trans-1,4-bis(iodomethyl)cyclohexane

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: The study was conducted in compliance with GLP and according to the guideline EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria).

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

EU Method B.13/14 (Mutagenicity - Reverse Mutation Test Using Bacteria)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

trans-1,4-bis(iodomethyl)cyclohexane

Cas Number:

83447-96-5

Molecular formula:

C8H14I2

IUPAC Name:

trans-1,4-bis(iodomethyl)cyclohexane

Method

Target gene:

Strain Mutation Type

TA 1535 hisG46 Base pair
TA 100 hisG46 Base pair
TA 1537 hisC3076 Frameshift
TA 98 hisD3052 Frameshift
WP211vrA trpE Base pair

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Details on mammalian cell type (if applicable):

All used bacterial strains are defective in DNA-repair (uvrB and uvrA respectively). The
Salmonella strains also have a defective lipopolysaccharide barrier on the cell wall (rfa) .
These properties confer extra sensitivity to DNA damage and also greater permeability to
large moleclules. The strains TA 98 and TA 100 also contain a plasmid (pKM1O1 ) which
enhances the error prone repair and confers ampicillin resistance.

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9

Test concentrations with justification for top dose:

1000, 500, 100, 50, 10 and 5 µg/plate for all strains and for TA1537 additionally 1µg/plate to reach 5 concentrations without cytotoxicity.

Vehicle / solvent:

Dimethyl sulfoxide

Controls

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

Dimethyl sulfoxide

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

2-nitrofluorene

sodium azide

methylmethanesulfonate

other: 2-aminoanthracene

Details on test system and experimental conditions:

The test item was tested in the plate incorporation test at concentrations of 1000, 500, 100, 50, 10 and 5 µg/plate in the experiments without and
with metabolic activation because the test item precipitated to a solid precipitate in the top agar at the concentration of 5000 µg/plate. In
the pre-incubation test the test item was tested at concentrations of 1000, 500, 100, 50, 10 and 5 µg/plate in the experiments without and with
metabolic activation as weIl as. An additional experiment with metabolic activation in the pre-incubation test in the bacterial strain TA 1537 was
carried out at the concentration of 1 µg/plate to reach five concentration levels without cytotoxicity.

Evaluation criteria:

A test is considered to be positive if the test item induces dose related increases in numbers of
revertants scored in two separate experiments and these increases are deemed to be of
biological relevance. Reproducible increases at one experimental point may also be indicative
of a positive response. For a biologically relevant response the number of revertants is
expec ted to be at least the double of the spontaneous reversion.
A test item producing neither a dose related and reproducible increase in the number of
revertants nor a reproducible positive response at any experimental point is considered to be
non-mutagenic in this test system.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

1000 and 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 98

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 100

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

1000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 5

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Applicant's summary and conclusion

Conclusions:

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

Based on the results of the reported study it is concluded that P2 does not induce gene mutation in Salmonella typhimurium and Escherichia coli
under the experimental conditions described.
Therefore, P2 is considered to be non-mutagenic in the bacterial reverse mutation test.

Executive summary:

At the concentration of 5000µg/plate solid precipitation was observed and therefore the concentration range was limited to 1000µg/plate at which for all Salmonella typhimurium cytotoxicity was observed (for TA1537 already at 500µg/plate). All strains were tested with and wuithout metabolic activation and there was no relevant increase in the number of revertants seen at any concentration tested in this experiment. Hence the substance is considered to be non mutagenic in the bacterial strains used in this study.