PESTICIDE RESIDUES IN FOOD - 1984
Sponsored jointly by FAO and WHO
EVALUATIONS 1984
The monographs
Data and recommendations of the joint meeting
of the FAO Panel of Experts on Pesticide Residues
in Food and the Environment and the
WHO Expert Group on Pesticide Residues
Rome, 24 September - 3 October 1984
Food and Agriculture Organization of the United Nations
Rome 1985
DIPHENYLAMINE
Explanation
Diphenylamine (DPA) was first evaluated by the Joint Meeting in
1969 (FAO/WHO, 1970). No-effect levels in mice, rats and dogs were
determined and an ADI was estimated on the basis of the no-effect
level observed in the two-year dog study. New toxicological data on
DPA were considered by the Joint Meeting in 1976 (FAO/WHO, 1977) and
an ADI was estimated from a no effect level demonstrated in a six-
month mouse study. The Joint Meeting in 1982 (FAO/WHO, 1983)
recommended the ADI be replaced by a temporary ADI based on concern
for nephrotoxic effects in new-born rats, primarily from impurities in
the commercial grade DPA. Clarification of the toxic impurities as
well as additional toxicity data have been submitted and are reviewed
in this monograph addendum.
EVALUATION FOR ACCEPTABLE DAILY INTAKE
TOXICOLOGICAL STUDIES
Special Study on Teratogenicity
Rabbit
Groups of pregnant New Zealand White rabbits (16 per group) were
administered purified Diphenylamine (99.9 percent purity) via gastric
intubation at dose levels of 0, 33, 100 and 300 mg/kg bw/day on
gestation days 7 to 19 inclusive. On gestation day 29 does were
sacrificed and the foetuses were removed by caesarean section and
examined for visceral and skeletal anomalies. Green discolouration of
the urine was observed in all dose groups, but in particular at the
100 and 300 mg/kg dose levels. At 300 mg/kg mean food consumption was
reduced and mean body weight was depressed. There were no other signs
of toxicity or mortality considered treatment-related. Pregnancy rate
was unaffected by treatment. Macroscopic findings were unremarkable
at terminal necropsy. Litter size, litter weight, pre- and post-
implantation loss and mean foetal weight were not affected by DPA.
There were no malformations or anomalies observed which are considered
compound-related. The incidence of visceral and skeletal anomalies was
unaffected by treatment at doses of DPA up to and including 300 mg/kg
bw (Edwards, et al., 1983).
Acute Toxicity
DPA (99.9 percent purity) was slightly toxic to rats via
the oral route, with LD50 2.48 and 2.96 g/kg for females and males,
respectively. It is not acutely toxic via the dermal route in rabbits,
with no mortality evidenced at 2 g/kg.
TABLE 1. Acute Toxicity of DPA in Animals
Species Sex Route LD50 Reference
Rat M Oral 2.96 g/kg Spanjers & Til, 1982
F Oral 2.48 g/kg Spanjers & Til, 1982
Rabbit M Dermal >2 g/kg van Beek & Bruijntjes, 1982
TABLE 2. Special Studies on Mutagenicity
Test System Test Object Concentration Purity Results References
of DPA used
Host mediated S. Typhimurium 1 450 to 2 900 Not Negative Braun,
Assay TA 1950 u moles/kg given Schoenich &
Zeibarth,
1977
Ames Test S. typhimurium 100 ug/plate Not Negative Ferretti,
(with and without TA 1538 given Lu & Liu,
metabolic 1977
activation)
Ames Test S. typhimurium 3 u moles/plate Not Negative Florin, et
(with and TA 98 given al., 1980
without TA 100
metabolic TA 1535
activation) TA 1537
Mouse Lymphoma Mouse ID50 (Molarity) Not No Amacher,
Forward Mutation L5178Y/TK given mutagenic Paillet &
Assay cells activity Turner,
reported 1979
Special Studies on Eye and Skin Irritation
DPA instilled in the eyes of rabbit at 0.1g undiluted produced
only slight iritis and moderate conjunctivitis, which cleared after
ten days (van Beek, 1982a). Application of 0.5g DPA undiluted to
intact and abraded skin sites of rabbits produced only very slight
primary skin irritation (van Beek, 1982b).
Special Studies on the Identification of Toxic Impurities in
Commercial Grade DPA
Further evaluation of the amount of impurities present in
commercial grade DPA demonstrate that the concentration of each
impurity (2-aminobiphenyl, 4-aminobiphenyl and aniline) varies
considerably with the source or brand of DPA, along with batch-to-
batch variations in each type. Analytical results from different
sources of DPA indicate the primary amine impurities can vary from
3 to 7 ppm. Information presented demonstrates that purification of
DPA is now feasible both technically and economically, resulting in a
purity of 99.9 percent (Pennwalt, 1984).
COMMENTS
Diphenylamine was last evaluated in 1982 when a temporary ADI was
assigned.
Diphenylamine of 99.9 percent purity was not teratogenic to New
Zealand White rabbits at doses up to and including 300 mg/kg bw/day,
and was not mutagenic. Studies on the three main aromatic amine
impurities showed that the concentration of each varies considerably
among sources, brands and even batches. As the purification of
Diphenylamine is now feasible, an ADI for Diphenylamine of 99.9
percent purity was estimated.
Level Causing no Toxicological Effect
Mouse: 10 ppm in the diet equivalent to 1.5 mg/kg bw.
Rat: 100 ppm in the diet equivalent to 5 mg/kg bw.
Dog: 100 ppm in the diet equivalent to 2.5 mg/kg bw.
Estimate of Acceptable Daily Intake for Humans
0 - 0.02 mg/kg bw (for 99.9 percent pure material)
FURTHER WORK OR INFORMATION
Desirable:
Further observations in humans.
REFERENCES
Amacher, D.E., Paillet, S.C., & Turner, G.N. Utility of the mouse
1979 lymphoma L5178Y/TK assay for the detection of chemical
mutagens. Banbury Report, 2: 277-293. Submitted by Pennwalt
Corporation, USA, to WHO.
Beek, L. van. Eye irritation test with Diphenylamine in albino
1982a rabbits. Institute Civo-Toxicology and Nutrition TNO,
Netherlands. Submitted by Pennwalt Corporation, The
Netherlands, to WHO. (Unpublished)
Beek, L. van. Primary skin irritation test with Diphenylamine in
1982b albino rabbits. Institute Civo-Toxicology and Nutrition TNO,
Netherlands. Submitted by Pennwalt Corporation, The
Netherlands, to WHO. (Unpublished)
Beek, L. van & Bruijntjes, J.P. Acute dermal toxicity study with
1982 diphenylamine in albino rabbits. Institute Civo-Toxicology
and Nutrition TNO, Netherlands. Submitted by Pennwalt
Corporation, The Netherlands, to WHO. (Unpublished)
Braun, R., Schoneich, J., & Zeibarth, D. In vivo formation of
1977 N-nitroso compounds and detection of their mutagenic
activity in the host-mediated assay. Cancer Res. 37:
4672-4679. Submitted by Pennwalt Corporation, USA, to WHO.
Edwards, J.A., Leeming, N.M., Clark, R. & Offer, J.M. Effect of
1983 Diphenylamine on pregnancy of the New Zealand White rabbit.
Huntingdon Research Centre, Huntingdon. Submitted by
Pennwalt Corporation, USA, to WHO.
Ferretti, J., Lu, W. & Liu, M. Mutagenicity of benzidine and related
1977 compounds employed in the detection of hemoglobin. Am. J.
Clin. Pathol., 67 526-527. Submitted by Pennwalt
Corporation, USA, to WHO.
Florin, I., Rutberg, L., Curvall, M. & Enzell, C.R. Screening of
1980 tobacco smoke constituents for mutagenicity using the Ames'
test. Toxicology, 18: 219-232. Submitted by Pennwalt
Corporation, USA, to WHO.
Pennwalt Corporation. Analysis of impurities in technical
1984 Diphenylamine. Submitted by Pennwalt Corporation, USA, to
WHO. (Unpublished)
Spanjers, M.Th, & Til, H.P. Determination of the acute oral toxicity
1982 of Diphenylamine in rats. Institute Civo-Toxicology and
Nutrition TNO, Netherlands. Submitted by Pennwalt
Corporation, The Netherlands, to WHO. (Unpublished)
See Also:
Toxicological Abbreviations
Diphenylamine (ICSC)
Diphenylamine (FAO/PL:1969/M/17/1)
Diphenylamine (Pesticide residues in food: 1976 evaluations)
Diphenylamine (Pesticide residues in food: 1979 evaluations)
Diphenylamine (Pesticide residues in food: 1982 evaluations)
Diphenylamine (Pesticide residues in food: 1984 evaluations)