Methomyl

Developmental toxicity / teratogenicity

Administrative data

Endpoint:

developmental toxicity

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

comparable to guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Materials and methods

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

The purpose of this study was to determine the toxicity and the teratogenic potential of the test substance in the rabbit. Solutions of the test substance in deionized water were administered by gavage on days 7 through 19 of presumed gestation to artificially inseminated rabbits at dosages of 0 (vehicle), 2, 6 and 16 mg/kg/day at a dosage volume of 5 mL/kg/day. Rabbits were observed for signs of agent effect, abortion or delivery. Body weights were recorded. Does which aborted were killed, examined, and observed uterine contents were recorded. Does which were found dead were similarly examined. On day 29 of presumed gestation, surviving female rabbits were killed, the abdomen of each was opened, and the uterus wag examined. Each fetus or delivered pup was weighed and subsequently examined for gender and for gross external, visceral and skeletal alterations.

GLP compliance:

yes

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:

Methomyl
Lot#: INX-1179-255
Purity: 98.7%

Test animals

Species:

rabbit

Strain:

New Zealand White

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Dutchland Laboratories, Inc., Swampbridge Road, Box 139A, Denver, Pennsylvania
- Age at study initiation: Approximately 28 weeks
- Weight at study initiation: 3.14 to 4.71 Kg
- Housing: Female rabbits were housed in vertical order according to dosage and insemination groups. Housing units consisted of eight individual stainless steel cages (5 square feet of floor area with 16 inch height per individual cage).
- Diet: Certified rabbit chow #5322, approximately 180 g each per day
- Water: ad libitum
- Acclimation period: Approximately 7 weeks

ENVIRONMENTAL CONDITIONS
- Temperature: 17.7-22.2°C
- Humidity: 30-80%
- Air changes: 12-15 per hr
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

other: deionized water

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Solutions of the test substance in reverse osmosis processed deionized water (R.O. water) a were prepared daily prior to use. Concentrations, calculated on the basis of the active ingredient were selected to provide 5 mL/kg dosage volume to rabbits in all dosage groups.

VEHICLE
- Concentration in vehicle: 0.4, 1.2 and 3.2 mg/mL for dosages of 2, 6 and 16 mg/kg respectively

Analytical verification of doses or concentrations:

no

Details on mating procedure:

- Impregnation procedure: Artificial insemination
- Verification of same strain and source of both sexes: yes
- Day of artificial insemination is referred to as day 0 of pregnancy

Duration of treatment / exposure:

GD 7 to GD 19

Frequency of treatment:

daily

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

2 mg/kg bw/day

Doses / concentrations 2

Dose / conc.:

6 mg/kg bw/day

Doses / concentrations 3

Dose / conc.:

16 mg/kg bw/day

No. of animals per sex per dose:

20

Control animals:

yes, concurrent vehicle

Details on study design:

- Dose selection rationale: Dosages evaluated in this study were selected on the basis of an initial toxicity assessment and a pilot evaluation.
In the initial toxicity assessment, one female rabbit (not inseminated) was administered a single 24 mg/kg/day dosage of the test substance to determine if a rabbit could tolerate the agent at the maximum dosage proposed for the preliminary study. In the preliminary study four artificially-inseminated rabbits were assigned to each dosage group, and the test substance was administered at dosages of 8, 16 or 24 mg/kg/day on days 7 through 19 of presumed gestation.
The 24 mg/kg/day dosage was lethal. Two rabbits in this dosage group died on day 7 of presumed gestation at approximately 10 and 30 minutes post dosage. Dosage-dependent clinical signs observed for surviving rabbits included ataxia, decreased motor activity, body jerks, tremors, vocalization (16 and 24 mg/kg/ day dosage groups), salivation, clonic convulsions, dyspnea, pin-point pupils, tales and reluctance to use hindlimbs (24 mg/kg/day dosage group). Pregnancy occurred in 4, 2, 1 and 3 rabbits in the control, low, middle and high dosage groups, respectively. Of these, one control rabbit aborted, and two high dosage group rabbits died. Maternal body weight and caesarean-sectioning data were inconclusive. The single surviving pregnant rabbit in the high dosage group had a larger number of resorptions and fewer live fetuses, as compared with control group rabbits, a possible agent-related effect. All fetuses from litters of does given test substance appeared normal at gross external examination. Two fetuses, one from each of two control litters, were hydrocephalic (domed heads).

Examinations

Maternal examinations:

OBSERVATIONS: Female rabbits were observed for physical signs and/or general appearance several times during the prestudy period and on days 0, 3 and 6 of presumed gestation. Observations for physical signs of agent effect, abortion and/or viability were made several times each day during the administration period (days 7 through 19 of presumed gestation) and daily during the post administration period (days 20 through 29 of presumed gestation).


BODY WEIGHT
- Time schedule for examinations: Day after arrival, once weekly during the acclimation period, on days 0 and 6 of presumed gestation, and daily during the administration period (days 7 through 19 of presumed gestation) and Post administration period (days 20 through 29 of presumed gestation) .

FOOD CONSUMPTION: During the study rabbits were observed for anorexia.

POST-MORTEM EXAMINATIONS
- Sacrifice on gestation day # 29
- Organs examined: Lungs, uterus and gross lesions

Ovaries and uterine content:

The ovaries and uterine content was examined after termination: Yes
Examinations included:
- Weight of each live fetus
- Number of corpora lutea
- Number of implantations
- Number and location of early and late resorptions
- Number and location of all live and dead fetuses
- Other: Any gross anomaly

Fetal examinations:

- External examinations: Yes
- Soft tissue examinations: Yes: [all per litter]
- Skeletal examinations: Yes: [all per litter]

Statistics:

Maternal physical sign data and the incidence of pregnancy and abortion were analyzed using Fisher's exact test.
The one-way analysis of variance was used to evaluate average body weights of rabbits assigned to the four dosage groups on days 0 and 6 of presumed gestation. Maternal body weight data for days 0, 7, 11, 15, 20, 24, 29 and 29C (corrected) of pregnancy a were analyzed using Bartlett's test of homogeneity of variances, and either the one-way analysis of variance or the Mann-Whitney U test. If Bartlett's test was positive (P ≤0.05), then the Mann-Whitney U test was used to analyze the data by testing each group individually against the control dosage group.
If Bartlett's test was negative (P >0.05), then the one-way analysis of variance was used to analyze the data. If the F-ratio was positive (P ≤0.05), then Dunnett's test was used to test each group individually against the control dosage group.
Data obtained during Caesarean-sectioning and for malformations and variations was evaluated using Jonckheere's test with the population of affected fetuses per litter considered to be the basic experimental unit. If Jonckheere's test was positive (P ≤0.05), and if there were fewer than 75% ties in a group, then the Mann-Whitney U test was used to analyze the data by testing each group individually against the control dosage group. If Jonckheere' s test was positive (P ≤0.05), and if there were more than 75% ties in a group, then Fisher's exact test was used with fixed point censoring to evaluate the data.

Results and discussion

Results: maternal animals

General toxicity (maternal animals)

Clinical signs:

effects observed, treatment-related

Description (incidence and severity):

CNS stimulation occurred for high dosage group rabbits and was considered to be an effect of the test substance. Dosage-dependent increases (P =0.047 to P <0.0001) in incidence of tremors, hyperactivity, body jerks, excess salivation, aggressive behavior (stamping of hind feet), hyperpnea, and convulsions were observed for high dosage group rabbits, as compared with control. Dosage-dependent increased incidences were also observed for ataxia, impaired or lost righting reflex, self-inflicted or spontaneous lesions, dyspnea, hyper sensitivity, pin-point pupils and vocalization. These incidences were not significantly different (P >0.05) from those observed for control rabbits.

Mortality:

mortality observed, treatment-related

Description (incidence):

Test substance related deaths occurred for one rabbit in the 6 mg/kg/day dosage group and 7 (P =0.003) rabbits in the 16 mg/kg/ day dosage group. These deaths generally followed observed evidence of central nervous system (CNS) stimulation.

Body weight and weight changes:

effects observed, treatment-related

Description (incidence and severity):

A small, dosage-related increase (P <0.05) in maternal body weight gain occurred between days 0 and 29 of gestation; however, only the gain of the high dosage group differed significantly from the control value. This difference was due to the decreased incidence of anorexia of the dams of the high dosage group during the latter portion of gestation.

Gross pathological findings:

effects observed, treatment-related

Description (incidence and severity):

Postmortem observations included a perforated stomach and evidence of excess salivation in the middle dosage group rabbit. Excess salivation was also observed for high dosage group rabbits, and the incidence was significantly increased (P =0.008), as compared with the control group.

Other effects:

effects observed, treatment-related

Description (incidence and severity):

As compared with the control group, the incidence of anorexia was significantly decreased (P =0.009) for rabbits in the high dosage group. This observation was considered to be a rebound in feed consumption after the dosage period. During the last part of gestation, spontaneous anorexia did occur among the control, low and middle dosage group rabbits.

Maternal developmental toxicity

Number of abortions:

no effects observed

Description (incidence and severity):

The incidence of pregnancy and number of corpora lutea per dam were similar for all dosage groups.

Pre- and post-implantation loss:

no effects observed

Description (incidence and severity):

The incidence of implantations per dam were similar for all dosage groups.

Total litter losses by resorption:

no effects observed

Description (incidence and severity):

The incidence of resorptions per dam were similar for all dosage groups.

Changes in number of pregnant:

no effects observed

Description (incidence and severity):

The incidence of pregnancy per dam were similar for all dosage groups.

Effect levels (maternal animals)

Remarks on result:

other: maternal toxicity was expressed as death and CNS stimulation

Results (fetuses)

Fetal body weight changes:

effects observed, non-treatment-related

Description (incidence and severity):

A dosage-related increase in average fetal body weight occurred; the difference from the control value was statistically significant only for the middle (P =0.025) and high (P =0.003) dosage groups. This increase in fetal body weight was not considered to be an adverse effect, since it was not observed to be due to the presence of edema. It probably was due to increased feed consumption of the dams during the latter part of gestation.

Reduction in number of live offspring:

no effects observed

Changes in sex ratio:

no effects observed

Changes in litter size and weights:

no effects observed

External malformations:

no effects observed

Skeletal malformations:

no effects observed

Visceral malformations:

no effects observed

Details on embryotoxic / teratogenic effects:

Administration of 2, 6 and 16 mg/kg/day dosages of the test substance to does did not result in adverse effects on embryo/ fetal viability or fetal sex ratio. Similarly, no gross external, soft tissue or skeletal malformations, developmental variations or variations due to retarded development observed for fetuses were attributed to administration of the test substance to the does.

Effect levels (fetuses)

Key result

Remarks on result:

other: Administration of aqueous solutions of the test substance to pregnant rabbits at dosages of 2, 6 and 16 mg/kg/day on days 7 through 19 of gestation was neither teratogenic nor embryo-fetal toxic, even in the presence of maternal toxicity.

Overall developmental toxicity

Key result

Developmental effects observed:

no

Applicant's summary and conclusion

Conclusions:

Administration of aqueous solutions of the test substance to pregnant rabbits at dosages of 2, 6 and 16 mg/kg/day on days 7 through 19 of gestation was neither teratogenic nor embryo-fetal toxic, even in the presence of maternal toxicity.

Executive summary:

The purpose of this study was to determine the embryofetal toxicity and the teratogenic potential of the test substance in the rabbit. Solutions of the test substance in reverse osmosis processed deionized water were administered by gavage on days 7 through 19 of presumed gestation to artificially-inseminated rabbits which had been randomly assigned to each of four dosage groups (20 rabbits/group).

The test substance as administered to rabbits at dosages of 0 (vehicle), 2, 6 and 16 mg/kg/day (calculated as the active ingredient) at a dosage volume of 5 mL/kg/day.

Agent-related deaths occurred for one rabbit in the 6 mg/kg/day dosage group and seven (P =0.003) rabbits in the 16 mg/kg/ day dosage group. These deaths generally followed observed evidence of central nervous system (CNS) stimulation.

One low dosage group rabbit aborted on day 27 of gestation, and one middle dosage group rabbit delivered on day 29 of gestation. These occurrences were considered to be spontaneous.

CNS stimulation occurred for high dosage group rabbits and was considered to be an effect of the test agent. Dosage-dependent increases (P =0.047 to P <0.0001) in incidence of tremors, hyperactivity, body jerks, excess salivation, aggressive behavior (stamping of hind feet), hyperpnea, and convulsions were observed for high dosage group rabbits, as compared with control. Dosage-dependent increased incidences were also observed for ataxia, impaired or lost righting reflex, self-inflicted or spontaneous lesions, dyspnea, hyper sensitivity, pin-point pupils and vocalization. These incidences were not significantly different (P >0.05) from those observed for control rabbits.

Postmortem observations included a perforated stomach and evidence of excess salivation in the middle dosage group rabbit. Excess salivation was also observed for high dosage group rabbits, and the incidence was significantly increased (P =0.008), as compared with the control group.

As compared with the control group, the incidence of anorexia was significantly decreased (P =0.009) for rabbits in the high dosage group. This observation was considered to be a rebound in feed consumption after the dosage period. During the last part of gestation, spontaneous anorexia did occur among the control, low and middle dosage group rabbits.

A small, dosage-related increase (P <0.05) in maternal body weight gain occurred between days 0 and 29 of gestation; however, only the gain of the high dosage group differed significantly from the control value. This difference was due to the decreased incidence of anorexia of the dams of the high dosage group during the latter portion of gestation.

A significant decrease (P =0.024) in maternal body weight was observed among the dams in the 2 mg/kg/day dosage group on days 0 to 7, as compared with that for the control group. This decrease was not test substance-related, since it occurred before the dosage period.

The incidence of pregnancy and number of corpora lutea, implantations and resorptions per dam were similar for all dosage groups.

A dosage-related increase in average fetal body weight occurred; the difference from the control value was statistically significant only for the middle (P =0.025) and high (P =0.003) dosage groups. This increase in fetal body weight was not considered to be an adverse effect, since it was not observed to be due to the presence of edema. It probably was due to increased feed consumption of the dams during the latter part of gestation.

Administration of 2, 6 and 16 mg/kg/day dosages of the test substance to does did not result in adverse effects on embryo/ fetal viability or fetal sex ratio. Similarly, no gross external, soft tissue or skeletal malformations, developmental variations or variations due to retarded development observed for fetuses were attributed to administration of the test substance to the does.

On the basis of these data, administration of aqueous solutions of the test substance to pregnant rabbits at dosages of 2, 6 and 16 mg/kg/day on days 7 through 19 of gestation was neither teratogenic nor embryo-fetal toxic, even in the presence of maternal toxicity. The maternal toxicity was expressed as death and CNS stimulation.