Nonan-2-one

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / micronucleus study

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Justification for type of information:

Data is from peer reviewed publication

Data source

Reference

Reference Type:

publication

Title:

Evaluation of the genotoxic potential of some microbial volatile organic compounds (MVOC) with the comet assay, the micronucleus assay and the HPRT gene mutation assay

Author:

Ludwika Kreja, Hans-Joachim Seidel

Year:

2002

Bibliographic source:

Mutation Research 513 (2002) 143–150

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: as below

Principles of method if other than guideline:

To evaluate the clastogenic effect of 2-Nonanone in Chinese hamster V79 cells by Micronucleus test.

GLP compliance:

not specified

Type of assay:

in vitro mammalian cell micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

Nonan-2-one

EC Number:

212-480-0

EC Name:

Nonan-2-one

Cas Number:

821-55-6

Molecular formula:

C9H18O

IUPAC Name:

nonan-2-one

Details on test material:

- Name of test material (as cited in study report): 2-Nonanone
- Molecular formula (if other than submission substance): C9H18O
- Molecular weight (if other than submission substance): 142.24g/mol
- Substance type: Organic
- Physical state: Liquid
Purity – No data available.
- Impurities (identity and concentrations):

Specific details on test material used for the study:

- Name of test material: 2-Nonanone
- IUPAC name: nonan-2 -one
- Molecular formula: C9H18O
- Molecular weight: 142.24g/mol
- Substance type: Organic
- Physical state: No data
- Purity: No data available
- Impurities (identity and concentrations): No data

Method

Target gene:

No data

Species / strain

Species / strain / cell type:

Chinese hamster lung fibroblasts (V79)

Details on mammalian cell type (if applicable):

- Type and identity of media:V79 Chinese hamster cell line were maintained in 75 cm2 tissue culture flasks at 37◦C in a humidified atmosphere with 5% CO2 in MEM Eagle medium (V79 cells) supplemented with 10% heat-inactivated fetal calf serum (FCS), 2mM l-glutamine, 100 IU/ml penicillin and streptomycin.
- Properly maintained: Yes
- Periodically checked for Mycoplasma contamination: No data available
- Periodically checked for karyotype stability: No data available
- Periodically "cleansed" against high spontaneous background: No data available

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

No data

Metabolic activation:

without

Metabolic activation system:

Aroclor 1254-induced rat liver S9 fraction

Test concentrations with justification for top dose:

0, 6 or 14 mM

Vehicle / solvent:

DMSO
- - Justification for choice of solvent/vehicle: The chemical was soluble in DMSO

Controls

Untreated negative controls:

yes

Remarks:

DMSO

Negative solvent / vehicle controls:

yes

True negative controls:

not specified

Positive controls:

yes

Remarks:

With and without S9 metabolilc activation system

Positive control substance:

cyclophosphamide

methylmethanesulfonate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium
- Cell density at seeding (if applicable): 100000 cells

DURATION
- Preincubation period: Not applicable
- Exposure duration: 4 hrs
- Expression time (cells in growth medium): 24 hrs
- Selection time (if incubation with a selection agent): No data
- Fixation time (start of exposure up to fixation or harvest of cells): No data

SELECTION AGENT (mutation assays): No data

SPINDLE INHIBITOR (cytogenetic assays): No data

STAIN (for cytogenetic assays): No data

NUMBER OF REPLICATIONS: Two to four independent experiments were performed

METHODS OF SLIDE PREPARATION AND STAINING TECHNIQUE USED:

NUMBER OF CELLS EVALUATED: 1000 cells/culture

NUMBER OF METAPHASE SPREADS ANALYSED PER DOSE (if in vitro cytogenicity study in mammalian cells): No data

CRITERIA FOR MICRONUCLEUS IDENTIFICATION: Micronuclie identification was done as per the criteria given by Miller et al

DETERMINATION OF CYTOTOXICITY
- Method: mitotic index; cloning efficiency; relative total growth; other: No data
- Any supplementary information relevant to cytotoxicity: No data

OTHER EXAMINATIONS:
- Determination of polyploidy: No data
- Determination of endoreplication: No data
- Methods, such as kinetochore antibody binding, to characterize whether micronuclei contain whole or fragmented chromosomes (if applicable): No data

- OTHER: No data

Rationale for test conditions:

No data

Evaluation criteria:

The test compound was classified as a clastogen when it was able to enhance the spontaneous MN frequency at least three-fold or higher over that of the control for at least one dose tested.

Statistics:

Micronuclei (MN) were analyzed in 1000 cells/culture. To show reproducibility of the results two to four independent experiments were performed and the mean MN frequency per 100 cells ± S.D. was calculated.

Results and discussion

Test results

Species / strain:

Chinese hamster lung fibroblasts (V79)

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Remarks:

A level of 0.5% spontaneous MN was found in the non-exposed control sample

Positive controls validity:

valid

Remarks:

a concentration-dependent significant increase of micronuclei was found after MMS exposure. After cyclophosphamide treatment used as a positive control for S9-mix metabolic activation, MN frequency was increased from 0.3 to 4.8%

Additional information on results:

No data

Any other information on results incl. tables

Table: Clastogenic effects of 2- Nonanone

Substance	Concentration (mM)	MN frequency (%)
		Without S9	With S9
Control non-exposed		0.5±0.3
MMS	0.25	3.4 ± 1.1
	0.5	11.8 ± 4.4
Cyclophosphamide	0.1	0.3	4.8
2- Nonanone	6	0.5 ± 0.2
	14	0.5 ± 0.1

Applicant's summary and conclusion

Conclusions:

2-Nonanone failed to induce clastogenic effects in Chinese hamster V79 cells in absence of S9 metabolic activation system by Micronucleus test and hence is considered to be negative for gene mutation in vitro.

Executive summary:

In Genetic toxicity in vitro study for 2-Nonanone ( IUPAC name: nonan-2 -one) in Chinese hamster V79 cells by Micronucleus test, V79 Chinese hamster cells were cultivated on microscope slides in quadripermdishes (Heraeus Instruments, Germany). A total of 10⁵cells were seeded in each chamber and cultivated 24 h before treatment. The medium was then substituted by 6ml of fresh medium containing the test compound at doe level of 6 or 14 mM and the cells were incubated for 4 h. After treatment the cultures were washed twice with medium and incubated further for 24 h. Micronuclei (MN) were analyzed in 1000 cells/culture. To show reproducibility of the results two to four independent experiments were performed and the mean MN frequency per 100 cells ± S.D. was calculated. The test compound was classified as a clastogen when it was able to enhance the spontaneous MN frequency at least three-fold or higher over that of the control for at least one dose tested. A level of 0.5% spontaneous MN was found in the non-exposed control sample. However, a concentration-dependent significant increase of micronuclei was found after MMS exposure (t-test; P < 0.001). After cyclophosphamide treatment used as a positive control for S9-mix metabolic activation, MN frequency was increased from 0.3 to 4.8%. 2-Nonanone failed to induce clastogenic effects in Chinese hamster V79 cells in the absence of S9 metabolic activation system by Micronucleus test and hence is considered to be negative for gene mutation in vitro.