FINAL
Umted States ECAO-CIN-426
Environmental Protection Fahnuru
Agency reuruary.
IvEPA Research and
Development
DRINKING WATER CRITERIA DOCUMENT
FOR TOXAPHENE
Prepared for
OFFICE OF DRINKING WATER
Prepared by
Environmental Criteria and Assessment Office
Office of Health and Environmental Assessment
U.S. Environmental Protection Agency
Cincinnati, OH 45268
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DISCLAIMER
This document has been reviewed In accordance with the U.S. Environ-
mental Protection Agency's peer and administrative review policies and
approved for publication. Mention of trade names or commercial products
does not constitute endorsement or recommendation for use.
11
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FOREWORD •
Section 1412 (b)(3)(A) of the Safe Drinking Water Act, as amended In
1986, requires the Administrator of the Environmental Protection Agency to
publish maximum contaminant level goals (MCLGs) and promulgate National
Primary Drinking Water Regulations for each contaminant, which, in the
judgment of the Administrator, may have an adverse effect on public health
and which 1s known or anticipated to occur 1n public water systems. The
HCLG 1s nonenforceable and 1s set at a level at which no known or antici-
pated adverse health effects In humans occur and which allows for an
adequate margin of safety. Factors considered In setting the MCLG Include
health effects data and sources of exposure other than drinking water.
This document provides the health effects basis to be considered In
establishing the MCLG. To achieve this objective, data on pharmacoklnetlcs,
human exposure, acute and chronic toxldty to animals and humans, epidemi-
ology and mechanisms of toxldty are evaluated. Specific emphasis Is placed
on literature data providing dose-response Information. Thus, while the
literature search and evaluation performed 1n support of this document has
been comprehensive, only the reports considered most pertinent In the deri-
vation of the MCLG are cited 1n the document. The comprehensive literature
data base 1n support of this document Includes Information published up to
1984; however, more recent data may have been added during the review
process.
When adequate health effects data exist. Health Advisory values for less
than lifetime exposures (1-day, 10-day and longer-term, -10X of an
Individual's lifetime) are Included 1n this document. These values are not
used 1n setting the MCLG, but serve as Informal guidance to municipalities
and other organizations when emergency spills or contamination situations
occur.
Michael B. Cook
Director
Office of Drinking Water
111
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DOCUNENT DEVELOPMENT
Annette M. Gatchett, B.S., Document Manager
Environmental Criteria and Assessment Office, Cincinnati
U.S. Environmental Protection Agency
Helen H. Ball, M.S., Project Officer
Environmental Criteria and Assessment Office, Cincinnati
U.S. Environmental Protection Agency
Authors
Shane Que Hee, Ph.D.
Martha Radlcke, Ph.D.
University of Cincinnati
Department of Environmental Health
Cincinnati, OH
Reviewers
M.S.
Randall J.F. Bruins,
Frank M1nk, Ph.D.
Environmental Criteria and
Assessment Office, Cincinnati
U.S. Environmental Protection Agency
William Marcus, Ph.D.
Jennifer Orme, M.S.
Office of Drinking Water
Washington, DC
Fumlo Matsumura, Ph.D.
Pesticide Research Center
Michigan State University
East Lansing, MI
(Contract 68-03-3234; Eastern
Research Group)
William E. Pepelko, Ph.D.
Carcinogen Assessment Group, OHEA
U.S. Environmental Protection Agency
Washington, DC
James WUhey, Ph.D.
Sir Frederick Banting Research Center
Bureau of Chemical Safety
Ottawa, Ontario
Canada
(Contract 68-03-3234; Eastern
Research Group)
James 0. Pierce
University of Southern California
Cooperative Agreement
No. CR 813569-01-0
Editorial Reviewers
Erma Durden, B.S.
Judith Olsen, B.A.
Environmental Criteria and
Assessment Office, Cincinnati
U.S. Environmental Protection Agency
Document Preparation
Technical Support Services Staff: C. Cooper, P. Daunt, C. Fessler
B. Zwayer, Environmental Criteria and Assessment Office, Cincinnati
K. Mann,
1v
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TABLE OF CONTENTS
I. SUMMARY ........................... i.i
II. PHYSICAL AND CHEMICAL PROPERTIES ............... II-l
PRODUCTION AND USAGE ..................... II-8
SPECTROSCOPIC PROPERTIES ................... II-9
ANALYSIS ...... ..................... 11-12
SUMMARY ........................... II-15
III. TOXICOKINETICS ........................ III-l
ABSORPTION ............... ' ........... III-l
ORAL ............................. III-l
INHALATION .......................... III-5
DERMAL ............................ III-5
DISTRIBUTION ......................... TII-5
DISTRIBUTION IN ANIMAL TISSUES ................ III-6
Birds ...................... ..... III-6
Mammals ......................... III-9
DISTRIBUTION IN HUMAN TISSUES . ............... 111-19
METABOLISM .......................... 111-19
ELIMINATION ......................... 111-25
SUMMARY ........................... 111-31
IV. HUMAN EXPOSURE ........................ IV-1
V. HEALTH EFFECTS IN ANIMALS .................. V-l
ACUTE AND SUBACUTE TOXICITY ........... . ..... V-l
CHRONIC TOXICITY ....................... V-18
ENZYME EFFECTS ........................ V-26
TERATOGENICITY AND REPRODUCTIVE EFFECTS ........... V-37
Mammals ......................... V-37
Birds .......................... V-44
MUTAGENICITY ......................... V -47
CARCINOGENICITY ....................... V-50
SUMMARY ..... ..................... . V-65
VI. HEALTH EFFECTS IN HUMANS ................... VI-1
ACUTE TOXICITY ........................ VI-1
SUBCHRONIC AND CHRONIC TOXICITY ............... VI-5
SUMMARY ...................... ..... VI-7
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Page
VII. MECHANISMS OF TOXICITY VII-1
INTRODUCTION VII-1
ENZYMATIC EFFECTS VII-2
MUTAGENIC ACTIVITY VII-7
SUMMARY VII-8
VIII. QUANTIFICATION OF TOXICOLOGICAL EFFECTS VIII-1
INTRODUCTION. ... VIII-1
NONCARCINOGENIC EFFECTS . VIII-6
QUANTIFICATION OF NONCARCINOGENIC EFFECTS , VIII-12
Derivation of 1-Day HA VIII-12
Derivation of 10-Day HA VIII-13
Derivation of Longer-Term HA VIII-14
Assessment of Lifetime Exposure and Derivation
of a DWEL VIII-14
CARCINOGENIC EFFECTS VIII-14
QUANTIFICATION OF CARCINOGENIC EFFECTS VIII-20
EXISTING GUIDELINES, RECOMMENDATIONS AND STANDARDS VIII-23
SPECIAL GROUPS AT RISK VIII-26
Sensitive Subpopulatlon VIII-26
Interactions VIII-27
IX. REFERENCES. IX-1
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LIST OF TABLES
No. Title Page
II-l Nomenclature, Indexing Terms and Synonyms Currently
Used for Toxaphene II-3
II-2 Identified Toxic Components of Toxaphene II-4
II-3 Physical Properties of the Degradation Products
of Toxaphene. . 11-6
II-4 Chemical Information Regarding Toxaphene Mixtures 11-10
II-5 Examples of Analytical Methods for Toxaphene 11-13
III-l Absorption Data for All Routes for Toxaphene III-2
III-2 Distribution of Toxaphene Continuously Administered
Orally to Black Ducks from Birth by Mixing Toxaphene 1n
Propylene Glycol, which Constituted IX of the Duck Mash . . 111-10
III-3 Percent Uptake of Radioactivity 1n Various Rat Tissues
and Organs Following a Single Dose of "Cl-Toxaphene
(20 mg/kg) Administered 1n 0.5 ml Peanut 011/Green
Acacia 111-12
III-4 Distribution of Radioactivity 1n Various Rat Tissues
and Organs at Day 14 Following a Single Dose of
36Cl-Toxaphene, 14C-Toxaphene, Toxicant A and B
Administered 1n Corn 011 by Stomach Intubation III-14
III-5 Distribution and Metabolism of Toxicant B (51775-36-1)
at 7 and 72 Hours After Stomach Intubation of Sprague-
Dawley Rats at 3.1 mg/kg bw 111-16
III-6 Distribution of Radioactivity 1n Various Female Sprague-
Dawley Rat Tissues and Organs Following a Single Dose of
2.6 ing 14C-Toxaphene/kg bw Dose to Four Pregnant Rats
Administered In Olive 011 (0.1 mi) by Oral Gavage 111-18
III-7 Identified Metabolites of Toxaphene Components 111-22
III-8 Elimination of 14C-Toxaphene, Toxicant A and
Toxicant B Administered to Sprague-Dawley Rats
by Stomach Intubation 1n Corn 011 Carrier 111-28
III-9 Experimental or Calculated Half-Life of Toxaphene
Elimination 1n Various Species 111-32
V-l Acute Oral Toxlclty of Technical Toxaphene
(CAS RN 8001-35-2) to Laboratory Mammals. V-2
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No. ' Title Page
V-2 Acute Dermal LDsg Values for Toxaphene 1n Laboratory
Mammals V-5
V-3 Acute Toxlclty of Technical Toxaphene Components
Administered IntraperHoneally to Mice V-6
V-4 Residue Concentrations In Dead Birds and Animals V-10
V-5 Acute and Subchronlc Oral Toxldty of Toxaphene V-13
V-6 Chronic Toxlclty of Toxaphene to Laboratory Mammals
at Low Dietary Levels V-19
V-7 National Cancer Institute Chronic Feeding Study V-22
V-8 Incidence of Tumors In Male Rats Fed Toxaphene V-53
V-9 Incidence of Tumors In Female Rats Fed Toxaphene V-54
V-10 Incidence of Tumors 1n Male Mice Fed Toxaphene In
the Diet V-56
V-ll Incidence of Tumors In Female Mice Fed Toxaphene
1n the Diet V-57
V-12 Survival of Mice Fed Toxaphene 1n the Diet V-59
V-13 Hepatocellular Tumor Incidence In B6C3F1 Mice Fed
Toxaphene In the Diet V-61
V-14 Variation 1n Doses of Toxaphene Used 1n NCI (1979)
Study of CarclnogenlcHy V-69
VI-1 Case Studies of Toxaphene Poisoning 1n Humans 1n which
Ingestlon Was the Primary Route of Entry VI-3
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LIST OF ABBREVIATIONS
ATP
bw
CNS
DNA
dw
DWEL
GC
GI
HA
l.p.
1.v.
LD50
LOAEL
MEL
MFO
NADPH
NOAEL.
NOEL
ppm
RfD
s.c.
TLC
TLV
Toxicant A
Toxicant A-l
Toxicant A-2
Toxicant Ac
Toxicant B
TWA
ww
Adenoslne trlphosphate
Body weight
Central nervous system
DeoxyMbonuclelc add
Dry weight
Drinking water equivalent level
Gas chromatography
Gastrointestinal
Health advisory
IntrapeMtoneal
Intravenous
Lethal dose to 50% of the recipients
Lowest-observed-adverse-effect level
Minimal effect level
Mixed function oxldase
N1cot1nam1de adenlne dlnucleotlde phosphate dehydrogenase
No-observed-adverse-effect level
No-observed-effect level
Parts per million
Reference dose
Subcutaneous
Thin-layer chromatography
Threshold limit value
Subfractlon of toxaphene separated chromatographlcally
A toxic component of Toxicant A
A toxic component of Toxicant A
A contaminant of Toxicant A
Subfractlon of toxaphene separated chromatographlcally
Time-weighted average
Wet weight
1x
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I. SUMMARY
Toxaphene 1s a broad spectrum, chlorinated terpenold pesticide Intro-
duced In the United States as a contact Insecticide In 1948. Together with
methoxychlor, toxaphene has been the chief replacement for DOT when use of
DOT was banned. On May 25, 1977, the U.S. EPA Issued a notice of rebuttable
presumption against registration and continued registration of pestlcldal
products containing toxaphene. A further notice was Issued 1n the Federal
Register In 1982. Registration for use after December 31, 1983 for scabies
treatment of beef cattle and sheep 1n vat dips and spray dips was still
allowed under specified conditions. Toxaphene use to control sporadic
Infestations of army worms, cutworms and grasshoppers on cotton, corn and
small grains was also permitted under specific conditions. Uses for control
of mealybugs and pineapple gummosIs moths on pineapples, and weevils In
bananas are allowed only In the Virgin Islands and Puerto R1co. Sale and
distribution of existing toxaphene stockpiles was allowed until December 31,
1986 with the use of toxaphene under limited conditions only. In addition
to the previous uses, It may be applied to soybeans and peanuts for sickle-
pod control. Its sale and distribution for use as an Insecticide 1n no-till
corn and dry and southern peas was permitted until December 31, 1986. These
administrative decisions were made as a result of the toxic effect of toxa-
phene on wildlife, aquatic organisms, nontarget organisms, and the potential
oncogenldty of toxaphene In humans.
A variety of guidelines or standards have been published to protect
against toxaphene 1n aquatic media. The national Interim primary drinking
01990 1-1 02/25/87
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water standard Is 5 yg/i. The FOA mandated level for bottled water Is
also 5 yg/l. These standards are based upon organoleptlc effects.
The water quality criterion for the protection of human health was
originally set at 0.5 wg/l for a 10"5 cancer risk level. This cri-
terion was subsequently revised to 7.1 ng/l for a risk of 10"5. If con-
sumption of aquatic organisms 1s considered alone, the corresponding risk
level 1s 7.3 ng/l. These criteria were based on the Induction of hepato-
cellular tumors In mice.
Toxaphene 1s a mixture {average molecular weight 414) of at least 177
compounds, mostly chlorinated camphenes, which has been used as a broad
spectrum contact pesticide (e.g., In soybean and cotton fields, and for
ectoparasites). It Is produced Industrially by the reaction of camphene and
chlorine 1n carbon tetrachlorlde solvent 1n the presence of ultraviolet
light to produce a product of highly variable quality. Toxaphene can be
provided as a 40X dust concentrate, as a liquid formulation containing 10%
xylene, or as emulslflable concentrates. It 1s extensively co-formulated
with other pesticides (e.g., DDT, parathlon, endrln).
The vapor pressure, densUy, solubility and other physical properties
vary according to the quality of the product. Toxaphene Is nonpolar and Is
soluble In nonpolar solvents; It 1s less soluble In polar solvents. It
adsorbs to soils, and partitions readily Into n-octanol. Because toxaphene
1s aliphatic, It Is a poor absorber of ultraviolet light above wavelengths
of 250 nm. Gas chromatography with electron capture detection or chemical
01990 1-2 02/25/87
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1on1zat1on mass spectrometrlc detection are commonly used to measure toxa-
phene levels In biological samples. In a few cases, 'H- and 13C-nuclear
magnetic resonance techniques have been used to assess the "fingerprints" of
toxaphene fractions.
Toxaphene readily dehydrochloMnates on heating above 120°C, but the
toxaphene subfractlons Toxicants A and B appear to be stable on gas chroma-
tography. DehydrochloMnatlon also occurs In the presence of alkali (used
as an analytical method), of reduced hematln, or Iron compounds. Reductive
dechlorlnatlon occurs on ultraviolet light Irradiation (230-290 nm) of
toxaphene In organic solvents, preferentially at the C. and Cr positions.
Most of the analytical techniques used to detect toxaphene metabolites
have employed thin-layer chromatography as the end step. Toxaphene 1n
environmental samples and tissues often does not have the chromatographk
pattern of a toxaphene standard so that qualitative Identification by gas
chromatography occurs primarily through retention time; quantltatlon 1s
generally accomplished by the method of alkaline dehydrochlorlnatlon,
extraction and chromatography (Gomes method, detection limit 1 ng). Metabo-
lites are generally Isolated by thin-layer chromatography or liquid chroma-
tography.
Though no quantitative data are available, the symptoms of toxicosis as
well as the presence of residues and metabolites after exposure 1n animals,
birds and humans Indicate that toxaphene can be absorbed through the skin,
the lungs, and the gut.
01990 1-3 02/25/87
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In White Leghorn chickens, accumulation of toxaphene residues occurs at
doses of >12.5 mg toxaphene/kg bw/day. In ducks, brain residues were
detected at concentrations >10 ppm 1n the feed. Extensive dechloMnatlon of
toxaphene by chlorinated organic residues occurs mostly 1n the fat. Doses
less than those resulting 1n accumulation can be tolerated for months with
IHtle effect.
In mammals, storage of toxaphene 1n the fat has been reported In sheep,
steers and dairy cows. Accumulation of toxaphene 1n sheep and steers
occurred at administered concentrations between 10 and 100 mg toxaphene/kg
feed/day. In contrast, dogs appear to store toxaphene derivatives In the
brain. In male or female rats, fat levels of toxaphene were relative to
concentrations >21 ppm In the diet. Studies using labeled toxaphene
revealed rapid dechlorlnatlon and subsequent elimination of toxaphene.
Toxicant A and Toxicant B at doses <19 mg toxaphene/kg bw administered by
gavage to male rats using corn oil as a carrier. Toxaphene derivatives were
measured 1n the blood, fat, liver and kidneys. When male rats were
chronically dosed at 2.4 mg toxaphene/kg bw/day, plateau levels were found
In the liver and brain after 1, 3 and 6 months. Transplacental transfer as
well as blood-brain barrier transfer occurred In rats. The adrenal gland,
carcass, cecum and abdominal fat of female rats contained toxaphene deriva-
tives 3 days after an acute dosing with 2.6 mg toxaphene/kg bw. Extensive
dechlorlnatlon and rapid elimination of metabolites also occurred in guinea
pigs, hamsters, rabbits, mice and monkeys.
01990 1-4 02/18/87
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In rats, successful detoxification appeared to require NAOPH and oxygen.
Toxicants A and B undergo reductive dechlorlnatlon, dehydrochlorlnatlon, and
vicinal chloride elimination. Involvement of the MFC. system was suggested
by the observation of Type I binding spectra with the hepatic cytochrome
P-450 of rats, mice, sheep and rabbits, as well as the demonstrated enhanced
toxldty to mice of toxaphene 1n the presence of plperonyl butoxlde. There
also appeared to be a detoxification pathway dependent upon glutathlone.
Toxicant 8 resisted metabolism more than Toxicant C. Glucuronlde (major)
and sulfate (minor) conjugates have been detected as metabolites. Toxicant
C also produced one primary and four secondary alcohols, whereas reductive
dechlorlnatlon predominated for Toxicant B. These data suggested that
toxaphene components were not necessarily metabolized at the same loci In
the mlcrosomal system.
Elimination of products derived from toxaphene has been demonstrated to
occur In the feces, urine and expired air of rats. The fecal route appears
to be slightly more predominant than the urinary route. Milk and eggs may
also contain residues. The species order of decreasing elimination effi-
ciency for Toxicant B Is as follows: monkey, rat, hamster > mouse, rabbit,
guinea pig > chicken.
Acute oral exposure to toxaphene gave LD.- values ranging from 7.5-10
mg/kg bw 1n female monkeys to 75-500 mg/kg 1n rabbits. The LD5Q values
resulting from dermal application of toxaphene are generally higher than
those observed after oral exposure; they range from -250 mg/kg bw to -4000
mg/kg bw with both values being reported for rabbits. The vehicle used with
the pesticide may affect the toxic response. Specific components of
01990 1-5 02/25/87
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technical grade toxaphene are more toxic than the complex mixture and, as
reported for mice, L05Q values for several components varied from 2.5
to >100 mg/kg bw. Brain levels of toxaphene may be Indicative of acute
tox1c1ty. In swine, >4 mg/kg ww (brain tissue) constituted a lethal level,
and 2 mg/kg was associated with clinical signs of toxldty.
Subchronlc oral doses of toxaphene In laboratory animals resulted In few
clinical signs of poisoning. Lethality responses varied In mice fed 1280
ppm toxaphene In the diet for 6 weeks. No observable adverse effects were
seen In rats following 1ngest1on of feed containing 189 ppm toxaphene.. At
concentrations <189 ppm In the diet, subcllnlcal lesions such as decreased
bile production and questionable liver pathology were reported. In mice
receiving 100 or 200 ppm toxaphene 1n the diet, humoral antibody production
(IgG antibody formation) was suppressed; however, cell-mediated Immune
responses were not affected.
Inhaled toxaphene mist concentrations as high as 500 mg toxaphene/m3
for 3 weeks caused no mortality 1n rats and rabbits; but at 12 mg toxaphene
dust/m3 for 3 months, mortality occurred 1n rats, dogs and guinea pigs.
Long-term exposures of animals to dietary levels of toxaphene are sum-
marized In Chapter V. In a number of studies, no adverse effects were
reported among the parameters monitored In each experiment. In several
lifetime studies In rats, no effects were reported for dietary concentra-
tions between 10 and 100 ppm toxaphene; other studies at 100 ppm toxaphene
level reported liver pathology. The lowest dietary level producing liver
damage In rats was 5 ppm for 2 years. In addition to liver pathology, a
01990 1-6 02/25/87
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variety of adverse effects were reported from both Vn vivo and in vUro
studies and Included kidney pathology, decreased bile flow, decreased
survival, Inhibition of Intestinal transport of glucose, Inhibition of
gluconeogenesls. Inhibition of mltochondrlal enzymes, Inhibition of
Nah/Kf-ATPase brain and kidney activity, and Inhibited Mg2f-ATPase
activity In mouse kidney, liver and brain.
Toxaphene Induced the mlcrosomal MFO system; treatment of animals j_n
vivo with a cytochrome P-450 system Inhibitor Increased toxaphene toxldty.
No effects on parental animals or offspring were noted 1n a throe-
generation study In which Sprague-Oawley rats were fed dietary levels of 25
or 100 ppm toxaphene. In a study using CO rats, maternal mortality was 31%
at 35 mg toxaphene/kg bw. There was also a dose-related decrease 1n
maternal weight gain and fetal weight (15, 25 or 35 mg/kg bw). No adverse
effects were reported In the offspring of mixed-strain white rats dosed with
4 mg/kg bw during the entire pregnancy. In hamsters given the same dose,
toxaphene was reported to be teratogenlc but the anomaly was not Identified.
More sensitive endpolnts In rats were examined by several Investigators.
Toxaphene, at 12 mg/kg bw administered orally to pregnant rats, depressed
chollnesterase activity In fetal cardiac neural structures and delayed
cardiac neural differentiation. At 25 mg/kg bw, the only significant
differences noted were decreases 1n alkaline phosphatase activity and total
protein 1n the kidney. There was no effect at 12.5 mg/kg bw on the rat
01990 1-7 02/25/87
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kidney. In two separate experiments there was IHtle evidence of behavorlal
abnormalities 1n the offspring of toxaphene treated dams except for delayed
righting ability (6 mg/kg and 50 vg/kg bw).
Toxaphene given to pregnant mice at 15 and 25 mg/kg bw had no adverse
effect on parameters examined In offspring. However, a dose of 35 mg
toxaphene/kg bw Induced encephaloceles In five Utters of 90 treated dams.
In a 5- to 6-generat1on study where toxaphene (25 ppm) was given 1n the
diet, little or no adverse effects were observed In mice. No effects were
noted on the anatomical development of the fetal guinea pig from pregnant
females given 15 mg/kg bw from days 21-35 of gestation.
Toxaphene appears to have a low teratogenlc potential unless doses are
large enough to Induce maternal toxIcHy. In the studies reported here, the
lowest dose producing an effect was 50 vg/kg bw (rats) given In the diet
from day 5 of gestation through termination of the study. Effects Included
significant decreases 1n overall swimming ability and righting reflex of
young pups when toxaphene was administered before postnatal day 16. In this
study, however, the precise dose to the offspring Is uncertain since they
will receive toxaphene transplacentally, 1n the milk and In the feed.
Toxaphene 1s mutagenlc 1n the Salmonella/mlcrosomal reverse mutation
assay using strains TA98 and TA100. Results of assays of toxaphene compo-
nents for their mutagenlc potential Indicated that mutagenlclty resided in
the polar fraction. Mutagenlclty was decreased by the addition of a liver
mlcrosome, S9, an active HFO (these results support the Jm vivo observation
that Inhibitors of cytochrome P-450 Increased toxldty). The mutagenlc
activity of commercial preparations varies.
01990 1-8 02/18/87
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Studies of toxaphene cardnogenkHy are reported In Chapter V. In an
NCI carcinogenic study many doses were lowered because of the overt toxlclty
seen 1n rats and mice Ingesting feed containing toxaphene. The concurrent
numbers of control animals were low (10/sex/group) and historical controls
were used 1n order to match the number of treated animals per group (50/sex/
group). The range of doses given to the animals made H difficult to use
time-weighted averages of doses as a basis on which to estimate risk for
humans. It was concluded that under the conditions of the bloassay, toxa-
phene was carcinogenic 1n male and female B6C3F. mice, as evidenced by an
Increased Incidence of hepatocellular carcinomas 1n a dose-related manner.
In a separate study using B6C3F, mice of both sexes, fed 7, 20 and 50
mg toxaphene/kg diet, the carclnogenlcUy of toxaphene was demonstrated at
much lower doses. After 18 months of toxaphene Ingestlon and 6 months on a
control diet, significantly Increased Incidences of hepatocellular car-
t,
dnomas were detected 1n males. The response 1n females was less pro-
nounced. Another study reported only one tumor, a subcutaneous neurosarcoma
In rats exposed to 1000 ppm toxaphene 1n the diet.
Based on the positive carcinogenic evidence of toxaphene from the NCI
mouse and rat studies and the other mouse study, 1t can be concluded that
toxaphene 1s carcinogenic 1n at least two species of laboratory animals.
Using the EPA criteria for classifying cardnogenldty data, the animal
evidence would be considered "sufficient".
01990 1-9 04/02/87
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Toxaphene has caused convulsions and nausea In humans when exposure has
occurred by Ingestlon, skin contact or Inhalation. The acute oral L05Q
was estimated from accidental poisonings to vary between 29 and 100 mg
toxaphene/kg bw. Mixtures of toxaphene with DOT (2:1, by weight) are more
acutely toxic than either component alone. The estimated hazardous dermal
dose (applied to the skin) 1s -660 mg/kg bw for liquid toxaphene. Allergic
bronchopneumonla has been observed after acute Inhalation of toxaphene
sprays. Women have been shown to suffer a significant Incidence of chromo-
somal aberrations In lymphocytes.
Dermal doses of 300 mg/day for 30 days, and Inhaled doses of 0.4 mg/m3
for 10 minutes on each of 15 days and 250 mg/m3 for 30 minutes (-41
mg/day) on each of 13 days caused no atypical subjective or clinical effects.
Studies In the workplace are confounded because exposure to many
chemicals had occurred 1n all the reported epidemlologic studies. There are
no porphyrlnogenlc or sympathotonlc effects mentioned In the available
literature.
Human data are Inadequate to assess the carcinogenic potential for
toxaphene.
Animal studies were used to derive the 1- and 10-day HAs as a result of
the lack of human data. The recommended 1-day health advisory (HA) for
toxaphene In drinking water 1s 0.5 mg/l for a 10 kg child. A NOAEL of 5
mg/kg bw based upon the onset of convulsions In dogs receiving a single dose
of toxaphene was used to derive this HA. The recommended 10-day HA for
01990 1-10 02/18/87
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toxaphene In drinking water Is 0.04 mg/i for a 10 kg child. A LOAEL of 4
mg/kg bw based upon minimal kidney and liver pathology 1n dogs was used to
derive this MA.
There are no acceptable studies 1n the available literature for the
derivation of a longer-term HA or a lifetime DWEL.
The recommended criterion for chronic exposure to toxaphene 1n drinking
water, not considering Intake from other sources, Is 3.1, 0.31 and 0.031
vg/4, to maintain Individual cancer risk less than 10~4, 10~5 and
10~6, respectively. This criterion Is based upon an Increase 1n hepato-
cellular tumors 1n male mice fed 7, 20 and 50 ppm toxaphene In the diet.
The sufficient level of carcinogenic evidence In experimental animals,
together with the Inadequate level of human evidence meets the IARC Group 28
criteria for weight of evidence that a chemical Is likely to be a human
carcinogen. The assignment of a Group 28 designation by IARC means that the
chemical 1s probably carcinogenic 1n humans. Applying the criteria
described In the U.S. EPA's guidelines for assessment of carcinogen risk,
toxaphene may be classified as Group B2: probable human carcinogen, meaning
there 1s Inadequate evidence from human studies and sufficient evidence from
animal studies.
01990 1-11 05/12/87
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II. PHYSICAL AND CHEMICAL PROPERTIES
Toxaphene Is a broad spectrum, chlorinated terpenold pesticide. Intro-
duced as a contact Insecticide Into the United States In 1948 (Brooks,
1974). Together with methoxychlor, 1t was and has been the chief replace-
ment for DOT after DOT was banned. On May 25, 1977, the U.S. EPA Issued a
notice of rebuttable presumption against registration and continued regis-
tration of pestlddal products containing toxaphene (Federal Register,
1977). A further notice was Issued In the Federal Register (1982). Regis-
tration for use after December 31, 1983 for scabies treatment of beef cattle
and sheep In vat dips and spray dips was still permitted under specified
conditions. Toxaphene use to control sporadic Infestations of army worms,
cutworms and grasshoppers on cotton, corn and small grains was also
permitted under certain restrictions (Federal Register, 1982). The use of
toxaphene to control mealybugs, gummosls moths on pineapples, and weevils In
bananas 1s allowed only 1n the Virgin Islands and Puerto R1co. Depletion of
existing toxaphene stockpiles was allowed until December 31, 1986 under
limited conditions only. In addition to the previous uses, It may be
applied to soybeans and peanuts for slcklepod control. Its sale and dis-
tribution for use as an Insecticide In no-till corn and dry and southern
peas was permitted until December 31, 1986 (Federal Register, 1982). These
administrative decisions were made as a result of the toxic effect of
toxaphene on wildlife, aquatic organisms, nontarget organisms, and potential
oncogenlclty of toxaphene 1n humans.
Toxaphene can be produced Industrially by the reaction between technical
grade camphene and chlorine (In carbon tetrachloMde solvent) In the
02000 II-l 02/25/87
-------
presence of ultraviolet light (mercury arc radiation) for 15-30 hours until
the final product has a chlorine content between 67 and 69% (SHtlg, 1977).
The chemical quality of the final product 1s highly variable (IUPAC, 1979).
The many synonyms of toxaphene are provided In Table II-l. Toxaphene Is
known as chlorophene or polychlorocamphene In the Russian literature
(Melnlkov, 1971). Synonyms, chemical names and Indexing terms for the major
toxic components of toxaphene are given 1n Table II-2. The most toxic
Ingredients to mammals 1n technical toxaphene are Toxicant A and Toxicant B
(Table II-2). each consisting of -2-6X of the mixture (Metcalf, 1981;
Khalifa et al., 1974). Toxicant A consists of three toxic components, Toxi-
cant A-l (CAS No. 58002-18-9) and Toxicant A-2 (CAS No. 58002-19-0), and a
contaminant, Toxicant Ac (CAS No. 66860-80-8) (Chandurkar et al., 1978).
Synonyms, chemical names and Indexing terms for the major metabolites and
degradation products of toxaphene are provided 1n Chapter III.
Since the technical grade product 1s produced by free radical reactions
Initiated by ultraviolet light, toxaphene Is a complex mixture of poly-
chlorinated camphenes and bornanes with an average empirical formula of
C,nHinClQ, and an average molecular weight of 414. In fact, over 177
i u i u o
Incompletely characterized components have been separated (Holmstead et al.,
1974). Technical grade toxaphene 1s an amber, waxy solid with a mild
terpene odor, a softening range of 70-95°C, a vapor pressure of 0.17-0.40 mm
Hg at 25°C, and a density of 1.66 g/ml at 27°C (Brooks, 1974). The solu-
bility 1n water has been variously reported as -3 mg/i (Brooks, 1974) and
0.5 mg/i at 25°C (Paris et al.. 1977). Differing values for vapor pres-
sure, solubilities and other physical constants are to be expected since
02000 II-2 02/25/87
-------
TABLE II-l
Nomenclature, Indexing Terms and Synonyms Currently Used for Toxaphenea
CAS RN 8001-35-2°
Empirical Formula:
Toxaphene (DOT)C (SCI & 9CI)d«e
Agrlclde maggot killer (f)
Alltex
ATHox
Camphechlor
Camphochlor
Chem-Phene
Chlorinated camphene
Chlorinated camphene, tech
Chlor chem t-590
Compound 3956
Estonox
ENT 9,735
Fasco-Terpene
Genlphene
Gy-Phene
Hercules 3956
Kamfochlor
M 5055
Mellpox
Motox
NCI-C 00259
Octachlorocamphene
Penphene
Phenaclde
Phentox
Polychlorcamphene
Polychlorocamphene
PCHK
Strobane T
Synthetic 3956
Technical chlorinated
camphene (67-69%) chlorine)
Toxadust
Toxafeen (Dutch)
Toxakll
Toxaphen (German)
Source: SANSS data base, June 1983
bChem1cal Abstracts Service Registry Number
C0epartment of Transportation
dE1ghth Collective Index, Chemical Abstracts
eN1nth Collection Index, Chemical Abstracts
02000
II-3
02/03/87
-------
o
ro
O
lABlt 11-2
Identified loxtc Components of loxaphene
CAS RN
tv
Cheated! Abstracts
(9CI)
Synonyas
Reference
51775-36-1
C10HnCl7
52819-39-3
57028-55-4
57981:- 30-3
58002-18-9
CIOHIOC'8
58002-19-0
66860-80-8
70940-13-5
Btcyclo|2.2.1)heptane.2.2.5.6 tetrachloro
1.7.7.-trls(chloro«elhyl)-.<5 endo.6 exo)
Blcyclo(2.2.1]heptane.2.3.3.5.6-pentachloro-
7.7-bls(chloroM>thyl)-l (dtchloro«ethyl)-
(2-endo.5-exo.6-exo)-
Blcyclo[2.2.1]heptane.2.2.5.6 tetrachloro
1,7-bts(chloroMthyl) -7-(dtchloroaethyl) -
Btcyclo|2.2.1)heptane.2.5.6-trlchloro-3.
3-bls(chloroaethyl)-2-(dlchloroM>thyl)
(exo.exo.exo)-
Blcyclo)2.2.1]heptane.2.2.5.6-tetrachloro-
1.7-bts(chloroaethyl 7 (dlchloro«ethyl)-
(5-endo.6-exo.7-antl)-
Blcyclol2.2.1]heptane.2.2.5.6 tetrachloro
1.7-bls(chlor«Mthyl-7-(dlchlorowtbyl)-
(5-endo.6-exo.7-syn)-
Blcyclo[2.2.1Jheptane.2.3.5.6 tetrachloro-
7-(chloroMthyl)-l-7-bl$(dtchlor(MKthyl)-
(2-endo.3-exo.5-endo.6-exo.7-syn)-
Blcyclo[2.2.1Jheptane.2.3.3.5.6-pentachloro-
2.2.S endo.6 exo.8.9.10
heptachlorobornane;
loxaphene toxicant B
loxaphene toxicant C*;
2-endo.3.3.5.6-exo.8.9.
10.10-nonachlorobornane
Toxic fraction A; 2.2.
5-endo.6-exo.8.9.9.10-
octachlorobornane
2.5.6-6X0.8.8.9.10-
heptachlorodlhydro-
caophene
2.2.5 endo.6 exo.8.8.9.10
octachlorobornane;
loxaphene toxicant A-l
2.2.S endo.6-exo.8.9.9.10
octachlorobornane;
Toxaphene toxicant A-2
Toxaphene toxicant Ac
loxaphene toxicant C4;
?,3,3-endo:5:6-exo,a,9
10.10-nonachlorobornane
Clark and Natsuwira. 1979;
Saleh and Caslda. 1978;
Saleh et al.. 1979;
Chandurkar and NatsuMira.
1979a.b
Chandurkar and Matsuaura.
1979a.b
Clark and HatsuMira. 1979
Swanson et al.. 1978
Pollock and Kllgoie. 1980
Pollock and Ktltjore. 1980
Chandurkar et al.. 1978
Clark and NalsuMira. 1979
•Proposed structures have not been confirmed ,
CAS RN = Chemical Abstracts Service Registry Nunber; Nf -- Holecular Fornula; 9C1 -- Ninth Collective Index. Chealtal Abstracts
O
CJ
-------
toxaphene Is a complex mixture and the quality of the commercial product Is
so variable. Thus, the lower chlorinated components will be more water
soluble, and more volatile than those with a higher chlorine content. Odor
detectlbUHy by one Individual of toxaphene dissolved In water has been
found to be 0.14 mg/i at 60°C (Slgworth, 1965).
Selber (1982) noted the occurrence of differential volatilization.
Toxaphene Is mlsdble In acetone, benzene, carbon tetrachlorIde, ethylene
dlchlorlde, toluene and xylene; It Is soluble (g/100 mt at 27°C) In tur-
pentine (350-400), kerosene (>280), and fuel oil (250-275) (Brooks, 1974).
Solubilities In alcohols such as Isopropanol (15-18) and 95X ethanol (10-13)
are generally lower. The heat of fusion 1s 0.39 cal/g; the specific heat Is
0.258 cal/g/°C at 41°C; the viscosity Is 1.4 poise at 100'C (Brooks, 1974).
The octanol/water coefficient was variously found to be -3300 (Paris et al.,
1977) and 825 (Sanborn et al., 1976). The diffusion coefficient of toxa-
phene 1n water at 15°C has been estimated to be 5.6x10"* cm2 sec"1
(Velth and Lee, 1971). The KQC value at 25°C Is 98,600 (McDowell et al.,
1981). The physical properties of these products are provided In Table II-3.
Toxaphene dehydochloMnates when heated above 120°C, or exposed to
ultraviolet light (mercury arc) or Intense sunlight (Brooks, 1974).
Toxaphene (20 mg of a 90% toxaphene/lOX xylene liquid) when placed In a
sealed tube for 15 minutes will produce different IR spectra at 250°C;
volatilization occurs above 125°C (Kennedy et al., 1978). When heated In an
open crucible In a muffle furnace for 45 minutes, the toxaphene/xylene
02000 11-5 02/25/87
-------
o
g TABLE 11-3
o
Physical Properties of the Degradation Products of Toxicant Bd
CAS RN Purity mp Retention Ttmeb *»N1-EC Response Molecular
(X) (°C) (minutes) Relative to Hlrexc Formula
57981-29-0 99 155-156 14.00 23 C10H12C16
64618-63-9 99 107-108 11.15 23 C10H10C16
7 65620-64-6 90 146-148 13.23 13 CloHl2n6
o>
66157-70-8 99 100-101 8.45 11 ClOHllc15
Diagnostic
CI/NS Peaks
M-C1. M-HC1.
M-C1-HC1
HO. H-C1,
H-HC1
H-C1-HC1
H-C1. M-HC1,
M-C1-HC1
M*l. M-C1.
H-HC1
aSource: Saleh and Caslda. 1978
bThe retention time of Nlrex on an open tubular column at 200°C was 45.50 minutes; the retention time of
toxicant B was 20.41 minutes, and Its relative EC-response was 51.
CEC = Electron Capture; Nlrex response Is taken to be 100.
CAS RN - Chemical Abstracts Service Registry Number
o
CO
CD
-------
mixture showed a weight loss of 94.2X at 400°C and 99.9% at 1000°C. Notice-
ably different gas chromatograms were noted at 300°C for the open crucible
experiments. At 400°C, the major gaseous products were tentatively Identi-
fied as hydrochloric add, chlorine, phosgene, carbon tetrachlorlde, chlori-
nated allphatlcs containing two carbons, carbon dioxide, carbon monoxide and
water. In catalytic hydrodechlorlnatlon, the gemlnal chlorines are more
readily attacked than the other chlorines (Kranlch et al., 1978). Thus, <2%
of toxaphene degradation products containing more than five chlorines
remained after 19 hours of reaction (at 100'C, 50 bar pressure); a 10 g
toxaphene 1n ethanol per gram N1 on Kleselguhr was used as a catalyst.
Toxicants A and B appear to be stable under the conditions of gas chromato-
graphy (Khalifa et al., 1974).
DehydrochloMnatlon reactions are accelerated In the presence of a-lkalls
and Iron compounds (Brooks, 1974), that 1s, when toxaphene Is refluxed In
alcoholic potassium hydroxide (Archer and Crosby, 1966; Gomes, 1977; Crist
et al., 1980), or when toxaphene 1s In the presence of hematIn/sodium thlo-
sulfate (Khalifa et al., 1976).
The colorlmetMc technique formerly used depended upon the reaction of
toxaphene with dlphenylamlne and zinc chloride at 205°C to give a chromo-
phore with ax of 640 nm (Graupner and Dunn. 1960).
max
Studies on Toxicant B were reported by Saleh and Caslda (1978). Toxi-
cant B undergoes reductive dechlorlnatlon at the germinal dlchloro group to
yield 2-endo,5-endo,6-exo,8,9,10-hexachlorobornane and 2-exo,5-endo,6-
exo,8,9,10-hexachlorobornane 1n many systems.
02000 II-7 02/25/87
-------
Production and Usage
Toxaphene production In the United States was 18.4 Mkg In 1978, and
accounted for more than 13% of total cyclic pesticide production. Three
manufacturers who reported their production or sales figures for that year
were Tenneco Chemicals, Inc., Hercules, Inc. and Vertac, Inc. Vlcksburg
Plant (USITC, 1979). The United States toxaphene consumption for the years
1978, 1979 and 1980 was 15.5, 13.5 and 13 Mkg, respectively (FAO, 1982). In
1980, -1 Mkg was applied to soybean acreage. This 1980 usage represented
23.8% of the total Insecticides applied to this crop (Elchers and SerleUs,
1982). During fiscal year 1980 toxaphene was also used by the U.S. Forest
Service for tick and lice control on 3030 cattle. A total of 1607 acres
(650 ha) were Involved In this process (USOA, 1981).
Primary utilization occurs 1n agricultural crop applications, especially
cotton (IUPAC, 1979). It 1s also extensively used for the control of exo-
parasltes on livestock. The application rates vary from 1.2-9.6 kg/m3
water (IUPAC, 1979). The cumulative world usage of toxaphene from 1946 to
1974 was -450,000 tons {IUPAC, 1979).
Technical grade toxaphene can be supplied as a 40% dust concentrate and
as a viscous liquid containing 10% xylene (Brooks, 1974), with the liquid
being vulnerable to degradation by metals, such as Iron and aluminum.
Emulslf1able concentrates containing 4, 6 or 8 Ib. of toxaphene/U.S. gallon
(0.48, 0.72 or 0.96 kg/i) are formulated usually with xylene, 0.5%
eplchlorohydrln and an appropriate emulslfler (1f water Is the carrier),
Uettable powders (40% toxaphene content), dusts (5-20%) and granules (5 and
20%) are also used (Brooks, 1974; IUPAC, 1979). Toxaphene has also been
02000 11-8 02/25/87
-------
coformulated with other pesticides [e.g. DOT, Maneb, Z1neb, dioxathlon,
dlazlnon, malathlon, methylparathlon, endrln, endosulfan, Isobornyl thlo-
cyanoacetate plus related toxaphene terpenolds (ThanH'e), pyrethrlns, and
plperonyl butoxlde] (Brooks, 1974). A selected list of recently used
coformulatlons Is provided 1n Table II-4. In addition to such uses toxa-
phene has been used as a protective agent to reduce phytotoxlc effects of
such other pesticides as phosphorothlolates (Aller and Hansen, 1981), to
provide broad spectrum pestlddal properties In combination with such herbi-
cides as benazolln (Anonymous, 1982), and to produce more potent 1nsect1-
ddal mixtures. I.e. with AmHraz (Kerry and Welghton, 1979).
Spectroscoplc Properties
Toxaphene components are not aromatic and, therefore, would be expected
to absorb light at wavelengths below 250 nm.
Infrared analysis 1s still used for qualitative Identification. There
are prominent absorption bands 1n the 6-8 ym region and the specified
absorbence at 7.2 vin of 0.0177 (maximum) 1s utilized (Brooks, 1974). The
Infrared spectrum of toxaphene was reported by Archer and Crosby (1966).
Since toxaphene Is a complex mixture, mass spectroscopy (MS) of the mixture
Is not diagnostic. However, upon gas chromatography/mass spectroscopy
(GC/MS), the presence of the most toxic components can be found 1f adequate
resolution can be obtained on the gas chromatographlc column. The electron
Impact and chemical 1on1zat1on mass spectra of Toxicant A and B (see Table
II-2) were recorded 1n Holmstead et al. (1974). Because the compounds are
02000 11-9 02/25/87
-------
1ABLE II 4
Cheated! Information Regarding loxaphene Mixtures
CAS RN
Molecular forMula
ChcMtcal Na«es
SynonyMS
o
u>
OD
8073-68-5
8077-22-3
37271-99-1
37272-06-3
39295-90-4
39295-93-7
39295-94-8
39400-89-0
39474-75 4
50641-56-0
55267-24-8
58386-60 0
C4HgN202S-Unknown
C|4H9C1.,.CHH,UIIOSPS
Unknown
C8HiQNOsPS-Unknown
Unknown
Unknown
C)4HMM04PS.Unknown
.C5H,2M03
PS2-Unknown
C)oH)4MOi,PS. Unknown
c16H15cl3°2*Unknown
Cj0H)906PS2-Unknown
HhanlMldothlotc actd.N ((d*lnocdrbonyl)oxy)-.
•ethyl ester. «»xt. with toxaphene (9C1)
Phosphorothlotc acld.O.O-
-------
1AHI t 11 -4 (cunl.)
rsj
CAS HN
Molecular foraula
Chemical Nanes
Synonym
O
CJ
CO
58703-96-1
58934-38-6
64034-61-3
67872-25-7
70028-45 4
70162-18-4
73755-37 0
7/5I8-58-?
77518-59-3
77837-51-5
77837-52 6
C,4H,Cl5.C,0Hi4IIOs
PS-Unknown
CH4M20-Unknown
C8H|oMOsPS-Unknown
C4H7Br2Cl204P-
Unknown
Unknown
«0SPS.Unknown
c25H22c1N03*Unknown
Phosphorothlulc actd.O.O-dtethyl-0-(4-n1tro-
phenyl)ester. •III. with toxaphene and 1.1'-
(2.?.^-lrlcbloroethylldene)bls(4-chloroben2ene)
(9C1)
Cyclohexane.l.2.3.4.5.6-hexachloro-.(l alpha.
2 alpha. 3 beta. 4 alpha. 5 alpha. 6 beta)-.
•Ixt. with toxaphene (9CI)
Urea. alxt. with toxaphene (9CI)
Phosphorothlolc acid. 0.0 dlethyl 0 (3.5.6
trtchloro 2 pyrtdtnyl)eiter. atxt. with 0.0
dimethyl-0-(4-nltrophenyl)phosphorothloate
and toxaphene (9C1)
Phosphoric acld.l.2-dlbroMo-2.2-dtchloroethyl
dl«ethyl ester. *lxt. with toxaphene (9C1)
Nethanl«)da«tde. M' <2.4 dl«ethylphenyl) M
(112.4 -dlaethylpheny 1) l>lno)aethyl) -N-Mthyl -.
•lit. with toxaphene (9C1)
Phosphorothlolc acid. 0.0 dlaethyl 0 (4-nltro-
phenyl)ester. alxt. with M' (4 chloro 2-
•ethylphenyH-M.N-dtaethylaethanlMtdaatde
and toxaphene (9CI)
Phosphorothlolc acid. 0-ethyl-S-propy1-0-
(2.4.6-trlchlorophenyl)ester. «lxt. with
toxaphene (9CI)
Phosphorothtolc acid. 0-|4 bro«o 2-chloro-
phenyl)-0-ethyl-S-propyl ester, alxt. with
toxaphene (9CI)
Ben/enedcetIc acId.4 -chloro-aIpha-(1 -
•ethylethyl)-.cyano(3-phenoxyphenyl)iMthyl
ester. «lxt. with toxaphene (9C1)
Cyc lopropanerar boxy lie acld.3-(2.2-dlchloro-
etheny1)-2.2 dlaelhy1 -.(3-phenoxypheny1)
•ethyl ester, ntxt. with toxaphene (9C1)
Hexatox
TXL
loxaphene-naled
•txture
CAS RN =- Che«lcdl Abstracts Service Registry Number; 9CI -- Ninth Collective Index. Chenlca) Abstracts
-------
allcycllc, the mass electron Impact spectra have extremely weak or non-
existent parent Ion peaks, the major Intense peaks being In the low m/e
range. Thus, chemical 1on1zat1on MS after GC 1s more useful than electron
Impact MS (Holmstead et al., 1974).
Although capillary GC showed Toxicant A was one peak, 90 and 270 MHz
'H-NMR spectroscopy revealed 1t to consist of two major components
(Matsumura et al., 1975). The structure of Toxicant Ac was deduced pri-
marily through Its 'H-NMR spectra (Chandurkar et al., 1978), since Us gas
chromatographlc properties were similar to those of components A-l and A-2.
Saleh and Caslda (1978) recorded the chemical 1on1zat1on/mass spectra
(CI/MS) of the major In vitro and U> vivo degradation products of Toxicant B
(see Chapter III). The chemical 1on1zat1on/mass spectrometrlc (CI-MS) data
can be found In Table III-7.
The 'H-NMR spectra of Toxicant C and Us major metabolite (CR 70459-
31-3) (Table III-7) was reported by Chandurkar and Matsumura (1979b).
Analysis
The major recent analytical methods for toxaphene In a variety of
matrices are summarized In Table II-5. Most of the methods employ extrac-
tion from the suitably prepared substrate, a Florlsll clean-up step, and
electron capture/gas chromatography (EC/GC), or GC/MS. In the case of
tissue analysis, a silica gel clean-up step 1s also often utilized before
EC/GC or GC/MS (Haseltlne et al., 1980). A nonpolar phase like OV-101 Is
generally utilized for gas chromatography.
02000 11-12 02/25/87
-------
1ABLE II 5
ExaMples of Analytical Methods for Toxaphene
o
0
o
l~i
1
CJ
G»
00
Substrate
PrlMry sludge
(20 g)
Duck tissues
(brain, liver.
blood, eggs and
carcasses)
Soil (1 g)
Bovine deftbrtnated
whole blood
circulated through
a per fust on
apparatus (2 at)
HI Ik (20 M)
Butter (1 g fat)
Heat (10 g)
Method
Extraction; concentration; florlsll coluwi cleanup; EC/GC
Extraction; florlstl coluwi cleanup; silicic acid separa-
tion; EC/GC (Gel peraeatton chroaatography for blood and
young carcasses)
Florlsll extraction and cleanup;
Dehydrochlorlnatlon with KOH;
GC
I. Agitation with H?0 and hexane;
centrifuge; hexane extraction;
GC
II. Agitation with 88X f orate acid
and hexane; solvent phase shaken
with I^COj; hexane extraction; GC
HI. Agitation with 88X foralc acid;
florlsll extraction and cleanup;
GC
5X Dlethyl ether -hexane elutton; GPC solvent Isolation;
acid oxidation; hexane extraction; florlsll coluwi
cleanup; EC/GC
5X Olethyl ether -hexane elutlon; GPC solvent Isolation:
acid oxidation; hexane extraction; florlsll coluwi
cleanup; EC/GC
5X Dtethyl ether -hexane elutlon; GPC solvent Isolation;
acid oxidation; hexane extraction; florlstl coluwi
cleanup; EC/GC
Concentration Recoveries Reference
Added
50 Mg 92»27X Rodrlquei et al.. 1982
84.2X
91»7X
15 t,g 89»28X
89»19X
93»22X
10-20 ppa 113X Haselttne et al.. 1980
(average) :
1.0 Mg 84. 9X Crist et al.. 1980
0.5 1,9 71. 4X
0.1 M9 71. IX
20 tig/at 73. 4X Nalortno et al.. 1980
20 Mg/*t 71.7X
20 Mg/*i 103.4X
0.3-15 Mg/at 96. 8-102. IX
(range)
0.5-5.0 ppa 78-88X Boshoff and Pretortus.
(range) 1979
0.5-5.0 ppa 79-86X
(range)
0.5 5.0 ppa 76-79X
(range)
-------
Since toxaphene 1s a complex mixture, the chromatographlc pattern of
standards may not match that of the residual toxaphene remaining In environ-
mental samples, since differential volatilization, sorptlon, solub1T1zat1on
or mlcroblal degradation may have occurred that changed the diagnostic
pattern. However, certain components of toxaphene are more toxic than
others; these components can be selectively quantHated by the methods given
In Table II-.5. GC/MS techniques are Invariably the ultimate forms of
analysis after a EC/GC screening step, with chemical lonlzatlon MS being
preferred since electron Impact MS produces no or very weak molecular Ions.
Some of the older methods for detecting toxaphene such as total organic
chloride, coloMmetrlc methods, or bloassays using Insects, were generally
nonspecific. Since toxaphene Is an extremely complex mixture, and not very
heat stable, the question of thermal degradation on GC columns remains a
vexing one. Although there are three major chromatographlc peaks (Gomes,
19.77), the use of a flash heater filled with various reagents In the Injec-
tion port produced substantial changes 1n the chromatogram (Mlnyard and
Jackson, 1965). Treatment of toxaphene residues with alcoholic potassium
hydroxide followed by column chromatography (detection to 1 ng) causes
dehydrohalogenatlon with the appearance of one large single peak (Archer and
Crosby, 1966; Gomes, 1977; Crist et al., 1980). Recoveries from soil are
claimed to average between 71 and 85X at levels from 0.1-1 ppm toxaphene In
soil, although the method did not appear to be useful for rat adipose tissue
(Crist et al., 1980).
02000 11-14 02/25/87
-------
Reviews by Selber (1982) and Pollock and KHgore (1978a) highlight the
difficulties of separating toxaphene from other chlorinated hydrocarbons,
and pesticides.
Analysis for toxaphene metabolites has Involved extraction techniques
followed by thin-layer chromatography (TLC) (Chandurkar and Matsumura,
1979a,b). Pollock and Kllgore (1980) used TLC to circumvent artifacts
caused by thermal degradation on GC columns.
Summary
Toxaphene Is a mixture (average molecular weight 414) of at least 177
compounds, mostly chlorinated camphenes, which has been used as a broad
spectrum contact pesticide (e.g., In soybean and cotton fields, and for
ectoparasites). It Is produced Industrially by the reaction of camphene and
chlorine (In carbon tetrachloMde solvent) 1n the presence of ultraviolet
light to produce a product of highly variable quality. Toxaphene can exist
as a 40% dust concentrate, a 9:1 liquid mixture containing 10% xylene or as
emulslflable concentrates. It 1s extensively coformulated with other pesti-
cides (e.g., DDT, parathlon, endrln).
The vapor pressure, density, solubility and other physical properties
vary according to the quality of the product. Toxaphene Is nonpolar and so
Is mlsdble In nonpolar solvents, but less soluble In polar solvents. It
adsorbs avidly to soils and partitions readily Into n-octanol. Toxaphene,
being aliphatic, Is a poor absorber of ultraviolet light above wavelengths
of 250 nm. Gas chromatography with electron capture or chemical 1on1zat1on
02000 11-15 02/25/87
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mass spectrometMc detection are the most commonly used analytical methods.
'H- and 13C-nuclear magnetic resonance techniques may be used to assess
the "fingerprints" of toxaphene fractions.
Toxaphene readily dehydrochloMnates when heated above 120°C, but Toxi-
cants A and B appeared stable In gas chroroatography. Dehydrochlorlnatlon
also occurs In the presence of alkali (used as an analytical method), of
reduced hematln, and of Iron compounds. Reductive dechlorInatlon occurs on
exposure to ultraviolet light In the radiation range (230-290 nm) of toxa-
phene In organic solvents, preferentially at the C« and C? positions.
Most of the analytical techniques to detect toxaphene metabolites have
utilized thin-layer chromatography as the end step. Toxaphene In environ-
mental samples and tissues often does not have the chromatographlc pattern
of a toxaphene standard so that recognition by gas chromatography occurs
primarily by retention time, and quantHatlon (detection to 1 ng) Is gener-
ally accomplished by the method of alkaline dehydrochlorlnatlon, extraction
and chromatography (Gomes, 1977). Metabolites are generally Isolated by
thin-layer chromatography or liquid chromatography.
02000 11-16 02/25/87
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III. TOXICOKINETICS
This chapter refers only to data relevant to warm-blooded vertebrates.
Residue estimation In tissues after toxaphene exposure 1s generally based on
the comparison of EC/GC traces of a test sample with the pattern of stan-
dards. Systematic errors In residue levels may exist because of the differ-
ential behavior of the >177 components of toxaphene relative to metabolism
can render such comparisons Inappropriate.
Absorption
Toxaphene 1s absorbed through the skin (especially 1f mixed with
xylene), the lung, and the gut (IUPAC, 1979; IARC, 1979). The rates of
absorption have not been documented. The available data are summarized In
Table III-l.
Oral
One Incident that Illustrates toxaphene absorption Involved toxaphene
poisoning of the Tule Lake Basin of Northeastern California and Southern
Oregon (Keith, 1965). Toxaphene, -2.2 kg/ha, was used between 1956 and 1963
In areas on and off the Tule Lake and Lower Klamath Refuges, with heavy use
occurring between 1957 and 1960 (>1000 ha treated/year) In Modoc County.
Though many pesticides. Including DOT, were also used 1n the area, the
wildlife bird mortalities coincided with the significant use of toxaphene.
Toxaphene was found 1n the carcasses and tissues of dead birds In 1960-1961
(0.3-15 mg/kg whole carcasses or organ composites) (see Chapter V for
discussion on residue levels as linked to mortality). Keith (1965) also
reported on the mortality of other birds (Robinson, 1959) In Nebraska and In
Montana upland that were associated with toxaphene application.
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TABLE III-l
Absorption Data for All Routes for Toxaphene
Absorption
Route
Oral
Species
birds
birds
bald
eagle
birds
range
birds
Exposure
2.2 kg/ha
unknown
unknown
unkown
unknown
Effect
kills In N.E. California
and southern Oregon;
residues In carcasses
kills 1n Nebraska
and Montana; residues
In carcasses
death; residues 1n
carcasses
kill; residues In
carcasses
kill; residues In
carcasses
Reference
Keith, 1965
Robinson, 1959
Barbehenn and
Relchel, 1981
Pollock and
Kllgore, 1978a
HcEwen et al . ,
1972
birds
raccoon
horse
pig
unknown
unknown
unkown
unknown
kills 1n California,
South Dakota, Texas
and Arizona
kill; residues In
carcass
poisoning, residues
poisoning; residues
cattle
various
poisoning; residues
In tissues, milk
U.S. EPA, 1977
U.S. EPA, 1977
Mount and
Oehme, 1981
Buck et al.,
1976
Mount et al.,
1980
D1P1etro and
Hallburton,
1979
Claborn et
al., 1963
Zwelg et al.,
1963
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TABLE III-l (cont.)
Absorption
Route Species Exposure
Effect
Reference
Oral (cont.) cattle
various
Oral
Inhalation
Dermal
humans
humans
animals
humans
humans
animals
humans
pig
animals
unknown
unknown
various
unknown
various
various
unknown
(Undane/
toxaphene)
various
various
poisoning; residues In
tissues, milk
three fatal; two
poisoning
poisoning from
contaminated fish;
8 fatalities;
44 poisonings
1059 values
bilateral hllar adeno-
pathy with fine mlHary
lesions on lungs; acute
pneumonia; blood
eos1nophH1a; serum
globulins Increased
no toxaphene residues
(<0.1 mg/kg blood)
L050
Illness
nervous system
disorder
1050 values
Steele et al.,
1978
Steele and
Mount, 1980
Steele et
al., 1980
McGee et al.,
1952
U.S. EPA, 1977
U.S. EPA,
1977,1980
see Table V-l
Warrakl, 1963
U.S. EPA, 1980
Boots Hercules
Agrochemlcals,
Inc., n.d.
Pollock, 1958
D1P1etro and
Hallburton,
1979
see Table V-2
02010
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Toxaphene residues have been measured 1n the tissues of dead wild birds
(U.S. EPA, 1979a). The carcass/brain residue ratio In bald eagles was 6.7,
similar to that of ds-nonachlor, PCBs, oxychlordane and ds-chlordane In
the same species (Barbehenn and Relchel, 1981). The ubiquity of the occur-
rence of toxaphene residues In birds Implies that toxaphene absorption
occurred by the oral and possibly percutaneous routes. In fact, Pollock and
KHgore (1978a) attributed two bird kills to eating of contaminated fish,
and direct toxaphene exposure. McEwen et al. (1972) noted no change In bird
numbers 1 week after spraying toxaphene on a range land, but during the 2nd
week a significant decrease In bird numbers was observed and several bird
carcasses contained 0.1-9.6 mg toxaphene/kg ww. Live birds In the area had
carcass residues of around 0.4-1.0 mg/kg ww. Other cases of bird kills are
summarized 1n Table III-l.
Poisoning Incidents In domestic animals-also demonstrate that absorption
can occur by the oral route (see Table III-l). The U.S. EPA (1977) docu-
mented 38 specific cases of toxaphene exposure to wildlife and livestock
between 1966 and 1975, Including 10 for cattle, 3 for swine, 2 for horses
and 1 for sheep. Animal oral L05Q values In toxlclty experiments depend
on the vehicle; I.e.. readily absorbed oils like corn oil and peanut oil
elicit low LD5Q values, whereas carriers not readily absorbed like kero-
sene yield higher LD5Q values (U.S. EPA, 1980) (see Chapter V).
Evidence for oral absorption In humans resulting In toxaphene poisoning
1s summarized 1n Table III-l and treated fully 1n Chapter VI.
02010 III-4 02/25/87
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Inhalation
The relevant evidence for absorption by the Inhalation route Is summa-
rized 1n Table IH-l.
Dermal
Data In animal experiments (IUPAC, 1979) Indicate that toxaphene can be
absorbed through the skin. However, the toxldty 1s generally an order of
magnitude less than acute oral ID™ values using the same carrier. The
LD5Q values are also very vehicle-dependent (see Chapter V). The values
for the rabbit range from <250 mg/kg for toxaphene applied 1n solution to
<10QO-2000 mg/kg for toxaphene applied as a dust.
01P1etro and HalVburton (1979) reported an episode of toxaphene toxico-
sis In swine after topical application 36 hours earlier with -10 times the
prescribed toxaphene dose (300 mJU of 61X toxaphene stock In 4 i of
H_0) for sarcoptlc mange.
Distribution
Since there are over 177 components of toxaphene that do not metabolize
In the same manner, nor at the same rate; their Identification can be diffi-
cult as a result of the change of the diagnostic EC/GC chromatogram from
that of a reference chromatogram. This problem has been observed 1n milk
analysis (Cairns at al., 1981) and analysis of bird organs (Haseltlnc et
al., 1980). One solution 1s to dechloMnate the remaining toxaphene. This
type of analysis has been performed by Chandurkar and Matsumura (1979b) for
Toxicants B and C In rats and by Saleh and Caslda (1978) for Toxicant B In
rats. Another method 1s to study the metabolism of only the more toxic
02010 III-5 02/25/87
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components of toxaphene. This has been performed by Chandurkar and
Matsumura (1979b), and Saleh and Caslda (1978). A third approach Is to
utilize radlolsotopes. This has been done by Bush et al. (1978) using
broiler chickens, by Chandurkar and Matsumura (1979a), Ohsawa et al. (1975),
and Crowder and Dlndal (1974) 1n rats. However, chromatographlc confirma-
tion Is still necessary for the labeling experiments since the position of
the label defines the significance of the results. Unfortunately, no single
study has compared all methods simultaneously.
In general, toxaphene appears to be rapidly metabolized and Us metabo-
lites quickly excreted In most species, with fat as the preferred tissue of
storage (IUPAC, 1979).
Distribution 1n Animal Tissues
Birds. A number of studies In avlan species such as ring-necked
pheasants (Genelly and Rudd, 1956a) and White Leghorn chickens (Bush et al.,
1977, 1978), suggest that birds can metabolize toxaphene adequately, as long
as the doses are not too high and they can tolerate steady Ingestlon of the
chemical for a period of months without serious effects at the lower doses
(Genelly and Rudd, 1956b; Page et al., 1978), with the exception of altered
cartilaginous structures (Page et al., 1978). Young white pelicans and
young water-fowl are more susceptible to toxaphene (Keith, 1966) (see
Chapter V).
Concentrations of 0, 25, 100, 200 and 300 ppm toxaphene 1n the form of
dietary mash were fed to adult ring-necked pheasants (10/sex) for up to 94
days (Genelly and Rudd, 1956a). The mean doses for 100 and 300 ppm levels
02010 III-6 02/25/87
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were 4.74 and 9.12 mg toxaphene/blrd/day, respectively. Organic chlorine
residues were found 1n fat for all doses >4.74 mg/b1rd/day, and 1n liver at
the 9.12 mg/b1rd/day dose. One bird at the 4.74 mg/b1rd/day dose showed
high residues In the testes.
Toxaphene at 0, 0.5, 5, 50 or 100 ppm diet was fed to five groups of 90
1-day-old female White Leghorn chickens per group for up to 34 weeks (Bush
et al., 1977). The pesticide, as Tox-SOl-6, containing 59% active Ingre-
dient, was first dissolved Into corn oil and then Into a formulated diet.
The level of toxaphene In excisable adipose tissue of 8-week-old birds
Increased with Increasing dietary toxaphene as described by the following:
mg toxaphene/kg body fat = 3.4 F + 33.5 r2 = 0.725
where F 1s the level of toxaphene In the feed In ppm. This amount accounts
for only 50% of the total toxaphene Ingested. Accumulation In 32-week-old
birds was less than In 8-week-old birds despite continued exposure. Bush et
al. (1979) suggested that White Leghorns were able to metabolize or excrete
toxaphene at a rate comparable with dally Intake. Residues were found In
the eggs of bird receiving dietary levels >5 ppm.
Toxaphene-»*Cl was utilized In a study by Bush et al. (1978) to assess
the distribution of the labeled pesticide In broiler chickens. Concentra-
tions of 0, 0.22, 0.40, 2.16 and 3.82 ppm toxaphene-3*Cl were administered
to 1-day-old unsexed broiler chicks (three replicates of 30 birds each).
Feed and water were supplied ad libitum under a regime of continuous
Incandescent light.
02010 III-7 02/25/87
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For toxaphene (T) In tissue (mg/kg ww) versus toxaphene In feed {ppm)
after 8 weeks of continuous feeding (F), the following relationships were
found:
Adipose: T = 4.081 F + 0.765 r2 = 0.96
Gizzard: T = 0.1588 F * 0.0140 r2 = 0.7484
Heart: T = 0.1659 F * 0.0565 r2 = 0.9305
Kidney: T = 0.0942 F *• 0.0400 r2 = 0.7512
Liver: T = 0.0355 F + 0.0625 r2 = 0.4528
Breast Muscle: T = 0.0472 F > 0.0018 r2 = 0.6269
Leg Muscle: T = 0.1307 F * 0.0241 r2 = 0.8669
The Intercepts In these equations are probably not different from zero.
Only In adipose tissue did appreciable amounts of toxaphene accumu-
late, but this still represented only 5% of the consumed toxaphene.
One-year-old black ducks of Anas rubMpes were fed dietary levels of 0,
10 or 50 ppm toxaphene with the toxaphene dissolved In a volume of propylene
glycol equivalent to IX of the total diet (Haseltlne et al., 1980). After
hatching of eggs, the residues 1n ducklings fed on the same diets as their
mothers were monitored at days 14 and 84 post-hatching. At day 14, the
ducklings contained 6.3 and 20.8 mg toxaphene/kg bw for the 10 and 50 ppm
dietary levels, respectively. At day 84, female ducks contained 7.7 and
28.0 mg/kg carcass. GC chromatograms of body residues were similar to the
toxaphene standards.
02010 III-8 02/04/87
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Haseltlne et al. (1980) also performed a one-generation study on the
distribution of toxaphen_fi In black ducks (15 pairs/group). Toxaphene was
administered to newly hatched ducks (F. generation) continuously over 19
months 1n propylene glycol admixed so that the carrier constituted IX of the
duck mash feed. All ducklings obtained from the Fl and Fl breedings
3D
were fed the toxaphene containing diets until 12 weeks of age. The techni-
cal toxaphene concentrations 1n the diet were 0, 10 or 50 ppm. Toxaphene
residues 1n liver, brain, blood and carcass were quantified by direct com-
parison of the total area under the GC curve with the total area under the
standard toxaphene curve (Table III-2) or by comparing the area under..two
characteristic peaks with the area under the same peaks 1n the standard.
The two methods did not agree, Indicating that differential metabolism of
the toxaphene components had occurred. Liver residues for the 50 ppm group
at month 19 constituted only 12% of the cumulative dietary levels. Brain
residues were higher \n males (1-3 mg/kg ww) than 1n females (-0.5 mg/kg ww)
exposed to 50 ppm. Blood levels of the 50 ppm group were not significantly
higher than In the 10 ppm dosed birds. No brain residues were noted at the
10 ppm level. Carcass levels were higher for the 50 ppm treatment compared
with 10 ppm.
Mammals. Lackey (1949a) administered doses of 4 g toxaphene 1n corn
oil/day 1n a gelatin capsule to dogs and determined that brain tissues (and
not fat) at day 106 after treatment showed Increased organic chloride levels
relative to unexposed animals. This was Interpreted by the author to sig-
nify a storage of toxaphene or chlorinated derivatives 1n the brain but not
In the fat. In sheep, steers and dairy cows, storage of toxaphene 1n fat
did occur on repeated exposure (D1ephu1s and Dunn, 1949; Conley, 1952).
02010 III-9 02/04/87
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I—«
I—*
*—•I
1ABLE III-2
Distribution of Toxaphene Continuously Administered Orally to Black Ducks from Birth
by Nixing Toxaphene In Propylene Glycol, which Constituted IX of the Duck Nash*
Dose (ppra
toxaphene In
the diet)
10
50
Nean Concentration (rag toxaphene/kg
Age
Week 2
Week 12
Nonth 19
Week 2
Week 12
Nonth 19
Gender
N/F
N/F
M (6)
F (6)
N/F
N/F
H (6)
F (6)
Generation
Fla
Fib
Fla
Fib
FO
FO
Fla
Fib
Fla
Fib
FO
FO
Carcass
6.3*0.3
3.1*1.3
7.7*0.9
3.U0.2
3.1*0.6
3.0±0.04
20.8*3.0
7. OM. 4
28.0*3.5
11.6i3.4
28.0*5.7
11.U1.4
Liver
NH
NN
NN
NN
0.40*0.08
0.40.*0.07
NN
NN
NN
NN
6.1*1.3
2. 2^0. 4
Brain
NN
NN
NN
NN
<0.1
<0.1
NN
NN
NN
NN
1.4*0.6(5)
0.68±0.45
ww) In
Blood
NN
NN
NN
NN
0.14*0
0.18M)
NN
NN
NN
NN
0.50*0
0.29^0
I
.01
.02
.16
.03
•Source: Haseltlne et al.. 1980
NN = Not measured
CO
-------
Feeding toxaphene at 10 ppm In the diet of sheep and calves for 28 days
revealed no detectable storage In the fat (Claborn, 1956). Feeding cattle
25 and 100 ppm 1n the diet for 16 weeks resulted In the appearance of. 12 and
38 mg toxaphene/kg omental fat, respectively. Eight weeks after feeding
stopped, storage In the 100 ppm diet group diminished to 0.5 mg/kg ww.
Dermal spraying (0.5X solution) led to the appearance of toxaphene residues
In cattle fat of 2.5 mg/kg ww after a single spray (Claborn, 1956). Analy-
sis of fat 4 weeks after 1-2 sprayings of 0.5X solution at 2-week Intervals
showed toxaphene present 1n renal fat. Six weeks after the last spraying,
no toxaphene could be detected In fat. Roberts and Radeleff (1960) sug-
gested that meat from toxaphene-treated swine 1s safe for human consumption
If the animals are slaughtered at least 6 weeks after being sprayed once or
twice with a 0.05X solution of toxaphene.
Clapp et al. (1971) reported the results of a 12-week feeding study with
12 male and 12 female rats provided with feed containing 0, 2.33, 7, 21, 63
and 189 ppm toxaphene. TLC was used to assess levels 1n omental fat and
liver. Levels In .the liver were greatest at week 4, but fat levels
increased continuously, although male rats tended to have maximum fat levels
at week 8. Below the 21 ppm dose, residues 1n fat, liver and carcass were
similar. Above this dose, fat residues were proportional to dose.
Crowder and Dlndal (1974) studied the distribution of 20 mg technical
36C1-toxaphene/kg bw administered orally by stomach Intubation 1n 0.5 ma.
of a peanut oil/gum acacia solution to 30-day-old Holtzman albino rats. The
uptake of the label In various tissues from 3 hours to 20 days (Table III-3)
showed that the highest concentrations In most tissues occurred 1n 12 hours.
02010 III-ll 02/04/87
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o
ro
TABLE 111-3
Percent Uptake of Radioactivity In Various Rat Tissues and Organs following a Single
Dose of »*Cl-Toxaphene (20 mg/kg) Administered In O.S ML Peanut Oil/Green Acacia*
Tissue
Blood Supernatant
Blood Cells
StoMch
Liver
Kidney
fat
Testes
Brain
Muscle
Saall Intestine
First 2 CM
Last 2 CM
Large Intestine
Esophagus
Splten
X Tissue Total
Feces
Urine
X Excretion Total
Total X Recovered
0.125
0.64
3.1
3.7
0.33
O.OS
0.14
0.02
0.03
0.93
0.06
0.10
0.19
0.04
0.04
9.4
9.4
0.25
1.2
0
19
1.1
0.13
0.15
0.08
0.06
1.6
0.34
0.34
0.60
0
0.06
24.3
24
0.50
2.4
0
77
2.3
0.43
0.86
0.28
0.23
5.3
0.34
0.28
1.2
0.04
0.08
90.9
91
1
1.3
0.06
2.0
0.50
0.10
0.57
0.06
0.05
1.3
0.05
0.13
0.19
0.03
0.05
6.4
24
1.5
25
31
2
0.60
0.90
0.63
0.31
0.03
0.31
0.04
0.04
0
0.01
0.01
0.08
0.01
0.02
3.0
7.5
3.2
11
14
Days
3
0.36
2.6
0.61
0
0.03
0.18
0.03
0
0.65
0
0.15
0.02
0.01
0
4.6
1.3
2.9
4.1
8.7
Post-Dosing
456
0
0
0.
0.
0.
0.
0.
0
2.
0
0
0.
0.
0.
3.
1.1 1.
2.4 1.
3.5 2.
6.
.
-
39
01
01
18
03
-
4
-
-
03
02
03
1
1 1.2
8 1.2
9 2.4
0
7
0.18
0
0.16
0
0
0.02
0.02
0
0.40
0
0
0.04
0
0.24
1.1
0.69
1.2
1.8
2.9
8 9
0.09
1.1
0.12
0.48
0.03
3.7
0.06
0.01
0.14
0.84
0
0
0.03
0 0.06
6.6
0.27 0.31
0.54 0.72
0.8 1.0
7.6
20
0.06
1.2
0
0
0 ,
0.03
0
0
o.ai
0.09
0
0
0
0
2.2
•Source: Crowder and Dlndal. 1974.
o
*•
\
CD
-------
All blood levels of labeled toxaphene peaked after 3 days. The data may
Indicate the distribution of toxaphene metabolites 1f metabolism Is fast,
rather than the distribution of toxaphene Itself. In addition, If extensive
dechlorlnatlon occurred the labeled Cl detected might represent Inorganic Cl
rather than organic CT. The former appeared to be the case for the feces
and urine. It appears that the 36C1 1s rapidly secreted with the stomach
acids within 0.50 days after dosing, which Implied that extensive metabolism
had occurred.
Male Sprague-Dawley rats (250-290 g) were administered the 3«C1-
labeled compounds by stomach Intubation at 0 and -13 mg toxaphene/kg bw with
0.15 ml corn oil as the vehicle and with another 0.10 ml as rinse
(Ohsawa et al., 1975). 14C-toxaphene was similarly administered at 0,
8.5, 12.7 and 19.0 mg/kg. In the same study 14C-labeled Toxicants A and B
were administered to 200-225 g rats at 0 or 0.84 and 0 or 2.6 mg/kg bw,
respectively, using 0.25 mi corn oil as the vehicle and also as rinse.
The results given In Table III-4 show that toxaphene and Toxicants A and B
were rapidly eliminated (high excretion percent) as metabolites, and that
toxaphene did not remain as such In tissues to any great extent (14C- and
36C1-toxaphene levels were not comparable). Most of the label resided In
blood, fat, liver and kidneys. The 36Cl-results are generally consistent
with those of Crowder and Olndal (1974) although detectable levels In the
blood and muscle were not observed. At day 14 between 2.2 and 6.6% of the
3«Cl-label should be found In the tissues (Crowder and Olndal, 1974) and
>53X should have been excreted. Since 59X of the 19 mg 14C-toxaphene/kg
dose of toxaphene was excreted to day 14 1n the Ohsawa et al. (1975) study,
results of both studies are compatible. Toxicant A and B appeared to be
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o
I\J
o
o
TABLE 1114
Distribution of Radioactivity in Various Rat Tissues and Organs at Day 14 Following a Single Dose
of a'Cl-Toxaphene, 14C-Toxaphene, Toxicant A and B Administered In Corn Oil by Stomach Intubation3
Tissue
Blood
Liver
Kidney
Fat
Bone
Brain
Heart
Lung
Nuscle
Spleen
Testes
Urine0
Feces0***
Breath »«C02C
X Excreted0
19 rag/kg
NR
0.30
NR
0.78
NR
NR
NR
NR
NR
NR
NR
31.8
27.1
NR
58.9
14C-Toxaphene
12.7 mg/kg
NR
NR
NR
NR
NR
NR
NR
NR
NR
NR
NR
25.4
31.7
NR
57.1
8.5 mg/kg
0.14
0.12
0.17
0.52
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
0.02-0.09
21.3
34.7
1.2
57.2
a*Cl-loxaphene
14.2 mg/kg
NR
NR
NR
NR
NR
NR
NR
NR
NR
NR
NR
49.1
26.9
NR
76.0
Levels (mg
Toxicant A
NR
NR
NR
NR
NR
NR
NR
NR
NR
NR
NR
28.3
47.8
1.8
68.5
label/kq uw)
Toxicant Bb
0.07
0.12 '
0.09
NR
<0.03
<0.03
<0.03
<0.03
<0.03
<0.03
<0.03
26.7
47.8
0.7
75.2
in
\
CD
aSource: Ohsawa et al.. 1975
bAt day 9
cPercent cumulative
dMethanol soluble fraction only
NR = Not reported
-------
quickly dechlorlnated. l4C-Tox1cant 8 was hardly detectable In most
tissues at day 9 except for blood, liver and kidney {Ohsawa et al., 1975).
These results are somewhat different from those described 1n the earlier
report of Clapp et al. (1971) In that no deposition 1n fat was observed, but
the TLC technique utilized by the latter authors may not have been as
specific as the techniques utilized by Crowder and Dlndal (1974) and Ohsawa
et al. (1975).
Hale albino Sprague-Dawley rats were separately administered toxaphene
(13 mg/kg bw); 14C-toxaphene (1.5 mg/kg bw) and 3.1 mg/kg bw each of
Toxicant B; 2-exo,5-endo,6-exo,8,9,10-hexachlorobornane (CAS RN 57981-29-0);
and 2-endo,5-endo,6-exo,8,9,10-hexachlorobornane (CAS RN 65620-64-6); a
mixture of the latter two compounds (0.52/0.95 mg/kg bw, respectively) using
soybean oil (0.15 mi) as a vehicle for administration by stomach Intuba-
tion; and 0.1 mi for the rinse (Saleh and Caslda, 1978). GC analysis
confirmed that all of these compounds underwent dechlorlnatlon, and that
chromatograms derived from samples of fat did not resemble the toxaphene
standard. This was not the case for the liver and feces. The amounts of
Toxicant 8 and Us metabolites 1n fat and liver at 7 and 72 hours after oral
Intubation are provided In Table III-5.
Peakall (1979) administered an acute single dose of 120 mg toxaphene/kg
bw orally 1n a gelatin capsule to male rats. Four animals were sacrificed
at 1, 5 and 15 days. The liver residues were 2.3, 4.2 and 5.7 mg/kg ww,
respectively. The residues 1n brain were below detection limits, 2 and 2.7
mg/kg ww, respectively. When fed 2.4 mg toxaphene/kg bw/day, four animals
02010 111-15 02/04/87
-------
o
o
o
TABLE II1-5
Distribution and Metabolism of Toxicant B (51775-36-1) at 7 and 72 Hours
After Stomach Intubation of Sprague-Oawley Rats at 3.1 rog/kg bw*
uQ/kg ww of CAS RN
Tissue
Fat
Liver
Tine (hours)
post dosing
7
72
7
72
Number
of Rats
3
5
3
5
51775-36-1
453*285
335±44
8.6*4.1
17. 3H1. 7
65620-64-6
8±?
13*5
5.9H.9
2.0^0.6
57981-29-0
14il3
34±8
10.2i2.3
9.2i7.0
1
64618-63-9
15*9
14j;2
3.4±0.9
0.8±0.6
'Source: Saleh and Caslda. 1978
o
00
-------
were sacrificed at 1, 3 and 6 months. The "Mver residues were 28, 26 and 26
rag/kg ww, respectively. Brain residues were 9.3, 13 and 12 mg/kg ww,
respectively. These results are suggestive that steady state plateay levels
can be reached 1n liver and brain 1n chronic studies.
In another study, virgin female Sprague-Oawley rats (318+34 g) were used
and pregnant animals were acutely administered 2.6 mg 14C-toxaphene/kg bw
1n olive oil (0.1 ml) as a vehicle for oral gavage (Pollock and Hill-
strand, 1982). Rat chow and water were available ad libitum. The amount of
label 1n various organs, excreta, fetus and umbilical cord was measured.
The results are provided 1n Table III-6. The urinary excret.lon of the label
by pregnant animals up to day 5 compared well with that of nonpregnant
females (Pollock and Hlllstrand, 1982). The fecal excretion (28.3%) was
slightly lower than that found 1n nonpregnant animals (35.7%). The levels
In the fat for pregnant animals at day 5 (7.1-7.8 mg toxaphene/kg ww) also
agreed well for levels 1n nonpregnant animals after 7 days (6.4 mg toxa-
phene/kg ww). All other tissues contained <10% of the level In fat. The
carcass, cecum, small Intestine and adrenal gland contained levels >0.5 mg
toxaphene/kg ww at day 5. There were Indications that metabolites resided
In fat deposits and 1n the fetus, but these were not specifically Identi-
fied. Transplacental transfer (<1% of the administered dose) also occurred,
as did blood-brain barrier transfer. From Table III-6, the adrenal gland,
carcass, cecum, abdominal fat, GI tract contents, kidney, large Intestine,
liver, ovaries and small Intestines each contained over 1 mg toxaphene/kg ww
at day 1. At day 3, only the adrenal gland, carcass, cecum, abdominal fat,
GI tract contents and small Intestines contained residues >0.5 mg toxa-
phene/kg ww.
02010 111-17 02/04/87
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TABLE III-6
Distribution of_Rad1oact1v1ty 1n Various Female Sprague-Dawley
Rat Tissues and Organs Following a Single Dose of 2.6 mg
14C-Toxaphene/kg bw Dose to Four Pregnant Rats
Administered 1n OUve 011 (0.1 mi) by Oral Gavaged
14C-labeled Material (pg Toxaphene
Equivalent/kg Wet Weight) at
Tissue
Adrenal gland
Brain
Carcassb
Cecum
Fat (abdominal)
Fetus
GI tract0
Heart
Kidney
Large Intestine
Liver
Lungs
Ovaries
Placenta
Small Intestine
Spleen
Stomach
Umbilical
Uterus
Ur1ned
Fecesd'e
Day 1
1092
185
1532
6354
7208
84
6534
349
1195
2757
1210
735
1054
191
6890
171
709
207
154
(8.9+2.0)
(9.3+4.8)
Day 3
577
63
588
1225
8474
30
1995
90
317
370
397
413
k 175
70
629
49
327
145
99
18.3
23.8
Day 5
625-731
57-62
489-924
1012-1136
7107-7845
27-29
NM
71-92
303-347
204-428
363-384
339-407
469-484
47-56
486-1185
50-73
180-351
134-155
56-216
22.0
28.3
aSource: Pollock and Hlllstrand, 1982
^Contains blood and subcutaneous fat
cContents of GI tract at termination
^Percent cumulative for four animals per day
emethanol soluble radioactivity
NM = Not measured
02010 111-18 02/12/85
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Distribution In Human Tissues
Apart from the Instances noted In the absorption section, no residue
data, or metabolites of toxaphene have been found 1n human tissue, even
though 44 cases (Including deaths) Involving exposure of humans were docu-
mented between 1966 and 1975 In the United States (U.S. EPA, 1977). In an
occupational study, exposure of 54 workers primarily by the Inhalation or
dermal routes did not generate detectable levels of toxaphene 1n the blood
(>0.1 mg/kg blood) 1n spite of the fact that 49 of the personal air samples
showed detectable amounts of toxaphene (U.S. EPA, 1980).
The above findings signify that toxaphene may either be quickly metabo-
lized and transported In another form In humans, or that no absorption has
occurred. In view of the many human and animal poisoning Incidents Involv-
ing toxaphene, H 1s more likely that toxaphene 1s metabolized quickly and
then transported.
Metabolism
Toxaphene undergoes fast reductive dechlorlnatlon, dehydrochlorlnatlon
and hydroxylatlon 1n mammals. Little Is known about the mechanism of
transport.
Toxaphene was thought to be slowly detoxified In such animals as sheep,
steers and cows by excretion of ethereal sulfate and "glucuronate" (Conley,
1952). Crowder and Dlndal (1974) In their distribution study of
"Cl-toxaphene reported 1n the section on "Distribution In Animal Tissues"
observed much redistribution of »*C1 label with time. Host of the
3*Cl-labeled toxaphene was excreted (see Table III-3) within 6-7 days and
little remained In the tissues.
02010 111-19 02/04/87
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Ohsawa et al. (1975) confirmed the observations of Crowder and Dlndal
(1974) with 3«Cl-toxaphet)e (see Table III-4). The only Identified metabo-
lite was chloride Ion that appeared almost entirely In the urine and
accounted for about half of 3*Cl-toxaphene eliminated. The dechlorlnatlon
occurred mostly within two days. 14C-toxaphene distribution as well as
that of Toxicants A and B showed only a small amount unmetabollzed In the
feces (3X for 14C-toxaphene) and that the metabolites probably Included
acidic materials partially or completely dechloMnated, and 14CO? (2% of
14C-toxaphene). The ease of dechlorlnatlon appears to be shared by all
components of toxaphene, even Toxicants A and B.
Khalifa et al. (1976) examined the reactions of toxaphene and Toxicants
A and B with two Iron (II) protoporphyrln systems, hematln reduced with
sodium thlosulfate and mlcrosomal cytochrome P-450 reduced with NADPH. They
found that toxaphene reacts with reduced hematln 1n neutral aqueous medium
to cleave about half of the C-C1 bonds, yielding derivatives with shorter
retention times on gas chromatography and of reduced sensitivity for detec-
tion by electron capture. The system also converts Toxicants A and B to
products formed by reductive dechlorlnatlon, dehydrochlorlnatlon and a
combination of these reactions. Extensive metabolism of toxaphene and
Toxicants A and B by rat liver mlcrosomes required NADPH.
From these results It Is clear that many toxaphene components, Including
Toxicants A and B, are converted by one or both of these Iron (II) protopor-
phyMns to more polar derivatives (as evidenced by TLC). The studies with
02010 111-20 02/25/87
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reduced hematlh establish that toxaphene undergoes extensive dechlorInatlon
and that the products from Toxicants A and B determined by GC-CI-MS are
formed by two or more of the following: reductive dechlor1nat1on, dehydro-
chlorlnatlon and vicinal chloride elimination.
Toxaphene has been shown to yield type I binding spectra with hepatic
cytochrome P-450 of rats, mice, sheep and rabbits, which suggests that
toxaphene may serve as a substrate for the hepatic mlcrosomal MFO system
(Kulkarnl et al., 1975). Type II binding has not been observed.
In another rat metabolism study, Chandurkar and Matsumura (1979a) found
that dechloMnatlon and oxldatlve degradation by MFO Involving cytochrome
P-450 are active degradation mechanisms for toxaphene.
Toxaphene metabolism 1n chickens and 6 species of mammals (guinea pig,
hamster, rabbit, mouse, rat, monkey) Involves extensive dechloMnatlon; and,
1n the case of rats, ~50% of the dose 1s excreted as chloride 1on (Saleh and
Caslda, 1979; Saleh et al., 1977, 1979).
Saleh and Caslda (1978) showed that the major products of Toxicant B
produced by rat liver mlcrosomes are CAS RN 57981-29-0 and 65620-64-6 In a
2:1. Table III-7 lists synonyms, chemical names and Indexing terms for the
major metabolites and degradation products of Toxicant B. These products
were not detected under aerobic conditions or under aerobic or anaerobic
conditions 1f NADPH was absent. These products were also found (see Table
III-5) 1n fat, liver and feces 1n addition to CAS RN 64618-63-9 after oral
02010 111-21 02/04/87
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1ABIE 111-7
Identified Metabolites of loxaphene Components
o
to
o
CAS RH
NT
Chemical Abstracts Indexing lerm (9C1)
Synonym
•nls
Reference
$7981 WO
o
o
*.
\
GO
64618-63-9
CIOMIOC'6
6S620-64-6
66157-70-8
C10H11C15
70459-31 3
Blcyclo[2.2.1)heptane.2.3.6 trtchloro
l.7.7.-trls(chloromethyl) ,(2-exo.3 endo.
6-exo)-
or
2-exo.S-endo.6-exo.8.9.10-hexachloro-
bornane
Blcyclo(2.2.1]hept-2-ene,2.5.6-trIchloro-
1.7.7-trts(chloromethyl)-.5 (endo.6 exo)-
or
2.5-endo.6-exo.8.9.10-hexachloroborn-
2.3-cne
Blcyclo(2.?.l)heptane.?.3.6. trlchloro 1.
7.7 trts(chloromethyl) ,(2-exo.3 endo.
6-endo)-
or
2-endo.S-endo.6-6X0.8.9.10-hexachloro-
bornane
Trlcyclol2.2.1.02.*]heptane.3.4-dlchloro-
1.7.7-trls(chloromethyl)-.
or
2.5-endo.8.9,10-pentachloroiricyclene
Blcyclo)2.2.1Jheptane.2.3.5.5 tetrachloro
7.7-bts(chloromethyl)-l-(d1chloromethyl)-
(exo.exo)-
or
3.3.5.6-exo-8.9.10.10-octachlorobornane
Reductive dechlor(nation of toxaphene
toxicant B In: reduced heaatln system.
bovine rumen flutd. rat liver micro-
somes . and rats Jn vivo
fecal metabolite of toxaphene toxicant
B In: chicken, mouse, rat. hamster.
guinea pig. rabbit and monkey
Reductive dehydrochlorInatton of
toxaphene toxicant B In: reduced
hemattn system and rats \n vivo
Fecal metabolite of toxaphene toxicant
B In: chicken, mouse, rat. hamster.
guinea pig. rabbit and monkey
Reductive dechiorInatton of toxaphene
toxicant B In: reduced hemattn system.
bovine rumen fluid, rat liver micro-
somes, and rats In vivo
Fecal metabolite of toxaphene toxicant
B In: chicken, mouse, rat. hamster.
guinea pig. rabbit and monkey
Reduced hemattn conversion of
toxaphene toxicant B
loxaphene toxicant C metabolite In rat
liver In vitro
Saleh and Caslda. 1978
Saleh et al.. 1979
Saleh and Caslda. 1978
Saleh et al. 1979
Saleh and Caslda. 1978
Saleh et al.. 1979
Saleh and Caslda. 1978
Chandurkar and Hatsumura. 1979b
CAS RN = Chemical Abstracts Service Registry Number; Nf -- Molecular lornula; 9CI -- Ninth Collective Index. Chemical Abstracts
-------
administration by gavage of 3.1 mg Toxicant B/kg bw to male albino Sprague-
Oawley strain rats. The-rat1o of CAS RN 57981-29-0 to 65620-64-6 was 1.8 In
fat, and 1.7 1n the liver 7 hours after administration'. At 72 hours, the
ratios were 2.6 and 4.6, respectively, and 2.5 1n the feces. At least 7.4%
of the administered dose was eliminated after 72 hours 1n rats by reductive
dechlorlnatlon at the germinal dlchloro group. The chromatograms of the
residues In fat almost conformed to the toxaphene standard but chromatograms
of liver and feces samples did not. The latter samples, however, did con-
tain toxaphene components.
Chandurkar and Matsumura (1979a) reported that In the presence of NAOPH,
the liver mlcrosomal fraction of rats metabolized 3*Cl-toxaphene and
14C-toxaphene after 2 hours at 37°C to the extent of 22 and 9.5%, respec-
tively. In the presence of glutatMone (GSH), the extent of metabolized
3*C1 and 14C toxaphene were 11 and 4.8%, respectively; and with cystelne
as a cofactor, the respective values were 10 and 3.6%. Inhibitors of NADPH
dependent mixed-function oxldases and of glutath1one-S-transferases
Inhibited conversion of toxaphene to polar water-soluble metabolites. The
results depended on which fraction was utilized from rat liver homogenates.
For example, one NAOPH dependent mixed-function oxldase Inhibitor caused
100% Inhibition of conversion by a lOO.OOOxg supernatant fraction compared
with 53% utilizing a 20,000xg supernatant fraction from the same homogen-
ate. When Toxicants B (Gig) and C (Clg) were Investigated In the
20,000xg supernatant under the same conditions as toxaphene, the extents of
metabolism were ~73 and 56%, respectively In the presence of NAOPH; the
respective figures for glutathlone were 31 and 47%. Whereas the extent of
Toxicant C metabolism In the presence of NAOPH was complete within 1 hour,
02010 111-23 02/25/87
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the rate of Toxicant 8 conversion was constant with time to 3 hours. The
control showed an ability to metabolize Toxicant B and C without added
NADPH, Toxicant C being more prone 1n this regard. TLC then EC/GC revealed
that different components In toxaphene were metabolized by the NADPH system
as compared with the GSH system. Most of these metabolites were more polar
than the parent compounds {TLC data), some being hydroxyllc (esterlf1cat1on
data), and some being still chlorinated (EC/GC data). Treatment of water
soluble 14C-toxaphene metabolites with B-glucuron1dase 1n the presence of
NADPH revealed an Increase of 16.IX 1n ether extractable material originally
present as glucuronldes. Smaller Increases of 7.8 and 8.6X were shown In
the presence of B-sulfatase and dilute hydrochloric add, respectively.
Addition of UDP-g1ucuron1c add to a toxaphene/NADPH system Increased the
extent of metabolism 8-fold, but not 1n the additional presence of a NADPH
dependent mixed-function oxldase Inhibitor. These results show that ox 1 da-
tive metabolism Is Important for toxaphene. The water-soluble metabolites
of GSH-fort1f1ed Incubates are probably GSH-conjugates formed Involving loss
of a chlorine from the original toxaphene components.
In another report of Chandurkar and Matsumura (1979b), using the same
20,000xg supernatant system fortified with NADPH (see Chandurkar and
Matsumura, 1979a), the major dechlorlnated metabolite of Toxicant C In vitro
was shown to have a molecular formula of c-|QHincl8 ^MS data). This
compound has been designated as 3,3,5,6-exo-8,9,10,10-octachlorobornane (CAS
RN 70459-31-3) (see Table III-7). Oxidation products of Toxicants B and C
were not tertiary alcohols, but 4 products of Toxicant C were secondary
alcohols, and one was a primary alcohol. No Toxicant B metabolites were
primary alcohols. None of the secondary alcohols were dlols. NADPH caused
02010 111-24 02/04/87
-------
more Toxicant B metabolism than for Toxicant C, but no alcohol metabolites
of Toxicant B could be-found, thus being suggestive of the occurrence of
reductive dechloMnatlon rather than hydroxylatlve dechlorlnatlon. This may
Indicate different loci for metabolism of Toxicants B and C.
Pollock and Hlllstrand (1982) reported that the metabolites measured In
the fetus are apparently different from those measured In the fat of
,Sprague-Dawley dams based on TLC analysis. No metabolite Identification was
reported.
Elimination
In a study where White Leghorn chickens were fed 5, 50 or 100 ppm
toxaphene 1n the diet for 34 weeks (Bush et a!.. 1977), the half-life of
toxaphene 1n adipose tissue was 42, 25 and 20 days, respectively.
To determine the length of time necessary for depletion of 36Cl-toxa-
phene residues from adipose tissue of broiler chickens, 15 birds from each
of the replicates (45 birds/treatment group) were fed toxaphene-frce feed
from week 6 onwards (Bush et al., 1978). The half-life of toxaphene In
adipose tissue was found to be 2.66, 2.76, 2.47 and 2.5 weeks for birds fed
0.22, 0.40, 2.16 and 3.82 ppm toxaphene In the diet, respectively. Data
were obtained from three birds, each sacrificed at Intervals of 2 weeks for
10 weeks.
Black ducklings were fed dietary levels of 0, 10 and 50 ppm technical
grade toxaphene In propylene glycol (1% of a commercial duck feeder mash)
until the egg laying season (1976) was over (Haseltlne et al., 1980). A
02010 111-25 02/25/87
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select number of eggs (Fg generation) were allowed to hatch, and the F,
generation ducklings were_s1m1larly exposed up to the next egg laying season
(1977). In 1976, the six eggs from the FQ generation ducks samples con-
tained 4.4^0.4 and 23.5^1.9 mg/kg ww for the 10 and 50 ppm concentrations,
using total area GC estimation. In 1977, the respective figures for the two
levels were Increased to 5:6^0.6 and 29.5+2.0 mg/kg ww. In 1977 the levels
In both the eggs and adult carcases were similar except 1n females fed 50
ppm toxaphene. Carcass levels for females were 11.U1.4 compared with egg
levels of 29.5^2.0. The Investigators suggest that residues are not concen-
trated In eggs, since eggs contained a similar toxaphene load as carcasses.
Both adults and young birds can metabolize toxaphene and excrete enough that
little accumulation takes place In the organs.
The half-life of 14C- or 3*Cl-labeled toxaphene In rats after single
oral doses appears to be less than a week, with most of the elimination
occurring 1n the urine and feces (Crowder and Olndal, 1974; Ohsawa et a "I.,
1975). Only a small portion of the urine and fecal metabolites are elimi-
nated as glucuronld-e or sulfate conjugates (Chandurkar, 1977), In contrast
to early findings by Conley (1952).
Ohsawa et al. (1975) found that 49% of »*Cl-label was eliminated In
the urine 14 days after administration of 14.2 mg 3«Cl-toxaphene/kg bw
(see Table III-4); In the same time 25.4% of the label from a 12.7 mg
14C-toxaphene/kg bw dose was eliminated 1n the urine. The bulk of the
"Cl-label (90%) was chloride 1on. After 14 days, 27% of "Cl-label was
found 1n the feces. Thus, 1n 14 days, 76% of 3*Cl-label was found 1n the
urine and feces. Subfractlonated toxaphene gave essentially the same
02010 111-26 02/04/87
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results. Unmetabollzed toxaphene comprised 3.4% of the feces residues, and
around 27% 1n urine and-feces appear to be organic* that are dechloMnatlon
products. Between 5 and 10% are completely dechlorlnated.
The data for Toxicants A and 8 are compared with that for 14C-toxa-
phene In Tables III-4 and III-8. In general, Toxicants A and B appear more
readily eliminated but their dosages were lower than for toxaphene Itself,
and only one animal was tested. Unmetabollzed compound constituted only 8.1
(Toxicant A) and 2.6% (Toxicant B) of fecal residues, respectively.
In a similar set of experiments (Saleh and Caslda, 1978), a 1.5 mg
14C-Tox1cant B (CAS No. 51775-36-1)/kg bw dose was given to Sprague-Dawley
rats by stomach tube with soybean oil as the vehicle. Only 0.2^0.1% of
Unmetabollzed Toxicant B was found In the feces at 0-72 hours, along with
2.U0.6, 5.3+1.6 and 1.0*0.3% of the administered label as CAS RN
65620-64-6, 57981-29-0 and 64618-63-9, respectively. The combination CAS RN
65620-64-6 and 57981-29-0 constituted 7.4% of the administered label, and
constituted products of reductive dechlor1nat1on at the germinal dlchloro
group. When a mixture of CAS RN 65620-64-6 and 57981-29-0 (0.95/0.52 mg/kg)
was administered to the same strain of rat, the feces contained 45-47% of
the Unmetabollzed compounds collected up to 72 hours. The feces contained
many peaks of shorter retention time than those 1n toxaphene standards.
Chandurkar and Natsumura (1979a) also utilized four male Sprague-Dawley
albino rats to administer a dose of 15 mg 14C-toxaphene/kg bw by a stomach
tube (0.25 ml of corn oil). Fifty-seven percent of the label was excreted
In the feces. Only 9% of the administered dose was found 1n the urine. On
02010 111-27 02/25/87
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TABLE III-8
Elimination of 14C-Toxaphene, Toxicant A and Toxicant B Administered
to Sprague-Qawley Rats by Stomach Intubation 1n Corn 011 Carrier3
Label Eliminated
(Percent of Administered Dose)
Toxaohene
Sample
Urine
Feces
Breath
1*C02
Totals
Time After
Treatment
(days)
0-1
1-2
2-14
0-1
1-2
2-14
0-1
2-14
(8.5 mg/kg)
3 rats
10.6
6.3
4.4
34. 7b
0.8
0.4
57.2
(19 mg/kg)
2 rats
11.3
11.8
8.7
27. lb
NM
NM
58.9
Toxicant A
(0.84 mg/kg)
1 rat
18.9
6.0
3.4
27.4
6.0
5.0
0.8
1.0
68.5
Toxicant 8
(2.6 mg/kg)
1 rat
15.7
6.2
4.8
32.3
9.7
5.9
0.5
0.2
75.2
aSource: Ohsawa et al.t 1975
blt Is not clear 1f these figures are totals for days 0-14 or the same
measurement for each time after treatment.
NM = Not measured
02010
111-28
02/25/87
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the average, 8.9% of the urine metabolites and 0.7% of the fecal metabolites
were glucuronldes; 9.5%_jof the urinary and 7.5X of the fecal metabolites
were add-hydrolyable products; only a small percentage (0.7%) of the total
fecal metabolites are sulfate conjugates.
A single spray treatment of cows with 2 quarts of 0.5% toxaphene sus-
pended 1n water did not produce detectable toxaphene In the milk (Lelghton
et al., 1952). Spraying cows twice at 3-week Intervals with 2 quarts of
0.5% toxaphene emulsion or suspension resulted In peak levels of 0.51-0.82
mg/i milk 1 or 2 days after the second spraying. Twenty-one days after
spraying had ceased, the toxaphene concentrations returned to preexposure
levels (Claborn, 1956). Spraying cows under exaggerated conditions (twice
dally for 21 days with 57 mg of a 2X oil solution of toxaphene) resulted In
peak residues of 0.11-0.50 mg/l milk after -3 days. Twenty-one days after
spraying ceased, the concentration diminished to 0.06 mg/i milk (Claborn,
1956).
Keating (1979) reported on toxaphene residues In cow milk from cows
dipped once or twice weekly 1n AU1k [0.25% w/w toxaphene plus 0.03% w/v
D1oxath1on (Delnav)]. Residues ranged from 27-45 mg/kg milk fat, with maxi-
mum residues found one day after dipping. When dipping was discontinued for
a cow dipped In AH1k once weekly, the levels decreased to 5 mg/kg milk fat
after 19 days. Even at 32 days, toxaphene could still be detected (3.1
mg/kg).
Pollock and Hlllstrand (1982) found that excretion In urine and feces
for pregnant rats (Sprague-Oawley) 1s very similar to that In virgin female
02010 111-29 02/04/87
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rats, although there was a large weight difference. The results for
pregnant females are provided In Table III-6, which Indicate a half-life of
-5 days.
Toxaphene was administered to cows continuously at Increasing dally
concentrations ranging from 1.7-12 ppm diet. Toxaphene levels were first
detected 1n the milk of cows In the 4 ppm group after 3 weeks. Excretion
equalled 7-8 mg toxaphene/l milk 1n the 12 ppm group. When exposure was
discontinued, the concentration 1n milk diminished promptly (Shaw, 1947).
In cows fed contaminated hay for 16 weeks, the toxaphene levels )n milk
(50-410 mg toxaphene/l) were proportional to the amount In the diet, with
the average excretion ranging from 2.3-18.2 mg toxaphene/l milk (Bateman
et al., 1953). Claborn (1960) fed four groups of three cows each 20, 60,
100 and 140 ppm toxaphene In the diet, respectively. The excretion was
proportional to the toxaphene concentration. Once exposure ceased, residues
diminished to 13-17% within 1 week, followed by a slow excretion phase
probably related to the mobilization of fat reserves.
Cows were fed 0-20 ppm toxaphene 1n the diet for 77 days; milk samples
were analyzed twice weekly (Zwelg et al., 1963). The results confirmed
previous findings at higher feed levels that toxaphene residues declined
rapidly after toxaphene exposure had terminated. Residues ranged from
0.043-0.179 mg toxaphene/l milk, and were concentration-dependent.
Following cessation of exposure, residues 1n milk decreased to undetectable
levels after 2 weeks for levels lower than 10 ppm toxaphene. For the 20 ppm
level, residues were detected 30 days after feeding contaminated diet was
halted.
02010 111-30 02/04/87
-------
In another study Involving 12 cows fed 5 ppm toxaphene 1n the diet
(Cairns et al., 1981), toxaphene metabolites and components were found In
milk fat with the EC/GC chromatograms being enriched In the components of
shorter GC retention time. Six milk samples contained residues estimated by
CI/SIM/GC/MS to range from between 7 and 280 mg/l milk. An EC/GC study
using toxaphene standards estimated the range to be 10-620 mg toxaphene/l
milk. The noncorrespondence of residues to Intake may reflect the time of
sampling since this was not specified. However, this does Illustrate that
the analytical methods for toxaphene may yield different answers when
enrichment or depletion of selected compounds 1n toxaphene occurs.
A summary of elimination half-life data for various animal species 1s
provided 1n Table III-9. Saleh and Caslda (1979) reported that quick metab-
olism and elimination occurs 1n rats, mice, guinea pigs, hamsters, rabbits
and chickens, with the monkey metabolizing toxaphene most extensively and
rapidly, and chickens the least and slowest. The order of decreasing rate
of metabolism 1s as follows: monkeys, rats, hamsters > mice, rabbits,
guinea pigs > chickens for a dose of Toxicant B at 3 mg/kg bw. The data for
toxaphene In Table III-9 are In general agreement with the elimination data
for Toxicant B, with half-lives being <1 week for all mammals, Including man.
Summary
. Though no precise quantitative data are available, the symptoms of
toxicosis as well as the presence of residues and metabolites after exposure
1n animals, birds and humans Indicate that toxaphene can be absorbed through
the skin (especially 1f mixed with xylene), the lung, and the gut.
02010 111-31 02/04/87
-------
TABLE III-9
Experimental or Calculated Half-Life of Toxaphene Elimination In Various Species
Animal
Broiler chicken
(Hubbard-Hubbard)
Cattle
Cow
~ Rat (Holtzman)
i
Hale rat
(Sprague-Dawley)
Male rat
(Sprague-Dawley)
Male rat
(Sprague-Dawley)
Female rat
( Sprague-Dawley )
o Humans
^>
o
Dosea
(carrier)
0.22-3.82°
100° for 16 weeks
57 mg of 2X oil
solution sprayed
twice dally for
21 days
20 (peanut oil/
green acacia)
8.5-19
(corn oil)
0.52-0.95
(soybean oil)
15 mg
(corn oil)
2.6
(olive oil)
Catfish fillet
52 mg/kg fillet
Compound
»*C1 -label
Unlabeled
Unlabeled
•*Cl-label
"Cl-label
••Cl-label
Toxicants A.B
"Cl-label
Toxicant B
"Cl-label
"Cl-label
Unlabeled
Route Half-Life
(days)
adipose (18.2H)
adipose 5-6(calc.)
milk 21
feces; 6-7
urine
feces; <14
urine
feces 3
feces 4-5
feces; 5
urine
blood 6-7
Reference
Bush et al.. 1978
Claborn. 1956
Claborn. 1956
Crowder and Dlndal.
1974
Ohsawa et al.. 1975
Saleh and Caslda.
1978
Chandurkar and
Hatsumura. 1979a
Pollock and
Hlllstrand. 1982
U.S. EPA. 1978
CD
aln mg toxaphene/kg body unless specified otherwise
bppm diet
-------
In birds (ring-necked pheasants, ducks and leghorn layers), toxaphene
can be tolerated for months with the most sensitive effect being altered
cartilaginous structures. Extensive dechlorlnatlon of toxaphene occurs with
chlorinated organic residues occurring mostly In the fat. White Leghorns
were able to metabolize or. excrete toxaphene at a rate comparable with dally
Intake. In ducks, brain residues appeared at doses >10 ppm In the feed.
In mammals, storage of toxaphene 1n the fat has been reported In sheep,
steers and dairy cows. Accumulation In cattle occurred at exposures to
25-100 ppm toxaphene In the feed for 16 weeks. In contrast, dogs appeared
to store toxaphene derivatives preferentially 1n the brain. In male or
female rats, fat levels of toxaphene were relative to concentrations >21 ppm
1n the diet. Studies with labeled toxaphene revealed quick dechlorlnatlon
and subsequent elimination of toxaphene, Toxicant A and Toxicant B at doses
<19 mg toxaphene/kg bw administered by gavage to male rats using corn oil as
the carrier. Toxaphene derivatives resided In the blood, fat, liver and
kidneys. When male rats were chronically dosed with 2.4 mg toxaphene/kg
bw/day, plateau levels were found In the liver and brain after 1, 3 and 6
months. Transplacental transfer as well as blood-brain barrier transfer
occurred 1n rats. The adrenal gland, carcass, cecum and abdominal fat of
female rats contained toxaphene derivatives 3 days after acute dosing with
2.6 mg toxaphene/kg bw. Extensive dechlorlnatlon and rapid elimination of
metabolites occurred also 1n guinea pigs, hamsters, rabbits, mice and
monkeys. No residue data or metabolites of toxaphene In humans have been
reported 1n the available literature.
02010 111-33 02/25/87
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In rats, successful detoxification appears to require NADPH and oxygen.
Toxicants A and B underwent reductive dechloMnatlon, dehydrochloMnatlon,
and vicinal chloride elimination. Involvement of the MFO system Is sug-
gested by the observation of Type I binding spectra with the hepatic cyto-
chrome P-450 of rats, mice, sheep and rabbits, as well as the enhanced tox-
Idty to mice of toxaphene In the presence of plperonyl butoxlde. There
also appeared, to be a detoxification pathway dependent on GSH. Toxicant B
resisted metabolism more than Toxicant C. Glucuronlde (major) and sulfate
(minor) conjugates have been detected as metabolites. Toxicant C also pro-
duced one primary and four secondary alcohols whereas reductive dechlorlna-
tlon predominated for Toxicant B. These data suggest that toxaphene com-
ponents are not metabolized necessarily at the same loci In the mlcrosomal
system.
Elimination of products derived from toxaphene has been demonstrated to
occur In the feces, urine and 1n expired air of rats. The fecal route
appears to be the more predominant route than In urine. Milk and eggs also
showed residues. The species order of decreasing elimination efficiency for
Toxicant B Is as follows: monkey, rat, hamster > mouse, rabbit, guinea pig
> chicken.
02010 111-34 02/25/87
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IV. HUNAN EXPOSURE
This chapter will be submitted by the Science and Technology Branch,
Criteria and Standards Division, Office of Drinking Hater.
02020 IV-1 02/25/87
-------
IV. SOURCES OF HUMAN EXPOSURE - TOXAPHENE
This chapter summarizes the available data on use,
environmental fate, and occurrence to characterize the
potential for exposure to toxaphene. A more extensive
discussion of the information on toxaphene in the environment
is presented in the draft document entitled "Occurrence of
Pesticides in Drinking Water, Food and Air" (U.S.EPA, 1987).
Humans may be exposed to toxaphene from a variety of
sources, including drinking water, food and ambient air. •
Individual exposure to toxaphene will vary widely based on
factors such as where a person lives, works and travels,
and what a person eats. Intake of toxaphene will be affected
by age, weight-and lifestyle.
The Exposure Estimates section of this chapter presents
available information on the range of human exposure to and
intake of toxaphene from drinking water, food and ambient
air. It is not possible to provide an estimate of the number
of individuals experiencing specific exposures from these
sources. However, this section provides some insight into
the sources' relative contributions.
IV-1
-------
A. Use/Environmental Fate
Toxaphene is a pesticide that was widely used during the
1960s and 1970s primarily on agricultural crops and livestock
In 1974, domestic production of the compound was estimated
at 103 million pounds, with approximately 74 million pounds
applied for agricultural uses (U.S. EPA, 1977). By 1982,
domestic usage had declined to about 16 million pounds, with
only 5.9 million pounds used on field crops (USDA, 1983).
In November of the same year, EPA published in the Federal
Register its intent to cancel or restrict registrations for
products containing toxaphene. All major uses of toxaphene
were cancelled (47 FR 53784)(USEPA 1982). In addition, all
existing stocks of the compound could be used in certain
specified situations until December 31, 1986. Formulations
unsuitable for conversion to uses in any of the specified
situations were discontinued after December 31, 1983.
Toxaphene is no longer commercially available (Berg, 1986).
Once released into the environment, toxaphene is very
persistent in both soil and surface waters. Toxaphene is
relatively insoluble in water and binds readily with soil,
consequently it is fairly resistant to leaching from the soil
column. Biodegradation is not a factor in removing toxaphene
IV-2
-------
from aerobic soils, with reported half-lives of up to 20
years. However, biodegradation in anaerobic soils can remove
up to 50 percent of the compound in 6 weeks. Volatilization is
the only important removal process for toxaphene in shallow
soils. The persistence of low levels of toxaphene in surface
waters over several years suggests that volatilization,
biodegradation, hydrolysis, and adsorption to sediments are
not rapid removal processes (Callahan et al., 1979). Toxaphene
is a mixture of a number of chlorinated compounds; most of
these compounds are expected to bioaccumulate.
B. Occurrence
Drinking Water
Under the interim drinking water regulations, all
community drinking water suppliers are required to monitor
for toxaphene. Since the regulations took effect, no system
has reported a violation of the 5 ug/L interim standard (FRDS,
1984).
The 1978 National Rural Water Survey, as well as several
regional surveys, provide information on the occurrence of
toxaphene in public drinking water supplies. According to
IV-3
-------
available information/ toxaphene concentrations did not
exceed the minimum quantification limit of 0.17 ug/L in the
71 groundwater systems tested (U.S. EPA, 1984). Regional
studies of toxaphene in the 1970's have generally reported
that toxaphene, when it was detected, did not occur at levels
greater than 0.1 ug/L (U.S. EPA, 1987). Current toxaphene
concentrations in drinking water supplies are believed to be
even lower than in the 1970's, due to the restrictions placed
on the use of the compound in 1982 and its later removal from
production.
Diet
Although toxaphene is no longer used, the pesticide is
still expected to occur at low levels in many foods because
of its past widespread use and persistence in the environment.
Toxaphene has been detected in the FDA market basket surveys
(FDA, 1986). Surveys conducted between 1982-1985 reported
finding daily intakes of toxaphene for 25-30 year old males
and females of 0.576 and 0.369 ug/day, respectively. Dietary
levels are expected to decrease in the future because of the
discontinuation of the use of toxaphene.
IV-4
-------
Air
According to U.S. EPA (1980) and Arthur et a).. (1976),
particularly high toxaphene concentrations were reported, in
several agricultural regions of the southern United States in
the early and mid 1970's, with levels ranging up to 1.747 ug/m
The highest reported toxaphene level was 8.7 ug/m^ in Arkansas
in 1970 (Kutz et al., 1976). In the 1970's, toxaphene was
widely used as an agricultural insecticide, and the data from
the 1970's reflect this use. Due to the discontinuation of
toxaphene in the last 5 years, ambient air levels today are
likely to be very low.
C. Exposure Estimates
The following table summarizes the current exposure
levels of toxaphene in drinking water, food and air. The
drinking water levels are based on the range of the levels
found in the regional studies in the 1970's. The estimated
daily intake levels were made based on the assumption of an
intake of 2 liters per day of drinking water. Current levels
are expected to be lower. The dietary exposures are taken
from an average of several total diet studies taken from
different regions of the United States from 1982 to 1985.
IV-5
-------
This number is a reasonable estimate of the average dietary
intake of adult males between the ages of 25-30 years old.
Some individuals would be expected to have a higher intake
than the average.
IV-5
Exposure Estimates for Toxaphene
Reported Exposure
Levels (low-high)
Est imated
Adult Intake
Drinking Water
Diet
Air
0-<0.1 ug/L
Negligible
0-<0.2 ug/day
0.6 ug/day
Negligible
The current available information on occurrence of
toxaphene is insufficient to determine the national distribution
of intake by any of the three routes. However, EPA believes
that intakes from diet will generally be less than 0.6 ug/day
and that air exposure to toxaphene is expected to be negligible.
If toxaphene does occur in drinking water at levels of more
than a few tenths of a ug/L, it is likely to be the major
source of intake. However, for the majority of individuals
in the United States diet is the major source.
IV-6
-------
D. REFERENCES
Arthur, R.D., J.D. Cain, and B.F. Barrentine. 1976.
Atmospheric levels of pesticides in the Mississippi
delta. Bull. Environ. Contain. Toxicol. 15 (2 ): 129-134 .
Berg, G.L. (ed.). 1986. Farm chemicals handbook. Meister
Publishing Co., Willoughby, OH.
Callahan, M., et al. 1979. Water-related environmental
fate of 129 priority pollutants. Final Report. EPA-
44014-79129a. Office of Water Planning and Standards.
U.S. Environmental Protection Agency, Washington, D.C.
FDA. 1986. Food and Drug Administration. Memorandum from
E. Gunderson, Division of Contaminants Chemistry, Center
for Food Safety and Applied Nutrition, Washington, D.C.
to Dr. Paul S. Price, Office of Drinking Water, U.S.
Environmental Protection Agency, November 6, 1986.
Washington, D.C.
FRDS. 1984. Federal Reporting Data System. Computer
printout, dated April 4, 1984, containing data on organic
chemical MCL violations, FY 1979-1983. U.S. Environmental
Protection Agency, Washington, D.C.
Kutz, F.W., A.R. Yobs, and H.S.C. Yang. 1976. National
pesticide monitoring programs. In: R.E. Lee (ed.).
Air Pollution from Pesticides and Agricultural Processes.
CRC Press, pp. 95-136, Cleveland, OH.
USDA. 1983. U.S. Department of Agriculture. Inputs.
Outlook and situation. Washington, DC: Economic Research
Service, U.S. Department of Agriculture. IOS-2.
U.S. EPA. 1977. U.S. Environmental Protection Agency.
Toxaphene: Position Document 1. Washington, D.C.
Special Pesticide Review Division, U.S. Environmental
Protection Agency. EPA/SPRD-80/55. Washington, D.C,
U.S. EPA. 1980. U.S. Environmental Protection Agency.
Ambient water quality criteria for toxaphene.
EPA-440/5-80-076. Office of Water Regulations and
Standards, U.S. Environmental Protection Agency.
Washington, D.C.
IV-7
-------
U.S. EPA. 1982. U.S. Environmental Protection Agency.
Decision document on toxaphene. Office of Pesticide
Programs, U.S. Environmental Protection Agency,
Washington, D.C. November 29, 1982. 47 FR 53784.
U.S. EPA. 1984. U.S. Environmental Protection Agency.
Rural water survey. Computer data provided by
Department of Sociology, Cornell University, Ithaca,
NY.
U.S. EPA. 1987. U.S. Environmental Protection Agency.
Occurrence of pesticides in drinking water, food and
air. Office of Drinking Water, U.S. Environmental
Protection Agency, Washington, D.C.
IV-8
-------
V. HEALTH EFFECTS IN ANIMALS
Acute and Subchronlc Toxldtv
Current studies of acute toxldty resulting from exposure of experimen-
tal animals to toxaphene can be divided Into two main groups: exposure to
unfractlonated (technical grade) toxaphene and exposure to specific toxa-
phene components or subfractlons, e.g., Toxicants A and 8. Tables V-l, V-2
and V-3 summarize the L0c0 values observed 1n each of these experimental
situations. Greater than 10-fold differences 1n toxldty have been docu-
mented for various toxaphene fractions or components that differed from each
other 1n chemical composition, polarity and solubility (Pollock and KHgore,
1978a,b).
Information on the acute oral toxldty of unfractlonated toxaphene to
laboratory animals 1s summarized In Table V-l. In cases of acute Intoxica-
tion, toxaphene like most chlorinated hydrocarbon Insecticides, appears to
act as a CMS stimulant. However, unlike DOT, toxaphene does not signifi-
cantly affect conduction 1n the rat superior cervical ganglion (WhHcomb and
Santoludto, 1976). Effects of toxic exposures In humans (hypersens1t1v1ty,
tremors and convulsions) are similar to those observed In both rats and dogs
(Lehman, 1951). Along with convulsions, hyperreflexla has also been noted
In dogs (Lackey, 1949a), and rats (Boyd and Taylor, 1971).
It Is Important to note that humans who Ingest a protein-deficient diet
may represent a susceptible population. Rats fed a protein-deficient diet
were more susceptible to toxaphene poisoning than rats fed laboratory chow,
LD5Q, 80+19 and 220*33 mg/kg bw, respectively (Boyd and Taylor, 1971).
02030 V-l 02/14/85
-------
o
co
o
TABLE V-l
Acute Oral ToxicIty of Technical Toxaphene (CAS RN 8001-35-2) to Laboratory Namnals
Species
Vehicle
(mg/kg bw)
Reference
Rats:
Unspecified strain
Wtstar. male,
(3-4 weeks. 50-60 g)
low protein diet
optimal diet
Sherman, male.
(>90 days. >175 g)
Sherman, female.
(>90 days. >200 g)
Sprague-Dawley. male
(24 days. 70 g)
Unspecified strain.
male
o
ro
CD
en
Mite
corn oil
cottonseed oil
peanut oil
peanut oil
peanut oil
corn oil
olive oil (l.p.)
unspecified
corn oil
unspecified oil
60
220 t 33a
80 * 19
293 *• 31a
Lehman. 1948; Boots Hercules
Agrochemlcals. Inc.. n.d.
Boyd and Taylor. 1971
90(67-122)b Galnes. 1960
80(70-91)b Galnes. 1960
40 Shelanskl and Gellhorn. n.d.
120-125 Shelanskl and Gellhorn. n.d.
228 Pollock et al.. 1983
270 Kuz'mlnskaya et al.. 1980
112 Boots Hercules Agrochemlcals.
Inc.. n.d.
80 Rico. 1961
-------
TABLE V-l (cont.)
o
ro
o
CO
Species Vehicle
Nice
Swlss-Uebstci , male corn oil
Cats peanut oil
unspecified oil
Dogs corn oil
kerosene
peanut oil
deodorized kerosene
< Rabbits peanut oil
^ deodorized kerosene
Guinea pigs corn oil
kerosene
unspecified oil
Hamsters:
Syrian Golden, male unspecified
L050
(rag/kg bw)
112
25 40C
100
20-25
>400
>30C
>450C
75-100c
250-500C
270
365
80
200
Reference
Fattah and Crowder. 1980
Treon et al.. 1950
Rico. 1961
Lackey. 1949a
Lackey. 1949a
Treon et al., 1950
Treon et al.. 1950
Treon et al.. 1950
Treon et al.. 1950
Boots Hercules Agrochemtcals,
Inc.. n.d.
Boots Hercules Agrochenilcals.
Inc.. n.d.
Rico. 1961
Cabral et al.. 1979
rvi
er
GO
and female. 6 weeks
Calves:
Mixed-breed. 136-232 kg
60X solution,
unspecified. (Cooper
Tox Emulslftable
Concentrate) diluted
with 2 l water
62
Steele et al.. 1980
-------
TABU V-l (cont.)
IV)
o
Species
Cattle
Sheep
Goat
Horse
Monkey: female
Pheasant:
Unspecified strain
Mallard:
Anas platyrhynchos
Hen:
Unspecified strain
Vehicle
grain
xylcne
xylene
unspecified
peanut oil saline
(l-v.)
unspecified
unspecified
unspecified
L°50
(nig/kg bw)
144
200
200
80C
7.5-10
40
71
100
Reference
Boots Hercules Agrochemlcals.
Inc., n . tl .
Boots Hercules Ayrochemlcals,
I nc . , n . d .
Boots Hercules Agrochenitcals.
Inc., n.d.
Rico. 1961
Treon et al.. 1950
Netcalf. 1981
Hudson et al.. 1972
Rico. 1961
aStandard error of the mean
b95X confidence Interval
cN1n1mum lethal dose
oo
-j
-------
TABLE V-2
Acute Dermal 1050 Values for Toxaphene In Laboratory Mammals
Species
Rats
Sherman, male
(>90 days, >175 g)
unfasted
Sherman, female,
(>90 days, >175 g)
Rats
Rats, male
Rabbits
Vehicle
xylene
xylene
xylene
unspecified
dust
L050
(mg/kg bw)
1075
(717-1613)*
780
(600-1014)*
930
940
-4000
Reference
Galnes, 1960,
1969
Galnes, 1960,
1969
Boots Hercules
Agrochemlcals,
Inc., n.d.
Ku?'m1nskaya
et al., 1980
Boots Hercules
Rabbits
peanut oil
-250
Agrochemlcals,
Inc., n.d.
Boots Hercules
Agrochemlcals,
Inc., n.d.
*95X confidence Interval
02030
V-5
02/12/85
-------
o
Gi
O
TABLE V-3
Acute Toxlctty of Technical Toxaphene Components Administered IntraperHoneally to Ntce
Toxaphene Component
(CAS RN)
Species/Strain
Vehicle
1050
(mg/kg bw)
Reference
i
0>
2.?.5-endo-6-exo-8.
9,10-heptachloro-
bornane
(51775-36-1)
2.2.S-endo-6-exo-8.
9,10-heptachloro-
bornane
(51775 36-1)
2.2.S-endo-6-exo-8.
9,9,10-octachloro-
bornane
(58002-19-0)
Technical toxaphene
mice/Albino
male
mice/
unspecified
mice/
unspecified
mlce/SwIss-
Uebster. male
rats/Sprague-
Dawley. male
unspecified
unspecified
unspecified
dimethyl
sulfoxlde
olive oil
75 Saleh et al.. 1977
6.6 Khalifa et al.. 1974
3.1 Khalifa et al.. 1974
33(27-41)* Pollock and Kllgore. 1980
228 Pollock et al.. 1983
*95X confidence limits
o<
V.
00
-------
Clinical signs of depression and stimulation of the CNS were the same In
both groups; however, signs appeared earlier and at lower concentrations 1n
protein-deficient rats. Pathological effects Included cloudy swelling and
congestion of the kidneys, fatty degeneration and necrosis of the liver, and
decreased spermatogenesls. Mehendale (1978) reported that toxaphene (100
ppm In the diet for 8 days) Inhibited hepatoblHary function In rats.
The acute dermal toxldty of toxaphene Is summarized 1n Table V-2.
Toxaphene appears to be somewhat less toxic when administered dermally
although toxldty appears to be carrier-dependent. In rats the ratio of
dermal to oral LD5Qs 1s 12 (1075/90) for males and 9.8 (780/80) for
females (Galnes, 1960, 1969). Dermal LDc-s for rats range from 780-1075
mg/kg bw (Galnes, 1960, 1969; Boots Hercules Agrochemlcals, Inc., n.d.).
A 40% technical toxaphene dusi (3-4 g/ma) killed around half of an
exposed group of rats after 1 hour (Boots Hercules Agrochemlcals, Inc.,
n.d.). Assuming that a 200 g rat Inhales 200 ml of air per minute and
absorbs 100% of the Inhaled dose, a 3 g/m3 concentration for 60 minutes
would equal 72 mg/kg bw.
Repeated (14 applications) administration of a 20% solution of toxaphene
1n kerosene to the eyes of rabbits (0.01 ml) and guinea pigs (0.05 ml)
caused mild Irritation to the eyelids with loss of hair of the eyelids.
There was no Injury to the eye and condition on the Hds cleared completely
In 10 days (Boots Hercules Agrochemlcals, Inc., n.d.).
02030 V-7 02/14/85
-------
Hyde et al. (1978) compared the acute electrophyslologlcal effects of
representative organochloMne and organophosphate Insecticides on laboratory
rats. Concurrent neurological, respiratory and cardiac Involvement were
primary concerns. Each of 14 male Sprague-Dawley rats (240-280 g) was
equipped with dural electrodes to record spontaneous cortical electro-
encephalograms. The animals were given a minimum of 7 days to recover from
electrode Implantation and were randomly assigned to 1 of 7 treatment
groups, 2 rats/group. Treatments consisted of separate lethal 1.p. Injec-
tions of each of three organochlorlne pesticides, Including 70 mg toxa-
phene/kg bw and three organophosphates emulsified 1n a Tween-80 1soton1c
saline solution. Controls received 0.5 ml of the vehicle only. Total
percentage changes 1n cardiac and respiratory rates during exposure to toxa-
phene (expressed as percentage change from pretreatment to late stage of
toxlclty) were ECG rates, +-17.5 and respiration, +56.5. The distortion of
spontaneous cortical potentials recorded during toxaphene exposure revealed
a cerebral sensitivity.
Toxaphene 1s used extensively for external parasite control In domestic
animals. The maximal nontoxlc concentration for topical application In
adult swine 1s a 1.5X solution. 01P1etro and Hallburton (1979) reported
signs of a CNS disorder In about 40 of a herd of around 150 feeder pigs.
Thirty-six hours earlier the affected animals had been treated for sarcoptlc
mange with a toxaphene mixture applied at -10 times the manufacturer's
labeled dosage (300 ml of 61% toxaphene stock In 4 i H?0). One day
after the treated pigs were sprayed with warm water, severely affected pigs
had Improved markedly. By day 5 the animals appeared normal.
02030 V-8 02/14/85
-------
Deaths and abnormal behavior In a herd of 50 crossbred beef cows In good
body condition were attributed to an Ingestlon of toxaphene-contamlnated
feed (Braun et al., 1980). Levels of 11-35 mg toxaphene/kg hay were
detected 1n four samples from pastures where the animals grazed.
With the objective of determining tissue levels of toxaphene following
exposure to a single oral dose of toxaphene, Steele et al. (1980) exposed
heifer calves ranging 1n weight from 136-232 kg to the following doses In
mg/kg bw (number of animals exposed to each dose 1n parentheses): 50 (6),
100 (7), 150 (6). Toxaphene originally available as a 61% solution was
diluted with -2 l of water. Surviving animals were sacrificed after 7
days. In the low dosage group, there were 2/6 deaths after 4 days; In the
100 mg/kg exposed group there were 6/7 deaths within 4.5-42 hours, and In
the high dose group, there were 5/6 deaths within 21 hours to 5 days. Clin-
ical signs of toxIcHy Included apprehension and hyperexcHabUHy, followed
by anterior, then posterior, muscle faslculatlons. Terminal stages were
characterized by generalized muscle twitching and clon1c-ton1c convulsions.
When toxaphene tissue levels were determined, 1t was observed that while
kidney and brain toxaphene levels were not dlagnostkally significant, liver
residues of toxaphene >4 mg/kg ww were associated with lethality for the
7-day observation period (Steele et al., 1980). By contrast, brain levels
of toxaphene may be Indicative of acute toxldty; as a result of Inadvertent
dermal exposure In swine, >4 mg/kg ww In the brain was found to constitute a
lethal level, and 2 mg/kg was associated with clinical signs of toxldty
(OlPletro and Hallburton, 1979). A summary Is provided In Table V-4. Acute
Intoxication of horses has also been reported following Ingestlon of alfalfa
contaminated with DDT and toxaphene (level not reported) (Nazarlo and Capel-
laro, 1980).
02030 V-9 02/14/85
-------
TABLE V-4
Residue Concentrations 1n Dead Birds and Animals
Animal
Organ Residue Range3
(mg/kg ww)
Reference
Female bat, Tadarlda
brasH1ens1s Hexlcana)
Rabbit (2/31 )b
Deer (3/22)b
Heifer calves (6/7)b
Swine
carcass
carcass
carcass
liver
brain
1.2-12.4
1.7-8.7
4 mg/kg was
critical level for
acute exposure
4 mg/kg was
critical level for
acute exposure
Geluso et al.,
1981
Causey et al.,
1972
Causey et al.,
1972
Steele et al.,
1980
DIPIetro and
Hallburton, 1979
Bald eagle (8/49)b
( 20/49 )b
(7/50)b
(ll/50)b
(13/69)b
(22/69)b
Great blue heron (9/35)
(6/36)b
Cattle egret (3/3)b
(2/3)b
Great egret (1/1 )b
(1/1 )b
brain
carcass
brain
carcass
brain
carcass
brain
carcass
brain
carcass
brain
carcass
0.13-1.2 (1975) Kaiser et al.,
0.06-2.5 (1975) 1980
0.05-0.31 (1976)
0.05-0.55 (1976)
0.06-2.7 (1977)
0.06-0.88 (1977)
0.17-0.82 Ohlendorf
(1972-1978) et al., 1981
0.11-0.50
(1972-1978)
0.28-0.36 (1978)
0.11-0.16 (1978)
0.54 (1978)
0.58 (1978)
aRange In positives
bPos1t1ves 1n total birds and animals examined
02030
V-10
02/14/85
-------
The acute toxldty of toxaphene components In mite Is listed In Table
V-3. In male Sprague-Dawley 24-day-old rats, the less polar toxaphene frac-
tion, Rf 0.35-0.85, and a more polar fraction, R 0-0.57, 1s Included In
this table and exhibited LD5Q values of 217 and 266 mg/kg bw, respective-
ly, compared with toxaphene, R, 0-0.85, LD5Q 228 mg/kg. The route of
administration was 1.p., and the solvent used 1n the toxldty determination
was olive oil (Pollock et al., 1983).
In a comparative study certain polychlorobornanes that are known to be
components of toxaphene have been examined for structure-toxldty relation-
ships (Saleh et al., 1977; Saleh and Caslda, 1979); using 2,2,5-endo, 6-exo.
8,9,10-heptachlorobornane as a standard with a relative toxldty of 100
(LDcQ 1.p. 75 mg/kg bw 1n male albino mice). An enhanced acute toxldty
was associated with the Introduction of a single chloro group In the 8- or
9-pos1t1on, and to a lesser extent In the 5-exo-position. Pretreatment of
mice with plperonyl butoxlde (which modifies cytochrome P-450-med1ated oxl-
datlve or reductive detoxification mechanisms) was associated with lower
acute LD5Q values (2- to 8-fold) for the heptachlorobornane compound and a
mixture of Its 8- or 9-monochloro derivatives. Acute toxldty was reduced
relative to the standard when monochloro groups were Introduced 1n either
the 3-exo or I0-pos1t1ons, but the toxldty of the 10-chloro derivative was
also enhanced 2- to 8-fold by pretreatment of mice with plperonyl butoxlde.
Dehydrochlorlnatlon of the hexachlorobornenes with apparent removal of the
6-chloro group yielded a hexachlorobornane with only slightly Increased
acute toxldty relative to the parent compound.
02030 V-ll 02/14/85
-------
LD5Q values for 20-29 g male Swiss-Webster mice were compared for
toxaphene and for three toxaphene fractions dissolved 1n dimethyl sulfoxlde,
and administered 1.p. Only one fraction, with an L05Q of 20 mg/kg bw
(19-21, 95X confidence Intervals), was more toxic than toxaphene (33 mg/kg)
Itself (Pollock and Kllgore, 1980).
Table V-5 summarizes the effects of acute and subchronlc oral adminis-
tration of toxaphene to laboratory mammals. Except for convulsions observed
1n dogs given 10 mg/kg bw/day (Lackey, 1949a), few of the exposures detailed
In Table V-5 resulted 1n clinical signs of toxaphene poisoning.
Lackey (1949a) exposed dogs of an unreported breed and sex both acutely
and subchronlcally to toxaphene. Dosing was performed by stomach tube with
the toxaphene dissolved 1n corn oil or kerosene for acute experiments and
corn oil administered as a single dose dally 1n a gelatin capsule for longer
term exposures. None of the three animals receiving a single dose of 5
mg/kg bw 1n corn oil developed convulsions as compared with 4/5 treated with
10 mg/kg. At doses >15 mg/kg the convulsions were accompanied by Increasing
mortality. Of those treated with toxaphene dissolved 1n kerosene, the
minimum dose at which animals had convulsions was 75 mg/kg; none were
recorded for two dogs dosed with 25 mg/kg or one dog receiving 50 mg/kg.
These effects may actually Indicate a possible vehicle effect.
Dally treatment with 5 mg/kg toxaphene resulted 1n the onset of convul-
sions after a few days. For subchronlc treatment the dosage was therefore
decreased to 4 mg/kg bw. Two dogs were treated dally at this level for 44
days and two for 106 days. One dog treated for 106 days and both dogs
02030 V-12 02/14/85
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TABLE V 5
Acute and Subchronlc Oral Toxtctty of Toxaphene
C3
CO
O
Species
Nice, albino and
wild strains
Albino rats
Rats
Rats
Rats. SherMan.
* Mile and feMale.
- 100,
Rats
Rats. Osborne-
Nendel
Rats. Osborne-
Nendel
NuMber of Vehicle
AnlMals
5-18 diet
12N. 12F diet
at each
level
12N. 12F diet
unknown NS
6N. 6F diet
at each
level
4 diet
SN. SF acetone
at each
level
SN. SF acetone
at each
level
Duration
Several weeks
or Months
4. 8 and 12
weeks •
12 weeks
7 Months
2-9 Months
8 days
6 weeks
6 weeks
Dose
(Mg/kg/day or
ppM In the diet)
SO Mg/kg/day
(250-480)*
2.33
7.0
21.0
63.0
189.0
189*
1.2-4.8 Mg/kg/day
SO and 200*
100*
160*
320*
640*
1280*
2S60*
1280*
2S60*
SI 20*
Response
Changes In blood cheMlstry
and urine protein
No effect on weight.
liver cell histology
No apparent adverse effects
Temporary change In blood
cheMlstry
Questionable liver pathology
Decreased bile production
No deaths and Mean weight gains
unaffected (1280). 2 deaths
(both F) and Mean weight gains
of survivors not adversely
affected (2S60)
Two deaths (IN. IF) at 2S60
Reference
BluMler. 197S
Clapp et al. ,
1971
Clapp et al..
1971
Crebenyuk. 1970
Ortega et al..
19S7
Hehendale. 1978
NCI. 1979
NCI. 1979
CO
-------
1ABU tf-S (cont.)
CJ
O
Species
Nice. B6C3F1
Dogs
Nuaber of Vehicle
AnlMls
5H. 5F acetone
»t each
level
3 corn oil
S corn oil
2 corn oil
Duration
6 weeks
1 exposure
1 exposure
44 days
Dose
(•g/kg/day or
PPM In the diet)
40*
80*
160*
320*
640*
1280*
S wi/kg/day
10 ag/kg/day
4 Mg/kg/day
Response
? deaths (1 H. 1 F) and mean
weight gains unaffected (320).
6 deaths (4 H. 2 f ) (640)
No convulsions
Convulsions In 4/5 animals
Questionable liver pathology:
Reference
NCI. 1979
Lackey. 1949a
Lackey. It49a
Lackey. 1949a
corn oil
106 days
4 ag/kg/day
renal tubular degeneration;
sporadic convulsions
Questionable liver pathology:
renal tubular degeneration;
sporadic convulsions
Lackey. 1949a
•pp» toxaphene In the diet
NS > not specified; N - male; f - feaale
CO
-------
treated for 44 days lost weight during the course of exposure. Convulsions
were reported to occur tfnly occasionally. There were no changes In complete
blood counts throughout the experiment and no gross pathological changes
were found. Upon hlstopathology the liver showed generalized hydropic
degenerative changes, which appeared to be reversible, and degenerative
changes In the parenchyma.
In both the Lackey (1949a) study (using dogs) and the Ortega et al.
(1957) study (using rats) changes 1n liver histology were noted 1n the
animals dosed with toxaphene. Morphologically, these changes appeared as
vacuoles of plasma with occasional red blood cells appearing within hepatic
cells. This condition, referred to as hydropic accumulation, Is distinct
from fatty degeneration. In neither rats nor dogs was hydropic accumulation
associated with the destruction of hepatic cells. Ortega et al. (1957) also
noted occasional masses of red blood cells Invading the cytoplasm of liver
cells In areas of centrolobular-cell hypertrophy and marg1nat1on was
observed In 50% of the rats fed a diet containing 200 ppm toxaphene from 2-9
months. Similarly, 27% of rats fed 50 ppm showed this same effect. No con-
trol rats were examined however.
In addition to liver damage, Lackey (1949a) documented widespread
degeneration of the tubular epithelium of the kidney, occasionally accom-
panied by Inflammation of the pelvis of the kidney for dogs fed 4 mg/kg
bw/day for 44 and 106 days. Similar pathological changes were seen In dogs
and rabbits who survived prolonged dermal exposures [200-600 mg/kg bw for up
to 32 days (dogs); 100-1000 mg/kg bw for up to 30 days (rabbits)] to toxa-
phene (Lackey, 1949b). For dogs, the lowest lethal dose was 600 mg toxa-
02030 V-15 02/26/87
-------
phene/kg bw; for rabbits, the lowest lethal dose was 600 mg/kg bw when dis-
solved In mineral oil or dimethyl phthalate, with the highest, nonlethal dose
being 500 mg/kg bw when applied as a dust for 5-14 applications {Lackey,
1949b). Ortega et al. (1957), however, did not note any pathological
changes attributable to toxaphene 1n the kidneys of rats fed a 50 or 200 mg
toxaphene/kg diet for up to 9 months.
Allen et al. (1983) studied the effects of Ingested toxaphene on the
Immune response of female Swiss-Webster mice. A total of 130 8-week-old
mice were given feed containing 10, 100 or 200 ppm toxaphene for 8 weeks
(acetone solvent). Humoral antibody production, IgG antibody formation, was
suppressed 1n mice receiving the 100 and 200 ppm doses. Cell-mediated
Immune responses were not affected by toxaphene treatment.
Clapp et al. (1971) fed technical grade toxaphene to groups of 12 male
and 12 female albino rats (144 total) 1n their diets for 4, 8- and 12 weeks
at levels of 0, 2.33, 7, 21, 63 and 189 ppm. Four animals from each group
were sacrificed at each time Interval. Two male rats died during the test
period but their deaths did not appear to be caused by toxaphene levels In
the diets. Toxaphene had no measurable adverse effects on physical appear-
ance, gross pathology, weight gain or liver cell histology of any of the
experimental animals.
Cattle and sheep have been fed toxaphene at concentrations as high as
320 ppm In hay for 134-151 days. Central nervous stimulation with muscle
tremors occurred In steers at the highest dose, but not In sheep receiving a
similar dose. There appeared to be no abnormal hlstopathology or blood
chemistry (Boots Hercules Agrochemlcals, Inc., n.d.).
02030 V-16 02/26/87
-------
Guinea pigs suffered 73X mortality when solutions of 20% toxaphene \n
mineral oil, equivalent to a dose of 332 mg toxaphene/kg bw/day, was admin-
istered for 14 days (Boots Hercules Agrochemlcals, Inc., n.d.).
Feeding studies were conducted to estimate the maximum tolerated doses
of toxaphene In Osborne-Mendel rats and B6C3F1 mice for a National Cancer
Institute study of cardnogenlclty (NCI, 1979). Toxaphene was dissolved In
acetone and added to the feed. Corn oil was also added as a dust suppres-
sant 1n an amount equal to 2% of the final weight of feed. Treatment
groups, consisting of 5 male and 5 female animals 1n each group, were given
food with or without toxaphene for 6 weeks, and they were observed for
another 2 weeks.
For rats, toxaphene was added to the feed In 2-fold Increasing concen-
trations, ranging from 160-2560 ppm feed. A second study was performed on
male and female rats at toxaphene levels ranging from 1280-5120 ppm to con-
firm the results and to extend the concentration range of the first study.
At 1280 ppm 1n the first and second studies, there were no deaths among
rats and mean weight gains of both sexes were comparable to controls (see
Table V-5). This Is a NOAEL for mortality over a 6-week period. At 2560
ppm, two female rats died In the first study but mean weights of the surviv-
ors were not adversely affected. During the second study, one male and one
female died at 2560 ppm. On the basis of these results, the low and high
doses for the chronic studies were set at 1280 and 2560 ppm for males, and
640 and 1280 ppm for females.
02030 V-17 02/26/87
-------
In the same 1979 NCI subchronlc study, mice were given feed containing
40-1280 ppm toxaphene 1n_ the diet. Four males and two females died at 640
ppm, and one male and one female given 320 ppm died. Mean weight gains of
mice given 320 ppm were comparable with those of controls. On the basis of
these results the low and high doses for the chronic studies were set at 160
and 320 ppm for males and females, respectively. The 160 ppm diet dose can
be regarded as the NOAEL In mice with respect to mortality over a 6-week
period.
Toxaphene aerosols 1n the form of dusts are more toxic to rats than In
the form of mists. M1st concentrations as high as 500 mg toxaphene/m3
caused no mortality In rats and rabbits over a 3-week period. However, no
rats survived dust concentrations of 250 mg/ma for 1 week. Rats, dogs and
guinea pigs were killed at 12 but not at 4 mg dust/m3 over a 3-month
period of exposure. Some surviving female rats exhibited slight focal
hepatic cell necrosis (Boots Hercules Agrochemlcals, Inc., n.d.).
Chronic ToxIcUy
Long-term exposures to low dietary levels of toxaphene are summarized In
Table V-6. All studies note some form of liver pathology 1n rats at levels
>100 ppm 1n the diet. At this same dose, cytoplasmlc vacuollzatlon similar
to that seen following subchronlc oral exposure was noted by Kennedy at al.
(1973). Lehman (1952) also reported fatty degeneration of the liver 1n rats
fed 100 ppm. With a 25 ppm diet, Fltzhugh and Nelson (1951) observed
Increased liver weight with minimal liver cell enlargement. Unpublished
studies on rats, dogs and monkeys by Boots Hercules Agrochemlcals, Inc.
(n.d.) are In general agreement with the above published reports.
02030 V-18 02/14/85
-------
TABLE V-6
0
IVi
o
co
o
Chronic Toxlclty of
Duration
Species of Feeding
(number of animals)
Toxaphene to Laboratory
Toxaphene
Concentration
(ppm diet)
Rats. 3 generations 25
Sprague-Dawley (42 weeks for FQ
(64N. 128F. and 39 weeks for 100
weanling) Fj and F2>
to
o
Rats lifetime
Rats lifetime
Dogs 2 years
2 years
25
100
25
100
(20N. 20F)
1000-1500
(20M. 20F for
each dose)
5,10.20
200
(IF. IN)
Hammals at Low Dietary Levels
Response Reference
No effect Kennedy et al..
1973 |
Liver pathology
No effect Lehman. 1952
Liver pathology
Liver pathology Fltzhugh and
Nelson. 1951
No effects
Slight liver damage. CNS
stimulation
No effect on organ Boots Hercules
weights, histology Agrochemicals.
or clinical tests Inc.. n.d.
Moderate liver degen-
eration
03
in
-------
1ABLE V 6 (cont.)
ru
O
Duration
Species of Feeding
Dogs (6) 1360 days
(3.7 years)
up to
1260 days
Toxaphene
Concentration
(ppro diet)
200* (4)
(5 mg/kg/day)
400* (2)
(10 mg/kg/day)
Response Reference
Liver necrosis Boots Hercules
AgrochemUals.
Inc.. n.d.
One death after 33 days
Bobvhtte quail
Collnus vlrglnlanus
male
Unite Leghorn
Chicks, female
138-171 days
up to 50 weeks
10.0
50.0
0.5
5.0
50.0
100.0
138 days SOX errors In
behavioral tests
171 days-same as
controls
Occasional deformation
of the cartilaginous
region of the keel.
Increased growth of
cartilage.
Sternal deformation and
mild nephrosls at 5. 50
and 100.
Renal lesions at 50. 10Q
Decreased body weight
Kreltzer. 1980
Bush et al.,
1977
S 'Administered In capsules containing toxaphene In corn oil
\
£ H -- male; F = female
\
03
-------
Kuz'mlnskaya and Ivan1tsk11 (1979, 1981) showed that a single dose to
rats, one-half of the LD5Q of toxaphene (120 mg/kg bw), decreased the
amount of adrenaline 1n adrenal glands and Increased adrenaline and nor-
adrenallne 1n the urine during the first 24 hours after administration of
the pesticide. A chronic study was Initiated using albino male rats
(200-250 g) that were given dally doses of toxaphene (2.4 mg/kg bw) up to 6
months. The number of animals per group was not reported. Chronic adminis-
tration altered catecholamlne metabolism and breakdown. There were disturb-
ances 1n catecholamlne metabolism In toxaphene treated animals 1n the brain
and the heart that may account for the clinically observed damage to the CNS
and the myocardium.
The NCI (1979) conducted a chronic study with Osborne-Mendel rats and
B6C3F1 mice to determine the possible carclnogenldty of toxaphene. Toxa-
phene was added to feed as described,. 1n the subchronlc effects section.
Fifty animals of each gender constituted a treatment group of rats or mice.
Ten untreated animals of each gender were matched controls and data from 45
*
or 40 untreated animals from similar bloassays were pooled for statistical
evaluations.
Groups of 50 mice/sex were administered toxaphene for 80 weeks and
observed until sacrifice at 90-91 weeks (NCI, 1979) (Table V-7). Low-dose
males and females received 160 ppm In the feed for 19 weeks followed by 80
ppm for 61 weeks; high-dose males and females, 320 ppm for 19 weeks, follow-
ed by 160 ppm for 61 weeks. TWA doses were 99 or 198 ppm 1n the diet for
both males and- females. Mean body weights attained by high-dose male mice
were lower than those of matched controls, but weights of other exposure
02030 V-21 02/26/87
-------
TABLE V-7
National -Cancer Institute Chronic Feeding Study3
Duration Toxaphene Time-Weighted
Species of Feeding Concentration Average Doseb
(weeks) (ppm diet) (ppm diet)
Response
High-dose
Females
Low-dose
High-dose
19
61
19
61
19
61
19
61
Rats,
Osborne-Hendel
High-dose
2
53
25
2
53
25
160
80
320
160
99
198
160
80
320
160
99
198
1280
640
320
2560
1280
640
556
1112
Mean body weights
unaffected
Mean body weights
adversely affected;
several animals died
before week 19; dose-
related decrease In
survival
Mean body weights
unaffected
Mean body weights
unaffected; several
animals died before
week 19; dose-related
decrease 1n survival
Mean body weights
unaffected
HyperactlvHy (week 2);
generalized body
tremors (week 53);
mean body weights
unaffected; no dose-
related decrease 1n
survival
02030
V-22
02/26/87
-------
TABLE V-7 (cont.)
Duration Toxaphene Time-Weighted
Species of Feeding Concentration Average Dose''
(weeks) (ppm diet) (ppm diet)
Response
Females
Low-dose
High-dose
55
25
55
25
640
320
1280
640
540
1080
Mean body weights
adversely affected
Mean body weights
adversely affected;
generalized body
tremors (week 53); no
dose-related decrease
In survival
aSource: NCI, 1979
bT1me-weighted average dose
Ifdose In ppm x no. of weeks at that dose)
z(no. of weeks receiving each dose)
02030
V-23
02/26/87
-------
groups were essentially unaffected by toxaphene. Several animals died In
the high-dose groups before week 19 when doses were lowered. Following
this, the exposed mice were generally comparable with controls 1n appearance
and behavior during the first year of the study. During the second year,
abdominal dlstentlon was observed 1n all dosed groups but predominantly In
the high-dose males. Other clinical signs Included alopecia, diarrhea,
rough hair coats and dyspnea. From weeks 60-76 the low-dose males appeared
hyperexcltable. After week 75 there were dose-related differences 1n sur-
vival. Because of the varied dose regimens, valid NOAEL data cannot be cal-
culated.
Groups of 50 rats of each gender were administered toxaphene for 80
weeks (NCI, 1979) and then observed until survivors were sacrificed at
108-110 weeks (see Table V-7). Low-dose males were given 1280 ppm 1n the
feed for 2 weeks, 640 ppm for 53 weeks and 320 ppm for 25 weeks, which was
reported as a TWA of 556 ppm. High-dose males were administered 2560 ppm In
the feed for 2 weeks, 1280 ppm for 53 weeks and 640 ppm for 25 weeks, or a
TWA, of 1112 ppm. Low-dose females received 640 ppm 1n the feed for 55 weeks
and 320 ppm for 25 weeks, for a TWA of 540 ppm. High-dose females received
1280 ppm 1n the feed for 55 weeks, and 640 ppm for 25 weeks, for a TWA of
1080 ppm.
Mean body weights of the low- and high-dose female rats were lower than
those of the matched controls throughout most of the bloassay study period,
whereas weights of low- and high-dose males were essentially unaffected.
During the first 16 weeks, dosed males were generally comparable with con-
trols 1n appearance and behavior, with the exception of high-dose males
02030 V-24 02/26/87
-------
that appeared hyperactive during week 2 when the doses for male rats were
reduced. At week 53, tn-e concentration of toxaphene In the feed was reduced
because a majority of the high-dose males and females developed generalized
body tremors. Dose-related decreases 1n survival rates were not observed.
Clinical signs usually associated with aging were observed earlier In dosed
rats than 1n controls and were as follows: alopecia, diarrhea, dyspnea,
pale mucous membranes, rough hair coats, dermatitis, ataxla, leg paralysis,
eplstaxls, hematurla, abdominal dlstentlon and vaginal bleeding. Two
females, one high-dose and one low-dose, had Impaired equilibrium. Again,
no valid NOAEL data 1s available for this study.
Bush et al. (1977) added graded levels of toxaphene In corn oil to the
diets of female White Leghorn chicks from 1 day of age In order to achieve
doses of 0, 0.5, 5, 50 and 100 ppm diet. Each treatment consisted of 90
randomly selected birds'(30 birds 1n each of three replicates). Levels up
to 100 ppm diet did not produce symptoms of toxldty throughout the 50-week
study. Mortality was <5X In all groups. Body weights at 6 and 30 weeks
were significantly decreased when the birds were fed 100 ppm. There were no
significant treatment-related changes 1n heart, liver, gizzard or kidney
weights at 4 or 8 weeks. Necropsy of 31-week-old birds fed 5, 50 and 100
ppm toxaphene 1n the diet revealed sternal deformation resembling osteoma-
lada. Occasional keel deformation Involving the cartilaginous tissue as
well as an apparent Increase 1n growth of cartilage was found In birds fed
0.5 ppm toxaphene In the diet. Further studies were planned to determine
whether the backbone of the chicken as well as other skeletal structures
were affected by toxaphene. Hlstopathologlcal examination of organs of
31-week-old birds showed mild nephrosls of the kidney In birds fed toxaphene
02030 V-25 02/26/87
-------
at 5, 50 and 100 ppm. Other observations Included cysts, occasional cel-
lular casts In the renrt tubules and accumulation of a brown granular pig-
ment 1n the cytoplasm of some tubular epithelial cells.
Kreltzer (1980) determined whether behavioral effects 1n birds would be
produced by toxaphene at levels below those that produced overt signs of
Intoxication. Adult male bobwhlte quail Collnus v1rq1n1anus were fed toxa-
phene dissolved In propylene glycol and blended Into the feed at levels of
10 and 50 ppm. An equal amount of propylene glycol was added to the feed of
controls. There were four controls and four birds per treatment level For
each of two tests (with different pairs of patterns) that measured perform-
ance on nonspatlal discrimination reversal tasks. Birds were Initially
exposed at 3 days of age for 138 and 171 days before tests 1 and 2, respec-
tively. The treated birds 1n test 1 had SOX more errors than the controls
(p<0.02). There were no significant differences In the performances of
birds fed the two toxaphene levels. In test 2, after 171 days of exposure,
treated birds performed as well as the controls, Indicating an accomodatlon
to the chemical.
Enzyme Effects
Toxaphene appears to Induce the mlcrosomal MFO system as evidence by the
shortening of pentobarbHal sleeping time at 50 mg toxaphene/kg bw (Schwabe
and HendUng, 1967). A dose of 100 mg toxaphene/kg bw has been shown to
decrease the barbiturate sleeping time by -20% for up to 20 days after
exposure (Ghazal, 1965). Toxaphene will also Initiate MFO activity 1n whole
liver homogenates from rats pretreated with toxaphene (Klnoshlta et al.,
1966). Effects 1n male rats were more pronounced than females. Toxaphene
02030 V-26 02/26/87
-------
also appears to stimulate liver mlcrosomal metabolism of estrone and testos-
terone. For example, an 1.p. dose of 120 mg/kg bw to male rats caused a
484% Increase 1n testosterone metabolism by 5 days after dosing and a 19%
decrease 1n plasma testosterone, the latter level returning to control
levels by the 15th day after dosing 1n spite of elevated mlcrosomal enzyme
activity by 532% (Peakall, 1976). For female rats pretreated at 25 mg/kg
bw/day for 7 days, estrone metabolism Increased by 155% (Welch et al.,
1971). Since toxaphene 1s also known to bind to mammalian cytochrome P -450
to give a typical type I difference spectrum (Kulkarnl et al., 1975), these
effects may be attributed to Induction of hepatic cytochrome P-450.
The toxlclty of Toxicant B to mice Is Increased by a factor of 2-8 by
administration of plperonyl butoxlde that Inhibits the cytochrome system
(Saleh et al., 1977; Turner et al., 1977), which Indicated to the authors
that cytochrome P-450-med1ated detoxification mechanisms were Important to
explain the mode of action of toxaphene toxldty (see Chapter VII). The rat
liver mlcrosomal system also mediates reductive dechloMnatlon of Toxicant A
and B (Khalifa et al., 1976; Saleh and Caslda, 1978). NADPH and relatively
anaerobic conditions were required. However, significant nonenzymlc reac-
tion 1n these systems may also occur as demonstrated by the ability of
reduced hematln to cause reductive dechlorlnatlon (Saleh and Caslda, 1978,
1979). The metabolites of these nonenzymlc systems have already been Iden-
tified In Chapter III, and reductive dechlorlnatlon and dehydrochlorlnatlon
occur. Germinal and vicinal halogenated aliphatic organlcs are particularly
susceptible.
02030 V-27 02/26/87
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Trottman and Desalah (1980) fed male rats (Sprague-Dawley 175 g;
5-6/group) diets containing 0, 50, 100, 150 and 200 ppm toxaphene In the
diet for 14 days causing Increased liver weights at the 200 ppm dose and
decreased thymus weight at the 150 and 200 ppm dose. In addition, enhance-
ment occurred for liver hydroxylatlons of pentobarbltal 1n a dose dependent
manner, and for aniline and ethylmorph1ne-N-demethylase activity In a non-
dose-dependent manner 1n the postmltochondrlal supernatant fraction.
Decreased sleeping time following pentobarbltal administration also
occurred, but was not dependent on dose. Hepatic mlcrosomal protein content
was Increased 20-35% 1n comparison with control values. In the liver mlcro-
somal fraction, the activity of NAOPH-cytochrome c-reductase was Increased
20-50% at doses >150 ppm. Cytochrome P-450 levels were Increased 15-27% at
doses >50 ppm. There were no effects on the activity of NADPH-dehydrogenase
or of the levels of cytochrome b.. The binding of both aniline and'hexo-
barbltal to cytochrome P-450 was Increased by 55-210% at all doses. All of
these results show that specific functions of the MFO system of the liver,
I.e., cytochrome P-450 and NAOPH cytochrome c-reductase (the sites that are
sensitive to aniline and hexobarbltal Interactions), are affected by toxa-
phene.
A toxaphene formulation was subfractlonated on a silica gel column Into
64 fractions of varying polarity (Pollock et al., 1983). The most highly
toxic components to Sprague-Oawley rats were found 1n the nonpolar frac-
tions. Young Sprague-Dawley rats (24 days; 70+6 g) were administered two
fractions, one constituting 73% (the nonpolar fraction) and the other 27%
(the polar fraction) at 25 mg/kg bw as well as unfractlonated toxaphene at
doses of 0, 5, 25 and 100 mg/kg bw by 1.p. Injection using olive oil
02030 V-28 02/26/87
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carrier. Liver weights Increased after administration of toxaphene doses of
25 mg/kg bw or more 1n_a dose dependent manner, as also did the cytochrome
P-450 level, amlnopyrlne demethylase activity and aldrln epoxldatlon.
Mlcrosomal protein was Increased only at the 100 mg/kg dose. The protein
content In the postmltpchondMal supernatant was not affected by toxaphene.
Both polar and nonpolar fractions caused the same results as toxaphene
alone. The "no significant Inductive effect" level for 1.p. administered
toxaphene Is between the 5 and 25 mg/kg dosages.
Other enzyme systems are affected by toxaphene. One major system
Involves the metabolism of sugar to produce high energy compounds. At 330
vM, toxaphene Inhibited the in vitro activity of mltochondrlal sucdnoxl-
dase In beef heart by 76% and the NADH-ox1dase system by 96% (Pardlnl et
al., 1971). At 100 yM, the active transport of glucose through Isolated
mice Intestine was Inhibited (Guthrle et al., 1974). Doses of 1.2 mg toxa-
phene/kg bw/day for 6 months or an acute 120 mg toxaphene/kg bw dose to rats
altered lactate dehydrogenase (LOH) spectra 1n the liver and blood
(Kuz'mlnskaya and Alekhlna, 1976). In the liver, the acute dose caused a
45% Inhibition of total LDH activity for 15 days postexposure. Increases In
the Isozymes, LOH-1, LDH-2 and LOH-3 were observed, LDH-4 only transiently
decreasing and LOH-5 not at all. In the blood, the acute dose affected only
LDH-1. Chronic exposures decreased total LOH activity 1n the blood (30%)
and liver (45%). In the liver, LOH-3 Increased after 3 months and LOH-5
decreased after 6 months. In the blood, LDH-1 and LDH-2 activities were
depressed and LDH-5 activity elevated after 6 months. Depressed LDH activi-
ties, were noted 1n the liver (17%), kidneys (18%) and serum (20%) of rats
given 35 mg toxaphene/kg bw/day for 6 months (Gertlg and Nowaczyk, 1975).
02030 V-29 02/14/85
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In the kidney, there was a 17X decrease 1n LDH-1. Decreases of 50% In both
serum alkaline phosphata_se and liver glutamate dehydrogenase also occurred
after the 6-month exposure.
Peakall (1979), who gave male rats an acute 120 mg toxaphene/kg bw dose
by capsule, showed that levels of pyruvate and lactic add In blood plasma
at 1, 5 and 15 days were not affected. This was so even when rats given 1.2
mg toxaphene/kg bw/day were observed at 1, 3 and 6 months. A steady-state
level of toxaphene residues 1n the liver occurred after 1 month of chronic
dosage; In the brain, this took 1-3 months. The author Implied that the
rats had to be stressed 1n some way to obtain the results of Kuz'mlnskaya
and Alekhlna (1976) and Gertlg and Nowaczyk (1975). Peakall himself, how-
ever, gave no data on strain of rat, weight, gender or age.
Technical grade toxaphene was used 1n two studies by Srebocan et al.
(1978, 1980a) on the enzymatic regulation of carbohydrate metabolism In
poultry. In the 1978 study, groups of 1-day-old N1ck chick cockerels (10
birds) were fed for 2 weeks on a formulated diet containing 0, 0.1, 0.5, 1,
5, 10, 50 or 100 ppm toxaphene (using sunflower oil to dissolve the pesti-
cide and to mix H with the diet). Inhibition of enzymes of the gluconeo-
genlc pathway 1n the liver was observed at 1 ppm. Enzymes Involved 1n pyru-
vate metabolism were Inhibited between 20 and 50% (pyruvate carboxylase, EC
6.4.1.1J and phosphoenolpyruvate carboxyklnase, EC 4.1.1.32). There was
not, however, a dose response. Fructose-l,6-d1phosphatase (EC 3.1.3.11) and
glucose-6-phosphatase (EC 3.1.3.9) activities, and glucose levels were not
significantly affected at p<0.05. Pyruvate levels were Increased by toxa-
phene 1ngest1on. Based on studies with DDT, the authors suggested that
02030 V-30 02/14/85
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the effects of a number of chlorinated pesticides were adrenocortkal ly
mediated. In the Srebocan et al. (1980a) study, the same experimental
protocol was utilized except that toxaphene was given at 5 ppm 1n the diet
separately or 1n combination with heat or starvation stress together with
the appropriate controls. While toxaphene alone caused an accumulation of
pyruvate (40%), heat stress alone also caused an accumulation of pyruvate
(60%) and Increased the activities of fructose-1,6-d1phosphatase (15%) and
glucose-6-phosphatase (17%); the combined heat and toxaphene treatment
caused accumulation of pyruvate (17%) and Increased the activity of glucose-
6-phosphatase (17%). The activities of pyruvate carboxylase, phosphpenol-
pyruvate carboxyklnase, and the level of glucose were not significantly
affected by any of the treatments. For starvation stress/toxaphene experi-
ments, all the toxaphene treatments (5 and 10 ppm diet) lowered the activi-
ties of all the above enzymes and Increased the accumulation of pyruvate
(70-80% excess). However control results were not consistent with those for
the 5 ppm dose already quoted above. Starvation stress caused accumulations
of pyruvate (20%) and enhanced fructose-1,6-dlphosphatase activity (100%).
The combination experiments led to Inhibition of glucose-6-phosphatase
(35-40%) and glucose depletion (35-40%) for the 5 and 10 ppm doses, and
Inhibition of pyruvate carboxylase (40%) and phosphoenolpyruvate carboxy-
klnase (60%). Fructose-1,6-dlphosphatase levels 1n the toxaphene and toxa-
phene/starvatlon experiments were not different from controls, whereas a
large elevation occurred from starvation stress alone. These results Imply
that stringent quality assurance of the environment and diets must be main-
tained during toxlcologlcal studies.
02030 V-31 02/14/85
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In an abstract, Desalah et al. (1979) reported that male Sprague-Oawley
rats fed 0, 25, 50 and" 75 ppm toxaphene In the diet for 8 weeks showed
marked dose-dependent Increases 1n liver homogenate glucose-6-phosphatase
and fructose-l,6-d1phosphatase In the liver mlcrosomal fraction, both being
maximally stimulated by 70%. It was concluded that toxaphene affected the
process of gluconeogenesls.
Alekhlna and Kuz'mlnskaya (1980) determined LDH activity and changes In
LDH Isoenzymes In the liver and myocardium of "white" male rats (200-250 g),
dosed perorally with 2.7 mg toxaphene/kg bw/day, percutaneously at 9.4 mg
toxaphene/kg bw, and a combination group, all treatments being over 4
months. LDH activity In myocardium after 4 months Increased 11-34% regard-
less of the mode of application, though the combination group gave the high-
est Increase compared with the single treatments (16-18%). LOH activity
Increased (32%) 1n the liver only for the combination treatment. When pro-
vided orally, toxaphene caused Increases In LDH-1 (65%) and a decrease In
LOH-5 (27%); 1n the myocardium, decreases occurred In LOH-3 (35%) and In
LDH-4 (70%). When provided percutaneously, Increases 1n liver LOH-3 (64%)
and LDH-4 (81%) were found as well as In myocardium (18 and 62%, respective-
ly). LDH-5 was decreased (28%) only 1n the liver. For the combined treat-
ment, the LOH-5 content 1n the liver remained decreased (25%), but LOH-3 and
LDH-4 were Increased by 24 and 140%, respectively. Although these changes
In myocardium were postulated by the authors to be due to toxaphene residue
deposition, this was not demonstrated directly.
02030 V-32 ' 02/14/85
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Male albino Sprague-Dawley rats (300-350 g, 8/group) treated l.p. with a
single 40 mg toxaphene/kg bw/day 1n vegetable oil carrier (0.5 ml) showed
decreased (19%) plasma cholesterol only 60 days posttreatment, with no
effect on plasma trlglycerldes, normalized liver weights, mlcrosomal pro-
tein, or cytochrome P-450 (Ishlkawa et al., 1978).
In another series of experiments, Kuz'mlnskaya et al. (1980) determined
the oral and dermal toxlclty of toxaphene 1n male rats separately and then
Investigated concomitant application of oral and dermal doses. Endpolnts of
toxlclty Included the activity of marker enzymes of mitochondria, mlcrosomes
and lysosomes 1n fractions of liver homogenates. Toxaphene applied In vivo
Increased 1n 24 hours the activity of the liver mltochondrlal enzymes suc-
clnlc dehydrogenase 175% and cytochrome oxldase 1000%. Mlcrosomal glucose-
6-phosphatase activity was Increased 41% and lysosomal add phosphatase
activity was Increased 46%. The dose (or doses) that produced these
responses was not clearly Identified nor was the vehicle reported. The
doses (mg/kg bw) toxaphene used In this study were as follows: oral, 67.5,
38.5 and 6.75; dermal, 235, 134 and 23.5. The greatest Increase 1n enzyme
activity was seen 24 hours after application.
Hehendale (1978) reported a decrease of biliary flow and decreased
l4C-1m1pram1ne excretion from the liver of male Sprague-Dawley rats dosed
at 100 ppm toxaphene In the diet for 8 days. Trottman and Desalah (1979)
studied ATPase activities 1n the brain, kidney and liver of male ICR mice
(30 g each) in vivo and \n vitro. For the in vivo studies, toxaphene was
provided at 0, 10, 25 and 50 mg/kg bw/day over 3 days by corn oil (0.3 ml)
for 4-6 mice 1n each group. Enzyme activities were measured 36 hours after
02030 V-33 02/14/85
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the last dosage. For Jm vitro studies, toxaphene at 7.5, 15.0 and 30 WM
Inhibited brain (52-56%) and kidney (42-46%) Naf/Kf-ATPase activities,
and the ol1gomydn-sens1t1ve Mg2f-act1v1ty In brain (53-61%), kidney
(37-65%) and liver (20-62%). The Inhibition of Ol1gomyc1n-sens1t1ve Mg2t-
ATPase activity was dose-dependent 1n the kidney and liver preparations.
The ol1gomydn-1nsens1t1ve Mg f-ATPase activity was also Inhibited In
brain (29-45%), kidney (47-50%) and liver (24-51%), the Inhibition being
almost dose-dependent for brain and liver. For In vivo studies, Na*/Kf-
and ol1gomyc1n-sens1t1ve and Insensitive Mg *-ATPase activities In toxa-
phene-treated mouse brain were not altered significantly. Kidney ATPases
were significantly decreased (down to 42% for Na*"/Kf-ATPase activity;
35% for ol1gomydn-1nsens1t1ve Mg *-ATPase) In a dose-dependent manner
except for ol1gomyc1n-sens1t1ve Mg -ATPase. A dose-dependent decrease In
the ollgomycln-sensUlve Mg *-ATPase activity (down to 35%) was found In
the liver, but, 1n contrast with the kidney, kall doses Increased the
ol1gomyc1n-1nsens1t1ve component (over 60%). The differences between the In
vivo and ^n vitro results for the brain may arise from the fact that toxa-
phene may be metabolized extensively 1n in vivo studies before 1t reaches
the liver (perhaps by reduced hematln or another nonenzymatlc process),
kidney or brain. Rapid metabolism may also account for the effects on
ATPases 1n kidney and liver. The crucial presence or absence of toxaphene
after addition In the \t± vitro or ^n vivo studies was not directly measured.
The Naf/K*-ATPase Is postulated to be Involved In active 1on transport;
the ol1gomydn-sens1t1ve Mg *-ATPase In mitochondria Is believed to be
Involved 1n oxldatlve phosphorylatlon. The Interference of toxaphene with
the production of ATP may cause an Impairment of transport processes In the
liver and kidney, and may be related to Impairment of biliary excretion.
02030 V-34 02/14/85
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In another study (Trottman and Oesalah, 1983), adult male Sprague-Dawley
rats (175-200 g; 6/grouji) were fed 0, 50, 100, 150 and 200 ppm toxaphene In
powdered chow for 8 weeks. Toxaphene Inhibited NaVKf- and Mg2f-
ATPases of synaptosomes in vitro In a dose-dependent manner. The IC.Q
values were about 30 yM toxaphene for Naf/Kf-ATPase and 15 yM toxa-
M
phene for the Mg *-ATPase. Fifty percent Inhibition of ouabaln and dopa-
mlne binding occurred around 150 and 200 yM toxaphene, respectively. For
^n vivo studies, no dose responses were observed for decrease In enzyme
activities (30-40X decreases for doses >100 ppm) even though all ATPase
activities were decreased. In fact, there were no significant changes for
ID. v^vo studies Involving ouabaln and dopamlne binding to synaptosomes.
Toxaphene was postulated by the authors to have metabolized rapidly J_n vivo
and not reached the brain to explain the difference In the in v 1 vo and in
vitro results. Naf/K*-ATPase has been shown to be a receptor of ouabaln
and perhaps dopamlne. Toxaphene Itself Interferes in vitro but not in vivo,
and caution therefore must be exercised 1n the use of in vitro tests 1n
place of in vivo tests. Fattah and Crowder (1980) also tested the effects
of toxaphene in vivo and in vitro on the plasma membrane ATPase of kidney,
brain and liver from mice. Male Swiss Webster mice (35 per treatment and
control groups) were administered by oral gavage an LD5Q (112 mg/kg bw)
dose of toxaphene 1n corn oil, or corn oil alone. Kidney ATPases were more
sensitive to toxaphene than the liver and brain enzymes. The Na*/K*-
ATPase was Inhibited significantly only 1n the kidney, and the Mg f-ATPase
was Inhibited significantly 1n all three tissues. Twenty mice were used to
study the effects of hi vitro toxaphene exposure of tissues. Each tissue
was tested with four concentrations of toxaphene, 10~«, 10~5, 10"6 and
02030 V-35 02/14/85
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TO"7 M, and two controls. Three samples for each treatment were run In
each of the three tissues. As observed following in vivo exposure, toxa-
phene significantly Inhibited the Naf/K*-ATPase activity only In the
kidney homogenates, In this case with a dose effect. Mg2*-ATPase was
significantly reduced 1n all tissues; In the Hver homogenates only 10"4 M
toxaphene significantly Inhibited activity.
Parvu et al. (1980) reported In an abstract that toxaphene given orally
to white Ulstar rats at 4 mg/kg bw/day for 30 days caused I1p1d accumulation
In the liver, depletion of free fatty adds, and an Increase of "liver
cholesterol.
Toxaphene may also affect VHamln A storage. A concentration of 100 ppm
toxaphene In the diet for 72 days to 28-day female weanling rats (Phillips
and Hatlna, 1972) was followed by mating with untreated males. At 20 days
postpartum, liver Vitamin A levels relative to controls were depressed by
11% (p<0.01) In newborn rats, but not 1n dams.
Alterations In clinical chemistry have also been seen In subchronlc oral
toxaphene exposures. Mice with no clinical signs of Intoxication evidenced
consistent Increases 1n serum add phosphatase, glutamlc pyruvlc trans-
amlnase and gamma-glutamyl transpeptldase activities, along with Increased
neutrophll counts and changes 1n urine protein (Blumler, 1975). At a much
lower dally dose, rats had only a transient Increase 1n serum alkaline phos-
phatase during the fifth month of 1ngest1on and showed no variation In urine
hlppurlc add (Grebenyuk, 1970). Increases 1n all of the above enzyme
activities are consistent with the mild liver pathology associated with
subchronlc toxaphene exposure.
02030 V-36 02/26/87
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Teratoqenldtv and Reproductive Effects
Hamma1s. The multtgeneratlon reproductive effects of toxaphene and
two other pesticides were Investigated by Kennedy et al. (1973). two con-
trol groups and two test groups of Sprague-Dawley albino rats consisted of 8
male and 16 female animals each. The treated groups started receiving 25 or
100 ppm toxaphene 1n the diet (adjusted to contain 2X corn oil) added to
commercial chow at 28 days of age. FQ parental animals were exposed to
toxaphene for 42 weeks and then sacrificed. F, and F. parents were
exposed for 39 weeks and then sacrificed. Animals were first mated at 100
days of age. First litters were reduced to 10/Htter on postpartum day 5
and retained for 21 days until weaning. After a 10-day rest, the parental
animals were mated again to obtain second Utters. At weaning of the second
Utters, 8 males and 16 females were randomly selected from each group as
parental animals for the next generation. This procedure was continued
through three successive 2-l1tter generations. Toxaphene had no effect on
Utter size, pup survival or weanling body weights. Parental animals at 100
ppm showed liver changes consisting of slight cytoplasmlc fatty vacuollza-
tlon In 63% of animals examined. This change was not observed at 25 ppm and
was not accompanied by adverse effects on growth, mortality, organ weights
(liver, kidney, spleen, gonads, heart, brain, adrenal glands, thyroid gland)
or among the corresponding organ-to-body weight ratios or reproductive
capacity. No differences were observed among test and control groups of
F- parental animals with regard to total, free and esterlfled cholesterol
concentrations, hematologlc parameters, urine analysis and other clinical
chemical findings. Tumor Incidence was not affected and there was no
evidence of teratogenldty among test progeny. The no-effect level was
judged to be 25 ppm 1n the diet for toxaphene (1.25 mg/kg by assuming food
Intake equal to 5X of body weight).
02030 V-37 02/14/85
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Chernoff and Carver (1976) used rather high doses of toxaphene to study
fetal tox1c1ty of the pesticide 1n rats. CO rats were administered toxa-
phene 1n corn oil (0.1 ml) by gastric Intubation on days 7 through 16 of
gestation at doses of 15, 25 or 35 mg/kg bw. The number of rats per group
were as follows: controls, 33 rats; 15 and 25 mg/kg, 39 rats/group; 35
mg/kg, 16 rats. At 35 mg toxaphene/kg, toxldty was evidenced by 31X
maternal mortality. There was also a dose-related reduction In weight gain
of dams (p<0.001) at 15 and 25 mg/kg/day and fetal weight (p<0.001) at 25 mg
toxaphene/kg/day. Even though there was significant maternal toxldty In
all treated groups, there were no dose related changes 1n fetal mortality or
1n the occurrence of anomalies.
The rat fetus was shown to contain a small amount of 14C-toxaphene
when assayed 5 days after an oral dose of 2.6 or 6.5 mg/kg maternal body
weight onv day 15 of gestation (Pollock and HUlstrand, 1982). Fetal tissues
contained 28 yg/kg wet weight; In comparison, maternal fat contained 7.48
mg/kg tissue.
Mixed-strain white rats and golden hamsters were used by Martson and
Shepel'skaya (1980b) to evaluate the embryotox1c1ty of toxaphene. Rats were
given 4 mg toxaphene/kg bw/day by gastric Intubation during the period of
organogenesls (days 6-15) and during the entire pregnancy (1-31 days) and to
hamsters on days 7-11 and 1-15 of gestation. In rats, no adverse effects
were noted on development, fetal weight, ratio of males to females, or the
number of fetal deaths. In hamsters, the only toxaphene-related effect was
reported as an Increase In an unidentified developmental anomaly.
02030 V-38 02/14/85
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Badaeva (1979, 1981} examined the effects of toxaphene on chollnesterase
activity 1n cardiac neural structures of dams and future offspring on days
14 and 20 of gestation. A group of 12 pregnant animals was perorally admin-
istered dally doses of 12 mg toxaphene/kg bw. It was found that the number
of dead embryos was higher when toxaphene was administered throughout gesta-
tion than when administered between the 6th and 14th day of gestation. When
given throughout gestation, the amount of toxaphene found In the maternal
heart was higher on the 20th day of gestation (14 yg/g), uterus (9.7
yg/g), brain and spinal cord (4.3 yg/g) than on the 14th day. The
number of pups per Utter (6-8) was less 1n treated groups than 1n control
groups (10-11); birth weight of pups was depressed and Increased mortality
was noted 1n the first week postpartum. In treated embryos, chollnesterase
activity along the myocardlal vessels and cells of the atrloventrlcular node
was almost absent at 14 days of gestation; at 20 days, there was a delay In
differentiation of fetal cardiac neural elements and a continued depression
of chollnesterase activity. On the 20th day, 50-60% of the cells of the
cardiac ganglion of pregnant treated animals exhibited decreased cytoplasmlc
chollnesterase activity.
Toxaphene was one of five compounds administered to pregnant 90-day-old
CD rats during the period of organogenesls (7-16 days of gestation), and the
effects on organ differentiation were determined In day 21 fetuses (Kavlock
et al., 1982). Two dose levels of toxaphene 1n corn oil, 12.5 and 25 mg/kg
bw/day, administered perorally to a group of five animals, had no effect on
the average number of Implants, fetal mortality or fetal weight. No soft
tissue abnormalities were, observed. There was no statistical difference
between the following parameters 1n treated and untreated rats: brain
02030 V-39 02/14/85
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weight, total DNA and total protein; lung weight and primary surfactant
materials; liver weight "and total glycogen; and kidney weight. There was a
statistically significant decrease 1n alkaline phosphatase activity of the
kidney at 25 mg toxaphene/kg bw and 1n total protein 1n the kidney at both
doses.
Behavioral effects of low levels of toxaphene, and Us toxic components,
Toxicants A and B, were studied 1n juvenile albino rats exposed perlnatally
(Olson et al., 1980). Toxaphene, or Toxicant A or B, was given dally 1n the
feed, starting on day 5 of gestation. Pups were exposed to dietary levels
after weaning until the study was terminated at 3 months postpartum: toxa-
phene 50 yg/kg bw, Toxicants A and B, 2.0 yg/kg bw. Because sufficient
supplies of Toxicant A could not be procured. Toxicant B was substituted for
Toxicant A when supplies were depleted (rats were 40 days old). Offspring
from 12 mothers made up three treatment groups of 16 each and one control
group of 15. Behavioral testing began when the offspring were 7 days old.
Two different behavioral test periods were studied: early development with
pups being tested on postnatal days 7 through 17 (swimming and righting
reflex); motivational, learning and retention tests on days 70 through 90
(symmetrical maze). In their early development, treated animals showed
retarded maturation based on their performance In overall swimming ability
1n comparison with control animal performance (p<0.0001 on day 10). By day
16 all groups displayed normal swimming ability. Inferior swimming ability
may reflect an ability of toxaphene compounds to affect early functional and
behavioral development of the CNS In the Immature rat. Toxaphene-fed
animals constituted the only group to exhibit overall retarded righting
ability 1n tests conducted from days 7 through 17. On day 15 the order of
02030 V-40 02/14/85
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decreasing superiority was as follows: controls, Toxicant A, Toxicant 8 and
toxaphene (p<0.0001) with each group being significantly superior to the one
following. Inability of the Immature rat to metabolize the simple mixture
may account for the Increased toxlclty of toxaphene. The test of motivation
revealed no significant differences between any of the groups. In the maze
retention test. Toxicant A animals had no difficulty 1n learning the test
problems but. were Inferior to the other groups In retaining that knowledge.
Behavioral changes were observed at dosages far lower than those used In any
previously reported study.
In another behavioral study, Crowder et al. (1980) also reported a sig-
nificant difference In the length of time for treated animals to become
positive In the righting reflex. In the Crowder et al. (1980) study, three
pregnant rats received dally oral doses of 6 mg/kg bw 1n 0.1 mi corn oil
from day 7 of gestation until parturition. Testing methods Included a
simple two-choice maze, motor skills, an open field test and other behavior-
al tests.
Investigators In Russia found no effect on behavioral responses of
1-month-old pups exposed during days 6-15 of gestation (Hartson and
Shepel 'skaya, 1980a). The dose was reported as the 0.1 L05Q given by
gastric Intubation to six pregnant animals; however, the LD50 for the
mixed strain rats was not given. In this same study, 6 male and 6 female
rats received 4 mg toxaphene/kg bw by gastric Intubation dally for 10 weeks
before mating. Hales and females receiving toxaphene were bred with
untreated rats. At the age of 2 months, offspring were monitored for
behavioral response. There were no toxic effects observed In the offspring
02030 V-41 02/14/85
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as judged by locomotion, grooming and rearing activities. The authors con-
cluded that toxaphene did not have major gonadotoxlc effects. There was,
however, an Increase In the number of sperm with abnormal morphology
(Martson and ShepeVskaya, 1980b).
OlPasquale (1977) examined the effects of toxaphene on fetal guinea pig
development. In this study, toxaphene was administered orally to pregnant
females at a dose of 15 mg/kg bw from day 21 to day 35 of gestation. No
effects were noted on the anatomical development of the fetus. There was
some fetotoxldty as shown by adverse effects on collagen-related structures
of the fetus. This was attributed to a functional deficiency of vitamin C
related to MFO Induction. Maternal guinea pigs showed a slight loss of body
weight, but no effects attributable to toxaphene exposure were seen on
maternal liver weight or mortality.
CD-I albino mice were administered toxaphene In corn oil by gastric
Intubation during the period of organogenesls, days 7 through 16 of gesta-
tion (Chernoff and Carver, 1976). The number of mice per group and dally
doses of toxaphene were as follows: 75 mice, 0 toxaphene; 26 mice, 15 mg/kg
bw; 45 mice, 25 mg/kg bw; 90 mice, 35 mg/kg bw. Mice were killed on day 18
of gestation and fetuses weighed and prepared for soft tissue or skeletal
analysis. In the 35 mg/kg group there was 8% maternal mortality. There was
a dose-related (p<0.01) reduction 1n the average maternal weight gain and a
dose-related Increase 1n the I1ver-to-body weight ratio. There were no
significant dose-related responses In fetal mortality, fetal weight, number
of caudal or sternal ossification centers, or Incidence of supernumerary
02030 V-42 02/14/85
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ribs. Five Utters from dams receiving the high toxaphene close had one or
more fetuses with encephaloceles; none were observed 1n controls or other
treated groups.
KepHnger et al. (1970) conducted a 5-6 generation study of five pesti-
cides Including toxaphene and combinations of pesticides using Swiss white
mice. Offspring were exposed in utero. by the mother's milk, and 1n the
diet after pups were weaned. Toxaphene (25 ppm 1n the diet) was fed to mice
during the entire study Including mating, gestation and lactation. Animals
not used for subsequent breeding were sacrlfled for chemical analyses and
histology. Eighteen 120-day-old mice, 4 males and 14 females, made up each
group. For each successive generation, parental groups were remated to pro-
duce second Utters and the next parental groups were selected from second
Utters when 120 days of age. Little or no adverse effect was noted through
five generations of mice fed 25 ppm toxaphene In the diet Including the fol-
lowing parameters for each generation: Utter size, embryonic death, sur-
vival and body weight. There were no statistically significant differences
between several Indices calculated by the Investigators Including viability,
lactation, survival and reproduction. H1stolog1cally there was evidence of
fatty changes In the liver, especially In the central zone. There was
little or no damage to the brain.
Twenty-eight compounds of known teratogenlc potential were assayed by an
In vivo teratology screening procedure (Chernoff and Kavlock, 1982). Toxa-
phene In corn oil was administered by oral gavage (75 mg/kg bw) to 25 gravid
CD-I mice on days 8-12 gestation, the period of organogenesls. Two mice
died before term and only 11 were pregnant. Dams were allowed to give
02030 V-43 02/14/85
-------
birth, and Utter size and weight on postpartum days 1 and 3 were compared
with concurrent controls. Results Indicated significantly reduced maternal
weight change and reduced fetal weights on postpartum day 1. There was no
difference 1n the body weights of treated versus control pups on postpartum
day 3.
Allen et al. (1983) studied the effect of toxaphene on Immune responses
of offspring of mice exposed orally 3 weeks before breeding, and during.
gestation and lactation. Toxaphene In acetone was mixed Into ground rat
pellets to attain a concentration of 10, 100 or 200 ppm. Transplacental/
lactation exposure to toxaphene produced some degree of Immunosuppresslon In
the pups, even though overt toxlclty of the Insecticide was not apparent at
the lower dose. At 10 ppm there was a significant depression of macrophage
phagocytosis (p<0.05) but no effect on hypersensUlvHy responses and anti-
body tlters to bovine serum albumin. Relative degree of Immunosuppresslon
was greatest In macrophages followed by humoral Immunity; cell-mediated
Immunity was least affected.
Birds. The effects of toxaphene have been examined 1n a number of
bird species. White leghorn female chicks, 1 day of age, were fed diets
containing 0, 0.5, 5, 50 and 100 ppm toxaphene (Bush et al., 1977). Each
treatment group contained 90 randomly selected hens that were bred at 23
weeks of age. Even at 100 ppm, toxaphene did not significantly alter egg
production, natchabllHy or fertility. Arscott et al. (1976) added 0, 10
and 100 ppm toxaphene to the feed of white leghorn layers (48/group) for 24
weeks. Except for a slight decrease 1n egg production, no adverse effects
were observed on fertility, hatchabllHy and survival of progeny.
02030 V-44 02/26/87
-------
Three groups of 15 pairs of 1-year-old black ducks (Anas rubrlpes) were
fed ad libitum w1h commercial duck breeder mash coated with 0, 10 or 50 ppm
toxaphene beginning 90 days prior to laying (Mehrle et al, 1979). Toxaphene
was dissolved In propylene glycol equivalent to IX of the total diet.
Reproductive success of all birds was followed throughout the breeding
season. Toxaphene did not affect reproduction or survival. Ducklings were
fed starter mash containing the same concentration of toxaphene as the diet
the parents received. Duckling growth, as Indicated by weight, was
decreased significantly 5 and 14 days after hatching In the group fed 50
ppm, and was Increased significantly at 28 and 42 days after hatching 1n the
group fed a diet containing 10 ppm toxaphene In the diet. Duckling weight
was not altered by toxaphene In measurements made more than 42 days after
hatching. Backbone development was Impaired by toxaphene within 14 days
after hatching and collagen was decreased significantly 1n the cervical
vertebrae of ducklings fed 50 ppm. In contrast to the effects on vertebrae,
toxaphene exposure did not alter tibia development.
Forty-five pairs of 1-year-old black ducks (Anas rubrlpes) were placed
on a diet of duck breeder mash containing 0, 10 or 50 ppm toxaphene (Heinz
and Flnley, 1978). Toxaphene was dissolved 1n propylene glycol before
Incorporation Into the feed at 1 part/99 parts feed. About 3 months later,
hens began to lay eggs and the eggs were Incubated. Ducklings were brooded
by hens and ate a diet of duck starter mash containing the same concentra-
tions of toxaphene as those fed the hens. At 5 days of age, ducklings were
tested for avoidance behavior. There were no significant differences 1n
avoidance behavior between control ducklings and those treated with 10 or 50
ppm toxaphene.
02030 V-45 02/14/85
-------
In a similar study conducted over a 19-month period, which Included two
breeding seasons, three Tets of 15 pairs of black ducks (Anas rubrlpes) were
given 0, 10 or 50 ppm toxaphene In dry mash (Haseltlne et al., 1980). Toxa-
phene was dissolved In propylene glycol before Incorporation Into the feed
at IX of feed by weight. Adult survival was not affected, but the weights
of treated males were depressed during the summer months. At the end of the
second breeding season females Ingesting 10 or 50 ppm had larger livers than
controls. Drakes fed 50 ppm In the diet exhibited elevated brain weights
that were not apparent 1n 50 ppm females. In both years of the study,
breeding pairs fed 50 ppm toxaphene In the diet appeared hyperactive but
caging facilities did not allow for testing of this possibility. Eqg
production, fertility, hatchabllHy, eggshell thickness, growth and survival
of young did not vary significantly with toxaphene 1ngest1on In either
breeding season. The mean number of days required to complete a clutch was
lower 1n birds fed toxaphene than In controls. Clutches o'f hens fed 50 ppm
toxaphene showed Improved hatching success In the second year of the study
compared with' the first year.
Toxaphene was Injected at levels of 0.1, 0.5, 1.0, 5.0 and 10 mg/kg egg
weight Into groups of 20 fertile Nick chick eggs on day 0 of their Incuba-
tion until day 14 (Srebocan et al., 1980b). The Injection was made 1n the
air sac and contained 0.1 ml of the toxaphene solution In sunflower oil.
The following assays were carried out on nine or more embryo Hvers per
treatment: pyruvate, pyruvate carboxylase, phosphoenolpyruvate carboxy-
klnase, fructose-1,6-d1phosphatase, glucose-6-phosphatase and glucose. The
effect on carbohydrate metabolism was similar In all groups. Increased
pyruvate concentration, decreased glucose concentration and decreased enzyme
02030 V-46 02/26/87
-------
activities, particularly 1n the embryos Injected with the higher doses,
Indicated that toxaphene" significantly Inhibited the gluconeogenlc pathway
In the embryonic liver. Lower reproductive success might also cause
embryonic death or a lowered vitality of hatched chicks.
Hoffman and Eastln (1982) determined the effects of externally treating
mallard (Anas platyrhychos) eggs with formulations and concentrations of
toxaphene similar to those In field applications. Toxaphene, In aqueous
emulsion applied on day 3 after the eggs were placed In an Incubator,
resulted In significant mortality (p<0.01) even at one-half the field level
of application (12.5 Ib/acre at 100 gal/acre); the IC™ was 108 Ib/acre
(121 kg/hectare). At concentrations greater than the LC5Q there were
abnormal survivors that exhibited dislocation of Joints. Treatment on day 8
at 5 times the field level resulted 1n embryotoxlc effects Including 53%
mortality, a reduction In growth, and an Increased Incidence of abnormal
survivors (18%), some with joint defects. When toxaphene was applied In an
oil vehicle either at day 3 or day 8, the LC^s were greater than for the
concentrated formulation (66 Ib/A). Treatment on day 8 resulted In growth
reduction (p<0.05). Higher concentrations resulted 1n brain, bill and Joint
defects. Exposures up to 5 times the field level did not grossly affect
cervical vertebrae as revealed by alizarin red staining.
Hutaqenldty
The National Toxicology Program (NTP, 1983) reported toxaphene to be
mutagenlc when assayed by the Salmonella/mlcrosomal reverse mutation assay.
Hill (1977), summarizing tests done by Litton Blonetlcs for Hercules, Inc.,
Indicated that toxaphene was directly mutagenlc only for Salmonella typhl-
02030 V-47 02/14/85
-------
muMum strains TA98 (which detects frameshlft mutagens) and TA100 (non-
specific). By contrast,-a "high temperature" toxaphene (high boiling compo-
nents) was mutagenlc only when rat Hver preparations (S9) were Incorporated
1n the assay to provide for metabolism of the compounds to biologically
active forms. Hooper et al. (1979) used Salmonella mutagenlclty assays to
Identify toxaphene components of potential carcinogenic hazard. TA100
proved to be the most sensitive Indicator of toxaphene mutagenlclty and H
was determined that mutagenlclty resided 1n the polar fraction. Mutagenlc
activity was reduced by 50% upon addition of rat or mouse S9 to the assay
plates. A major toxic component of toxaphene. Toxicant B, was not mutagenlc
for any of five Salmonella strains encompassing three DNA sHes. Treatment
of toxaphene with ethanollc KOH 1n molar ratios of 1:1 or 1:10 for 24 hours
at 25°C resulted In dechlorlnatlon of the major
-------
Griffin and H111 (1978) described use of an in vitro DNA breakage a-ssay
utilizing purified covalent closed circular ONA molecules of plasmld ColEl.
Breakage rates were determined after analysis by alkaline sucrose gradient
centrlfugatlon. Breakage rates significantly greater than control were
obtained with three mutagenlc alkylatlng agents and 4/11 pesticides tested
(dexon, dlchlorvos, malathlon and methyl parathlon). Toxaphene Incubated at
0.1 mg/mt 1n hexane with plasmld DNA for an unspecified amount of time
caused no Increase In breaks.
Epstein et al. (1972) used a modified dominant lethal assay In mice to
evaluate the mutagenlc potential of a variety of chemical agents Including
toxaphene. In this study, four groups of male ICR/Ha Swiss mice were given
toxaphene either IntraperHoneally (single doses of 36 or 180 mg/kg) or
orally (five doses of 8 or 16 mg/kg/dose). After dosing, the treated males
were mated to throe untreated females/week over an 8-week period. Females
were dissected 13 days following the midweek of their caging with the males.
Based on measurements of early fetal deaths per pregnancy and the percentage
of females with early fetal deaths, there was no significant difference
between control and toxaphene treated groups. Thus, 1n this strain of mice
toxaphene apparently does not produce chromosomal abnormalities In sperm
that preclude zygote development.
Sobtl et al. (1983) assayed organochloMne pesticides for their ability
to cause sister chromatld exchanges (SCE) 1n a human lymphold cell line,
LAZ-007, of B cell origin. Significant Increases 1n numbers of SCE were
observed with toxaphene concentrations of TO"5 and 10"* M. SCE
frequency, however, did not reach the level of 2 times control frequency
02030 ' V-49 02/14/85
-------
that has been recommended as the criterion for a positive response (Latt et
al., 1981). Samosh (T974) reported Increased frequency of chromosomal
aberrations 1n lymphocyte cultures obtained from eight women occupatlonally
exposed 1n a spraying operation to 2 kg/ha (13.IX 1n exposed vs. 1.6% \n
control). Aberrations consisted of acentric fragments and chromatld
exchanges. This 1s by contrast to a U.S. EPA (1978) study that found no
significant differences In rates of chromosomal aberrations In leukocytes
between groups of Individuals occupatlonally exposed to toxaphcne and
unexposed groups.
Cardnoqenlclty
The most definitive studies of toxaphene carc1nogen1c1ty were performed
by Tracor JHco Co. under contract from the National Cancer Institute (NCI,
1979) 1n spite of the fact they were not conducted 1n accordance with NCI
guidelines (control groups 'contained only 10 animals each; pair-feeding was
also not done). In this study both genders of Osborne-Mendel rats and
B6C3F1 mice of age 35 days were used. Each experimental group contained 50
animals of each gender, and the pesticide was added to the diet as an
acetone solution, 2% corn oil also being added as a dust suppressant. The
high and low male rat diets Initially contained 2560 and 1280 ppm toxaphcne,
respectively, and the corresponding diets for females contained 1280 and 640
ppm, respectively. In the case of mice the corresponding high and low dose
figures were 320 and 160 ppm diet for both sexes. Owing to overt toxlclty
these concentrations were later lowered. In male rats the high dose was
lowered to 1280 ppm at 2 weeks, and to 640 ppm at 55 weeks after Initiation
of the study. The low dose was similarly lowered to 640 ppm diet after 2
weeks and 320 ppm 55 weeks after feeding had begun. In the case of females,
02030 V-50 02/26/87
-------
both the doses were halved after 55 weeks, and for both sexes pesticide
treatment was discontinued after 80 weeks. Subsequently the animals were
fed control diets without corn oil for 20 weeks and then control diets for
an additional 8 weeks. In male and female mice both doses were halved 19
weeks after treatment was Initiated; toxaphene treatment discontinued after
80 weeks, and animals were fed control diets without corn oil for 7 weeks
then diets with 2% corn oil for an additional 3-4 weeks.
Animals that died during the study, and also all animals surviving at
the termination of the study, were submitted to pathologic evaluation of
major tissues, major organs and all gross lesions. In the male rats, 90% of
the high dose, 94% of the low dose and all of the control group lived until
at least week 52 of the study. In females 96% of the high dose, 92% of the
low dose and all 10 control animals survived beyond the 52nd week. Although
none of the tumors qbserved 1n treated animals were uncommon for the animal
strain used, certain tumors and hyperplastlc lesions were present with
higher Incidence In the treated animals. These Included thyroid folUcular
cell adenomas and carcinomas (7/41 low-dose, 9/35 high-dose and 1/7 control
males; 1/43 low-dose, 7/42 high-dose and 0/6 control females) and hyper-
plaslas (3/41 low-dose, 3/35 high-dose and 0/7 control males; 5/43 low-dose,
3/42 high-dose and 0/6 control females). The thyroid folUcular-cell hyper-
plaslas were observed only In the treated animals and were at relatively low
Incidences. Therefore It could not be concluded that Increased Incidences
of neoplasms or prollferatlve lesions were a result of toxaphene treatment.
These authors regarded adenomas as neoplastlc lesions; however, this is
controversial. Taking thyroid folllcular cell adenomas and carcinomas
02030 V-51 02/14/85
-------
together, a statistically significant Increase was found for the high-dose
group compared with th.fi matched controls for both male and female rats
(Tables V-8 and V-9, respectively). In comparison with values from histor-
ical controls from the same laboratory, statistical significance was also
found for the above lesions. In the female rats there was also a statisti-
cally significant Increase 1n the cumulative Incidence of tumors of the
pituitary (chromophobe adenomas, chromophobe carcinomas or adenomas) In the
high dose compared with the control group, although H 1s warranted only to
refer to the carcinomas as neoplastlc lesions. Due to the high spontaneous
Incidences seen In controls, NCI does not conclude that an association
exists between toxaphene administration and an Increased Incidence of
pituitary tumors. From the above study the authors concluded that toxaphene
administration was associated with an Increase In thyroid tumor Incidence.
Following an Independent examination of h1sto1og1cal preparations,
Reuber (1979) came to similar qualitative conclusions, reporting 10, 15 and
28% thyroid adenoma and 6, 18 and 18% thyroid carcinoma Incidence 1n control
male rats or 1n groups Ingesting the low or high dose of toxaphene. respec-
tively. In females, the corresponding figures were 6, 18 and 19% for
adenoma, and 10, 9 and 20% for carcinoma Incidence. In addition, Reuber
(1979) claimed Increases In benign and malignant tumor Incidence In
endocrine organs, reproductive system and mammary glands In toxaphene-
treated rats. Until differences between Reuber's criteria and those of
others are resolved, however, 1t will be difficult to draw conclusions from
his findings.
02030 V-52 02/14/85
-------
ro
CJ
O
1ABIE V-8
Incidence of lumors In Hale Rats fed loxaphene*
Dose
(ppm diet)
0 Hatched
Low
High
Number
Initial
10
SO
SO
of Animals
Examined
9
47
45
Skin Hemato-
potetlc
0/10 0/10
2/50(4X)d 0/SO(OX)
1/4S(2X) 2/45J4X)
Kidney Pituitary
0/9 0/7
1/45(2X) 0/42
1/4S(4X) 1/31 (3X)
Adrenal
HAL1GNAN1
0/9
1/4K2X)
0/37
Thyroid
0/7
1/41(2X)
2/35 (W)
Prostate
0/9
0/37
1/35 (3X)
Liver'1 Husculo-
skeletal
1/9(1 IX) 0/10
6/44 (14X) 0/50
4/45(9X) 1/45(2X)
Heso-
thelloma
0/10
0/50
0/45
Nervous
System
0/9
0/50
0^45
Totalc
0
7
10
0 Hatched
Low
High
10
SO
0/10
0/9
0/9
BENIGN
l/7(14X)e 2/9(22X) l/7(14X)f
2/7(29X)9 2/9(22X)
47 2/50(4X) 3/4S(7X) 2/45(4X) 1/42J2X)* 4/41(10X)n
12/42(29X)9
0/9
0/37
1/26(4X)J
SO 4S 1/4S(2X) 3/42(7X) 2/4S(4X) 4/31(13X)9 3/37(8X)n 7/3S(20X)f*k 0/3S
1/37I3X)1
0/9
0/44
0/4S
0/10
0/50
0/10
0/50
0/9
2/SO(4X) 27
1/4S(2X) 1/4S(2X) 4/4S(2X) 20
00
'Source: NCI. 1979
Liver * neoplasttc nodules
clotal = total anlMls with tuw>rs
tf Muaber of anlaals bearing iumor stated
NuHber of anlMls with tissue stated examined Microscopically
folltcular cell adenoma
^Chromophobe adenoma
Cortical adenoma
C-cell adenoma
Adenoma of parathyroid
Adenoma » carcinoma Incidence significant (Eisner exact text, p-0.008) compared with matched controls
Pheoc hr omoc y t oma
-------
TABLE V 9
Incidence of Tiwors In Female Rats Fed Toxaphene*
IVJ
o
OJ
0 Niwber of Anlaals
Dose Skin Lung Liver Pituitary Adrenal Thyroid MaMury Uterus Ovary Kidney Tolalb
(ppadlet) Initial Examined
0 Hatched 10 10 0/10 0/9 1/10J10X) 0/8
Low SO 49 l/50(2X)e 0/46 4/42(10X)d 0/41
MALIGNANT
0/8 1/6(17*) 0/10 0/9 0/8 0/8 1
0/44 0/43 1/50(2*) 1/41(2*) 0/40 0/49 4
ifl
High
0 Matched
Low
High
SO
10
SO
so
49
10
49
49
1/42(2X)<
3/49(6X) 1/48(2X) 4/40(10X) l/39(3X)f 2/43(5*) 0/42
BENIGN
0/10 0/9 0/10 1/8(13X)9 0/8 0/6
2/8(25*)'
0/50 0/46 1/46(2*)" 15/49(37*)' 3/44(7*)J 1/43(2*)
3/49(6*) 0/45
1/10(10X)
2/50(4X)9 9/50°
l/SO(2X)k
10/50(20*)!
1/36(3X) 0/481
0/8
0/18
1/50(2X) 0/49
28
1/49(2X) 0/48 1/40|3*)h 18/39(46*)' 4/43(9X)J 7/42(17X)n 1/49(2X)« 1/45«2X)J 0/36 1/48(2X) 36
4/39(10X)f>« 2/49(4X)k 5/45(llX)«
10/49(20*)'
'Source: NCI. 1979
bTotal - total antMls with tuaors
c Number of antaals bearing tuaor stated
NiMber of anlaals with tissue stated examined Microscopically
dNeoplasttc nodule In the liver of low dose
eHepatocellular caret now In the liver of low dose
*lotal tumor Incidence of tuwtrs In pituitary (p»0.012. Cochran-Arattage test) coapared with pooled controls
"Bile duct haMrtoM
^Chroaophobe ademxu
JCorttcal adnoaa
kflbroM
^Flbroadenoaa
Pancreatic llpoaa
"Eolllcular cell adenoM Incidence significant (p=0.021. fisher exact test) compared with pooled controls
°StroMl polyp
CO
-------
In the case of mice, greater toxldty of toxaphene was reported (NCI,
1979). As In the case "of rats, survival 1n treated groups was high; 90-98%
of treated and control animals survived beyond the 52nd week of the study.
Of the tumors appearing In treated animals, none were observed In greater
Incidence compared with controls with the exception of those In the liver.
Hepatocellular carcinomas occurred 1n treated mice only, with Incidences of
69% and 98% In males (Table V-10) at the low and high doses, and 10% and 69%
In females (Table V-ll) at the low and high doses, respectively. These
neoplasms were not observed In control animals of either sex, but hepatic
nodules were observed 1n 20% of matched-control males, though not In
females. On the basis of these findings the authors concluded that toxa-
phene caused Increases In the Incidence of hepatocellular carcinomas and
neoplastlc nodules, and hence was carcinogenic In B6C3F1 mice.
Reexam1nat1on of the hlstologlcal sections prepared In the above study
led Reuber (1979) to come to qualitatively similar, but quantitatively dif-
ferent conclusions. In this regard he reported liver carcinoma Incidences
of 20, 78 and 100% In male mice Ingesting zero, low or high doses of toxa-
phene and 0, 31 and 89% respective Incidences 1n females. In addition one
male mouse Ingesting the low concentration developed an anglosarcoma of the
liver, and one female an adenocardnoma of the salivary gland. In addition,
Reuber reported Incidences of 0, 12 and 31% of stromal cell carcinoma 1n the
uterus of females In control, low-dose and high-dose groups, respectively,
the latter Incidence being the only statistically significant result. No
mention was made of these carcinomas 1n the original NCI (1979) report.
02030 V-55 02/26/87
-------
O
CO
1ABIE V 10
Incidence of luaors In Hale Nice fed loxaphene In the Diet'
I
en
Dose
(•g/kg diet) Initial Examined
0 Hatched 10 10
Low 50 50
High 50 46
Blood Liver Total*
HAL1GNAN1
0/1 Od 2/10(20X) 0
2/50(4X) 6/49(1 2X)e 37
34/49 (69X)r
0/50 45/46(98X)f 45
Lungc
1/10(10X1
1/49(2X)
2/46(4X)
fye
BENIGN
0/10
1/50(2X)
0/50
Totalb
1
2
2
•Source: NCI. 1979
bToUl . total an tails with tutors
cAlveolar/bronchlolar adenoaa
«• Nuaber of anlaals bearing iumor stated
NiMber of anlMls with tissue stated exaatned Microscopically
"12K were described Is have "p.ecplasttc nodules.* assuaed to be Ml!«nani. 69X hepatocellular circlnona and ?X
'incidence of hepatocellular carcinomas In low and high dose groups significant (p<0.001. fisher exact test) compared with Matched or pooled
controls.
CO
-------
INJ
o
00
o
1ABLE V 11
Incidence of Tutors In female Mice fed loxaphene In the 0»eta
Dose
(•g/kg diet)
0 Hatched
Low
High
'Source: NCI.
blotal . total
Number of
Initial
10
SO
SO
1979
antMls with
AnlMls
HeMtopolettc Liver Total0 Salivary Mammary Uterus Harder Ian Totalb
Examined System Gland
I
ML1GNAN1 BENIGN
10 0/10* 0/9 0 0/8 0/10 0/10 0/10 1
49 1/50(2X) S/49(10X)d 6 1/46(?X) 1/SO(2X) 2/48(4%) 0/SO 4
13/49(?7X)d
49 0/SO 34/49(b«)*.f 34 0/47 0/SO 0/44 1/50(2X) 1
6/49(1 W)«.f
tumors
c Number of antMls bearing tumor stated
NuMber «f anlaals with tissue stated exanlned Microscopically
d27X were reported to have "neoplastlc nodules.* (ascribed Malignant) and 10X hepatocellular carclnona
e!2X were reported to have "neoplastlc nodules.* and 69X hepatocellular carclnotu
'incidence of hepatocellular carctnotaas significant (p<0.001. fisher exaci iesij coapared with aatched or pooled tontro! groups
oo
tn
-------
In addition to the previous study, a number of other much less complete
studies on toxaphene carcinogenic1ty were conducted. In a study conducted
by the U.S. FDA (Nelson, 1949), groups of 12 male and 12 female rats assumed
to be of the Osborne-Mendel strain Ingested 0, 25, 100, 400 or 1600 ppm
toxaphene 1n the diet for 107 weeks. Further groups of 5 male and 5 female
rats wore given 0, 40, 200 or 1000 ppm toxaphene and sacrificed after 56
weeks. Tissues were sectioned only from the first set of groups, however,
and Incomplete histology was performed on other groups. Only one neoplasm
was noted, a subcutaneous neurosarcoma In a male rat fed the 1000 ppm do-se
level, although liver hyperplasla and hyperplastlc nodules were reported.
Thyroid hyperplasla was also noted In treated animals, especially In males.
In the groups constituting the 107-week experiment, 2/3 female and 2/2
male rats Ingesting 1600 ppm toxaphene developed hepatic carcinomas; 1/3
female rats In each of the 400 and 1600 ppm groups developed hyperplastlc
nodules. No Incidence of either of the above tumors was observed In control
or other treatment groups (Nelson, 1949).
A further study demonstrating a statistically-significant Increase In
hepatic neoplasms was also done (Litton Blonetlcs, Inc., 1978). Fifty-four
weanling B6C3F1 mice were randomly assigned to each of four groups. Toxa-
phene (X-16189-49), as well as the dose levels, were selected and provided
by Hercules Inc. The animals were administered toxaphene at 0, 7, 20 and 50
ppm levels 1n the diet for 18 months and surviving mice were kept on the
control diet for another 6 months. All survivors were killed 24 months
after the Initiation of treatment. Table V-12 shows the effects of
toxaphene on survival. The survival In both treated and control groups was
similar.
02030 V-58 02/26/87
-------
TABLE V-12
Survival of Mice fed Toxaphene 1n the 01eta
Sex
Hale
Female
Dose
(ppra)
0
7
20
50
0
7
20
50
No. at
Start
54
54
54
54
54
54
54
53
1-26
0
0
0
lb
0
0
1
0
Weeks -
1n a Hor
27-52
0
0
1
0
0
0
1
0
Died or Killed
1bund Condition
53-78
1
0
3
0
0
2
2
0
79-105
10
8
6
7
10
7
8
9
aSource: Litton B1onet1cs, Inc., 1978
bM1ss1ng
02030
V-59
02/26/87
-------
The hlstopathologlcal evaluation data presented 1n Table V-13 shows a
statistically significant-Incidence of hepatocellular tumors (hepatocellular
carcinomas and hepatocellular adenomas) In male mice fed 50 ppm toxaphene In
the diet as compared with the controls. However, hepatocellular tumor Inci-
dence was not statistically significant 1n females as compared with controls.
Toxaphene was administered to 50 weanling ARS Golden Syrian hamsters of
each sex and each dose at levels of 0, 100, 300 and 1000 ppm based on a
subchronlc study (LHton B1onet1cs, Inc., 1979). The hamsters, obtained
from the Sprague-Oawley laboratory In Madison, HI, were randomly assigned to
each group. The toxaphene was mixed 1n corn oil and blended with the food
that was available ad libitum. The animals were observed during a 15-day
acclimation period before Initiation of the study and dally afterward for
general condition and mortality Treatment was continued for 18 months for
the females and 21.5 months for the males who showed a high survival at 18
months.
Animals were observed until spontaneous death, unless moribund, or
surviving until termination of the experiment. Moribund animals and those
surviving until termination were sacrificed. All animals were necropsled.
The postmortem observations Included a thorough external examination and
collection of major organs and representative tissues from all animals. All
collected tissues from control and high-dose animals, and target organs and
gross abnormalities from the low- and Intermediate-dose animals, were
evaluated h1stopatholog1cally. Blood smears were taken from all animals at
sacrifice, but were not examined unless necessary for definition of specific
lesions or diseases. A compound-related lowered body weight was observed 1n
02030 V-60 02/26/87
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TABLE V-13
Hepatocellular Tumor Incidence In B6C3F1 Mice fed Toxaphene In the Diet*
Hepatocellular Tumor Incidence
Dose
(ppm)
0
7
20
50
Hales
10/53 (19%)
10/54 (19%)
12/53 (23X)
18/51 (35X)
Females
2/53
2/53
4/53
6/52
(4%)
(4%)
(8%)
(12%)
*Source: Litton B1onet1cs, Inc., 1978
02030
V-61
02/26/87
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high-dose males until 12 months of study. Similar effects were noted In
males receiving 300 ppm "from months 2-7 and for a brief period In females
receiving 1000 ppm. At terminal sacrifice, an Increase 1n liver weight was
observed 1n males at the 1000 ppm dose, which correlated to the pathologic
finding of megahepatocytes In 6 of 21 males observed. A decrease In heart
weight was noted In males at both the 300 and 1000 ppm dose levels. A
decrease In the thyroid gland weights of the 1000 ppm females was also noted.
Only the liver changes were supported by hlstopathology and were there-
fore judged to be treatment-related. These liver changes are similar to
those accompanying an Induction of liver enzymes after administration of
chlorinated hydrocarbons and other compounds. Microscopic evaluation of the
tissues did not reveal any Incidence of a tumorlgenlc effect related to
toxaphene. Even when the Incidence of lymphoretlcular neoplasms of all
types was combined, no difference was detected between controls and dosed
animals. Under conditions of this study, toxaphene was not carcinogenic.
It Is likely that the maximum tolerated dose was not employed In these
studies (Litton Blonetlcs, Inc., 1979). In the subchronlc study, the
maximum tolerated dose estimate was based on evidence of liver toxldty
only. No hamster at any dose level died during the study; no changes In
general appearance, condition or behavior among experimental animals was
observed; and measurements of mean body weights and food consumption
Indicated no compound-related change. In the chronic study, a dose-related
decrease In body weight was observed, but H was temporary. The weight of
treated animals equaled or surpassed that of controls by the end of 1 year
for all treatment groups. Treated animals also survived for a longer period
02030 V-62 02/26/87
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than controls. A comparison of the number of animals 1n each treatment
group surviving to termination of the experiment using an X2 test For
homogeneity revealed significant differences for both males (p<0.05) and
females (p<0.001).
Intragastrlc Injection of toxaphene 1n sunflower oil solution )n a dose
of 50 mg/kg bw twice-weekly for 10 weeks was reported to give no excess
tumor Incidence In mice (18 g Initial weight) (Oldenko et al., 1978). The
mouse strain was apparently Indeterminate (stated as nonlinear), and both
sexes were used. Although the sites of the tumors 1n each sex were not
reported, the "tumor formation frequencies" of 12.5X for treated, and 8.9%
for control groups were reported to be not significantly different 60 weeks
after the start of the experiment. Of toxaphene-treated mice, 8 bore
pulmonary adenomas, 1 adenocarclnoma, and 1 lymphogranulomatosls. In the
control group, five mice had pulmonary adenomas. In a parallel study In
rats, animals of 80 g weight were Injected twice-weekly for 16 weeks with 80
mg toxaphcne/kg bw 1n sunflower oil. No primary data were presented but
toxaphene Injection was reported not to Increase the frequency of tumor
formation compared with controls, or to Induce differences In tumor distri-
bution relative to gender. Both control and treated animals were reported to
develop leukemia and lung tumors by the age of 2.
In a study on the effects of toxaphene on spontaneous and BaP-1nduced
cardnogenesls In female A/J mice (TMolo et al., 1982), the pesticide was
added to the diet In a fashion such that control diets contained 5% corn oil
and experimental diets 5% corn oil and 100 or 200 ppm technical grade toxa-
phene. Mice were 9 weeks old on Initiation of the study, and were maintain-
ed on these diets for 12 or 20 weeks. Following the 12-week experiment
02030 V-63 02/26/87
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essentially no tumors of any kind were found In the forestomach, lung,
liver, glandular stomachr Intestinal tract or spleen from control or experi-
mental groups. When mice were Intubated with 3 mg BaP In 0.25 ml corn oil
on days 7 and 21 by oral Intubation, 100 ppm toxaphene In the diet had no
effect on BaP-1nduced forestomach tumorlgenesls, but 200 ppm caused a stat-
istically significant decrease In tumor Incidence. In the case of the lung,
200 ppm was without effect, but 100 ppm resulted In decreased tumor (presum-
ably adenoma) Incidence slgnlfIcant.ly compared with BaP alone. Concurrent-
ly, the same dietary concentrations of toxaphene were found to significantly
Increase control and BaP-1nduced hydroxylase activities In the liver, but to
reduce them 1n lung (TMolo et al., 1982).
In the 20-week portion of this experiment BaP-treated mice Ingested
toxaphene at 100 ppm 1n the diet; a significant decrease 1n the number of
lung tumors per mouse was observed compared with mice treated with BaP
alone. In mice treated with the pesticide alone, the Incidence of tumors
was low and Identical to untreated controls. The Incidence of forestomach
tumors In the animals receiving BaP and toxaphene was reported to be too
large to quantify accurately. Similar effects were seen on BaP-hydroxylase
activity after 20 weeks of toxaphene Ingestlon to those observed after a
12-week feeding period (TMolo et al., 1982).
In order to assess the carcinogenic activity of toxaphene In vitro, the
effect of the pesticide at concentrations of 0.3-100 yM on C3H10T1/2CL8
cell transformation Induced by 4 yM BaP was Investigated (Nesnow and
Leavitt, 1979). Of a number of agents tested, toxaphene was least active
but was reported to Increase the BaP Induced transformation frequency In a
dose-related manner. No primary data were presented.
02030 V-64 02/26/87
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Summary
Acute oral exposure of mammals and birds to toxaphene resulted 1n.LD5Q
values ranging from 7.5-10 mg/kg bw In female monkeys to 75-500 mg/kg In
rabbits (see Table V-l). The LD5Q values resulting from dermal applica-
tion of toxaphene are generally higher than those observed after oral
exposure and range from -250 mg/kg bw to -4000 mg/kg, both values being
reported for rabbits (see Table V-2). Toxaphene fractions or components
differed 1n chemical composition, polarity and solubility leading to
>10-fold differences In toxldty. The vehicle 1n which the pesticide Is
solublUzed for administration also appears to have an effect on the toxic
response. Specific components of technical toxaphene are more toxic than
the complex mixture, and as reported for mice, ID,- values for several
components varied from 2.5 to >100 mg/kg bw (see Table V-3). Brain levels
of toxaphene may be Indicative of acute toxldty (see Table V-4). In swine,
>4 mg/kg wet weight (brain tissue) constituted a lethal level, and 2 mg/kg
was associated with clinical signs of toxldty.
Subchronlc oral doses of toxaphene 1n laboratory animals resulted In few
clinical signs of poisoning. Responses varied from lethality In several
mice given 1280 ppm In the diet for 6 weeks to no apparent adverse effects
In rats following 1ngest1on of feed containing 189 ppm toxaphene (see Table
V-5). At doses <189 ppm In the diet, subcllnlcal toxldty was reported such
as decreased bile production and questionable liver pathology. In mice
receiving 100 or 200 ppm toxaphene 1n the diet, humoral antibody production
(IgG antibody formation) was suppressed; however, cell-mediated Immune
responses were not affected.
02030 V-65 02/26/87
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Toxaphenc aerosols In the form of dusts are apparently more toxic to
rats than toxaphene mists. M1st concentrations as high as 500 mg toxa-
phene/ma for 3 weeks caused no mortality 1n rats and rabbits; at 12 mg
toxaphene dust/ma for 3 months, mortality occurred 1n rats, dogs and
guinea pigs. These results may be a function of particle size.
Long-term .exposures of animals to dietary levels of toxaphene are sum-
marized 1n Tables V-6 through V-ll. In a number of studies (see Table V-6),
no adverse effects were reported among the parameters monitored 1n each
experiment. In several lifetime studies In rats, no effects were reported
for doses between 10 and 100 ppm toxaphene 1n the diet; other studies at 100
ppm reported liver pathology. The lowest dietary level producing liver
damage In dogs was 5 ppm In the diet for 2 years. In addition to liver
pathology, a variety of adverse effects are reported from both J[n vivo and
in vitro studies: kidney pathology, decreased bile flow, decreased
survival, Inhibition of Intestinal transport of glucose, Inhibition of
gluconeogenesls, Inhibition of mitochondria! enzymes, Inhibition of
* *5
Na'/K'-ATPase brain and kidney activity and Mg -ATPase activity
Inhibited 1n mouse kidney, liver and brain.
Toxaphene Induces the mlcrosomal MFO system, and treatment of an animal
Ifl v^vo wlth a cytochrome P-450 system Inhibitor Increases toxaphene
toxlclty.
In a 3-generatlon study with 25 or 100 ppm toxaphene In the feed of
Sprague-Dawley rats, no effects on parental animals or offspring could be
detected. With CD rats, maternal mortality was 31% at 35 mg toxaphene/kg bw
02030 V-66 02/26/87
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and there was a dose related decrease 1n maternal weight gain and fetal
weight (15, 25 or 35 mg/kg bw). No adverse effects were reported In the
offspring of mixed-strain white rats given 4 mg/kg bw during the entire
pregnancy. In hamsters given the same dose, the authors reported toxaphcne
to be teratogenlc but did not Identify the anomaly.
More sensitive endpolnts In rats were examined by several Investigators.
Toxaphene (oral), at 12 mg/kg bw given to pregnant rats, depressed
chollnesterase activity 1n fetal cardiac neural structures and delayed
cardiac neural differentiation. At 25 mg/kg bw, of the many biochemical
parameters examined, the only significant difference was noted in the kidney
(decrease In alkaline phosphatase activity and total protein). There was no
effect at 12.5 mg/kg bw on the rat kidney. In two separate experiments,
there was little evidence of behavorlal deficits In the offspring of
toxaphene treated'dams except for retarded righting ability (6 mg/kg and 50
ug/kg bw).
Toxaphene, given to pregnant mice (15 mg/kg and 25 mg/kg bw) had no
adverse effect on parameters examined 1n offspring. In this same study the
high dose (35 mg/kg bw) Induced encephaloceles 1n five Utters of 90 treated
dams. A 5-6 generation study with toxaphene (25 ppm) In the diet resulted
In little or no adverse effect In mice. No effects were noted on the
anatomical development of the fetal guinea pig from pregnant females given
15 mg/kg bw from days 21-35 of gestation.
02030 V-67 02/26/87
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Toxaphene appears to have a low teratogenlc potential unless doses are
high enough to Induce maternal toxldty. In the studies reported here, the
lowest dose producing an effect was 50 yg/kg bw (rats) given In the diet
from day 5 of gestation through termination of the study; effects Included
significant decreases 1n overall swimming ability and righting ability of
young pups when toxaphene was administered before postnatal day 16.
Toxaphene Is mutagenlc 1n the Salmonella/mlcrosomal reverse mutation
assay using strains TA98 and TA100. Results of assays of toxaphene compo-
nents for their mutagenlc potential Indicated that mutagenlclty resided In
the polar fraction. Mutagenlclty was decreased by the addition of active
MFOs, S9; this agrees with the in vivo observation that Inhibitors of cyto-
chrome P-450 Increased toxldty. The mutagenlc activity of commercial prep-
arations varies.
Studies of toxaphene cardnogenlclty are reported 1n Tables V-8 through
V-14. In one study with cardnogenlclty as the endpolnt (NCI, 1979), many
doses were lowered because of overt toxldty 1n the rats and mice Ingesting
toxaphene containing feed. The concurrent numbers of control animals were
low (10/sex/group) and historical controls were used 1n order to match the
number of treated animals per group (50 of each sex/group). The range of
doses given to the animals makes 1t difficult to use THAs of doses as a
basis on which to estimate risk for humans (Table V-14). It was concluded
that, under the conditions of the bloassay, toxaphene Is carcinogenic In
male and female B6C3F1 mice, causing Increased Incidences of hepatocellular
carcinomas 1n a dose-related manner. The results also revealed that toxa-
phene 1s carcinogenic for the thyroid of male and female Osborrne-Mendel rats.
02030 V-68 04/02/87
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TABLE V-14
Variation 1n Ooses of Toxaphene Used In NCI (1979)
Study of Cardnogenldty
Time-Weighted
Species Sex Average Dose Range of Dose
(ppm In feed) (ppm In feed)
Rata
Mouseb
male
female
male
female
male
female
1112
556
1080
540
198
99
640-2560
320-1280
640-1280
320-640
160-320
80-160
Increased Incidence of thyroid tumors 1n both males and females at the
high doses.
Increased Incidence of hepatocellular carcinoma In males and females at
both doses.
02030 V-69 02/26/87
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In another study with B6C3F1 jnlce of both sexes, using 7, 20 and 50 ppm
toxaphene In the diet, the cardnogen1c1ty of toxaphene was demonstrated at
these much lower doses. After 18 months of toxaphene Ingestlon and 6 months
on a control diet, there were Increased Incidences of hepatocellular
carcinoma 1n males. The response 1n females was less pronounced.
Reports from the U.S. FDA study (Osborne-Mendel rats) Included one sub-
cutaneous neurosarcoma In a male rat given 1000 mg toxaphene/kg diet, liver
hyperplasla and hyperplastlc nodules, and thyroid hyperplasla.
A significant Increase 1n hepatocellular tumors was detected In mice In
two separate studies. When adenomas and carcinomas are combined, a statis-
tically significant Increase In thyroid tumors was noted In both male and
female rats along with a significant Increase In the cumulative Incidence of
pituitary tumors 1n female rats. These findings are considered sufficient
evidence to classify toxaphene as an animal carcinogen. Using the IARC
criteria for classifying the carcinogenic evidence with "sufficient" animal
data, toxaphene 1s ranked 1n the IARC 2B group, meaning that toxaphene Is
probably carcinogenic 1n humans. Classification according to the EPA
guidelines for carcinogens classifies toxaphene 1n group B2: Probable Human
Carcinogen, meaning that there 1s sufficient evidence of cardnogenldty
from animal studies and Inadequate or no data from ep1dem1olog1c studies.
02030 V-70 04/02/87
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VI. HEALTH EFFECTS IN HUMANS
Acute Tox1c1ty
Toxaphene, like most Insecticides, Is poisonous by swallowing, skin con-
tact, or Inhalation (Boots Hercules Agrochemlcals, Inc., n..d.). The fol-
lowing properties are cited 1n Boots Hercules Agrochemlcals, Inc., undated.
Emulsions or solutions In vegetable oils facilitate skin or gastrointestinal
absorption. Solutions 1n kerosene are hazardous, but absorption 1s much
more "erratic." When toxaphene Is 1n the form of a powder, dermal absorp-
tion Is negligible, but Inhalation exposure of resplrable particles may be
considerable. Toxaphene poisoning In humans 1s manifested by diffuse stimu-
lation of the central nervous system resulting 1n salivation, restlessness,
hyperexc1tabH1ty, muscle tremors or spasms, generalized convulsions, and
sometimes loss of consciousness. Nausea and vomiting may follow oral 1nges-
tlon. Clonlc convulsions may also occur and can be prevented by barbitu-
rates (I.e. sodium pentobarbHal) administered preferably by 1.v. Toxaphene
has a fairly long duration of action (I.e. over several hours) and a long-
acting barbiturate such as phenobarbltal can be used after Initial control
of convulsions. In lethal cases, convulsions continue until death, which Is
ultimately caused by respiratory failure.
The IUPAC (1979) has estimated an acute oral LD,Q dose of 60 mg toxa-
phene/kg bw. Hayes (1975) reported a level of 78 ppm 1n the liver of a dead
person poisoned with toxaphene as well as 10 ppm In blood. Conley (1952)
estimated an acute lethal dose to be between 2 and 7 g/person or 29-100
mg/kg for a 70 kg man.
02040 VI-1 02/14/85
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Fatal Intoxications of humans have been caused by "cotton dust" contain-
ing 5X DOT, 10% toxapherre and 40X sulfur. The "subacute fatal human dose"
for this mixture Is reported to be 0.75 g DDT * 1.5 g toxaphone (Heyridrlckx,
1960).
Several cases of death from toxaphene poisoning have been recorded
(Hayes, 1963). A 9-month-old child poisoned with a 2:1 mixture of toxa-
phene:DOT died after convulsions and respiratory arrest. Toxaphene was
detected 1n the liver (7.9 ppm), kidney (6.8 ppm) and brain (14 ppm). The
ratio of toxaphene:DDT In the brain and liver was 10:1, and In the kidneys,
3:1 (Haun and Cueto, 1967). Four other cases of acute poisoning In
children, 3 of them fatal, have been reported (McGee et al., 1952). Table
VI-1 summarizes the findings of this study. No residue data were reported.
Recovery of humans from poisoning occurred at doses of 10-47 mg/kg bw.
Vomiting was Induced so 1t Is not clear how much toxaphene was absorbed.
Poisoning by prolonged skin contact with a Undane/toxaphene solution
has been reported by Pollock (1958). The patient was semiconscious, dis-
oriented and stuporous, nauseous and had vomited many times. Severe
epigastric pain, severe aches, and pains of the extremities were also
reported. Eventually stiff neck, dehydration, and Irregular pulse develop-
ed. Treatment was with penicillin, chloromycetln and digitalis. The
atypical symptoms may be related to the presence of Undane or the dermal
portal of entry. No residue data were given. Hayes (1963) estimated a
hazardous dermal dose to be 46 g. For a 70 kg man, this Is equivalent to
660 mg/kg or about 10 times that of an oral dose.
02040 VI-2 02/25/87
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o
ro
O
I
CJ
1ABIE VI-)
Case Studies of Toxaphene Poisoning In Humans In which Ingest ton Has the Primary Route of Entry*
Case No.
Sub)ect(s)
Mature of
toxaphene
Dose
Time to react
symptoms
Symptoms
Outcome
Time of death
or recovery
1 2
Male. 2 years Male. 4 years
8 Moths
Max Emulsion In
water
Unknown Unknown
7 hours 2 hours
Convulsions Convulsions
2-5 Minute
Intervals
Death Death
9.S hours 6 hours
3
Hale. 1 year
S Months
60X in
solvents
Unknown
MS
Convulsions
Intermittent
Death
11 hours
4
Male. 2 years
20X In solution
Unknown
MS
Convulsions. Intermittent
•lid cerebral excitement;
aimless jerking motion
and excessive muscular
tensions of extremities.
marked pharyngeal and
laryngeal spasms; labored
respiration; cyanosis
Recovery
12 hours
5
Female. 49 years
Female. 20 years
Female. 16 years
Female. 12 years
Residue of spray
In food
9.5 47 mg/kg
1.5-4 hours
Nausea; vomiting
convulsions
Recovery
12 hours
6
Hale, adult
Male, young
i
Residue of spray In
food
Unknown
4 hours
No nausea; spontaneous
vomiting; convulsions.
Jerking and transitory
movements; muscular
rigidity; periods of
unconsciousness;
amnesla(?)
Recovery
24 hours
•Source: HeGee et al.. 1952
NS ~ Not specified
'CO
-------
Allergic bronchopneumonla was Immediately observed In two workers who
had just used toxaphene, sprays (Warrakl, 1963). One had been exposed to
other pesticides for 4 years and showed marked bilateral hllar. lymph-
adenopathy with fine mlHary opacities heavily distributed over both lungs
which, after treatment with dlhydrostreptomycln and 1son1az1d, regressed.
The other man also showed similar symptoms, and no past pesticide exposure
was evident apart from that to toxaphene. The m111ary shadows regressed
when the man was treated with streptomycin, 1son1az1d, prednlsone and cor-
tisone. Both cases evidenced the following: sudden exertlonal dyspnea,
tachycardia and weakness with low blood pressure; extensive mlHary shadows.
In both lungs; blood eos1noph111a; high serum globulins; reversibility of
X-ray shadows, symptoms and physical signs to normal on treatment with cor-
tisone under cover of antltuberculosls drugs; and acute pneumonia symptoms.
Eight women working In an area that had been sprayed with 2 kg toxa-
phene/ha by aircraft showed a higher Incidence of chromosome aberrations
(acentric fragments and chromatld exchanges), as observed In lymphocyte cul-
tures, compared with an unspecified number of control Individuals: 13.1%
vs. 1.6X (Samosh, 1974).
The sequelae observed after a pesticide fire In which -80 fire fighters
were potentially heavily exposed to toxaphene amongst many pesticides
(Mellus and Schulte, 1981) were generally vague (tension, depression, hyper-
exc1tab1l1ty, fatigue, and confusion were significant relative to the 24*
fire fighters who reported acute symptoms of nausea, dizziness, headache at
the time of the fire).
02040 VI-4 02/14/85
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Subchronlc and Chronic Toxlcltv
Application of an aerosol spray containing toxaphene to the skin of 50
human subjects dally for 30 days at a dose of 300 mg/day produced no toxic
manifestations. Fifty human volunteers who Inhaled 0.4 mg/m3 of toxaphene^
aerosol for 10 minutes a day for 15 days had no subjective or objective
effects. Assuming alveolar ventilation of 5.4 i/m1nute the dally dose
equalled -0.02 mg. A mist containing 250 mg/m3 toxaphene was Inhaled by
25 humans for 30 minutes each day (-41 mg/day) for 13 days and they showed
no evidence of local or systemic toxic manifestations (Shelanskl, 1947).
Many studies exist that Involve long-term toxaphene exposure to workers.
Most of them do not detail exposure data, Involve many other pesticides, and
do not attempt to correlate serum levels of toxaphene or Us metabolites to
known symptoms of toxaphene so that the results, while suggestive, are very
often not definitive. Examples of studies of this kind follow. One was a
health study detailing an Increased Incidence of lung cancer (10 observed
vs. 0.54 expected) among 285 pesticide applicators (Barthel, 1976). Another
was a survey Involving 321 exposed and 46 nonexposed males 1n Kuwait In-
which the 260 exposed to organochloMnes reported Increased sputum and
hyperreflexla compared with controls (El-Desouky et al., 1978). A third
study was a nonsignificant case-control study of the association between the
yearly rate of fatal aplastlc anemia and the use of many pesticides In the
USA between 1950 and 1975 (Wang and Grufferman, 1981); a fourth Investiga-
tion was a cancer morbidity study (Barthel, 1981) In 1658 male workers of at
least 5 years experience as agricultural plant protection workers or plant
protection agronomists In the German Democratic Republic between 1948 and
1972. Toxaphene exposures occurred after 1954. An Incidence of 169 mallg-
02040 VI-5 02/14/85
-------
nant neoplasms was found. The ones with a significant excess compared with
the general population^were bronchial carcinoma, 59 (35%) to 42 (24%),
respectively, of which 22 were undlfferentlated and small-cell carcinomas,
14 were squamous epithelium carcinomas, 3 were adenocarclnomas, 2 were solid
carcinomas and one was an alveolar cell carcinoma. The average latency
period for the lung cancers was calculated to be 17^6 years (6-29 years) and
these cases had been exposed on the average for 14+5 years (6-23 years).
Smoking was not a significant factor 1n these cases. Toxaphene was thought
not to be a significant factor 1n these Incidences.
In a survey of 199 employees who worked or had worked with toxaphene
between 1949 and 1977, with exposures ranging from 6 months to 26 years-
(mean 5.23 years), 20 employees died, one with cancer of the colon. None of
these deaths could be attributed to toxaphene (IARC, 1979).
No Increased porphyrln production was found 1n the urine of 45 workers
exposed dally to a variety of pesticides, predominantly parathlon, toxa-
phene, DDT and dleldrln. The qualitative method used was capable of detect-
ing 0.4 yg porphyr1ns/ml. No correlations were found between serum
pesticide levels and urinary excretion of ALA, PBG and CCA. Parathlon
depressed plasma or red cell ChE or both In five workers. These findings
suggest that ordinary occupational exposure to the pesticides noted above
has no strong porphyrlnogenlc or sympathotonlc effects (Embry et al., 1972).
02040 VI-6 02/14/85
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Summary
Toxaphene has caused convulsions and nausea In humans when exposure has
occurred by swallowing, skin contact or Inhalation. The acute oral L0,n
has been estimated In one study to be 29-100 mg/kg, and 60 rng toxaphene/kg
bw In another. Mixtures of toxaphene with DOT (2:1, by weight) are more
acutely toxic than either component alone, the subacute fatal human doses
being 0.75 g DDT and 1.5 g toxaphene. The liver, brain and kidney can show
residues. The estimated hazardous dermal dose Is -660 mg/kg bw for liquid
toxaphene. Allergic bronchopneumonla has been observed after acute Inhala-
tion of toxaphene sprays.
Dermal doses of 300 mg/day for 30 days, and Inhaled doses of 0.4 mg/m3
for 10 minutes on each of 15 days, and 250 mg/ma for 30 minutes (-41
mg/day) on each of 13 days caused no atypical subjective or clinical effects.
Studies In the workplace are confounded because exposure to many
chemicals occurred 1n all the reported studies. There are no porphyrlnogen-
1c or sympathotonlc effects mentioned 1n the available literature.
Human data are Inadequate to assess the human carcinogenic potential for
toxaphene.
02040 VI-7 02/25/87
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VII. MECHANISMS OF TOXICITY
Introduction —•
Toxaphene toxldty 1s Influenced by many factors: age, gender, species,
state of mlcrosomal-enzyme systems, presence of starvation or of other
chemicals that may affect cytochrome P-450 levels, and the type of carrier
utilized In toxldty experiments (see Chapter V).
In all cases the obvious acute effects In humans and animals (saliva-
tion, hyperexcltabUHy, behavioral changes, muscle spasms, convulsions)
Indicate that the central nervous system Is the chief target system. The
mechanism of the effect 1s still unknown. Other target organs affected
acutely Include the kidneys (cloudy swelling, congestion, renal tubular
degeneration), the liver (fatty degeneration, necrosis, Inhibition of bile
flow and biliary function), the testes (decreased spermatogenesls) and the
skeleton (altered cartilaginous structures In birds and In guinea pig
fetuses).
At least In Swiss-Webster albino mice (Saleh et al., 1977; Saleh and
Caslda, 1979), enhanced acute toxldty (oral LD5Q values) was associated
with Introduction of a single chloro-group In the 8- or 9-pos1t1on and to a
lesser extent 1n the 5-exo-pos1t1on. A decrease In toxldty for hepta-
chlorobornanes was noted for single chloro-groups In the 3-exo or 10-posl-
tlons. For the nonachlorobornanes, additional chloro-groups In the 3-exo.
10-posltlons or 1n the 8, 10-posltlons caused compounds to be much less
acutely toxic. The hexachlorobornane produced by removal of a chlorine at
the 6-pos1t1on from the heptachlorobornane was not much more toxic than the
heptachlorobornane Itself.
02050 VII-1 02/12/87
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Enzvmatlc Effects
Khalifa et al. (197tr) and Chandurkar and Matsumura (1979a) found that
toxaphcne and Toxicants A and B could be reductlvely dechlorlnated or dehy-
drochloMnated quickly or their vicinal chloride eliminated by liver mlcro-
somal preparations In vitro as well as nonenzymlcally by reduced hematln
(Khalifa et al., 1976). Clearly, certain toxaphene components will be
metabolized faster than others. |£ vitro metabolism of toxaphene by Iso-
lated mlcrosomal preparations from mice required NAOPH and relatively
anaerobic conditions (Khalifa et al., 1976). The fact that toxaphcne gave
type I binding spectra with the hepatic cytochrome P-450 of rats, mice,
sheep and rabbits Is suggestive that toxaphene components are substrates for
the hepatic mlcrosomal mixed-function oxldase system (Kulkarnl et al.,
1975). This Is also supported by the following: the toxlclty of toxaphene
can be potentiated by plperonyl butoxlde (Turner et al., 1977); toxaphene
causes shortened pentobarbUal sleeping times In rats (Schwabe and Wendllng,
1967; Trottman and Desalah, 1980); shortened N-methyl cyclohexenylymethyl
barbiturate sleeping times In rats have been measured (Ghazal, 1965);
transient decreases In (Peakall, 1976) and estrone (Welch et al., 1971)
metabolism; enhanced mlcrosomal enzyme activity (Peakall, 1976; Trottman and
Desalah, 1980; Pollock et al., 1983); Increases for pentobarbUal hydroxyla-
tlon (Trottman and Desalah, 1980), amlnopyrlne demethylase and aldrln epoxl-
datlon (Pollock et al., 1983) and the level of cytochrome P-450 (Pollock et
al., 1983) and NAOPH cytochrome c-reductase (Trottman and Desalah, 1980)
being dose-dependent. The "no significant hepatolnductlve effect" level for
1.p. administered toxaphene Is between 5 and 25 mg/kg bw for these enzymes
(Pollock et al., 1983).
02050 VII-2 02/25/87
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Saleh and Caslda (1978) showed that the major "products of Toxicant 8
produced by rat liver Tnlcrosomes under anaerobic conditions were CAS RN
57981-29-0 and 65620-64-6 In a 2:1 ratio; these products were not produced
under aerobic or anaerobic conditions 1f NADPH was absent. These products
were found directly 1n fat, liver and feces of adult male Sprague-Oawley
rats as well as CAS RN 64618-63-9 after a gavage dose of 3.1 mg Toxicant
B/kg bw. Chandurkar and Matsumura (1979a) reported that In addition to a
pathway requiring NAOPH, there appeared to be a pathway utilizing gluta-
thlone, as found by EC/GC product analysis and Inhibitor experiments using
3*C1- and 14C-toxaphene, and Toxicants B and C as substrates. In addi-
tion, the NADPH pathway metabolized Toxicant C much more rapidly than
Toxicant B. Glucuronldes, sulfonates and add hydrolysable conjugates were
also found. The addition of UDP-glucuron1c acid to toxaphene/NADPH system
Increased the extent of metabolism by 8-fold, but not when Sesamex (an
Inhibitor of NADPH) was also present, also showing probable Involvement of
the glucuronlc add metabolic pathway. The water soluble metabolites found
when glutathlone (GSH) was added were thought to be GSHconjugates formed
Involving loss of a chloride. Chandurkar and Matsumura (1979a) also Identi-
fied the major dechlorlnatlon product of Toxicant C to be CAS RN 70459-31-3,
and 4 secondary alcohols (hydroxyls at C2t C~, C^, C, positions) and
one primary alcohol. Oxidation products of Toxicant B were not alcohols.
Thus, different components In toxaphene, not surprisingly, are metabolized
differently. Ionic-chlorine Is the major excretion product. In both feces
and urine (Crowder and Dlndal, 1974; Ohsawa et al., 1975), and small amounts
of urine and fecal metabolites are glucuronldes or sulfate conjugates
(Chandurkar, 1977). The small amounts of conjugates may support the obser-
vation by Mehendale (1978) that toxaphene Inhibits hepatob111ary function,
at least In rats.
02050 VII-3 02/25/87
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Mehrle et al. (1979) proposed that the skeletal effects In birds could
be explained by a functional deficiency of vitamin C 1n the backbone result-
ing from shunting vitamin C to the liver to enable the organism to detoxify
toxaphene In the liver by mixed function oxldase activity, as Is also
proposed for the "broken back syndrome" caused by toxaphene for fish (IUPAC,
1979). Birds capable of synthesizing vitamin C 1n the liver (e.g., ravens,
sparrows) should not be so susceptible as those that cannot (e.g. ducks,
geese, quail, turkeys, doves, pigeons). This has not yet been shown experi-
mentally however.
DIPasquale (1977) showed that T5 mg toxaphene/kg bw/day (day 21 to day
35 of gestation) Increased collagen-containing structures In guinea pig
fetuses, and this was thought to be due to a functional deficiency of
vitamin C related to maternal mixed-function oxldase Induction.
How the vitamin C regulatory shunt Interacts with the hepatic NADPH and
glutathlone systems 1n the metabolism of toxaphene 1s still unknown, and It
1s unclear whether there 1s a link between events 1n the liver and those
Induced 1n the central nervous system.
Delchmann and KepHnger (1970) noted that pretreatment of rats with
aldrln or dleldrln raised 96-hour ID.g values 2-fold, and 3-fold after
pretreatment with p,p-DOT. Since these compounds are all known to Induce
hepatic mixed function oxldase activities, this may mean that compounds that
Induce this system will lower the acute toxldty of toxaphene. This was
also so when toxaphene was administered together with parathlon, dlazlnon or
02050 VII-4 02/12/87
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trlthion with which toxaphene Is often co-formulated (KepHnger and Dekh-
mann, 1967). The potenilatlon of toxaphene toxlclty with piperonyl butoxlde
(Turner et al., 1977; Saleh and Caslda, 1978) and perhaps the effect of
starvation (Boyd and Taylor, 1971; Srebocan et al., 1980a) may be mediated
through inhibition of the activity of mlcrosomal enzymes although this has
not been shown experimentally.
Sgbacute administration of toxaphene also appears to affect enzymatic
regulation of carbohydrate metabolism In young poultry (Srebocan et al.,
1978, 1980b). Thus, pyruvate:COp Upase (ADP) and GTP:oxaloacetate car-
boxylase (transphosphorylatlng) In cockerel livers were significantly
decreased by doses >5 mg/kg diet after 2 weeks of exposure. The authors
evoked adrenocortlcal mediation as the mechanism. The activities of beef
heart mltochondrlal sucdnoxldase and NAOH-ox1dase were Inhibited in vitro
t>y 330 yM toxaphene (Pardlnl et al., 1971). GuthMe et al. (1974)
reported that at 100 yMr the active transport of glucose through Isolated
mouse Intestine was Inhibited. The in vitro Inhibition of brain, kidney and
f*
liver Na/K-ATPases (IC5Q« 30 WM) and o11gomydnsens1t1ve (icco»
H
15 yM) and Insensitive Mg *-ATPase activities from male mice also Impli-
cates an Inhibition of membrane transport systems (Trottman and Oesalah,
1979, 1983; Fattah and Crowder, 1980). Subacute blood chemistry phenomena
Include: Increased serum add phosphatase, glutamlc pyruvlc transamlnase,
gamma-glutamyl transpeptldase activities, and Increased neutrophll counts
for dosed mice (Baumler, 1975). This behavior Is consistent with mild liver
pathology.
02050 VII-5 02/25/87
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The major enzymlc effect caused by chronic exposure appears to be unbal-
anced sugar metabolism.-At a dose to rats of 35 mg toxaphene/kg bw/day for
6 months (Gertlg and Nowaczyk, 1975), depressed (17-20%) LOH activities
occurred 1n the liver, kidney and serum, as well as decreased (50%) serum
alkaline phosphatase and liver glutamate dehydrogenase. Peakall (1979)
showed that levels of pyruvlc and lactic acids In blood plasma were not
affected even after 6 months at a dose of 2.4 mg toxaphene/kg bw/day and
postulated that the effects on the LOH system came from "rats stressed 1n
some way." However, Desalah et al. (1979) found dosedependent Increases In
liver mlcrosomal glucose-6-phosphatase and fructosel,6-d1phosphatase after 8
weeks of dosing Sprague-Oawley rats with doses of 0, 25, 50 and 75 mg toxa-
phene/kg diet. This was Indicative of changes In the process of gluconeo-
genesls. Alekhlna and Kuz'mlnskaya (1980) found that LOH activities 1n
myocardium Increased after 4 months of dosing "white rats" with 2.7 mg
toxaphene/kg bw/day, and In the liver only after both oral and percutaneous
exposure (9.4 mg/kg bw/day). Ishlkawa et al. (1978) reported that a single
l.p. treatments of 40 mg toxaphene/kg bw/day caused decreased plasma choles-
terol at day 60 with no effects on plasma trlglycerldes, liver weights,
mlcrosomal protein or cytochrome P-450 In old Sprague-Dawley rats. No dose-
related effects were observed on brain NaVKf-ATPases or ollgomycln-
sensltlve and -Insensitive Mg *-ATPases for doses of 0, 50, 100, 150 and
200 mg toxaphene/kg chow for 8 weeks to adult Sprague-Oawley rats, a 30-40%
decrease being observed for doses above 50 mg/kg diet. Since in vitro tests
showed dose-effect relationships, H was postulated that 1n vivo toxaphene
metabolized before It reached the brain. This was evident also for those
enzyme activities In the brain of male ICR rats dosed at 0. 10 25 and 50
mg/kg bw/day over 3 days, though the activities of kidney ATPases except for
02050 VII-6 02/26/87
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the ol1gomyc1n-sens1t1ve Mg f-ATPase were decreased In a dose-dependent
manner as was liver- ollgomycIn-sensitive Mg2*-ATPase (Trottman and
Desalah, 1979). Parvu et al. (1980) observed that 4 mg toxaphene/kg bw/day
for 30 days to Wlstar rats caused I1p1d accumulation In the Hver, depletion
of free fatty adds, and Increased liver cholesterol.
These data reveal that the major target organs for chronic toxldty
effects are, 1n order of decreasing Importance: liver (probably by Imbal-
ances 1n sugar metabolism and ATPase activity depression), kidney (probably
by kidney ATPases), heart (catecholamlne metabolism distortion; probable
ATPase effects), the central nervous system (catecholamlne metabolism
distortion; generalized tremors 1n chronically dosed Osborne-Hendel rats),
and the Immune system (premature aging effects In mice and rats). In birds,
the skeleton Is probably as Important a target organ as the liver (though
this may also occur In young mammals) because of the Inability of some birds
to synthesize vitamin C In the liver necessary for detoxifying the adminis-
tered toxaphene.
Hutagenlc Activity
Toxaphene, and specifically a polar subfractlon, Is a direct-acting
mutagen. It Is not clear from the literature what the Identities are of the
compounds 1n the polar subfractlon, or whether, like most of the nonpolar
components, they are metabolized so quickly that In fact metabolites are the
direct-acting mutagens. Clarification of this mechanism Is an Important
research need. Toxaphene has been found to be carcinogenic 1n two animal
species, Osborne-Mendel rats and B6C3F1 mice In studies described In
Tables V-8 to V-ll (NCI, 1979). The molecular events and the role of the
02050 VII-7 02/26/87
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polar fraction have not been formulated. While rats tend to show malignan-
cies of the endocrine system (the thyroid and the pituitary 1n females) mice
showed cancers only In the liver. This does suggest the link between the
liver and central nervous system could be by effects on the endocrine system.
Summary
Figure VII-1 summarizes the mechanisms underlying the acute to chronic
effects of toxaphene.
02050 VII-8 02/25/87
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f»t
t^Cftl
ton-polar cMponwu
*.|. TMlCMIt* M.C
I. C«trt1 iwovi tjtlM tffKU
t. (IdMf
1. lt»» 4
taut
I.
f«t«M»
*•
tak*-*.^
[ tlnUttloM c«n>n»U» j-
r»»t w
UkOl
llwr*
(rmlrw .
N«iO. »1U«1n C
Mm
teduert k«Mt1n
froducts M t r««
rtduetta* dtdil
M •
dtdilortattlM,
dilwld* «1talMt1on,
* l«i. kjrdr«jrlttt»t
NtU6o11Ui «ur««
«• f»t
k1l« flw k«t
jdviulv
ftft •llBlMtlW (UK-
lift* »1tt1« 1 M* 111
MMr»1 ip*rt trm 1« ktrdi).
Cklvld* 1» «J» ncrtttw
ulfttM vt i
T
S*«prt«
ATTutt (MdXMt
(rmturt mtM)
Oltttrt*
SMkdirwIc/dirwU tfTtcU
(§Ot1d»t1f1idJ_
Dtr«et-4CtlH
1.
Wt tff*ct«4 *.|. tfrv,
ktrt, e«i»4l iwe«t tyttn.
lyitM, kltfnty, MdecrlM ifiitt* (T)
l* t«IUB. AtUtOi
C«nc*rt AMTVM
nuiunr
f»ti)
tol«)
tolMtU i
ICMCV M t Mtol
tgCltt «tf <• M
FIGURE VII-1
Summary of Mechanisms Underlying Acute to Chronic Effects of Toxaphene
a!0 mg/kg diet for sheep, steers, cows; 4.74 ing/female ring-necked pheas-
ant/day; 12.5 mg/kg bw/day for female White Leghorns; 21 mg/kg diet for
rats, 2.4 mg/kg bw/day causes residue steady state 1n liver after 1 month
and 3 months for brain
bNo significant Inductive effect on affected liver enzymes 1s between 5
and 25 mg/kg bw for rats.
cl-3 mg/kg bw/day
02050
VII-9
07/26/84
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VIII. QUANTIFICATION OF TOXICOLOGICAL EFFECTS
Introduction
The quantification of toxicologlcal effects of a chemical consists of
separate assessments of noncarclnogenlc and carcinogenic health effects.
Chemicals that do not produce carcinogenic effects are believed to have a
threshold dose below which no adverse, noncarclnogenlc health effects occur,
while carcinogens are assumed to act without a threshold.
In the quantification of noncarclnogenlc effects, a Reference Dor
(RfD), [formerly termed the Acceptable Dally Intake (ADI)] 1s calculated.
The RfD 1s an estimate (with uncertainty spanning perhaps an order magni-
tude) of a dally exposure to the human population (Including sensitive
subgroups) that 1s likely to be without an appreciable risk of deleterious
health effects during a lifetime. The RfD 1s derived from a no-observed-
adverse-effect level (NOAEL), or lowest-observed-adverse-effect level
(LOAEL), Identified from a subchronlc or chronic study, and divided by an
uncertainty factor(s) times a modifying factor. The RfD 1s calculated as
follows:
IfD . '"0>a °r L°"L' «,A9
[Uncertainty Factor(s) x Modifying Factor]
Selection of the uncertainty factor to be employed In the calculation of
the RfD 1s based upon professional judgment, while considering the entire
data base of toxicologlcal effects for the chemical. In order to ensure
that uncertainty factors are selected and applied 1n a consistent manner.
02060 VIII-1 02/26/87
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the U.S. EPA (1986a) employs a modification to the guidelines proposed by
the National Academy of-Sc1ences (NAS, 1977, 1980) as follows:
Standard Uncertainty Factors (UFs)
Use a 10-fold factor when extrapolating from valid experimental
results from studies using prolonged exposure to average healthy
humans. This factor 1s Intended to account for the variation
1n sensitivity among the members of the human population. [10H]
Use an additional 10-fold factor when extrapolating from valid
results of long-term studies on experimental animals when
results of studies of human exposure are not available or are
Inadequate. This factor 1s Intended to account for the uncer-
tainty 1n extrapolating animal data to the case of humans.
[10A]
Use an additional 10-fold factor when extrapolating from less
than chronic results on experimental animals when there Is no
useful long-term human data. This factor 1s Intended to
account for the uncertainty In extrapolating from less than
chronic NOAELs to chronic NOAELs. [10S]
Use an additional 10-fold factor when deriving an RfD from a
LOAEL Instead of a NOAEL. This factor Is Intended to account
for the uncertainty In extrapolating from LOAELs to NOAELs.
[10L]
Modifying Factor (MF)
Use professional Judgment to determine another uncertainty
factor (MF) that Is greater than zero and less than or equal to
10. The magnitude of the MF depends upon the professional
assessment of scientific uncertainties of the study and data
base not explicitly treated above, e.g., the completeness of
the overall data base and the number of species tested. The
default value for the MF 1s 1.
The uncertainty factor used for a specific risk assessment Is based
principally upon scientific Judgment rather than scientific fact and
accounts for possible 1ntra- and Interspedes differences. Additional
considerations not Incorporated In the NAS/ODW guidelines for selection of
an uncertainty factor Include the use of a less than lifetime study for
deriving an RfD, the significance of the adverse health effects and the
counterbalancing of beneficial effects.
02060
VIII-2
02/26/87
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From the RfD, a Drinking Water Equivalent Level (DUEL) can be calcu-
lated. The DUEL represents a medium specific (I.e., drinking water)
lifetime exposure at which adverse, noncarclnogenlc health effects are not
anticipated to occur. The DUEL assumes 100% exposure from drinking water.
The DWEL provides the noncarclnogenlc health effects basis for establishing
a drinking water standard. For Ingestlon data, the DWEL 1s derived as
follows:
(RfD) x (Body weight In kg) /B
— — = mci/8.
Drinking Water Volume In i/day "
where:
Body weight = assumed to be 70 kg for an adult
Drinking water volume = assumed to be 2 i/day for an adult
In addition to the RfD and the DWEL, Health Advisories (HAs) for expo-
sures of shorter duration (1-day, 10-day and longer-term) are determined.
The HA values are used as Informal guidance to municipalities and other
organizations when emergency spills or contamination situations occur. The
HAs are calculated using an equation similar to the RfD and DWEL; however,
the NOAELs or LOAELs are Identified from acute or subchron'ic studies. The
HAs are derived as follows:
HA (NOAEL or
(UF) x ( i/day)
Using the above equation, the following drinking water HAs are developed
for noncarclnogenlc effects:
1. 1-day HA for a 10 kg child Ingesting 1 I water per day.
2. 10-day HA for a 10 kg child Ingesting 1 I water per day.
3. Longer-term HA for a 10 kg child Ingesting 1 l water per day.
4. Longer-term HA for a 70 kg adult Ingesting 2 l water per day.
02060 VIII-3 02/26/87
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The 1-day HA calculated for a 10 kg child assumes a single acute
exposure to the chemical and 1s generally derived from a study of <7 days
duration. The 10-day HA assumes a limited exposure period of 1-2 weeks and
Is generally derived from a study of <30 days duration. The longer-term HA
Is derived for both the 10 kg child and a 70 kg adult and assumes an
exposure period of ~7 years (or 10% of an Individual's lifetime). The
longer-term HA Is generally derived from a study of subchronlc duration
(exposure for 10X of animal's lifetime).
The U.S. EPA categorizes the carcinogenic potential of a chemical, based
on the overall we1ght-of-ev1dence, according to the following scheme:
Group A: Human Carcinogen. Sufficient evidence exists from
epidemiology studies to support a causal association between
exposure to the chemical and human cancer.
Group B: Probable Human Carcinogen. Sufficient evidence of
carclnogenlclty 1n animals with limited (Group 81) or Inade-
quate (Group 82) evidence 1n humans.
Group C: Possible Human Carcinogen. Limited evidence of
cardnogenldty 1n animals 1n the absence of human data.
Group 0: Not Classified as to Human Cardnogenldty. Inade-
quate human and animal evidence of cardnogenldty or for which
no data are available.
Group E: Evidence of Noncardnoqen1c1ty for Humans. No
evidence of cardnogenldty In at least two adequate animal
tests In different spedes or 1n both adequate ep1demtolog1c
and animal studies.
If toxlcologlcal evidence leads to the classification of the contaminant
as a known, probable or possible human carcinogen, mathematical models are
used to calculate the estimated excess cancer risk associated with the
Ingestlon of the contaminant In drinking water. The data used In these
02060 VIII-4 02/26/87
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estimates usually come from lifetime exposure studies using animal. In
order to predict the MsV for humans from animal data, animal doses must be
converted to equivalent human doses. This conversion Includes correction
for noncontlnuous exposure, less than lifetime studies and for differences
1n size. The .factor that compensates for the size difference Is the cube
root of the ratio of the animal and human body weights. It Is assumed that
the average adult human body weight 1s 70 kg and that the average water
consumption of an adult human 1s 2 l of water per day.
For contaminants with a carcinogenic potential, chemical levels are
correlated with a carcinogenic risk estimate by employing a cancer potency
(unit risk) value together with the assumption for lifetime exposure from
Ingestlon of water. The cancer unit risk 1s usually derived from a linear-
ized multistage model with a 95X upper confidence limit providing a low dose
estimate; that Is, the true risk to humans, while not Identifiable, Is not
likely to exceed the upper limit estimate and, 1n fact, may be lower.
Excess cancer risk estimates may also be calculated using other models such
as the one-hit, Welbull, loglt and probH. There Is little basis In the
current understanding of the biological mechanisms Involved 1n cancer to
suggest that any one of these models Is able to predict risk more accurately
than any other. Because each model Is based upon differing assumptions, the
estimates derived for each model can differ by several orders of magnitude.
The scientific data base used to calculate and support the setting of
cancer risk rate levels has an Inherent uncertainty that Is due to the
systematic and random errors In scientific measurement. In most cases, only
studies using experimental animals have been performed. Thus, there 1s
02060 VIII-5 02/26/87
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uncertainty when the data are extrapolated to humans. When developing
cancer risk rate levels,' several other areas of uncertainty exist, such as
the Incomplete knowledge concerning the health effects of contaminants In
drinking water, the Impact of the experimental animal's age, sex and
species, the nature of the target organ system(s) examined and the actual
rate of exposure of the Internal targets In experimental animals or humans.
Dose-response data usually are available only for high levels of exposure
and not for the lower levels of exposure closer to where a standard may be
set. When there 1s exposure to more than one contaminant, additional
uncertainty results from a lack of Information about possible synerglstlc or
antagonistic effects.
Noncardnoqenlc Effects
The effects of acute exposure to toxaphene, In humans as In animals,
Include salivation, hyperexc1tab1l1ty, behavioral changes, muscle spasms,
and convulsions, Indicating that the CNS 1s the critical target organ
system. In contrast, the liver and the Immune system appear to be the crit-
ical target sites In chronic or subchronlc exposure. Central nervous system
effects characteristic of acute Intoxication appeared during chronic expo-
sure only at dietary concentrations greatly 1n excess of those causing liver
damage In rats (Kennedy et a!., 1973).
Studies of the effects of long-term exposure of workers to toxaphene
have been reported. These are of little utility for standard development,
however, because exposure generally was poorly defined and often was to a
mixture of pesticides, and because no effort was made to correlate effects
with plasma or tissue levels of toxaphene or Us metabolites.
02060 VIII-6 02/26/87
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A few controlled subacute studies of human toxaphene exposure have been
reported. None of these-resulted In any observed toxldty. Application of
an aerosol spray containing toxaphene to the skin of 50 human subjects dally
for 30 days at a dose of 300 mg/day produced no toxic manifestations. Fifty
human volunteers who Inhaled 0.0004 mg/i of toxaphene aerosol for 10
minutes/day for 15 days had no subjective or objective effects. Twenty-five
humans Inhaling a mist containing 0.25 mg toxaphene/i of air for 30
minutes each day for 13 days showed no local or systemic toxic manifesta-
tions (Shelanskl, 1947).
Most acute studies to date have Involved attempts to define the LD,Q
for a variety of species. Lackey (1949a) also determined the threshold for
Initiation of convulsions In dogs. Single doses of toxaphene were admin-
istered In corn oil by stomach tube at doses varying from 5-50 mg/kg bw In 5
mg/kg .Increments. No convulsions or mortality occurred In three dogs dosed
with 5 mg/kg. Convulsions developed In 4/5 exposed to 10 mg/kg. Mortality
was first detected at the 15 mg/kg dose (2/8) with 6/8 developing convul-
sions. The LD5Q was estimated to be -25 mg/kg bw. If repeated doses of
toxaphene at 5 mg/kg bw were given, convulsions would develop regularly, but
only after several days. Four dogs dosed dally with 4 mg/kg bw, 2 dogs for
44 days and 2 for 106 days, survived although convulsions occurred occasion-
ally. In the subchronlc exposures, hydropic degeneration was seen In the
liver with degenerative changes 1n the tubular epithelium of the kidneys.
Olson et al. (1980) reported effects at much lower doses. Behavioral
effects were noted 1n offspring of rats given dally doses of 50 yg/kg from
day 5 of gestation until 70-90 days postpartum. Since the effects were gen-
erally transient and appeared to result from temporary delays In maturation,
02060 VIII-7 02/26/87
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It Is uncertain whether they can be classified as adverse. Secondly, since
the offspring were expoied In utero, by nursing and by toxaphene added to
the feed, 1t 1s uncertain what actual exposure levels were. Studies such as
those of Shaw (1947) Indicate that toxaphene may be concentrated 1n the milk.
There are a number of subchronlc and chronic studies of toxaphene tox-
Iclty 1n animals. Many of these document changes 1n activities of various
enzymes, such as lactate dehydrogenase Isozymes, serum alkaline phosphatase,
and tissue ATPases (Gertlg and Nowaczyk, 1975; Oesalah et al., 1979; Trott-
man and Desalah, 1979; Alekhlna and Kuz'mlnskaya, 1980). Although some of
these observations are suggestive of an effect on gluconeogenesls, they are
of doubtful significance since there 1s no direct proof that glucose metabo-
lism Is actually altered, at least In rats (Peakall, 1979).
There 1s some evidence that suggests that Upld metabolism may be
affected by toxaphene exposure. Ishlkawa et al. (1978) reported that a
single l.p. treatment of old Sprague-Oawley rats with 40 mg toxaphene/kg
bw/day resulted In a decrease In plasma cholesterol 60 days later with no
effect on plasma tMglycerldes or on liver weight. On the other hand, Parvu
et al. (1980) observed that administration of 4 mg/toxaphene/kg bw/day to
Ulstar rats for 30 days caused llpld accumulation In the liver, depletion of
free fatty adds, and an Increase In liver cholesterol.
Recently, Allen et al. (1983) concluded that the Immune system Is at
least as sensitive to the Ingestlon of toxaphene as the liver. Toxaphene
was added to the diet of female Swiss Webster mice at concentrations of 10,
100 and 200 ppm for 8 weeks. Among groups of 23-26 animals each, liver
weights were significantly (p<0.05) Increased 1n the 100 and 200 ppm groups.
02060 VI11-8 02/26/87
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Among groups of 10 animals each, IgG antibody tHers were also significantly
(p<0.05) decreased at the two higher doses. Cell-mediated Immune responses,
however, were not affected. Offspring of females fed toxaphone In the diet
from 3 weeks before mating until weaning were also tested at 8 weeks of age.
The cell-mediated Immune response was suppressed 1n the offspring of the 100
ppm group, while the phagocytlc ability of macrophages was significantly
reduced even In the 10 ppm group. The actual toxaphene dose cannot be
estimated accurately In the latter study since the offspring received
toxaphene transplacentally, 1n the milk and possibly even In the feed. It
does suggest, however, that neonates may be at greater risk than adults.
In most studies the critical target organ In rats and dogs chronically
exposed to toxaphene was the liver. At dietary concentrations of 100 ppm
toxaphene In the diet and above, all studies document some form of liver
pathology In- exposures lasting from 2 years (In rats and dogs) to lifetime
(In rats). Toxaphene at 25 ppm 1n the diet fed to rats over their lifetimes
caused an Increase 1n liver weight with minimal liver cell enlargement
(FUzhugh and Nelson, 1951). Over shorter periods of time (up to 12 weeks),
dietary toxaphene levels of up to 189 ppm had no measurable effects on male
or female albino rats (Clapp et al., 1971).
The NCI (1979) conducted a chronic study with Osborne-Mendel rats and
B6C3F1 mice to determine the possible cardnogenldty of toxaphene added to
the diet. Each treatment group consisted of 50 rats or mice of each gender,
and 10 untreated animals of each gender were matched controls with data from
40 or 45 untreated animals from similar bloassays pooled for statistical
evaluations.
02060 VIII-9 02/26/87
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Groups of 50 mice of each gender were administered toxaphene for 80
weeks and observed untTT sacrifice at 90-91 weeks (NCI, 1979). Low-dose
males and females received 160 ppm In the diet for 19 weeks followed by 80
ppm for 61 weeks; high-dose males and females received 320 ppm for 19 weeks
followed by 160 ppm for 61 weeks. The TWA doses were 99 or 198 ppm for both
males and females. Mean body weights of high-dose male mice were lower than
those of matched controls but weights of other dose groups were essentially
unaffected by toxaphene. Several animals died before week 19 when doses
were lowered. Following this, the dosed mice were generally comparable with
controls In appearance and behavior during the first year of the study.
During the second year, abdominal detention was observed In all dosed
groups, but predominantly 1n the high-dose males. Other clinical signs
Included alopecia, diarrhea, rough hair coats, and dyspnea. From weeks
60-76 the low-dose males appeared hyperexdtable. After week 75 there were
dose-related differences 1n survival.
Groups of 50 rats of each gender were administered toxaphene for 80
weeks (NCI, 1979) and then observed until survivors were sacrificed at
108-110 weeks (see Table V-7). Low-dose males were given 1280 ppm in the
diet for 2 weeks, 640 ppm for 53 weeks and 320 ppm for 25 weeks for a TWA of
556 ppm. High-dose males received 2560 ppm In the diet for 2 weeks, 1280
ppm for 53 weeks and 640 ppm for 25 weeks for a TWA of 1112 ppm. Low-dose
females received 640 ppm 1n the diet for 55 weeks and 320 ppm for 25 weeks
for a TWA of 540 ppm. High-dose females received 1280 ppm for 55 weeks and
640 ppm for 25 weeks for a TWA of 1080 ppm.
02060 VIII-10 02/26/87
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Mean body weights of the low- and high-dose female rats were lower than
those of the matched controls throughout most of the study, whereas weights
of low- and high-dose males were essentially unaffected. During the first
16 weeks, dosed males were generally comparable with controls in appearance
and behavior, with the exception of high-dose males that appeared hyper-
active during week 2 when the doses for male rats were lowered. At week 53,
the concentration of toxaphene In the diet was reduced because a majority of
the high-dose males and females developed generalized body tremors. Dose-
related decreases In survival rates were not observed. Clinical signs
usually associated with aging were observed earlier In dosed rats than in
controls: alopecia, diarrhea, dyspnea, pale mucous membranes, rough hair
coats, dermatitis, ataxla, leg paralysis, eplstaxls, hematurla, abdominal
distentlon and vaginal bleeding. Two females, one high-dose and one low-
dose, had Impaired equilibrium.
In summary, chronic feeding studies suggest that there are no marked
species differences In sensitivity among rats, mice and dogs, although mice
may be slightly more sensitive than rats to frank effects of toxaphene. On
chronic exposure, a dietary level of about 25 mg/kg diet appears to be a
NOAEL, and a dietary level of 40 mg/kg diet to be a LOAEL, with liver
pathology being the critical toxldty endpolnt. Chronic dietary levels
above 100 mg toxaphene/kg diet are likely to be associated with frank
effects.
02060 VIII-11 02/26/87
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Quantification of Noncardnoqenlc Effects
Derivation of 1-Dav HA. While kidney and liver pathology as well as
Immune effects are the critical endpolnts for chronic exposure to toxaphene,
the nervous system Is generally more sensitive to acute exposure. The
available data suggest a rather sharp threshold for nervous system effects.
Lackey (1949a) reported convulsions 1n dogs exposed dally to toxaphene at 5
mg/kg/day after several days, while 4 mg/kg/day Induced few convulsions even
with much longer exposures. No convulsions were Induced \r\ dogs after a
single dose of 5 mg/kg bw, while 10 mg/kg Induced convulsions 1n 4/5
animals. The Lackey (1949a) study Is the only one available that accurately
defines a NOAEL for very short-term exposures. Dogs appear to be more
sensitive to the acute toxic effects of toxaphene than other laboratory
species (see Table V-l) with reported t-D50s as 1ow as 20-25 mg/kg bw.
There 1s little evidence that humans are more sensitive to toxaphene than
dogs. IUPAC (1979) has estimated an oral L05Q dose 1n humans of 60 mg/kg
bw, while Conley (1952) estimated the acute lethal dose In humans to range
from 29-100 mg/kg. Finally, the nervous system 1s the critical endpolnt for
acute toxldty of toxaphene. There 1s no evidence from the Lackey (1949a)
study or from the variety of teratology studies conducted that very short-
term exposures at the threshold level for convulsions result 1n kidney or
liver pathology. Using the NOAEL from the Lackey (1949a) study, the 1-day
HA Is derived as follows:
1 day HA child = a °'5 mg/l
where:
5 mg/kg = NOAEL for a single oral exposure to dogs (Lackey, 1949a)
10 kg = weight of protected Individual (child)
02060 VIII-12 02/26/87
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100 = uncertainty factor appropriate for use with a NOAEL from
animal data and to protect sensitive members of the
human population
1 i/day = assumed volume of water consumed dally by a 10 kg child
It 1s recommended that 0.5 mg/i for children be accepted as the 1-day HA.
Derivation of 10-Day HA. The majority of subacute exposures have been
conducted to evaluate the reproductive or teratologlc effects of toxaphene.
Chernoff and Carver (1976) reported maternal toxldty of rats and mice at
all doses used (15-35 mg/kg) during exposure from days 7-16 of gestation.
Kavlock et al. (1982) found Increased total protein In fetuses from dams
exposed to 12.5 and 25 mg/kg/bw on days 7-16 of gestation In rats. Dally
doses of 12 mg/kg In rats on days 1-2 and 6-14 of gestation resulted In
smaller litters and decreased chollnesterase activity along myocardlal
vessels and cells of the atrloventMcular node (Badaeva, 1979, 1981).
Toxaphene administered by Intubation 1n doses of 4 mg/kg/bw to rats and
hamsters on days 7-11 or 1-15 of gestation had no effect upon development,
fetal weight, ratios of males to females, the number of fetal deaths or
maternal toxldty (Martson and Skepel'skaya 1980b). The NOAEL for maternal
toxldty as well as standard reproductive and teratologlcal parameters in
laboratory animals appears to be 4 mg/kg bw. Results of the Lackey (1949a)
study, however, suggested the NOAEL may be slightly lower In dogs than In
rodents. Minimal kidney and liver pathology were reported In dogs exposed
to 4 mg/kg bw/day for up to 44 days. It Is uncertain 1f these effects occur
with 10 days of exposure. Occasional convulsions were also noted at this
dose level.
02060 VI11-13 02/26/87
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Using a LOAEL of 4 mg/kg bw/day from the Lackey (1949a) study, the
10-day HA 1s derived as follows:
10-day HA child = 4 ""I71"* x 10 ^ - 0.04
1000 x 1 I
where:
4 mg/kg = LOAEL from oral exposure to dogs (Lackey, 1949a)
10 kg = weight of protected Individual (child)
1000 = uncertainty factor appropriate for use with a LOAEL from
animal data and to protect sensitive members of the
human population
1 l = assumed volume of water consumed dally by a 10 kg child
It Is recommended that the 10-day HA of 0.04 mg/l for children derived
from the Lackey (1949a) study be accepted.
Derivation of Longer-Term HA. There are no acceptable studies in the
available literature for the derivation of a longer-term HA.
Assessment of Lifetime Exposure and Derivation of a DMEL. There are
no acceptable studies 1n the available literature for the derivation of a
lifetime OWEI.
Carcinogenic Effects
Although several epidemiology studies have been conducted attempting to
correlate cancer Incidence with exposure to organochlorlne pesticides
(Barthel, 1976, 1981; El-Oesouky et al., 1978; Wang and Grufferman, 1981),
toxaphene was not thought to be a significant factor 1n the results of any
of these studies. In a survey of 199 employees who worked or had worked
02060 VIII-14 02/26/87
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with toxaphene between 1949 and 1977 with exposures ranging from 6 months to
26 years (mean 5.23 yejrs), 20 employees died, one with cancer of the
colon. None of the deaths could be attributed to toxaphene (IARC, 1979).
On the basis of the available Information, the evidence for carclnogenldty
of toxaphene 1n humans must be classified as Inconclusive.
Several Investigations of the carcinogenic potential of toxaphene have
been carried out 1n mice and rats. While Interpretations of the results of
these studies have been subject to challenge, the conclusion Is that chronic
exposure to toxaphene can Induce hepatocellular carcinoma In both mice and
rats and shows some evidence of carcinogenic activity at other sites as well.
The most thorough of these studies were performed by Tracor JHco Co.
under contract to the NCI (1979). The diets and the adjustments made In
toxaphene concentrations In the diets have been described above under non-
carcinogenic effects. Treatment was discontinued after 80 weeks and the
studies were terminated 10-11 weeks later (mice) or 28 weeks later (rats).
All animals that died during the studies and all animals surviving at the
termination of the studies were submitted for pathologic evaluation. Of the
male rats, 90% of the high-dose, 94% of the low-dose and all of the control
group lived until at least week 52 of the study. Of the female rats, 96% of
the high-dose, 92% of the low-dose and all 10 controls survived beyond the
52nd week. Although none of the tumors observed 1n treated animals were
uncommon for the animal strain used, certain tumors and hyperplastlc tissues
were present with higher Incidence 1n the treated animals. These Included
thyroid folUcular cell adenomas and carcinomas, and hyperplaslas. Thyroid
folUcular cell adenomas and carcinomas combined were found to be elevated
02060 VIII-15 04/02/87
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statistically In both male and female high-dose groups compared with either
concurrent or h1stor1caT~controls from the same laboratory (see Tables V-8
and V-9). In the female rats there was also a statistically significant
Increase 1n the cumulative Incidence of tumors of the pituitary (chromophobe
carcinomas or adenomas) In the high-dose compared with the control group.
The authors concluded that toxaphene administration was associated with an
Increase 1n thyroid tumor Incidence. However, It should be noted that the
adenomas Included 1n these evaluations are not classifiable as neoplasms.
Following an Independent examination of these hlstologlcal preparations,
Reuber (1979) also concluded that toxaphene administration was associated
with an Increase In tumor Incidence even greater than that reported by NCI
(1979). Since It 1s uncertain why his Interpretation of the tissues differs
from the original NCI Interpretation, 1t Is difficult to draw conclusions
from this study.
In the mice as In the rats, survival 1n treated groups was high, 90-98%
of treated and control animals surviving beyond the 52nd week of the study.
Of the tumors appearing In treated animals, none were observed In greater
Incidence compared with controls with the exception of those In the liver.
Hepatocellular carcinomas occurred In treated mice only, with Incidences of
98% and 69% In males (see Table V-10) at the high and low doses, and 69% and
10% 1n females (see Table V-ll) at the high and low doses, respectively.
These neoplasms were not observed In control animals of either sex, but
hepatic nodules were observed In 20% of the matched-control males, though
not In females. On the basis of these findings, the authors concluded that
toxaphene caused Increases In the Incidence of hepatocellular carcinomas,
02060 VIII-16 02/26/87
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and hence, was carcinogenic In B6C3F1 mice. Upon examining the tissues,
Reuber (1979) came to a "qualitatively similar conclusion, although again he
reported a greater tumor Incidence than NCI (1979).
In addition to the previous study, a number of other much less complete
studies on toxaphone cardnogenlcHy were conducted. In a study conducted
by the FDA (Nelson, 1949), groups of 12 male and 12 female rats assumed to
be of the Osborne-Mendel strain were fed 0, 25, 100, 400 or 1600 ppm toxa-
phene 1n the diet for 107 weeks. Further, groups of 5 male and 5 female
rats were fed 0, 40, 200 or 1000 ppm and then sacrificed after 56 weeks.
Tissues were sectioned only from the first set of groups; however, an Incom-
plete histology was performed on other groups. Among the second set of
animal groups killed after 56 weeks, only one neoplasm was noted, a subcu-
taneous neurosarcoma In a male rat given 1000 mg toxaphene/kg. although
liver hyperplasla and hyper.plastlc nodules were reported. Thyroid hyper-
plasla was also noted In treated animals, especially In males.
In the groups used In the 107-week experiment, 2/3 female and 2/2 male
rats Ingesting 1600 mg toxaphene/kg developed hepatic carcinomas; 1/3 female
rats 1n each of the 400 and 1600 mg/kg dose groups developed hyperplastk
nodules. No Incidence of either of the above tumors was observed In control
or other treatment groups.
Another study In mice demonstrated a statistically significant Increase
In hepatic neoplasms (LHton Blonetlcs, Inc., 1978). Fifty-four weanling
B6C3F1 mice were randomly assigned to each of four groups. Toxaphene
(X-16189-49), as well as the dose levels, were selected and provided by
02060 VIII-17 02/26/87
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Hercules Inc. The animals were administered toxaphene at 0, 7, 20 and 50
ppm levels In the diet -for 18 months and surviving mice were kept on the
control diet for another 6 months. All survivors were killed 24 months
after the Initiation of treatment. Table V-12 shows the effects of
toxaphene on survival. The survival 1n both treated and control groups was
similar.
The hlstopathologlcal evaluation data presented In Table V-13 shows a
statistically significant Incidence of hepatocellular tumors (hepatocellular
carcinomas and hepatocellular adenomas) 1n male mice fed 50 ppm toxaphene In
the diet as compared with the controls. However, hepatocellular tumor Inci-
dence was not statistically significant 1n females as compared with controls.
Taken together, the results of these studies (LHton Blonetlcs, 1978;
NCI, 1979) demonstrate that toxaphene, when administered chronically to mice
at moderate- to high-dose rates, Induces hepatocellular carcinomas in
Osborne-Mendel rats and 1n B6C3F1 mice, and may Induce tumors at other sites
as well.
In another LHton Blonetlcs (1979) study, toxaphene was administered to
50 weanling ARS Golden Syrian hamsters of each sex and each dose at levels
of 0, 100, 300 and 1000 ppm based on a subchronlc study. The hamsters,
obtained from the Sprague-Oawley laboratory 1n Madison, MI, were randomly
assigned to each group. The toxaphene was mixed In corn oil and blended
with the food that was available ad libitum. The animals were observed
during a 15-day acclimation period before Initiation of the study and dally
02060 VI11-18 02/26/87
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afterward for general condition and mortality Treatment was continued for
18 months for the females and 21.5 months for the males who showed a high
survival at 18 months.
Animals were observed until spontaneous death, unless moribund, or
surviving until termination of the experiment. Moribund animals and those
surviving until termination were sacrificed. All animals were necropsled.
The postmortem observations Included, a thorough external examination and
collection of major organs and representative tissues from all animals. All
collected tissues from control and high-dose animals, and target organs and
gross abnormalities from the low- and Intermediate-dose animals, were
evaluated hlstopathologlcally. Blood smears were taken from all animals at
sacrifice, but were not examined unless necessary for definition of specific
lesions or diseases. A compound-related lowered body weight was observed In
high-dose males until 12 months of study. Similar effects were noted in
males receiving 300 ppm from months 2-7 and for a brief period In females
receiving 1000 ppm. At terminal sacrifice, an Increase In liver weight was
observed In males at the 1000 ppm dose, which correlated to the pathologic
finding of megahepatocytes 1n 6 of 21 males observed. A decrease In heart
weight was noted In males at both the 300 and 1000 ppm dose levels. A
decrease 1n the thyroid gland weights of the 1000 ppm females was also noted.
Only the liver changes were supported by hlstopathology and were there-
fore Judged to be treatment-related. These liver changes are similar to
those accompanying an Induction of liver enzymes after administration of
chlorinated hydrocarbons and other compounds. Microscopic evaluation of the
tissues did not reveal any Incidence of a tumorlgenlc effect related to
02060 VI11-19 02/26/87
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toxaphene. Even when the Incidence of lymphoretlcular neoplasms of all
types was combined, no ^difference was detected between controls and dosed
animals. Under conditions of this study, toxaphene was not carcinogenic.
It Is likely that the maximum tolerated dose was not employed In these
studies (LHton B1onet1cs, Inc., 1979). In the subchronlc study, the
maximum tolerated dose estimate was based on evidence of liver toxldty
only. No hamster at any dose level died during the study; no changes In
general appearance, condition or behavior among experimental animals was
observed; and measurements of mean body weights and food consumption
Indicated no compound-related change. In the chronic study, a dose-related
decrease 1n body weight was observed, but H was temporary. The weight of
treated animals equaled or surpassed that of controls by the end of 1 year
for all treatment groups. Treated animals also survived for a longer period
than controls. A comparison of the number of animals In each treatment
group surviving to termination of the experiment using an X2 test for
homogeneity revealed significant differences for both males (p<0.05) and
females (p<0.001). '
Quantification of Carcinogenic Effects
Since the results of two bloassays (NCI, 1979; LHton B1onet1cs, Inc.,
1978) were positive for cancer Induction, estimated risk levels for toxa-
phene In drinking water can be calculated using a linearized multistage
model as discussed 1n the appendices to the October 1980 Federal Register
notice regarding the availability of Water Quality Criteria Documents
(Federal Register, 1980). Both studies are considered suitable for the
02060 VIII-20 02/26/87
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derivation of a criterion. The Litton 81onet1cs study was selected because
H allowed derivation of -a slightly more conservative criterion. The animal
data used 1n the derivation are as follows:
Dose Incidence of Hepatocellular Carcinoma In Male Mice
(mq/kq/dav) . (No. Responding/No. Tested)
0.0 10/53
0.91 10/54
2.6 12/53
6.5 18/51
Length of exposure = 18 months
Length of study » 24 months
Duration of Lifetime = 24 months
Body weight = 0.030 kg
With these parameters, the carcinogenic potency factor for humans q *
1s 1.131 (mg/kg/dayr1. The upper-limit unit risk estimates from the
animal data are derived from a linearized multistage nonthreshold extrapola-
tion model which 1s 'currently programmed as GLOBAL 83. Justification for
Us use Is presented 1n EPA's Guidelines for Carcinogen Risk Assessment
(U.S. EPA, 1986b). While recognizing that alternative statistical modeling
approaches exist [e.g., One-hH, Welbull, Log-ProbH, and LogH models, and
maximum likelihood estimates], the range of risks described by using any of
these modelling approaches has Uttle biological significance unless data
can be used to support the selection of one model over another. In the
Interest of approach consistency and of providing an upper bound estimate
for the potential cancer risk, the Agency recommends the use of the linear-
ized multistage model. EPA considers this model and resulting risk estimate
to be an upper limit value 1n the sense that the true risk Is unlikely to be
higher and may be lower even zero. An established procedure does not yet
02060 VIII-21 12/07/87
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exist for making "most likely" or "best" estimates of risk within the range
of uncertainty derived by the upper and lower limit values. In order to
derive a drinking water concentration of toxaphene calculated to keep Indi-
vidual risk below 10~5, but not taking Into account 1ngest1on of seafood
or Intake of toxaphene by other sources or other routes, the following
equation can be used.
Hater Concentration = 10 ' x 70 kq = 0.31 vg/i
1.131 mg/kg x 2 I
where:
10~5 = risk Level
70 kg = body weight of adult human
1.131 mg/kg = carcinogenic potency factor (human q-)*)
2 l = dally water Intake for an average adult
For risk levels of 10~4, 10~s and 10~6, the corresponding exposure
levels are 3.1, 0.31 and 0.031 vg/l, respectively.
In summary, toxaphene has been shown to Induce cancer 1n two species of
animals, while evidence for cancer Induction 1n humans Is Inconclusive.
Toxaphene 1s, therefore, classified as an animal carcinogen, IARC Category
2B ranking, meaning that toxaphene 1s probably carcinogenic In humans.
Applying the criteria described 1n EPA's guidelines for assessment of
carcinogenic risk (U.S. EPA, 1986b), toxaphene may be classified as Group
B2: probable human carcinogen, meaning there 1s Inadequate evidence from
human studies and sufficient evidence from animal studies.
02060 VIII-22 05/12/87
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Existing Guidelines. Recommendations and Standards
Standards for toxaphene 1n air, water and food have been established or
recommended by many groups. However, most of these standards were set
before the results of the NCI (1979) or LHton Blonetlcs (1978) bloassays of
toxaphene for cardnogenldty were available. On May 25, 1977, the U.S. EPA
Issued a notice of rebuttable presumption against registration and continued
registration of pestlddal products containing toxaphene (Federal Register,
1977). A further notice was Issued 1n the Federal Register (1982) where
registration for use under specified conditions was required after December
31, 1983 for scabies treatment of beef cattle and sheep. Depletion of
existing toxaphene stockpiles was allowed until December 31, 1986 under
limited conditions only. Its sale and distribution for use as an Insecti-
cide In no-till corn and dry and southern peas was also permitted until
December 31, 1986 (Federal Register, 1982).
The ACGIH (1977a) established a TWA value of 500 wg/m* for toxaphene
In the air of the working environment. The ACGIH (1977b) based this stan-
dard on unpublished acute and chronic toxlclty studies conducted 1n the
1950s and on comparisons of the toxlclty of toxaphene with DOT and llndane.
In addition, this group set a tentative short-term exposure limit for
toxaphene of 1.0 mg/m* (ACGIH. 1977a).
The national Interim primary drinking water standard for toxaphene Is 5
yg/l (U.S. EPA, 1976). This standard 1s based on the reported organo-
leptlc effects of toxaphene at concentrations >5 yg/l (Slgworth, 1965).
A safe level of 25 w9/i was also calculated based on minimal or no
effects 1n rats after they were fed toxaphene at a concentration of 10 mg/kg
02060 VIII-23 04/06/87
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In the diet, which was estimated to give an average dally dose of 1 mg/kg bw
(Lehman, 1965). This latter calculation used the following assumptions:
weight of rat = 300 g
dally food consumption of rat = 50 g
weight of average human adult = 70 kg
average dally water Intake for man = 2 I
safety factor = 500
dietary Intake = trace (assume zero)
From these assumptions, the maximum safe dally dose for humans was esti-
mated to be 3.4 yg/kg/bw (U.S. EPA, 1976). It should be noted, however,
that the assumption of 50 g dally food consumption for a 300 g rat 1s prob-
ably excessively high. The FDA standard of 5 yg/t for bottled water
(Federal Register, 1979) 1s Identical to the U.S. EPA's standard for drink-
Ing water.
The National Ambient Mater Quality Criterion for the protection of human
health Is based upon levels that may result In Incremental Increases In can-
cer risk over the lifetime. For risk levels of 10~5, 10'* and 10~7,
the corresponding criteria are 7.1, 0.71 and 0.07 yg/l, respectively
(U.S. EPA, 1980; Federal Register, 1980). If the above estimates are made
for consumption of aquatic organisms only, excluding consumption of water,
the levels are 7.3, 0.73 and 0.07 wg/l, respectively. The criterion was
derived from the development of hepatocellular carcinomas and neoplastk
nodules In B6C3F1 male mice given several doses of toxaphene (Litton
B1onet1cs, 1978).
02060 VIII-24 04/02/87
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The NRC (1977) estimated the RfO of toxaphene for man at 1.25
This was based on a study by FHzhugh and Nelson (1951), In which rats evi-
denced Increased liver weight and hepatic cell enlargement after exposure to
toxaphene at 25 ppm In the diet for 2 years. In their estimation NRC (1977)
assumed the dally dose 1n rats during the FHzhugh and Nelson (1951) study
was equivalent to 1.25 mg/kg/bw, and the application of a safety factor of
1000 was appropriate. Then, assuming a human body weight of 70 kg and a
dally water consumption of 2 l, NRC (1977) set the suggested NOAEL from
water at 8.75 yg/l (assigning 20% of the total RfO to water) or 0.44
(assigning IX of the total RfO to water).
The U.S. EPA criterion for protection of freshwater life was determined
to be 0.013 yg/l as a 24-hour average, and the concentration should not
exceed 1.6 vg/i at any time. For saltwater aquatic life the criterion
1s 0.019 vg/l as a 24-hour average with a maximum of 0.120 yg/l
(Federal Register, 1980; U.S. EPA, 1980).
The International Joint Commission of the United States and Canada
(1977) has recommended a water standard of 0.008 yg/l for the protection
of aquatic life. This standard 1s based on the study by Mayer and Mehrle
(1976) In which they found that toxaphene at 0.039 yg/i caused a signif-
icant Increase 1n mortality and a significant decrease 1n growth In brook
trout fry over a 90-day period. The standard of 0.008 yg/l 1s obtained
by applying a safety factor of 5.
02060 VIII-25 04/02/87
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Special Groups at Risk
Sensitive Suboooulatlon. Children may form a sensitive subpopulatlon
for the toxic effects of toxaphene. Most fatal cases of toxaphene poisoning
have Involved children (Hayes, 1982). It Is possible that this 1s due to a
greater Incidence of high level exposure such as 1ngest1on of toxaphene
solutions. However, there Is also some evidence for Increased suscepti-
bility 1n young animals. OffpsMng of rats fed only 50 wg/kg bw/day, from
3 weeks before mating until weaning age, showed delayed behavioral develop-
ment (Olson et al., 1980).
Inadequate diet may result 1n Increased susceptibility to toxaphene.
Central nervous system effects appeared earlier and at lower concentrations
1n rats fed a protein deficient diet (Boyd and Taylor, 1971). Toxaphene-
Induced Inhibition of enzymes Involved 1n carbohydrate metabolism was
greater In fasted than In fed chicks (Srebocan et al., 1980a).
Individuals with a less Indudble xenoblotlc metabolizing enzyme system
may constitute a sensitive subpopulatlon. The Importance of the cytochrome
P-450 mediated detoxification mechanism was Indicated by the 2- to 8-fold
Increase 1n the toxldty of Toxicant B 1n mice after treatment with the
cytochrome P-450 Inhibitor plperonyl butoxlde (Saleh et al., 1977; Turner et
al., 1977).
Since toxaphene 1s a complex mixture of mostly chlorobornanes, differing
solubilities may affect the actual concentration of the more toxic compo-
nents 1n drinking water. The overall solubility of toxaphene Is quite low.
02060 VIII-26 04/02/87
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Brooks (1974) reported a mean of -3 mg/l, while Paris et al. (1977)
reported a solubility of 0.5 mg/l at 25°C. Almost all of the toxlclty In
toxaphene 1s due to Toxicants A and B (Hayes, 1982). Toxicant A, the larger
of these two fractions. Is less polar than the entire mixture {Pollock and
Kllgore, 1980) Indicating Its solubllty 1n water 1s probably less than toxa-
phene as a whole. Toxicant B 1s more polar and thus possibly more soluble,
but makes up only -1% of the total volume. Thus, 1t 1s unlikely that dif-
fering solubilities will constitute a major factor In the toxlclty of toxa-
phene 1n water. However, since the polar fractions are mutagenlc (Hooper et
al., 1979) the more water soluble components may be Important 1f mutagenesls
Is related to cancer.
Toxaphene has been detected 1n drinking water; 27/58 samples contained
toxaphene with 2 samples at levels >0.050 ppb (U.S. EPA, 1977). Toxaphene
In drinking water has not been detected In some environmental surveys
(Schafer et al., 1969; Schulze et al., 1973; Mattraw, 1975), although It has
been detected (up to 0.41 ppb) 1n drinking water after toxaphene spraying
(Nicholson, 1964). Toxaphene In rain (up to 0.5 ppb) has also been detected
(Williams and Bldleman, 1978; Harder et al., 1980; Munson, 1976; Bldleman
and ChMstenson, 1979), and this exposure route may be Important where rain
water 1s utilized as drinking water.
Interactions. Induction of hepatic mlcrosomal MFO systems appears to
account for mosy of the Interactions of toxaphene with other compounds.
In rats pretreated with aldrln or dVeldrln and evidencing Increased liver
0-dealkylase and 0-demethylase activities, toxaphene 96-hour LD5Q values
were -2 times higher (Indicating decreased toxldty) than those of rats
02060 VIII-27 04/02/87
-------
given no pretreatment. Similarly, pretreatment with p.p'-DDT, a known
Inducer of hepatic mlcrosomal MFO, resulted 1n a 3-fold Increase In the
96-hour LD50 of toxaphene in rats (Delchmann and KepHnger, 1970).
When administered by oral Intubation to rats, equltoxlc combinations
(LD5Qs) of toxaphene with parathlon, dlazlnon or trlthlon were less toxic
than would be expected, based on the assumption of addHWe toxldty and
similar action (KepHnger and Delchmann, 1967).
Toxaphene 1s commonly formulated with methyl parathlon, usually In a 2:1
mixture, respectively, for control of certain Insect pests of cotton.
Chlordlmeform Is often added to the first formulation for additional
control. Combinations of these pesticides were given to male Swiss mice by
gastric Intubation; 25 mg toxaphene/kg bw 1n 0.16 ml corn oil; 12.5 mg
methyl parath1on/kg * 25 mg toxaphene/kg; 12.5 mg methyl parath1on/kg <• 25
mg toxaphene/kg * 3.25 mg chlord1meform/kg (Crowder and WhUson, 1980). In
these excretion-retention studies, mortality occurred only In groups receiv-
ing formulations containing methyl parathlon all within 3 hours of dosing.
It appeared that methyl parathlon toxldty was slightly enhanced by chlor-
dlmeform and slightly lowered by toxaphene but the differences were not
significant.
The possible Interaction of methyl parathlon and toxaphene was also
examined on the behavior of offspring of Sprague-Oawley rats exposed to
pesticides on days 7 through 15 of pregnancy (Crowder et al., 1980).
Pregnant rats were given 1.0 mg/kg bw methyl parathlon or 1.0 mg/kg methyl
parathlon + 2.0 mg/kg toxaphene by oral gavage 1n 0.1 ml corn oil.
02060 VIII-28 04/02/87
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Toxaphen* alone was given from day 7 until parturition at 6 mg/kg/day. Rat
pups exposed In utero to these doses of the pesticides alone or combined
demonstrated few significant changes 1n learning ability as measured by a
simple two-choice maze, motor skills or behavior. The only significant
change was In the additional time required (4.3 days) for methyl parathlon-
toxaphene treated Utters to develop the righting reflex.
Trlolo et al. (1982) Investigated 1n A/J mice the effects of Ingested
toxaphene on the Incidence of lung tumors Induced by oral administration of
benzo(a)pyrene (BaP). Groups of 9-week-old mice (7-48/group) were fed a
diet containing toxaphene (1n corn oil) at 100 ppm for 12 weeks or 200 ppm
diet for 20 weeks. Two doses of BaP (3 mg each) were given by the oral
route: the first BaP dose on day 7 after the Initiation of the toxaphene
diet, and the second dose on day 21. Nice were sacrificed after 12 or 20
weeks* and prepared for tumor analysis and analysis of BaP hydroxylase
activity. Mice that received toxaphene 1n the diet alone, or toxaphene and
BaP, showed an Increase 1n BaP hydroxylase activity In the liver and a
decrease 1n enzyme activity 1n the lung. Inhibition of lung BaP hydroxylase
activity was paralleled by a reduction In BaP-1nduced lung tumors In mice
fed toxaphene.
Over a period of 60 days Bogachuk and Fllenko (1978) studied the effects
of vertical vibration, toxaphene, and a combination of these physical and
chemical factors In 80 Immature Inbred male rats of the Hlstar and August
strains. Rats were divided Into four groups consisting of 10 rats of each
strain; group 1, control; group 2, vertical vibration (frequency 50 hertz,
amplitude 1.25 mm, 30 m1n/day); group 3, 0.01 L05Q toxaphene perorally
02060 VI11-29 04/02/87
-------
(L050 not given); group 4, combined action of vibration and toxaphene at
the same levels as In groups 2 and 3. Experiments were conducted on every
group at the same time each day for 60 days and covered the period of rapid
growth of the animals. Decreased body weight, kidney weight, volume, linear
dimensions and thickness of the cortex were observed 1n the three treated
groups. The changes were most pronounced 1n the group exposed to the
combination of toxaphene and vibration; the combined effects appeared to be
additive.
Mount et al. (1980) reported an outbreak of toxaphene poisoning 1n 10 of
15 crossbred sows 1n mldgestatlon. The animals ate feed that was poured on
the wet floor of a holding pen treated a month earlier with a commercial
spray containing 45X toxaphene and 2% Undane. Within 30 minutes, 10 of the
animals developed an acute nervous disorder. Two sows died within 2 hours
from the onset of clinical signs. In the others, severity of signs
diminished without treatment after 2.5 hours. Toxaphene and Undane levels
(mg/kg wet weight) found 1n the tissue were: cerebrum, 2.0 and 0.02; cere-
bellum, 4.0 and 0.03; and serum (hemolyzed), 0.3 and 0.002, respectively.
Although the levels of Undane 1n the tissues were below levels associated
with clinical Intoxication, the presence of Undane may have contributed to
the severity of the signs.
Starvation may cause greater mortality to toxaphene than optimal diets.
Thus, rats fed a protein-deficient diet showed acute oral LD.g values of
80+19 mg/kg bw, whereas for animals fed standard laboratory chow the LD5Q
was 220*33 mg/kg. The LD5Q of rats fed a high protein diet was 293*31 mg
toxaphene/kg bw, Indicating a protective effect of dietary protein. CNS
02060 VIII-30 • 04/02/87
-------
effects appeared earlier and at lower concentrations than In control rats
(Boyd and Taylor, 1971). Toxaphene (5 or 10 ppm In the diet), administered
to chicks after 24 hours without food, caused lower activities of glucose-6-
phosphatase, pyruvate carboxylase and phosphoenolpyruvate carboxyklnase 1n
1-day-old N1ck chick cockerels than did toxaphene or starvation alone
(Srebocan et al., 1980a). Heat stress In poultry did not appear to Interact
significantly with toxaphene (Srebocan et al., 19806). Based on the absence
of toxaphene-lnduced changes 1n plasma levels of pyruvlc and lactic adds In
unstressed rats, Peakall (1979) Implied that rats had to be "stressed In
some way" to obtain depressed blood LDH activity as claimed by Kuz'mlnskaya
and Alekhlna (1976) and Gertlg and Nowaczyk (1975).
The toxldty of Toxicant B to mice Is Increased by a factor of 2-8 by
administration of plperonyl butoxlde (Saleh et al., 1977; Turner et al.,
1977), Indicative of the Importance of cytochrome P-450 mediated detoxifica-
tion mechanisms. This would Indicate that any chemical that depressed or
enhanced mlcrosomal function might affect the ultimate toxldty to toxa-
phene. It has been postulated that 1n young white Leghorn chickens the
detoxification systems are not yet fully Induced, making younger animals
more susceptible than older ones (Bush et al., 1977). Thus, age may also be
an Interacting variable, a fact also noted during behavioral testing (Olson
et al., 1980).
02060 VIII-31 04/02/87
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