Iso(C10-C14)alkyl (3,5-di-tert-butyl-4-hydroxyphenyl)methylthioacetate

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:

1989-07-04 to 1989-02-07

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: OECD guideline study, GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1989

Report date:

1989

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OTS 798.5395 (In Vivo Mammalian Cytogenics Tests: Erythrocyte Micronucleus Assay)

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: EEC (19 September 1984), Mutagenicity (Micronucleus Test). Official Journal of the European Communities No L 251 137-139.

GLP compliance:

yes

Type of assay:

micronucleus assay

Test material

Test material information

Constituent 1

Reference substance name:

Iso(C10-C14)alkyl (3,5-di-tert-butyl-4-hydroxyphenyl)methylthioacetate

EC Number:

404-800-4

EC Name:

Iso(C10-C14)alkyl (3,5-di-tert-butyl-4-hydroxyphenyl)methylthioacetate

Cas Number:

118832-72-7

Molecular formula:

C30H52O3S

IUPAC Name:

11-methyldodecyl 2-{[(3,5-di-tert-butyl-4-hydroxyphenyl)methyl]sulfanyl}acetate

Test animals

Species:

hamster

Strain:

other: Cricetulus griseus

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: CIBA-GEIGY Tierfarm, Sisseln
- Weight at study initiation: females 31-34 g, males 30-35 g (tolerability test), females 21-31 g, males 22-34 g (mutagenicity test).
- Assigned to test groups randomly: yes
- Housing: Individually
- Diet: ad libitum (NAFAG No. 924)
- Water: tap water, ad libitum
- Acclimation period: At least 3 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 15 - 23
- Humidity (%): 50 - 84
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

- Vehicle(s)/solvent(s) used: arachis oil

Details on exposure:

The preparation was administered orally to groups of 24 female and 24 male animals each in the negative and in the 5000 mg/kg bw dose group. The positive control group consisted of 8 female and 8 male animals. Treatment consisted of a single application. 16, 24 and 48 hours after application 8 female and 8 male animals per sampling time were sacrificed by dislocation of the cervical vertebrae.

Duration of treatment / exposure:

Single exposure

Frequency of treatment:

Once

Post exposure period:

16, 24, 48 hrs

Doses / concentrations

Remarks:

Doses / Concentrations:
5000 mg/kg bw
Basis:
nominal conc.

No. of animals per sex per dose:

In the tolerability test: 2 females and 2 males
In the mutagenicity test: 24 females and 24 males in the treatment groups and in the negative control groups. 8 females and 8 males in the positive control group

Control animals:

yes, concurrent vehicle

Positive control(s):

yes (CPA; Cyclophosphamide)

Examinations

Tissues and cell types examined:

Bone marrow was harvested from the shafts of both femurs with fetal calf serum. After centrifugation small drops of the sediment mixture were transferred on the end of a slide, spread out with the aid of a polished cover glass and air-dried. Within 24 hours, the slides were stained in undiluted May-Grünwald solution for 3 min then in May-Grünwald solution/water 1/1 for 2 min. After being rinsed in distilled water, the slides were left immersed in diluted Giemsa solution (16.6%), for 10 min. After rinsing with distilled water and air-drying, the slides were cleared in Xylene and mounted.

Details of tissue and slide preparation:

The slides of five animals from each sex showing the best differentiation between mature and polychromatic erythrocytes were selected for later scoring. The slides of five female and five male animals each of the negative control group and of the dosage group sacrificed at 16, 24 and 48 hours post treatment were examined. From the animals of the positive control group which were sacrificed 24 hours after application, the slides of five female and five male animals were scored. 1000 polychromatic erythrocytes per animal each were scored for the incidence of micronuclei. To determine the mitotic activity of the red compartment, the ratio of polychromatic to normochromatic erythrocytes was calculated for each animal by counting a total of 1000 erythrocytes. A low proportion of polychromatic erythrocytes is indicative for a mitosis inhibiting activity of the test substance.

Evaluation criteria:

- The quality of the slides must allow a clear differentiation between polychromatic and normochromatic erythrocytes.
- The result obtained with the positive control has to fulfill the criteria given for a positive response.

Statistics:

The significance of difference was assessed by X2-test.

Results and discussion

Test results

Sex:

male/female

Genotoxicity:

negative

Toxicity:

no effects

Vehicle controls validity:

valid

Negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RESULTS OF RANGE-FINDING STUDY
Three groups of four Chinese hamsters (two females and two males) are treated with three different doses, one receiving the maximum dose of 5000 mg/kg, or the highest applicable dose, and the other two doses of 1/5 and 1/25 of that amount respectively. The animals are treated with a single dose. The observation period corresponds to the interval between administration and sacrifice of the animals in the mutagenicity test, plus one day. In this experiment the dose of 5000 mg/kg was determined as the highest applicable in the mutagenicity assay

POSITIVE CONTROL
The positive control (cyclophosphamide, 64 mg/kg, sampling time 24 hours) yielded a marked increase of the percentage of micronucleated cells. Here the mean percentage of polychromatic erythrocytes with micronuclei was 1.00. In comparison with the negative control (0.08) this value is highly significant (p <0.05), confirming an appropriate sensitivity of the test system.

Any other information on results incl. tables

Experimental Result

Sacrifice	Treatment	Sex	polychromatic erythrocytes (average)	normochromatic erythrocytes (average)	ratio of p / n erythrocytes	number of polychromatic erythrocytes with micronuclei (average)	% of polychromatic erythrocytes with micronuclei (average)
16 h	Control	female	510	490	1	0.6	0.06
		male	603	397	1.5	0	0
	5000 mg/kg	female	548	452	1.2	0.6	0.06
		male	632	368	1.7	0.8	0.08
24 h	Control	female	512	488	1	1	0.1
		male	573	427	1.3	0.6	0.06
	5000 mg/kg	female	544	456	1.2	0.4	0.04
		male	657	343	1.9	0.4	0.04
48 h	Control	female	581	419	1.4	0.6	0.06
		male	648	352	1.8	0.2	0.02
	5000 mg/kg	female	485	515	0.9	0.8	0.08
		male	605	395	1.5	0.4	0.04
Positive Control
48 h	Control	female	512	488	1	1	0.1
		male	573	427	1.3	0.6	0.06
	64 mg/kg	female	523	477	1.1	12.6	1.26
		male	581	419	1.4	7.4	0.74

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information): negative

Executive summary:

The test item was assessed in the micronucleus assay for its potential to induce micronuclei in polychromatic erythrocytes (PCE) in the bone marrow cells of the Chinese hamster. The test item was administered orally to groups of 24 female and 24 male animals at 0 (vehicle control) or 5000 mg/kg bw. 16, 24 and 48 hours after application 8 females and 8 males per sampling time were sacrificed and bone-marrow smears were prepared. There was no significant increase in the number of micronucleated polychromatic erythrocytes in the animals treated with the dose of 5000 mg/kg bw of the test item as compared with the negative control animals at all three sampling times. The positive control group (cyclophosphamide) consisted of 8 female and 8 male animals and showed the appropriate response, thus confirming sensitivity of the test system. In conclusion, it can be stated that during the study described and under the experimental conditions reported, the test item did not induce micronuclei as determined by the micronucleus test in the bone marrow cells of the Chinese hamster.