Benzenepropanal, 2-methyl-4-(2-methylpropyl)-

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:

09 March 2015 to 23 March 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 2

Qualifier:

according to guideline

Guideline:

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

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

3-[2-methyl-4-(2-methylpropyl)phenyl]propanal

EC Number:

811-285-3

Cas Number:

1637294-12-2

Molecular formula:

C14H20O

IUPAC Name:

3-[2-methyl-4-(2-methylpropyl)phenyl]propanal

Test material form:

liquid

Specific details on test material used for the study:

Name (as stated in the report): GR-88-0778
Lot No: Batch 6
Aspect: Colourless liquid
Expiration date: March 29, 2017

Method

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

S9-mix

Test concentrations with justification for top dose:

In the first mutation assay, GR-88-0778 was tested up to concentrations of 5000 μg/plate in the absence and presence of 5% (v/v) S9-mix. GR-88-0778 precipitated on the plates at dose levels of 1600 μg/plate and upwards. Toxicity was observed in all tester strains, except in tester strain WP2uvrA in the presence of S9-mix. However since GR-88-0778 precipitated heavily on the plates at the test substance concentration of 5000 μg/plate, the number of revertants of this dose level could not be determined in tester strain WP2uvrA.In the second mutation assay, GR-88-0778 was tested at appropriate dose ranges in the absence and presence of 10% (v/v) S9-mix in the tester strains TA1535, TA1537, TA98, TA100 and WP2uvrA. Toxicity was observed in all tester strains, except in WP2uvrA in the presence of S9-mix. However since GR-88-0778 precipitated heavily on the plates at the test substance concentration of 5000 μg/plate, the number of revertants of this dose level could not be determined in tester strain WP2uvrA.

Vehicle / solvent:

GR-88-0778 was dissolved in dimethyl sulfoxide (DMSO, SeccoSolv, Merck, Darmstadt, Germany).

Controls

Controls 1

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

sodium azide

Remarks:

TA1535 without S9

Controls 2

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: ICR-191

Remarks:

TA1537 without S9

Controls 3

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

2-nitrofluorene

Remarks:

TA98 without S9

Controls 4

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

methylmethanesulfonate

Remarks:

TA100 without S9

Controls 5

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

WP2uvrA without S9

Controls 6

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-aminoanthracene

Remarks:

All strains with S9

Results and discussion

Test results

Key result

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

In the first mutation assay, toxicity was observed in all tester strains, except in tester strain WP2uvrA in the presence of S9-mix (tested up to concentrations of 5000 μg/plate).
In the second mutation assay, toxicity was observed in all tester strains, except in WP2uvrA in the presence of S9-mix.
In the third mutation experiment performed on strain TA98 without S9 mix toxicity was observed at the dose levels of 164 μg/plate and upwards.

Applicant's summary and conclusion

Conclusions:

All bacterial strains showed negative responses over the entire dose range, i.e. no significant dose-related increase in the number of revertants in two independently repeated experiments.
The negative and strain-specific positive control values were within the laboratory background historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly.
Based on the results of this study it is concluded that GR-88-0778 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.

Executive summary:

Evaluation of the mutagenic activity of GR-88-0778 in the Salmonella typhimurium reverse mutation assay and the Escherichia coli reverse mutation assay. GR-88-0778 was tested in the Salmonella typhimurium reverse mutation assay with four histidine-requiring strains of Salmonella typhimurium (TA1535, TA1537, TA100 and TA98) and in the Escherichia coli reverse mutation assay with a tryptophan-requiring strain of Escherichia coli (WP2uvrA). The test was performed in two independent experiments in the presence and absence of S9-mix (rat liver S9-mix induced by induced by Aroclor 1254). An additional experiment was performed with the tester strain TA98 in the absence of S9-mix. The study procedures described in this report were based on the most recent OECD, EC and MITI guidelines. Batch 6 of GR-88-0778 was a colourless to pale yellow liquid with a purity of 98.6%. The test substance was dissolved in dimethyl sulfoxide. In the first mutation assay, GR-88-0778 was tested up to concentrations of 5000 μg/plate in the absence and presence of 5% (v/v) S9-mix. GR-88-0778 precipitated on the plates at dose levels of 1600 μg/plate and upwards. Toxicity was observed in all tester strains, except in tester strain WP2uvrA in the presence of S9-mix. However since GR-88-0778 precipitated heavily on the plates at the test substance concentration of 5000 μg/plate, the number of revertants of this dose level could not be determined in tester strain WP2uvrA. In the second mutation assay, GR-88-0778 was tested at appropriate dose ranges in the absence and presence of 10% (v/v) S9-mix in the tester strains TA1535, TA1537, TA98, TA100 and WP2uvrA. Toxicity was observed in all tester strains, except in WP2uvrA in the presence of S9-mix. However since GR-88-0778 precipitated heavily on the plates at the test substance concentration of 5000 μg/plate, the number of revertants of this dose level could not be determined in tester strain WP2uvrA. Since in tester strain TA98 in the second experiment too many dose levels showed severe toxicity in the absence of S9-mix, a third mutation experiment was performed with this strain in the absence of S9-mix. GR-88-0778 was tested up to the dose level 1600 μg/plate. Toxicity was observed at the dose levels of 164 μg/plate and upwards. GR-88-0778 did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four tester strains (TA1535, TA1537, TA98 and TA100) and in the number of revertant (Trp+) colonies in tester strain WP2uvrA both in the absence and presence of S9-metabolic activation. These results were confirmed in an independently repeated experiment. In this study, the negative and strain-specific positive control values were within the laboratory historical control data ranges indicating that the test conditions were adequate and that the metabolic activation system functioned properly. Based on the results of this study it is concluded that GR-88-0778 is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay.