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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
Remarks:
Type of genotoxicity: chromosome aberration
Type of information:
experimental study
Adequacy of study:
key study
Study period:
10 June 1991 to 8 November 1991
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study followed OECD Guideline 474

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent

Test animals

Species:
mouse
Strain:
NMRI
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Strain specifics: Hoe: NMRKf (SPF71)
- Source: Hoechst AG, Kastengrund, SPF breeding colony
- Age at study initiation: 7 weeks
- Weight at study initiation: males: 25-35 g, mean 29.7 g, females: 22-28 g, mean 24.9 g
- Housing: in groups of five in Macrolon type 3 cages in fully air-conditioned room, softwood granulate
- Diet (e.g. ad libitum): rat/mouse diet, ad libitum
- Water (e.g. ad libitum): tap water, ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 22 ± 2 °C
- Humidity (%): 55 ± 10 %
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle(s)/solvent(s) used: sesame oil
- Concentration of test material in vehicle: 25% (w/v)
- Amount of vehicle (if gavage or dermal): 10 ml/kg body weight
- Purity: Oleum sesame Ph. Eur. III, Fa. Pharm. Fabrik GmbH, Frankfurt/Main, Germany
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
25% was the highest concentration achievable in sesame oil
Duration of treatment / exposure:
One single oral administration
Frequency of treatment:
See above
Post exposure period:
24 hours (dosed group, negative control and positive control), 48 hours (dosed group and negative control) and 72 hours (dosed group and negative control)
Doses / concentrations
Remarks:
Doses / Concentrations:
2500
Basis:
nominal conc.
mg/kg body weight
No. of animals per sex per dose:
5 males and 5 females (10 animals) per dose group
Control animals:
yes, concurrent vehicle
Positive control(s):
50 mg/kg body weight cyclophosphamide (endoxan, 5 males and 5 females)

Examinations

Tissues and cell types examined:
1000 polychromatic erythrocytes from the femoral bone marrow of each animal
Details of tissue and slide preparation:
TREATMENT AND SAMPLING TIMES ( in addition to information in specific fields):
Animals were killed 24, 48 or 72 h after application

DETAILS OF SLIDE PREPARATION:
Removal of femoral bones and bone marrow from the proximal ends flushed into centrifuge tube containing about 3 ml foetal bovine serum, centrifugation (5 min at 1200 rpm), one drop of thoroughly mixed sediment smeared on a cleaned slide, air-dried for approx. 24 h, staining (methanol, May-Grünwalds solution, Giemsa) and coating with Entellan

METHOD OF ANALYSIS:
Number of cells with micronuclei and ratio of polychromatic to normochromatic erythrocytes
Statistical evaluation (see below)
Evaluation criteria:
95% level of significance for comparisons
Statistics:
Wilcoxon (paired, one-sided, increase) for number of cells and Wilcoxon (paired, two-sided) for the ratio

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Toxicity:
no effects
Remarks:
red coloured faeces but free of clinical signs of toxicity after 48 hours
Vehicle controls validity:
valid
Positive controls validity:
valid
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
- Dose range: 2500 mg/kg body weight
- Clinical signs of toxicity in test animals: red coloured faeces
- Other: lethality: 0 out of 3 males and 0 out of 3 females

RESULTS OF DEFINITIVE STUDY
- Induction of micronuclei (for Micronucleus assay): no statistically significant increase of micronucleated polychromatic erythrocytes in dosed animals
- Ratio of PCE/NCE (for Micronucleus assay): the ratio remained essentially unaffected by the test compound
- Statistical evaluation: see above

Any other information on results incl. tables

Mean mutation indices in polychromatic erythrocytes:

Vehicle control group -- dose group -- positive control group

24 hours male: 1.0 -- 1.8 -- 23.5

24 hours female: 1.0 -- 1.3 -- 21.1

48 hours male: 1.0 -- 0.7 -- not done

48 hours female: 1.0 -- 0.3 -- not done

72 hours male: 1.0 -- 1.7 -- not done

72 hours female: 1.0 -- 1.0 -- not done

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Oral administration of the test item (Pigment Red 122) did not lead to a substantial increase of micronucleated polychromatic erythocytes and was not mutagenic in the in vivo mouse micronucleus test
Executive summary:

The test item was tested in the micronucleus test. The test compound was suspended in sesame oil and dosed once oral at 2500 mg per kg bodyweight to male and female mice, upon the results of the previously conducted dose range finding assay. According to the test procedure the animals were killed 24, 48 or 72 hours after administration.

EndoxanR was used as positive control substance and was administered orally at a dose of 50 mg per kg bodyweight.

The number of polychromatic and normochromatic erythrocytes containing micronuclei was not increased. The ratio of polychromatic/normochromatic erythrocytes in both male and female animals remained unaffectedby the treatment with the test item and was statistically not different from the control values.

EndoxanR induced in both males and females a marked statistically significant increase in the number of polychromatic cells with micronuclei, indicating the sensitivity af the system.

The results indicate that, under the conditions of the present study, the test item is not mutagenic in the micronucleus test.