4-isopropyl-1-methylcyclohexene

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2013

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

Screening Test

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

no guideline available

Version / remarks:

Guideline currently in draft with the OECD

Principles of method if other than guideline:

An equal volume of specialized growth medium containing BlueScreen HC cells was added to each well.
Microplates were covered with a breathable membrane and incubated at 37°C, 5% CO2 and 95% humidity. After 3 hours, cells were
washed with phosphate buffered saline, centrifuged, and placed in recovery medium. After an additional 45 hours of incubation, the
fluorescence and flash luminescence of each well was measured using a microplate reader. Cytotoxicity was measured by the lysis of
cells followed by staining with a fluorescent DNA binding stain to provide an estimate of relative cell density.

GLP compliance:

yes

Type of assay:

other: (GLuc-T01), which contain a Gaussia luciferase (GLuc)-based reporter system

Test material

Test material information

Constituent 1

Reference substance name:

4-isopropyl-1-methylcyclohexene

EC Number:

226-841-5

EC Name:

4-isopropyl-1-methylcyclohexene

Cas Number:

5502-88-5

Molecular formula:

C10H18

IUPAC Name:

1-methyl-4-(propan-2-yl)cyclohex-1-ene

impurity 1

Reference substance name:

rel-(1R,4R)-1-isopropyl-4-methylcyclohexane

Cas Number:

1678-82-6

Molecular formula:

C10H20

IUPAC Name:

rel-(1R,4R)-1-isopropyl-4-methylcyclohexane

impurity 2

Reference substance name:

rel-(1S,4S)-1-isopropyl-4-methylcyclohexane

Cas Number:

6069-98-3

Molecular formula:

C10H20

IUPAC Name:

rel-(1S,4S)-1-isopropyl-4-methylcyclohexane

Test material form:

liquid

Specific details on test material used for the study:

Colourless liquid

Method

Target gene:

(GLuc-T01), which contain a Gaussia luciferase (GLuc)-based reporter system

Species / strain

Species / strain / cell type:

human lymphoblastoid cells (TK6)

Details on mammalian cell type (if applicable):

TK6 cells (GLuc-T01), which contain a Gaussia luciferase
(GLuc)-based reporter system.

Additional strain / cell type characteristics:

not applicable

Cytokinesis block (if used):

No

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced rats

Test concentrations with justification for top dose:

2097.15 to 10000 μM based on cytoxicity

Vehicle / solvent:

DMSO

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

cyclophosphamide

Evaluation criteria:

Luminescence induction below threshold of 1.8.
Microsoft Excel was used to store raw data that were analyzed by BlueScreen HC software to produce a semi-quantitative assessment of cytotoxicity and genotoxicity.

Results and discussion

Test results

Key result

Species / strain:

human lymphoblastoid cells (TK6)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not applicable

Positive controls validity:

valid

Additional information on results:

In the absence of S9:
Dose: 10000 micromolar Effects: Cytotoxicity
Results: Luminescence induction below threshold of 1.8; value=0.0

In the presence of S9:
Dose: 10000 micromolar Effects: Cytotoxicity
Results: Luminescence induction below threshold of 1.5; value=0.0.

Remarks on result:

other: Negative

Applicant's summary and conclusion

Conclusions:

Absence of S9: For this protocol, the statistically defined threshold for a positive genotoxic result is 1.8 (i.e., 80% induction over and above the baseline for vehicle controls). The positive control passed the test criteria.
Presence of S9: For this protocol, the statistically defined threshold for a positive genotoxic result is 1.5 (i.e., 50% induction over and above the baseline for vehicle controls). The positive control passed the test criteria.

Executive summary:

Negative for genotoxicity in the absence and presence of S9.