4,8-dimethyl-4,9-decadienal

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

The mutagenic activity of Floral Super was evaluated in accordance with OECD 471 (1997) guideline and according to GLP principles. The test was performed in two independent experiments: at first a direct plate assay was performed and secondly a pre-incubation assay, both in the absence and presence of S9-mix. The dose levels were selected based on observed cytotoxicity in dose range finding tests in strain TA 100 (direct plate -/+ S9 and preincubation +S9 >= 164 µg/plate, preincubation -S9 17 and 52 µg/plate). Adequate negative and positive controls were included. The substance did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four S. typhimurium tester strains (TA1535, TA1537, TA98 and TA100) and E. coli tester strain (WP2uvrA), both in the absence and presence of S9 -metabolic activation. These results were confirmed in independently repeated experiments. Based on the results of this study it is concluded that Floral Super is not mutagenic in the Salmonella typhimurium reverse mutation assay and Escherichia coli reverse mutation assay.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Study period:

19 March, 2015 - 16 April, 2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Reliability 1 is assigned because the study is conducted according to OECD TG 471, in compliance with GLP, without deviations that influence the quality of the results.

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

(1997)

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Version / remarks:

(2008)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Target gene:

- S. typhimurium: Histidine gene
- Escherichia coli: Tryptophan gene

Species / strain / cell type:

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

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix induced by Aroclor 1254.

Test concentrations with justification for top dose:

Direct plate:
- Dose range finding test:
TA 100 and WP2uvrA (without and with S9): 1.7, 5.4, 17, 52, 164, 512, 1600 and 5000 µg/plate
- Experiment 1:
Due to cytotoxicity (in TA 100), the following dose levels were used:
TA 1535, TA 1537 and TA 98 (without S9): 0.54, 1.7, 5.4, 17, 52 and 164 μg/plate
TA 1535, TA 1537 and Ta 98 (with S9): 1.7, 5.4, 17, 52, 164 and 512 μg/plate

Preincubation:
- Dose range finding test:
TA 100 (without S9): 0.17, 0.54, 1.7, 5.4, 17, 52 and 164 µg/plate
TA 100 (with S9): 0.54, 1.7, 5.4, 17, 52, 164 and 512 µg/plate
WP2uvrA (without and with S9): 5.4, 17, 52, 164, 512, 1600 and 5000 µg/plate
- Experiment 1:
Due to cytotoxicity (in TA 100), the following dose levels were used:
TA 1535, TA 1537, TA 98 and TA 100 (without S9): 0.056, 0.18, 0.55, 1.7, 5.4, 17 and 52 μg/plate
TA 1535, TA 1537 and TA 98 (with S9): 0.55, 1.7, 5.4, 17, 52 and 164 μg/plate

Vehicle / solvent:

- Solvent used: DMSO
- Justification for choice of solvent: the test substance was found to be soluble in DMSO up to 5000 µg/plate

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

Remarks:

(100 µL/plate)

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 1: in agar (plate incorporation)
- Experiment 2: (independent repeat): preincubation

DURATION
- Preincubation period: 30 minutes
- Exposure duration: 48 hours

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in triplicate in each strain (in all experiments)

DETERMINATION OF CYTOTOXICITY
- Method: on the basis of a decline in the number of spontaneous revertants, a thinning of the background lawn or a microcolony formation

Evaluation criteria:

A test substance is considered positive (mutagenic) in the test if:
a) The total number of revertants in tester strain TA100 is greater than two (2) times the concurrent control, or the total number of revertants in tester strains TA1535, TA1537, TA98 or WP2uvrA is greater than three (3) times the concurrent control.
b) In case a repeat experiment is performed when a positive response is observed in one of the tester strains, the positive response should be reproducible in at least one follow up experiment.

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Toxicity was observed in all tester strains.

Vehicle controls validity:

valid

Positive controls validity:

valid

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Only in the preincubation assay without S9-mix at the concentration of 164 µg/plate and upwards.

Vehicle controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: In the direct plate assay precipitation (oily droplets) was observed at the concentration of 1600 and 5000 µg/plate. In the preincubation assay, precipitation was observed at concentrations of 512 μg/plate in tester strain TA100 and at 5000 μg/plate in tester strain WP2uvrA.

RANGE-FINDING/SCREENING STUDIES:
- Direct plate assay: In strain TA 100 toxicity (microcolonies) was observed at the concentration of 164 µg/plate and above, both in the absence and presence of S9. In tester strain WP2uvrA, no reduction of the bacterial background lawn and no biologically relevant decrease in the number of revertants were observed.
- Preincubation assay: In strain TA 100 toxicity was observed at the concentration of 164 µg/plate and above, in the presence of S9. In the absence of S9, toxicity was observed at concentration of 17 and 52 µg/plate. In tester strain WP2uvrA, toxicity was observed at the concentration of 164 µg/plate and above, in the absence of S9. In the presence of S9, no reduction of the bacterial background lawn and no biologically relevant decrease in the number of revertants were observed.

COMPARISON WITH HISTORICAL CONTROL DATA:
- 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.

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- Direct plate assay: In all strains toxicity (microcolonies) was observed at the higher doses (at >= 164 µg/plate), both in the absence and/or presence of S9.
- Preincubation assay: In all strains toxicity (microcolonies) was observed at the concentrations of 17 and 52 µg/plate, in the absence of S9. In the presence of S9, in all strains toxicity was observed at the concentration of 164 µg/plate.

Conclusions:

Interpretation of results (migrated information):
negative

Floral Super is not mutagenic in the Salmonella typhimurium reverse mutation assay and in the Escherichia coli reverse mutation assay performed according to OECD 471 (1997) and GLP principles.

Executive summary:

The mutagenic activity of Floral Super was evaluated in accordance with OECD 471 (1997) guideline and according to GLP principles. The test was performed in two independent experiments: at first a direct plate assay was performed and secondly a pre-incubation assay, both in the absence and presence of S9-mix. The dose levels were selected based on observed cytotoxicity in dose range finding tests in strain TA 100 (direct plate -/+ S9 and preincubation +S9 >= 164 µg/plate, preincubation -S9 17 and 52 µg/plate). Adequate negative and positive controls were included. The substance did not induce a significant dose-related increase in the number of revertant (His+) colonies in each of the four S. typhimurium tester strains (TA1535, TA1537, TA98 and TA100) and E. coli tester strain (WP2uvrA), both in the absence and presence of S9 -metabolic activation. These results were confirmed in independently repeated experiments. Based on the results of this study it is concluded that Floral Super is not mutagenic in the Salmonella typhimurium reverse mutation assay and Escherichia coli reverse mutation assay.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Additional information

Additional information from genetic toxicity in vitro:

Justification for selection of genetic toxicity endpoint
This study result of this study is reliable and adequate for covering this endpoint

Justification for classification or non-classification

Based on the results of the Ames test, Floral Super does not have to be classified for mutagenicity in accordance with Regulation (EC) No. 1272/2008 and Directive 67/548/EEC.