Registration Dossier

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:
From 15 February, 1983 to 18 March, 1983
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: Study was conducted according to the OECD guideline 474 and EU method B.12.

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1983
Report Date:
1983

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
Qualifier:
equivalent or similar to
Guideline:
EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
Deviations:
no
GLP compliance:
not specified
Type of assay:
micronucleus assay

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: slightly viscous solution
Details on test material:
- Name of test material (as cited in study report): Quaternary ammonium salts no. 1 (trimethylcocoammoniumchloride)
- Description: Clear, colourless, slightly viscous solution. This solution contained 67% water as the solvent.

Test animals

Species:
mouse
Strain:
CD-1
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River U.K. Limited, Margate, Kent
- Weight at study initiation: 18-21 g
- Assigned to test groups randomly: Yes
- Housing: Plastic disposable cage
- Diet: Spratt's Laboratory Diet number 1, ad libitum
- Water: Tap water, ad libitum
- Acclimation period: 10 d

ENVIRONMENTAL CONDITIONS
- Air changes (per hr): 30/h
- Photoperiod (hrs dark / hrs light): 12 h/12 h

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
1% methyl cellulose
Details on exposure:
Dilution of the original solution were made in aqueous 1% methyl cellulose to give the desired concentration of the test substance. The mice were starved overnight prior to dosing. All animals in all groups were dosed with the standard volume of 2 mL/10 g bw. The test substance and vehicle control were dosed by oral gavage. The positive control was dosed by intraperitoneal injection.
Duration of treatment / exposure:
Single treatment
Frequency of treatment:
Single
Doses / concentrations
Remarks:
Doses / Concentrations:
468 mg/kg bw
Basis:
nominal conc.
No. of animals per sex per dose:
45 males and 45 females
Control animals:
yes, concurrent vehicle
Positive control(s):
Mitomycin C was used. It was prepared as a solution in sterile 0.9% saline at a concentration of 0.4 mg/mL.

Examinations

Tissues and cell types examined:
The femurs were cleared of tissue and one epiphysis removed from each bone.
Details of tissue and slide preparation:
A direct bone marrow smear was made on to a slide containing a drop of calf-serum. One smear was made from each femur. The prepared smears were air-dried and fixed in methanol. After fixation, the smears were air-dried and stained using Giemsa's technique. After rinsing in buffered distilled water, the slides were air-dried and mounted with coverslips using DPX. The stained smears were examined by light microscopy to determine the incidence of micronucleated cells/1,000 polychromatic erythrocytes/animal. The ratio of polychromatic to nonchromatic erythrocyte (NCEs) for each animal was assessed by examination of at least 1,000 erythrocyte.
Evaluation criteria:
Reproducible and significant increase in the number of micronucleated NCEs in the test group over that of the control group.
Statistics:
Non-parametric methods by Hollander M and Wolfe DA.

Results and discussion

Test results
Sex:
male/female
Genotoxicity:
negative
Vehicle controls valid:
yes
Positive controls valid:
yes
Additional information on results:
RESULTS OF RANGE-FINDING STUDY
Probit analysis yielded an estimated LD10/3 of 468 mg/kg bw (LD50/3 was 884 mg/kg bw). A dose level of 468 mg/kg bw was chosen for the micronucleus test.

RESULTS OF DEFINITIVE STUDY- Induction of micronuclei (for Micronucleus assay): No significant difference as compared to controls.
- Ratio of PCE/NCE (for Micronucleus assay): The ratio of PCE to total erythrocytes remained essentially unaffected by the test substance
- Appropriateness of dose levels and route: Yes
- Statistical evaluation: Yes

Any other information on results incl. tables

No mortalities was seen in animals. Clinical signs included pilo-erection, hunched posture, lethargy and ptosis.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): negative
Under the test conditions, the test substance is considered to be non-mutagenic in the micronucleus test in mice.
Executive summary:

An OECD 474 study was conducted to assess the potential of C12-C18 TMAC to cause chromosomal damage (clastogenicity) in an in vivo mouse bone marrow micronucleus test. The test substance was suspended in 1% methyl cellulose and was administered as an oral dose of 468 mg/kg bw to male and female mice based on the results of a dose range finding assay. Following dosing, the animals were examined regularly and any clinical signs of reaction to the test substance were recorded. Five males and five females were sacrificed 24, 48 and 72 hours after dosing. The number of polychromatic erythrocytes containing micronuclei was not increased. The ratio of polychromatic erythrocytes to total erythrocytes in both male and female animals remained unaffected by the treatment with test substance. Under the test conditions, C12-C18 TMAC was not mutagenic in the mouse micronucleus test (Allenet al.,1983).