COUMAPHOS
EXPLANATION
First draft prepared by Dr S. Caroldi
University of Padua, Padua, Italy
The present Meeting reviewed available data evaluated at the 1987
Meeting, together with data submitted to the present Meeting.
Coumaphos has been evaluated toxicologically seven times previously,
in 1968, 1972, 1975, 1978, 1980, and 1987 (Annex 1, FAO/WHO, 1969ab,
1973ab, 1976a, 1979a, 1981ab, and 1987b). The temporary ADI was
withdrawn in 1980. This monograph addendum summarizes the data
submitted in 1987 and 1990.
EVALUATION FOR ACCEPTABLE INTAKE
TOXICOLOGICAL STUDIES
Long term/carcinogenicity studies
Fifty male and 50 female Wistar rats (Bor: WISW strain.
Winkelmann-Borcher) were treated with coumaphos (technical grade,
99.2% purity) at concentrations of 0, 1, 5, 25 ppm, equal to 0, 0.05,
0.25, and 1.22 mg/kg bw/day for males and 0, 0.07, 0.36, and 1.70
mg/kg bw/day for females (calculated as average daily intake througout
the duration of the study) for 24 months. Twenty additional rats per
sex per dose were treated for 12 months and then sacrificed for
intermediate investigations. Rats were four to five weeks old at the
beginning of the study. Coumaphos was mixed with the feed up to the
nominal concentrations. The actual content of coumaphos in the
formulations was checked every three months and were found to be
similar to the nominal levels.
In male rats, food and water intake and weight gain were not
influenced by coumaphos administration up to 25 ppm. In female rats
marginal reduction in food and water intake was observed at 25 ppm
coumaphos (about 12% for both food and water, the difference being
statistically significant only for water intake). Body weight gain
was reduced in females, compared to controls.
Neither sex showed any changes in the incidence of clinical
signs, or mortality rate at any dose level. At the end of the
treatment the mortality rates were 8, 16, 22, and 18% (males) and 23,
24, 7, and 18% (females) at 0, 1, 5, and 25 ppm coumaphos,
respectively. Differences in haematological, clinical chemistry and
urinalysis parameters were trivial and not related to coumaphos
administration. Cholinesterases were measured in plasma and
erythrocytes (method by Ellman et al., 1961) after 5, 14, 27, 53,
80, and 104 weeks and in brain after 53 and 104 weeks from the
beginning of the study. In males the 25 ppm dose reduced
cholinesterase activity in plasma by 27-43% while in females the
average inhibitions varied between 45-53%. The 25 ppm dose reduced
cholinesterase activity in erythrocytes by 16-28% in males and 18-34%
in females (range of average inhibitions at different times). At 5
ppm trivial and sporadic differences were detected. Brain
cholinesterase was significantly higher in all treated females after
53 weeks of treatment (normal at the end of the study) and in all
dosed males at the end of treatment (a marginal increase had been
detected after 53 weeks only in the group dosed with 5 ppm). The
increased activity of brain cholinesterase was dose related ranging
between 15-42% in females after 53 weeks and between 23-43% in males
at the end of the study.
Ophthalmological examination was performed at the beginning of
the study, after 12 months and at the end of the study. Pupillary
reflexes were evaluated together with the regions around the eyes and
the anterior segments. Refractive media of the eyes and fundus oculi
were observed after pupil dilatation. Scattered alterations were not
dose related.
Pathology was performed on all animals. Gross pathological
effects were comparable between groups. Organ weights (brain, heart,
lung, spleen, kidneys, adrenal glands, testicles and thyroid gland)
were not different at 12 and 24 months. In males treated with
coumaphos at 25 ppm, liver weight was lower than in controls at 12
months but not at 24 months.
There were no statistically significant differences in the
incidence of neoplasms. No non-neoplastic changes appeared to be
compound-related. The dose of 5 ppm coumaphos was tolerated without
damage under the conditions described. The NOAEL for coumaphos in
the diet in this study was 5 ppm, equal to 0.25 mg/kg bw/day and to
0.36 mg/kg bw/day for male and female rats, respectively. The study
indicates absence of any evidence of carcinogenicity under the test
conditions (Eiben, 1988).
Special studies on delayed neurotoxicity
Groups of 1-8 hens (Leghorn laying hens, 12 months old) received
single (50 or 100 mg/kg bw) or up to 90 daily (5 or 10 mg/kg bw) doses
of coumaphos (97.1% purity) by oral route in gelatin capsules.
Controls received empty gelatin capsules. Other hens received single
(50, 100, 500 mg/kg bw) or daily (100 mg/kg bw for either 30 or 90
days) dermal applications of coumaphos (dissolved in acetone and
applied on a 5.3 cm› area on the unprotected back of the neck).
All birds dosed orally with coumaphos showed acute cholinergic
symptoms despite atropine treatment. All 4 hens dosed with 100 mg/kg
bw died within 1-5 days. The single hen dosed with 50 mg/kg bw
survived the 30 day observation period. At 10 mg coumaphos/kg bw
daily 3/3 hens died because of acute toxicity while 2/3 hens survived
the 5 mg/kg bw coumaphos daily doses up to the scheduled 90 doses.
Animals dosed dermally with coumaphos did not show acute cholinergic
symptoms.
All hens which survived oral coumaphos and all hens which
received coumaphos dermally developed clinical signs of delayed
neuropathy. The severity of toxic effects (measured on a 1-4 scale)
was dose related varying between a minimum of gross ataxia (grade 2),
detectable after a single oral or dermal dose of 50 mg/kg bw of
coumaphos, and a maximum of complete paralysis (grade 4), reached
after 48-50 days of dermal applications of 100 mg coumaphos/kg bw.
Some improvement was detected in birds which received 90 doses (either
oral or dermal), after the treatment was stopped.
Animals dosed with coumaphos per os did not show
histopathological changes either in peripheral nerves or in spinal
cord (equivocal alterations were observed in the spinal cord of a
single hen). Histopatological changes were not detected in peripheral
nerves of dermally treated birds (except a single hen dosed daily with
100 mg coumaphos/kg bw which died after 49 doses). Unequivocal
alterations in spinal cord were detected in about 50% of hens dosed by
dermal route, the other 50% showing either equivocal alterations or
being unaffected.
It was concluded that dermally applied coumaphos is capable of
causing delayed neurotoxicity in adult hens. Acute toxicity of orally
administered coumaphos does not allow the administration of dosages
large enough to produce delayed neuropathy without killing the hens
(Abou-Donia et al., 1982).
Thirty hens (Leghorn, Brinkschulter, Senden, 12-18 months old)
were dosed orally with 50 mg/kg bw of coumaphos (Asuntol 99.1% purity
grade dissolved 2% v/v in deionized water and given by gavage)
followed, three weeks later, by a second oral dose of 100 mg/kg bw
Coumaphos. All birds were treated with atropine against cholinergic
toxicity. Five other hens were dosed orally with tri-ortho-cresyl
phosphate (TOCP, 375 mg/kg bw) as positive control and 6 hens received
vehicle only.
The animals dosed with coumaphos showed cholinergic symptoms
starting a few hours after dosing and 4/30 and 3/26 died after the
first and the second dose, respectively, in spite of atropine therapy.
Surviving hens were observed for abnormal gait until day 42 after
first dosing but none of them developed ataxia. Histopathology
performed in 6 hens randomly chosen from the 23 survivors showed no
signs of degeneration in peripheral nerves, lumbar spinal cord,
medulla oblongata and cerebellum. Scattered alterations observed in
thoracic and cervical spinal cord were not different from those
observed in controls.
Hens dosed with TOCP did not show acute cholinergic symptoms.
All of them developed delayed neuropathy starting on day 10, and
progressed to complete paralysis between day 16 and 20. Histopathology
(performed in 4 birds) showed marked degeneration in all observed
sections. Therefore, a double oral dose of coumaphos (50 + 100 mg/kg
p.o.), which resulted in severe cholinergic toxicity, did not cause
delayed neurotoxicity in hens (Flucke et al., 1987).
The effect of coumaphos administration on neuropathy target
esterase (NTE) was investigated in nervous tissue of hens. Single
doses of coumaphos (97.7% purity) were given to chickens (Lohmann
selected Leghorn, Brinkschulter, Senden, older than 9 months) either
by gavage (100 mg/kg bw to 20 birds in two different studies) or by
dermal application (500 mg/kg bw to 6 birds). Animals were killed 24
and 48 hours after dosing and NTE activity was measured in the brain
and spinal cord. NTE activities in nervous tissue of hens dosed with
coumaphos were compared with those measured in vehicle treated
controls. Other hens were dosed by gavage with TOCP (300 mg/kg bw),
a known inhibitor of NTE, and NTE was measured. All animals were
treated with atropine and pralidoxime against cholinergic toxicity.
Acute symptoms were recorded but not reported in the study.
Cholinesterase activities were not measured and no spontaneous deaths
occurred within 48 hours. NTE activity in nervous tissue of controls
was within the expected range in hens. As expected NTE inhibition was
between 80 and 90% in animals dosed with TOCP.
Forty-eight hours after 100 mg/kg bw coumaphos p.o., a single hen
showed about 60% inhibition of NTE in brain and spinal cord. Two other
hens showed NTE inhibition between 20 and 30% after 24 hours in both
tissues. In remaining hens NTE activity was normal or marginally
affected. After dermal application of 500 mg coumaphos/kg bw dermally
(area of application not reported) inhibition, of NTE was below 10% in
all birds (Bomann et al., 1989a, 1989b).
This study is insufficient to draw conclusions regarding
neurotoxicity for the following reasons: NTE was measured in brain
and spinal cord but not in peripheral nerves, the real target for
delayed neuropathy; NTE was measured only up to 48 hours which may not
be sufficient in the case of delayed absorption/disposal of coumaphos;
no hens were observed for symptoms of delayed neuropathy; and no
histological examination was performed at the time when the onset of
delayed neuropathy is expected.
Special study on haematological effects
Rat
The effects of the oral administration of a single dose of 0, 1,
3 and 10 mg/kg bw coumaphos (purity not given) in 1% tylose were
investigated on several haematological parameters in groups of 5 WISW
(SPF Cpb)/Winkelmann rats (sex not given). Analysis of blood samples
collected 90 min after dosing showed no treatment-related effects on
haematocrit, haemoglobin, thromboelastogram, platelet number and
aggregation, blood sedimentation rate, fibrinogen, thrombin and
thromboplastin time (Seuter and Perzborn, 1981).
Special study on urinary function
Rats
The effect of coumaphos on urine and electrolyte excretion was
studied in groups of male Bor: WISW (SPF Cpb)/Winkelmann rats
administered a single oral dose of 0, 1, 3 and 10 mg/kg bw coumaphos
(purity not given) suspended in a 1% methylcellulose solution. The
volume of urine and the amount of urinary K+ and Na+ excreted in
the 6 hours after dosing were measured. There was no significant
difference in the mean volume of urine excreted or in the amount of
either of the 2 electrolytes between any of the treated groups and the
controls (Garthoff, 1981a).
Observations in humans
The effects of the human exposure to coumaphos resulting from the
spraying of a 0.1% Asuntol emulsion for veterinary use in a piggery
was investigated in 4 human subjects (male, 30-50 years). These
subjects stayed in the piggery continuously for the first 2 hours of
the study and intermittently thereafter. No acetylcholinesterase
activity inhibition was observed in the whole blood of these subjects
or in plasma and erythrocytes of 6 treated pigs after 1, 2, 4, 6, or
24 hours after spraying. Measurement of coumaphos concentration in
the air at 50 and 150 cm above the floor showed that only small amount
of the compound was present during the first 50 minutes after spraying
(0.123 and 0.097 mg/m3, respectively). No detectable amount of
coumaphos could be found in the air after 135 minutes from the
spraying (Eben et al., 1981).
COMMENTS
The data base is incomplete in that the teratogenicity studies
were submitted too late for evaluation. Further, questions existed
with respect to the acceptability of a mouse reproduction study
reviewed by the JMPR in 1968. Consequently an ADI could not be
estimated.
Studies without which the determination of an ADI is impracticable
- Rat teratogenicity study
- Rabbit teratogenicity study
- Adequate multigeneration study
REFERENCES
Abou-Donia, M.B., Makkawy, H., and Graham, D.J., (1982). Coumaphos:
delayed neurotoxic effect following dermal administration in hens. J.
Toxicol. Environ. Health, 10: 87-99.
Bomann, W., Eben, A.. Asuntol (1989a). Investigations into the effect
on neurotoxic esterase (NTE) after dermal application to chickens.
Unpublished report No. 18415 from Bayer AG, Institute of Toxicology.
Submitted to WHO by Bayer.
Bomann, W., Eben, A.. Asuntol (1989b). Investigations into the effect
on neurotoxic esterase (NTE) after dermal application to chickens.
Unpublished report No. 18423 from Bayer AG, Institute of Toxicology.
Submitted to WHO by Bayer.
Eben, A., Machemer, L., Dorn, H. and Lorke, D., (1981). Influence of
a spray treatment with a 0.1% aqueous Asuntol emulsion on the operator
and the treated animals (pigs). Unpublished report No. 9860 from
Institute for Toxicology, Bayer AG. submitted to WHO by Bayer AG.
Eiben, R. (1988). Studies on chronic toxicity and carcinogenicity in
Wistar rats (administration with feed for 24 months). Unpublished
report No. 17131 from Bayer AG, Institute of Toxicology. Submitted to
WHO by Bayer.
Ellman, G.L., Courtney, K.D., Andres, V., Featherstone, M., 1961. A
new and rapid colorimetric determination of acetylcholinesterase
activity. Biochem. Pharmacol., 7: 88-95.
Flucke, W., Kaliner, G.. Asuntol-Wirkstoff (1987). An examination of
acute neurotoxicity after oral dosing in the hen. Unpublished report
No 15430 from Bayer AG, Institute of Toxicology. Submitted to WHO by
Bayer.
Garthoff B., (1981a). Bay B 9134: Study of diuretic action in rats.
Unpublished report No. 10072 from Dept. of Cardiovascular
Pharmacology, Pharmacological Institute, Bayer AG. Submitted to WHO
by Bayer AG.
Garthoff B., (1981b). Bay B 9134: Effect on hemodynamics and
myocardial contractility in anesthetized dogs after oral
administration. Unpublished report No. B/B 9134/002 from Dept. of
Cardiovascular Pharmacology, Pharmacological Institute, Bayer AG.
Submitted to WHO by Bayer AG.
Goodman D.G., Ward J.M., Squire R.A., Chu K.C., and Linhart M.S.,
(1979). Neoplastic and nonneoplastic lesions in aging F344 rats.
Toxicol. Appl. Pharmacol. 48: 237-248.
Green L.L., (1977). Coumaphos levels in tissues and feces from 13
week; 180 day; and 103 week studies. Unpublished report No. 76-36-
106002 from Gulf South Research Institute, New Iberia, Louisiana, USA.
Submitted to WHO by Tracor Jitco, Inc. Rockville, Maryland., USA.
Herbold B., (1981a). Asuntol: Salmonella/microsome test for the
detection of point mutagen effects. Unpublished report No. 10188 from
Institute of Toxicology, Bayer AG. Submitted to WHO by Bayer AG.
Herbold B., (1981b). P-O analogue of Asuntal: Salmonella/microsome
test for the detection of point mutagen effects. Unpublished report
No. 10094 from Institute for Toxicology, Bayer AG. Submitted to WHO
by Bayer AG.
Herbold B., (1981c). Chlorferron: Salmonella/microsome test for
point-mutagenic activity. Unpublished report No. 10095 from
Toxicological Institute, Bayer AG. Submitted to WHO by Bayer AG.
Herbold B., (1981d). Potasan: Salmonella/microsome test for the
detection of point mutagen effects. Unpublished report No. 10093 from
Institute of Toxicology, Bayer AG. Submitted to WHO by Bayer AG.
NCI, (1979a). Bioassay of coumaphos for posible carcinogenicity.
Veterinary Medical Review, 2: 179-181 (Abstract).
NCI, (1979b). Bioassay of coumaphos for possible carcinogenicity.
Unpublished report No. 96 from Carcinogenesis Testing Program,
Division of Cancer Cause and Prevention, National Cancer Institute,
National Institutes of Health, Bethesda, Maryland USA. Submitted to
WHO by Bayer AG.
Neuser V. and Jacobi H., (1981). Bay B 9134: Effect on central
nervous system. Unpublished report No. 10548 from Troponwerke GmbH
and Co. KG. Submitted to WHO by Bayer AG.
Seuter F. and Perzborn E., (1981). Bay B 9134: Haemopharmacological
investigations. Unpublished report No. 10070 from Institute for
Pharmacology, Bayer AG. Submitted to WHO by Bayer AG.
See Also:
Toxicological Abbreviations
Coumaphos (ICSC)
Coumaphos (FAO/PL:1968/M/9/1)
Coumaphos (WHO Pesticide Residues Series 2)
Coumaphos (WHO Pesticide Residues Series 5)
Coumaphos (Pesticide residues in food: 1978 evaluations)
Coumaphos (Pesticide residues in food: 1980 evaluations)
Coumaphos (Pesticide residues in food: 1983 evaluations)