1,6-dichlorohexane

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro cytogenicity / micronucleus study

Remarks:

Type of genotoxicity: chromosome aberration

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2012-05-24 to 2012-08.17

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: GLP study Guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2012

Report date:

2012

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

other: OECD Guideline 487 (In Vitro Mammalian Cell Micronucleus Test /MNvit)) 2010

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

in vitro mammalian cell micronucleus test

Test material

Test material information

Constituent 1

Reference substance name:

1,6-dichlorohexane

EC Number:

218-491-7

EC Name:

1,6-dichlorohexane

Cas Number:

2163-00-0

Molecular formula:

C6H12Cl2

IUPAC Name:

1,6-dichlorohexane

Test material form:

other: liquid

Method

Species / strain

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

- Type and identity of media: in Ham’s F-12K medium
- Periodically checked for Mycoplasma contamination: yes

Metabolic activation:

with and without

Metabolic activation system:

rat liver post-mitochondrial fraction (S9 mix) from Aroclor 1254 induced animals

Test concentrations with justification for top dose:

15.63, 31.3, 62.5, 125, 250 µg test item/mL

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO

Controls

Controls 1

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

mitomycin C

Remarks:

without metabolic activation (clastogen)

Controls 2

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

cyclophosphamide

Remarks:

with metabolic activation (clastogen)

Controls 3

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Colchicine

Remarks:

without metabolic activation (aneugen)

Details on test system and experimental conditions:

METHOD OF APPLICATION: in medium

DURATION
- Exposure duration: 4 and 20 hours (without S9 mix), 4 hours (with S9 mix, twice)
- Fixation time (start of exposure up to fixation or harvest of cells): 20 hours after the end of exposure

NUMBER OF REPLICATIONS: 2

NUMBER OF CELLS EVALUATED: 5 * 10000 cells

DETERMINATION OF CYTOTOXICITY
- Method: cytokinesis proliferation block index (CPBI)

OTHER EXAMINATIONS:
- Determination of relative frequencies of mononucleate, binucleate, and multi-nucleate cells: treatment of cultures with cytoB (Cytochalasin B)

Evaluation criteria:

The assay demonstrates its ability to reliably and accurately detect substances of known aneugenic and clastogenic activity, with and without metabolic activation.
Solvent/vehicle control and untreated cultures give reproducibly low and consistent micronuclei frequencies, typically 5 – 25 micronuclei per 1000 cells according to OECD 487. Data from negative and positive controls are used to establish historical control ranges. These values are used in deciding the adequacy of the concurrent negative/positive controls for an experiment i.e. the negative/positive control data must be within the historical ranges.

Statistics:

The assessment was carried out by a comparison of the samples with the positive and the vehicle control using a chi-square test corrected for continuity according to YATES (COLQUHOUN, 1971) as recommended by the UKEMS guidelines (The United Kingdom Branch of the European Environmental Mutagen Society: Report of the UKEMS subcommittee on guidelines for mutagenicity testing, part III, 1989: Statistical evaluation of mutagenicity data). However, the results of statistical testing were assessed with respect to dose-response relationship. Reproducibility and historical data were also be taken into consideration.

Results and discussion

Test results

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at the top concentration of 250 µg 1,6-Dichlorohexane/mL

Vehicle controls validity:

not examined

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

RANGE-FINDING/SCREENING STUDIES: In the preliminary experiment without and with metabolic activation was tested the concentrations of 0, 10, 25, 100, 250, 1000, 2500, 5000 µg test item/mL. In the experiment pronounced to complete cytotoxicity was noted starting at concentrations of 250 µg test item/mL.

COMPARISON WITH HISTORICAL CONTROL DATA: The historical control range of micronucleus frequency of the untreated controls was 1.0 to 9.0 micronuclei per 1000 binucleated cells.

ADDITIONAL INFORMATION ON CYTOTOXICITY: In the main stuy cytotoxicity was noted at the top concentration of 250 µg test item/mL in the experiment without and with metabolic activation.

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative with metabolic activation
negative without metabolic activation

Under the present test conditions, 1,6- dichlorohexane tested up to a cytotoxic concentration of 250 µg/mL in the absence and in the presence of metabolic activation employing two exposure times (without S9) and one exposure time (with S9) revealed no indications of mutagenic properties in the vitro micronucleus test.