3-anilino-5-methyl-5-(4-phenoxyphenyl)-1,3-oxazolidine-2,4-dione;famoxadone (ISO)

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1994

Report date:

1994

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

EPA OPP 84-2

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 4

Qualifier:

according to guideline

Guideline:

other: Japan Ministry of Agriculture, Forestry and Fisheries Testing Guidelines for Toxicology Studies, 59 NohSan No. 4200

GLP compliance:

yes

Type of assay:

mammalian erythrocyte micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

5-methyl-5-(4-phenoxyphenyl)-3-(phenylamino)-1,3-oxazolidine-2,4-dione

EC Number:

603-520-1

Cas Number:

131807-57-3

Molecular formula:

C22H18N2O4

IUPAC Name:

5-methyl-5-(4-phenoxyphenyl)-3-(phenylamino)-1,3-oxazolidine-2,4-dione

Test material form:

solid

Specific details on test material used for the study:

Substance name: DPX-JE874
Lot#: JE874-221
Purity: 97.4%

Test animals

Species:

mouse

Strain:

other: Crl:CD®_1(ICR)BR

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Inc., Saint Constant, Quebec, Canada
- Age at study initiation: Approximately 53 days old
- Weight at study initiation: Male: 28.5-34.2 g; Female: 24.0-28.1 g
- Assigned to test groups randomly: Yes
- Housing: Individually
- Diet: ad libitum
- Water: ad libitum
- Acclimation period: 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 23 ± 2°C
- Humidity (%): 50 ± 10%
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

Corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS: Immediately prior to dosing, suspensions of the test substance in the vehicle were prepared at concentrations of 200, 100, and 50 mg/mL. Uniformity was maintained during dosing by constant stirring. Single acute doses of the appropriate suspensions were administered by oral intubation at 25 mL/kg, yielding treatments of 5000, 2500, and 1250 mg test substance/kg body weight.

Duration of treatment / exposure:

Single dose

Frequency of treatment:

Single dose

Post exposure period:

24, 48, 72 hours

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

1 250 mg/kg bw (total dose)

Doses / concentrations 2

Dose / conc.:

2 500 mg/kg bw (total dose)

Doses / concentrations 3

Dose / conc.:

5 000 mg/kg bw (total dose)

No. of animals per sex per dose:

6 (high dose group); 5 (rest of the groups)

Control animals:

yes, concurrent vehicle

Positive control(s):

The CP was administered by intraperitoneal injection in a volume of 5 mL/kg. An 8.0 mg/mL CP solution was used, yielding a dose of 40 mg/kg.

Examinations

Tissues and cell types examined:

Bone marrow cells

Details of tissue and slide preparation:

TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields): Mice were treated with either the test substance, vehicle or positive indicator on the day following release from quarantine. Groups of 6 male and 6 female mice (high dose group) and 5 male and 5 female mice (negative control) were sacrificed approximately 24, 48, and 72 hrs post-dosing. For the intermediate and low dose groups, 5 males and 5 females were sacrificed 24 hrs post-dosing. A positive indicator group of 5 male and 5 female mice was concurrently treated and sacrificed approximately 24 hrs post-dosing.

DETAILS OF SLIDE PREPARATION: Immediately after sacrifice, marrow from both femurs of each animal was aspirated and flushed into approximately 3 mL prewarmed (37°C) fetal bovine serum. The marrow was collected by centrifugation (approximately 200 x g, 5 min). Most of the supernatant was removed and the cells were resuspended in the remaining 1-2 drops of serum. A Miniprep® automatic blood smearing instrument was used to prepare marrow smears. At least 3 slides per animal were prepared and fixed in absolute methanol for 8 min. Slides were stained for 3 min in 0.0125 mg/mL acridine orange in phosphate buffer (pH 7.4). Prior to scoring, a coverslip was floated on each slide using phosphate buffer.

Statistics:

Data for the proportion of PCEs and MNPCEs among 2000 erythrocytes (PCE frequency and MNPCE frequency, respectively) were transformed prior analysis using the arcsin square root function. This transformation appropriate for proportions since the distribution of the transformed data more closely approximates a normal distribution than do the nontransformed proportion. Transformed data for PCE or MNPCE frequency were analyzed separately for normality of distribution using the Shapiro-Wilkes test. If results indicated that the transformed values for PCE:MNPCE frequency normally distributed in both sexes, parametric methods (viz., Analysis of Variance (AN0VA) and Dunnett test) were used. If there was nonnormality in either sex, nonparametric methods (viz., Kruskal-Wallis test and Mann-Whitney U tests) were used for that variable using nontransformed proportions. Positive indicator data were not included in evaluating normality of distribution. Weight gain data were assumed to be normally distributed and were analyzed by ANOVA. Data from each sex and sacrifice time were analyzed separately, and individual comparisons to the control were made using each animal as the experimental unit. All analyses conducted at a significance level of 5%. Positive indicator data were analyzed separately.

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

other: equivocal

Remarks:

equivocal based on the incidence of soft stools and diarrhoea which are likely to be related to the corn oil dosing vehicle (possibly affecting absorption of the test material in the treated mice).

Toxicity:

yes

Remarks:

wet perineum and/or soft stools, diarrhea

Vehicle controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

The test substance did not induce micronuclei in bone marrow cells of mice; the material is negative in this in vivo assay.

Executive summary:

This study was conducted following US EPA 84-2 and OECD guideline 474. The test substance evaluated for its ability to induce micronuclei in bone marrow polychromatic erythrocytes (PCEs) of Crl:CD®-1(ICR)BR mice. Acute doses of 0, 1250, 2500, and 5000 mg/kg administered by oral intubation to male and female mice. In the control and high dose groups, bone marrow smears were prepared approximately 24, 48, and 72 hrs after dosing. In the intermediate and low dose groups, prepared approximately 24 hrs after dosing. Two thousand PCEs per animal were scored for micronuclei.

No statistically significant increases in the frequency of micronucleated PCEs were observed in test substance treated mice at any dose level or sampling time. In addition, no statistically significant depression in the proportion of PCEs among 1000 erythrocytes were observed. In this assay, the test substance is negative.