Reaction mass of bis(N-decyl-N,N-dimethyldecan-1-aminium) carbonate and N-decyl-N,N-dimethyldecan-1-aminium hydrogen carbonate

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: GLP study according to EU- and/or OECD-guidelines

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Report date:

2004

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

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

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Test guideline 3

Qualifier:

according to guideline

Guideline:

EPA OPPTS 870.5100 - Bacterial Reverse Mutation Test (August 1998)

GLP compliance:

yes (incl. QA statement)

Remarks:

Safepharm Laboratories Ltd

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Test material form:

other: solid gel

Details on test material:

- Name of test material: Didecyldimethylammonium carbonate

Method

Target gene:

The study was designed to assess the mutagenic potential of the test material using a bacterial/microsome test system, by exposing histidine auxotrophs of Salmonella typhimurium to various concentrations of the test material.

Species / strain

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

Prior to master stains being used, characterization checks were carried out to determined the amino-acid requirement, presence of rfa, R factors, uvrB mutation and spontaneous reversion rate.
In this assay, overnight sub-cultures of the appropriate coded stock cultures were prepared nutrient broth (Oxoid Limited; lot number 301083 04/08) and incubated at 37 °C for approximately 10 hours. Each culture was monitored spectrophotometrically for turbidity with titres determined by viable count analysis on nutrient agar plates.

Metabolic activation:

with and without

Metabolic activation system:

Activated (phenobarbitone/beta-naphthoflavone, 80/100 mg/kg bw/day) rat liver microsomal S9 fraction

Test concentrations with justification for top dose:

- Concentrations used for cytotoxicity testing: 0, 0.15, 0.5, 1.5, 5, 15, 50, 150, 500, 1500 and 5000 µg/plate
- Concentrations used for genotoxicity testing:
without S9 mix: 0.15, 0.5, 1.5, 5, 15 and 50 µg/plate
with S9 mix: 0.5, 1.5, 5, 15, 50 and 150 µg/plate

Vehicle / solvent:

- Vehicle used: water

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

N-ethyl-N-nitro-N-nitrosoguanidine

benzo(a)pyrene

mitomycin C

other: 2-Aminoanthracene; 1,8-Dihydroxyanthraquinone

Details on test system and experimental conditions:

METHOD OF APPLICATION:
- In agar (plate incorporation)

DURATION
- Exposure duration: 48 hours

SELECTION AGENT (mutation assays):
- Histidine

NUMBER OF REPLICATIONS:
- 3

DETERMINATION OF CYTOTOXICITY
- Method: relative total growth

Evaluation criteria:

The test material should have induced a reproducible, dose-related and statistically significant increase in the revertant count in at least one strain of bacteria.

Statistics:

Dunnett's method of linear regression.

Results and discussion

Test results

Species / strain:

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

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

The test material was considered to be non-mutagenic under the conditions of this test.

Executive summary:

A study was carried out according to EU Method B.13/14, OECD Guideline 471 (Bacterial Reverse Mutation Assay) and EPA OPPTS 870.5100. Salmonella typhimurium strains TA1535, TAl537, TA102, TA98 and TA100 were treated with test material using the Ames plate incorporation method at six dose levels in triplicate, both with and without the addition of a rat liver homogenate metabolizing system (10 % liver S9 in standard co-factors). The dose range was determined in a preliminary toxicity assay ranged between 0.15 to 150 µg/plate in the first experiment. The experiment was repeated on a separate day using a similar dose range to Experiment 1, fresh cultures of the bacterial strains fresh test material formulations.

Additional dose levels were included to allow for test material induced toxicity, ensuring that a minimum of four non-toxic dose levels were achieved.

The vehicle (sterile distilled water) control plates gave counts of revertant colonies within the normal range. All chemicals used in the test induced marked increases in the frequency of revertant colonies, both with or without metabolic activation. Thus, sensitivity of the assay and the efficacy of the S9-mix were validated.

The test material caused a visible reduction in the growth of the bacterial background lawn to all of the tester strains initially at 50 and 15 µg/plate, with and without S9-mix, respectively. The test material was, therefore, tested up to the toxic limit. No test material precipitate was observed on the plates at any of the doses tested in either the presence or absence of S9-mix.

No significant increases in the frequency of revertant colonies were recorded for any the bacterial strains, with any dose of the test material, either with or without metabolic activation.

In conclusion the test material was considered to be non-mutagenic under the conditions this test.