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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
Study period:
13 March 1989 to 12 October 1989
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1989
Report date:
1989

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to guideline
Guideline:
OECD Guideline 474 (Mammalian Erythrocyte Micronucleus Test)
Qualifier:
according to guideline
Guideline:
EU Method B.12 (Mutagenicity - In Vivo Mammalian Erythrocyte Micronucleus Test)
GLP compliance:
yes
Type of assay:
mammalian erythrocyte micronucleus test

Test material

Constituent 1
Chemical structure
Reference substance name:
Cyclopentanone, 2-[(2R)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2S)-rel-
Molecular formula:
C14H24O1
IUPAC Name:
Cyclopentanone, 2-[(2R)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2S)-rel-
Constituent 2
Chemical structure
Reference substance name:
Cyclopentanone, 2-[(2S)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2R)-rel-
Molecular formula:
C15H24O
IUPAC Name:
Cyclopentanone, 2-[(2S)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2R)-rel-
Constituent 3
Chemical structure
Reference substance name:
Cyclopentanone, 2-[(2S)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2S)-rel-
Molecular formula:
C15H24O
IUPAC Name:
Cyclopentanone, 2-[(2S)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2S)-rel-
Constituent 4
Chemical structure
Reference substance name:
Cyclopentanone, 2-[(2R)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2R)-rel-
Molecular formula:
C15H24O
IUPAC Name:
Cyclopentanone, 2-[(2R)-2-[(1R)-4-methyl-3-cyclohexen-1-yl]propyl]-, (2R)-rel-
Test material form:
liquid
Specific details on test material used for the study:
Name (as stated in the report) : NECTARYL - LRG 1371
Identification of the test item for the study: 07683 D9 001
Aspect: colourless slightly viscous liquid
Batch Number: AS1241001

Test animals

Species:
mouse
Strain:
other: OFl mouse (IOPS Caw).
Details on species / strain selection:
The mouse is currently used in this type of test and is recommended by the different legislative authorities.
Sex:
male/female
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Iffa-Credo (69210 L'Arbresle - France).
- Age at study initiation: young adults, aged from 6 to 8 weeks
- Weight at study initiation: values between 28.3 g and 36.9 g for the males, and between 21.2 g and 28.8 g for the females.
- Cages : housed in groups of 5 (or 2 in the preliminary studies), by sex in translucent polypropylene VS cages (internal dimensions 205 x 115 x 129 mm).
- Air changes: at least 10 per hour
- Temperature: 22, 3° C
- Humidity: 30 to 70% R.H
- Lighting: artificial, 12 hours out of 24 (photoperiod = 07,30 - 19,30).
- Hygiene: cages changed for each study.

Administration / exposure

Route of administration:
other: Oral route (test substance and negative control) and intraperitoneal route (positive control).
Vehicle:
The test substance was diluted in sterile Codex liquid paraffin (Aguettant, Batch 6170 B, March 1993)
Details on exposure:
Preliminary study: 4 groups made up of 2 males and 2 females each treated respectively at the dose levels of 9600*, 4800, 960 and 192 mg/kg. A complementary study was performed at the dose levels of 4800 and 960 mg/kg to confirm the clinical observations.
Main study: The dose level of 4800 mg/kg was administered in the main study
Duration of treatment / exposure:
Oral administration: 10 ml/kg of live body weight
Frequency of treatment:
The test substance was administered once only
Post exposure period:
Observations were carried out within the first hour and then 15 minutes, 24, 48 and 72 hours after administration.
Doses / concentrations
Dose / conc.:
4 800 mg/kg bw/day (nominal)
Remarks:
Dose conc of the main study was determined from the complementary study where no mortality occured at the dose level of 4800 mg/kg bw
No. of animals per sex per dose:
5 males and 5 females per group / 1 dose concentration per group
Control animals:
yes
yes, concurrent vehicle
Positive control(s):
A solution of cyclophosphamide (Sigma, Batch 114 F 0393) in sterile water for injection (Aguettant, Batch 85456, July 1991) was prepared. Used concentration: 10 mg/ml

Examinations

Details of tissue and slide preparation:
For each animal both femurs were removed. The femoral bone marrow was extracted with the aid of syringe, and suspended in foetal calf serum. After centrifugation and resuspension the smears were prepared (2 slides per animal). The slides were stained with May-Grunwald- Giemsa. Microscopic examination of the slides was performed. For each animal 1000 erythrocytes were examined (500 cells per slide, 2 slides). A count of the number of polychromatophile erythrocytes bearing micronuclei was performed. At the same time the ratio of normochromatophile erythrocytes to polychromatophile erythrocytes was established by the observation of 400 cells per animal (200 cells per slide, 2 slides).

Results and discussion

Test results
Key result
Sex:
male/female
Genotoxicity:
negative
Toxicity:
yes
Remarks:
see "additional information on results" field below
Vehicle controls validity:
not examined
Remarks:
test item was administered pure
Negative controls validity:
valid
Positive controls validity:
valid
Additional information on results:
The normochromatic / polychromatic erythrocytes ratio, is enhanced by a factor of 2 to 3 in the animals treated by the test substance. This observation shows the slightly toxic effect of the test substance on the bone marrow cells.

Applicant's summary and conclusion

Conclusions:
No significant increase of the number of polychromatophile erythrocytes bearing micronuclei was observed in the animals treated with the test substance, at the dose level of 4800 mg/kg and killed 24, 48 or 72 hours after administration.
In contrast, significant results were observed in the animals treated with the positive control. The normochromatic / polychromatic erythrocytes ratio, is enhanced by a factor of 2 to 3 in the animals treated by the test substance. This observation shows the slightly toxic effect of the test substance on the bone marrow cells.
From these experimental results, it may be concluded that the test substance does not induce any mutagenic effect in the mouse.
Executive summary:

The mutagenic potential of the test substance was tested in the mouse using the micronucleus test OECD Guideline No. 474. The test substance was administered by the oral route at the dose level of 4800mg/kg as described in the preliminary study.

A negative control (administration of vehicle) and a positive control (administration of a standard mutagenic substance) were included in this study.

The animals (5 males and 5 females per group) were killed 24, 48 or 72 hours after the administration of the test substance. For each animal an examination of 1000 polychromatophile erythrocytes obtained from the femoral bone marrow was performed. The frequency of micronuclei was calculated for each animal and for each group.

Under the experimental conditions employed,the test substance NECTARYL did not induce mutagenic effects in the mouse.