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Toxicological information

Genetic toxicity: in vivo

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Administrative data

Endpoint:
in vivo mammalian germ cell study: cytogenicity / chromosome aberration
Type of information:
other: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
weight of evidence
Study period:
1985
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Remarks:
The reliability of the source study was established to be R2: Justification for read-across is detailed at section 13.
Justification for type of information:
Justification for read-across is detailed at section 13.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1985
Report Date:
1985

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
GLP compliance:
not specified
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
- Source: Charles River GmbH, WIGA, D-8741 Sulzfeld, FRG
- Weight at study initiation: mean 26.7 g
- Assigned to test groups randomly: yes
- Housing: individually in Makrolon cages, type MI
- Diet: Standardized pelleted feed (Kliba Haltungsdiät, Klingentalmühle AG, CH-4303 Kaiseraugst, Switzerland); ad libitum.
- Water: drinking water from bottles; ad libitum.
- Acclimation period: about one week.

ENVIRONMENTAL CONDITIONS
- Temperature: 20 - 24 °C
- Humidity: 30 - 70 %
- Photoperiod (hrs dark / hrs light): 12 / 12

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
- Vehicle/solvent used: CMC (carboxymethyl cellulose)
- Concentration of test material in vehicle: 8.5, 17, 34 g/100 ml
- Amount of vehicle: 20 ml/kg bw
Details on exposure:
PREPARATION OF DOSING SOLUTIONS
All test substance formulations were prepared immediately before administration. The amount of substance or volume to be administered was related to the specific weight of the individual animals on the day of the experiment.
Duration of treatment / exposure:
16, 24, 48 h for the highest dose of test material; 24 h for all other dose groups and controls
Frequency of treatment:
Single application
Post exposure period:
16, 24, 48 h for the highest dose of test material; 24 h for all other dose groups and controls
Doses / concentrationsopen allclose all
Dose / conc.:
1 700 mg/kg bw/day (nominal)
Remarks:
#1
Dose / conc.:
3 400 mg/kg bw/day (nominal)
Remarks:
#2
Dose / conc.:
6 800 mg/kg bw/day (nominal)
Remarks:
#3
No. of animals per sex per dose:
5
Control animals:
yes, concurrent vehicle
Positive control(s):
- Substance: cyclophosphamide
- Route of administration: oral by gavage
- Doses / concentrations: 40 mg/kg bw

Examinations

Tissues and cell types examined:
Bone marrow of the two femora
Details of tissue and slide preparation:
CRITERIA FOR DOSE SELECTION
In the determination of the acute oral toxicity all animals survived the high dose of 6800 mg/kg body weight without any clinical signs or symptoms. A volume as high as 20 ml/kg body weight had to be selected in order to be able do administer this amount. Higher doses suspended in an 0.5 % aqueous CMC formulation led to a viscous mass which could no longer be administered.

DETAILS OF SLIDE PREPARATION
Smears were prepared using slides with ground edges, the preparations were dried in the air and subsequently stained. Staining in eosin and methylene blue solution for 5 minutes. Rinsed in aqua dest., then placed in fresh aqua dest. for 2 or 3 minutes. Staining in Giemsa solution for 12 minutes. After being rinsed twice in aqua dest. and clarified in xylene, the preparations were embedded in Entellan.

METHOD OF ANALYSIS
As a rule, 1000 polychromatic erythrocytes from each of the male and female animals of every test group are evaluated and investigated for micronuclei. The normochromatic erythrocytes (= normocytes), which occur, are also scored.
Evaluation criteria:
The increase in the micronucleus rate in polychromatic erythrocytes of treated animals as compared with the solvent control group provides an index of a chromosome breaking (clastogenic) effect or of a spindle activity of the substance tested.
Statistics:
Two statistical tests were used to answer the questions of whether there are significant differences between control group and dose group or between the individual dose groups concerning the rate of micronuclei in polychromatic erythrocytes: first, the exact test according to FISHER, which was applied to register significant differences between the relative frequencies of a characteristic of two groups, and, second, the asymptotic U test according to MANN-WHITNEY (rank test modified according to WILCOXON). The relative frequencies of cells with micronuclei per animal were use das a criterion of the rank determination for the U 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- Clinical signs of toxicity in test animals: The single oral administration in doses of 6800 mg/kg, 3400 mg/kg or 1700 mg/kg body weight was tolerated by all animals without any signs of toxicity.-Necropsy: The gross-pathological examination of the animals sacrificed at the end of the study did not reveal any changes of the internal organs which could the attributed to the test substance administered.

Any other information on results incl. tables

Results:

Substance Dose (mg/kg bw) Interval: 16 hours Interval: 24 hours Interval: 48 hours
Polychromatic erythrocytes investigated Normocytes / 10000 polychromatic erythrocytes Cells with micronuclei Polychromatic erythrocytes investigated Normocytes / 10000 polychromatic erythrocytes Cells with micronuclei Polychromatic erythrocytes investigated Normocytes / 10000 polychromatic erythrocytes Cells with micronuclei
per 1000 polychromatic erythrocytes per 1000 normochromatic erythrocytes per 1000 polychromatic erythrocytes per 1000 normochromatic erythrocytes per 1000 polychromatic erythrocytes per 1000 normochromatic erythrocytes
vehicle control, 0.5% CMC - 10000 3485 1.8 0
Säurebraun 6229 6800 10000 2988 1.4 1.34 10000 4349 1.4 1.61 10000 4359 1.3 1.61
Säurebraun 6229 3400 - 10000 3506 1.4 0.86
Säurebraun 6229 1700 - 10000 4307 1.9 0.46
Cyclophospamide 40 - 10000 5623 23.4 1.78

Applicant's summary and conclusion

Conclusions:
No induction of chromosome breaking (clastogenic) effect or a spindle activity in polychromatic erythrocytes of the bone marrow of the femora mice.
Executive summary:

The substance was tested for chromosome aberration potential following OECD 474, by oral administration.

The test substance, under the experimental conditions, did not induce chromosome breaking (clastogenic) effect or a spindle activity in polychromatic erythrocytes of the bone marrow of the femora mice.