DIPHENYLHYDRAZINE Ambient Water Quality Criteria Criteria and Standards Division Office of Water Planning and Standards U.S. Environmental Protection Agency Washington, D.C. ------- 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. ------- 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 A-l ------- 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. A-2 ------- 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. A-3 7 ------- Windholtz, M., ed. 1976. The Merck Index. Merck and Co, Inc., Rahway, N.J. A-4 ------- 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. B-l ------- 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. B-2 ------- 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. B-3 ------- 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 I ------- 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 ------- 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 Cfl ------- SALTWATER ORGANISMS Introduction No acute toxicity, chronic toxicity or residue data are t available for saltwater organisms and 1,2-diphenylhydrazine. B-7 ------- 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. B-8 ------- 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. B-9 ------- -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. C-l ------- 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). C-2 ------- 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 C-3 ------- Oi Nitrobenzene Alkaline "1"^ *Azobenzene *Azoxybenzene H H 1,2-diphenylhydrazine Figure 2. Synthesis of 1,2-diphenylhydrazine (Williams, 1959) *Carcinogenic C-4 ------- H H 1,2-diphenylhydrazine Diphenyline o-Benzidine o-Seraidine Figure 3. Benzidine Rearrangement of 1,2-diphenylhydrazine (Williams, 1959) *Carcinogenic c-5 ------- 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 C-6 ------- 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 C-7 ------- (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. C-8 ------- 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. ------- 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. C-10 ------- 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. C-ll ------- o I t-> 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 ------- o I 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 ------- 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. C-14 ------- 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. C-15 ------- 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 C-16 ------- 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. C-17 ------- 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. c-18 ------- 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. C-19 ------- 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. C-20 ------- 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. C-21 ------- 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. C-22 ------- 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. c-23 ------- 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 C-24 ------- |