PHORATE EXPLANATION The toxicology of phorate was considered by Joint Meetings in 1977, 1982 and 1983 (Annex 1, FAO/WHO, 1978a, 1983a, 1984). A toxicological monograph was prepared by the Joint Meeting in 1977 (Annex 1, FAO/WHO, 1978b) and a monograph addendum was prepared in 1982 (Annex 1, FAO/WHO, 1983b). Due to the lack of an appropriate delayed-neurotoxicity study, only a temporary acceptable daily intake of 0-0.002 mg/kg b.w. was allocated in 1982. In 1983, the required study was not available. The meeting extended the temporary ADI, however, to 1985. Since then, the required neurotoxicity study and additional mutagenicity studies have become available. These studies are reviewed in this monograph addendum. EVALUATION FOR ACCEPTABLE DAILY INTAKE BIOLOGICAL DATA Toxicological studies Special study on Neurotoxicity Phorate of 89.5% purity was dissolved in corn oil and orally administered by gavage to fasted white leghorn hens (22 to 23 months old). The single dose oral LD50 was determined to be 14.2 mg/kg b.w. Fifty fasted hens were then atropinized by intramuscular injection of 10 mg/kg b.w. of atropine sulfate and 1 hour later given single oral doses of phorate at a dosage level of 14.2 mg/kg b.w. (test group). An additional 15 fasted and atropinized hens were given corn oil only (vehicle control group) and 15 more non-atropinized hens were given tri-o-tolyl phosphate (TOTP) at a dosage level of 500 mg/kg b.w. (positive control group). Twenty-one days later, the dosing was repeated for all suviving hens in the test and vehicle control groups except that atropine sulfate was administered at a dosage level of 30 mg/kg b.w. All hens were observed daily for mortality, clinical signs and evidence of neurotoxic reactions. Body weights and food consumption were recorded every 3 days. Gross necropsies were performed on all hens that died during the study and on all hens that were sacrificed at termination of the study at 42 days. Hens sacrificed at termination of the study were perfused with 10% neutralized formalin. Brains, vertebral columns (with spinal cord in situ) and the entire lengths of right and left sciatic nerves were excised and fixed. Microscopic slides of neural tissue were prepared as follows: a sagittal section of the entire brain (corpus striatum, cerebellum, pons), a longitudinal and cross section of three levels of spinal cord (cervical, thoracic, lumbrosacral) and both sagittal and longitudinal sections of the right and left sciatic nerves. Sections were stained with hematoxylin and eosin and replicate sections with luxol fast blue. The tissues from 10 hens in each of the test, vehicle-control and positive-control groups were histopathologically examined. None of the 15 vehicle-control hens died during the 42-day study. All 15 positive-control hens were sacrificed in extremis on day 16 of the study after clinical signs of neurotoxicity, first observed on day 11, progressively became more severe. These signs included generalized weakness, ataxia and paralysis of the legs and wings. Of the 50 test hens, 27 died within 24 hours of the first dose of phorate and 13 more within 24 hours after the second dose. Ten test hens survived to termination of the study at 42 days. Vehicle control and test hens displayed slight generalized weakness of the limbs, lasting about 2 hours, shortly after each treatment with atropine sulfate. Test hens had slightly more severe reactions and, in addition, slight to moderate ataxia for up to 2 hours after dosing with phorate. No clinical signs of delayed neurotoxicity, however, were observed in any vehicle-control or test hens. Mean body-weight gains of test hens were greater at 0 to 21 days and less at 21 to 42 days than those of vehicle-control hens. Mean feed consumption of test hens was less at 0 to 21 days and greater at 21 to 42 days than that of vehicle-control hens. Gross necropsies were negative for any effects attributable to the test material. Histopathological examination of neural tissues from positive- control hens revealed treatment-related lesions involving the brain, spinal cord, and/or sciatic nerves in all 10 hens. Generally mild to moderate axonal degeneration was observed in the brains of 4/10 hens, in the spinal cords of 10/10 hens, and in the sciatic nerves of 10/10 hens. In addition, demyelination was observed in the spinal cords of 7/10 hens and in the sciatic nerves of 7/10 hens. Schwann cell hyperplasia was also observed in the sciatic nerves of 3/10 hens. These lesions were compatible with a TOTP-induced delayed neuro-toxic response. Minimal to mild focal lesions of axonal degeneration of the sciatic nerves were noted in 3/10 test hens. No axonal degeneration was noted in any vehicle-control hens. The axonal degeneration observed in 3/10 test hens was associated with interstitial infiltrations of lymphoid cells, which was also observed in additional test and vehicle-control hens. This syndrome, which was distinctly different than that observed in the positive-control hens, was ascribed to lesions of naturally occurring disease (i.e. Marek's disease) and was not considered to be related to the test material. Treatment of hens with phorate did not induce clinical or histopathological signs indicative of acute delayed neurotoxicity (Fletcher, 1984). Special studies on mutagenicity Technical grade phorate of unspecified purity was tested in the mutagenicity studies reported in Table 1. Table 1. Results of mutagenicity assays with phorate Study type Dosage level and/or Results Reference conditions Bacteria tests Reverse mutation, dosage levels up to 1000 negative Simmon et al., S. typhimurium, mg/plate* 1977 strains TA1535, TA1537, TA1538, & TA100. E. coli, strain WP2 Preferential 1 mg (on filter disc)/ negative Simmon toxicity, E. coli, plate, w/o metabolic et al., 1977 strains p3478, activation W3110. B. subtilis, strains M45, H17 Yeast tests Mitotic recombination, 5% w/v for 4 hours negative Simmon S. cerevisiae D3 incubation followed by et al., 1977 plating* Cultured mammalian cells Unscheduled DNA dosage levels up to negative Simmon synthesis, human 10-3 M* et al., 1977 fibroblast (WI-38) cells In vivo study Dominant lethal, 0, 5, 10, & 20 mg/kg/ negative Simmon male mice day in the diet for et al., 1977 7 weeks, followed by weekly matings for 8 weeks * without and with metabolic activation COMMENTS Phorate did not induce clinical or histophathological signs of neurotoxicity in a neurotoxicity study in hens. No evidence of mutagenic potential was observed in a series of mutagenicity studies. Since all required toxicity studies on phorate have been submitted and evaluated, the Meeting estimated an ADI for phorate. TOXICOLOGICAL EVALUATION LEVEL CAUSING NO TOXICOLOGICAL EFFECT Rat: 1 ppm in the diet, equivalent to 0.05 mg/kg b.w. Dog: 0.01 mg/kg b.w./day ESTIMATE OF ACCEPTABLE DAILY INTAKE FOR MAN 0-0.0002 mg/kg b.w. FURTHER WORK OR INFORMATION DESIRABLE Observations in man. REFERENCES Fletcher, D.W. 42-Day neurotoxicity study with phorate in mature white 1984 leg horn chickens. Unpublished report from Bio-Life Associates, Ltd., BLAL No. 83 DN 103. AC Protocol No. 981-84-114. Submitted to WHO by American Cyanimid Company. Simmon, V.F., Mitchell, A.D. & Jorgenson, T.A. Evaluation of selected (1977) pesticides as chemical mutagens in in vitro and in vivo studies. Unpublished report from Stanford Research Institute, SRI report No. LSU-3493, for U.S. Environmental Protection Agency, EPA report No. EPA-600/1-77-028. Submitted to WHO by American Cyanamid Company).
See Also: Toxicological Abbreviations Phorate (ICSC) Phorate (Pesticide residues in food: 1977 evaluations) Phorate (Pesticide residues in food: 1982 evaluations) Phorate (Pesticide residues in food: 1984 evaluations) Phorate (Pesticide residues in food: 1994 evaluations Part II Toxicology) Phorate (Pesticide residues in food: 1996 evaluations Part II Toxicological)