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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian somatic cell study: cytogenicity / erythrocyte micronucleus
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1982
Report Date:
1982

Materials and methods

Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
yes
Remarks:
(only 3 animals each sex instead of 5; no data on test material purity, animal source and acclimatisation period; proportion polychromatic erythrocytes/normochromatic erythrocytes was not documented, only 1000 cells per slide were counted)
GLP compliance:
no
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Physical state: Solid

Test animals

Species:
hamster, Chinese
Strain:
not specified
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Weight at study initiation: Males 24-32 g, females 21-31 g
- Fasting period before study: No data.
- Housing: Individual caging
- Diet: Standard diet (NAFAG No. 924)
- Water: Tap water, ad libitum

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 22
- Humidity (%): 59 - 70
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle used: Arachid oil
- Amount of vehicle: 20 ml/kg bw
Duration of treatment / exposure:
2 consecutive days
Frequency of treatment:
1 application daily
Post exposure period:
24 h
Doses / concentrationsopen allclose all
Dose / conc.:
750 mg/kg bw/day (actual dose received)
Dose / conc.:
1 500 mg/kg bw/day (actual dose received)
Dose / conc.:
3 000 mg/kg bw/day (actual dose received)
No. of animals per sex per dose:
3
Control animals:
yes, concurrent vehicle
Positive control(s):
Cyclophosphamide
- Route of administration: Orally (gavage)
- Doses / concentrations: 128 mg/kg bw in 20 ml/kg bw arachid oil

Examinations

Tissues and cell types examined:
Bone marrow was harvested from the shafts of both femurs.
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION:
The oral LD50 was found to be > 3000 mg/kg bw in Chinese hamsters of either sex (CIBA-Geigy Ltd., No. 820742, 1982, see chapter 7.2.1)

DETAILS OF SLIDE PREPARATION:
Bone marrow was harvested from the shafts of both femurs. In a siliconised pipette filled with approx. 0.5 µl rat serum the bone marrow was drawn up. In order to receive a homogeneous suspension the content of pipette was aspirated gently about three times. Small drops of the mixture were transferred on the end of a slide, spread out by pulling it behind a polished cover glass and the preparations were air-dried. Three hours later, the slides were stained in undiluted May-Grünwald solution for 2 minutes then in May-Grünwald solution/water 1/1 for 2 minutes and then in Giemsa's, 40 % for 20 minutes. After being rinsed in methanol 55 % for 5-8 seconds and washed off twice in water, they were left immersed in water for approx. 2 minutes. After rinsing with distilled water and air-drying, the slides were cleared in Xylol and mounted in Eukitt.

METHOD OF ANALYSIS:
The slides of 3 female and 3 male animals each of the negative control group, the positive control group and of the groups treated with various doses of the test substance were examined. 1000 bone marrow cells each were scored per animal and the following anomalies were registered:
a) Single Jolly bodies, b) fragments of nuclei in erythrocytes, c) micronuclei in erythroblasts, d) micronuclei in leucopoietic cells, e) polyploid cells.
Statistics:
The significance of difference was assessed by χ2 -test.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Vehicle controls validity:
valid
Negative controls validity:
not examined
Positive controls validity:
valid
Additional information on results:
RESULTS OF DEFINITIVE STUDY
In all dosage groups the percentage of cells displaying anomalies of nuclei did not differ significantly from the negative control.By contrast, the positive control (cyclophosphamide, 128 mg/kg) yielded a marked increase of the percentage of cells with anomalies. Here the mean percentage of anomalies was 8.72, whereas the negative control yielded a percentage of 0.10. The difference is highly significant (p<0.05).

Any other information on results incl. tables

Tab. 1: The effect of the test substance and cyclophosphamide on bone marrow cells of Chinese hamster (animals sacrificed 24 hours after second application)

f = female, m = male

 

concentration

Animal No.

Sex

Single Jolly bodies

Fragments of nuclei in erythrocytes

Micronuclei in erythrocytes

Micronuclei in Leucopoietic cells

Polyploid cells

Total

 

 

Control (Arachid oil)

 

1

f

0.3

 

 

 

 

0.3

2

f

 

 

 

 

 

0.0

3

f

 

 

 

 

 

0.0

4

m

0.1

 

 

 

 

0.1

5

m

 

 

 

 

 

0.0

6

m

0.2

 

 

 

 

0.2

 

 

Cyclophosphamide

 

 

128 mg/kg bw

1

f

10.3

1.4

1.9

0.5

 

14.1

2

f

8.6

0.7

2.3

0.3

0.1

12.0

3

f

7.5

0.9

1.2

0.5

0.1

10.2

4

m

3.4

0.2

0.7

0.2

 

4.5

5

m

3.1

0.1

1.3

0.4

 

4.9

6

m

4.9

0.4

1.3

 

 

6.6

 

 

 

 

 

 

 

 

Test substance

 

 

750 mg/kg bw

1

f

 

 

 

 

 

0.0

2

f

0.2

 

 

 

 

0.2

3

f

0.3

 

 

 

 

0.3

4

m

0.2

 

 

 

 

0.2

5

m

0.1

 

 

 

 

0.1

6

m

 

 

 

 

 

0.0

 

 

1500 mg/kg bw

1

f

0.2

 

 

 

 

0.2

2

f

 

 

 

 

 

0.0

3

f

0.1

 

 

 

 

0.1

4

m

 

 

 

 

 

0.0

5

m

0.1

 

 

 

 

0.1

6

m

 

 

 

 

 

0.0

 

 

3000 mg/kg bw

1

f

0.1

 

 

 

 

0.1

2

f

0.2

 

 

 

 

0.2

3

f

0.1

 

 

 

 

0.1

4

m

 

 

 

 

 

0.0

5

m

0.1

 

 

 

 

0.1

6

m

0.2

 

 

 

 

0.3

Applicant's summary and conclusion

Conclusions:
Under the conditions of this experiment, no evidence of mutagenic effects was obtained in Chinese hamsters treated with the test item.