Propanoic acid, 3-[[bis(2-methylpropoxy)phosphinothioyl]thio]-2-methyl-

Genetic toxicity: in vivo

Administrative data

Endpoint:

in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

May 10, 2000 - June 13, 2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

Guideline study conducted according to GLP. Deviations from the protocol were stated (example given: relative humidity under which the experiment was conducted ranged between 18-70 % and not between 30-70 % as described in the protocol) which however, did not affect the validity of the experiment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2000

Report date:

2000

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Version / remarks:

(1997)

Deviations:

yes

Remarks:

Deviations from the protocol were stated (example given: relative humidity under which the experiment was conducted ranged between 18-70 % and not between 30-70 % as described in the protocol) which however, did not affect the validity of the experiment.

GLP compliance:

yes (incl. QA statement)

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

-

EC Number:

434-070-2

EC Name:

-

Cas Number:

268567-32-4

Molecular formula:

C12 H25 O4 P S2

IUPAC Name:

3-{[bis(2-methylpropoxy)(sulfanylidene)-λ⁵-phosphanyl]sulfanyl}-2-methylpropanoic acid

Test material form:

liquid: viscous

Test animals

Species:

mouse

Strain:

NMRI

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd. Biotechnology & Animal Breeding Division CH-4414 Füllinsdorf
- Age at study initiation: 8 - 10 weeks
- Weight at study initiation: 33 +/- 2.7 g (males), 28.4 +/- 1.8 g (females)
- Assigned to test groups randomly: yes
- Housing: individually in Makrolon type-I cages
- Diet (ad libitum): pelleted standard diet, ALTROMIN 1324
- Fasting period before study: 18 h
- Water (ad libitum): tap water
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 21 +/- 4
- Humidity (%): 18 - 70
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle: corn oil
- Concentration of test material in vehicle: In a pretest 2000 mg/kg bw, 1800 mg/kg bw in main study
- Application volume: 10 mL/kg bw

Duration of treatment / exposure:

After single oral administration the following dose levels of the test item were investigated:
24 h preparation interval: 450, 900 and 1800 mg/kg bw
48 h preparation interval: 1800 mg/kg bw

Frequency of treatment:

Single oral administration

Post exposure period:

MAIN STUDY:
450, 900 and 1800 mg/kg bw: 24 h
1800 mg/kg bw: 48 h

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

450 mg/kg bw/day (actual dose received)

Remarks:

observation period 24 h

Doses / concentrations 2

Dose / conc.:

900 mg/kg bw/day (actual dose received)

Remarks:

observation period 24 h

Doses / concentrations 3

Dose / conc.:

1 800 mg/kg bw/day (actual dose received)

Remarks:

observation period 24 h and 48 h

No. of animals per sex per dose:

Six animals per sex per dose (5 per sex were evaluated)

Control animals:

yes, concurrent no treatment

Positive control(s):

- Positive Control: 40 mg/kg bw Cyclophosphamide
- Volume administered: 10 mL/kg bw
- Route of administration: single oral application

Examinations

Tissues and cell types examined:

Bone marrow was harvested from the femurs. 2000 PCE per animal were used to determine the incidence of micronucleated PCE. Normochromatic erythrocytes (NCE) as well as polychromatic erythrocytes (PCE) were counted in the same sample for determination of the PCE/NCE-ratio.

Details of tissue and slide preparation:

CRITERIA FOR DOSE SELECTION:
In a pre-test with 2000 mg/kg bw a female died 24 h after application of CG 37-1586 per gavage. In a second pre-test with 4 animals (2 males, 2 females) no mortality was observed within 48 h after administration of 1800 mg/kg bw test article. Therefore, 1800 mg/kg bw CG 37-1586 was selected as maximum tolerated dose level.

DETAILS OF SLIDE PREPARATION:
Bone marrow was harvested from the shafts of femurs. Preparations were air-dried and stained in May-Grünwald/Giemsa's. Cover slips were mounted in Eukitt.

Evaluation criteria:

In this report, a test article was classified as mutagenic if it induces either a dose-related increase in the number of micronucleated polychromatic erythrocytes or a statistically significant and biologically relevant positive response.

A test item producing neither a dose-related increase in the number of micronucleated polychromatic erythrocytes nor a statistically significant and biologically relevant positive response is considered non-mutagenic.

Statistics:

The significance of difference was assessed by the non-parametric Mann-Whitney test.

Results and discussion

Test results

Key result

Sex:

male/female

Genotoxicity:

negative

Remarks:

PCEs with micronuclei 0.03, 0.05, 0.035 % after 24 h at 450, 900 and 1800 mg/kg bw, respectively versus 0.05 % in control. After 48 h at 1800 mg/kg bw the percentage was 0.015 %. Positive control (24 h) showed 1.67 % micronucleated PCE.

Toxicity:

yes

Remarks:

The number of NCE increased slightly at the dose level of 1800 mg/kg bw (2027 NCE/2000 PCE after 24 h; 2117 NCE/2000 PCE after 48 h) in comparison to vehicle control (1732 NCE/2000 PCE).

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Any other information on results incl. tables

In a pre-test with 2000 mg/kg bw a female died 24 h after application of the test article per gavage. In a second pre-test with 4 animals (2 males, 2 females) no mortality was observed within 48 h after administration of 1800 mg/kg bw test article. Therefore, 1800 mg/kg bw test material was selected as maximum tolerated dose level. However, animals at this dose level showed clinical signs of toxicity (reduction of spontaneous activity, eyelid closure, apathy, abdominal position). During main study one female died in the highest dose level within 24 h. It was replaced with a satellite animal so that the overall number of animals was sufficient.

No statistically significant increase in the frequency of micronuclei at any preparation interval and dose level after administration of the test item was seen. The mean values of micronuclei observed after treatment with the test item were below or as high as the value of the vehicle control group and within the historical control data range.

The percentage of micronucleated cells in the high dose group at the 48 h sampling time was below the historical data range. However, the authors stated that such values have been obtained in previous studies and do not affect the validity of the study.

 

The positive control (cyclophosphamide) showed a statistically significant increase of induced micronucleus frequency.

Summary of Micronucleus Test Results

Test group	Dose mg/kg bw	Sampling time (h)	PCEs with micronuclei (%)	Range*	PCE/NCE
Vehicle	0	24	0.050	0 - 3	2000 / 1732
Test item	450	24	0.030	0 - 2	2000 / 1638
Test item	900	24	0.050	0 -3	2000 / 1917
Test item	1800	24	0.035	0 - 2	2000 / 2027
Cyclo phosphamide	40	24	1.670	7 - 52	2000 / 1909
Test item	1800	48	0.015	0 - 2	2000 / 2117

* Number of micronucleated PCEs

Applicant's summary and conclusion

Conclusions:

Under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test with bone marrow cells of the mouse. Therefore, the test item is considered to be non-mutagenic in this micronucleus assay.