Methomyl

Repeated dose toxicity: oral

Administrative data

Endpoint:

sub-chronic toxicity: oral

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference 1

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Reference 2

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Reference 3

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Reference 4

Reference Type:

study report

Title:

Unnamed

Year:

1979

Report date:

1979

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Groups of male and female, Fischer 344 rats were administered test substance admixed in the diet at levels of 100-600 ppm (males) and 135-810 ppm (females) for 13 weeks; 50 ppm (males) and 68 ppm (females for 4 weeks followed by 800 ppm (males) and 1080 ppm (females) for 9 weeks; or 20 ppm (males) and 27 ppm (females) for 9 weeks followed by 1000 ppm (males) and 1500 ppm (females) for 4 weeks. The animals were observed for clinical observations, clinical pathology, gross pathology and histopathology.

GLP compliance:

no

Limit test:

no

Test material

Test material information

Constituent 1

Reference substance name:

Methomyl

EC Number:

240-815-0

EC Name:

Methomyl

Cas Number:

16752-77-5

Molecular formula:

C5H10N2O2S

IUPAC Name:

(E)-[1-(methylsulfanyl)ethylidene]amino N-methylcarbamate

Test material form:

solid: crystalline

Specific details on test material used for the study:

- Substance name: S-Methyl N-[(methylcarbamoyl)oxy]thioacetimidate
- Substance ID: SD 14999-Tech
- Purity: Not reported

Test animals

Species:

rat

Strain:

Fischer 344

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Breeding Laboratories, Portage, Michigan
- Age at study initiation: males: 6 weeks, females: 7 weeks
- Housing: housed individually
- Diet: ad libitum except for 18 hour fast periods required prior to clinical pathology procedures and necropsy
- Water: ad libitum
- Acclimation period: 14 days

ENVIRONMENTAL CONDITIONS
- All animals were housed in an isolated room with filtered 100% fresh air supply and controlled temperature and humidity
- Photoperiod: 12 hrs dark / 12 hrs light

Administration / exposure

Route of administration:

oral: feed

Vehicle:

other: acetone, corn oil, diet

Details on oral exposure:

- DIET PREPARATION
- Rate of preparation of diet (frequency): Weekly
- Mixing appropriate amounts with (Type of food): Purina Rodent Chow® #1500
- Storage temperature of food: Room temperature until use, but no longer than one week
- Preparation method: The appropriate amount of the test substance was dissolved in a small quantity of acetone and combined with an amount of corn oil such that when the mixture was added to the rodent chow, a 2% w/w level of corn oil in the ration resulted. Feed was weighed into the stainless steel bowl of a Hobart mixer and the test substance/acetone/corn oil mixture was slowly added while the mixer was running. A small acetone rinse was used to achieve a quantitative transfer. The diets were mixed for one hour to assure homogeneity of the mix and to allow the acetone to evaporate. The control diet was prepared in a similar manner but without addition of the test substance.

Analytical verification of doses or concentrations:

no

Duration of treatment / exposure:

13 Weeks

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

20 ppm

Remarks:

Weeks 1-9 (males); Group DE2

Doses / concentrations 2

Dose / conc.:

27 ppm

Remarks:

Weeks 1-9 (females); Group DE2

Doses / concentrations 3

Dose / conc.:

1 000 ppm

Remarks:

Weeks 10-13 (males); Group DE2

Doses / concentrations 4

Dose / conc.:

1 500 ppm

Remarks:

Weeks 10-13 (females); Group DE2

Doses / concentrations 5

Dose / conc.:

50 ppm

Remarks:

Weeks 1-4 (males); Group DE3

Doses / concentrations 6

Dose / conc.:

68 ppm

Remarks:

Weeks 1-4 (females); Group DE3

Doses / concentrations 7

Dose / conc.:

800 ppm

Remarks:

Weeks 5-13 (males); Group DE3

Doses / concentrations 8

Dose / conc.:

1 080 ppm

Remarks:

Weeks 5-13 (females); Group DE3

Doses / concentrations 9

Dose / conc.:

100 ppm

Remarks:

males; Group DE4

Doses / concentrations 10

Dose / conc.:

200 ppm

Remarks:

males; Group DE5

Doses / concentrations 11

Dose / conc.:

400 ppm

Remarks:

males; Group DE6

Doses / concentrations 12

Dose / conc.:

600 ppm

Remarks:

males; Group DE7

Doses / concentrations 13

Dose / conc.:

135 ppm

Remarks:

females; Group DE4

Doses / concentrations 14

Dose / conc.:

270 ppm

Remarks:

females; Group DE5

Doses / concentrations 15

Dose / conc.:

540 ppm

Remarks:

females; Group DE6

Doses / concentrations 16

Dose / conc.:

810 ppm

Remarks:

females; Group DE7

No. of animals per sex per dose:

20

Control animals:

yes, plain diet

Examinations

Observations and examinations performed and frequency:

CAGE SIDE OBSERVATIONS:
- Time schedule: Twice daily

DETAILED CLINICAL OBSERVATIONS:
- Time schedule: Twice daily

BODY WEIGHT:
- Time schedule for examinations: Once weekly

HAEMATOLOGY:
- Time schedule for collection of blood: At pretest and termination (from tip of the tail) and at Day 42 (from orbital sinus)
- Animals fasted: Yes
- How many animals: 10 animals/sex/dose

CLINICAL CHEMISTRY:
- Time schedule for collection of blood: At pretest and termination (from inferior vena cava) and at Day 42 (from orbital sinus)
- Animals fasted: Yes
- How many animals: 10 animals/sex/dose

URINALYSIS: Yes / No / Not specified
- Time schedule for collection of urine: An 18 hour urine sample was obtained from 10 animals/sex/group at 45 days and at end of the study
- Animals fasted: Yes

OTHER: Plasma and red blood cell cholinesterase activities were determined in blood samples taken shortly before the beginning of dosing and Weeks 1, 3, 6 and 13 . All blood samples were obtained from the orbital sinus and enzyme activity was determined by a modified Ellman method. Brain cholinesterase activity was determined at termination.

Sacrifice and pathology:

GROSS PATHOLOGY: After 90 days of treatment, rats were fasted overnight and then sacrificed by exposure to chloroform vapor. Rats were incised to expose the contents of the abdominal, pelvic and thoracic cavities and a thorough visual examination was made of all organs and body tissues in situ. The following organs and tissues were examined for gross pathologic changes: brain, pituitary, eyes, nasal cavity, salivary gland (submaxillary), thyroid, trachea, esophagus, thymus, heart, lungs, liver, spleen, pancreas, stomach, duodenum, jejunum, ileum, colon, rectum, adrenals, kidneys, mesenteric lymph node, bladder, prostate, testes/epididymides, ovaries/with fallopian tubes, uterus, skeletal muscle, sternebrae (bone/marrow), spinal cord, unusual lesions.


HISTOPATHOLOGY: All organs, tissued and unusual lesions along with the carcass were placed in 10% formalin for preservation and possible subsequent histopathologic examination. The specimens from the organs and tissues were paraffin embedded, sectioned at 6 microns and stained with hematoxylin and eosin.
Groups DE1 and DE3 rats: brain, spinal cord (2 levels), eyes, pituitary, thyroid, adrenals, salivary gland, lymph node, esophagus, trachea, thymus, heart, lungs (2 sections), liver (2 sections), spleen, kidneys, urinary bladder, stomach, pancreas, small intestines (3 levels), large intestine (2 levels), testes/epididymides, prostate, ovaries/fallopian tubes, uterus/cervix, skeletal muscle, bone marrow (sternum or femur, after proper decalcification), nasal cavity (after proper decalcification), and unusual lesions.
Groups DE2, DE5, and DE7 rats: brain, heart, liver (2 sections), spleen, kidneys, testes/epididymides, ovaries/fallopian tubes, uterus/cervix, bone marrow (sternum or femur, after proper decalcification), and unusual lesions.
Groups DE4 and DE6 rats: heart, liver (2 sections), kidneys, bone marrow (sternum or femur, after proper decalcification), and unusual lesions.

Statistics:

Differences between control and treated group values for weekly body weights and change in body weights, hematologic, serum chemistry, cholinesterase and organ weight data were analyzed for statistical significance by the method of Dunnett. This is a multiple comparison procedure for comparing several treatments simultaneously with a control.

Results and discussion

Results of examinations

Clinical signs:

effects observed, non-treatment-related

Description (incidence and severity):

No consistent or apparent treatment related clinical effects were observed during the 90-day treatment period.

Mortality:

mortality observed, non-treatment-related

Description (incidence):

Animals at 135, 270, and 540 ppm were found dead during Week 2. The cause of the deaths were due to gastritis, enteritis and/or colitis, gastrointestinal distress. Within 2 days of water imbition for urinalysis, the female rats at 1500, 1080, 135, 270, 540, 810 ppm were found dead. These rats all had watery fluid in the pleural cavity and their lungs were darkened and congested. All rats apparently had esophageal ruptures due to attempts at regurgitation after water loading. Other rats found dead during Week 7 included the animals from 27, 800, 1080, 135, 540, 810 ppm groups. Based on the gross and/or microscopic findings, these deaths appear to be primarily related to systemic infections which are probably attributable to the water loading that occurred earlier in the Week. The animal at 810 ppm had diarrhea prior to death, dark fluid in the stomach and yellowish fluid and gas in the intestinal tract, indicative of enteritis and/or colitis. None of the deaths appear to be directly attributable to the test substance administration.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

Significant differences that appear to be directly related to test substance administration include the following: less body weight gain during Week 3 and a significantly smaller value for the mean body weights during Weeks 3-6 for the 540 ppm; smaller mean body weight values during Week 7-12 for the 810 ppm females; decreased body weight gains for male and female rats and a smaller value for the mean body weight of the females from 1080 ppm group during Week 5, which coincides with the exposure level being increased from 50 and 65 ppm to 800 and 1080 ppm for the males and females, respectively; and decreased body weight gains for the males and females in 1500 ppm group during Week 10 and smaller mean values for body weights of the males during Weeks 10 and 11 (at 1000 ppm) and the females during Weeks 10-13. This latter finding in this group coincides with an increase in dosage at the beginning of Week 10 from 20 and 27 ppm to 1000 and 1500 ppm for the males and females, respectively.

Food consumption and compound intake (if feeding study):

effects observed, non-treatment-related

Description (incidence and severity):

Statistical evaluation of the data revealed several significant (p ≤0.05) differences between treated and control animals in estimated food consumption at various weeks throughout the study. However, many of these differences, both increases and decreases were not consistent nor apparent treatment related changes.

Haematological findings:

effects observed, treatment-related

Description (incidence and severity):

After 6 weeks of the test substance administration, there was an apparent treatment related decrease in hemoglobin in the male rats with significant differences noted between control and Groups 800/1080, 200/270, 400/540, and 600/810 ppm. In addition, the female rats at 810 ppm had a significantly lower mean value for erythrocytes and an increased mean MCV value. Groups at 200/270 and 600/810 ppm also had elevated mean values for MCV. The females at 1500 ppm had a significantly lower mean value for neutrophils with a concomitantly higher mean value for lymphocytes. This latter finding was considered to be incidental or spontaneous since it did not occur in other female groups or at the Week 13 evaluation. Platelet counts did not differ between treated and control groups.

After 13 weeks of treatment, the mean hemoglobin values at 800 and 400 ppm males, and 1500, 1080, 540, 810 ppm females were significantly less than the corresponding control values. However, the males at 600 ppm had a significantly elevated mean value for hemoglobin and MCV. In addition, the mean erythrocyte values for males and females and the mean hematocrit value for females at 600/810 ppm were significantly less than the corresponding control values. 1500 ppm females and 100 ppm males had significantly elevated platelet counts. 1000, 800, and 600 ppm males and 1500, 1080 and 270, 540, 810 ppm females had significantly greater values for reticulocyte counts than did the corresponding control groups.

Clinical biochemistry findings:

effects observed, non-treatment-related

Description (incidence and severity):

After 6 weeks of treatment, significant differences included smaller group mean glucose values for 800, 400, and 600 ppm males, smaller mean values of urea for the females of 1080 and 810 ppm, greater mean values of K+ for males at 100, 200, 400, 600 ppm and females at 540 ppm, and an elevated mean value for direct bilirubin for males at 400 ppm. The elevated K+ levels are probably of little importance since the K+ differences are slight (0.5 mEq/L) and the absolute values are well within the reference range (4.1-7.7). The elevated direct bilirubin value at 400 ppm appears to be an incidental or spurious finding since differences were not observed in male animals receiving higher doses of test substance or in any of the female groups.

Significant differences noted for serum chemistry variables determined at termination include elevated mean values of urea at 1080 ppm males and 1500 ppm females and smaller mean glucose values for the females at 1500, 1080, 540, and 810 ppm. The following significant differences between control and treated values were also noted but are considered to be spontaneous or incidental and not related to test substance administration: Elevated mean values of total protein and globulin for the females at 135, 270, and 810 ppm; smaller mean values of total bilirubin at 1080 and 540 ppm females; an increased mean value of direct bilirubin and a smaller mean value of cholesterol at 800 ppm males; an elevated mean K+ level at 135 ppm males; and an elevated activity level of SGPT at 270 ppm females. These differences were slight and/or inconsistent between dose groups or sex.

Urinalysis findings:

effects observed, non-treatment-related

Description (incidence and severity):

There were no consistent or distinct treatment related differences between treated and control values from the samples obtained during Week 7 or 13.

Organ weight findings including organ / body weight ratios:

effects observed, treatment-related

Description (incidence and severity):

Both the absolute and relative weights of the spleens from the male and female rats at 1000/1500, 800/1080, and 600/810 ppm were significantly greater than the corresponding control values. In addition, the relative weights of the spleens from the female rats at 540 ppm were significantly greater than controls. The absolute and relative weights of the ovaries (with fallopian tubes) were significantly smaller in the 1500 ppm females than control values. The absolute and relative liver weights at 1080 ppm females were significantly larger than control weights and the relative liver weights of the females at 1500, 540, and 810 ppm were significantly greater than relative control weights. In addition, the females at 1500, 1080, and 810 ppm had significantly greater values for relative kidney weights when compared to the relative kidney weights of control females.

Gross pathological findings:

effects observed, non-treatment-related

Description (incidence and severity):

There were no consistent or apparent changes indicative of a treatment related effect.

Histopathological findings: non-neoplastic:

effects observed, non-treatment-related

Description (incidence and severity):

No distinct or consistent compound related microscopic changes were found in the tissue sections evaluated.

Other effects:

effects observed, treatment-related

Description (incidence and severity):

Decreases in plasma cholinesterase activity was noted for females receiving 810 ppm or greater of the test substance but similar changes were not observed in the male rats. In contrast, there was an apparent treatment related decrease in cholinesterase activity for RBCs from the high dose (800 ppm) males at termination, but similar changes were not observed in the female rats. Statistical analysis of the brain cholinesterase activity data did not reveal any statistically significant differences between treated and control values.

Effect levels

1

Key result

Dose descriptor:

other: MTD

Effect level:

> 1 000 ppm

Based on:

test mat.

Sex:

male/female

Basis for effect level:

body weight and weight gain

clinical biochemistry

haematology

organ weights and organ / body weight ratios

2

Key result

Dose descriptor:

LOEL

Effect level:

540 ppm

Basis for effect level:

body weight and weight gain

3

Key result

Dose descriptor:

LOEL

Effect level:

270 other: 200/270 ppm

Sex:

male

Basis for effect level:

haematology

Target system / organ toxicity

Critical effects observed:

no

Applicant's summary and conclusion

Conclusions:

MTD (Rat): >1000 ppm

Executive summary:

Groups of male and female rats were administered the test substance in the diet at levels of 100-600 ppm (males) and 135-810 ppm (females) for 13 weeks; 50 ppm (males) and 68 ppm (females for 4 weeks followed by 800 ppm (males) and 1080 ppm (females) for 9 weeks; or 20 ppm (males) and 27 ppm (females) for 9 weeks followed by 1000 ppm (males) and 1500 ppm (females) for 4 weeks.

No consistent or apparent test substance related clinical effects were noted during twice daily observations of the rats or 13 weeks. There were transient changes in weekly body weights observed for male and female rats receiving 1000 ppm or greater of test substance in the diet.

Clinical pathologic changes indicative of a systemic effect include decreases in hemoglobin and increases in reticulocytes at levels of 400 ppm (males) and 540 ppm (females) or greater and decreased RBC counts in males receiving 600 ppm and females receiving 810 ppm for 13 weeks. Decreased glucose levels were noted for male rats given 400 ppm or greater of test substance for 6 weeks but not after 13 weeks. Similar changes were noted in female rats after receiving 540 or 810 ppm for 13 weeks, or 68 ppm for 4 weeks followed by 1080 ppm for 9 weeks, or 27 ppm for 9 weeks followed by 1500 ppm for 4 weeks. No distinct or apparent treatment related differences were observed in the results of the urinalyses conducted during Week 7 or at termination. Decreases in plasma cholinesterase activity was noted for females receiving 810 ppm or greater of test substance but similar changes were not observed in the male rats. In contrast, there was an apparent treatment related decrease in cholinesterase activity for RBCs from the high dose (800 ppm) males at termination, but similar changes were not observed in the female rats.

Spleen weights, absolute and/or relative, were increased in males receiving 600 ppm or greater and in females receiving 540 ppm or greater of test substance. Relative weights of the livers and kidneys were increased for females receiving test substance at and above 540 and 810 ppm, respectively. No distinct or consistent test substance related microscopic changes were found in tissue sections evaluated.

Based on the data obtained in this study, the maximum tolerated dose (MTD) should be greater than 1000 ppm for both sexes and there were no apparent differences indicative of a real sex difference in response to the test substance administration.