United States Office of Water EPA 440/5-80-040
Environmental Protection Regulations and Standards October 1980
Agency Criteria and Standards Division
Washington DC 20460
C.I
&EPA Ambient
Water Quality
Criteria for
Dichlorobenzidine
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AMBIENT WATER QUALITY CRITERIA FOR
DICHLOROBENZIDINE
Prepared By
U.S. ENVIRONMENTAL PROTECTION AGENCY
Office of Water Regulations and Standards
Criteria and Standards Division
Washington, D.C.
Office of Research and Development
Environmental Criteria and Assessment Office
Cincinnati, Ohio
Carcinogen Assessment Group
Washington, D.C.
Environmental Research Laboratories
Corvalis, Oregon
Duluth, Minnesota
Gulf Breeze, Florida
Narragansett, Rhode Island
Rcr ',:ccr'.v-*-1 P-otr.otton A^orcT
::,:;! GCXi
i
-------
DISCLAIMER
This report has been reviewed by the Environmental Criteria and
Assessment Office, U.S. Environmental Protection Agency, and approved
for publication. Mention of trade names or commercial products does not
constitute endorsement or recommendation for use.
AVAILABILITY NOTICE
This document is available to the public through the National
Technical Information Service, (NTIS), Springfield, Virginia 22161.
11
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FOREWORD
Section 304 (a)(l) of the Clean Water Act of 1977 (P.L. 95-217),
requires the Administrator of the Environmental Protection Agency to
publish criteria for water quality accurately reflecting the latest
scientific knowledge on the kind and extent of all identifiable effects
on health and welfare which may be expected from the presence of
pollutants in any body of water, including ground water. Proposed water
quality criteria for the 65 toxic pollutants listed under section 307
(a)(l) of the Clean Water Act were developed and a notice of their
availability was published for public comment on March 15, 1979 (44 FR
15926), July 25, 1979 (44 FR 43660), and October 1, 1979 (44 FR 56628).
This document is a revision of those proposed criteria based upon a
consideration of comments received from other Federal Agencies, State
agencies, special interest groups, and individual scientists. The
criteria contained in this document replace any previously published EPA
criteria for the 65 pollutants. This criterion document is also
published in satisifaction of paragraph 11 of the Settlement Agreement
in Natural Resources Defense Council, et. alI. vs. Train, 8 ERC 2120
(D.D.C. 1976), modified, 12 ERC 1833 (D.D.C. 1979).
The term "water quality criteria" is used in two sections of the
Clean Water Act, section 304 (a)(l) and section 303 (c)(2). The term has
a different program impact in each section. In section 304, the term
represents a non-regulatory, scientific assessment of ecological ef-
fects. The criteria presented in this publication are such scientific
assessments. Such water quality criteria associated with specific
stream uses when adopted as State water quality standards under section
303 become enforceable maximum acceptable levels of a pollutant in
ambient waters. The water quality criteria adopted in the State water
quality standards could have the same numerical limits as the criteria
developed under section 304. However, in many situations States may want
to adjust water quality criteria developed under section 304 to reflect
local environmental conditions and human exposure patterns before
incorporation into water quality standards. It is not until their
adoption as part of the State water quality standards that the criteria
become regulatory.
Guidelines to assist the States in the modification of criteria
presented in this document, in the development of water quality
standards, and in other water-related programs of this Agency, are being
developed by EPA.
STEVEN SCHATZOW
Deputy Assistant Administrator
Office of Water Regulations and Standards
111
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ACKNOWLEDGEMENTS
Aquatic Life Toxicology:
William A. Brungs, ERL-Narragansett
U.S. Environmental Protection Agency
David J. Hansen, ERL-Gulf Breeze
U.S. Environmental Protection Agency
Mammalian Toxicology and Human Health Effects:
Havish Sikka (author)
Syracuse Research Corporation
Steven D. Lutkenhoff (doc. mgr.)
ECAO-Cin
U.S. Environmental Protection Agency
Jerry F. Stara (doc. mgr.), ECAO-Cin
U.S. Environmental Protection Agency
Edward Calabrese
University of Massachusetts
Herbert Cornish
University of Michigan
Alfred Garvin
University of Cincinnati
Norman E. Kowal, HERL
U.S. Environmental Protection Agency
Larry K. Lowry
National Institute for Occupational
Safety and Health
Roy E. Albert*
Carcinogen Assessment Group
U.S. Environmental Protection Agency
H.T. Appleton
Syracuse Research Corporation
Douglas L. Arnold
Health and Welfare
Canada
Richard A. Carchman
Medical College of Virginia
Patrick Durkin
Syracuse Research Corporation
Ernest Foulkes
University of Cincinnati
Frank Gostomski
Criteria and Standards Division
U.S. Environmental Protection Agency
Roman W. Kuchuda
U.S. Environmental Protection Agency
Frank Stern
National Institute for Occupational
Safety and Health
Technical Support Services Staff: D.J. Reisman, M.A. Garlough, B.L. Zwayer,
P. A. Daunt, K.S. Edwards, T.A. Scandura, A.T. Pressley, C.A. Cooper,
M.M. Denessen.
Clerical Staff: C.A. Haynes, S.J. Faehr, L.A. Wade, D. Jones, B.J. Bordicks,
B.J. Quesnell,, T. Highland, R. Rubinstein.
*CAG Participating Members:
Elizabeth L. Anderson, Larry Anderson, Dolph Arnicar, Steven Bayard,
David L. Bayliss, Chao W. Chen, John R. Fowle III, Bernard Haberman,
Charalingayya Hiremath, Chang S. Lao, Robert McGaughy, Jeffrey Rosen-
blatt, Dharm V. Singh, and Todd W. Thorslund.
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TABLE OF CONTENTS
Criteria Summary
Introduction A-l
Aquatic Life Toxicology B-l
Introduction B-l
Effects B-l
Residues B-l
Criteria B-l
References B-3
Mammalian Toxicology and Human Health Effects C-l
Exposure C-l
Ingestion from Water C-l
Ingestion from Food C-l
Inhalation C-3
Dermal C-5
Pharmacokinetics C-5
Absorption C-5
Distribution C-5
Metabolism C-6
Excretion C-7
Effects C-9
Acute, Subacute and Chronic Toxicity C-9
Synergistic and/or Antagonistic Compounds C-10
Teratogenicity C-10
Mutagenicity C-ll
Carcinogenicity C-13
Summary C-21
Criterion Formulation C-22
Existing Guidelines and Standards C-22
Current Levels of Exposure and Special
Groups at Risk C-22
Basis and Derivation of Criterion C-23
References C-26
Appendix C-31
Summary and Conclusions Regarding the Carcinogenicity
of 3,3'-Dichlorobenzidine C-31
Summary of Pertinent Data C-33
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CRITERIA DOCUMENT
DICHLOROBENZIDINE
CRITERIA
Aquatic Life
The data base available for dichlorobenzidines and freshwater organisms
is limited to one test on bioconcentration of 3,3'-dichlorobenzidine, and no
statement can be made concerning acute or chronic toxicity.
No saltwater organisms have been tested with any dichlorobenzidine, and
no statement can be made concerning acute or chronic toxicity.
Human Health
For the maximum protection of human health from the potential
carcinogenic effects due to exposure of dichlorobenzidine through ingestion
of contaminated water and contaminated aquatic organisms, the ambient water
concentrations should be zero based on the non-threshold assumption for this
chemical. However, zero level may not be attainable at the present time.
Therefore, the levels which may result in incremental increase of cancer
risk over the lifetime are estimated at 10 , 10 , and 10 . The
corresponding recommended criteria are 0.103 yg/1, 0.010 pg/1, and 0.001
pg/1, respectively. If the above estimates are made for consumption of
aquatic organisms only, excluding consumption of water, the levels are 0.204
vg/1, 0.020 ug/1, and 0.002 yg/l, respectively.
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INTRODUCTION
Dichlorobenzidine (^,4'-d>amino-3,3'-dich'lorobipheny1 or 3,3'-dichloro-
benzidine) (DCB) is used in the production of dyes and pigments and as a
curing agent for polyurethanes. The molecular formula of dichlorobenzidine
is ci2H10C12N2 and the molecu1ar weight is 253.13 (Stecher, 1968).
DCB forms brownish needles with a melting point of 132 to 133"C (Pollock
and Stevens, 1965). It is readily soluble in alcohol, benzene, and glacial
acetic acid (Stecher, 1968), slightly soluble in HC1 (Radding, et al. 1975),
and sparingly soluble in water (0.7 g/1 at 15"C) (Stecher, 1968). When com-
bined with ferric chloride or bleaching powder, a green color is produced
(Pollock and Stevens, 1965).
The affinity of DCB for suspended particulates in water is not clear;
its basic nature suggests that it may be fairly tightly bound to humic mate-
rials in soils (Radding, et al. 1975). Soils may be moderate to long term
reservoirs.
Pyrolysis of DCB will most likely lead to the release of HC1. Because
of the halogen substitution, DCB compunds probably biodegrade at a slower
rate than benzidine alone. The photochemistry of DCB is not completely
known. DCB may photodegrade to benzidine (Sikka, et al. 1978).
Assuming the clean air concentrations of ozone (2 X 10~* M) and an
average atmospheric concentration of hydroxyl radicals (3 X 10~*5 M), the
half-life for oxidation of DCB by either of these chemical species is on the
order of one and one to 10 days, respectively. Furthermore, assuming a rep-
resentative concentration of 10 M for peroxy radicals in sunlit oxygen-
ated water, the half-life for oxidation by these species is approximately
100 days, given the variability of environmental conditons (Radding, et al.
1975).
A-l
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REFERENCES
Pollock, J.R.A. and R. Stevens (eds.) 1965. Dictionary of Organic Com-
pounds. Eyre and Spottiswoode, London.
Radding, S.B., et al. 1975. Review of the environmental fate of selected
chemicals. U.S. Environ. Prot. Agency, Washington, D.C.
Sikka, H.C., et al. 1978. Fate of 3,3'-dichlorobenzidine in aquatic envi-
ronments. EPA 600/3-8-068. U.S. Environ. Prot. Agency.
Stecher, P.G. (ed.) 1968. The Merck Index. 8th ed. Merck and Co., Rah-
way, New Jersey.
A-2
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Aquatic Life Toxicology*
INTRODUCTION
The data base for^dichlorobenzi dines and freshwater and saltwater or-
ganisms is limited to a bioconcentration and depuration study with the blue-
gill and 3,3'-dichlorobenz1dine (Appleton and Sikka, 1980).
EFFECTS
Residues
The apparent equilibrium bioconcentration factors for the bluegill dur-
ing tests of from 96 to 168 hours were from 114 to 170 for edible flesh and
495 to 507 for whole body (Table 1). An initial rapid rate of elimination
was followed by a low or negligible rate, with appreciable residues remain-
ing after 14 days in clean water.
CRITERIA
The data base available for dichlorobenzidines and freshwater organisms
is limited to one test on bioconcentration of 3,3'-dichlorobenzidine and no
statement can be made concerning acute or chronic toxicity.
No saltwater organisms have been tested with any dichlorobenzidine and
no statement can be made concerning acute or chronic toxicity.
*The reader is referred to the Guidelines for Deriving Water Quality Cri-
teria for the Protection of Aquatic Life and Its Uses in order to better un-
derstand the following discussion and recommendation. The following tables
contain the appropriate data that were found in the literature, and at the
bottom of each table are calculations for deriving various measures of tox-
icity as described in the Guidelines.
B-l
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Table 1. Residues for dlchlorobenzldlne (Appleton A Slkka, 1980}
Bloconcentratlon Duration
Species Tissue CheatcaI Factor (days}
FRESHWATER SPECIES
Bluegill, whole body 3,3'-dlchloro- 495-507 4-7
Lepomls macrochlrus benzldine
Bluegill, edible flesh 3.3»-dlchloro- 114-170 4-7
Lepomls macrochlrus benzldine
CO
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REFERENCES
Appleton, H.T. and H.C.iSikka. 1980. Accumulation, elimination, and metabol-
ism of dichlorobenzidine in the bluegill sunfish. Environ. Sci. Technol.
14: 50.
B-3
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Mammalian Toxicology and Human Health Effects
EXPOSURE
Ingestion from Water
To date, few systematic measurements of DCS in water supplies
have been undertaken. In one instance/ analysis of purge wells and
seepage water near a waste disposal lagoon receiving DCB-manufac-
ture wastes showed levels of DCS ranging from 0.13 to 0.27 mg/1.
High levels of benzidine (up to 2.5 mg/1) were also seen, which may
have arisen from photodegradation of DCS (Sikka, et al. 1978) ,
since benzidine is no longer manufactured in the U.S. Several
other dichlorobenzidine isomers were also detected at levels from 1
to 8 mg/1. The use of lagoons to handle DCB-containing wastes
might lead to contamination of ground water and pose a threat to
persons relying on nearby wells for drinking water.
Takemura, et al. (1965) analyzed the water of the Sumida River
in Tokyo during 1964. This river receives the waste effluents of
several dye and pigment factories. The presence of DCS was demon-
strated by thin layer chromatography. Although levels of DCS it-
self were not quantified, colorimetric analysis revealed that total
aromatic amine content of the water (including benzidine, dichloro-
benzidine, o<-naphthylamine, and ^-naphthylamine) reached levels
up to 0.562 mg/1. The authors suggested that the presence of the
free amines might be due to chemical reduction of the azo-dyes by
the high levels of H2S and S02 in the river.
Ingestion from Food
Few studies have attempted to identify DCS as a contaminant of
human food. Since DCB has never had an application as an agricul-
C-l
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tural or food chemical, the most likely source of dietary DCB would
be through consumption of contaminated fish.
A bioconcentration factor (BCF) relates the concentration of a
chemical in aquatic animals to the concentration in the water in
which they live. The steady-state BCFs for a lipid-soluble com-
pound in the tissues of various aquatic animals seem to be propor-
tional to the percent lipid in the tissue. Thus, the per capita
ingestion of a lipid-soluble chemical can be estimated from the per
capita consumption of fish and shellfish, the weighted average per-
cent lipids of consumed fish and shellfish, and a steady-state BCF
for the chemical.
Data from a recent survey on fish and shellfish consumption in
the United States were analyzed by SRI International (U.S. EPA,
1980). These data were used to estimate that the per capita con-
sumption of freshwater and estuarine fish and shellfish in the
United States is 6.5 g/day (Stephan, 1980). In addition, these
data were used with data on the fat content of the edible portion of
the same species to estimate that the weighted average percent
lipids for consumed freshwater and estuarine fish and shellfish is
3.0 percent.
A measured steady-state bioconcentration factor of 500 was
obtained for 3,3'-dichlorobenzidine using bluegills (Appleton and
Sikka, 1980). Since bluegills from another source contained an
average of 4.8 percent lipids (Johnson, 1980), these, bluegills
probably contained about the same percent lipids. An adjustment
factor of 3.0/4.8 = 0.625 can be used to adjust the measured BCF
from the 4.8 percent lipids of the bluegill to the 3.0 percent
C-2
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lipids that is the weighted average for consumed fish and shell-
fish. Thus, the weighted average bioconcentration factor for 3,3'-
dichlorobenzidine and ttfe edible portions of all freshwater and
estuarine aquatic organisms consumed by Americans is calculated to
be 500 x 0.625 - 312.
No DCS was detected in fish sampled from the vicinity of a
DCB-contaminated waste lagoon using analytical methods with sensi-
tivity of 10 to 100 yg/kg (G. Diachenko, personal communication).
Inhalation
The physical properties of DCB (low volatility/ large crystal
structure) probably minimize the risk of exposure of general popu-
lations to DCB through inhalation of air contaminated through in-
dustrial processes. However, inhalation might represent a major
source of occupational exposure under sub-optimum working condi-
tions. Akiyama (1970) examined the exposure of workers to DCB in a
pigment plant in Japan and determined that during the addition of
DCB to reaction vessels for synthesis of DCB pigments, the concen-
tration of DCB in air reached 2.5 mg/100 m in 10 minutes of charg-
ing of reaction vessels and decreased to 0.2 mg/100 m3 within 20
minutes. The distance of the sampling device from the operation
was not specified. Also, the amount of total aromatic amines was
elevated in exposed personnel (presumably due to the presence of
DCB) . The mean urinary concentrations of aromatic amines in process
workers charging the reaction vessels with DCB and plant laboratory
workers were 20.1 ppm and 21.1 ppm, respectively. Levels were only
14.5 ppm in workers who dried and cracked the pigments, 12.7 ppm in
office clerks, and 13.6 ppm in controls (medical students). Al-
C-3
-------
though the concentrations detected were highly variable (i.e., the
mean 20.1 ppra from the charging personnel was derived from data
ranging from 48.5 td 10 ppm), it is possible that the elevated
levels result from DCS exposure, since Akiyama claims that few or
no precautions were taken to prevent exposure, particularly on hot
days. It is uncertain whether the amines entered the body through
respiration or through dermal absorption.
Gerarde and Gerarde (1974) reported on an industrial process
in which both DCB and the DCB diarylide pigments were manufactured.
Most steps in the process were performed in closed systems, and the
DCB was handled in a salt form in a slurry (ca. 80 percent water
content). DCB dust was said not to be a problem. The possibility
that DCB contamination exposure could, however, occur is indicated
by the statement that "...the floor and accessible surfaces contam-
inated with the slurry were usually hosed down to prevent accumula-
tion of dried material...." Also, an outbreak of dermatitis in the
plant was attributed to a process change in DCB production. In
utilizing DCB in pigment production, the major sources of potential
exposure are listed as the weighing process and charging of the
tanks. Prior to May 1973, operators wore gloves and goggles but
not dust face masks. DCB was manufactured in this plant from 1938
to 1957. Thereafter, DCB was purchased from an outside supplier.
On-site inspection of three DCB utilizing plants showed that two of
the plants posed relatively low exposure potential which was due to
use of metal reactors and protective arrangements at the point of
tank charging. However, in the third plant, chemicals were dumped
into open reaction vessels from an elevated platform, posing an
C-4
-------
enhanced potential for exposure. Therefore, a great deal of varia-
bility concerning the exposure of individuals to DCS may exist
among various operations/
Dermal
Because of large particle size and increased usage of closed
systems and protective clothing, dermal absorption of DCB probably
represents a relatively minor route of DCB exposure in humans at
present. However, Meigs, et al. (1954) presented some experimental
evidence that under certain environmental conditions favoring moist
skin conditions, such as high relative humidity and high air tem-
perature, the dermal absorption by humans of benzidine and possibly
other congeners such as DCB may be enhanced.
PHARMACOKINETICS
Absorption
Virtually no information exists that quantifies the degree and
rate of absorption of DCB in experimental animals or in humans,
although Meigs, et al. (1954) detected DCB in the urine of DCB pro-
cessing and manufacturing workers.
Distribution
A detailed distribution study of DCB in rats, monkeys, and
dogs given 0.2 mg/kg of C-DCB by intravenous injection was re-
ported by Kellner, et al. (1973). The results indicate a rather
general distribution within the body after a 14-day observation
period with highest levels found in the livers of all three species.
The bile of monkeys and the lungs of dogs showed significant
levels of radio-activity.
C-5
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Metabolism
DCB metabolites have not been detected in the urine of dogs
jt
administered DCB orally or by intraperitoneal injection (Sciarini
and Meigs, 1961; Gerarde and Gerarde, 1974).
Kellner, et al. (1973) examined the urine of a Rhesus monkey
14
given 0.2 mg/kg C-DCB intravenously and found that in the first
14
four hours following injection, about one-third of the urinary C
was unchanged DCB, with another third identified as mono-N-acetyl
DCB, based on chromatographic properties. The remainder of the
14
urinary C was not recoverable via ether extraction at pH 11. At
later intervals, mostly metabolites were excreted, with nonextract-
14
able C comprising the majority of this material.
No ortho-hydroxy metabolites of DCB were detected in the urine
of human subjects after oral dosing (Gerarde and Gerarde, 1974).
Aksamitnaia (1959) reported that prolonged ingestion (7.5 to
8.5 months) of small doses or a single large dose of DCB in rats led
to the appearance of four transformation products, including benzi-
dine and possibly glucuronide conjugates. This conclusion may be
tenuous because analysis was done by paper chromatography (one sol-
vent system) without benefit of radiotracer techniques, and the
products were not quantified or further characterized. DCB was
never detected in the urine in any of the experiments; by-products
were seen only after seven months of chronic DCB ingestion.
In a study of the bioconcentration of DCB in bluegill sunfish,
over one-half of the DCB residues in the fish were in the form of a
conjugate which, under very'mildly acidic conditions, hydrolyzed to
reform free DCB (Sikka, et al. 1978).
C-6
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Hirai and Yasuhira (1972) noted that DCB was not oxidized by
cytochrome c, whereas benzidine and other derivatives were oxi-
dized.
The majority of information available at present suggests that
DCB is resistant to metabolism, with the exception of certain con-
jugative mechanisms and possibly certain bioactivation steps. Ring
chlorination of benzidine probably blocks ring hydroxylation reac-
tions of DCB for both electronic and steric reasons (Shriner, et
al. 1978).
Excretion
The excretion of DCB and metabolites following a 0.2 mg/kg
14
intravenous dose of C-DCB was studied by Kellner, et al. (1973)
in rats, dogs, and monkeys. With all species, measurable elimina-
tion had ceased within seven days of administration (Table 1) .
Fecal excretion was the predominant route of elimination in rats
and dogs, and possibly in monkeys.
Sciarini and Meigs (1961) also noted a preponderance of fecal
elimination of DCB in dogs. Finally, Gerarde and Gerarde (1974)
cite an unpublished study utilizing human volunteers which con-
cluded that DCB is excreted largely by the fecal route in man as
well as in dogs.
Insufficient data is available to assess the ability of the
body to accumulate significant burdens of DCB through repeated
exposures.
C-7
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TABLE 1
14 14
Excretion of C-DCB in Rats, Dogs and Monkeys Administered C-DCB*
Species
o Rat
CD
Dog
Monkey 1
Monkey 2
Interval
(day)
Urine
*
0-6 18+4
0-7 8+6
0-7 27
0-7 37
Elimination of Total
Feces
*
79 + 12
84 + 11
46
26
fc50
Phase IV
(hr)
45
--
--
--
Dose
Feed
*
1
5
21
20
Administered
Balance
0-7 day
98 + 12
97 + 8
94
83
Residues
%
2
3
*Source: Kellner, et al. 1973
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EFFECTS
Acute, Subacute, and Chronic Toxicity
Gaines and Nelso*n (19'77) reported the acute oral toxicity of
DCB to male and female mice. The LD^Q (mg/kg/day) for DCB given
daily for seven days was 352 for female mice (slope = 27.39) and
386 for male mice (slope = 23.15). The single dose LD50 (mg/kg)
was 488 for females and 676 for males.
Gerarde and Gerarde (1974) listed results of several toxico-
logical studies with DCB. DCB-dihydrochloride failed to produce
skin irritation in rabbits at an unspecified dose. An intradermal
dose of 700 mg/kg also gave a negative reaction. One hundred mg of
DCB-free amine placed in the conjunctival sac of the eye of a rab-
bit gave a negative reaction, while 20 mg of DCB dihydrochloride
produced erythema, pus, and opacity of the eye, giving a score of
84 of a possible 110 in one hour according to the method of Draize.
The oral LD5Q was given as 7.07 g/kg in albino rats for DCB free
amine, and 3.82 g/kg in male and female Sprague-Dawley rats for DCB
dihydrochloride. For topical application to skin, an LD_Q of 8
g/kg in male and female rats was seen. Pliss (1959) noted that rats
given 120 mg of DCB subcutaneously exhibited a state of excitation
with short-lived convulsions.
No human fatalities resulting from exposure to DCB have been
reported.
Ten rats exposed to a concentrated atmospheric dust of DCB
dihydrochloride for 14 days showed, upon autopsy, slight to moder-
ate pulmonary congestion and one pulmonary abcess (Gerarde and
Gerarde, 1974). An irritant effect from HC1 cannot be discounted
in the study.
C-9
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Freeman, et al. (1973) noted that DCB was cytotoxic to embry-
onic rat cells in culture at concentrations of 5 ppm or greater.
No mortalities tfere obtained in inhalation- studies where rats
were exposed to a concentrated atmosphere of concentrated DCB dihy-
drochloride dust for 14 days, or to 355 mg DCB free amine for 2
hours daily for 7 days (Gerarde and Gerarde,4 1974).
Gerarde and Gerarde (1974) listed the principal reasons for
visits to a company medical clinic by employees working with DCB.
These were as follows: (1) gastro-intestinal upset, (2) upper
respiratory infection, (3) sore throat, (4) caustic burns,
(5) headache, (6) dizziness, and (7) dermatitis. The only illness
apparently directly related to DCB was dermatitis. An outbreak of
dermatitis was attributed to a manufacturing process change which
led to small amounts of DCB-free base in the isolated DCB sulfate
salt. Two cases of acute cystitis were found in the medical record
review of the workers. One was of infectious origin and the other
related to the presence of renal calculi. Cystoscopic examination
of three other workers with urinary system symptoms revealed two
had renal calculi, and another had cystitis cystica.
Synergistic and/or Antagonistic Compounds
No data are available concerning compounds which synergize or
antagonize the toxicity of DCB.
Teratogenicity
No information is available defining the teratogenic potential
of DCB. While perhaps not directly relevant to the question of
DCB-induced teratogenesis, several studies summarized in the fol-
lowing discussion show that DCB can cross the placental barrier and
can also affect developmental systems.
C-10
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DCS has been demonstrated to significantly increase the inci-
dence of leukemia in the offspring of pregnant female mice given
comparatively low d6ses (ca. 8-10 mg) of DCB by subcutaneous injec-
tion in the last week of gestation (Golub, et al. 1974) . This could
have been due to postnatal transfer of DCB to the young through
lactation. However, transplacental effects of DCB have also been
observed. Shabad, et al. (1972) and Golub (1969) noted that kidney
tissue taken from embryos of pregnant female mice treated with DCB
exhibited altered behavior in organ culture, including increased
survival and hyperplastic changes in epithelium not seen in con-
trols.
The degree of exposure of pregnant women to DCB is probably
low. The work force involved in the manufacture and utilization of
DCB is predominantly or totally male. Maclntyre (1975) lists five
women, all between the ages of 20 and 34 years, as having been DCB
service or production workers in a plant in Great Britain. The
same area of the plant employed 217 men.
Mutagenicity
Garner, et al. (1975) compared the relative mutagenicity of
benzidine, DCB, and other analogs in the bacterial mutagenesis sys-
tem developed by Ames, et al. (1973) , utilizing the Salmonella
typhimurium tester strain TA1538, an indicator of frameshift muta-
genesis. The relevant data are summarized in Table 2. These re-
sults show that DCB is considerably more potent as a frameshift
mutagen in this system than is benzidine. Also, a low degree of
mutation is elicited by DCB.but not by benzidine in the absence of
the S-9 activation enzyme system.
C-ll
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TABLE 2
Mutagenicity of DCB in the Ames Assay*
Compound
3,3 '-Dichlorobenzidine
3,3 '-Dichlorobenzidine
Sulfate salt,
technical grade
Benzidine
Dimethyl sulfoxide
(control)
yg Chemical/ „ q#*
Plate s~9
50 +
100 +
50
100
50 +
100 +
50
100
50 +
100 +
50
100
+
Revertants/
Plate
3,360
7,520
114
131
5,490
8,350
127
129
430
640
5
15
16
8
*Source: Garner, et al. 1975
**S-9 is the NADPH-fortified rat liver activation enzyme preparation,
+ signifies preparation present; -, preparation absent.
C-12
-------
Similar observations were made by Lazear and Louis (1977),
utilizing an enzyme activation system obtained from the livers of
male mice and Ames tester strain TA98 (an indicator of frameshift
mutation). As before, DCB was much more mutagenic than benzidine
and, unlike benzidine, retained an appreciable mutagenic activity
without the liver enzymes. DCB was also slightly mutagenic towards
tester strain TA100, indicating base-pair substitution mutation,
Carcinogenicity
Stula, et al. (1975) maintained 50 male and 50 female rats on
a dietary level of DCB of 1,000 mg/kg. The average 50 percent sur-
vival was 356 days, with average days on the test of 349 days for
females and 353 days for males. The range of days on the test was
118-486 days for males and 143-488 days for females. The rats were
38 days old at the start of the assay and were apparently autopsied
at time of death or after 486-488 days (not specified) . The re-
sults of this study are listed in Table 3.
In addition to the cancers listed in Table 3, the occurrence
of malignant lymphoma was elevated over controls but not at statis-
tically significant (p«<0.05) levels. No bladder cancer was noted.
In a recent study, Stula, et al. (1978) reported on the induc-
tion of both papillary transitional cell carcinomas of the urinary
bladder and hepatic carcinomas in female beagle dogs. An oral dose
of 100 mg DCB was administered to the experimental animals, three
times per week for six weeks, then five times per week continuously
for periods up to 7.1 years. DCB was found to be carcinogenic at
statistically significant -levels (p<.025). The incidences of
hepatic carcinomas were 4/5 and 0/6 in DCB-treated and control
C-13
-------
TABLE 3
Induction of Cancer in Male and Female Rats
by 1,000 ppm Dietary DCBa
o
i
No. of Cancers
Type of Cancer
Maiqmary adenocarcinoma
Granulocytic leukemia
Zymbal's gland carcinoma
Male0
DCB
7b
9b
8b
Vehicle
control
0
2
0
Female0
DCB
26b
0
1
k-
Vehicle
control
%
3
0
0
aSource: Stula, et al. 1975
Significantly greater than controls at p<0.05
•
The number of animals examined histologically was 44 each for male and female.
-------
groups, respectively. The incidences of urinary bladder carcinomas
were 5/5 and 0/6, respectively (Table 4).
For 12 months,-* 6 times weekly, Pliss (1959) added 0.5 to 1.0
ml of a 4.4 percent suspension of DCB to the feed of rats of both
sexes of a strain assumed by Pliss to have a low spontaneous tumor
rate. Each rat received a total dose of 4.53 g. Neoplasms were
detected in 22 of 29 (75.8 percent) surviving animals. Tumors,
primarily carcinomas, were observed in a broad spectrum of organs
including mammary gland, Zymbal's gland (sebaceous gland of the
external auditory meatus), bladder, skin, small intestine, liver,
thyroid gland, kidney, hematopoietic (lymphatic) system, and sali-
vary glands.
An assay of DCB carcinogenicity was also done with mice
(Pliss, 1959). The mice received 0.1 ml of a 1.1 percent DCB sus-
pension in their food for 10 months, receiving a total dose of
127.5 to 135 mg DCB. Hepatic tumors were found in 4 of 18 mice
surviving after 18.5 months (22.2 percent). A sebaceous gland car-
cinoma and a lung adenoma were also seen.
The Pliss studies show that DCB may possess carcinogenic
activity in both rats and mice. However, the massive and apparent-
ly acutely-toxic dose levels employed, the uncertain purity of the
commercial product used, the virtual lack of dose-response data,
and the lack of adequate controls limit the studies' utility for
assessing human health hazards.
Carcinogenicity assays were also performed using rats and mice
which received DCB by subcutaneous injection (Pliss, 1959, 1963).
However, these studies are not considered here because of the
C-15
-------
Table 4.
Summary of Grpss^ Pathology and Microscopic Pathology
I
1
!
i
•
f.
f
r
1
U
SES S
*• H-.* *
ft* *M 4M«J 2
S o ~ at, "
I 2 u *'u I
3 b S ti S *
** C»OSS fATHOLOCV
*•* C 1.* Liven pale fatty appearance
451 t l.t Numerous grey firm nodule*, up to 2.J
cm. In liver, lung, urlnery bladder,
kidney, heart, lymph nodes, temporal
auscles. uterus, and gall bladder.
*•' S ' J.I One pink raised Irregular nodule on
urothellal surface of urinary bladder.
foci, up to 1.5 en, u liver.
*•• S 7.1 Six raised gray nodules, up to 2.S cm,
on urothellel surface of urinary
bladder; numerous palo brown foci, up
to 2.S cm. In liver.
J" S 7.1 Several raised gray nodules, up to 4.0
aa». on urothellal surface of urinary
bladder. Numerous nodules. up to 7.0
eo. In liver with adhesions to gall
bladder and pancreas.
**l S 7.1 Six raised fray nodules, up to 4.0 mm,
on urotliellal surface of urinary
bladder. Numerous gray, hemorrhaglc ,
cystic nodulis. up to 2.S cm. In liver.
*" 1 1.1 Hauaeryt 2 nodules () cm); liver 1 pale
nodule (2 sa»); lung! multiple nodule
(1 cm); spleen! 2 nndules it nm)
°M S (.0 Hannaryi nodule (2 cm); adrenal cortex!
nodule (i mm); liven 2 pale nodulee
(12 mm)
•00 S t.O Hanuryi nodule (t mm)] spleeni
liven pale firm streaks, nodular
'51 f t.O All sun>ary glands enlarged, contained
milk; spleeni nodule (10 m>); vaginal
one ralced nodule (2 ma); livari
. multiple pale noJules (l.i cm)
"° S 1.0 Mammary: nodule (S mm); vaginal several
firm nodules (2 mm)
Ml S t.O Ham»aryi 2 nodules (1 mm) firm; kldneyi
several gray nodules in cortex (2 m»)|
lung: diffuse areas of firmness: liven
seversl gray nodules (i mm)
1 •
Nul<- 1 Alt du)>s had |H>nrxlnMl jl disease wilh loss ol some leellt
2 1 IH- organs exan.mod hisloloKically included, brain, spinal < ord. heart, aorla. kinB. Irachca. ovary, ulc-rus. vaBina. esophagus stomach small
inluiiute. cecum. large intt-sline, bone marrow, spleen. Ihyrnus. liver, panireas. salivary gland, pituitary, thyioid. parathyroid! adrenal kidiu-y
eye. urinary bladder and all gross lesions.
Source: Stula, et al. 1978.
-------
irrelevancy of the subcutaneous route of administration to human
exposure.
Griswold, et al. (1968) examined the potency of cancer induc-
tion by DCS, benzidine, and other compounds, using induction of
mammary cancer in young female Sprague-Dawley rats as the major
index. Forty-day-old female Sprague-Dawley rats were given 30 mg
of DCB every three days for 30 days by gavage and were then observed
for nine months. Under the conditions of this assay, DCB was
ineffective as a mammary carcinogen but benzidine was highly effec-
tive at lower doses.
Sellakumar, et al. (1969) maintained male and female hamsters
for an unspecified length of time on a diet containing 0.1 percent
(1,000 ppm) of DCB. With 30 animals of each sex, no cancer was
observed. However, at 0.3 percent dietary DCB, four transitional
cell bladder carcinomas, some liver tumors, and diffuse chronic
intrahepatic obstructing cholangitus were seen. At 0.1 percent in
the diet of benzidine, many liver tumors were obtained but no blad-
der cancer was found.
DCB was also found to produce transformation in cultured rat
embryo cells infected with Rauscher leukemia virus (Freeman, et al.
1973). The index of transformation was the development of macro-
scopic foci of spindle cells, lacking polar orientation and contact
inhibition. Cells from typical foci were tumorigenic when trans-
planted into newborn Fisher rats, although this transplantability
was not quantitated. DCB-induced transformation was seen at a con-
centration of 5 ppm in the medium, but not at 1 ppm. Levels of 10
ppm or higher were cytotoxic. This in_ vitro test system detected
C-17
-------
transformation-activity in 6 of 7 aromatic amines characterized as
active in vivo carcinogens, 1 of 2 aromatic amines classed as weak
in vivo carcinogens, and K) of 3 aromatic amines classed as non-car-
cinogenic in vivo. DCB was classed by the authors as a weak car-
cinogen.
The history of human industrial experience with DCB has been
summarized and analyzed by Gerarde and Gerarde (1974) and Rye, et
al. (1970) in the United States; by Maclntyre (1975) and Gadian
(1975) in Great Britain; and by Akiyama (1970) in Japan. The con-
census of these authors, achieved through epidemiological studies,
is that there is no evidence that DCB itself has induced bladder
cancer, the characteristic lesion induced by benzidine, naphthyla-
mine, and other carcinogenic aromatic amines used in the dye and
pigment industry. The case for DCB carcinogenicity has been made
largely on the basis of its structural similarity to benzidine and
its tumorigenicity in several species of animals (Maclntyre, 1975).
One problem associated with epidemiological studies of DCB effects
in humans is that the population which has been exposed only to DCB
is small. Many workers have also handled benzidine or other car-
cinogens. Also, the characteristic latency period for induction of
bladder cancer by chemicals is quite long, exceeding 16 years for
benzidine (Haley, 1975), and may not have elapsed for many workers.
Finally, most of these studies have focused solely upon bladder
cancer as the disease of interest. As discussed below, this ap-
proach may be misleading and fallacious in view of the pattern of
DCB carcinogenesis in anima-ls and the nature of cancer observed in
DCB process workers.
C-18
-------
Gadian (1975) examined the health records of 59 workers at a
dyestuff plant in Great Britain who were exposed from 1953 through
1973 to DCB only and compared them to those working with both ben-
zidine and DCB, and to unexposed populations. This time was justi-
fied as the average latency period for chemically-induced bladder
cancer in humans (ca. 18 years) . It was calculated that the DCB
process worker was actually exposed to DCB for a maximum of 10
hours per work week. Men whose total DCB exposure was less than 245
hours (six months' full-time work) were excluded from the study,
leaving 35 segregated DCB workers. These 35 workers, representing
a total of 68,505 hours of DCB exposure, had no urinary tract
tumors, no other tumors, and two deaths from other causes (coronary
thrombosis, cerebral hemorrhage). In contrast, among 14 mixed ben-
zidine and DCB workers with 16,200 hours exposure (approximately 60
percent worked with benzidine, 40 percent worked with DCB), three
men developed tumors of the bladder, and one man developed carcino-
ma of the bronchus. One death from coronary thrombosis occurred.
Since the use of benzidine ceased in 1964, the mixed group had a
longer time to develop tumors than the DCB-segregated group.
Therefore, the DCB-alone hours worked during the same period (1953-
1964) as the mixed group was 31,945 hours. These results, while
admitting that the population studied was small, were taken as evi-
dence that DCB can be safely used if the provisions of the Carcino-
genic Substances Regulations are observed.
Maclntyre (1975) also surveyed the health history of a DCB-
utilizing plant in Great Britain. It was noted that the vast
majority (209 out of 217) of production and service workers had
C-19
-------
received first exposure to DCB less than 20 years before the time
of the report, indicating that the latent period for tumor forma-
tion might not have elapaed. Only 3 of the 217 exposed workers were
deceased. The causes of death were amytrophic lateral sclerosis
(age 55 years, 15 years of DCB exposure, 39 years since first ex-
posed) , carcinoma of the lung (age 61 years, one year of DCB expo-
sure, 12 years since first exposed), and pneumonia (age 70 years,
10 years of DCB exposure, 43 years since first exposed). Three
other employees who had not been exposed to DCB died of bronchial
carcinoma. All employees exposed to DCB since 1965 have received
cytological testing twice yearly, with all tests proving negative.
A 1974 meeting of occupational physicians is also cited, stating
that in Europe approximately 1,000 persons have been exposed to DCB
with a zero incidence of bladder cancer.
Gerarde and Gerarde (1974) reported the results of an epidemi-
ological study of workers exposed to DCB in manufacture and utili-
zation in a plant in the United States. A survey of the number of
DCB-exposed workers who developed neoplasms and the type of neo-
plasm was presented. These included lung cancer (2 workers), leu-
kemia-bone marrow (1) , lipoma (6) , rectum-papilloma (3) , sigmoid
colon carcinoma (2), prostate carcinoma (1), breast muscle myoblas-
toma (1), and skin basal cell epithelioma (1). A total of 17 work-
ers of the total of 207 workers surveyed had developed neoplasms.
The etiology of bladder cancer was discussed and the data
treated using several epidemiological and statistical approaches.
Accordingly, if DCB were as. potent as benzidine as a bladder car-
cinogen and the latent period long enough, a total of 22 cases of
C-20
-------
bladder cancer out of 163 DCS production workers would have been
observed, whereas none were seen. The possible induction by DC8 of
tumors at sites other than the bladder was not considered.
Summary
Based upon existing data, there is little doubt that DCB is
carcinogenic in several animal species including rats, mice, ham-
sters, and dogs. According to current methodology, the experi-
mental evidence serves as an indication that a potential carcino-
genic risk is posed to man. DCB induces tumors in a variety of
tissues in animals, with mammary, hematopoietic, and skin (Zymbal's
gland) tissue being the most affected. Many of the tumors have
been characterized as malignant.
C-21
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CRITERION FORMULATION
Existing Guidelines and Standards
The American Corffereruce of Governmental Industrial Hygienists
(ACGIH, 1977) has recommended that no exposure to DCS by any route
should be permitted, because of a demonstrated high carcinogenic
response in animals. Strict regulations have recently been promul-
gated by the Occupational Safety and Health Administration to mini-
mize or eliminate occupational exposure to DCB (29 CFR 1910). To
date, no standards have been placed on permissable levels of DCB in
the environment or in food.
Current Levels of Exposure and Special Groups at Risk
It is estimated that between 250 and 2,500 workers receive
exposure to DCB in the U.S., compared to 62 for benzidine (Fish-
bein, 1977). Given the stringent precautions which must be taken
in the manufacture and use of DCB, the level of exposure may be
minimal at present, although no data is available. However, past
exposure of individuals working without benefit of protective mea-
sures must present a cause for concern. In addition, the general
population may receive exposure to DCB through contaminated drink-
ing water or food (fish), although there is no significant evidence
for this at the present.
Additional groups that may be at risk include workers in the
printing or graphic arts professions handling the DCB-based azo
pigments. DCB may be present as an impurity in the pigments, and
there is very limited evidence to suggest that DCB may be metabo-
lically liberated from the azp pigment. More information is needed
on the levels of exposure to and metabolism of these pigments.
C-22
-------
Basis and Derivation of Criterion
The safe dose of DCS in water was calculated from the carcino-
genicity assays, using a*linearized multistage model described in
the Human Health Methodology Appendices to the October 1980 Federal
Register notice which announced the availability of this document.
The calculation assumes a risk of 1 in 100,000 of developing cancer
as a result of daily consumption of 2 liters of water and 6.5 g DCB-
contaminated fish or shellfish having a bioconcentration factor of
312. Although several carcinogenicity studies are available for
use in calculating a criterion for DCB in drinking water, only the
work of Stula and coworkers (1975, 1978) was considered, since the
studies by Pliss (1959, 1963) lack appropriate control data. More
specifically, the data on induction of hepatic carcinomas in female
beagle dogs (Stula, et al. 1978) were chosen as a base for the cal-
culation. Based on these data, a DCB criterion of 0.103 ug/1 is
judged to be adequate to protect the population consuming the
water. This dose is low from an occupational viewpoint and should
justify efforts to eliminate exposure of workers to DCB.
Under the Consent Decree in NRDC v. Train, criteria are to
state "recommended maximum permissible concentrations (including
where appropriate, zero) consistent with the protection of aquatic
organisms, human health, and recreational activities." DCB is sus-
pected of being a human carcinogen. Because there is no recognized
safe concentration for a human carcinogen, the recommended concen-
tration of DCB in water for maximum protection of human health is
zero.
C-23
-------
Because attaining a zero concentration level may be infeasible
in some cases and in order to assist the Agency and states in the
possible future development of water quality regulations, the con-
centrations of DCB corresponding to several incremental lifetime
cancer risk levels have been estimated. A cancer risk level pro-
vides an estimate of the additional incidence of cancer that may be
expected in an exposed population. A risk of 10~ for example,
indicates a probability of one additional case of cancer for every
100,000 people exposed, a risk of 10 indicates one additional
case of cancer for every million people exposed, and so forth.
In the Federal Register notice of availability of draft ambi-
ent water quality criteria, the U.S. EPA stated that it is consid-
ering setting criteria at an interim target risk level of 10" ,
10" , or 10~ as shown in the table below.
Exposure Assumptions Risk Levels and Corresponding Criteria(1)
(daily intake)_ - _
10~7 10"6 10"5
2 1 of drinking
water and consumption 0.001 ug/1 0.010 ug/1 0.103 ug/1
of 6.5 g of fish
and shellfish (2)
Consumption of fish 0.002 ug/1 0.02 ug/1 0.204 ug/1
and shellfish only.
(1) Calculated by applying a linearized multistage model as men-
tioned above. Appropriate bioassay data used in the calcula-
tion are presented in the Appendix. Since the extrapolation
model is linear at low doses, the additional lifetime risk is
directly proportional to the water concentration. Therefore,
water concentrations corresponding to other risk levels can be
derived by multiplying or dividing one of the risk levels and
C-24
-------
corresponding water concentrations shown in the table by fac-
tors such as 10, 100, 1,000, and so forth.
(2) Fifty percent o£ DCB exposure results from the consumption of
aquatic organisms which exhibit an average bioconcentration
potential of 312-fold. The remaining 50 percent of DCB expo-
sure results from drinking water.
Concentration levels were derived assuming a lifetime exposure
to various amounts of DCB, (1) occurring from the consumption of
both drinking water and aquatic life grown in water containing the
corresponding DCB concentrations and, (2) occurring solely from
consumption of aquatic life grown in the waters containing the cor-
responding DCB concentrations.
Although total exposure information for DCB is discussed and
an estimate of the contributions from other sources of exposure can
be made, this data will not be factored into the ambient water
quality criteria formulation because of the tenuous estimates. The
criteria presented, therefore, assume an incremental risk from
ambient water exposure only. Care must be taken to remember that
the proposed criterion is derived from animal experiments using
pure DCB. In the environment, DCB undergoes degradation to other
possibly toxic compounds such as benzidine. The possible addition-
al risk posed by these breakdown products should be considered in
the overall assessment of DCB.
C-25
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REFERENCES
Akiyama, T. 1970. ~*The investigation on the manufacturing plant of
organic pigment. Jieki. Med. Jour. 17: 1.
Aksamitnaia, I.A. 1959. Some data on transformation products of
3,3-dichlorobenzidine excreted with rat urine. Vopr. Onkol.
5: 18.
American Conference of Governmental Industrial Hygienists. 1977.
Documentation of the threshold limit values for substances in work-
room air. 3rd ed. Cincinnati, Ohio.
Ames, B.N., et al. 1973. Carcinogens are mutagens: A simple test
system combining liver homogenates for activation and bacteria for
detection. Proc. Natl. Acad. Sci. 70: 2281.
Appleton, H.T. and H.C. Sikka. 1980. Accumulation, elimination,
and metabolism of dichlorobenzidine in the bluegill sunfish. Envi-
ron. Sci. Technol. 14: 50.
Pishbein, L. 1977. Potential industrial carcinogens and mutagens.
Off. Toxic Subst., U.S. Environ. Prot. Agency, Washington, D.C.
Freeman, A.E., et al. 1973. Transformation of cell cultures as an
indication of the carcinogenic potential of chemicals. Jour. Natl.
Cancer Inst. 51: 799.
C-26
-------
Gadian, T. 1975. Carcinogens in industry, with special reference
to dichlorobenzidine. Chem. Ind. 19: 821.
Gaines, T.B. and C.J. Nelson. 1977. In; 5th Anniversary Report,
Natl. Center for Toxicol. Res. (FOA). 77: 1029.
Garner, R.C., et al. 1975. Testing of some benzidine analogues
for microsomal activation to bacterial mutagens. Cancer Lett.
1: 39.
Gerarde, H.W. and D.F. Gerarde. 1974. Industrial experience with
3,3'-dichlorobenzidine: An epidemiolgoical" study of a chemical
manufacturing plant. Jour. Occup. Med. 16: 322.
Golub, N.I. 1969. Transplacental action of 3,3'-dichlorobenzidine
and orthotolidine on organ cultures of embryonic mouse kidney tis-
sue. Bull. Exp. Biol. Med. 68: 1280.
Golub, N.I.r et al. 1974. Oncogenic action of some nitrogen com-
pounds on the progeny of experimental mice. Bull. Exp. Biol. Med.
78: 62.
Griswold, D.P., et al. 1968. The carcinogenicity of multiple
intragastric doses of aromatic and heterocyclic nitro or amino
derivatives in young female Sprague-Dawley rats. Cancer Res.
28: 924.
C-27
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Hirai, K. and Y. Yasuhira. 1972. Mitochondrial oxidation of 3,3'-
diaminobenzidine and related compounds, and their possible relation
to carcinogenisis. QANN. , 63: 665.
Johnson, K. 1980. Memorandum to D. Kuehl. U.S. EPA. March 10.
Kellner, H.M., et al. 1973. Animal studies on the kinetics of
benzidine and 3,3'-dichlorobenzidine. Arch. Toxicol. 31: 61.
Lazear, E.J. and S.C. Louis. 1977. Mutagenicity of some congeners
of benzidine in the Salmonella typhimurium assay system. Cancer
Lett. 4: 21.
Maclntyre, I. 1975. Experience of tumors in a British plant han-
dling 3,3'-dichlorobenzidine. Jour. Occup. Med. 17: 23.
Meigs, J.W., et al. 1954. Skin penetration by diamines of the ben-
zidine group. Arch. Ind. Hyg. Occup. Med. 9: 122.
Pliss, G.B. 1959. Dichlorobenzidine as a blastomogenic agent.
Vopr. Onkol. 5: 524.
Pliss, G.B. 1963. On some regular relationships between carcino-
genicity of aminodiphenyl derivatives and the structure of sub-
stance. Acta Unio. Int. Contra. Cancrum. 19: 499.
C-28
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Rye, W.A., et al. 1970. Facts and myths concerning aromatic dia-
mine curing agents. Jour. Occup. Med. 12: 211.
Sciarini, L.J. and J.W.'Meigs. 1961. Biotransformation of the
benzidines. III. Studies on diorthotolidine, dianisidine, and
dichlorobenzidine: 3,3'-disubstituted congeners of benzidine
(4,4'-diamino-biphenyl). Arch. Environ. Health. 2: 108.
Sellakumar, A.R., et al. 1969. Aromatic amines carcinogenicity in
hamsters. Proc. Am. Assoc. Cancer Res. 10: 78.
Shabad, L.M., et al. 1972. Transplacental effects of some chemi-
cal compounds on organ cultures of embryonic kidney tissue. Cancer
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Shriner, C.R., et al. 1978. Reviews of the environmental effects
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U.S. Environ. Prot. Agency, Washington, B.C.
Sikka, H.C., et al. 1978. Fate of 3,3'-dichlorobenzidine in
aquatic environments. U.S. Environ. Prot. Agency, EPA 600/3-8-068.
Stephan, C.E. 1980. Memorandum to J. Stara. U.S. EPA. July 3.
Stula, E.F., et al. 1975. Experimental neoplasia in rats from
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C-29
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Stula, E.F., et al. 1978. Liver and urinary bladder tumors in dogs
from 3,3'-dichlorobenzidine. Jour. Environ. Pathol. Toxicol.
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Takemura, N. , et al. 1965. A survey of the pollution of the Sumida
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C-30
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APPENDIX
Summary and Conclusions Regarding the
Carcinogenicity of 3,3'-Dichlorobenzidine (DCS)*
-*
3,3'-Dichlorobenzidine (DCB) is used as an intermediate in the
synthesis of dyes and pigments. It is structurally related to car-
cinogenic aromatic amines, which have been used in the dye and pig-
ment industries.
Five epidemiological studies of employees handling DCB in
chemical plants in the United States, Great Britain, and Japan have
provided no evidence of DCB-induced cancers. However, investiga-
tive problems associated with these studies, such as too short a
follow-up time and small sample size, make them unreliable as the
sole basis for making conclusions about human cancer risks from
DCB.
DCB has induced carcinomas in three species of experimental
animals receiving oral doses of the chemical. Dogs (female) devel-
oped papillary transitional cell carcinomas of the urinary bladder
and hepatocellular carcinomas. Hamsters developed transitional
cell bladder carcinomas, liver cell, and cholangiomatous tumors.
Rats developed mammary adenocarcinomas (male and female), granulo-
cytic leukemia (males), and Zymbal's gland carcinomas (males).
Two studies of the mutagenicity of DCB showed that it was
mutagenic in two Salmonella typhimurium tester strains (TA1538,
TA98) in the presence and absence of an S-9 liver enzyme system.
DCB also transformed cultured rat embryo cells, and the transformed
cells were tumorigenic when transplanted into newborn rats.
*This summary has been prepared and approved by the Carcinogens
Assessment Group, U.S. EPA, on June 15, 1979.
C-31
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The carcinogenic, mutagenic, and transforming activities of
DCB in laboratory organisms and its chemical similarity to benzi-
dine, a human bladder carcinogen, are strong evidence that it is
likely to be a human carcinogen.
The water quality criterion for DCB is based on the induction
of hepatic carcinomas in female beagle dogs, given an oral dose of
100 mg 3,3'-dichlorobenzidine, three times per week for six weeks,
then five times per week continuously for up to 7.1 years (Stula,
et al. 1978). The concentration of DCB in water, calculated to
keep the lifetime cancer risk below 10" , is 0.103 ug/1.
C-32
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Summary of Pertinent Data
The water quality criterion for DCB is based on the induction
of hepatic carcinomas in female beagle dogs, given an oral dose of
100 mg DCB, three times per week for six weeks, then five times per
week continuously for periods up to 7.1 years (Stula, et al. 1978).
The criterion was calculated from the following parameters:
Dose Incidence
(mg/kg/day) (No. responding/No, tested)
0 0/6
7.36 4/5
le = 2,593 days w = 11.391 kg
Le = 2,593 days R = 312 I/kg
L = 3,159 days
With these parameters the carcinogenic potency factor for humans,
q,*, is 1.692 (mg/kg/day)~ . The resulting water concentration for
DCB, calculated to keep the individual lifetime cancer risk below
10~5, is 0.103 yg/1.
C-3 3 * U. S GOVERNMENT PRINTING OFFICE : 1980 720-016/4362
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