3-(p-methoxyphenyl)-2-methylpropionaldehyde

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:

06 Jul 2004 to 15 Jul 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2004

Report date:

2004

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

July 21 1997

GLP compliance:

yes (incl. QA statement)

Remarks:

Cytotest Cell Research GmbH (RCC-CCR), In den Leppsteinswiesen 19, D-64380, Rossdorf

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

3-(p-methoxyphenyl)-2-methylpropionaldehyde

EC Number:

226-749-5

EC Name:

3-(p-methoxyphenyl)-2-methylpropionaldehyde

Cas Number:

5462-06-6

Molecular formula:

C11H14O2

IUPAC Name:

3-(4-methoxyphenyl)-2-methylpropanal

Test material form:

liquid

Method

Target gene:

Histidine gene

Species / strain

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and TA 102

Metabolic activation:

with and without

Metabolic activation system:

rat liver S9 induced with phenobarbital and ß-Napthoflavone

Test concentrations with justification for top dose:

Pre-experiment
3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate
The pre-experiment is reported as main experiment l, since the following criteria are met: Evaluable plates (>0 colonies) at five concentrations or more in all strains used.

Main experiment I (plate incorporation test): 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate
Main experiment II (pre-incubation assay) without S9 mix: 1; 3; 10; 33; 100; 333; 1000; and 2500 µg/plate
Main experiment II (pre-incubation assay) with S9 mix: 3; 10; 33; 100; 333; 1000; 2500; and 5000 µg/plate

Vehicle / solvent:

ethanol (>99%)

Controls

Untreated negative controls:

yes

Remarks:

untreated

Negative solvent / vehicle controls:

yes

Remarks:

100 µL ethanol

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: see section "Any other information on materials and methods incl. tables"

Details on test system and experimental conditions:

METHOD OF APPLICATION:
Experiment I: in agar (plate incorporation)
Experiment II: preincubation

DURATION:
- Preincubation period: 60 minutes
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS: triplicates

OTHER EXAMINATIONS:
The colonies were counted using the AUTOCOUNT (Artek Systems Corporation, BIOSYS GmbH, D-61184 Karben). The counter was connected to an IBM AT compatible PC with printer which printed out both, the individual and mean values of the plates for each concentration together with standard deviations and enhancement factors as compared to the spontaneous reversion rates. Due to reduced background growth, the colonies were partly counted manually.

Evaluation criteria:

Acceptability of the Assay:
The Salmonella typhimurium reverse mutation assay is considered acceptable if it meets the following criteria:
- regular background growth in the negative and solvent control
- the spontaneous reversion rates in the negative and solvent control are in the range of our historical data
- the positive control substances should produce a significant increase in mutant colony frequencies

Evaluation of Results:
A test item is considered as a mutagen if a biologically relevant increase in the number of revertants exceeding the threshold of twice (strains TA 98, TA 100, and TA 102) or thrice (strains TA 1535 and TA 1537) the colony count of the corresponding solvent control is observed.
A dose dependent increase is considered biologically relevant if the threshold is exceeded at more than one concentration.
An increase exceeding the threshold at only one concentration is judged as biologically relevant if reproduced in an independent second experiment.
A dose dependent increase in the number of revertant colonies below the threshold is regarded as an indication of a mutagenic potential if reproduced in an independent second experiment. However, whenever the colony counts remain within the historical range of negative and solvent controls such an increase is not considered biologically relevant.

Statistics:

A statistical analysis of the data is not required

Results and discussion

Test results

Key result

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 98, TA 100, TA 102

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:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitation of the test item occurred up to the highest investigated dose.

RANGE-FINDING/SCREENING STUDIES:
To evaluate the toxicity of the test item a pre-experiment was performed with strains TA 1535, TA 1537, TA 98, TA 100, and TA 102. Eight concentrations were tested for toxicity and mutation induction with each 3 plates. The pre-experiment is reported as main experiment l, since the following criteria are met: Evaluable plates >0 colonies) at five concentrations or more in all strains used.

HISTORICAL CONTROL DATA
- Positive historical control data: The historical range of positive controls was exceeded in strain TA 1537 (experiment I) with metabolic activation. This effect indicates the sensitivity of the strains rather than compromising the assay.
- Negative (solvent/vehicle) historical control data: In experiment I, with metabolic activation, the number of colonies did not quite reach the lower limit of our historical control data in the solvent control of strain TA 102. Since this deviation is rather small, this effect is judged to be based upon biological fluctuations and has no detrimental impact on the outcome of the study.

Any other information on results incl. tables

Distinct toxic effects, evident as a reduction in the number of revertants (below the factor of 0.5) were observed at the following concentrations (µg/plate):

 

Strain	Experiment I		Experiment Il
	without S9 mix	with S9 mix	without S9 mix	with S9 mix
TA 1535	1000 - 5000	1000 - 5000	1000, 2500	2500, 5000
TA 1537	1000 - 5000	5000	1000, 2500	2500, 5000
TA 98	2500, 5000	5000	2500	2500, 5000
TA 100	1000- 5000	5000	1000, 2500	2500, 5000
TA 102	333 - 5000	333 - 5000	1000, 2500	1000 - 5000

Applicant's summary and conclusion

Conclusions:

The test substance had, under the present test conditions, no ability to induce mutations in the bacterial reverse mutation assay according to OECD 471.

Executive summary:

The mutagenic activity of the test substance was evaluated in accordance with OECD 471 (1997) and GLP principles. The test was performed according to the plate incorporation (experiment 1) and pre-incubation method (experiment 2), in the absence and presence of S9-mix. In the pre-experiment the concentration range of the test item was 3- 5000 µg/plate. The pre-experiment is reported as experiment I since evaluable plates (> 0 colonies) were obtained at five concentrations or more in all strains used. In both experiment 1 and 2, reduced background growth was observed with and without S9 mix at higher concentrations in all strains (Salmonella typhimurium strains TA 1535, TA 1537, TA 98, TA 100, and TA 102). No substantial increase in revertant colony numbers of any of the five tester strains was observed following treatment with the test substance at any dose level, neither in the presence nor absence of metabolic activation (S9 mix). There was also no tendency of higher mutation rates with increasing concentrations in the range below the generally acknowledged border of biological relevance. Adequate vehicle and positive controls were included. Based on the results of this study it is concluded that the test substance is not mutagenic in the bacterial reverse mutation assay.