DIPHENYLHYDRAZINE
Ambient Water Quality Criteria
Criteria and Standards Division
Office of Water Planning and Standards
U.S. Environmental Protection Agency
Washington, D.C.
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CRITERION DOCUMENT
1,2-DIPHENYLHYDRAZINE
CRITERIA
Aquatic Life
For 1,2-diphenylhydrazine the criterion to protect freshwater
aquatic life as derived using the Guidelines is 17 ug/1 as a 24-
hour average and the concentration should not exceed 38 ug/1 at
any time.
For saltwater aquatic life, no criterion for 1,2-diphenyl-
hydrazine can be derived using the Guidelines, and there are
insufficient data to estimate a criterion using other procedures.
Human Health
For the maximum protection of human health from the potential
carcinogenic effects of exposure to 1,2-diphenylhydrazine through
ingestion of water and contaminated aquatic organisms, the ambient
water concentration is zero. Concentrations of 1,2-diphenylhydra-
zine estimated to result in additional lifetime cancer risks rang-
ing from no additional risk to an additional risk of 1 in 100,000
i
are presented in the Criterion Formulation section of this docu-
ment. The Agency is considering setting criteria at an interim
target risk level in the range of 10~5, 10~~6, or 10"? with cor-
responding criteria of 0.4 ug/1/ 0.04 ug/1 and 0.004 ug/1/ re-
spectively.
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Introduction
_^MHMH^»WB«^_^«V^^BB ^
Diphenylhydrazine exists as an assymmetrical isomer, 1,1-
diphenylhydrazine, and a symmetrical isomer, 1,2-diphenylhy-
drazine (hydrazobenzene). The hydrochloride of 1,1-diphenyl-
hydrazine is used as a reagent for the sugars, arabinose and
lactose (Windholtz, 1976). 1,2-Diphenylhydrazine is used in
organic synthesis (Bennett, 1974) and has a major use as the
starting material in the production of benzidine, for use in
dyes (Lurie, 1964).
No data were found on the environmental presence or per-
sistence of diphenylhydrazines, except for one report of de-
tection in drinking water at a concentration of 1 ug/1 (U.S.
EPA, 1975). 1,1- and 1,2-diphenylhydrazines have been char-
acterized as slightly soluble and insoluble in water, respec-
tively (Windholtz, 1976; Bennett, 1974). No quantitative data
were found for the water solubilities and vapor pressures of
these compounds; consequently, no predictions can be made
about their presistence in water,.
Since no data are available on levels of diphenylhydra-
zines in surface waters, ambient water levels cannot be com-
pared to the levels of these chemicals determined to be toxic
in laboratory studies. Experimental data indicate that 1,2-
diphenylhydrazine is toxic to freshwater aquatic organisms and
reported LC50 values range from 0.27 to 4.1 mg/1 (U.S. EPA,
1978). Rats and mice developed a variety of malignancies
after dermal exposures to 1,2-diphenylhydrazine (Spitz, et al.
1950; Pliss, 1974). Oral doses of 1,2-diphenylhydrazine did
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not produce malignancies (Marhold, et al. 1968). The docu-
mented carcinogenicity of 1,2-diphenylhydrazine is of primary
concern to industrial workers, who are exposed to higher con-
centrations than other segments of the population.
Although there are few data describing the fate of di-
phenylhydrazines in water, the carcinogenic effects of 1,2-di-
phenylhydrazine warrant its regulation, to protect humans and
aquatic organisms from possible water-related hazards associ-
ated with this chemical.
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REFERENCES
Bennett, H., ed. 1974. Concise chemical and technical dic-
tionary. Chemical Publishing Co. Inc., New York.
Keinath, T.M. 1976. Benzidine: Wastewater treatment tech-
nology. Prepared for Off. Water Plann. Stand. U.S. Environ.
Prot. Agency.
Lurie, A.P. 1964. Benzidine. P. 408. Ijn Kirk-Othmer Ency-
clopedia of Chemical Technology. 2nd ed. Vol. 3.
Marhold, J., et al. 1968. The possible complexity of di-
phenyline in the origin of tumors in the manufacture of ben-
zidine. Neoplasma 15: 3.
Pliss, G. 1974. Carcinogenic properties of hydrazobenzene.
Vop. Onkol. 20: 53.
Spitz, S., et al. 1950. The carcinogenic action of benzi-
dine. Cancer Sept: 789.
U.S. EPA. 1975. Preliminary assessment of suspected carcino-
gens in drinking water. Off. Tox. Subs. Washington, D.C.
U.S. EPA. 1978. In-depth studies on health and environmental
impacts of selected water pollutants. Contract No. 68-01- .
4646.
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Windholtz, M., ed. 1976. The Merck Index. Merck and Co,
Inc., Rahway, N.J.
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AQUATIC LIFE TOXICOLOGY*
FRESHWATER ORGANISMS
Introduction
Toxicity tests with the bluegill and Daphnia magna have been
conducted using static procedures, and the results demonstrated
4
adverse effects as low as 251 ug/1.
Acute Toxicity
The unadjusted 96-hour LC50 for the bluegill is 270 ug/1
(Table 1), and when this concentration is adjusted for testing
' V
procedures and species sensitivity a Final Fish Acute Value of 38
ug/1 is obtained.
Daphnia magna is less sensitive with an unadjusted 48-hour
EC50 of 4,100 ug/1 (Table 2). The resultant Final Invertebrate
j
Acute Value of 170 ug/1 is higher than that value for fish, so the
latter, 38 ug/1, becomes the Final Acute Value.
*The reader is referred to the Guidelines for Deriving Water
;
Quality Criteria for the Protection of Aquatic Life [43 FR 21506
(May 18, 1978) and 43 FR 29028 (July 5, 1978)] in order to better
understand 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 the calcula-
tions for deriving various measures of toxicity as described in
the Guidelines.
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Chronic Toxicity
The chronic effects of 1,2-diphenylhydrazine on Daphnia magna
have been determined (U.S. EPA, 1978) and adverse effects were
observed at 420 ug/1 (Table 3). No detectable effects occurred ,at
150 ug/1. The latter concentration is 0.037 of the 48-hour EC50
(Table 2). A Final Invertebrate Chronic Value of 49 ug/1 is
obtained from these results and, since there are no data on
chronic toxicity to fish, plant effects, or Residue Limited
Toxicant Concentration, this concentration also becomes the Final
Chronic Value.
Residues
No measured steady-state bioconcentration factor (BCF) is
available for 1,2-diphenylhydrazine. A BCF can be estimated using
the octanol-water coefficient of 870. This coefficient is used to
derive an estimated BCF of 100 for aquatic organisms that contain
about 8 percent lipids. If it is known that the diet of the
wildlife of concern contains a significantly different lipid
content, an appropriate adjustment in the estimated BCF should be
X
made.
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CRITERION FORMULATION
Freshwater-Aquatic Life
Summary of Available Data
The concentrations below have been rounded to two significant
figures.
Final Fish Acute Value = 38 ug/1
Final Invertebrate Acute Value = 170 ug/1
Final Acute Value = 38 ug/1
Final Fish Chronic Value = not available
Final Invertebrate Chronic Value = 49 ug/1
Final Plant Value - not available
Residue Limited Toxicant Concentration » not available
Final Chronic Value = 49 ug/1
0.44 x Final Acute Value = 17 ug/1
The maximum concentration of 1,2-diphenylhydrazine is the
Final Acute Value of 38 ug/1 and the 24-hour average concentration
is 0.44 times the Final Acute Value. No important adverse effects
on freshwater aquatic organisms have been reported to be caused by
concentrations lower than the 24-hour average concentration.
CRITERION: For 1,2-diphenylhydrazine the criterion to
protect freshwater aquatic life as derived using the Guidelines is
17 ug/1 as a 24-hour average and the concentration should not
exceed 38 ug/1 at any time.
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Table 1 Freshwater fish acute values for 1,2-diphenylhydrazine (U.S. EPA, 1978)
Adjusted
Bioassay Test Time LC50 LC50
Organise Method* Cone.** (fare)- (uq/11 (ug/ll
Bluegill. S U 96 270 150
Lepomis macrochirus
* S = static
** U «• unmeasured
Geometric mean of adjusted values = 150 ug/1 = 38 pg/1
00
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09
I
tn
Table 2. Freshwater invertebrate acute values for 1,2-diphenylhydrazine (U.S. EPA, 1978)
Organism
Cladoceran,
Daphnia magna
fiioassay
MetiiOQ*
S
Test
Cone .**
U
Time
(nts)
48
Adjusted
LC50 LCbO
(Uq/i) (Uq'l)
4,100 3,470
* S = static
** U = unmeasured
Geometric mean of adjusted values = 3,470 pg/1 A. - 170 wg/1
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Table 3. Freshwater invertebrate chronic values for 1,2-diphenylhydrazine (U.S. EPA, 1978)
\
Chronic
Limits Value
Organism Teat* lug/11 (yg/1)
Cladoceran, LC 150-420 251
Daphnla magna
* LC » life cycle or partial life cycle
Geometric mean of chronic value
Lowest chronic value =251 Mg/1
251
Geometric mean of chronic values - 251 ug/1 - * 49 wg/1
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SALTWATER ORGANISMS
Introduction
No acute toxicity, chronic toxicity or residue data are
t
available for saltwater organisms and 1,2-diphenylhydrazine.
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CRITERION FORMULATION
Saltwater-Aquatic Life
No saltwater criterion can be derived for 1,2-diphenylhy-
drazine using the Guidelines because no Final Chronic Value for
either fish or invertebrate species or a good substitute for
either value is available, and there are insufficient data to
estimate a criterion using other procedures.
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1,2-DIPHENYLHYDRAZINE
REFERENCES
U.S. EPA. 1978. In-depth studies on health and environmental
impacts of selected water pollutants. Contract No. 68-01-
4646.
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-Mammalian Toxicology and Human Health Effects
EXPOSURE
Introduction
1,2-diphenylhydrazine (DPH) is a precursor in the manu-
facture of benzidine, an intermediate in the production
of dyes. If2-diphenylhydrazine is used in the synthesis
of phenylbutazone, a potent anti-inflammatory (anti-arthritic)
drug (Wenner, 1967).
The commercial production of 1,2-diphenylhydrazine
per se in 1977 was in excess of 1000 Ibs. (SRI, 1977).
However, this figure is probably an underestimate of the
amount of diphenylhydrazine that was actually available.
Diphenylhydrazine is produced in several synthetic processes
as an intermediate and a contaminant but there is no way
of estimating these quantities which are substantial.
Industrial exposure to 1,2-diphenylhydrazine is prim-
arily limited to workers in the dye manufacturing industry
and in the pharmaceutical industry.
Derivatives of hydrazine are reported to be hepatotoxic,
hemolytic, convulsants, and irritants. They are absorbed
from all routes (Button, 1967).
The basis for concern over 1,2-diphenylhydrazine includ-
es: (1) its presence in drinking water (highest concentra-
tion reported was 1 pg/1 (Ippb), U.S. EPA, (1975); (2) the likely
production of benzidine from DPH in the stomach due to gas-
tric acidity (IARC, 1972); (3) the documented carcinogenicity
of hydrazine and selected substituted hydrazines (IARC,
1974); (4) the increased incidence of bladder cancer among
workers involved in the manufacture of dyes (Wynder, et al.
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1963); Anthony, et al. 1970); (5) the carcinogenicity of
azobenzene, a metabolite of DPH, as well as the established
carcinogenicity of benzidine to which DPH is converted.
The structure and physical data for 1,2-diphenylhydra-
zine are presented in Figure 1.
The primary commercial method of production of this
compound is the reduction of nitrobenzene by catalysts such
j-3 +2
as Fe or Zn in alkaline solution. By this procedure,
one obtains nearly quantitative yields (Kirk-Othmer, 1963).
Figure 2 depicts this process as well as the by-products
of diphenylhydrazine, azobenzene and azoxybenzene.
The reaction of 1,2-diphenylhydrazine with acid results
in the benzidine rearrangement (Kenner, 1968). This reaction
is presented in Figure 3. In addition to benzidine, other
products formed include diphenyline, o-benzidine, and o-
semidine. In the stomach, 1,2-diphenylhydrazine can be
converted into benzidine, a human carcinogen (Haley, 1975;
IARC, 1972).
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Synonyms - Hydrazobenzene
Symmetrical diphenylhydrazine
N,N' Diphenylhydrazine
N N' - Bianiline
1,1' Hydrazodibenzene
Cas No. 530-50-7(b)
Molecular weight = 184.24
Melting Point = 131°C
Boiling Point = 220°C
Solubility = Slightly soluble in water
Very soluble in benzene, ether, alcohol
Figure 1. 1,2-diphenylhydrazine: Chemical and Physical Properties3
(a) from CRC Handbook - 59th Ed.
(b) DHEW report, 1978
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Oi
Nitrobenzene
Alkaline
"1"^
*Azobenzene
*Azoxybenzene
H H
1,2-diphenylhydrazine
Figure 2. Synthesis of 1,2-diphenylhydrazine (Williams, 1959)
*Carcinogenic
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H H
1,2-diphenylhydrazine
Diphenyline
o-Benzidine
o-Seraidine
Figure 3. Benzidine Rearrangement of 1,2-diphenylhydrazine
(Williams, 1959)
*Carcinogenic
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Ingestion from Water and Foods
DPH was not detected in the finished water of any of
ten cities selected for a detailed EPA study in 1975. However,
the same study demonstrated the presence of DPH in drinking
water in concentrations up to 1 jug/1 (1 ppb).
' There are no available data identifying DPH as a direct
or indirect food additive, or as a naturally occurring consti-
tuent of any food.
Inhalation, Dermal, and Other Sources
Workers involved in the manufacture of dyes, certain
Pharmaceuticals and chemicals, laboratory workers, and work-
ers in forensic medicine risk occupational exposure to 1,2-
diphenylhydrazine. Both inhalation and dermal contact are
possible routes of exposure in these settings. However,
no experimental data are available to quantitate either the
dermal or inhalation exposure risks to this population.
; PHARMACOKINETICS
Absorption, Excretion, and Distribution
There are no available data on the absorption or excre-
tion of 1,2-diphenylhydrazine by mammals. The administration
of DPH by various routes results in systemic effects and
the presence of DPH metabolites in the urine. This indicates
that it is absorbed.
Metabolism
The metabolism of 1,2-diphenylhydrazine in the rat
is presented in Figure 4 (Williams, 1959). The 1,2-diphenyl-
hydrazine was administered to rats orally (200, 400 mg/kg),
i.p. (200 mg/kg), intratracheally (5, 10 mg/kg), and i.v.
(4, 8 mg/kg). Urines were analyzed chromatographically
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Route
p.o.
i.p.
i.t.a
i. v.
Dose (mg/kg)
200, 400
100, 200
5, 10
4, 8
Solvent
oil
DMSOb
water
DMSO
NH,
Figure 4. 1,2-diphenylhydrazine Metabolites in Rat Urine
(Williams, 1959)
intratracheally
Dimethyl sulfoxide
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(TLC) and a scheme proposed (Figure 4). Benzidine was identi-
fied as a metabolite. The metabolites detected were not
dependent upon the dose or the route of administration.
EFFECTS
Acute/ Sub-acute, and Chronic Toxicity
Two studies reporting DPH oral LD^Q'S were identified.
Marhold, et al. (1968) used ten male Wistar rats and adminis-
tered DPH as a five percent aqueous suspension. The LD
reported was 959 mg/kg. In the Registry of Toxic Effects
of Chemical Substances, 1977 (RTECS) the oral LD5Q is listed
as 301 mg/kg. No details were given. The basis for this
difference is not known. Liver damage is an important feature
of hydrazine toxicity, particularly after chronic exposure,
but Sutton (1967) noted that phenylhydrazines cause prominent
kidney damage and more diffuse liver damage in animals.
Two rat studies were reported in Reg. Tox. Eff. Chem.
Subst. (1977) . There were no data available on the strain,
sex, doses, or solvents used for these studies. One rat
study reported an oral TDLo of 396 gm/kg. Neoplasms were
seen by 53 weeks. In the second rat study, a total dose
of 16 mg/kg DPH was administered subcutaneously and neoplasms
were reported after 52 weeks.
Two mouse studies were reported in Reg. Tox. Eff. Chem.
Subst. (1977). When 1,2-diphenylhydrazine was applied topi-
cally for 25 weeks (total dose of 5,280 mg/kg) neoplasms
were found. Subcutaneous injection of DPH (8,400 mg/kg)
resulted in neoplasms after 38 weeks. These results are
presented in Table 1.
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TABLE 1
1,2-diphenylhydrazine Toxicity Data
o
Species/Strain
Rat/Wistar
Rat/N.O.S.d
Rat/N.O.S.
Rat/N.O.S.
Mouse/N.O.S.
Mouse/N.O.S.
Sex No. Dose Solvent Cone Route T.I.a Duration
(mg/kg)
M 10 N.O.S. water 5% p.o. U>SQ 959 N.O.S.
(Suspension)
N.O.S. N.O.S. N.O.S. N.O.S. N.O.S. p.o. U>50 301 N.O.S.
N.O.S. N.O.S. N.O.S. N.O.S. N.O.S. p.o. TDLoe 53 wks.
39,600
N.O.S. N.O.S. N.O.S. N.O.S. N.O.S. s.c.C TDLo 52 wks.
16,000
N.O.S. N.O.S. N.O.S. N.O.S. N.O.S. skin TDLo 25 wks.
5,280
N.O.S. N.O.S. N.O.S. N.O.S. N.O.S. S.C. TDLo 38 wks.
Effects
N.O.S.
N.O.S.
Neoplasms
Neoplasms
Neoplasms
Neoplasms
References
Mar hold, et al.
(1968)
RTECSb
RTECS
RTECS
RTECS
RTECS
8,400
(a) Toxicity Index
(b) Registry of Toxic Effects of Chemical Substances, 1977
(c) Subcutaneous
(d) Not Otherwise Specified
(e) TDLo-Toxic Dose Low - the lowest dose of a substance introduced by any route, other than
inhalation, over any given period of time and reported to produce any toxic effect in
humans or to produce carcinogenic, teratogenic, mutagenic, or neoplastigenic effects in
humans of animals.
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No epidemiological studies have been carried out on
the effects of DPH in humans.
Synergism and/or Antagonism
Marhold, et al. (1968) demonstrated that diphenyline,
a product of benzidine rearrangement, acts synergistically
with benzidine to produce tumors. Genin (1975) showed a
synergistic effect of hydrazobenzene and benzidine sulfate.
Kulyanskii, et al. (1976) observed that benzidine sulfate-
hydrazobenzene or benzidine sulfate-dianisidine sulfate
mixtures when administered subcutaneously to rats increased
the incidence of bladder cancer and decreased the latent
period for tumor development when compared with the carcino-
genic activity of the individual compounds. The authors
emphasize the importance of preventing the possible exposure
of industrial workers to combinations of 1,2-diphenylhydra-
zine and benzidine during the manufacture of benzidine-sulfate,
Teratogenicity
There are no studies available on the teratogenicity
of DPH.
Mutagenicity
Sieler (1977) studied the incorporation of H-thymidine
into testicular DNA using the technique developed by Friedman
and Staub (1976). OPH was administered i.p. in a dose of
100 mg/kg and an inhibitory effect on testicular DNA synthesis
was observed. Seiler implied from this study that DPH has
a mutagenic potential.
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Carcinogenicity
There are several reports on the possible carcinogeni-
city of DPH. The most comprehensive study was reported
in 1978 (NCI, 1978). Technical grade DPH* was fed as a
dietary admixture to rats (Fischer 344) and mice (B6C3F1)
of both sexes. Dietary concentrations (DPH) fed to rats
and mice are indicated in Tables 2 and 3. DPH was fed to
mice and rats for 78 weeks followed by observation periods
of 17 to 96 weeks and 28 to 109 weeks, respectively. Controls
consisted of 47 to 50 animals of each sex. The results
of this study are summarized in Tables 2 and 3. There were
differences in the nature and organ distribution of tumors
between sexes and species. DPH was carcinogenic to Fischer
344 rats of both sexes and caused significant increases
in hepatocellular carcinoma at 5 pg/kg/day and 18.8 jig/kg/day;
Zymbal's gland squamous-cell tumors in male rats at 18.8
jug/kg/day and neoplastic liver nodules in female rats at
7.5 jug/kg/day. Female mice showed an increase in hepatocelluar
carcinomas only at 3.75 /ig/kg/day. DPH was not carcinogenic
in B6C3F1 male mice.
120 - 124 Cf K & K labs.
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to
TABLE 2
Carcinogenicity of 1,2-diphenylhydrazine in Mice*
Mf» ^ Ir r- r>£ C^nl. r.
Sex f
50
50
male
50
50
50
50
female
47
50
Dose Treated Observed Hepatocellular
Post treatment Carcinomas
LD control
HD control
0.75 /ig/kg/day
3.75 jug/kg/day
LD control
HD control
0.375 /ig/kg/day
3.75 /ig/kg/day
0
0
78
78
0
0
78
78
95
96
17
17
96
96
17
18
12/50
6/48
11/47
8/46
2/47
1/50
4/39
20/43 p<0.001
Pulmonary
Carcinomas
5/50
5/49
1/47
0/46
2/46
3/50
3/38
2/40
*Data taken from NCIf 1978
LD = Low dose
HD = High dose
Dose = DPH in diet
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TABLE 3
Carcinogenic!ty of 1,2-diphenylhydrazine in Rats
Sex I
SO
49
male
SO
SO
SO
50
female
50
50
Effects
Weeks — —
Dose Treated Observed Hepatocellular Zymbal's Gland
Carcinoma
Ld control
HD control
5 ug/kg/day
18. B ug/kg/day
LD control
HO control
3 ug/kg/day
7.5 ug/kg/day
0
0
78
78
0
0
78
78
108
109
29
28
109
109
30
29
0/47
1/48
5/49 p 0.031
31/49 p 0.001
Hepatic Neoplastic
0/47
0/50
0/50
6/50 p 0.013
0/47
1/48
2/50
7/49 p 0.007
Nodules
0/48
0/50
1/50
0/50
Pulmonary
Carcinomas
1/47
1/48
3/49
1/48
Mammary carcinomas
1/48
0/50
3/50
6/50 p<0.013
*Data taken from NCI, 1978.
LD = Low dose
HD = High dose
Dose = % DPH in diet
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In an additional study by Pliss in 1974, the carcino-
genic properties of DPH were studied over a period of 588
days in rats (N = 163) and C 57 mice (N = 110). DPH was
suspended in sunflower seed oil and administered by s.c.
injection (40 mg/wk/rat and 5 mg/wk/mouse), and by addition
to food (30 mg/5 times/wk), or application to the skin
(30 mg/5 times/wk/rat and 2 mg/3 times/wk/mouse).
TABLE 4
*
Carcinogenicity of DPH in Mice and Rats*
Effects
Species Route % Tumor Incidence Tumors
mice s.c. * 36.6
PO 50
epicutaneous 22.2
r habdomyosarcoma
pulmonary adenoma,
leukemia, liver
skin, lung, liver
rats
s.c.
22.6
uterus, mammary,
Zymbal's gland, liver
spleen, lymphoid leukemia
The data summarized in Table 4 indicate that
DPH produces a wide variety of tumors in both mice and rats.
Data taken from Pliss, 1974.
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In contrast to the NCI (1978) study and the report
by Pliss (1974), Marhold, et al. (1968) and Spitz (1950)
did not find any significant increase in carcinogenicity
by DPH. These latter two studies were difficult to interpret
due to the lack of specific information on the purity of
DPH, experimental design or statistical analysis. The Pliss
study (1974) should be used in a cautious manner in indicting
DPH as a carcinogen. The author indicates that animals
had to be added to the study in order to replace animals
afflicted with a parasitic infection. In addition, although
the tumor incidence is given for DPH treated animals, the
incidence of tumors in control animals is not presented
except in the case of the epicutaneous administration of
DPH. Values of 17 percent vs. 22.2 percent for control
and DPH groups, respectively, are presented but no statistical
analysis of these incidences is given. The NCI (1978) report
stands in marked contrast to the other published studies
on the carcinogenecity of DPH. It represents the only reli-
able study and indicates that DPH is carcinogenic.
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CRITERION FORMULATION
Existing Guidelines and Standards
No existing guidelines or standards were found for
1,2-diphenylhydrazine.
Current Levels of Exposure
No information is available on the concentration of
1,2-diphenylhydrazine in the atmosphere.
1,2-Diphenylhydrazine has been found to be present
in drinking water at levels of 1 ;ug/l = 1 ppb (U.S. EPA,
1975).
1,2-Diphenylhydrazine has not been found to be a natural
constituent of food.
Special Groups at Risk
Manufacturers of dyes and Pharmaceuticals are subject
N
to occupational exposure. Groups working in the laboratory
and forensic medicine may also be subject to 1,2-diphenyl-
hydrazine exposure.
Basis and Derivation of Criterion
An evaluation of the subacute, acute and chronic toxicity,
with the exception of carcinogenicity is impossible because
of only scanty data. No current guidelines or standards
presently exist for DPH. Diphenylhydrazine has been shown
to produce carcinogenic responses in rats and mice (NCI,
1978; Pliss, 1974). Since the NCI (1978) study represents
the only report in which all the data can be analyzed, it
will be used as a basis for formulating a criterion.
More specifically, the data on the induction of cancer
in male and female rats and female mice were chosen for
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analysis because they all had significantly increased tumor
formation following DPH treatment (i.e. dietary). The respec-
tive criterion levels obtained from applying the standard
water quality dose extrapolation/criteria calculation method-
ology is given in Table 5.
TABLE 5
1,2-Diphenylhydrazine Induction of Tumors in Mice and Ratsa
Species Sex Estimated Criterion Level
at 10~3 Risk0
Mouse Female 1.43 /ig/1
Rat Female, liver carcinoma 1.32 jug/1
mammary carcinoma 1.32 /ag/1
Male, zymbol gland tumor 5.14 >ug/l
liver carcinoma 0.38/ig/l
*Data taken from NCI, 1978. (Tech No. 92, 1978)
Calculated by applying a modified "one-hit" extrapolation
model described in the Federal Register 1062-5, 1979.
It can be seen that male rats appear to have the lowest
tolerance for DPH.
Under the Consent Decree in NRDC vs. 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." DPH is suspected of being a human carcinogen.
Because there is no recognized safe concentration for human
carcinogens, the recommended concentration of DPH in water
for maximum protection of human health is zero.
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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 concentrations of DPH corresponding to
several incremental lifetime cancer risk levels have been
estimated. A cancer risk level provides 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
ambient water quality criteria, EPA stated that it is consid-
ering setting criteria at an interim target risk level of
10~5, 10"6 or 10~7 as shown in the table below.
Exposure Assumptions Risk Levels and Corresponding Criteria
0 10~7 10"6 IGf5
2 liters of drinking water 0 4 ng/1 40 ng/1 400 ng/1
and consumption of 18.7
grams fish and shellfish (2)
Consumption of fish and 0 .019 jug/1 0.19 jug 1.9 >ug/l
shellfish only.
(1) Calculated by applying a modified "one-hit" extrapolation
model described in the PR 15926, 1979. Appropriate
bioassay data used in the calculation of the model
are presented in Appendix I. Since the extrapolation
model is linear to low doses, the additional lifetime
risk is directly proportional to the water concentration.
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Therefore, water concentrations corresponding to other
risk levels can be derived by multiplying or dividing
one of the risk levels and corresponding water concen-
trations shown in the table by factors such as 10,
100, 1000 and so forth.
(2) Twenty-one percent of the DPH exposure results from
the consumption of aquatic organisms which exhibit
an average bioconcentration potential of 29 fold.
The remaining percent of DPH exposure results from
drinking water.
Concentration levels were derived assuming a lifetime
exposure to various amount of DPH, (1) occurring from the
consumption of both drinking water and aquatic life grown
in water containing the corresponding DPH concentrations
and, (2) occurring solely from consumption of aquatic life
grown in the waters containing the corresponding DPH concen-
trations.
-. Although a total exposure evaluation for DPH is desirable
i
there is no data to support a total exposure analysis.
The criteria presented, therefore, assume an incremental
risk from assumed ambient water exposure only.
For DPH the case for criterion development is based
upon the existence of carcinogenicity responses in animals
(rats and mice).
Because of the lack of investigations for other chronic
and acute responses, there is no information on other effects
in either human or animal systems. Thus, the criterion
proposed should be considered as precautionary until further
studies can be used in the overall toxicity evaluations.
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REFERENCES
Anthony, A.M., et al. 1970. Tumors of the urinary bladder:
an analysis of the occupations of 1030 patients in Leeds,
England. Jour. Natl. Can. Inst. 45: 879.
CRC. 1978. Handbook of chemistry and physics. 59th ed. CRC
Press. West Palm Beach, Fla.
Friedman, M., and J. Staub. 1976. Inhibition of mouse testicular
DNA synthesis by mutagens and carcinogens as a potential
mammalian assay for mutagenesis. Mutat. Res. 37: 67.
Genin, V.A. 1975. Increase in carcinogenic activity during
joint effect of hydrazobenzene and benzidine sulfate. Gig.
Tr. Prof. Zabol. 6: 28.
Haley, T.J. 1975. Benzidine revisited: a review of the litera-
ture and problems associated with the use of benzidine and
its congeners. Clin. Tox. 8: 13.
International Agency for Research on Cancer. 1972. Aromatic
amines. Monog. on the evaluation of carcinogenic risk of
chemicals to man. 1: 69.
International Agency for Research on Cancer. 1974. Hydrazine
and its derivatives. Monog. on the evaluation of carcinogenic
risk of chemicals to man. 4: 81.
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Kenner, J. 1968. Benzidine rearrangement. Nature. 219: 153.
Kulyanskii, et al. 1976. Kirk-Othmer Encyclopedia of Chemical
Technology. 1963. 2nd ed. Vol. 3. New York: Interscience.
p. 408.
Marhold, J. Jr., et al. 1968. The possible complicity of
diphenyline in the origin of tumors in the manufacture of
benzidine. Neoplasma 15: 3.
NCI Publication NO. (NIH) 78-1342. 1978. Bioassay of hydrazo-
benzene for possible carcinogenicity.
Pliss, G.B. 1974. Carcinogenic properties of hydrazobenzene.
Vop. Onkol. 20: 53.
Registry of Toxic Effects of Chemical Substances. II: 1977.
i
Sieler, J.P. 1977. Inhibition of testicular DNA synthesis
by chemical mutagens and carcinogens. Preliminary results
in the validation of a novel short term test. Mutat. Res.
46: 305.
Spitz, S. 1950. The carcinogenic action of benzidine. Cancer.
3: 789.
Stanford Research Institute. 1977. 1977 Directory of chemical
producers, U.S.A. Menlo Park, Calif.
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Button, W.L. 1967. Heterocyclic and miscellaneous nitrogen
compounds. Industrial Hygiene and Toxicology. Vol. II:
2171. Toxicology, 2nd ed. F.A. Patty, ed. New York: Interscierice
Publishers.
U.S. EPA. 1975. Preliminary assessment of suspected carcinogens
in drinking water. Rep. to Congress. 11.
Wenner, W. 1967. Malonic acid and derivatives in Kirk-Othmer
Encyclopedia of Chemical Technology. 2nd ed. Vol. 12: 857.
New York: Interscience Publishers.
Williams, R. 1959. Detoxication Mechanisms. New York: John
Wiley and Sons. p. 480.
Wynder, E.L., et al. 1963. An epidemiological investigation
of cancer in the bladder. Cancer 16: 1388.
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APPENDIX I
Summary and Conclusions Regarding the
Carcinogenicity of 1,2,-Diphenylhydrazine*
I,2-Diphenylhydrazine is used primarily in dye manu-
facturing industries as a precursor in the synthesis of
benzidine. There are no data showing carcinogenic effects
of 1,2-diphenylhydrazine in humans. However, two studies
have shown that 1,2-diphenylhydrazine is carcinogenic in
mice and rats via subcutaneous and oral routes of administra-
tion. Male rats, receiving dietary concentrations of 0.03
percent 1,2-diphenylhydrazine, developed hepatocellular
carcinomas and squamous cell carcinomas of the Zymbal glands.
Female rats, receiving 0.01 percent 1,2-diphenylhydrazine
in the diet, developed neoplastic nodules of the liver and
mammary adenocarcinomas. Female mice, exposed to 0.04 percent
1,2-diphenylhydrazine in the diet, developed hepatocellular
carcinomas.
The carcinogenic responses induced in male and female
rats and female mice constitute substantial evidence that
1,2-diphenylhydrazine is likely to be a human carcinogen.
The water quality criterion for 1,2-diphenylhydrazine
is based on the induction of hepatocellular carcinoma and
neoplastic nodules in male Fischer 344 rats, exposed to
a time-weighted average concentration of 0.03 percent (300
ppm) 1,2-diphenylhydrazine in the diet for 78 weeks (NCI,
1978). The concentration of 1,2-diphenylhydrazine in water
calculated to keep the lifetime cancer risk below 10 is
0.40 micrograms per liter.
*This summary has been prepared and approved by the Carcinogens
Assessment Group of EPA on June 15, 1979.
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Summary of Pertinent Data
The water quality criterion for 1,2-diphenylhydrazine
is based on the induction of hepatocellular carcinomas and
neoplastic nodules in male Fischer 344 rats, exposed to
0.03 percent (300 ppm) 1,2-diphenylhydrazine in the diet
ad_ libitum for 78 weeks (NCI, 1978). The incidence of hepato-
cellular carcinomas and neoplastic nodules was 37/49 and
1/48 in the treated and control groups, respectively. The
criterion was calculated from the following parameters:
nt =37 d* = 15 mg/kg/day
Nt = 49 F = .0187 kg/day
nc = 1 R = 29
Nc = 48 W = 0.375 kg
Le = 104 weeks
le = 78 weeks
L = 104 weeks
Based on these parameters, the "one-hit" slope (BH) is 0.715
(mg/kg/day) . The resulting water concentration of 1^2-
diphenylhydrazine, calculated to keep the individual lifetime
cancer risk below 10" , is 0.40 >ig/l.
* The dose (expressed as mg/kg (body weight) /day) is
based on the assumption that the amount of diet consumed
by rats each day was five percent of their body weights0
0.05 x 0.375 kg = 0.01875 kg diet/day
0.01875 kg diet/day x 300 mg/kg = 5.625 mg 1,2 DPH/day
5.625 mg 1,2 DPH/day/0.375 kg = 15 mg/kg/day
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