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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian germ cell study: cytogenicity / chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other:

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1980
Report date:
1980

Materials and methods

Test guideline
Qualifier:
equivalent or similar to guideline
Guideline:
OECD Guideline 478 (Genetic Toxicology: Rodent Dominant Lethal Test)
GLP compliance:
no
Type of assay:
rodent dominant lethal assay

Test material

Constituent 1
Chemical structure
Reference substance name:
4-methylstyrene
EC Number:
210-762-8
EC Name:
4-methylstyrene
Cas Number:
622-97-9
Molecular formula:
C9H10
IUPAC Name:
1-methyl-4-vinylbenzene
Test material form:
liquid
Specific details on test material used for the study:
Lot/batch No.: 03248001-B1 & 03248001-B2

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals or test system and environmental conditions:
- Source: Charles River Breeding Farms, Quebec, Canada
- Age at study initiation: 10-12 weeks
- Weight at study initiation: Males, 265-370 g; Females, not specified
- Assigned to test groups: Randomly; basis not specified
- Housing: Males, 3 per cage during acclimation; one per cage thereafter. Females, 5 per cage during acclimation; 2-4 per cage after mating. Plastic cages with hardwood chip bedding
- Diet: Certified laboratory rodent chow, ad libitum
- Water: ad libitum
- Acclimation period: 10-14 days
- Temperature: Approximately 23 °C
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle: Olive oil, for controls only

Details on exposure:
Three groups of 10 males were given the test substance by oral gavage for 5 consecutive days at doses of 0.15, 0.5 or 1.5 mL/kg/day. A further group of 10 males was given olive oil (negative control) by oral gavage at 1.0 mL/kg/day for 5 consecutive days. A fifth group of 10 males was given a single dose of the positive control (triethylenemelamine) at 0.5 mg/kg by intraperitoneal injection. Three days after the last dose, each male was mated with 2 virgin females over a 5-day period. Males were then allowed to rest for 2 days. Subsequently, the mating process was repeated with additional virgin females until the males had been mated for 7 weeks, 2 females per week.
14-15 days from the mid-point of the mating period, the females were sacrificed by CO2 asphyxiation. The abdominal cavity was exposed and the membrane removed from each ovary. The corpora lutea were counted for each ovary and recorded separately. The uterine contents were examined and the number of live and dead implants was recorded for each uterine horn. The criteria for a living foetus were viability at necropsy, general colour, and absence of maceration; a dead implant was that present at the implantation site as a non-viable foetus reduced in size or as a necrotic embryonic mass. Males were sacrificed after completion of the mating period, but no necropsies were performed.
Duration of treatment / exposure:
Males only, 5 days
Frequency of treatment:
Males, once daily for 5 days (positive control, one dose only)
Post exposure period:
Males mated for 7 weeks post-exposure
Doses / concentrationsopen allclose all
Dose / conc.:
0.15 other: mL/kg bw/day
Dose / conc.:
0.5 other: mL/kg bw/day
Dose / conc.:
1.5 other: mL/kg bw/day
No. of animals per sex per dose:
5 groups of 10 males (Control, 3 doses of test substance, positive control), plus females for mating.
Control animals:
yes, concurrent vehicle
Positive control(s):
Triethylenemelamine
- Justification for choice of positive control(s): Not specified, but included in OECD 478
- Route of administration: Intraperitoneal injection, once only
- Doses / concentrations: 0.5 mg/kg bw in distilled water (concentration not given)

Examinations

Evaluation criteria:
Criteria for determination of a valid test:
Female Sprague-Dawley rats mated with negative control males must show an average of no less than 8 implantations per pregnant female. Females mated to positive control males must exhibit severe foetal damage, i.e. a statistically significant reduction in implantations relative to the negative controls and a statistically significant increase in females with 2 or more dead implants relative to the negative controls. This damage must be noted between weeks 2 and 7 of the spermatogenic cycle.
Statistics:
Dominant lethality was evaluated according to the following criteria, which were analysed statistically by recognised methods:
Fertility Index (the number of fertile matings)
Total implantations per pregnant female
Number of corpora lutea per pregnant female
Pre-implantation losses per pregnant female (difference between the number of corpora lutea and the number of total (live and dead) implantations
Dead implants per pregnant female
The proportion of pregnant females with one or more dead implants, and with two or more dead implants
Dead implants per total implants
Live implants per pregnant female

Results and discussion

Additional information on results:
Other than body weight losses / reduced weight gain in males given the test substance, particularly at 0.5 and 1.5 mL/kg/day, there were no remarkable clinical findings. The pregnancy data for females mated with males previously dosed with the positive control, triethylenemelamine, revealed expected findings, including: markedly reduced implantations for matings performed in weeks 1-4, and, for matings performed in weeks 1-5, markedly increased pre-implantation losses, increased number of dead implants per pregnancy, increased proportion of pregnant females with one/two or more dead implants, increased number of dead implants per total implants, and reduced number of live implants per pregnant female. Comparison of the pregnancy data for females mated with males previously dosed with the test substance with that of females mated with males given the negative control did not reveal any differences attributable to treatment with the test substance.

Applicant's summary and conclusion

Conclusions:
Under the study conditions, treatment of males with the test substance and their subsequent mating over a 7 week period (covering the spermatogenic cycle) with untreated virgin females did not result in any evidence of mutagenic potential.
Executive summary:

A study was conducted to determine the cytogenicity and chromosome aberration potential of the test substance according to a method similar to OECD Guideline 478. Three groups of 10 male Sprague Dawley rats were given the test substance by oral gavage for 5 consecutive days at doses of 0.15, 0.5 or 1.5 mL/kg bw/day. A further group of 10 males was given olive oil (negative control) by oral gavage at 1.0 mL/kg bw/day for 5 consecutive days. A fifth group of 10 males was given a single dose of the positive control (triethylenemelamine) at 0.5 mg/kg bw by intraperitoneal injection. Three days after the last dose, each male was mated with 2 virgin females over a 5 d period. Males were then allowed to rest for 2 d. Subsequently, the mating process was repeated with additional virgin females until the males had been mated for 7 weeks, 2 females per week. Fourteen to fifteen days from the mid-point of the mating period, the females were sacrificed by CO2 asphyxiation. The abdominal cavity was exposed and the membrane removed from each ovary. The corpora lutea were counted for each ovary and recorded separately. The uterine contents were examined and the number of live and dead implants was recorded for each uterine horn. The criteria for a living foetus were viability at necropsy, general colour, and absence of maceration; a dead implant was that present at the implantation site as a non-viable foetus reduced in size or as a necrotic embryonic mass. Males were sacrificed after completion of the mating period, but no necropsies were performed. The pregnancy data for females mated with males previously dosed with the positive control revealed expected findings. Comparison of the pregnancy data for females mated with males previously dosed with the test substance with that of females mated with males given the negative control did not reveal any differences attributable to treatment with the test substance. Under the study conditions, treatment of males with the test substance and their subsequent mating over a 7 week period (covering the spermatogenic cycle) with untreated virgin females did not result in any evidence of mutagenic potential (Putman, 1980).