the database
for these materials. U.S. EPA's Estimation Program Interfact Suite (EPISuite) (U.S. EPA.
2012c). the Organisation for Economic Co-operation and Development (OECD) QSAR toolbox
(OECD. 2017). and the VEGA (Benfenati et ai. 2013) models, all publically available
computerized QSAR models, were investigated.
U.S. EPA's EPISuite programs are based on a fragment contribution (additive)
method, which allows complex structure to be estimated based on smaller building blocks, such
as a carbon-chorine group. Fragment methods tend to overestimate a value as a fragment
increases in frequency. For example, EPISuite produces a log Kow estimate for the Cl-8
congener (CASRN 142534-71-2; T2) of 5.69, in good agreement with the experimental value of
5.52. However, the Cl-9 congener (CASRN 154159-06-5;
2,2,5,5,8,9,9,10,10-nonachlorobornane) results in an estimated value of 7.72, which is not in
good agreement with its experimental value of 5.96. The fragment method does not account for
differences in positional, spatial, or electronic (e.g., dipole) relationships. Thus, all congeners
with seven chlorines return the same Kow value, all of those with eight chlorines return another
value, and all with nine yet another, and so on. Although this method provides reasonable results
(except for the higher congeners), it does not provide any information to discriminate between
the various congeners possessing the same degree of chlorination.
The other models that were evaluated also employ fragment-based techniques; however,
they are alert based and not group-contribution based. In other words, they may trigger an alert
if a single structural alert is identified. Thus, if the model issues an alert for a carbon-chlorine
bond, it may issue the same alert whether six, seven, or eight chlorines are identified. This
happens when the model is trained on only a few compounds that are analogs to the compounds
being evaluated, which is the case for the toxaphene congeners. The chemical space they occupy
is not well represented in the model-building exercises, so the models are expected to give results
with a high degree of uncertainty. This is consistent with the results provided by the VEGA
models. Using the Cl-8 congener and the Cl-9 congener, as above, VEGA provided estimates for
10 models associated with human health endpoints (mutagenicity, cancer, reproductive and
developmental toxicity, and skin sensitization). The Cl-8 and Cl-9 congeners showed the exact
same results for all 10 models. Moreover, 7 of the 10 models reported the results as "not
reliable" or missing critical aspects. Similar results were provided by the OECD QSAR toolbox,
which listed the same alerts for both the Cl-8 and Cl-9 congeners for the cancer/genotoxicity
([poly] halogen and aliphatic halogen) and mutagenicity (halogen) endpoints, for example.
Based on this initial exercise, U.S. EPA concluded that (1) the chemical space associated
with the toxaphene congeners is not sufficiently represented in the training sets to provide a basis
for meaningful or reliable predictions of congener toxicity based on structure and (2) the
techniques do not discriminate between the toxicity of Cl-8 and Cl-9 congeners, and are unlikely
to do so for any congeners having the same degree of chlorination. As a result of this exercise,
QSAR approaches to predict toxicity of toxaphene congeners and mixtures were not pursued
further.
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Considerations of Alternative Approaches to Deriving Screening Subchronic and Chronic
p-RfDs for Weathered Toxaphene Mixtures
Approaches to deriving screening subchronic and chronic p-RfDs for weathered
toxaphene mixtures were considered, based on the available data. In one approach, the
10% benchmark dose lower confidence limit human equivalent dose (BMDLio [HED]) of
0.0092 mg/kg-day for cytoplasmic vacuolation (all severity grades) in the thyroid of F0 male
S-D rats following 25-29 weeks of dietary exposure (Chu et al.. 1988) is used as the point of
departure (POD), and a screening p-RfD for weathered toxaphene mixtures is therefore derived
as follows:
Screening Subchronic p-RfD = BMDLio (HED) UFc
for Weathered Toxaphene = 0.0092 mg/kg-day ^ 300
based on Technical Toxaphene = 3 x 10"5 mg/kg-day
Table A-2 summarizes the uncertainty factors for this screening subchronic p-RfD for
weathered toxaphene mixtures.
Table A-2. Uncertainty Factors for a Screening Subchronic p-RfD for Weathered
Toxaphene Mixtures Based on Cytoplasmic Vacuolation (All Severity Grades) in the
Thyroid of F0 Male S-D Rats Following 25-29 Weeks of Dietary Exposure
UF
Value
Justification
UFa
3
A UFa of 3 (10°5) is applied to account for uncertainty in characterizing the toxicokinetic or
toxicodynamic differences between rats and humans following TT exposure. The toxicokinetic
uncertainty has been accounted for by calculating an HED through application of a DAF as outlined
in the U.S. EPA's Recommended Use of Body Weight3''4 as the Default Method in Derivation of the
Oral Reference Dose (U.S. EPA, 2011b).
UFd
10
A UFd of 10 is applied because of the limited available data comparing the toxicity of TT and
samples of weathered toxaphene. Given the wide range in relative potencies of weathered
toxaphene samples and TT in the limited number of available comparisons, a UFd of 10 is applied
to protect against the possibility that any specific type or sample of weathered toxaphene may be
more toxic than technical toxaphene.
UFh
10
A UFh of 10 is applied for intraspecies variability to account for human-to-human variability in
susceptibility in the absence of quantitative information to assess the toxicokinetics and
toxicodynamics of TT in humans.
UFl
1
A UFl of 1 is applied for LOAEL-to-NOAEL extrapolation because the POD is a BMDL.
UFS
1
A UFS of 1 is applied because the principal study is greater than subchronic duration (25-29 wk).
UFC
300
Composite UF = UFA x UFD x UFH x UFL x UFS.
BMDL = benchmark dose lower confidence limit; DAF = dosimetric adjustment factor; HED = human equivalent
dose; LOAEL = lowest-observed-adverse-effect level; NOAEL = no-observed-adverse-effect level; POD = point of
departure; p-RfD = provisional reference dose; S-D = Sprague-Dawley; TT = technical toxaphene;
UF = uncertainty factor; UFa = interspecies uncertainty factor; UFC = composite uncertainty factor; UFD = database
uncertainty factor; UFH = intraspecies uncertainty factor; UFL = LOAEL-to-NOAEL uncertainty factor;
UFS = subchronic-to-chronic uncertainty factor.
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The BMDLio (HED) of 0.0092 mg/kg-day for cytoplasmic vacuolation (all severity
grades) in the thyroid of F0 male S-D rats following 25-29 weeks of dietary exposure (Chu et
al, 1988), used as the POD for the screening sub chronic p-RfD for weathered toxaphene
mixtures, is also used as the POD for the screening chronic p-RfD with the same composite
uncertainty factor (UFc) as follows:
Screening Chronic p-RfD = BMDLio (HED) UFc
for Weathered Toxaphene = 0.0092 mg/kg-day ^ 300
based on Technical Toxaphene = 3 x 10~5 mg/kg-day
Other approaches have been proposed in the literature for deriving an oral reference dose
(RfD) for weathered toxaphene using the data provided in Besselink et al. (2008). Besselink et
al. (2008) described an approach to deriving p-RfDs using a dai 1 y-dose-adj ustm ent of the
12.5 mg CLE/kg-week rat NOAEL noted above (12.5 mg CLE/kg-week x l week/7 days
= 1.79 mg/kg-day). In a similar approach, Simon and Manning (2006) proposed the mass (or
concentration) of p-26 (B8-1413), p-50 (B9-1679), and p-62 (B9-1025) as the dose metric for the
CLE test substance in the Besselink et al. (2008) study. These congeners were selected by
Simon and Manning (2006) to represent weathered toxaphene for deriving an RfD based on
analyses of toxaphene congeners in human serum and breast milk samples. Samples were
collected in five studies and show that >90% of the total congeners detected was the sum of only
three, p-26, p-50, and p-62 (£3PC). This suggests that humans are only exposed to these three
congeners due to rapid metabolic disposition and elimination of other congeners.
Comparatively, the Besselink et al. (2008) dosimetric approach based on total dose of
weathered toxaphene is more straightforward, involves fewer assumptions, and is better
supported by the data than the Simon and Manning (2006) approach, as detailed below:
1. The central assumption of the Simon and Manning (2006) approach that humans are
primarily exposed to the Y.3PC persistent congeners detected in human serum and breast
milk samples is highly uncertain. This is because the central assumption is based only on
detected levels in serum or breast milk samples [examination of toxaphene residues in
other human tissues was not conducted in the five studies cited by Simon and Manning
(2006)1, and the analytical techniques employed may not have been designed to detect
lower chlorinated toxaphene congeners with five or six chlorines per molecule. There are
no data to directly support the idea of rapid metabolic disposition and elimination of other
congeners in humans. There are, however, data suggesting the opposite (i.e., a shift away
from highly chlorinated to lesser-chlorinated congeners) by biological weathering in fish
(Besselink et al., 2008).
2. The X3PC content of the CLE test substance represented a very low percentage of its
total toxaphene mass and does not account for other toxaphene congeners in the CLE
sample that may be toxic. Simon and Manning (2006) estimated from chromatograms
presented in the Besselink et al. (2008) study report that the Y3PC percent content in the
CLE test substance was only between 0.2-0.3%. The chromatogram of the CLE sample
shows a complex profile of toxaphene congeners that is decidedly different from the
technical toxaphene fed to the cod, with enrichment of peaks in the lower chlorination
range of the chromatogram (Besselink et al 2008). There is evidence that toxaphene
mixtures enriched in congeners with <7 chlorines per molecule can be more toxic than
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mixtures enriched in congeners with >7 chlorines, including p-26, p-50, and p-62 (Kapp
et al., 2006). Not including these lower-chlorinated toxaphene congeners in deriving a
toxicity value for weathered toxaphene mixtures may miss a considerable toxic fraction
of whatever specific mixture of weathered toxaphene is being considered. This
possibility would lead to an underestimation of hazard potential or risk if the exposure
assessment were only based on measurements of £3PC content in environmental media
(e.g., fish as food, sediments, water) and the weathered toxaphene mixture to which the
target population was exposed was enriched in other toxic congeners, including those
with <7 chlorines.
The screening p-RfDs derived above using the Chu et al. (1988) study on technical
toxaphene are based on a dose metric of milligrams total toxaphene residues per kilogram body
weight per day. Applying the screening p-RfDs for weathered toxaphene to estimate health risks
would require chemical analyses for total toxaphene residues detected in environmental samples
to estimate daily intakes in exposed populations. Confidence is low in this derivation due to the
uncertainties discussed above and follow from the small amount of data comparing relative
toxicities of technical toxaphene with weathered toxaphene mixtures or among weathered
toxaphene mixtures of differing chemical composition. However, there is greater confidence in
the derivation based on technical toxaphene than a derivation based on the CLE study because
there is greater confidence in the POD used in the former. The POD for technical toxaphene was
derived from a high-quality study that administered technical toxaphene via the oral route and
established a lowest-observed-adverse-effect level (LOAEL) and a NOAEL for a sensitive
critical effect (histopathological thyroid lesions) supported by related findings in other animal
and human studies, as well as a comprehensive evaluation of other endpoints in the database as a
whole. In contrast, the POD for CLE was derived from a single study in rats that administered
CLE via a s.c. injection route (which imparts inherent uncertainty when extrapolating to an oral
dose), evaluated a single, less sensitive endpoint (liver tumor promotion), and failed to find any
effect supported in the toxicological database for technical toxaphene. Taken together,
derivation of a CLE-based screening p-RfD is precluded.
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APPENDIX B. DATA TABLES
Table B-l. Liver Endpoints in Male Mice Fed Technical Toxaphene in Diet for 14 Days3
Parameterb
Dose Group (mg/kg-d [ppm in food])
0
1.8
(10 ppm)
7.3
(40 ppm)
15
(80 ppm)
29.6
(160 ppm)
60.1
(320 ppm)
Range-Finding 14-D Study (B6C3Fi)
Relative liver weight
(% BW) (n= 12)
4.81 ±0.18
4.78 ±0.25
(-0.6%)
5.32 ±0.26
(+11%)
6.05 + 0.37*
(+26%)
6.55 + 0.32*
(+36%)
7.43 + 0.59*
(+54%)
Absolute liver weight (g)
(« = 12)
1.51 ±0.08
1.51 ±0.06
(0%)
1.72 ±0.21
(+14%)
1.77 + 0.17*
(+17%)
1.96 + 0.14*
(+30%)
2.05 + 0.15*
(+36%)
Serum ALT (IU/L)
(n = 3-5)
30.8 ±6.13
29.3 ±4.11
(-5%)
37.0 ±8.49
(+20%)
34.7 + 5.69
(+13%)
40.5 + 2.12
(+32%)
52.0 + 9.9*
(+69%)
BrdU labeling index in liver
(%)c (n = 5)
1.2 ±0.0
1.0 ±0.2
(-17%)
3.3±0.7
(+175%)
6.3 + 0.6
(+425%)
11.1 + 0.9*
(+825%)
17.6 + 3.6*
(+1,370%)
CAR Knockout 14-D Study (C57BL/6 and CAR )
Paramaterb
Dose Group (mg/kg-d [ppm in food])
0
(10 ppm)
(40 ppm)
(80 ppm)
(160 ppm)
60.4
(320 ppm)
Absolute liver weight:
Wild-type (C57BL/6) mice
1.54 ±0.04
NT
NT
NT
NT
1.96 + 0.14*
(+27%)
Absolute liver weight:
CAR~/_ mice
1.65 ±0.23
NT
NT
NT
NT
1.57 + 0.45
(-5%)
Relative liver weight:
Wild-type (C57BL/6) mice
5.80 ±0.21
NT
NT
NT
NT
9.09 + 0.45*
(+57%)
Relative liver weight:
CAR~/_ mice
5.91 ±0.26
NT
NT
NT
NT
6.35 + 0.9
(+7%)
"Wang et al. (2015).
bValues denote mean ± SD (% change from control) or SE as indicated, except where noted.
°Values denote mean ± SE (% change from control) digitally extracted from graphically presented data using
Grablt! software.
* Statistically significantly different from control at p< 0.05 using one-way ANOVA with Dunnett's post hoc test
as reported in the study.
ALT = alanine aminotransferase; ANOVA = analysis of variance; BrdU = bromodeoxyuridine; BW = body weight;
CAR = constitutive androstane receptor; NT = not tested; SD = standard deviation; SE = standard error.
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Table B-2. Liver Endpoints in Male B6C3Fi Mice Fed Technical Toxaphene in Diet for up
to 28 Days3'b
Time point
Dose Group (mg/kg-d [ppm in food])
0
0.6 (3 ppm)
5.9 (32 ppm)
60.3 (320 ppm)
Relative Liver Weight (g; n = 12)
D 7
D 14
D 28
4.95 ±0.31
5.25 ±0.33
5.28 ±0.27
5.28 ± 0.39 (+7%)
5.36 ± 0.36 (+2%)
5.27 ± 0.24 (-0.2%)
5.36 ± 0.73 (+8%)
5.57 ± 0.33 (+6%)
5.58 ±0.16 (+6%)
7.46 + 0.53* (+51%)
7.94 + 0.56* (+51%)
8.37 + 0.64* (+59%)
Absolute Liver Weight (% BW; n = 12)
D 7
D 14
D 28
1.31 ± 0.10
1.49 ±0.13
1.57 ±0.10
1.40 ±0.13 (+7%)
1.46 ±0.12 (-2%)
1.60 ±0.12 (+2%)
1.39±0.28 (+6%)
1.59 ±0.12 (+7%)
1.72 ±0.11 (+10%)c
1.93 + 0.21* (+47%)
2.07 + 0.23* (+40%)
2.34 + 0.26* (+49%)
Serum ALT (IU/L; n = 12)
D 7
D 14
D 28
38.67 ± 18.85
39.25 ± 17.03
59.83 ±37.82
25.92 ±4.12 (-33%)
32.08 ± 10.74 (-18%)
34.55 ± 6.80 (-42%)
41.33+ 17.1 (+7%)
33.73 + 6.48 (-14%)
43.36 + 7.20 (-28%)
66.58 + 14.89* (+72%)
77.83 + 37.46* (+98%)
100.67 + 38.36* (+68%)
BrdU Labeling Index in Liver (%; n = 5)d
D 7
D 14
D 28
0.9 ±0.3
1.2 ±0.6
0.9 ±0.2
0.9 ± 0.3 (0%)
1.5 ±0.3 (+25%)
1 ± 0.7 (+11%)
3.3 + 1.7 (+267%)
2.4 + 0.3 (+100%)
1.7+ 0.7 (+89%)
10.7 + 2.4* (+1,089%)
12.5 + 4.3* (+942%)
3.9 + 1.7* (+333%)
MDA Concentration in Liver (nmol/mg protein; n = 7)d
D 7
D 14
D 28
9.6 ±0.8
9.6 ±0.8
8.6 ±0.7
9.9 ± 0.5 (+3%)
9.3 ± 0.5 (-3%)
8.7 ± 0.7 (+1%)
10.7 + 0.4 (+12%)
9.5 + 1.2 (-1%)
9.5 + 0.5 (+11%)
12.2+ 1.1* (+27%)
12.8 + 0.6* (+33%)
11.6 + 0.4* (+35%)
"Wang et al. (2015).
bValues denote mean ± SD (% change from control) except where noted.
Increase was only 9.6% and not deemed to be biologically significant (i.e., >10%).
dValues denote mean ± SE digitally extracted from graphically presented data using Grablt! software.
* Statistically significantly different from control at p< 0.05 using one-way ANOVA with Dunnett's post hoc test
as reported in the study.
ALT = alanine aminotransferase; ANOVA = analysis of variance; BrdU = bromodeoxyuridine; BW = body weight;
MDA = malondialdehyde; SD = standard deviation; SE = standard error.
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Table B-3. Anti-SRBC Response (IgM and IgG) and Antitetanus Toxoid Response (IgG)
in Cynomolgus Monkeys Exposed to Encapsulated Technical Toxaphene for up to
75 Weeks3'b
Response
Weeks
Postimmunization
Dose Group (mg/kg-d)
0
0.1
0.4
0.8
Mean Log2 Titers ± SE (% change)0
Females (n = 10/group)
Anti-SRBC
primary
response
(IgM)
1
2
3
4
7.1 ±0.35
6.4 ±0.31
5.3 ±0.34
4.9 ±0.41
6.4 ±0.54 (-9.9)
5.2 ±0.73 (-18.8)
4.5 ±0.64 (-15.1)
4 ±0.61 (-18.4)
5.2±0.79* (-26.8)
4.6 ±0.78 (-28.1)
3.8 ±0.85 (-28.3)
3.2 ±0.63* (-34.7)
3.7+ 0.83* (-47.9)
3 + 0.88* (-53.1)
3 + 0.75* (-43.4)
2.8+ 0.61* (-42.9)
Anti-SRBC
secondary
responsed
(IgM)
5
6
7
8
6.1 ±0.43
4.7 ±0.34
4.3 ±0.21
3.9 ±0.28
4.6 ± 0.62 (-24.6)
3.8 ±0.42 (-19.1)
3.3 ±0.47 (-23.3)
2.8 ±0.53 (-28.2)
4.4 ± 0.48* (-27.9)
4.4 ± 0.52 (-6.4)
2.8 ±0.55* (-34.9)
2.6 ±0.64 (-33.3)
4.1 + 0.69* (-32.8)
3.8 + 0.57 (-19.1)
3.1 + 0.46* (-27.9)
2.3+ 0.58* (-41)
Anti-SRBC
(IgG)
1
2
3
6 ± 1.12
7.3 ± 1
7.4 ±0.9
3.5 ±0.75 (-41.7)
5.6 ± 1.01 (-23.3)
5.4 ± 1.07 (-27)
4.5 ± 1.13 (-25)
5.5 ± 1.19 (-24.7)
5.2 ± 1.17 (-29.7)
3.3 + 0.87 (-45)
3.6 + 0.91* (-50.7)
4.2 + 0.84* (-43.2)
Antitetanus
toxoid (IgG)
Pre immune
1
2
3
4
5
-1.54 ±0.13
-1.03 ±0.13
0.75 ±0.1
1.01 ±0.07
1.11 ±0.07
0.85 ±0.08
-1.41 ±0.11 (-8.4)
-0.89 ±0.08 (-13.6)
0.81 ±0.06 (+8)
1.05 ±0.07 (+4)
1.16 ±0.06 (+4.5)
0.89 ± 0.05 (+4.7)
-1.62 ±0.12 (+5.2)
-1.04 + 0.1 (+1)
0.84 + 0.04 (+12)
1.1 + 0.07 (+8.9)
1.21 + 0.07 (+9)
0.95 + 0.07 (+11.8)
-1.54 + 0.17(0)
-1.21+ 0.17 (+17.5)
0.45 + 0.08* (-40)
0.68+ 0.08* (-32.7)
0.84+ 0.09* (-24.3)
0.87 + 0.08 (+2.4)
Anti-SRBC
primary
response
(IgM)
1
2
3
Males (n = 5, controls; n = 4,0.8 mg/kg-d)
9 ±0.32
8 ±0.32
7.4 ±0.4
ND
ND
ND
ND
ND
ND
6.5 + 0.29* (-27.8)
5.25 + 0.63* (-34.4)
4.75+ 0.85* (-35.8)
aTryphonas et al. (2001).
bImmune testing occurred between exposure Weeks 33-46 for levels of immune components in blood,
Weeks 44-53 for SRBC responses, Weeks 53-63 for tetanus toxoid responses, and Weeks 66-70 for
pneumococcus responses. No exposure-related effects on blood immune components or antipneumoccocus
responses were found.
°% change = ([exposure mean - control mean] + control mean) x 100.
Secondary response (Weeks 5-8) indicates responses following a second exposure to SRBC antigen administered
during Week 4.
* Statistically significantly different from control value (p < 0.05 with Bonferroni adjustment) as determined by
study authors.
IgG = immunoglobulin G; IgM = immunoglobulin M; ND = no data; SE = standard error; SRBC = sheep red blood
cell.
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Table B-4. Relative Liver Weight and Serum ALP in Beagle Dogs Administered Technical
Toxaphene in Gelatin Capsules (Corn Oil Vehicle) for 13 Weeks3
Dose Group (mg/kg-d)
0
0.2
2.0
4.5
Parameterb
Males
Relative liver weight (% B W)
3.7 ±0.54
3.6 ±0.46 (-3%)
3.6 ±0.21 (-3%)
4.4 ± 0.45* (+19%)
Serum ALP (13 wk) (mlU/mL)
70 ±26
73 ± 49° (+4%)
94 ± 47 (+34%)
168 ± 66* (+140%)
Parameterb
Females
Relative liver weight (% B W)
3.0 ±0.6
3.4 ±0.4 (+13%)
3.6 ±0.31* (+20%)
3.9 ±0.46* (+30%)
Serum ALP (13 wk) (mlU/mL)
74 ±22
153 ± 80° (+107%)
145 ± 95° (+96%)
185 ± 68*° (+150%)
aChu et al. (1986).
bValues denote mean ± SD (% change from control) (n = 6 unless noted).
°n = 5.
* Significantly different from controls (p < 0.05) using one-way ANOVA with Dunnett's post hoc test as reported
in the study.
ALP = alkaline phosphatase; ANOVA = analysis of variance; BW = body weight; SD = standard deviation.
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Table B-5. Incidences of Non-neoplastic Lesions in Beagle Dogs Administered Technical
Toxaphene in Gelatin Capsules (Corn Oil Vehicle) for 13 Weeks"
Lesionb
Dose Group (mg/kg-d)
0
0.2
2.0
4.5
Males
Liver:
Accented zonation
Periportal eosinophilia
Cytoplasmic vacuolation
Increased cytoplasmic density
2/6 (33)
1/6 (17)
0/6 (0)
1/6 (17)
2/6 (33)
3/6 (50)
0/6 (0)
3/6 (50)
1/6 (17)
4/6 (67)
2/6 (33)
3/6 (50)
3/6 (50)
6/6* (100)
2/6 (33)
6/6* (100)
Kidney:
Glomerular adhesions
1/6 (17)
3/6 (50)
2/6 (33)
4/6 (67)
Thyroid:
Reduced follicle size/follicular collapse
Increased epithelial height
Reduced colloid density
0/6 (0)
0/6 (0)
0/6 (0)
0/6 (0)
1/6 (17)
1/6 (17)
1/6 (17)
3/6 (50)
1/6 (17)
3/6 (50)
2/6 (33)
2/6 (33)
Lesion
Females
Liver:
Accented zonation
Periportal eosinophilia
Cytoplasmic vacuolation
Increased cytoplasmic density
1/6 (17)
2/6 (33)
0/6 (0)
2/6 (33)
2/6 (33)
1/6 (17)
0/6 (0)
1/6 (17)
3/6 (50)
4/6 (67)
0/6 (0)
4/6 (67)
4/6 (67)
6/6 (100)
0/6 (0)
5/6 (83)
Kidney:
Glomerular adhesions
0/6 (0)
1/6 (0)
3/6 (50)
1/6 (17)
Thyroid:
Reduced follicle size/follicular collapse
Increased epithelial height
Reduced colloid density
0/6 (0)
0/6 (0)
0/6 (0)
4/6 (67)
4/6 (67)
5/6 (83)*
5/6* (83)
4/6 (67)
5/6* (83)
3/6 (50)
2/6 (33)
2/6 (33)
aChu et al. (1986).
bValues denote number of animals showing changes/total number of animals examined (% in parentheses).
* Statistically significantly different from control at p< 0.05, as calculated for this review (Fisher's exact test,
two-tailed).
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Table B-6. Relative Liver Weight in S-D Rats Fed Technical Toxaphene in Food for
13 Weeks"
Parameterb
Dose Group (mg/kg-d [ppm in food])
Males
0
0.35 (4 ppm)
1.8 (20 ppm)
8.6 (100 ppm)
45.9 (500 ppm)
Relative liver weight
(% BW)
3.8 ±0.27
3.7 ±0.29
(-3%)
3.9 ±0.29
(+3%)
3.9 ± 0.18
(+3%)
4.5 ±0.58*
(+18%)
Parameterb
Females
0
0.5 (4 ppm)
2.6 (20 ppm)
12.6 (100 ppm)
63 (500 ppm)
Relative liver weight
(% BW)
3.4 ±0.32
3.6 ± 0.31
(+6%)
3.7 ±0.22
(+9%)
3.7 ±0.36
(+9%)
4.5 ± 0.20*°
(+32%)
aChu et al. (1986).
bValues denote mean ± SD (% change from control) (n = 9 or 10).
°SD is reported in the study as 20; this is assumed to be typographical error.
* Significantly different from controls at p < 0.5 using one-way ANOVA with Dunnett's post hoc test as reported in
the study.
ANOVA = analysis of variance; BW = body weight; S-D = Sprague-Dawley; SD = standard deviation.
Table B-7. Incidences of Non-neoplastic Lesions in S-D Rats Fed Technical Toxaphene in
Food for 13 Weeks"
Dose Group (mg/kg-d [ppm in food])
Males
Lesionb
0
0.35 (4 ppm)
1.8 (20 ppm)
8.6 (100 ppm)
45.9 (500 ppm)
Liver:
Accented zonation
Minimal to mild
0/10 (0)
1/10 (10)
3/10 (30)
5/10* (50)
6/10* (60)
Moderate to severe
0/10 (0)
0/10 (0)
1/10 (10)
0/10 (0)
4/10 (40)
All severity
0/10 (0)
1/10 (10)
4/10 (40)
5/10f (50)
10/10f (100)
Anisokaryosis
Minimal to mild
2/10 (20)
4/10 (40)
7/10 (70)
9/10* (90)
3/10 (30)
Moderate to severe
0/10 (0)
0/10 (0)
1/10 (0)
0/10 (0)
7/10* (70)
All severity
2/10 (20)
4/10 (40)
8/10f (80)
9/10f (90)
10/10f (100)
Nuclear necrosis
Minimal to mild
0/10 (0)
0/10 (0)
5/10* (50)
2/10 (20)
9/10* (90)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
0/10 (0)
0/10 (0)
5/10f (50)
2/10 (20)
9/10f (90)
Kidney:
Tubular injury-primary
Minimal to mild
0/10 (0)
2/10 (20)
8/10* (80)
8/10* (80)
1/10 (10)
Moderate to severe
1/10 (10)
2/10 (20)
2/10 (20)
2/10 (20)
9/10* (90)
All severity
1/10 (0)
4/10 (40)
10/10f (100)
10/10f (100)
10/10f (100)
Tubular necrosis
Minimal to mild
0/10 (0)
0/10 (0)
4/10 (40)
7/10* (70)
9/10* (90)
Moderate to severe
0/10 (0)
0/10 (0)
1/10 (10)
0/10 (0)
1/10 (10)
All severity
0/10 (0)
0/10 (0)
5/10f (50)
7/10f (70)
10/10f (100)
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Table B-7. Incidences of Non-neoplastic Lesions in S-D Rats Fed Technical Toxaphene in
Food for 13 Weeks"
Dose Group (mg/kg-d [ppm in food])
Thyroid:
Reduced follicular size
Minimal to mild
4/10 (40)
2/10 (20)
5/10 (50)
6/9 (67)
3/9 (33)
Moderate to severe
0/10 (0)
0/10 (0)
1/10 (10)
1/9(11)
2/9 (22)
All severity
4/10 (40)
2/10 (20)
6/10 (60)
7/18 (39)
5/9 (56)
Increased epithelial
height
Minimal to mild
5/10 (50)
7/10 (70)
3/10 (30)
0/9 (0)
0/9 (0)
Moderate to severe
2/10 (20)
3/10 (30)
7/10 (70)
9/9* (100)
9/9* (100)
All severity
7/10 (70)
10/10 (100)
10/10 (100)
9/9 (100)
9/9 (100)
Cytoplasmic vacuolation
Minimal to mild
1/10 (10)
2/10 (20)
2/10 (20)
5/9 (56)
5/9 (56)
Moderate to severe
1/10 (10)
1/10 (10)
2/10 (20)
4/9 (44)
2/9 (22)
All severity
2/10 (20)
3/10 (30)
4/10 (40)
9/9f (100)
7/9f (78)
Reduced colloid density
Minimal to mild
6/10 (60)
7/10 (70)
3/10 (30)
1/9 (11)
0/9 (0)
Moderate to severe
1/10 (10)
3/10 (30)
7/10* (70)
8/9* (89)
9/9* (100)
All severity
7/10 (70)
10/10 (100)
10/10 (100)
9/9 (100)
9/9 (100)
Females
Lesionb
0
0.5 (4 ppm)
2.6 (20 ppm)
12.6 (100 ppm)
63 (500 ppm)
Liver:
Accented zonation
Minimal to mild
0/10 (0)
4/10 (40)
5/10 (50)
7/10* (70)
3/10 (30)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
7/10* (70)
All severity
0/10 (0)
4/10 (4)
5/10f (50)
7/10f (70)
10/10f (100)
Anisokaryosis
Minimal to mild
0/10 (0)
6/10* (60)
9/10* (90)
9/10* (90)
3/10 (30)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
1/10 (10)
7/10* (70)
All severity
0/10 (0)
6/10f (60)
9/10f (90)
10/10f (100)
10/10f (100)
Nuclear necrosis
Minimal to mild
0/10 (0)
0/10 (0)
3/10 (30)
3/10 (30)
7/10* (70)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
0/10 (0)
0/10 (0)
3/10 (30)
3/10 (30)
7/10f (70)
Kidney:
Tubular injury-primary
Minimal to mild
0/10 (0)
2/10 (20)
1/10 (10)
6/10* (60)
10/10* (100)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
0/10 (0)
2/10 (20)
1/10 (10)
6/10f (60)
10/10f (100)
Tubular necrosis
Minimal to mild
0/10 (0)
8/10* (80)
9/10* (90)
8/10* (80)
8/10* (80)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
0/10 (0)
8/10f (80)
9/10f (90)
8/10f (80)
8/10f (80)
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Table B-7. Incidences of Non-neoplastic Lesions in S-D Rats Fed Technical Toxaphene in
Food for 13 Weeks"
Dose Group (mg/kg-d [ppm in food])
Thyroid:
Reduced follicular size
Minimal to mild
0/10 (0)
5/10* (50)
5/10* (50)
7/10* (70)
5/10* (50)
Moderate to severe
0/10 (0)
1/10 (10)
3/10 (30)
2/10 (20)
5/10* (50)
All severity
0/10 (0)
6/10f (60)
8/10f (80)
9/10f (90)
10/10f (100)
Increased epithelial
height
Minimal to mild
0/10 (0)
6/10* (60)
10/10* (100)
9/10* (90)
8/10* (80)
Moderate to severe
0/10 (0)
3/10 (30)
0/10 (0)
1/10 (10)
2/10 (20)
All severity
0/10 (0)
9/10f (90)
10/10f (100)
10/10f (100)
10/10f (100)
Cytoplasmic vacuolation
Minimal to mild
0/10 (0)
8/10* (80)
8/10* (80)
8/10* (80)
3/10 (30)
Moderate to severe
0/10 (0)
2/10 (20)
0/10 (0)
1/10 (10)
7/10* (70)
All severity
0/10 (0)
10/10f (100)
8/10f (80)
9/10f (90)
10/10f (100)
Reduced colloid density
Minimal to mild
0/10 (0)
9/10* (90)
9/10* (90)
10/10* (100)
3/10 (30)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
7/10* (70)
All severity
0/10 (0)
9/10f (90)
9/10f (90)
10/10f (100)
10/10f (100)
aCfau et at (1986).
bNumber of animals with lesion/number of animals examined microscopically (corresponding %).
* Statistically significantly (p < 0.05) different from control within the same severity group, as calculated for this
review (Fisher's exact test, two-tailed).
f Statistically significantly (p < 0.05) different from control; total number of lesions (minimal to mild + moderate to
severe) compared with the total number of lesions in the control, as calculated for this review (Fisher's exact test,
two-tailed).
S-D = Sprague-Dawley.
Table B-8. Relative Liver Weight in Male S-D Rats Fed Technical Toxaphene
in Diet for 9 Weeks"
Dose Group (mg/kg-d [ppm in food])
Parameterb
0
2.6 (30 ppm)
25.8 (300 ppm)
Relative liver weight (% B W)
4.5 ±0.1
5.0 ±0.1 (+11%)
5.6 ±0.1* (+24%)
''Roller et al. (1983).
bValues denote mean ± SE (% change from control).
* Significantly different from controls (p < 0.0001) using ANOVA and least square means as reported in the study.
ANOVA = analysis of variance; BW = body weight; S-D = Sprague-Dawley; SE = standard error.
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Table B-9. Liver-Weight Changes in Female Swiss-Webster Mice Fed Technical
Toxaphene in Diet for 8 Weeks3
Dose Group (mg/kg-d [ppm in food])
Parameterb
0
1.9 (10 ppm)
19.1 (100 ppm)
39.2 (200 ppm)
Absolute liver weight (g)
1.76
1.78 (+1%)
2.01 (+14%)
2.24 (+27%)
Relative liver weight (g/g BW)
0.062
0.061 (-2%)
0.072* (+16%)
0.089* (+44%)
"Allen et al. (1983).
Presented as mean (% change from control) (n = 23-26).
* Statistically significantly different from control at p < 0.5 based on /-test as reported in the study.
BW = body weight.
Table B-10. Incidence Data for Liver Tumors in B6C3Fi Mice Fed Technical Toxaphene
in Diet for 18 Months"'b
Dose Group (mg/kg-d [ppm in food])
Tumor Type
0
0.91 (7 ppm)
2.6 (20 ppm)
6.5 (50 ppm)
Males
Early deaths
Hepatocellular adenomas
0/9
0/7
0/8
0/5
Hepatocellular carcinomas
2/9
2/7
2/8
1/5
Terminal sacrifice
Hepatocellular adenomas
3/44
0/47
2/45
11/46
Hepatocellular carcinomas
5/44
9/47
10/45
11/46
Total for livers with hepatocellular tumors
10/53**
10/54
12/53
18/51*
Females
Early deaths
Hepatocellular adenomas
0/7
0/7
0/9
0/7
Hepatocellular carcinomas
0/7
0/7
0/9
0/7
Terminal sacrifice
Hepatocellular adenomas
1/46
1/46
1/43
3/45
Hepatocellular carcinomas
1/46
1/46
3/43
2/45
Total for livers with hepatocellular tumors
2/53
2/53
4/52
6/52
"Litton Bionetics (1978) as reported by U.S. EPA (1980).
bMice were fed technical toxaphene in food for 18 months, followed by a 6-month observation period. Reported
incidence data are for mice with tumors at the terminal sacrifice and for mice that died earlier.
* Significantly different from control by Fisher's exact test as reported by U.S. EPA (1980).
~~Significant trend by Cochran-Armitage trend test as reported by U.S. EPA (1980).
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Table B-ll. Incidence of Non-neoplastic Lesions in Osborne-Mendel Rats and B6C3Fi
Mice Fed Technical Toxaphene in the Diet for 80 Weeks3
Dose Group (mg/kg-d [TWA concentration in food, ppm])
Male Rats
Lesionb
0
38.9 (556 ppm)
77.88 (1,112 ppm)
Liver
Degeneration, ballooning
Necrosis, focal
Fatty metamorphosis
0/9 (0)
0/9 (0)
0/9 (0)
1/44 (2)
0/44 (0)
2/44 (5)
1/45 (2)
1/45 (2)
4/45 (9)
Kidney
Multiple cysts
Chronic inflammation
Cystic degeneration
0/9 (0)
4/9 (44)
0/9 (0)
1/45 (2)
9/45 (20)
1/45 (2)
0/45 (0)
20/45 (44)
0/45 (0)
Thyroid
Follicular cyst
C-cell hyperplasia
Follicular hyperplasia
0/7 (0)
0/7 (0)
0/7 (0)
2/41 (5)
2/41 (5)
3/41 (7)
1/35 (3)
1/35 (3)
3/35 (9)
Female Rats
Lesionb
0
41.6 (540 ppm)
83.29 (1,080 ppm)
Liver
Fatty metamorphosis
Basophilic cytoplasmic change
1/10(1)
0/10 (0)
2/42 (5)
0/42 (2)
2/40 (5)
1/40 (3)
Kidney
Chronic inflammation
1/8 (13)
8/49 (16)
2/48 (4)
Thyroid
Follicular cyst
C-cell hyperplasia
Follicular hyperplasia
0/6 (0)
0/6 (0)
0/6 (0)
0/43 (0)
4/43 (9)
5/43 (12)
2/42 (5)
2/42 (5)
3/42 (7)
Female Mice
Lesionb
0
17 (99 ppm)
34.2 (198 ppm)
Liver
Chronic inflammation
Fatty metamorphosis
0/9 (0)
0/9 (0)
1/49 (2)
0/49 (0)
0/49 (0)
1/49 (2)
Thyroid
Follicular hyperplasia
0/7 (0)
0/45 (0)
1/38 (3)
''NCI (1979).
bValues denote number of animals showing lesion/total number of animals examined (% in parentheses). No
statistically significantly increased incidences of any non-neoplastic lesions were found in exposed groups,
compared with controls. Incidence data in table are for lesions in liver, kidney, or thyroid with nonzero incidences
in exposed groups.
TWA = time-weighted average.
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Table B-12. Statistically Significant Incidence Data for Tumors in Osborne-Mendel Rats
and B6C3Fi Mice Fed Technical Toxaphene in Diet for 80 Weeks3'b'c
Tumor Type
Dose Group (mg/kg-d [ppm in food])
Male Rats
Matched Control
Pooled Control
38.9 (556 ppm)
77.88 (1,112 ppm)
Liver
Neoplastic nodules
Thyroid
Follicular-cell carcinoma or adenoma
1/9
1/7
1/52
2/44
6/44**
7/41
4/45
9/35**
Female Rats
Matched Control
Pooled Control
41.6 (540 ppm)
83.29 (1,080 ppm)
Pituitary
Chromophobe adenoma, adenoma
(NOS), or carcinoma
Thyroid
Follicular cell adenoma
3/8***
0/6***
17/51***
15/41
1/43
23/39**
7/42**
Male Mice
Matched Control
Pooled Control
17 (99 ppm)
34.0 (198 ppm)
Liver
Hepatocellular carcinoma
Hepatocellular carcinoma or adenoma
0/10***
2/10***
4/48***
7/48***
34/49*/**
40/49*/**
45/46*/**
45/46*/**
Female Mice
Matched Control
Pooled Control
17 (99 ppm)
34.2 (198 ppm)
Liver
Hepatocellular carcinoma
Hepatocellular carcinoma or adenoma
0/9***
0/9***
0/48***
0/48***
5/49*/**
18/49*/**
34/49*/**
40/49*/**
"NCI (1979).
bOnly data for significantly increased incidences or increased dose trends are reported in this table.
°Rats were fed technical toxaphene in food for 80 weeks. Groups of 10 matched rats/sex served as nonexposed
concurrent controls and up to 52 control rats/sex or 48 control mice/sex were pooled from other studies started
within a 5-month period of the start of the toxaphene study.
* Significantly different from matched control by Fisher's exact test as reported by NCI (1979).
~~Significantly different from pooled control by Fisher's exact test as reported by NCI (1979).
"""~Significant trend by Cochran-Armitage trend test as reported by NCI (1979).
NOS = not otherwise specified.
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Table B-13. Body-, Liver-, and Kidney-Weight Changes in S-D Rats Fed Technical
Toxaphene in Diet for up to 34 Weeks"'b
Dose Group (mg/kg-d [ppm in diet])
Parameter0
F0 Males
0
0.36 (4 ppm)
1.8 (20 ppm)
9.2 (100 ppm)
45 (500 ppm)
Body-weight gain (g)
529 ± 66
551 ±71
(+4%)
480 + 58
(-9%)
528 + 74
(-0.2%)
520 + 41
(-2%)
Absolute liver weight (g)
21.6 ±2.9
22.8 ±4.4
(+6%)
20.1 + 2.6
("7%)
21.2 + 3.6
(-2%)
25.5 + 3*
(+18%)
Relative liver weight (% B W)
3.6 ±0.47
3.6 ±0.52
(0%)
3.6 + 0.42
(0%)
3.5 + 0.57
(-3%)
4.2 + 0.4*d
(+17%)
Absolute kidney weight (g)
1.7 ± 0.15
1.9 ±0.21
(+12%)
1.7 + 0.19
(0%)
1.8 + 0.16
(+6%)
2 + 0.22*
(+18%)
Relative kidney weight (% BW)
0.28 ±0.03
0.29 ±0.03
(+4%)
0.31 + 0.04
(+11%)
0.3+0.03
(+7%)
0.33 + 0.03*
(+18%)
Parameter0
F0 Females
0
0.36 (4 ppm)
1.9 (20 ppm)
8.5 (100 ppm)
46 (500 ppm)
Body-weight gain (g)
237 ±32
249 ± 54
(+5%)
240 + 35
(+1%)
235 + 34
(-0.8%)
239 + 35
(+0.8%)
Absolute liver weight (g)
11.6 ± 2
12.9 ±2
(+11%)
13.8 + 3.3*
(+19%)
13.6 + 2.7*
(+17%)
16.6 + 3.2*
(+43%)
Relative liver weight (% B W)
3.7 ±0.48
4 ±0.57
(+8%)
4.4 + 0.92*
(+19%)
4.4 + 0.59*
(+19%)
5.3+0.84*
(+43%)
Absolute kidney weight (g)
1.1 ±0.09
1.1 ±0.09
(0%)
1.2 + 0.14
(+9%)
1.2 + 0.18
(+9%)
1.1 + 0.09
(0%)
Relative kidney weight (% BW)
0.36 ±0.03
0.36 ±0.04
(0%)
0.38 + 0.03
(+6%)
0.37 + 0.04
(+3%)
0.37 + 0.03
(+3%)
Parameter0
Fla Adult Males
0
0.29 (4 ppm)
1.4 (20 ppm)
7.5 (100 ppm)
37 (500 ppm)
Body-weight gain (g)
526 ± 53
500 ± 57
(-5%)
507 + 43
(-4%)
544 + 64
(+3%)
471 +68*
("11%)
Absolute liver weight (g)
20 ±2.5
22 ±3.1
(+10%)
21+4.1
(+5%)
21+2.4
(+5%)
24 + 3.6*
(+20%)
Relative liver weight (% BW)e
1.7 ±0.19
3.6±0.49
(+112%)
3.4 + 0.62
(+100%)
3.4 + 0.31
(+100%)
4.2 + 0.38*
(+147%)
Absolute kidney weight (g)
1.7 ± 0.19
1.8 + 0.23
(+6%)
1.8 + 0.15
(+6%)
2.0 + 0.2*
(+18%)
2.0 + 0.33*
(+18%)
Relative kidney weight (% BW)
0.28 ±0.02
0.3 + 0.04
(+7%)
0.3 + 0.02
(+7%)
0.31 + 0.02
(+11%)
0.34 + 0.05*
(+21%)
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Table B-13. Body-, Liver-, and Kidney-Weight Changes in S-D Rats Fed Technical
Toxaphene in Diet for up to 34 Weeks"'b
Dose Group (mg/kg-d [ppm in diet])
Parameter0
Fla Adult Females
0
0.38 (4 ppm)
1.9 (20 ppm)
9.4 (100 ppm)
49 (500 ppm)
Body-weight gain (g)
254 ±61
240 ± 29
(-6%)
245 ± 32
(-4%)
246 ± 42
(-3%)
216 ±26*
(-15%)
Absolute liver weight (g)
12 ± 1.7
13.7 ± 1.9
(+14%)
12.8 ± 1.6
(+7%)
13 ±2.2
(+8%)
15.7 ±2.2*
(+31%)
Relative liver weight (% B W)
3.9 ±0.93
4.1 ±0.47
(+5%)
3.9 ±0.43
(0%)
4.1 ±0.54
(+5%)
5.2 ±0.76*
(+33%)
Absolute kidney weight (g)
1.1 ± 0.18
1.2 ± 0.11
(+9%)
1.1 ± 0.14
(0%)
1.2 ±0.16
(+9%)
1.1 ±0.12
(0%)
Relative kidney weight (% BW)
0.35 ±0.08
0.36 ±0.04
(+3%)
0.35 ±0.04
(0%)
0.38 ±0.05
(+9%)
0.36 ±0.04
(+3%)
aChu et al. (1988).
bF0 rats were exposed to toxaphene between 25-29 weeks depending upon length of mating time, which was
described as "up to 3 weeks" and GD 0. F1 rats were exposed for a total of 34 weeks (3 weeks in utero, 3 weeks
via lactation, followed by 28 weeks via diet).
Data reported as mean ± SD (% change from control).
dStudy reports SD as 4.0; this is assumed to be a typographical error.
eThe control relative liver weight for Fla male rats was unusually low, compared with control means for
Fla females (3.9), F0 males (3.6), and F0 females (3.7). It is likely that this reflects a typographical error (note that
the values are the same as the control absolute kidney weight reported in a neighboring column in the study) and
that the calculated percent change values are erroneous.
* Statistically significantly different from control at p< 0.05, using one-way ANOVA with Duncan's multiple range
test, as reported in the study.
ANOVA = analysis of variance; BW = body weight; GD = gestation day; S-D = Sprague-Dawley; SD = standard
deviation.
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Table B-14. Incidences of Non-neoplastic Lesions in Parental S-D Rats Fed Technical
Toxaphene for 25-29 Weeks"'b
Dose Group (mg/kg-d [ppm in diet])
F0 Males
Lesion0
0
0.36 (4 ppm)
1.8 (20 ppm)
9.2 (100 ppm)
45 (500 ppm)
Thyroid:
Follicle collapse/angularity
Minimal to mild
0/12 (0)
1/10 (10)
3/10 (30)
3/11 (27)
6/13* (46)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/11 (0)
0/13 (0)
All severity
0/12 (0)
1/10 (10)
3/10 (30)
3/11 (27)
6/13 f (46)
Cytoplasmic vacuolation
Minimal to mild
0/12 (0)
3/10 (30)
8/10* (80)
6/11* (55)
12/13* (92)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
1/11 (9)
0/13 (0)
All severity
0/12 (0)
3/10 (30)
8/10f (80)
7/1 If (64)
12/13f (92)
Reduced colloid density
Minimal to mild
5/12 (42)
8/10 (0)
10/10* (100)
9/11* (82)
3/13 (23)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
2/11 (18)
9/13* (69)
All severity
5/12 (42)
8/10 (80)
10/10f (100)
11/1 If (100)
12/13f (92)
Colloid inspissation
Minimal to mild
0/12 (0)
4/10* (40)
8/10* (80)
4/11 (36)
5/13* (38)
Moderate to severe
0/12 (0)
1/10(1)
0/10 (0)
0/11 (0)
7/13* (54)
All severity
0/12 (0)
5/10f (50)
8/10f (80)
4/11 (36)
12/13f (92)
Liver:
Increased cytoplasmic density
Minimal to mild
0/12 (0)
4/10* (40)
2/10 (20)
2/11 (18)
12/13* (92)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
1/12 (0)
4/10f (40)
2/10 (20)
2/11 (18)
12/13f (92)
Increased cytoplasmic homogeneity
Minimal to mild
0/12 (0)
0/10 (0)
0/10 (0)
1/11 (9)
8/13* (62)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/11 (0)
4/13 (31)
All severity
0/12 (0)
0/10 (0)
0/10 (0)
1/11 (9)
12/13f (92)
Anisokaryosis
Minimal to mild
2/12 (17)
2/10 (20)
4/10 (40)
3/11 (27)
11/13* (85)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/11 (0)
0/13 (0)
All severity
2/12 (17)
2/10 (20)
4/10 (40)
3/11 (27)
ll/13f (85)
Kidney:
Primary tubular injury
Minimal to mild
0/12 (0)
4/10* (40)
5/10* (50)
3/11 (27)
11/13* (85)
Moderate to severe
0/12 (0)
3/10 (30)
0/10 (0)
0/11 (0)
2/13 (15)
All severity
0/12 (0)
7/10f (70)
5/10f (50)
3/11 (27)
13/13f (100)
Anisokaryosis
Minimal to mild
1/12 (8)
2/10 (20)
5/10* (50)
0/11 (0)
5/13 (38)
Moderate to severe
0/12 (0)
1/10 (10)
0/10 (0)
1/11 (9)
1/13 (8)
All severity
1/12 (8)
3/10 (30)
5/10f (50)
1/11 (9)
6/13 f (46)
Pyknosis
Minimal to mild
0/12 (0)
2/10 (20)
2/10 (20)
1/11 (9)
5/13* (38)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/11 (0)
0/13 (0)
All severity
0/12 (0)
2/10 (20)
2/10 (20)
1/11 (9)
5/13f (38)
Interstitial sclerosis
Minimal to mild
0/12 (0)
2/10 (20)
3/10 (30)
2/11 (18)
5/13* (38)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/11 (0)
1/13 (8)
All severity
0/12 (0)
2/10 (20)
3/10 (30)
2/11 (18)
6/13 f (46)
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Table B-14. Incidences of Non-neoplastic Lesions in Parental S-D Rats Fed Technical
Toxaphene for 25-29 Weeks"'b
Dose Group (mg/kg-d [ppm in diet])
F0 Females
Lesion
0
0.36 (4 ppm)
1.9 (20 ppm)
8.5 (100 ppm)
46 (500 ppm)
Thyroid:
Follicle collapse/angularity
Minimal to mild
5/17 (29)
6/10 (60)
8/10* (80)
7/10 (70)
9/17 (65)
Moderate to severe
0/17 (0)
1/10 (10)
0/10 (0)
0/10 (0)
1/17 (12)
All severity
5/17 (29)
7/10 (70)
8/10f (80)
7/10 (70)
10/17 (59)
Increased epithelial height
Minimal to mild
12/17 (71)
10/10 (100)
10/10 (100)
7/10 (70)
17/17* (100)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
3/10 (30)
0/17 (0)
All severity
12/17 (71)
10/10 (100)
10/10 (100)
10/10 (100)
17/17"f* (100)
Cytoplasmic vacuolation
Minimal to mild
2/17 (12)
4/10 (40)
4/10 (40)
6/10* (60)
9/17* (53)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
1/10 (10)
0/17 (0)
All severity
2/17 (12)
4/10 (40)
4/10 (40)
7/10f (70)
9/17f (53)
Liver:
Increased cytoplasmic vacuolation
Minimal to mild
1/17 (6)
0/10 (0)
0/10 (0)
0/10 (0)
9/17* (53)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/17 (0)
All severity
1/17 (6)
0/10 (0)
0/10 (0)
0/10 (0)
9/17f (53)
Increased cytoplasmic density
Minimal to mild
10/17 (59)
5/10 (50)
6/10 (60)
7/10 (70)
9/17 (53)
Moderate to severe
0/17 (0)
0/10 (0)
2/10 (2)
1/10 (10)
8/17* (47)
All severity
10/17 (59)
5/10 (50)
8/10 (80)
8/10 (80)
17/17"f* (100)
Increased cytoplasmic homogeneity
Minimal to mild
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
12/17* (71)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
5/17 (18)
All severity
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
17/17!" (100)
Anisokaryosis
Minimal to mild
4/17 (24)
8/10* (80)
10/10* (100)
7/10* (70)
14/17* (82)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
3/17(18)
All severity
4/17 (24)
8/10f (80)
10/10f (100)
7/10f (70)
17/17!" (100)
Kidney:
Primary tubular injury
Minimal to mild
0/17 (0)
0/10 (0)
5/10* (50)
6/10* (60)
17/17* (100)
Moderate to severe
0/17 (0)
1/10 (10)
0/10 (0)
0/10 (0)
0/17 (0)
All severity
0/17 (0)
1/10 (10)
5/10f (50)
6/10f (60)
17/17!" (100)
aChu et al. (1988s).
bF0 rats were exposed to toxaphene between 25-29 weeks depending upon length of mating time, which was
described as "up to 3 weeks" and GD 0.
°Number of animals with lesion/number of animals examined microscopically (corresponding %).
* Statistically significantly different from control at p< 0.05, compared with control (within the same severity
group), as calculated for this review (Fisher's exact test).
f Statistically significantly different from control at p< 0.05, total number of lesions (minimal to mild + moderate
to severe) compared with the total number of lesions in the control, as calculated for this review (Fisher's exact
test).
GD = gestation day; S-D = Sprague-Dawley.
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Table B-15. Incidences of Non-neoplastic Lesions in F1 S-D Rats Fed Technical
Toxaphene in Diet for 34 Weeks3'b
Dose Group (mg/kg-d [ppm in diet])
Fla Adult Males
Lesion0
0
0.29 (4 ppm)
1.4 (20 ppm)
7.5 (100 ppm)
37 (500 ppm)
Thyroid:
Reduction of follicle size
Minimal to mild
3/12 (25)
5/10 (50)
5/10 (50)
4/10 (40)
5/13 (38)
Moderate to severe
0/12 (0)
1/10 (10)
1/10 (10)
5/10* (50)
4/13 (31)
All severity
3/12 (25)
6/10 (60)
6/10 (60)
9/10f (90)
9/13 f (69)
Follicle collapse/angularity
Minimal to mild
0/12 (0)
3/10 (30)
5/10* (50)
3/10 (30)
5/13* (38)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
0/12 (0)
3/10 (30)
5/10f (50)
3/10 (30)
5/13f (38)
Increased epithelial height
Minimal to mild
5/12 (42)
9/10* (90)
10/10* (100)
8/10 (80)
9/13 (69)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
2/10 (20)
1/13 (8)
All severity
5/12 (42)
9/10f (90)
10/10f (100)
10/10f (100)
10/13 (77)
Reduced colloid density
Minimal to mild
6/12 (50)
8/10 (80)
10/10* (100)
6/10 (60)
7/13 (54)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
3/10 (30)
3/13 (23)
All severity
6/12 (50)
8/10 (80)
10/10f (100)
9/10 (90)
10/13 (77)
Colloid inspissation
Minimal to mild
1/12 (8)
1/10 (10)
3/10 (30)
5/10* (50)
6/13* (46)
Moderate to severe
0/12 (0)
4/10* (40)
3/10 (30)
4/10* (40)
2/13 (15)
All severity
1/12 (8)
5/10f (50)
6/10f (60)
9/10f (90)
8/13 f (62)
Liver:
Increased cytoplasmic vacuolation
Minimal to mild
3/12 (25)
9/10* (90)
7/10 (70)
10/10* (100)
7/13 (54)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/10 (0)
3/13 (23)
All severity
3/12 (25)
9/10f (90)
7/10 (70)
10/10f (100)
10/13"f* (77)
Increased cytoplasmic density
Minimal to mild
0/12 (0)
3/10 (30)
3/10 (30)
4/10 (40)
9/13* (69)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/10 (0)
1/13 (8)
All severity
0/12 (0)
3/10 (30)
3/10 (30)
4/10 (40)
10/13"f* (77)
Increased cytoplasmic
homogeneity
Minimal to mild
0/12 (0)
6/10* (60)
7/10* (70)
4/10* (40)
4/13* (31)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/10 (0)
6/13* (46)
All severity
0/12 (0)
6/10f (60)
7/10f (70)
4/10f (40)
10/13!" (77)
Anisokaryosis
Minimal to mild
0/12 (0)
1/10 (10)
6/10* (60)
3/10 (30)
6/13* (46)
Moderate to severe
0/12 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
0/12 (0)
0/10 (0)
6/10f (60)
3/10 (30)
6/13 f (46)
Kidney:
Primary tubular injury
Minimal to mild
3/12 (25)
4/10 (40)
5/10 (50)
5/10 (50)
9/13* (69)
Moderate to severe
0/12 (0)
1/10 (10)
1/10 (10)
0/10 (0)
1/13 (8)
All severity
3/12 (25)
5/10 (50)
6/10 (60)
5/10 (50)
10/13!" (77)
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Table B-15. Incidences of Non-neoplastic Lesions in F1 S-D Rats Fed Technical
Toxaphene in Diet for 34 Weeks3'b
Dose Group (mg/kg-d [ppm in diet])
Fla Adult Females
Lesion0
0
0.38 (4 ppm)
1.9 (20 ppm)
9.4 (100 ppm)
49 (500 ppm)
Thyroid:
Reduction of follicle size
Minimal to mild
1/17 (6)
3/10 (30)
3/10 (30)
6/10* (60)
5/17 (29)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
1/10 (10)
2/17 (12)
All severity
1/17 (6)
3/10 (30)
3/10 (30)
7/10f (70)
7/17f (41)
Follicle collapse/angularity
Minimal to mild
0/17 (0)
3/10* (30)
7/10* (70)
2/10 (20)
4/17 (24)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/17 (0)
All severity
0/17 (0)
3/10f (30)
7/10f (70)
2/10 (20)
4/17 (24)
Increased epithelial height
Minimal to mild
1/17 (6)
9/10* (90)
9/10* (90)
9/10* (90)
7/17* (41)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
2/17 (12)
All severity
1/17 (6)
9/10f (90)
9/10f (90)
9/10f (90)
9/17f (53)
Cytoplasmic vacuolation
Minimal to mild
1/17 (6)
5/10* (50)
8/10* (80)
9/10* (90)
6/17 (35)
Moderate to severe
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
0/10 (0)
All severity
1/17 (6)
5/10f (50)
8/10f (80)
9/10f (90)
6/17 (35)
Reduced colloid density
Minimal to mild
1/17 (6)
4/10* (40)
8/10* (80)
9/10* (90)
8/17* (47)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
1/17 (6)
All severity
1/17 (6)
4/10f (40)
8/10f (80)
9/10f (90)
9/17f (53)
Colloid inspissation
Minimal to mild
0/17 (0)
2/10 (20)
2/10 (20)
8/10* (80)
3/17(18)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
1/17 (6)
All severity
0/17 (0)
2/10 (20)
2/10 (20)
8/10f (80)
4/17 (24)
Papillary proliferation
Minimal to mild
0/17 (0)
2/10 (20)
2/10 (20)
8/10* (80)
3/17(18)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
1/17 (6)
All severity
0/17 (0)
2/10 (20)
2/10 (20)
8/10f (80)
4/17 (24)
Liver:
Increased cytoplasmic density
Minimal to mild
1/17 (6)
7/10* (70)
7/10* (70)
8/10* (80)
8/17* (47)
Moderate to severe
0/17 (0)
1/10 (10)
0/10 (0)
0/10 (0)
2/17 (12)
All severity
1/17 (6)
8/10f (80)
7/10f (70)
8/10f (80)
10/17"f* (59)
Increased cytoplasmic
homogeneity
Minimal to mild
0/17 (0)
0/10 (0)
0/10 (0)
3/10* (30)
9/17* (53)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
1/17 (6)
All severity
0/17 (0)
0/10 (0)
0/10 (0)
3/10f (30)
10/17"f* (59)
Anisokaryosis
Minimal to mild
0/17 (0)
4/10* (40)
3/10* (30)
4/10* (40)
7/17*f (41)
Moderate to severe
0/17 (0)
0/10 (0)
0/10 (0)
0/10 (0)
2/17 (12)
All severity
0/17 (0)
4/10f (40)
3/10f (30)
4/10f (40)
9/17f (53)
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Table B-15. Incidences of Non-neoplastic Lesions in F1 S-D Rats Fed Technical
Toxaphene in Diet for 34 Weeks3'b
Dose Group (mg/kg-d [ppm in diet])
Fib Male Pups
Fib Female Pups
Total Fib Pups
46d
46d
46d
Lesion0
0
(500 ppm)
0
(500 ppm)
0
(500 ppm)
Thyroid:
Reduction in follicle size
Minimal to mild
0/5 (0)
2/5 (40)
0/4 (0)
3/5 (60)
0/9 (0)
5/10* (50)
Moderate to severe
0/5 (0)
0/5 (0)
0/4 (0)
0/5 (0)
0/9 (0)
0/10 (0)
All severity
0/5 (0)
2/5 (0)
0/4 (0)
3/5 (60)
0/9 (0)
5/10f (50)
Increased epithelial height
Minimal to mild
0/5 (0)
3/5 (60)
1/5 (20)
3/5 (60)
1/10 (10)
6/10* (60)
Moderate to severe
0/5 (0)
0/5 (0)
0/5 (0)
0/5 (0)
0/10 (0)
0/10 (0)
All severity
0/5 (0)
3/5 (60)
1/5 (20)
3/5 (60)
1/10 (10)
6/10f (60)
Liver:
Accentuated zonation
Minimal to mild
0/5 (0)
4/5* (80)
0/5 (0)
5/5* (100)
0/10 (0)
9/10* (90)
Moderate to severe
0/5 (0)
1/5 (20)
0/5 (0)
0/5 (0)
0/10 (0)
1/10 (10)
All severity
0/5 (0)
5/5 "f (100)
0/5 (0)
5/5 "f (100)
0/10 (0)
10/10f (100)
Increased portal density
Minimal to mild
0/5 (0)
5/5* (100)
0/5 (0)
5/5* (100)
0/10 (0)
10/10* (100)
Moderate to severe
0/5 (0)
0/5 (0)
0/5 (0)
0/5 (0)
0/10 (0)
0/10 (0)
All severity
0/5 (0)
5/5 "f (100)
0/5 (0)
5/5 "f (100)
0/10 (0)
10/10f (100)
Kidney:
Anisokaryosis
Minimal to mild
2/5 (40)
2/5 (40)
0/5 (0)
5/5* (100)
2/10 (20)
7/10 (70)
Moderate to severe
0/5 (0)
0/5 (0)
0/5 (0)
0/5 (0)
0/10 (0)
0/10 (0)
All severity
2/5 (40)
2/5 (40)
0/5 (0)
5/5 "f (100)
2/10 (0)
7/10 (70)
Timetal. (1988).
bAdult F1 rats were exposed for a total of 34 weeks (3 weeks in utero, 3 weeks via lactation, followed by 28 weeks
via diet).
°Number of animals with lesion/number of animals examined microscopically (corresponding %).
dFl pup dose based on that administered to F0 dams.
* Statistically significantly different from control at p < 0.05. compared with control (within the same severity
group), as calculated for this review (Fisher's exact test).
f Statistically significantly different from control at p< 0.05, total number of lesions (minimal to mild + moderate
to severe) compared with the total number of lesions in the control, as calculated for this review (Fisher's exact
test).
S-D = Sprague-Dawley.
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Table B-16. Developmental Effects in Rats and Mice Exposed to Technical Toxaphene in
Corn Oil by Gastric Intubation during GDs 7-16a
Reproductive Parametersb
Exposure Group (mg/kg-d)
0
15
25
35
CD Stock Charles River Rats
Conception
Pregnancies brought to term0
Mortality (dams)
29/33 (88%)
0/33 (0%)
31/39 (79%)
2/39 (5%)
25/39* (64%)
3/39 (8%)
4/16* (25%)
5/16* (31%)
Maternal observations
Average-weight gain (g)
Average liver/body weight (%)
110 ±23
4.9 ±0.4
86 ± 19* (-22%)
5.2 ± 0.5 (+6%)
80 ± 26* (-27%)
5.0 ± 0.03 (+2%)
65 + 40* (-41%)
4.6 + 0.8 (-6%)
Fetal observations
Average implants
Average mortality
Average body weight (g)
Average number of sternal
ossification centers
Average number of caudal
ossification centers
9.3 ±2.7
10.7 ±9.8
4.19 ±0.32
5.4 ±0.06
4.6 ±0.6
9.5 ± 1.9 (+2%)
7.6 ± 6.6 (-29%)
3.99 ±0.39 (-5%)
5.1 ±0.6* (-6)
4.5 ± 0.6 (-2%)
10.6 ± 2.5 (+14%)
10.7 ± 18.1 (0%)
3.71±0.32* (-12%)
4.9 ± 0.6* (-9%)
4 ± 0.6* (-13%)
9.5 + 1.9 (+2%)
8.5 + 6 (-21%)
3.8 + 0.78 (-9%)
5.1 + 0.5 (-6%)d
4.4 + 1.1 (-4%)
Reproductive Parametersb
CD-I Stock Charles River Mice
Conception
Pregnancies brought to term0
Mortality (dams)
45/75 (60%)
1/75 (1%)
16/26 (62%)
0/26 (0%)
32/45 (71%)
0/45 (0%)
61/90 (68%)
7/90 (8%)e
Maternal observations
Average-weight gain (g)
Average liver/body weight (%)
5.9 ±2.5
6.5 ±0.6
4.6 ± 1.7 (-22%)
8.0 ± 0.9* (+23%)
4.6 ± 1.6* (-22%)
8.1 ±0.6* (+25%)
3.5+ 2.1* (-41%)
8.6 + 0.7* (+32%)
Fetal observations
Average implants
Average mortality (%)
Average body weight (g)
Average number of sternal
ossification centers
Average number of caudal
ossification centers
Incidence of litters with
encephaloceles (number of fetuses)
11.4 ± 2.3
8.3 ±8.3
1.13 ±0.27
6.0 ±0.2
6.4 ±2.3
0/45 (0)
12.1 ± 1.7 (+6%)
12.6 ± 13.4
1.19 ±0.24 (+5%)
5.6 ±0.1 (-7%)
5.7 ±2.2 (-11%)
0/16 (0)
12.3 + 2 (+8%)
16.6+ 14.2
1.2 + 0.22 (+6%)
5.9 + 0.3 (-2%)
6.2 + 2.3 (-3%)
0/32 (0)
12.6+1.2 (+11%)
10.2+10
1.17 + 0.21 (+4%)
5.8 + 0.6 (-3%)
6.3 + 2.7 (-2%)
5/61° (11)
aChemoff and Carver (1976).
Presented as mean ± SD (% change), except for incidence data.
°Number full term pregnancies/number inseminated (%).
dAvailable copy of this report did not have an asterisk for this group, but given the statistical significance of the
means and SD of the other groups, this was likely significantly different from control.
eFisher's exact test indicated a marginal (p = 0.07) increase compared with control incidence.
* Statistically significantly different from control at p < 0.05. using the Mann-Whitney "U" test for continuous
variables as reported by Chemoff and Carver (1976) or a two-tailed Fisher's exact test for incidence data for this
assessment.
GD = gestation day; SD = standard deviation.
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APPENDIX C. BENCHMARK DOSE MODELING RESULTS
SELECTION OF DATA SETS FOR BMD MODELING
Based on the review of the available laboratory animal toxicity studies, the liver, kidney,
thyroid, and immune system are sensitive noncancer toxicity targets of technical toxaphene. To
provide a common basis for comparing potential points of departure (PODs) and critical effects
for a subchronic provisional reference dose (p-RfD) for technical toxaphene (i.e., comparing
benchmark dose [BMD] and benchmark dose lower confidence limits [BMDLs] among the most
sensitive endpoints), data sets from available studies with multiple exposure levels for each of
these sensitive toxicity targets were selected for BMD analysis (see Table 4A and the
"Derivation of Provisional Values" section in the main body of this report).
MODELING PROCEDURE FOR DICHOTOMOUS DATA
The BMD modeling of dichotomous (quantal) data was conducted with U.S. EPA's
Benchmark Dose Software (BMDS, Version 2.5). For these data, all of the dichotomous models
(i.e., Gamma, Multistage, Logistic, Log-Logistic, Probit, Log-Probit, and Weibull) available
within the software were fit using recommended parameter constraints and a benchmark
response (BMR) of 10% extra risk [as outlined in the Benchmark Dose Technical Guidance; U.S.
EPA (2012b VI. For all technical toxaphene-induced dichotomous effects modeled, a BMR other
than 10% is not supported by the statistical and biological characteristics of the data sets
(i.e., there are no developmental/fetal effects or epidemiological effects). Adequacy of model fit
was judged based on the goodness-of-fitp-walue (p > 0.1), magnitude of scaled residuals in the
vicinity of the BMR, and visual inspection of the model fit. Among all models providing
adequate fit, the lowest BMDL was selected if the BMDL estimates from different models varied
>threefold; otherwise, the BMDL from the model with the lowest Akaike's information criterion
(AIC) was selected as the best BMDL estimate for this data set.
In addition, in the absence of a mechanistic understanding of the biological response to a
toxic agent, data from exposures much higher than the study's lowest-observed-adverse-effect
level (LOAEL) do not provide reliable information regarding the shape of the response at low
doses. Such exposures, however, can have a strong effect on the shape of the fitted model in the
low-dose region of the dose-response curve. Thus, if the lack of fit is due to characteristics of
the dose-response data for high doses, then the Benchmark Dose Technical Guidance document
allows for data to be adjusted by eliminating the high-dose group (U.S. LP A. 2012b). Because
the focus of BMD analysis is on the low-dose regions of the response curve, elimination of the
high-dose group(s) is deemed reasonable.
MODELING PROCEDURE FOR CONTINUOUS DATA
The BMD modeling of continuous data was conducted with U.S. EPA's BMDS
(Versions 2.5 and 2.6). For these data, all continuous models available within the software were
fit using recommended parameter constraints and a standard BMR of 1 standard deviation (SD)
relative risk unless a biologically determined BMR was available, as outlined in the Benchmark
Dose Technical Guidance (U.S. EPA. 2012b). An adequate fit was judged based on the
goodness-of-fit p-w alue (p > 0.1), magnitude of the scaled residuals in the vicinity of the BMR,
and visual inspection of the model fit. In addition to these three criteria forjudging adequacy of
model fit, a determination was made on whether the variance across dose groups was
homogeneous. If a constant/homogeneous variance model was deemed appropriate based on the
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statistical test provided by BMDS (i.e., Test 2), the final BMD results were estimated from a
constant/homogeneous variance model. If the test for homogeneity of variance was rejected
(p < 0.1), the model was run again while modeling the variance as a power function of the mean
to account for this nonconstant/nonhomogeneous variance. If this nonconstant/nonhomogeneous
variance model did not adequately fit the data (i.e., Test 3;p<0. 1), the data set was considered
unsuitable for BMD modeling. Among all models providing adequate fit, the lowest BMDL was
selected if the BMDL estimates from different models varied >threefold; otherwise, the BMDL
from the model with the lowest AIC was selected as the best BMDL estimate for this data set.
As described above for dichotomous data, if none of the models fit and lack of fit is due
to characteristics of the dose-response data for high doses, then the Benchmark Dose Technical
Guidance document allows for data to be adjusted by eliminating the high-dose group(s) (U.S.
EPA, 2012b). Because the focus of BMD analysis is on the low-dose regions of the response
curve, elimination of the high-dose group(s) is deemed reasonable.
BMD MODELING TO IDENTIFY POTENTIAL PODS FOR THE DERIVATION OF A
SUBCHRONIC PROVISIONAL REFERENCE DOSE
Decreased Mean Primary Anti-SRBC IgM Response in Female Cynomolgus Monkeys
Administered Technical Toxaphene in Oral Capsules for 75 Weeks (Trypfaonas et al., 2001)
The procedure outlined above was applied to the data for decreased mean primary
anti sheep red blood cell (SRBC) immunoglobulin M (IgM) response (measured 1 week
postimmunization) in female cynomolgus monkeys administered technical toxaphene via oral
capsule for 75 weeks (Tryphonas et al. 2001) (see Table C-l). Table C-2 summarizes the BMD
modeling results. The constant variance model did not fit the variance data, but the nonconstant
variance model did. With the nonconstant variance model applied, all models except the Hill
model provided adequate fit to the means. The Power and Polynomial 2- and 3-degree models
converged onto the Linear model. BMDLs for models providing adequate fit were not
sufficiently close (differed by >threefold), so the model with the lowest BMDL was selected
(Exponential Model 4; the Exponential Model 5 converged onto the Exponential Model 4).
Thus, the BMDLi sd of 0.02 mg/kg-day from this model is selected for this endpoint
(see Figure C-l and the BMD text output for details).
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Table C-l. Mean Primary Anti-SRBC IgM Response in Female Cynomolgus Monkeys
Administered Technical Toxaphene Orally in Capsules for 75 Weeks
(Measured 1 Week Postimmunization)3
HED (mg/kg-d)
0
0.05
0.2
0.4
Sample size
10
10
10
10
Mean
7.1
6.4
5.2
3.7
SEM
0.35
0.54
0.79
0.83
SDb
1.11
1.71
2.5
2.62
aTryphonas et al. (2001).
bCalculated using U.S. EPA BMDS (Version 2.5).
BMDS = Benchmark Dose Software; HED = human equivalent dose; IgM = immunoglobulin M; SD = standard
deviation; SEM = standard error of the mean; SRBC = sheep red blood cell.
Table C-2. BMD Model Predictions for Decreased Anti-SRBC Primary Response (IgM) in
Female Cynomolgus Monkeys Administered Technical Toxaphene Orally in Capsules for
75 Weeks (Measured 1 Week Postimmunization)
Model
Variance
/j-Valuc11
Means
/j-Valuc11
AIC
BMDisd
(mg/kg-d)
BMDLisd
(mg/kg-d)
Nonconstant variance
Exponential (model 2)b
0.48
0.61
96.66
0.13425
0.07516
Exponential (model 3)b
0.48
0.61
96.66
0.13425
0.07516
Exponential (model 4)b c
0.48
0.93
97.68
0.07426
0.02438
Exponential (model 5)b
0.48
0.93
97.68
0.07426
0.02438
Hill
0.48
NAd
99.68
0.07145
0.02043
Linear"1
0.48
0.41
97.45
0.17225
0.10993
Polynomial (2-degree)6
0.48
0.41
97.45
0.17225
0.10993
Polynomial (3-degree)e
0.48
0.41
97.45
0.17225
0.10993
Powerb
0.48
0.41
97.45
0.17225
0.10993
aValues <0.10 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Selected model.
dDegrees of freedom for Test 4 [means /?-value| are <0; the x2 test for fit is not valid.
"Coefficients restricted to be negative.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDL = benchmark dose lower confidence limit;
IgM = immunoglobulin M; NA = not applicable; SD = standard deviation; SRBC = sheep red blood cell.
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Exponential 4 Model, with BMR of 1 Std. Dev. for the BMD and 0.95 Lower Confidence Limit for the BMDL
8
7
3
2
Exponential 4
BMDL
BMD
0 0.05 0.1 0.15 0.2 0.25 0.3 0.35 0.4
dose
10:29 01/23 2017
Figure C-l. Exponential Model 4 for Anti-SRBC Primary Response (IgM) in Female
Cynomolgus Monkeys Administered Technical Toxaphene Orally in Capsules for
75 Weeks (Measured 1 Week Postimmunization) (Tryphonas et al., 2001)
Text Output for Figure C-l:
Exponential Model. (Version: 1.10; Date: 01/12/2015)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/exp_Tryphonas_2 001_IgM_monkey_lwk_HEDs_Exp-Mod
elVariance-BMRlStd-Down.(d)
Gnuplot Plotting File:
Mon Jan 23 10:29:08 2017
BMDS Model Run
The form of the response function by Model:
Model 2
Model 3
Model 4
Model 5
Y[dose] = a * exp{sign * b * dose}
Y[dose] = a * exp{sign * (b * dose)Ad}
Y[dose] = a * [c-(c-l) * exp{-b * dose}]
Y[dose] = a * [c-(c-l) * exp{-(b * dose)Ad}]
Note: Y[dose] is the median response for exposure
sign = +1 for increasing trend in data;
dose;
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sign = -1 for decreasing trend.
Model 2 is nested within Models 3 and 4.
Model 3 is nested within Model 5.
Model 4 is nested within Model 5.
Dependent variable = Mean
Independent variable = Dose
Data are assumed to be distributed: normally
Variance Model: exp(lnalpha +rho *ln(Y[dose]))
The variance is to be modeled as Var(i) = exp(lalpha + log(mean(i)) * rho)
Total number of dose groups = 4
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
MLE solution provided: Exact
Initial Parameter Values
Variable
lnalpha
rho
a
b
c
Model 4
5.31043
-2.39187
7.455
7.05264
0. 472677
1 Specified
Parameter Estimates
Variable Model 4
lnalpha
rho
a
b
c
6.80044
-3.27847
7.16005
6.36769
0.559175
Std. Err.
5 . 5848 6e-121
1.828
0.348381
5 .37507
0.173055
Table of Stats From Input Data
Dose
0
0.05
0.2
0.4
10
10
10
10
Obs Mean
7.1
6.4
5.2
3.7
Obs Std Dev
1.11
1.71
2.5
2. 62
Dose
0
0.05
0.2
0.4
Estimated Values of Interest
Est Mean Est Std Scaled Residual
7.16
6.299
4.887
4.251
1.189
1.4 67
2.224
2 .796
-0.1597
0.2169
0.445
-0.6232
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Other models for which likelihoods are calculated:
Model A1: Yij = Mu(i) + e(ij)
Var{e(ij)} = SigmaA2
Model A2 : Yij = Mu(i) + e(ij)
Var{e(ij)} = Sigma(i)^2
Model A3: Yij = Mu(i) + e(ij)
Var{e(ij)} = exp(lalpha + log(mean(i)) * rho)
Model R: Yij = Mu + e(i)
Var{e(ij)} = Sigma^2
Likelihoods of Interest
Model Log(likelihood) DF AIC
A1 -47.14702 5 104.294
A2 -43.09597 8 102.1919
A3 -43.83786 6 99.67571
R -54.27915 2 112.5583
4 -43.84127 5 97.68254
Additive constant for all log-likelihoods = -36.76. This constant added to the
above values gives the log-likelihood including the term that does not
depend on the model parameters.
Explanation of Tests
Test 1: Does response and/or variances differ among Dose levels? (A2 vs. R)
Test 2: Are Variances Homogeneous? (A2 vs. Al)
Test 3: Are variances adeguately modeled? (A2 vs. A3)
Test 6a: Does Model 4 fit the data? (A3 vs 4)
Test
Test 1
Test 2
Test 3
Test 6a
Tests of Interest
-2*log(Likelihood Ratio)
22.37
8.102
1.484
0.006824
D. F.
6
3
2
1
p-value
0. 001039
0.04395
0.4762
0.9342
The p-value for Test 1 is less than .05. There appears to be a
difference between response and/or variances among the dose
levels, it seems appropriate to model the data.
The p-value for Test 2 is less than .1. A non-homogeneous
variance model appears to be appropriate.
The p-value for Test 3 is greater than .1. The modeled
variance appears to be appropriate here.
The p-value for Test 6a is greater than .1. Model 4 seems
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to adequately describe the data.
Benchmark Dose Computations:
Specified Effect = 1.000000
Risk Type = Estimated standard deviations from control
Confidence Level = 0.950000
BMD = 0.0742627
BMDL = 0.0243837
Increased Incidence of Primary Tubular Injury (All Severity Grades) in the Kidney of
Male S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks (Chu et al.,
1986)
The procedure outlined above was applied to the data for increased incidence of primary
tubular injury (all severity grades) in the kidney of male Sprague-Dawley (S-D) rats
administered technical toxaphene in the diet for 13 weeks (Chu et al, 1986) (see Table C-3).
Table C-4 summarizes the BMD modeling results. All models provided adequate fit to the full
data set. BMDLs were considered to be sufficiently close (differed by
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Table C-4. BMD Modeling Results for Incidence of Primary Tubular Injury (All Severity
Grades) in the Kidney of Male S-D Rats Administered Technical Toxaphene in the Diet
for 13 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
1
25.96
0.06448
0.00989
Logistic
0.9998
23.98
0.03731
0.02100
LogLogistic0
1
25.96
0.07902
0.01903
LogProbit0
1
25.96
0.07164
0.01977
Multistage (l-degree)d
0.8291
25.24
0.01541
0.00841
Multistage (2-degree)d
0.9993
25.96
0.04100
0.00988
Multistage (3-degree)d
0.9999
25.96
0.04586
0.00988
Multistage (4-degree)d
0.9881
27.96
0.04586
0.00988
Probit6
1
23.96
0.03530
0.01994
Weibullb
1
25.96
0.04897
0.00989
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
"Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
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Probit Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
0.8
0.6
0.4
0.2
0 2 4 6 8 10 12
dose
16:01 01/23 2017
Figure C-2. Probit Model for Increased Incidence of Primary Tubular Injury (All Severity
Grades) in the Kidney of Male S-D Rats Administered Technical Toxaphene in the Diet for
13 Weeks (Chu et al.. 1986)
Text output for Figure C-2:
Probit Model. (Version: 3.3; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/pro_Chu_198 6_prim_tub_inj_male_rats_HEDs_Pro-B
MR10.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/pro_Chu_198 6_prim_tub_inj_male_rats_HEDs_Pro-B
MRlO.plt
Mon Feb 13 08:44:16 2017
BMDS Model Run
The form of the probability function is:
P[response] = CumNorm(Intercept+Slope*Dose) ,
where CumNormf .) is the cumulative normal distribution function
Dependent variable = Effect
Independent variable = Dose
Slope parameter is not restricted
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Total number of observations = 5
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial (and Specified) Parameter Values
background = 0 Specified
intercept = 0.440208
slope = 0.151431
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 -0.8
slope -0.8 1
Parameter Estimates
Interval
Variable
Limit
intercept
0.533694
slope
25.76
Estimate
-1.28325
-2.32927
11.4563
Std. Err.
-0.23723
7.29794
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-2.84743
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-9.98095
-9.98129
-30.5432
# Param's Deviance Test d.f. P-value
5
2 0.000688441 3 ]
1 41.1245 4 <.0001
AIC:
23.9626
Dose
Est. Prob.
Goodness of Fit
Expected
Observed
Size
Scaled
Residual
0.0000
0.0900
0.4600
2 .2000
11.8000
0.0997
0.4004
1.0000
1.0000
1.0000
0.997
4.004
10.000
10.000
10.000
1.000
4.000
10.000
10.000
10.000
10.000
10.000
10.000
10.000
10.000
0. 003
-0.003
0. 018
0. 000
0. 000
Chi^2 = 0.00
d.f. = 3
P-value = 1.0000
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Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0352965
BMDL = 0.0199366
Increased Incidence of Tubular Necrosis (All Severity Grades) in the Kidney of Male S-D
Rats Administered Technical Toxaphene in the Diet for 13 Weeks (Chu et al., 1986)
The procedure outlined above was applied to the data for increased incidence of tubular
necrosis (all severity grades) in the kidney of male S-D rats administered technical toxaphene in
the diet for 13 weeks (Chu et al. 1986) (see Table C-5). Table C-6 summarizes the BMD
modeling results. All models except the Logistic and Probit models provided adequate fit to the
full data set. BMDLs for models providing adequate fit were not sufficiently close (differed by
>threefold; i.e., LogLogistic vs. LogProbit), so the model with the lowest BMDL was selected
(LogLogistic). Thus, the BMDLio of 0.045 mg/kg-day from this model is selected for this
endpoint (see Figure C-3).
Table C-5. Incidence of Tubular Necrosis (All Severity Grades) in the Kidney of Male S-D
Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.090
0.46
2.2
11.8
Sample size
10
10
10
10
10
Incidence
0
0
5
7
10
"Chu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
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Table C-6. BMD Modeling Results for Incidence of Tubular Necrosis (All Severity
Grades) in the Kidney of Male S-D Rats Administered Technical Toxaphene in the Diet
for 13 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.4677
31.99
0.14578
0.09087
Logistic
0.0363
39.86
0.46924
0.29560
LogLogistic0'd
0.448
33.43
0.13829
0.04492
LogProbit0
0.4188
31.66
0.21139
0.13219
Multistage (l-degree)e
0.4677
31.99
0.14578
0.09087
Multistage (2-degree)e
0.4677
31.99
0.14578
0.09087
Multistage (3-degree)e
0.4677
31.99
0.14578
0.09087
Multistage (4-degree)e
0.312
33.99
0.14586
0.09087
Probit
0.0376
39.54
0.44182
0.29375
Weibullb
0.4677
31.99
0.14578
0.09087
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
"Betas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
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Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
1
0.8
0.6
0.4
0.2
0
0MDL
3MD
0
2
4
6
8
10
12
dose
16:08 01/23 2017
Figure C-3. LogLogistic Model for Increased Incidence of Tubular Necrosis (All Severity
Grades) in the Kidney of Male S-D Rats Administered Technical Toxaphene in the Diet for
13 Weeks (Chu et al.. 1986)
Text output for Figure C-3:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_kid_tublr_necro_male_rats_HEDs_Ln
1-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_kid_tublr_necro_male_rats_HEDs_Ln
1-BMR10-Restrict.pit
Mon Feb 13 08:47:32 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
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Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 0.190526
slope = 1.18187
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 0.23
slope 0.23 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
1.36091
slope
2.08521
Estimate
0
0.421479
1.32363
Std. Err.
NA
0.47931
0.388565
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-0.517951
0.562058
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-13.0401
-14.7155
-34.2965
# Param's
5
2
1
Deviance Test d.f.
3.35087
42.5128
P-value
0.3406
<.0001
AIC:
33.4311
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000 0.000 0.000 10.000 0.000
0.0900 0.0592 0.592 0.000 10.000 -0.793
121
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Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.138287
BMDL = 0.044 9213
FINAL
07-31-2018
0.4600
0.3529
3.529
5.000
10.000
0. 973
2.2000
0.8123
8.123
7.000
10.000
-0.910
11.8000
0.9756
9.756
10.000
10.000
0.500
Chi^2 = 2.65
d.f. = 3
P-
-value = 0.
, 4480
Increased Incidence of Minimal to Mild Tubular Necrosis (the Only Severity Grade
Observed) in the Kidney of Female S-D Rats Administered Technical Toxaphene in the
Diet for 13 Weeks CChu et al., 1986)
The procedure outlined above was applied to the data for increased incidence of minimal
to mild tubular necrosis in the kidney of female S-D rats administered technical toxaphene in the
diet for 13 weeks (Chit et al. 1986) (see Table C-7). Table C-8 summarizes the BMD modeling
results. None of the models provided adequate fit to the full data set or the data set with the
highest dose dropped. With the two highest doses dropped, only the LogLogistic model
provided adequate fit to the data. Thus, a BMDLio of 0.0012 mg/kg-day was calculated for this
endpoint (see Figure C-4). However, the modeling results for this endpoint are not considered
reliable because all response levels were considered in excess of the BMR, near maximal
responses, leaving no data to inform the shape of the dose-response curve in the low-dose region
and requiring extrapolation far below the observable range in order to estimate the BMDL (U.S.
EPA, 2012b).
Table C-7. Incidence of Minimal to Mild Tubular Necrosis (the Only Severity Grade
Observed) in the Kidney of Female S-D Rats Administered Technical Toxaphene in the
Diet for 13 Weeks"
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
8
9
8
8
aChu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
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Table C-8. BMD Modeling Results for Incidence of Minimal to Mild Tubular Necrosis (the
Only Severity Grade Observed) in the Kidney of Female S-D Rats Administered Technical
Toxaphene in the Diet for 13 Weeks
Model
X2 Goodness-of-Fit /j-Value11
AIC
BMDio (mg/kg-d)
BMDLio (mg/kg-d)
Gammab
0.0001
65.73
1.39491
0.53105
Logistic
0.0001
65.95
1.85515
0.81228
LogLogistic0
0
55.93
0.01208
0.00523
LogProbit0
0.0001
66.58
3.08714
0.83424
Multistage (l-degree)d
0.0001
65.73
1.39491
0.53105
Multistage (2-degree)d
0.0001
65.73
1.39491
0.53105
Multistage (3-degree)d
0.0001
65.73
1.39491
0.53105
Multistage (4-degree)d
0.0001
65.73
1.39491
0.53105
Probit
0.0001
65.99
1.99065
0.95831
Weibullb
0.0001
65.73
1.39492
0.53105
Highest dose dropped
Gammab
0.0002
51.76
0.16283
0.07286
Logistic
0.0002
52.79
0.29340
0.15043
LogLogistic0
0.0016
35.42
0.00733
0.00283
LogProbit0
0
52.83
0.19165
0.02691
Multistage (l-degree)d
0.0002
51.76
0.16283
0.07286
Multistage (2-degree)d
0.0002
51.76
0.16283
0.07286
Multistage (3-degree)d
0.0002
51.76
0.16283
0.07286
Probit
0.0002
52.93
0.32306
0.18394
Weibullb
0.0002
51.76
0.16283
0.07286
Two highest doses dropped
Gammab
0.0239
23.45
0.01453
0.00881
Logistic
0.0019
32.64
0.04609
0.02502
LogLogistic0'e
0.8038
18.89
0.00392
0.00117
LogProbit0
0.0939
21.23
0.01949
0.01035
Multistage (l-degree)d
0.0239
23.45
0.01453
0.00881
Multistage (2-degree)d
0.0239
23.45
0.01453
0.00881
Probit
0.0018
33.05
0.05261
0.03316
Weibullb
0.0239
23.45
0.01453
0.00881
"Values <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
"Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
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Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
T3
0.6
CD
(J
CD
<
£=
O
u
CD
0.4
Ll_
0.2
3MDLBMD
0
0.1
0.2
0.3
0.4
0.5
0.6
dose
16:18 01/23 2017
Figure C-4. LogLogistic Model for Incidence of Minimal to Mild Tubular Necrosis (the
Only Severity Grade Observed) in the Kidney of Female S-D Rats Administered Technical
Toxaphene in the Diet for 13 Weeks (Two Highest Doses Dropped) (Chu et al., 1986)
Text output for Figure C-4:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_kid_tublr_necro_female_rats_2hdd
HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_kid_tublr_necro_female_rats_2hdd
HEDs_Lnl-BMR10-Restrict.pit
Mon Feb 13 08:49:29 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 3
Total number of records with missing values = 0
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Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 3.09701
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
4 . 61616
slope
Estimate
0
3.3452
Std. Err.
NA
0.648458
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
2.07425
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-8 .25485
-8.44636
-20.527
# Param's Deviance Test d.f. P-value
3
1 0.383009 2 0.8257
1 24.5442 2 <.0001
AIC:
18.8927
Dose
Goodness of Fit
Est._Prob. Expected Observed Size
Scaled
Residual
0.0000
0.1100
0.5800
0.0000
0.7573
0.9427
0.000
7.573
9.427
0.000
8.000
9.000
10.000
10.000
10.000
0. 000
0.315
-0.581
Chi^2 = 0.44
d.f. = 2
P-value = 0.8038
125
Toxaphene
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FINAL
07-31-2018
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.00391702
BMDL = 0.00116777
Increased Incidence of Primary Tubular Injury (All Severity Grades) in the Kidney of
F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks (Chu et
al.. 1988)
The procedure outlined above was applied to the data for increased incidence of primary
tubular injury (all severity grades) in the kidney of F0 male S-D rats exposed to technical
toxaphene for 25-29 weeks (Chu et al. 1988) (see Table C-9). Table C-10 summarizes the
BMD modeling results. None of the models provided an adequate fit using the full dose range.
The data were not modeled with the highest dose dropped due to the lack of a statistically
significant increase in lesion incidence at the next lowest dose and the lack of a positive
dose-response trend in the remaining data. Thus, these data are not amenable to BMD modeling.
Table C-9. Incidence of Primary Tubular Injury (All Severity Grades) in the Kidney of
F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks"
HED (mg/kg-d)
0
0.10
0.47
2.4
12
Sample size
12
10
10
11
13
Incidence
0
7
5
3
13
"Chu et al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
126
Toxaphene
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07-31-2018
Table C-10. BMD Modeling Results for Incidence of Primary Tubular Injury (All Severity
Grades) in the Kidney of F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet
for 25-29 Weeks
Model
X2 Goodness-of-Fit
/>-Value"
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0043
59.62
4.07226
1.34026
Logistic
0.0009
62.39
0.88597
0.51514
LogLogistic0
0.0043
59.62
4.54597
1.62888
LogProbit0
0.0014
61.62
5.87284
NDr
Multistage (l-degree)d
0.0005
64.18
0.52257
0.27666
Multistage (2-degree)d
0.0022
60.92
1.93718
0.49432
Multistage (3-degree)d
0.0037
59.92
3.13691
0.55112
Multistage (4-degree)d
0.0042
59.69
4.11211
0.54808
Probit
0.0011
61.98
0.89367
0.52579
Weibullb
0.0014
61.62
7.59794
1.11709
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose confidence limit; NDr = not determined; S-D = Sprague-Dawley.
Increased Incidence of Primary Tubular Injury (All Severity Grades) in the Kidney of
F0 Female S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks (Cfau et
al.. 1988)
The procedure outlined above was applied to the data for increased incidence of primary
tubular injury (all severity grades) in the kidney of F0 female S-D rats exposed to technical
toxaphene for 25-29 weeks (Chu et al. 1988) (see Table C-l 1). Table C-12 summarizes the
BMD modeling results. All models except the Logistic, LogProbit, and Probit models provided
adequate fit to the full data set. BMDLs for models providing adequate fit were considered to be
sufficiently close (differed by
-------
FINAL
07-31-2018
Table C-ll. Incidence of Primary Tubular Injury (All Severity Grades) in the Kidney of
F0 Female S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks3
HED (mg/kg-d)
0
0.083
0.44
1.9
11
Sample size
17
10
10
10
17
Incidence
0
1
5
6
17
"Clin et al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
Table C-12. BMD Modeling Results for Incidence of Primary Tubular Injury (All Severity
Grades) in the Kidney of F0 Female S-D Rats Exposed to Technical Toxaphene in the Diet
for 25-29 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.3998
39.44
0.14019
0.08841
Logistic
0.0427
46.83
0.46504
0.29996
LogLogistic0
0.4386
41.09
0.08126
0.03541
LogProbit0
0.0975
43.85
0.22203
0.12513
Multistage (l-degree)d'e
0.3998
39.44
0.14019
0.08841
Multistage (2-degree)d
0.3998
39.44
0.14019
0.08841
Multistage (3-degree)d
0.3998
39.44
0.14019
0.08841
Multistage (4-degree)d
0.2564
41.44
0.14049
0.08843
Probit
0.0464
46.51
0.43295
0.29068
Weibullb
0.3998
39.44
0.14019
0.08841
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
"Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
128
Toxaphene
-------
FINAL
07-31-2018
Multistage Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Multistage
T3
0.6
CD
(J
CD
<
£=
O
u
CD
0.4
Ll_
0.2
BMDL
3MD
0
2
4
6
8
10
dose
15:24 03/07 2017
Figure C-5. Multistage (1-degree) Model for Increased Incidence of Primary Tubular
Injury (All Severity Grades) in the Kidney of FO Female S-D Rats Exposed to Technical
Toxaphene in the Diet for 25-29 Weeks (Chu et al., 1988)
Text output for Figure C-5:
Multistage Model. (Version: 3.4; Date: 05/02/2014)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/mst_Chu_198 8_prim_tub_inj_F0_female_rats_HEDs
Mstl-BMRIO-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/mst_Chu_198 8_prim_tub_inj_F0_female_rats_HEDs
Mstl-BMRIO-Restrict.pit
Tue Mar 07 16:00:05 2017
BMDS Model Run
The form of the probability function is:
P[response] = background + (1-background)*[1-EXP(
-betal*doseAl) ]
The parameter betas are not restricted
Dependent variable = Effect
Independent variable = Dose
129
Toxaphene
-------
FINAL
07-31-2018
Total number of observations = 5
Total number of records with missing values = 0
Total number of parameters in model = 2
Total number of specified parameters = 0
Degree of polynomial = 1
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial Parameter Values
Background = 0
Beta(l) = 9.36682e+018
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -Background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
Beta(1)
Beta (1) 1
Parameter Estimates
Interval
Variable
Limit
Background
Beta(1)
1.18918
Estimate
0
0.751553
Std. Err.
NA
0.223282
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
0.313928
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood) # Param'
-16.9124 5
-18.7185 1
-44.0798 1
Deviance Test d.f.
3.61219
54.3347
P-value
0. 461
<.0001
AIC:
39.437
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000 0.000 0.000 17.000 0.000
0.0830 0.0605 0.605 1.000 10.000 0.524
0.4400 0.2816 2.816 5.000 10.000 1.536
130
Toxaphene
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07-31-2018
1.9000 0.7602 7.602 6.000 10.000 -1.187
11.0000 0.9997 16.996 17.000 17.000 0.066
Chi^2 =4.05 d.f. = 4 P-value = 0.3998
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.14019
BMDL = 0.0884148
BMDU = 0.253985
Taken together, (0.0884148, 0.253985) is a 90 % two-sided confidence
interval for the BMD
Increased Incidence of Moderate to Severe Reduced Colloid Density in the Thyroid of Male
S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks CChu et al., 1986)
The procedure outlined above was applied to the data for increased incidence of moderate
to severe reduced colloid density in the thyroid of male S-D rats administered technical
toxaphene in the diet for 13 weeks (Chu et al. 1986) (see Table C-13). Table C-14 summarizes
the BMD modeling results. All models provided adequate fit to the full data set. BMDLs for
models providing adequate fit were not sufficiently close (differed by >threefold), so the model
with the lowest BMDL was selected (LogLogistic). Thus, the BMDLio of 0.013 mg/kg-day from
this model is selected for this endpoint (see Figure C-6).
Table C-13. Incidence of Moderate to Severe Reduced Colloid Density in the Thyroid of
Male S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.090
0.46
2.2
11.8
Sample size
10
10
10
9
9
Incidence
1
3
7
8
9
"Chu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
131
Toxaphene
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07-31-2018
Table C-14. BMD Modeling Results for Incidence of Moderate to Severe Reduced Colloid
Density in the Thyroid of Male S-D Rats Administered Technical Toxaphene in the Diet
for 13 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.5108
43.29
0.07792
0.04310
Logistic
0.2182
45.92
0.19529
0.11066
LogLogistic0'd
0.8764
43.59
0.03751
0.01258
LogProbit0
0.5115
43.05
0.11010
0.05486
Multistage (l-degree)e
0.5108
43.29
0.07792
0.04310
Multistage (2-degree)e
0.5108
43.29
0.07792
0.04310
Multistage (3-degree)e
0.5108
43.29
0.07792
0.04310
Multistage (4-degree)e
0.5108
43.29
0.07792
0.04310
Probit
0.2039
46.14
0.21292
0.13632
Weibullb
0.5108
43.29
0.07792
0.04310
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
"Betas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
132
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
1
0.8
0.6
0.4
0.2
0
3MDL3MD
0
2
4
6
8
10
12
dose
15:17 01/24 2017
Figure C-6. LogLogistic Model for Increased Incidence of Moderate to Severe Reduced
Colloid Density in the Thyroid of Male S-D Rats Administered Technical Toxaphene in the
Diet for 13 Weeks (Chu et al., 1986)
Text output for Figure C-6:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_red_coll_dens_male_rats_HEDs_Lnl-
BMRIO-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_red_coll_dens_male_rats_HEDs_Lnl-
BMR10-Restrict.pit
Mon Feb 13 08:51:24 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
133
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.1
intercept = 0.813584
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
background intercept slope
background 1 -0.23 0.23
intercept -0.23 1 0.51
slope 0.23 0.51 1
Parameter Estimates
Interval
Variable
Limit
background
0.283227
intercept
2.72806
slope
1.94179
Estimate
0.0997053
1.4565
1.11286
95.0% Wald Confidence
Std. Err. Lower Conf. Limit Upper Conf.
0.0936352 -0.0838163
0.648766 0.18494
0.422936 0.283917
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-18.6076
-18.7931
-32.6013
# Param's Deviance Test d.f. P-value
5
3 0.371014 2 0.8307
1 27.9873 4 <.0001
AIC:
43.5862
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0997 0.997
0.0900 0.3044 3.044
0.4600 0.6794 6.794
2.2000 0.9205 8.284
11.8000 0.9867 8.881
Chi^2 = 0.26 d.f. =2 P
1.000 10.000 0.003
3.000 10.000 -0.030
7.000 10.000 0.140
8.000 9.000 -0.350
9.000 9.000 0.348
value = 0.87 64
134
Toxaphene
-------
FINAL
07-31-2018
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0375081
BMDL = 0.0125836
Increased Incidence of Reduced Colloid Density (All Severity Scores) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks (Chu et al.,
1986)
The procedure outlined above was applied to the data for increased incidence of reduced
colloid density (all severity scores) in the thyroid of female S-D rats administered technical
toxaphene in the diet for 13 weeks (Chu et al. 1986) (see Table C-15). Table C-16 summarizes
the BMD modeling results. Only the LogLogistic model provided an adequate fit using the full
dose range. Thus, the BMDLio of 0.00051 mg/kg-day from this model is selected for this
endpoint (see Figure C-7). However, the modeling results for this endpoint are not considered
reliable because all response levels were far in excess of the BMR and at or near maximal
response at all doses tested, leaving no data to inform the shape of the dose-response curve in the
low-dose region and requiring extrapolation far below the observable range in order to estimate
the BMD (U.S. 1 PA. 2012b).
Table C-15. Incidence of Reduced Colloid Density (All Severity Scores) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
9
9
10
10
"Chu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
135
Toxaphene
-------
FINAL
07-31-2018
Table C-16. BMD Modeling Results for Incidence of Reduced Colloid Density
(All Severity Scores) in the Thyroid of Female S-D Rats Administered Technical
Toxaphene in the Diet for 13 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0047
22.83
0.01252
0.00761
Logistic
0.0054
32.75
0.04007
0.02119
LogLogistic0'd
0.8
16.33
0.00228
0.00051
LogProbit0
0.0393
19.39
0.01530
0.00763
Multistage (l-degree)e
0.0047
22.83
0.01252
0.00761
Multistage (2-degree)e
0.0047
22.83
0.01252
0.00761
Multistage (3-degree)e
0.0047
22.83
0.01252
0.00761
Multistage (4-degree)e
0.0047
22.83
0.01252
0.00761
Probit
0.0058
33.36
0.04831
0.03022
Weibullb
0.0047
22.83
0.01252
0.00761
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
"Betas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
136
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
0.8
0.6
0.4
0.2
Log-Logistic
3MDLBMD
09:14 02/13 2017
10
12
14
dose
Figure C-7. LogLogistic Model for Increased Incidence of Reduced Colloid Density (All
Severity Grades) in the Thyroid of Female S-D Rats Administered Technical Toxaphene in
the Diet for 13 Weeks (Chu et al., 1986)
Text output for Figure C-7:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_red_coll_dens_female_rats_HEDs_Ln
1-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_red_coll_dens_female_rats_HEDs_Ln
1-BMR10-Restrict.pit
Mon Feb 13 08:57:48 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
137
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 1.03449
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
5 .37241
slope
Estimate
0
3. 88693
1
Std. Err.
NA
0.757911
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
2.40146
Model
Full model
Fitted model
Reduced model
Analysis of Deviance Table
#
Log(likelihood)
-6.50166
-7.16625
-27.554
Param's
5
1
1
Deviance Test d.f.
1.32919
42.1047
P-value
0.8564
<.0001
AIC:
16.3325
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000
0.1100 0.8429
0.5800 0.9658
2.8100 0.9928
14.0000 0.9985
0.000 0.000
8.429 9.000
9.658 9.000
9.928 10.000
9.985 10.000
10.000 0.000
10.000 0.497
10.000 -1.147
10.000 0.270
10.000 0.121
138
Toxaphene
-------
FINAL
07-31-2018
Chi^2 = 1.65 d.f. = 4 P-value = 0.8000
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.00227868
BMDL = 0.000511344
Increased Incidence of Reduced Colloid Density (All Severity Grades) in the Thyroid of
Fla Female S-D Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks (Cfau et al.,
1988)
The procedure outlined above was applied to the data for increased incidence of reduced
colloid density (all severity grades) in the thyroid of Fla female S-D rats exposed to toxaphene
for 34 weeks (Chu et al.. 1988) (see Table C-17). Table C-18 summarizes the BMD modeling
results. None of the models provided an adequate fit using the full dose range. However, after
dropping the highest dose, the LogLogistic model provided an adequate fit to the data and a
BMDLio of 0.0075 mg/kg-day (see Figure C-8). However, the modeling results for this endpoint
are not considered reliable because all response levels were considered in excess of the BMR,
leaving no data to inform the shape of the dose-response curve in the low-dose region and
requiring extrapolation far below the observable range in order to estimate the BMD (U.S. EPA.
2012b).
Table C-17. Incidence of Reduced Colloid Density (All Severity Grades) in the Thyroid of
Fla Female S-D Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks"
HED (mg/kg-d)
0
0.089
0.44
2.2
11
Sample size
17
10
10
10
17
Incidence
1
4
8
9
9
Timet al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
139
Toxaphene
-------
FINAL
07-31-2018
Table C-18. BMD Modeling Results for Incidence of Reduced Colloid Density
(All Severity Grades) in the Thyroid of Fla Female S-D Rats Exposed to Technical
Toxaphene in the Diet for 34 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0
94.66
116,506
0.93269
Logistic
0
91.63
4.11944
1.70852
LogLogistic0
0
91.04
1.64893
0.247744
LogProbit0
0
92.41
8.70036
2.54749
Multistage (l-degree)d
0
91.44
3.09326
1.07897
Multistage (2-degree)d
0
91.44
3.09327
1.07897
Multistage (3-degree)d
0
91.44
3.09327
1.07897
Multistage (4-degree)d
0
91.44
3.09326
1.07897
Probit
0
91.63
4.12581
1.7431
Weibullb
0
91.44
3.09326
1.07897
Highest dose dropped
Gammab
0.0159
47.16
0.05652
0.03286
Logistic
0.0062
52.59
0.17106
0.09569
LogLogistic0'e
0.8344
41.90
0.01705
0.00753
LogProbit0
0.0091
45.76
0.06226
0.03488
Multistage (l-degree)d
0.0159
47.16
0.05652
0.03286
Multistage (2-degree)d
0.0159
47.16
0.05652
0.03286
Multistage (3-degree)d
0.0159
47.16
0.05652
0.03286
Probit
0.006
53.07
0.19812
0.12902
Weibullb
0.0159
47.16
0.05652
0.03286
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
140
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
T3
0.6
CD
(J
CD
<
£=
O
u
CD
0.4
Ll_
0.2
BMDL
3MD
0
0.5
1
1.5
2
dose
10:02 03/09 2017
Figure C-8. LogLogistic Model for Increased Incidence of Reduced Colloid Density (All
Severity Grades) in the Thyroid of Fla Female S-D Rats Exposed to Technical Toxaphene
in the Diet for 34 Weeks (Highest Dose Dropped) (Chu et al., 1988)
Text output for Figure C-8:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_red_coll_dens_Fla_female_rats_hig
h_drop_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_red_coll_dens_Fla_female_rats_hig
h_drop_HEDs_Lnl-BMR10-Restrict.pit
Thu Mar 09 10:02:15 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 4
Total number of records with missing values = 0
141
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.0588235
intercept = 1.62914
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
background intercept
background 1 -0.2 4
intercept -0.24 1
Parameter Estimates
Interval
Variable
Limit
background
0.17077
intercept
2.84544
slope
Estimate
0.0590057
1. 87423
1
Std. Err.
0.0570236
0.495527
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-0.0527585
0.903011
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-18.7882
-18.9519
-32.4821
# Param's
4
2
1
Deviance Test d.f.
0.327528
27.3879
P-value
0.8489
<.0001
AIC:
41.9039
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0590 1.003 1.000 17.000 -0.003
0.0890 0.4044 4.044 4.000 10.000 -0.028
142
Toxaphene
-------
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07-31-2018
0.4400 0.7567 7.567 8.000 10.000 0.319
2.2000 0.9386 9.386 9.000 10.000 -0.509
Chi^2 = 0.36 d.f. = 2 P-value = 0.8344
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0170526
BMDL = 0.00752682
Increased Incidence of Colloid Inspissation (All Severity Grades) in the Thyroid of F0 Male
S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks CChu et al., 1988)
The procedure outlined above was applied to the data for increased incidence of colloid
inspissation (all severity grades) in the thyroid of F0 male S-D rats exposed to technical
toxaphene for 25-29 weeks (Chu et al. 1988) (see Table C-19). Table C-20 summarizes the
BMD modeling results. None of the models provided adequate fit to the full data set or the data
set with the highest dose dropped. With the two highest doses dropped, all models except the
Logistic and Probit models provided adequate fit to the data. BMDLs for models providing
adequate fit were not sufficiently close (differed by >threefold), so the model with the lowest
BMDL was selected (LogLogistic). Thus, the BMDLio of 0.0050 mg/kg-day from this model is
selected for this endpoint (see Figure C-9). However, the modeling results for this endpoint are
not considered reliable because all response levels were considered in excess of the BMR,
leaving no data to inform the shape of the dose-response curve in the low-dose region and
requiring extrapolation far below the observable range in order to estimate the BMD (U.S. EPA.
2012b).
Table C-19. Incidence of Colloid Inspissation (All Severity Grades) in the Thyroid of
F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks"
HED (mg/kg-d)
0
0.10
0.47
2.4
12
Sample size
12
10
10
11
13
Incidence
0
5
8
4
12
"Cfauetal. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
143
Toxaphene
-------
FINAL
07-31-2018
Table C-20. BMD Modeling Results for Incidence of Colloid Inspissation (All Severity
Grades) in the Kidney of F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet
for 25-29 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0011
68.97
0.65277
0.34507
Logistic
0.0013
68.82
1.10538
0.66674
LogLogistic0
0.0007
69.28
0.25946
0.03458
LogProbit0
0.0005
70.76
8.07759
0.81332
Multistage (l-degree)d
0.0011
68.97
0.65277
0.34507
Multistage (2-degree)d
0.0004
70.79
1.27909
0.35179
Multistage (3-degree)d
0.0005
70.63
1.57717
0.35849
Multistage (4-degree)d
0.0005
70.59
1.62614
0.36007
Probit
0.0014
68.77
1.15310
0.75534
Weibullb
0.0011
68.97
0.65277
0.34507
Highest dose dropped
Gammab
0.0005
61.53
1.63456
0.27679
Logistic
0.0005
61.58
2.23688
0.48130
LogLogistic0
0.0005
61.46
1.04941
0.03266
LogProbit0
0.0005
61.71
94.47660
0.78750
Multistage (l-degree)d
0.0005
61.53
1.63458
0.27679
Multistage (2-degree)d
0.0005
61.53
1.63457
0.27679
Multistage (3-degree)d
0.0005
61.53
1.63457
0.27679
Probit
0.0005
61.57
2.20102
0.47730
Weibullb
0.0005
61.53
1.63459
0.27679
Two highest doses dropped
Gammab
0.4974
27.16
0.02386
0.01474
Logistic
0.0254
34.34
0.07512
0.04526
LogLogistic0'e
0.9874
25.90
0.01182
0.00502
LogProbit0
0.4656
27.20
0.03715
0.02209
Multistage (l-degree)d
0.4974
27.16
0.02386
0.01474
Multistage (2-degree)d
0.4974
27.16
0.02386
0.01474
Probit
0.0255
34.18
0.07327
0.04761
Weibullb
0.4974
27.16
0.02386
0.01474
"Values <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
"Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
144
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
1
0.8
0.6
0.4
0.2
0
BMDL
3MD
0
0.1
0.2
0.3
0.4
dose
10:00 03/13 2017
Figure C-9. LogLogistic Model for Increased Incidence of Colloid Inspissation (All Severity
Grades) in the Thyroid of FO Male S-D Rats Exposed to Technical Toxaphene in the Diet
for 25-29 Weeks (Two Highest Doses Dropped) (Chu et al., 1988)
Text output for Figure C-9:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_coll_insip_F0_male_rats_2_high_dr
op_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_coll_insip_F0_male_rats_2_high_dr
op_HEDs_Lnl-BMR10-Restrict.pit
Mon Mar 13 10:00:53 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 3
Total number of records with missing values = 0
145
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 2.21355
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
3.22046
slope
Estimate
0
2.24079
1
Std. Err.
NA
0. 499839
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
1.26113
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-11.9355
-11.948
-21.6149
# Param's
3
1
1
Deviance Test d.f.
0.0250626
19.3587
P-value
0.9875
<.0001
AIC:
25.8961
Dose
Goodness of Fit
Est._Prob. Expected Observed Size
Scaled
Residual
0.0000
0.1000
0.4700
0.0000
0.4846
0.8154
0.000
4.846
8.154
0.000
5.000
8.000
12.000
10.000
10.000
0. 000
0. 098
-0.126
Chi^2 =0.03
d.f. = 2
P-value = 0.9874
146
Toxaphene
-------
FINAL
07-31-2018
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0118193
BMDL = 0.00502303
Increased Incidence of Colloid Inspissation (All Severity Grades) in the Thyroid of
Fla Male S-D Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks CChu et al.,
1988)
The procedure outlined above was applied to the data for increased incidence of colloid
inspissation in the thyroid of Fla male S-D rats exposed to technical toxaphene for 34 weeks
(Chu et al. 1988) (see Table C-21). Table C-22 summarizes the BMD modeling results. None
of the models provided adequate fit to the full data set. With the highest dose dropped, all
models except the Logistic, LogProbit, and Probit models provided adequate fit to the data.
BMDLs for models providing adequate fit were not sufficiently close (differed by >threefold), so
the model with the lowest BMDL was selected (LogLogistic). Thus, the BMDLio of
0.0089 mg/kg-day from this model is selected for this endpoint (see Figure C-10). However, the
modeling results for this endpoint are not considered reliable because all response levels were
considered in excess of the BMR, leaving no data to inform the shape of the dose-response curve
in the low-dose region and requiring extrapolation far below the observable range in order to
estimate the BMD (U.S. EPA. 2012b).
Table C-21. Incidence of Colloid Inspissation (All Severity Grades) in the Thyroid of
Fla Male S-D Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks3
HED (mg/kg-d)
0
0.078
0.37
2.0
10
Sample size
12
10
10
10
13
Incidence
1
5
6
9
8
Timet al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
147
Toxaphene
-------
FINAL
07-31-2018
Table C-22. BMD Modeling Results for Incidence of Colloid Inspissation (All Severity
Grades) in the Thyroid of Fla Male S-D Rats Exposed to Technical Toxaphene in the
Diet for 34 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0028
78.43
2.42668
1.11047
Logistic
0.0034
77.16
0.51957
0.07874
LogLogistic0
0.0019
79.33
4.41833
1.28977
LogProbit0
0.0032
78.13
1.72534
0.66792
Multistage (l-degree)d
0.0032
78.13
1.72533
0.66792
Multistage (2-degree)d
0.0032
78.13
1.72532
0.66792
Multistage (3-degree)d
0.0032
78.13
1.72534
0.66792
Multistage (4-degree)d
0.0028
78.44
2.46806
1.17402
Probit
0.0032
78.13
1.72534
0.66792
Weibullb
0.0028
78.43
2.42668
1.11047
Highest dose dropped
Gammab
0.1411
48.68
0.07776
0.04044
Logistic
0.0815
50.37
0.17895
0.10154
LogLogistic0'e
0.5095
46.00
0.02129
0.00885
LogProbit0
0.0911
49.52
0.12050
0.04922
Multistage (l-degree)d
0.1411
48.68
0.07776
0.04044
Multistage (2-degree)d
0.1411
48.68
0.07776
0.04044
Multistage (3-degree)d
0.1411
48.68
0.07776
0.04044
Probit
0.0773
50.50
0.19227
0.12282
Weibullb
0.1411
48.68
0.07776
0.04044
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
148
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
T3
0.6
CD
(J
CD
<
£=
O
u
CD
0.4
Ll_
0.2
BMDL
3MD
0
0.5
1
1.5
2
dose
09:49 03/13 2017
Figure C-10. LogLogistic Model for Increased Incidence of Colloid Inspissation
(All Severity Grades) in the Thyroid of Fla Male S-D Rats Exposed to Technical
Toxaphene in the Diet for 34 Weeks (Highest Dose Dropped) (Chu et al., 1988)
Text output for Figure C-10:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_coll_insip_Fla_male_rats_high_dro
p_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_coll_insip_Fla_male_rats_high_dro
p_HEDs_Lnl-BMR10-Restrict.pit
Mon Mar 13 09:49:21 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 4
Total number of records with missing values = 0
149
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.0833333
intercept = 1.39858
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
background intercept
background 1 -0.4 9
intercept -0.49 1
Parameter Estimates
Interval
Variable
Limit
background
0.290309
intercept
2 .78408
slope
Estimate
0.102335
1.6521
1
Std. Err.
0.0959071
0.577555
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-0.0856397
0.520111
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-20.3545
-21. 0022
-29.1122
# Param's
4
2
1
Deviance Test d.f.
1.29543
17.5155
P-value
0.5232
0. 0005536
AIC:
46.0043
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.1023 1.228 1.000 12.000 -0.217
0.0780 0.3620 3.620 5.000 10.000 0.908
150
Toxaphene
-------
FINAL
07-31-2018
0.3700 0.6937 6.937 6.000 10.000 -0.643
2.0000 0.9215 9.215 9.000 10.000 -0.253
Chi^2 = 1.35 d.f. = 2 P-value = 0.5095
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0212942
BMDL = 0.00884992
Increased Incidence of Reduced Follicular Size (All Severity Grades) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks CChu et al.,
1986)
The procedure outlined above was applied to the data for increased incidence of reduced
follicular size in the thyroid of female S-D rats administered technical toxaphene in the diet for
13 weeks (Chit et al. 1986) (see Table C-23). Table C-24 summarizes the BMD modeling
results. Only the LogLogistic model provided adequate fit to the data. Thus, a BMDLio of
0.0052 mg/kg-day was calculated for this endpoint (see Figure C-l 1). However, the modeling
results for this endpoint are not considered reliable because all response levels were considered
in excess of the BMR, leaving no data to inform the shape of the dose-response curve in the
low-dose region and requiring extrapolation far below the observable range in order to estimate
the BMD (U.S. EPA. 2012b).
Table C-23. Incidence of Reduced Follicular Size (All Severity Grades) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
6
8
9
10
aChu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
151
Toxaphene
-------
FINAL
07-31-2018
Table C-24. BMD Modeling Results for Incidence of Reduced Follicular Size (All Severity
Grades) in the Thyroid of Female S-D Rats Administered Technical Toxaphene in the Diet
for 13 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.011
45.12
0.07840
0.03885
Logistic
0.0136
48.02
0.20145
0.10942
LogLogistic0'd
0.7933
33.39
0.01208
0.00523
LogProbit0
0
41.17
0.04871
0.02988
Multistage (l-degree)e
0.011
45.12
0.07840
0.03885
Multistage (2-degree)e
0.011
45.12
0.07840
0.03885
Multistage (3-degree)e
0.011
45.12
0.07840
0.03885
Multistage (4-degree)e
0.011
45.12
0.07840
0.03885
Probit
0.0127
48.39
0.23405
0.14672
Weibullb
0.011
45.12
0.07840
0.03885
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
"Betas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
152
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
1
0.8
0.6
0.4
0.2
0
3MDLBMD
0
2
4
6
8
10
12
14
dose
15:28 01/24 2017
Figure C-ll. LogLogistic Model for Increased Incidence of Reduced Follicular Size (All
Severity Grades) in the Thyroid of Female S-D Rats Administered Technical Toxaphene in
the Diet for 13 Weeks (Chu et al., 1986)
Text output for Figure C-ll:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_red_foll_size_female_rats_HEDs_Ln
1-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_red_foll_size_female_rats_HEDs_Ln
1-BMR10-Restrict.pit
Mon Feb 13 08:52:41 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
153
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 0.741985
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
3.16779
slope
Estimate
0
2.21871
1
Std. Err.
NA
0.484232
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
1.26964
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-14.985
-15.6956
-32.0518
# Param's
5
1
1
Deviance Test d.f.
1.42133
34.1336
P-value
0.8405
<.0001
AIC:
33.3913
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000
0.1100 0.5029
0.5800 0.8421
2.8100 0.9627
14.0000 0.9923
0.000 0.000
5.029 6.000
8.421 8.000
9.627 9.000
9.923 10.000
10.000 0.000
10.000 0.614
10.000 -0.365
10.000 -1.048
10.000 0.279
154
Toxaphene
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Chi^2 =1.69 d.f. = 4 P-value = 0.7933
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0120832
BMDL = 0.00523198
Increased Incidence of Minimal to Mild Follicle Collapse/Angularity (the Only Severity
Grade Observed) in the Thyroid of Fla Male S-D Rats Exposed to Technical Toxaphene in
the Diet for 34 Weeks CChu et al., 1988)
The procedure outlined above was applied to the data for increased incidence of minimal
to mild follicle collapse/angularity in the thyroid of Fla male S-D rats exposed to technical
toxaphene for 34 weeks (Chu et al. 1988) (see Table C-25). Table C-26 summarizes the BMD
modeling results. None of the models provided adequate fit to the full data set or the data set
with the highest dose dropped. With the two highest doses dropped, all models except the
Logistic and Probit models provided adequate fit to the data. BMDLs were considered to be
sufficiently close (differed by
-------
FINAL
07-31-2018
Table C-26. BMD Modeling Results for Incidence of Minimal to Mild Follicle
Collapse/Angularity (the Only Severity Grade Observed) in the Thyroid of Fla Male S-D
Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0223
72.33
5.5 x 1013
NDr
Logistic
0.0626
69.44
5.48264
2.32130
LogLogistic0
0.0619
69.36
4.15669
0.96097
LogProbit0
0.0615
69.61
7.18951
2.82028
Multistage (l-degree)d
0.0621
69.39
4.62114
1.42397
Multistage (2-degree)d
0.0621
69.39
4.62109
1.42397
Multistage (3-degree)d
0.0621
69.39
4.62111
1.42397
Multistage (4-degree)d
0.0621
69.39
4.62114
1.42397
Probit
0.0625
69.43
5.39480
2.24996
Weibullb
0.0621
69.39
4.62119
1.42397
Highest dose dropped
Gammab
0.0252
51.63
0.89025
0.23451
Logistic
0.0256
51.79
1.31883
0.48338
LogLogistic0
0.0249
51.50
0.62700
0.08991
LogProbit0
0.0255
52.30
44.40930
0.56758
Multistage (l-degree)d
0.0252
51.63
0.89025
0.23451
Multistage (2-degree)d
0.0252
51.63
0.89025
0.23451
Multistage (3-degree)d
0.0252
51.63
0.89025
0.23451
Probit
0.0255
51.78
1.27588
0.46317
Weibullb
0.0252
51.63
0.89025
0.23451
Two highest doses dropped
Gammab
0.4735
29.41
0.04343
0.02518
Logistic
0.0797
34.01
0.11959
0.07392
LogLogistic0'e
0.7476
28.64
0.03016
0.01360
LogProbit0
0.1584
31.21
0.06410
0.03927
Multistage (l-degree)d
0.4735
29.41
0.04343
0.02518
Multistage (2-degree)d
0.4735
29.41
0.04343
0.02518
Probit
0.0831
33.85
0.11075
0.07017
Weibullb
0.4735
29.41
0.04343
0.02518
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
156
Toxaphene
-------
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07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
0.7
0.6
0.5
T3
CD
(J
CD
<
£=
O
0.4
u
CD
Ll_
0.3
0.2
BMDL
3MD
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
dose
16:58 03/07 2017
Figure C-12. LogLogistic Model for Increased Incidence of Minimal to Mild Follicle
Collapse/Angularity (the Only Severity Grade Observed) in the Thyroid of Fla Male S-D
Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks (Two Highest Doses
Dropped) (Chu et al., 1988)
Text output for Figure C-12:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_foll_collap_ang_Fla_male_rats_2_h
igh_drop_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_foll_collap_ang_Fla_male_rats_2_h
igh_drop_HEDs_Lnl-BMR10-Restrict.pit
Wed Mar 08 08:46:54 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
157
Toxaphene
-------
FINAL
07-31-2018
Total number of observations = 3
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 1.31015
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
2.26316
slope
Estimate
0
1.30391
1
Std. Err.
NA
0.48942
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
0.344669
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood) # Param'
-13.0401 3
-13.3179 1
-17.9947 1
Deviance Test d.f.
0.555651
9.90922
P-value
0.7574
0.007051
AIC:
28.6359
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000 0.000 0.000 12.000 0.000
0.0780 0.2232 2.232 3.000 10.000 0.583
0.3700 0.5768 5.768 5.000 10.000 -0.492
158
Toxaphene
-------
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07-31-2018
Chi^2 = 0.58 d.f. = 2 P-value = 0.7476
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.030163
BMDL = 0.0136007
Increased Incidence of Increased Epithelial Height (All Severity Scores) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks (Cfau et al.,
1986)
The procedure outlined above was applied to the data for increased incidence of
increased epithelial height (all severity scores) in the thyroid of female S-D rats administered
technical toxaphene in the diet for 13 weeks (Chu et al. 1986) (see Table C-27). Table C-28
summarizes the BMD modeling results. All of the models provided an adequate fit using the full
dose range. BMDLs for models providing adequate fit were not sufficiently close (differed by
>threefold), so the model with the lowest BMDL was selected (LogLogistic). Thus, the BMDLio
of 0.00014 mg/kg-day from this model is selected for this endpoint (see Figure C-13). However,
the modeling results for this endpoint are not considered reliable because all response levels were
in excess of the BMR and at maximal response at all but the lowest dose tested, leaving no data
to inform the shape of the dose-response curve in the low-dose region and requiring
extrapolation far below the observable range in order to estimate the BMD (U.S. EPA. 2012b).
Table C-27. Incidence of Increased Epithelial Height (All Severity Scores) in the Thyroid
of Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
9
10
10
10
aChu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
159
Toxaphene
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07-31-2018
Table C-28. BMD Modeling Results for Incidence of Increased Epithelial Height (All
Severity Scores) in the Thyroid of Female S-D Rats Administered Technical Toxaphene in
the Diet for 13 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
1
10.50
0.01089
0.00257
Logistic
0.9556
11.09
0.03301
0.01377
LogLogistic0'd
1
10.50
0.07218
0.00014
LogProbit0
1
10.50
0.09035
NDr
Multistage (l-degree)e
1
8.50
0.00503
0.00257
Multistage (2-degree)e
1
10.50
0.00505
0.00258
Multistage (3-degree)e
1
12.50
0.00505
0.00258
Multistage (4-degree)e
0.9947
14.50
0.00505
0.00258
Probit
1
10.50
0.05945
0.01470
Weibullb
1
10.50
0.00503
0.00257
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
"Betas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; NDr = not determined; S-D = Sprague-Dawley.
160
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
0.8
0.6
0.4
0.2
Log-Logistic
'
3MDL
BMD
15:42 01/24 2017
10
12
14
dose
Figure C-13. LogLogistic Model for Increased Incidence of Increased Epithelial Height
(All Severity Grades) in the Thyroid of Female S-D Rats Administered Technical
Toxaphene in the Diet for 13 Weeks (Chu et al., 1986)
Text output for Figure C-13:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_incr_epith_ht_female_rats_HEDs_Ln
1-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_incr_epith_ht_female_rats_HEDs_Ln
1-BMR10-Restrict.pit
Mon Feb 13 08:54:11 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
161
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 1.15959
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 1
slope 1 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
8299.22
slope
3758.94
Estimate
0
25 .2192
10.4301
Std. Err.
NA
4221.51
1912.54
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-8248.78
-3738.08
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-3.25083
-3.25083
-26.3454
# Param's
5
2 6.
1
Deviance Test d.f.
5451e-008
46.1891
P-value
<.0001
AIC:
10.5017
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000 0.000 0.000 10.000 0.000
0.1100 0.9000 9.000 9.000 10.000 -0.000
162
Toxaphene
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07-31-2018
0.5800
1.0000
10.000
10.000
10.000
0. 000
2.8100
1.0000
10.000
10.000
10.000
0. 000
14.0000
1.0000
10.000
10.000
10.000
0. 000
Chi^2 = 0.00 d.f. = 3 P-value = 1.0000
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0721794
BMDL = 0.000141103
Increased Incidence of Cytoplasmic Vacuolation (All Severity Scores) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks CChu et al.,
1986)
The procedure outlined above was applied to the data for increased incidence of
cytoplasmic vacuolation (all severity scores) in the thyroid of female S-D rats administered
technical toxaphene in the diet for 13 weeks (Chu et al. 1986) (see Table C-29). Table C-30
summarizes the BMD modeling results. None of the models provided an adequate fit using the
full dose range. The data were not modeled with doses dropped due to a plateau of incidence
data at the lowest and highest doses tested. Thus, these data are not amenable to BMD modeling.
Table C-29. Incidence of Cytoplasmic Vacuolation (All Severity Scores) in the Thyroid of
Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks3
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
10
8
9
10
Timet al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
163
Toxaphene
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FINAL
07-31-2018
Table C-30. BMD Modeling Results for Incidence of Cytoplasmic Vacuolation
(All Severity Scores) in the Thyroid of Female S-D Rats Administered Technical
Toxaphene in the Diet for 13 Weeks
Model
X2 Goodness-of-Fit
/>-Value"
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0002
47.53
0.10944
0.04730
Logistic
0.0001
48.55
0.19616
0.09697
LogLogistic0
0
43.51
0.02725
0.01594
LogProbit0
0.0002
47.53
0.10944
0.04730
Multistage (l-degree)d
0.0002
47.53
0.10944
0.04730
Multistage (2-degree)d
0.0002
47.53
0.10944
0.04730
Multistage (3-degree)d
0.0002
47.53
0.10944
0.04730
Multistage (4-degree)d
0.0001
48.79
0.23013
0.13185
Probit
0.0002
47.53
0.10944
0.04730
Weibullb
0.0002
47.53
0.10944
0.04730
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
Increased Incidence of Cytoplasmic Vacuolation (All Severity Grades) in the Thyroid of
F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks CChu et
al.. 1988)
The procedure outlined above was applied to the data for increased incidence of
cytoplasmic vacuolation (all severity grades) in the thyroid of F0 male S-D rats exposed to
technical toxaphene for 25-29 weeks (Chu et al. 1988) (see Table C-31). Table C-32
summarizes the BMD modeling results. None of the models provided adequate fit to the full
data set or the data set with the highest dose dropped. With the two highest doses dropped, all
models provided adequate fit to the data. BMDLs for models providing adequate fit were not
sufficiently close (differed by >threefold), so the model with the lowest BMDL was selected
(LogLogistic). Thus, the BMDLio of 0.0092 mg/kg-day from this model is selected for this
endpoint (see Figure C-14).
164
Toxaphene
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07-31-2018
Table C-31. Incidence of Cytoplasmic Vacuolation (All Severity Grades) in the Thyroid of
F0 Male S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks"
HED (mg/kg-d)
0
0.10
0.47
2.4
12
Sample size
12
10
10
11
13
Incidence
0
3
8
7
12
"Clin et al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
Table C-32. BMD Modeling Results for Incidence of Cytoplasmic Vacuolation (All
Severity Grades) in the Thyroid of F0 Male S-D Rats Exposed to Technical Toxaphene in
the Diet for 25-29 Weeks
Model
X2 Goodness-of-Fit
/>-Value"
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0033
64.24
0.41622
0.21982
Logistic
0.0021
66.22
0.90629
0.52714
LogLogistic0
0.0629
53.06
0.04016
0.02005
LogProbit0
0.0016
65.75
0.63562
0.22562
Multistage (l-degree)d
0.0033
64.24
0.41620
0.21982
Multistage (2-degree)d
0.0033
64.24
0.41621
0.21982
Multistage (3-degree)d
0.0033
64.24
0.41620
0.21982
Multistage (4-degree)d
0.0033
64.24
0.41620
0.21982
Probit
0.002
66.45
1.01220
0.66379
Weibullb
0.0033
64.24
0.41620
0.21982
Highest dose dropped
Gammab
0.002
54.73
0.17615
0.09134
Logistic
0.0013
57.00
0.43085
0.25783
LogLogistic0
0.0396
45.21
0.03580
0.01745
LogProbit0
0.0004
56.72
0.23870
0.10203
Multistage (l-degree)d
0.002
54.73
0.17615
0.09134
Multistage (2-degree)d
0.002
54.73
0.17615
0.09134
Multistage (3-degree)d
0.002
54.73
0.17615
0.09134
Probit
0.0013
56.96
0.42618
0.26827
Weibullb
0.002
54.73
0.17615
0.09134
Two highest doses dropped
Gammab
0.9983
24.23
0.03038
0.01832
Logistic
0.1339
29.31
0.10075
0.05946
LogLogistic0'e
1
26.23
0.03925
0.00915
165
Toxaphene
-------
FINAL
07-31-2018
Table C-32. BMD Modeling Results for Incidence of Cytoplasmic Vacuolation (All
Severity Grades) in the Thyroid of FO Male S-D Rats Exposed to Technical Toxaphene in
the Diet for 25-29 Weeks
Model
X2 Goodness-of-Fit
/>-Value"
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
LogProbitd
0.9565
24.31
0.05096
0.03045
Multistage (l-degree)d
0.9983
24.23
0.03038
0.01832
Multistage (2-degree)d
0.9983
24.23
0.03038
0.01832
Probit
0.1442
29.07
0.09463
0.05930
Weibullb
0.9983
24.23
0.03038
0.01832
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
0.2
0
Log-Logistic
BMDL
0.2
0.3
0.4
dose
09:32 01/25 2017
Figure C-14. LogLogistic Model for Increased Incidence of Cytoplasmic Vacuolation
(All Severity Grades) in the Thyroid of FO Male S-D Rats Exposed to Technical Toxaphene
in the Diet for 25-29 Weeks (Two Highest Doses Dropped) CChu et al., 1988)
166 Toxaphene
-------
FINAL
07-31-2018
Text output for Figure C-14:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_thy_cyt_vac_F0_male_rats_2_high_d
rop_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_thy_cyt_vac_F0_male_rats_2_high_d
rop_HEDs_Lnl-BMR10-Restrict.pit
Mon Feb 13 09:24:54 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 3
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 2.47602
slope = 1.4433
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 0.91
slope 0.91 1
Parameter Estimates
95.0% Wald Confidence
Interval
Variable Estimate Std. Err. Lower Conf. Limit Upper Conf.
Limit
background 0 NA
167
Toxaphene
-------
FINAL
07-31-2018
intercept
4.87405
slope
2 .77232
2.47602
1.4433
1.22351
0.678086
0.0779842
0.114273
NA - Indicates that this parameter has hit a bound
implied by some inequality constraint and thus
has no standard error.
Warning: Likelihood for the fitted model larger than the Likelihood for the full
model.
Model
Full model
Fitted model
Reduced model
Analysis of Deviance Table
Log(likelihood) # Param's Deviance Test d.f.
-11.1127 3
-11.1127 2 -3.55271e-015 1
-20.5917 1 18.9581 2
P-value
<.0001
-1
AIC:
26.2253
Dose
Goodness of Fit
Est._Prob. Expected Observed Size
Scaled
Residual
0.0000
0.1000
0.4700
0.0000
0.3000
0.8000
0.000
3.000
8.000
0.000
3.000
8.000
12.000
10.000
10.000
0. 000
0. 000
0. 000
Chi^2 = 0.00
d.f. = 1
P-value = 1.0000
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0392465
BMDL = 0.00914841
Increased Incidence of Cytoplasmic Vacuolation (All Severity Grades) in the Thyroid of
F0 Female S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks (Cfau et
al.. 1988)
The procedure outlined above was applied to the data for increased incidence of
cytoplasmic vacuolation (all severity grades) in the thyroid of F0 female S-D rats exposed to
technical toxaphene for 25-29 weeks (Chu et al, 1988) (see Table C-33). Table C-34
summarizes the BMD modeling results. None of the models provided adequate fit to the full
data set. With the highest dose dropped, all models provided adequate fit to the data. BMDLs
for models providing adequate fit were not sufficiently close (differed by >threefold), so the
model with the lowest BMDL was selected (LogLogistic). Although the BMDLio of
0.037 mg/kg-day from this model is selected for this endpoint (see Figure C-15), it is considered
a borderline case for passing visual inspection of the model fit.
168
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Table C-33. Incidence of Cytoplasmic Vacuolation (All Severity Grades) in the Thyroid of
F0 Female S-D Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks3
HED (mg/kg-d)
0
0.083
0.44
1.9
11
Sample size
17
10
10
10
17
Incidence
2
4
4
7
9
"Clin et al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
169
Toxaphene
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07-31-2018
Table C-34. BMD Modeling Results for Incidence of Cytoplasmic Vacuolation (All
Severity Grades) in the Thyroid of F0 Female S-D Rats Exposed to Technical Toxaphene
in the Diet for 25-29 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0052
92.46
1.3 x 1014
NDr
Logistic
0.0357
88.06
3.37599
1.78465
LogLogistic0
0.0463
87.36
1.54574
0.42723
LogProbit0
0.0252
88.84
5.17203
2.17277
Multistage (l-degree)d
0.0399
87.76
2.42250
1.04272
Multistage (2-degree)d
0.0399
87.76
2.42250
1.04272
Multistage (3-degree)d
0.0399
87.76
2.42250
1.04272
Multistage (4-degree)d
0.0399
87.76
2.42250
1.04272
Probit
0.0359
88.05
3.33414
1.78355
Weibullb
0.0399
87.76
2.42255
1.04272
Highest dose dropped
Gammab
0.324
57.64
0.18336
0.09471
Logistic
0.2603
58.22
0.34526
0.21333
LogLogistic0'e
0.3954
57.19
0.10299
0.03668
LogProbitd
0.2238
58.46
0.35276
0.17064
Multistage (l-degree)d
0.324
57.64
0.18336
0.09471
Multistage (2-degree)d
0.324
57.64
0.18336
0.09471
Multistage (3-degree)d
0.324
57.64
0.18336
0.09471
Probit
0.2629
58.19
0.33560
0.21653
Weibullb
0.324
57.64
0.18336
0.09471
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; NDr = not determined; S-D = Sprague-Dawley.
170
Toxaphene
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Log-Logistic Model h Br 1R of 10% Extra Risk for the BMD and 0. 95 Lower Confidence Limnt for the BMDL
0.8
0-6
0-4
0.2
Log-Logistic
BMDL
9MD
0.5
15
15:51 03/07 2017
dose
Figure C-15. LogLogistic Model for Increased Incidence of Cytoplasmic Vacuolation
(All Severity Grades) in the Thyroid of FO Female S-D Rats Exposed to Technical
Toxaphene in the Diet for 25-29 Weeks (Highest Dose Dropped) CChu et al., 1988)
Text output for Figure C-15:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_thy_cyt_vac_F0_female_rats_high_d
rop_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_thy_cyt_vac_F0_female_rats_high_d
rop_HEDs_Lnl-BMR10-Restrict.pit
Tue Mar 07 16:17:01 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 4
Total number of records with missing values = 0
171
Toxaphene
-------
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07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.117 647
intercept = 0.140388
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
background intercept
background 1 -0.55
intercept -0.55 1
Parameter Estimates
Interval
Variable
Limit
background
0.34634
intercept
1.43443
slope
Estimate
0.172143
0.0758934
1
Std. Err.
0.0888777
0.693144
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-0.00205448
-1.28264
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-25.7265
-26.5956
-30.7564
# Param's
4
2
1
Deviance Test d.f.
1.7382
10.0599
P-value
0.4193
0. 01806
AIC:
57.1911
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.1721 2.926 2.000 17.000 -0.595
0.0830 0.2402 2.402 4.000 10.000 1.183
172
Toxaphene
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07-31-2018
0.4400 0.4386 4.386 4.000 10.000 -0.246
1.9000 0.7286 7.286 7.000 10.000 -0.203
Chi^2 = 1.86 d.f. = 2 P-value = 0.3954
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.102991
BMDL = 0.03667 66
Increased Incidence of Minimal to Mild Cytoplasmic Vacuolation (the Only Severity Grade
Observed) in the Thyroid of Fla Female S-D Rats Exposed to Technical Toxaphene in the
Diet for 34 Weeks (Cfau et al., 1988)
The procedure outlined above was applied to the data for increased incidence of minimal
to mild cytoplasmic vacuolation in the thyroid of Fla female S-D rats exposed to toxaphene for
34 weeks (Chit et al. 1988) (see Table C-35). Table C-36 summarizes the BMD modeling
results. None of the models provided an adequate fit using the full dose range. However, after
dropping the highest dose, the LogLogistic model provided an adequate fit to the data. Thus, a
BMDLio of 0.0059 mg/kg-day was calculated for this data set (see Figure C-16). However, the
modeling results for this endpoint are not considered reliable because all response levels were
considered in excess of the BMR, leaving no data to inform the shape of the dose-response curve
in the low-dose region and requiring extrapolation far below the observable range in order to
estimate the BMD (U.S. EPA. 2012b).
Table C-35. Incidence of Minimal to Mild Cytoplasmic Vacuolation (the Only Severity
Grade Observed) in the Thyroid of Fla Female S-D Rats Exposed to Technical Toxaphene
in the Diet for 34 Weeks"
HED (mg/kg-d)
0
0.089
0.44
2.2
11
Sample size
17
10
10
10
17
Incidence
1
5
8
9
6
aCfau et al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
173
Toxaphene
-------
FINAL
07-31-2018
Table C-36. BMD Modeling Results for Incidence of Minimal to Mild Cytoplasmic
Vacuolation (the Only Severity Grade Observed) in the Thyroid of Fla Female S-D Rats
Exposed to Technical Toxaphene in the Diet for 34 Weeks
Model
X2 Goodness-of-Fit
/j-Valuc11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0
94.16
2.1 x 1013
NDr
Logistic
0
92.16
1,100
3.46836
LogLogistic0
0
92.16
4.0 x 106
2.09089
LogProbit0
0
92.16
392.557
6.09914
Multistage (l-degree)d
0
91.97
NDr
NDr
Multistage (2-degree)d
0.0001
90.16
NDr
NDr
Multistage (3-degree)d
0.0001
90.16
NDr
NDr
Multistage (4-degree)d
0.0001
90.16
NDr
NDr
Probit
0
92.16
1,100
3.46537
Weibullb
0
92.16
1,100
NDr
Highest dose dropped
Gammab
0.0052
49.22
0.05563
0.03152
Logistic
0.0041
54.11
0.16463
0.09128
LogLogistic0'e
0.7355
42.48
0.01379
0.00593
LogProbitd
0.0007
47.76
0.05450
0.03032
Multistage (l-degree)d
0.0052
49.22
0.05563
0.03152
Multistage (2-degree)d
0.0052
49.22
0.05563
0.03152
Multistage (3-degree)d
0.0052
49.22
0.05563
0.03152
Probit
0.004
54.59
0.19185
0.12402
Weibullb
0.0052
49.22
0.05563
0.03152
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; NDr = not determined; S-D = Sprague-Dawley.
174
Toxaphene
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Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
0.6
o
oj
<
£Z
o
~o
0.4
0.2
3MD
0
0.5
1
1.5
2
dose
09:47 03/09 2017
Figure C-16. LogLogistic Model for Increased Incidence of Minimal to Mild Cytoplasmic
Vacuolation (the Only Severity Grade Observed) in the Thyroid of Fla Female S-D Rats
Exposed to Technical Toxaphene in the Diet for 34 Weeks (Highest Dose Dropped)
(Chu et al.. 1988)
Text output for Figure C-16:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_thy_cyt_vac_Fla_female_rats_high
drop_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_thy_cyt_vac_Fla_female_rats_high
drop_HEDs_Lnl-BMR10-Restrict.pit
Thu Mar 09 09:47:06 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
175
Toxaphene
-------
FINAL
07-31-2018
Total number of observations = 4
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.0588235
intercept = 1.65075
slope = 1
the user,
background
intercept
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -slope
have been estimated at a boundary point, or have been specified by
and do not appear in the correlation matrix )
background intercept
1 -0.24
-0.24 1
Interval
Variable
Limit
background
0.176568
intercept
3.08083
slope
Estimate
0.0609484
2.08662
1
Parameter Estimates
Std. Err.
0.0589907
0.507257
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-0.0546712
1.09242
Model
Full model
Fitted model
Reduced model
Analysis of Deviance Table
#
Log(likelihood)
-18.9895
-19.2407
-32.5673
Param's
4
2
1
Deviance Test d.f.
0.502347
27.1555
P-value
0.7779
<.0001
AIC:
42.4814
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
176
Toxaphene
-------
0.0000
0.0890
0.4400
2 .2000
Chi^2 =0.61
0.0609
0.4531
0.7934
0.9499
d.f. = 2
1.036 1.000 17.000
4.531 5.000 10.000
7.934 8.000 10.000
9.499 9.000 10.000
P-value = 0.7355
-0.037
0.298
0. 052
-0.722
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0137895
BMDL = 0.00593414
FINAL
07-31-2018
Increased Relative Liver Weight in Female Beagle Dogs Administered Technical
Toxaphene in Gelatin Capsules for 13 Weeks (Cfau et al., 1986)
The procedure outlined above was applied to the data for increased relative liver weight
in female Beagle dogs administered technical toxaphene in gelatin capsules for 13 weeks (Chu et
al. 1986) (see Table C-37). Table C-38 summarizes the BMD modeling results. The constant
variance model provided adequate fit to the data, and all models provided adequate fit to means.
However, all models besides the Hill model failed visual inspection of the model fit. While the
Hill model provided an adequate model fit, the BMD was more than an order of magnitude
higher than the corresponding BMDL. Thus, none of the modeling results for this endpoint are
considered reliable.
Table C-37. Relative Liver Weights of Female Beagle Dogs Administered Technical
Toxaphene in Gelatin Capsules for 13 Weeks"
HED (mg/kg-d)
0
0.1
0.82
1.8
Sample size
6
6
6
6
Mean
3.0
3.4
3.6
3.9
SD
0.60
0.40
0.31
0.46
aChu et al. (1986).
HED = human equivalent dose; SD = standard deviation.
177
Toxaphene
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07-31-2018
Table C-38. BMD Model Predictions for Relative Liver Weight in Female Beagle Dogs
Administered Technical Toxaphene by Gelatin Capsule for 13 Weeks11
Model
Variance
/>-Valucb
Mean
/>-Valucb
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Constant variance
Exponential (model 2)°
0.44
0.29
-9.71
0.82698
0.55252
Exponential (model 3)°
0.44
0.29
-9.71
0.82698
0.55252
Exponential (model 4)°
0.44
0.19
-8.47
0.36171
0.00076
Exponential (model 5)°
0.44
0.19
-8.47
0.36171
0.00115
Hillc
0.44
0.31
-9.16
0.07882
0.00322
Linear"1
0.44
0.31
-9.84
0.76956
0.48722
Polynomial (2-degree)d
0.44
0.31
-9.84
0.76956
0.48722
Polynomial (3-degree)d
0.44
0.31
-9.84
0.76956
0.48722
Power0
0.44
0.31
-9.84
0.76956
0.48722
Timetal. (1986).
bValues <0.10 fail to meet conventional goodness-of-fit criteria.
Coefficients restricted to be positive.
dPower restricted to >1.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit.
Increased Absolute Liver Weight in F0 Female S-D Rats Exposed to Technical Toxaphene
in the Diet for 25-29 Weeks (Cfau et al., 1988)
The procedure outlined above was applied to the data for increased absolute liver weight
in F0 female S-D rats administered technical toxaphene in the diet for 25-29 weeks (Chu et al.
1988) (see Table C-39). Table C-40 summarizes the BMD modeling results. The constant
variance model did not fit the variance data, but the nonconstant variance model did. With the
nonconstant variance model applied using the full dose range, none of the models provided
adequate fit to the means. However, after dropping the highest dose, the Exponential Model 4
provided an adequate fit to the data. Thus, a BMDLio of 0.028 mg/kg-day was calculated for this
data set (see Figure C-17).
178
Toxaphene
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07-31-2018
Table C-39 Absolute Liver Weight in F0 Female S-D Rats Exposed to Technical
Toxaphene in the Diet for 25-29 Weeks"
HED (mg/kg-d)
0
0.083
0.44
1.9
11
Sample size
23
20
20
17
18
Mean
11.6
12.9
13.8
13.6
16.6
SD
2.0
2.0
3.3
2.7
3.2
"Clin et al. (19881.
HED = human equivalent dose; S-D = Sprague-Dawley; SD = standard deviation.
Table C-40. BMD Model Predictions for Absolute Liver Weight in F0 Female S-D Rats
Exposed to Technical Toxaphene in the Diet for 25-29 Weeks"
Model
Variance
/>-Valucb
Mean
/>-Valucb
AIC
BMD io
(mg/kg-d)
BMDLio
(mg/kg-d)
Constant variance
Exponential (model 2)°
0.05
0.07
298.07
3.87089
3.02370
Exponential (model 3)°
0.05
0.07
298.07
3.87089
3.02370
Exponential (model 4)°
0.05
0.06
298.82
1.45755
0.52458
Exponential (model 5)°
0.05
0.06
298.82
1.45755
0.52458
Hillc
0.05
0.06
298.65
1.22112
0.17175
Linear"1
0.05
0.08
297.88
3.49605
2.62151
Polynomial (2-degree)d
0.05
0.08
297.88
3.49605
2.62151
Polynomial (3-degree)d
0.05
0.08
297.88
3.49605
2.62151
Power0
0.05
0.08
297.88
3.49605
2.62151
Nonconstant variance
Exponential (model 2)°
0.38
0.01
298.79
3.83243
2.91741
Exponential (model 3)°
0.38
0.01
298.79
3.83243
2.91741
Exponential (model 4)°
0.38
0.01
298.76
1.04659
0.03477
Exponential (model 5)°
0.38
0.01
298.76
1.04659
0.03477
Hillc
0.38
0.05
295.06
0.12875
0.04087
Linear"1
0.38
0.01
298.54
3.44816
2.50604
Polynomial (2-degree)d
0.38
0.01
298.54
3.44816
2.50604
Polynomial (3-degree)d
0.38
0.01
298.54
3.44816
2.50604
Power0
0.38
0.01
298.54
3.44816
2.50604
179
Toxaphene
-------
FINAL
07-31-2018
Table C-40. BMD Model Predictions for Absolute Liver Weight in F0 Female S-D Rats
Exposed to Technical Toxaphene in the Diet for 25-29 Weeks"
Model
Variance
/>-Valucb
Mean
/>-Valucb
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Nonconstant variance (highest dose dropped)
Exponential (model 2)°
0.43
0.003
238.52
1.64649
0.84705
Exponential (model 3)°
0.43
0.003
238.52
1.64649
0.84705
Exponential (model 4)c'e
0.43
0.41
229.44
0.09870
0.02834
Exponential (model 5)°
0.43
NAf
231.27
0.09592
0.02994
Hillc
0.43
NAf
231.27
0.08661
0.01660
Linear11
0.43
0.003
238.37
1.56086
0.76143
Polynomial (2-degree)d
0.43
0.003
238.37
1.56086
0.76143
Polynomial (3-degree)d
0.43
0.003
238.37
1.56086
0.76143
Power0
0.43
0.003
238.37
1.56086
0.76143
Timetal. (1988).
bValues <0.10 fail to meet conventional goodness-of-fit criteria.
Coefficients restricted to be positive.
dPower restricted to >1.
Selected model.
fDegrees of freedom for Test 4 [means />-valuc| are <0; the x2 test for fit is not valid.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; NA = not applicable; S-D = Sprague-Dawley.
180
Toxaphene
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Exponential 4 Model, with BMR of 0.1 Rel. Dev. for the BMD and 0.95 Lower Confidence Limit for the BMDL
15
14
12
11
Exponential 4
BMDL
BMD
0.5
1.5
dose
10:24 03/09 2017
Figure C-17. Exponential Model 4 for Increased Absolute Liver Weight in FO Female S-D
Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks (Chu et al., 1986)
Text output for Figure C-17:
Exponential Model. (Version: 1.10; Date: 01/12/2015)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/exp_Chu_198 8_abs_liv_F0_females_high_drop
_HEDs_Exp-ModelVariance-BMR10-Up.(d)
Gnuplot Plotting File:
Thu Mar 09 10:24:03 2017
BMDS Model Run
The form of the response function by Model:
Model 2
Model 3
Model 4
Model 5
Y[dose] = a * exp{sign * b * dose}
Y[dose] = a * exp{sign * (b * dose)Ad}
Y[dose] = a * [c-(c-l) * exp{-b * dose}]
Y[dose] = a * [c-(c-l) * exp{-(b * dose)Ad}]
Note: Y[dose] is the median response for exposure
sign = +1 for increasing trend in data;
sign = -1 for decreasing trend.
dose;
Model 2 is nested within Models 3 and 4.
Model 3 is nested within Model 5.
Model 4 is nested within Model 5.
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Dependent variable = Mean
Independent variable = Dose
Data are assumed to be distributed: normally
Variance Model: exp(lnalpha +rho *ln(Y[dose]))
The variance is to be modeled as Var(i) = exp(lalpha + log(mean(i)) * rho)
Total number of dose groups = 4
Total number of records with missing values = 0
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
MLE solution provided: Exact
Initial Parameter Values
Variable Model 4
lnalpha -11.1217
rho 5.04 09
a 11.02
b 0.881981
c 1.31488
d 1 Specified
Parameter Estimates
Variable Model 4
lnalpha
rho
a
b
c
-11.4297
5.1557
11.6628
7.87633
1.18504
Std. Err.
6.55605
2 .56976
0.384103
4.8735
0.0572076
Table of Stats From Input Data
Dose
0
0. 083
0.44
1.9
23
20
20
17
Obs Mean
11.6
12.9
13. 8
13. 6
Obs Std Dev
2
2
3.3
2.7
Dose
0
0. 083
0.44
1.9
Estimated Values of Interest
Est Mean Est Std Scaled Residual
11.66
12.7
13.75
13.82
1.854
2 .309
2.836
2.872
-0.1624
0.3903
0. 07337
-0.3171
Other models for which likelihoods are calculated:
Model A1: Yij = Mu(i) + e(ij)
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Var{e(ij)} = SigmaA2
Model A2: Yij
Var{e(ij)}
Model A3: Yij
Var{e(ij)}
Model R: Yij
Var{e(ij)}
Mu(i) + e(i j)
Sigma(i)A2
Mu(i) + e(i j)
exp(lalpha + log(mean(i)) * rho)
Mu + e(i)
SigmaA2
Model
A1
A2
A3
R
4
Likelihoods of Interest
Log (likelihood) DF
-112.2946 5
-108.5167 8
-109.3695 6
-117.1847 2
-109.7213 5
AIC
234.5893
233.0334
230.739
238.3694
229.4426
Additive constant for all log-likelihoods = -73.52. This constant added to the
above values gives the log-likelihood including the term that does not
depend on the model parameters.
Explanation of Tests
Does response and/or variances differ among Dose levels? (A2 vs. R)
Are Variances Homogeneous? (A2 vs. Al)
Are variances adeguately modeled? (A2 vs. A3)
Test 6a: Does Model 4 fit the data? (A3 vs 4)
Test
1:
Test
2 :
Test
3:
Test
Test 1
Test 2
Test 3
Test 6a
Tests of Interest
-2*log(Likelihood Ratio)
17.34
7.556
1.706
0.7036
D. F.
6
3
2
1
p-value
0.008125
0.05614
0.4262
0.4016
The p-value for Test 1 is less than .05. There appears to be a
difference between response and/or variances among the dose
levels, it seems appropriate to model the data.
The p-value for Test 2 is less than .1. A non-homogeneous
variance model appears to be appropriate.
The p-value for Test 3 is greater than .1. The modeled
variance appears to be appropriate here.
The p-value for Test 6a is greater than .1. Model 4 seems
to adeguately describe the data.
Benchmark Dose Computations:
Specified Effect = 0.100000
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Risk Type = Relative deviation
Confidence Level = 0.950000
BMD = 0.0987043
BMDL = 0.0283373
Increased Relative Liver Weight in Male B6C3Fi Mice Administered Technical Toxaphene
in the Diet for 28 Days (Wang et al., 2015)
The procedure outlined above was applied to the data for increased relative liver weight
in male B6C3Fi mice administered technical toxaphene in the diet for 28 days (Wang et al.
2015) (see Table C-41). Table C-42 summarizes the BMD modeling results. The constant
variance model did not fit the variance data. With the nonconstant variance model applied, all
models provided only a marginal fit to the data (variancep = 0.09). Thus, these data are not
amenable to BMD modeling.
Table C-41. Relative Liver Weight in Male B6C3Fi Mice Administered Technical
Toxaphene in the Diet for 28 Days"
HED (mg/kg-d)
0
0.1
0.85
8.51
Sample size
12
12
12
12
Mean
5.3
5.3
5.6
8.4
SD
0.27
0.24
0.16
0.64
aWang et al. (2015).
HED = human equivalent dose; SD = standard deviation.
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Table C-42. BMD Model Predictions for Increased Relative Liver Weight in Male B6C3Fi
Mice Administered Technical Toxaphene in the Diet for 28 Days
Model
Variance
/>-Value"
Mean
/j-Value11
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Constant variance
Exponential (model 2)b
<0.0001
0.86
-43.76
1.8
1.7
Exponential (model 3)b
<0.0001
0.86
-43.76
1.8
1.7
Exponential (model 4)b
<0.0001
0.80
-42.00
1.4
0.81
Exponential (model 5)b
<0.0001
NA°
-40.06
0.97
0.85
Hillc
<0.0001
NA°
-40.06
0.88
0.85
Linear11
<0.0001
0.97
-44.00
1.4
1.3
Polynomial (2-degree)d
<0.0001
0.97
-44.00
1.4
1.3
Polynomial (3-degree)d
<0.0001
0.97
-44.00
1.4
1.3
Powerb
<0.0001
0.97
-44.00
1.4
1.3
Nonconstant variance
Exponential (model 2)b
0.09
0.80
-63.27
1.8
1.6
Exponential (model 3)b
0.09
0.80
-63.27
1.8
1.6
Exponential (model 4)b
0.09
0.61
-61.47
1.4
1.0
Exponential (model 5)b
0.09
NA
-59.72
0.94
0.86
Hillb
0.09
NA
89.06
NDr
NDr
Linear11
0.09
0.88
-63.47
1.4
1.3
Polynomial (2-degree)d
0.09
0.65
-61.51
1.5
1.3
Polynomial (3-degree)d
0.09
0.65
-61.51
1.5
1.3
Powerb
0.09
0.66
-61.53
1.5
1.3
aValues <0.10 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
Degrees of freedom for Test 4 [means /?-value] are <0; the x2 test for fit is not valid.
Coefficients restricted to be negative.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; NA = not applicable; NDr = not determined.
Increased Mean Bromodeoxyuridine Labeling Index in the Liver of Male B6C3Fi Mice
Administered Technical Toxaphene in the Diet for 28 Days (Wang et al., 2015)
The procedure outlined above was applied to the data for increased mean
bromodeoxyuridine (BrdU) labeling index in the liver of male B6C3Fi mice administered
technical toxaphene in the diet for 28 days (Wang et al. 2015) (see Table C-43). Table C-44
summarizes the BMD modeling results. The constant variance model did not fit the variance
data. With a nonconstant variance model applied, the modeled variance fit the data; however,
none of the models provided an adequate fit to the means. The high dose was not dropped
because findings at the mid dose were not statistically significant. Thus, these data are not
amenable to BMD modeling.
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Table C-43. Mean BrdU Labeling Index in the Liver of Male B6C3Fi Mice Administered
Technical Toxaphene in the Diet for 28 Days"
HED (mg/kg-d)
0
0.1
0.85
8.51
Sample size
5
5
5
5
Mean
0.9
1
1.7
3.9
SEM
0.2
0.7
0.7
1.7
SDb
0.4
1.6
1.6
3.8
"Wang et al. (2015).
bCalculated using U.S. EPA BMDS (Version 2.5).
BMDS = Benchmark Dose Software; BrdU = bromodeoxyuridine; HED = human equivalent dose; SD = standard
deviation; SEM = standard error of the mean.
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Table C-44. BMD Model Predictions for Mean BrdU Labeling Index in the Liver of Male
B6C3Fi Mice Administered Technical Toxaphene in the Diet for 28 Days
Model
Variance
/>-Valuca
Mean
/>-Valuca
AIC
BMDisd
(mg/kg-d)
BMDLisd
(mg/kg-d)
Constant variance
Exponential (model 2)b
0.0004
0.85
53.76
7.0
5.4
Exponential (model 3)b
0.0004
0.85
53.76
7.0
5.4
Exponential (model 4)b
0.0004
1.00
55.44
2.9
0.46
Exponential (model 5)b
0.0004
NA°
57.44
2.8
0.46
Hillb
0.0004
NA°
57.44
3.1
0.30
Linear11
0.0004
0.90
53.64
6.0
3.6
Polynomial (2-degree)d
0.0004
0.90
53.64
6.0
3.6
Polynomial (3-degree)d
0.0004
0.90
53.64
6.0
3.6
Powerb
0.0004
0.90
53.64
6.0
3.6
Nonconstant variance
Exponential (model 2)b
0.42
0.03
45.71
4.7
3.0
Exponential (model 3)b
0.42
0.03
45.71
4.7
3.0
Exponential (model 4)b
0.42
0.02
45.93
0.83
0.002
Exponential (model 5)b
0.42
0.02
45.93
0.83
0.004
Hillb
0.42
0.06
44.34
0.05
NDr
Linear11
0.42
0.05
44.98
2.8
1.2
Polynomial (2-degree)d
0.42
0.05
44.98
2.8
1.2
Polynomial (3-degree)d
0.42
0.05
44.98
2.8
1.2
Powerb
0.42
0.05
44.98
2.8
1.2
aValues <0.10 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
Degrees of freedom for Test 4 [means /?-value] are <0; the x2 test for fit is not valid.
Coefficients restricted to be negative.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; BrdU = bromodeoxyuridine; NA = not applicable;
NDr = not determined; SD = standard deviation.
Increased Incidence of Accented Zonation (All Severity Scores) in the Liver of Female S-D
Rats Administered Technical Toxaphene in the Diet for 13 Weeks CChu et al., 1986)
The procedure outlined above was applied to the data for increased incidence of accented
zonation (all severity grades) in the liver of female S-D rats exposed to toxaphene for 13 weeks
(Chu et al. 1986) (see Table C-45). Table C-46 summarizes the BMD modeling results. All
models except the LogProbit, and Multistage 3- and 4-degree models provided adequate fit to the
full data set. BMDLs for models providing adequate fit were not sufficiently close (differed by
>threefold), so the model with the lowest BMDL was selected (LogLogistic). Thus, the BMDLio
of 0.024 mg/kg-day from this model is selected for this endpoint (see Figure C-18). However,
the modeling results for this endpoint are not considered reliable because all response levels were
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considered far in excess of the BMR, leaving no data to inform the shape of the dose-response
curve in the low-dose region and requiring extrapolation far below the observable range in order
to estimate the BMD (U.S. EPA, 2012b).
Table C-45. Incidence of Accented Zonation (All Severity Scores) in the Liver of Female
S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
4
5
7
10
Timetal. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
Table C-46. BMD Modeling Results for Incidence of Accented Zonation (All Severity
Scores) in the Liver of Female S-D Rats Administered Technical Toxaphene in the Diet for
13 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.1689
50.13
0.22549
0.11690
Logistic
0.1429
51.28
0.48347
0.29377
LogLogistic0'd
0.2561
46.26
0.04914
0.02418
LogProbit0
0.0357
54.87
1.90772
0.16655
Multistage (l-degree)e
0.1689
50.13
0.22549
0.11690
Multistage (2-degree)e
0.1689
50.13
0.22549
0.11690
Multistage (3-degree)e
0.0808
52.13
0.22731
0.11692
Multistage (4-degree)e
0.0818
52.12
0.22945
0.11708
Probit
0.1433
51.24
0.47437
0.30331
Weibullb
0.1689
50.13
0.22549
0.11690
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
"Betas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
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Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
T3
0.6
CD
(J
CD
<
£=
O
u
CD
0.4
Ll_
0.2
3MDLBMD
0
2
4
6
8
10
12
14
dose
14:44 06/28 2017
Figure C-18. LogLogistic Model for Increased Incidence of Accented Zonation (All Severity
Scores) in the Liver of Female S-D Rats Administered Technical Toxaphene in the Diet for
13 Weeks (Chu et al.. 1986)
Text output for Figure C-18:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_liv_acc_zon_female_rats_HEDs_Lnl-
BMRIO-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_liv_acc_zon_female_rats_HEDs_Lnl-
BMR10-Restrict.pit
Thu Aug 17 08:50:16 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
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Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 0.11496
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
1.65006
slope
Estimate
0
0.815959
1
Std. Err.
NA
0.42557
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
-0.018143
Model
Full model
Fitted model
Reduced model
Analysis of Deviance Table
#
Log(likelihood)
-19.7702
-22.1298
-34.6173
Param's
5
1
1
Deviance Test d.f.
4.71905
29.6942
P-value
0.3174
<.0001
AIC:
46.2595
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000
0.1100 0.1992
0.5800 0.5674
2.8100 0.8640
14.0000 0.9694
0.000 0.000
1.992 4.000
5.674 5.000
8.640 7.000
9.694 10.000
10.000 0.000
10.000 1.590
10.000 -0.430
10.000 -1.513
10.000 0.562
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Chi^2 = 5.32 d.f. = 4 P-value = 0.2561
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.049135
BMDL = 0.0241785
Increased Incidence of Anisokaryosis (All Severity Scores) in the Liver of Male S-D Rats
Administered Technical Toxaphene in the Diet for 13 Weeks (Chu et al., 1986)
The procedure outlined above was applied to the data for increased incidence of
anisokaryosis (all severity grades) in the liver of male S-D rats exposed to toxaphene for
13 weeks (Chu et al, 1986) (see Table C-47). Table C-48 summarizes the BMD modeling
results. All models provided adequate fit to the full data set. BMDLs for models providing
adequate fit were not sufficiently close (differed by >threefold), so the model with the lowest
BMDL was selected (LogLogistic). Thus, the BMDLio of 0.009 mg/kg-day from this model is
selected for this endpoint (see Figure C-19). However, the modeling results for this endpoint are
not considered reliable because all response levels were considered far in excess of the BMR,
leaving no data to inform the shape of the dose-response curve in the low-dose region and
requiring extrapolation far below the observable range in order to estimate the BMD (U.S. EPA.
2012b).
Table C-47. Incidence of Anisokaryosis (All Severity Scores) in the Liver of Male S-D Rats
Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.090
0.46
2.2
11.8
Sample size
10
10
10
10
10
Incidence
2
4
8
9
10
aChu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
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Table C-48. BMD Modeling Results for Incidence of Anisokaryosis (All Severity Scores) in
the Liver of Male S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks
Model
X2 Goodness-of-Fit
/>-Value"
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.3988
46.76
0.07856
0.04089
Logistic
0.2301
48.52
0.16410
0.08985
LogLogistic0'd
0.7753
46.56
0.02767
0.00904
LogProbit0
0.3857
46.21
0.10154
0.04679
Multistage (1-degree)6
0.3988
46.76
0.07856
0.04089
Multistage (2-degree)6
0.3988
46.76
0.07856
0.04089
Multistage (3-degree)6
0.3988
46.76
0.07856
0.04089
Multistage (4-degree)e
0.3988
46.76
0.07856
0.04089
Probit
0.2112
48.82
0.18718
0.11709
Weibullb
0.3988
46.76
0.07856
0.04089
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
eBetas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
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Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
1
0.8
0.2
0
Log-Logistic
10
12
14
dose
14:32 06/28 2017
Figure C-19. LogLogistic Model for Increased Incidence of Anisokaryosis (All Severity
Scores) in the Liver of Male S-D Rats Administered Technical Toxaphene in the Diet for
13 Weeks (Chu et al.. 1986)
Text output for Figure C-19:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_liv_aniso_male_rats_HEDs_Lnl-BMRl
O-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_liv_aniso_male_rats_HEDs_Lnl-BMRl
O-Restrict.pit
Thu Jun 29 13:25:35 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
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Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0.2
intercept = 0.913084
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
background intercept slope
background 1 -0.3 0.22
intercept -0.3 1 0.5
slope 0.22 0.5 1
Parameter Estimates
Interval
Variable
Limit
background
0.434678
intercept
3.05993
slope
1.93638
Estimate
0.19553
1.65094
1.07272
95.0% Wald Confidence
Std. Err. Lower Conf. Limit Upper Conf.
0.122016 -0.0436177
0.718885 0.241955
0.440654 0.209054
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-19.989
-20.2804
-32.0518
# Param's
5
3
1
Deviance Test d.f.
0.582793
24.1256
P-value
0.7472
<.0001
AIC:
46.5608
Dose
Est. Prob.
Goodness of Fit
Expected Observed Size
Scaled
Residual
0.0000
0.0900
0.4600
2 .2000
11.8000
0.1955
0. 4228
0.7537
0.9388
0.9892
1.955
4.228
7.537
9.388
9.892
2.000
4.000
8.000
9.000
10.000
10.000
10.000
10.000
10.000
10.000
0. 036
-0.146
0.340
-0.512
0.330
Chi^2 = 0.51
d.f. = 2
P-value = 0.7753
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Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0276729
BMDL = 0.00904366
Increased Incidence of Anisokaryosis (All Severity Scores) in the Liver of Female S-D Rats
Administered Technical Toxaphene in the Diet for 13 Weeks (Chu et al., 1986)
The procedure outlined above was applied to the data for increased incidence of
anisokaryosis (all severity grades) in the liver of female S-D rats exposed to toxaphene for
13 weeks (Chu et al. 1986) (see Table C-49). Table C-50 summarizes the BMD modeling
results. All models provided adequate fit to the full data set. BMDLs for models providing
adequate fit were not sufficiently close (differed by >threefold), so the model with the lowest
BMDL was selected (LogLogistic). Thus, the BMDLio of 0.003 mg/kg-day from this model is
selected for this endpoint (see Figure C-20). However, the modeling results for this endpoint are
not considered reliable because all response levels were considered far in excess of the BMR,
leaving no data to inform the shape of the dose-response curve in the low-dose region and
requiring extrapolation far below the observable range in order to estimate the BMD (U.S. EPA.
2012b).
Table C-49. Incidence of Anisokaryosis (All Severity Scores) in the Liver of Female S-D
Rats Administered Technical Toxaphene in the Diet for 13 Weeks"
HED (mg/kg-d)
0
0.11
0.58
2.81
14
Sample size
10
10
10
10
10
Incidence
0
6
9
10
10
"Chu et al. (1986).
HED = human equivalent dose; S-D = Sprague-Dawley.
195
Toxaphene
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07-31-2018
Table C-50. BMD Modeling Results for Incidence of Anisokaryosis (All Severity Scores) in
the Liver of Female S-D Rats Administered Technical Toxaphene in the Diet for 13 Weeks
Model
X2 Goodness-of-Fit
/>-Value"
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.7844
23.45
0.01918
0.01157
Logistic
0.1402
31.15
0.06062
0.03405
LogLogistic0'd
0.9758
24.28
0.01530
0.00295
LogProbit0
0.9035
22.80
0.02891
0.01635
Multistage (1-degree)6
0.7844
23.45
0.01918
0.01157
Multistage (2-degree)6
0.7844
23.45
0.01918
0.01157
Multistage (3-degree)6
0.7841
23.45
0.01917
0.01157
Multistage (4-degree)e
0.7844
23.45
0.01918
0.01157
Probit
0.1317
31.26
0.06381
0.04034
Weibullb
0.7844
23.45
0.01918
0.01157
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dSelected model.
eBetas restricted to >0.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
196
Toxaphene
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07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
1
0.8
0.2
0
Log-Logistic
3MDLBMD
10
12
14
dose
14:32 06/28 2017
Figure C-20. LogLogistic Model for Increased Incidence of Anisokaryosis (All Severity
Scores) in the Liver of Female S-D Rats Administered Technical Toxaphene in the Diet for
13 Weeks (Chu et al.. 1986)
Text output for Figure C-20:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_liv_aniso_female_rats_HEDs_Lnl-BM
RIO-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 6_liv_aniso_female_rats_HEDs_Lnl-BM
RIO-Restrict.pit
Thu Jun 29 13:42:27 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 5
Total number of records with missing values = 0
197
Toxaphene
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07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 1.16344
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 0.8 9
slope 0.89 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
5 .57647
slope
2 .51284
Estimate
0
3.20789
1.29317
Std. Err.
NA
1.20848
0.622291
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
0.839309
0.0735049
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-9.98095
-10.1422
-30.5432
# Param's Deviance Test d.f. P-value
5
2 0.322451 3 0.955E
1 41.1245 4 <.0001
AIC:
24.2843
Goodness of Fit
Scaled
Dose Est._Prob. Expected Observed Size Residual
0.0000 0.0000 0.000 0.000 10.000 0.000
0.1100 0.5875 5.875 6.000 10.000 0.081
198
Toxaphene
-------
Benchmark Dose Computation
Specified effect = 0.1
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0153027
BMDL = 0.00294711
FINAL
07-31-2018
0.5800
0.9244
9.244
9.000
10.000
-0.292
2.8100
0.9895
9.895
10.000
10.000
0.326
14.0000
0.9987
9.987
10.000
10.000
0.115
Chi^2 = 0.21
d.f. = 3
P-
-value = 0.
, 9758
Increased Incidence of Anisokaryosis (All Severity Grades) in the Liver of F0 Female S-D
Rats Exposed to Technical Toxaphene in the Diet for 25-29 Weeks CChu et al., 1988)
The procedure outlined above was applied to the data for increased incidence of
anisokaryosis (all severity grades) in the liver of female F0 S-D rats exposed to technical
toxaphene for 25-29 weeks (Chu et al. 1988) (see Table C-51). Table C-52 summarizes the
BMD modeling results. None of the models provided adequate fit to the full data set or the data
set with the highest dose dropped. With the two highest doses dropped, only the Logistic,
Multistage 1-degree, and Probit models provided adequate fit to the data. For these three
models, BMDLs were considered to be sufficiently close (differed by
-------
FINAL
07-31-2018
Table C-52. BMD Modeling Results for Incidence of Anisokaryosis (All Severity Grades)
in the Liver of F0 Female S-D Rats Exposed to Technical Toxaphene in the Diet for
25-29 Weeks
Model
X2 Goodness-of-Fit
/>-Valuca
AIC
BMDio
(mg/kg-d)
BMDLio
(mg/kg-d)
Gammab
0.0014
64.00
0.17918
0.07492
Logistic
0.0012
64.52
0.28285
0.13244
LogLogistic0
0
57.95
0.00956
0.00309
LogProbit0
0.0005
65.72
0.28642
0.07827
Multistage (l-degree)d
0.0014
64.00
0.17919
0.07492
Multistage (2-degree)d
0.0014
64.00
0.17919
0.07492
Multistage (3-degree)d
0.0004
66.00
0.18248
0.07495
Multistage (4-degree)d
0.0004
65.98
0.18653
0.07511
Probit
0.0012
64.57
0.31339
0.15386
Weibullb
0.0014
64.00
0.17919
0.07492
Highest dose dropped
Gammab
0.0004
63.97
0.18708
0.07521
Logistic
0.0004
64.50
0.29265
0.13279
LogLogistic0
0
57.75
0.01002
0.00318
LogProbit0
0.0002
65.53
0.37082
0.08156
Multistage (l-degree)d
0.0004
63.97
0.18708
0.07521
Multistage (2-degree)d
0.0004
63.97
0.18708
0.07521
Multistage (3-degree)d
0.0004
63.97
0.18708
0.07521
Probit
0.0003
64.57
0.31405
0.15387
Weibullb
0.0004
63.97
0.18708
0.07521
Two highest doses dropped
Gammab
NA
34.56
0.02677
0.00322
Logistic6
0.9949
32.56
0.01252
0.00715
LogLogistic0
NA
34.56
0.06050
0.00071
LogProbitd
NA
34.56
0.04308
0.00514
Multistage (l-degree)d
0.937
32.57
0.00646
0.00322
Multistage (2-degree)d
NA
36.56
0.03273
0.00322
Probit
1
32.56
0.01225
0.00759
Weibullb
NA
34.56
0.01513
0.00322
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
"Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; NA = not applicable; S-D = Sprague-Dawley.
200
Toxaphene
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FINAL
07-31-2018
Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Logistic
1
0.8
0.6
0.4
0.2
BMDL
3MD
0
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
0.4
0.45
dose
14:09 06/29 2017
Figure C-21. Logistic Model for Increased Incidence of Anisokaryosis (All Severity Grades)
in the Liver of FO Female S-D Rats Exposed to Technical Toxaphene in the Diet for
25-29 Weeks (Two Highest Doses Dropped) (Chu et al., 1988)
Text output for Figure C-21:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/1og_Chu_l98 8_liv_aniso_F0_female_rats_2_high_d
rop_HEDs_Log-BMR10.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/1og_Chu_l98 8_liv_aniso_F0_female_rats_2_high_d
rop_HEDs_Log-BMR10.pit
Thu Jun 29 15:31:24 2017
BMDS Model Run
The form of the probability function is:
P[response] = 1/[1+EXP(-intercept-slope*dose)]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is not restricted
Total number of observations = 3
Total number of records with missing values = 0
Maximum number of iterations = 5 00
201
Toxaphene
-------
FINAL
07-31-2018
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
Default Initial Parameter Values
background = 0 Specified
intercept = -0.36056
slope = 8 .12881
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept slope
intercept 1 -0.59
slope -0.59 1
Parameter Estimates
Interval
Variable
Limit
intercept
0.571708
slope
53.9345
Estimate
-1.17871
-2.29924
30.9053
Std. Err.
-0.0581829
11.7498
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
7.87617
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-14.2791
-14.2792
-24.9803
# Param's Deviance Test d.f. P-value
3
2 8.08097e-005 1 0.992E
1 21.4023 2 <.0001
AIC:
32.5583
Dose
Est. Prob.
Goodness of Fit
Expected Observed Size
Scaled
Residual
0.0000
0.0830
0.4400
Chi^2 = 0.00
0.2353
0.8000
1.0000
d.f. = 1
4.000 4.000 17.000
8.000 8.000 10.000
10.000 10.000 10.000
P-value = 0.9949
0. 000
-0.000
0. 006
Benchmark Dose Computation
Specified effect = 0.1
202
Toxaphene
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07-31-2018
Risk Type = Extra risk
Confidence level = 0.95
BMD = 0.0125152
BMDL = 0.00714717
Increased Incidence of Cytoplasmic Homogeneity (All Severity Grades) in the Liver of
Fla Male S-D Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks (Cfau et al.,
1988)
The procedure outlined above was applied to the data for increased incidence of
cytoplasmic homogeneity in the liver of Fla male S-D rats exposed to technical toxaphene for
34 weeks (Chu et al, 1988) (see Table C-53). Table C-54 summarizes the BMD modeling
results. None of the models provided adequate fit to the full data set or the data set with the
highest dose dropped. With the two highest doses dropped, only the LogLogistic model
provided adequate fit to the data. Thus, the BMDLio of 0.0039 mg/kg-day from this model is
selected for this endpoint (see Figure C-22). However, the modeling results for this endpoint are
not considered reliable because all response levels were considered far in excess of the BMR,
leaving no data to inform the shape of the dose-response curve in the low-dose region and
requiring extrapolation far below the observable range in order to estimate the BMD (U.S. EPA.
2012b).
Table C-53. Incidence of Cytoplasmic Homogeneity (All Severity Grades) in the Liver of
Fla Male S-D Rats Exposed to Technical Toxaphene in the Diet for 34 Weeks3
HED (mg/kg-d)
0
0.078
0.37
2.0
10
Sample size
12
10
10
10
13
Incidence
0
6
7
4
10
Timet al. (1988).
HED = human equivalent dose; S-D = Sprague-Dawley.
203
Toxaphene
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FINAL
07-31-2018
Table C-54. BMD Modeling Results for Incidence of Cytoplasmic Homogeneity (All
Severity Grades) in the Liver of Fla Male S-D Rats Exposed to Technical Toxaphene in
the Diet for 34 Weeks
Model
X2 Goodness-of-Fit /j-Value11
AIC
BMDio (mg/kg-d)
BMDLio (mg/kg-d)
Gammab
0.0038
74.49
1.05512
0.51671
Logistic
0.0039
74.54
1.51357
0.88846
LogLogistic0
0.0036
74.42
0.63692
0.17157
LogProbit0
0.0039
74.74
9.16632
1.00947
Multistage (l-degree)d
0.0038
74.49
1.05512
0.51671
Multistage (2-degree)d
0.0038
74.49
1.05512
0.51671
Multistage (3-degree)d
0.0012
76.49
1.06583
0.51671
Multistage (4-degree)d
0.0012
76.49
1.11203
0.51675
Probit
0.0039
74.53
1.53015
0.94297
Weibullb
0.0038
74.49
1.05512
0.51671
Highest dose dropped
Gammab
0.0012
60.44
1.12349
0.21385
Logistic
0.0012
60.49
1.47578
0.36399
LogLogistic0
0.0012
60.37
0.76576
0.04289
LogProbit0
0.0013
60.69
56.62090
0.55837
Multistage (l-degree)d
0.0012
60.44
1.12347
0.21385
Multistage (2-degree)d
0.0012
60.44
1.12347
0.21385
Multistage (3-degree)d
0.0012
60.44
1.12347
0.21385
Probit
0.0012
60.49
1.45752
0.36315
Weibullb
0.0012
60.44
1.12353
0.21385
Two highest doses dropped
Gammab
0.0795
32.17
0.02067
0.01298
Logistic
0.006
39.08
0.06406
0.03927
LogLogistic6
0.4858
29.01
0.00925
0.00393
LogProbitd
0.0505
32.49
0.02995
0.01812
Multistage (l-degree)d
0.0795
32.17
0.02067
0.01298
Multistage (2-degree)d
0.0795
32.17
0.02067
0.01298
Probit
0.006
38.96
0.06229
0.04069
Weibullb
0.0795
32.17
0.02067
0.01298
aValues <0.1 fail to meet conventional goodness-of-fit criteria.
bPower restricted to >1.
°Slope restricted to >1.
dBetas restricted to >0.
"Selected model.
AIC = Akaike's information criterion; BMD = benchmark dose; BMDio = 10% benchmark dose;
BMDLio = 10% benchmark dose lower confidence limit; S-D = Sprague-Dawley.
204
Toxaphene
-------
FINAL
07-31-2018
Log-Logistic Model, with BMR of 10% Extra Risk for the BMD and 0.95 Lower Confidence Limit for the BMDL
Log-Logistic
0.6
o
oj
<
£Z
o
~o
0.4
u_
0.2
BMDL
3MD
0
0.05
0.1
0.15
0.2
0.25
0.3
0.35
dose
14:42 06/30 2017
Figure C-22. LogLogistic Model for Increased Incidence of Cytoplasmic Homogeneity
(All Severity Grades) in the Liver of Fla Male S-D Rats Exposed to Technical Toxaphene
in the Diet for 34 Weeks (Two Highest Doses Dropped) (Chu et al., 1988)
Text output for Figure C-22:
Logistic Model. (Version: 2.14; Date: 2/28/2013)
Input Data File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_liv_cyto_homog_Fla_male_rats_2_hi
gh_drop_HEDs_Lnl-BMR10-Restrict.(d)
Gnuplot Plotting File:
C:/Users/swesselk/Desktop/BMDS2 601/Data/lnl_Chu_198 8_liv_cyto_homog_Fla_male_rats_2_hi
gh_drop_HED s_Lnl-BMR10-Restrict.pit
Fri Jun 30 14:42:15 2017
BMDS Model Run
The form of the probability function is:
P[response] = background+(1-background)/[1+EXP(-intercept-siope*Log(dose))]
Dependent variable = Effect
Independent variable = Dose
Slope parameter is restricted as slope >= 1
Total number of observations = 3
Total number of records with missing values = 0
205
Toxaphene
-------
FINAL
07-31-2018
Maximum number of iterations = 5 00
Relative Function Convergence has been set to: le-008
Parameter Convergence has been set to: le-008
User has chosen the log transformed model
Default Initial Parameter Values
background = 0
intercept = 2.33798
slope = 1
Asymptotic Correlation Matrix of Parameter Estimates
( *** The model parameter(s) -background -slope
have been estimated at a boundary point, or have been specified by
the user,
and do not appear in the correlation matrix )
intercept
intercept 1
Parameter Estimates
Interval
Variable
Limit
background
intercept
3.46616
slope
Estimate
0
2.48579
1
Std. Err.
NA
0.500198
NA
NA - Indicates that this parameter has hit a bound
implied by some ineguality constraint and thus
has no standard error.
95.0% Wald Confidence
Lower Conf. Limit Upper Conf.
1.50542
Analysis of Deviance Table
Model
Full model
Fitted model
Reduced model
Log(likelihood)
-12.8388
-13.5063
-21.6149
# Param's
3
1
1
Deviance Test d.f.
1.33505
17.5522
P-value
0.513
0. 0001544
AIC:
29.0126
Dose
Goodness of Fit
Est._Prob. Expected Observed Size
Scaled
Residual
0.0000
0.0780
0.3700
0.0000
0.4837
0.8163
0.000
4.837
8.163
0.000
6.000
7.000
12.000
10.000
10.000
0. 000
0.736
-0.950
Chi^2 = 1.44
d.f. = 2
P-value = 0.4858
206
Toxaphene
-------
FINAL
07-31-2018
Benchmark Dose Computation
Specified effect
Risk Type
Confidence level
BMD
BMDL
0.1
Extra risk
0. 95
0.0092511
0.00392959
207
Toxaphene
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FINAL
07-31-2018
APPENDIX D. REFERENCES
ACGIH (American Conference of Governmental Industrial Hygienists). (2001). Chlorinated
camphene. Documentation of the threshold limit values for chemical substances, 7th
edition. In Documentation of the threshold limit values and biological exposure indices
(7th ed.). Cincinnati, OH. http://www.acgih.org/forms/store/ProductFormPublic/2015-
tlvs-and-beis
ACGIH (American Conference of Governmental Industrial Hygienists). (2015). Chlorinated
camphene. 2015 TLVs and BEIs. Based on the documentation of the threshold limit
values for chemical substances and physical agents and biological exposure indices
[TLV/BE1], Cincinnati, OH. http://www.acgih.org/forms/store/ProductFormPublic/2015-
tlvs-and-beis
ACGIH (American Conference of Governmental Industrial Hygienists). (2017). 2017 TLVs and
BEIs: Based on the documentation of the threshold limit values for chemical substances
and physical agents & biological exposure indices. Cincinnati, OH.
http://www.acgih.org/forms/store/ProductFormPublic/2017-tlvs-and-beis
Alder. L; Yieth. B. (1996). A congener-specific method for the quantification of camphechlor
(toxaphene) residues in fish and other foodstuffs. Fresenius J Anal Chem 354: 81-92.
Allen. A I.: K oiler. I.I); Pollock. GA. (1983). Effect of toxaphene exposure on immune responses
in mice. J Toxicol Environ Health 11: 61-69.
http://dx.doi.org/10.1080/1528739830953032Q
Andrews. P; Head rick. K; Pi Ion. JC; Brvce. F; Iverson. F. (1996). Capillary GC-ECD and ECNI
GCMS characterization of toxaphene residues in primate tissues during a feeding study.
Chemosphere 32: 1043-1053. http://dx.doi.org/10.1016/0045-6535(96)00024-0
Andrews. P; Vetter. W. (1995). A systematic nomenclature system for toxaphene congeners Part
1: Chlorinated bornanes. Chemosphere 31: 3879-3886. http://dx.doi.org/10.1016/0Q45-
6535(95)00260-F
Angerhofer. D; Kimmel. L; Koske. G; Fingerling. G; Burhenne. J; Parlar. H. (1999). The role of
biotic and abiotic degradation processes during the formation of typical toxaphene peak
patterns in aquatic biota. Chemosphere 39: 563-568. http://dx.doi.org/10.1016/S0Q45-
6535(99)00121-6
Arnold, PL; Brvce. F; Baccanale. C; Hayward, S; Tanner, JR; MacLellan, E; Dearden, T; Fernie,
(2001). Toxicological consequences of toxaphene ingestion by cynomolgus (Macaca
fascicularis) monkeys. Part 1: Pre-mating phase. Food Chem Toxicol 39: 467-476.
http://dx.doi.org/10.1016/S0278-6915(00)00151-4
AT SDR (Agency for Toxic Substances and Disease Registry). (2014). Toxicological profile for
toxaphene [ATSDR Tox Profile], Atlanta, GA: U.S. Department of Health and Human
Services. http://www.atsdr.cdc.gov/ToxProfiles/tp94.pdf
Barbini. DA; Stefanelli. P; Girolimetti. S; Di Muccio. A; Dommarco. R. (2007). Determination
of toxaphene residues in fish foodstuff by GC-MS. Bull Environ Contam Toxicol 79:
226-230. http://dx.doi.org/10.1007/s00128-0Q7-9179-6
Bartos. T: Skarek. M; Cupr. P; Kosubova. P; Holoubek. I. (2005). Genotoxic activity of a
technical toxaphene mixture and its photodegradation products in SOS genotoxicity tests.
Mutat Res 565: 113-120. http://dx.doi.Org/10.1016/i.mrgentox.2004.09.007
Benfenati. E; Manganaro, A; Gini. G. (2013). VEGA-QSAR: AI inside a platform for predictive
toxicology. Popularize Artificial Intelligence 2013: Proceedings of the Workshop on
Popularize Artificial Intelligence (PAI 2013), December 5, 2013, Turin, Italy.
208
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