Benzenesulfonic acid, C10-60-alkyl derivs., barium salts

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

Study period:

The experimental phase of this study was performed between 14 January 1988 and 19 January 1988

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

Study conducted in compliance with agreed protocols, with some deviations from standard test guidelines and/or minor methodological deficiencies, which limit the reliability of the relevant results. The study report was conclusive, recognizing methodological limitations; the study was conducted under GLP conditions.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1988

Report date:

1988

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

yes

Remarks:

Single experiment and only four strains of Salmonella used.

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

61789-86-4

IUPAC Name:

61789-86-4

Constituent 2

Reference substance name:

Sulfonic acids, petroleum, calcium salts

EC Number:

263-093-9

EC Name:

Sulfonic acids, petroleum, calcium salts

IUPAC Name:

263-093-9

Details on test material:

Physical State: Liquid (viscous)
Color: Dark brown
Solubility: Tetrahydrofuran (THF) at 100 mg/ml
Storage Conditions: Room temperature
Safety Precautions: Avoid topical and respiratory contact

Method

Target gene:

Histidine for Salmonella.

Species / strain

Species / strain / cell type:

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

Details on mammalian cell type (if applicable):

Not applicable.

Additional strain / cell type characteristics:

not applicable

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254 treated male Sprague Dawley rats

Test concentrations with justification for top dose:

All strains except TA1537 without S9 (-S9) at 5000 1500, 500, 150, 50 and 15 pg/plate, and at 2500, 750, 250, 75, 25 and 7.5 pg/plate for TA1537 -S9.

Vehicle / solvent:

The test sample was dissolved in tetrahydrofuran (THF) at 100 mg/ml. All dilutions were made fresh in THF. A fresh stock solution was made for each
assay.

Controls

Controls 1

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA100

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene: 2 µg/plate

Remarks:

With S9 mix

Controls 2

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA1535

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene: 2 µg/plate

Remarks:

With S9 mix

Controls 3

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA1537

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: 2-Aminoanthracene: 2 µg/plate

Remarks:

With S9 mix

Controls 4

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA98

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Remarks:

Example:

Positive controls:

yes

Positive control substance:

benzo(a)pyrene

Remarks:

With S9 mix Migrated to IUCLID6: 2-Aminoanthracene: 2 µg/plate

Controls 5

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA98

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

no

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

Remarks:

without S9 mix Migrated to IUCLID6: 4-Nitrofluorene: 10 µg/plate

Controls 6

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA1537

Negative solvent / vehicle controls:

yes

Remarks:

Sterile distilled water.

True negative controls:

no

Positive controls:

yes

Positive control substance:

9-aminoacridine

Remarks:

without S9 mix Migrated to IUCLID6: 9-Aminoacridine: 50 µg/plate

Controls 7

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA100

Negative solvent / vehicle controls:

yes

Remarks:

Sterile distilled water.

True negative controls:

no

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

without S9 mix Migrated to IUCLID6: Sodium azide: 10 µg/plate

Controls 8

Untreated negative controls:

yes

Remarks:

Spontaneous mutation rates of TA1535

Negative solvent / vehicle controls:

yes

Remarks:

Sterile distilled water

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

N-ethyl-N-nitro-N-nitrosoguanidine

Remarks:

Without S9 mix Migrated to IUCLID6: Sodium azide: 10 µg/plate

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Preincubation period for bacterial strains: overnight incubation
- Exposure duration: Approximately 65 hrs
- Expression time (cells in growth medium): Not applicable
- Selection time (if incubation with a selection agent): Not applicable

NUMBER OF REPLICATIONS: Duplicate plating.

DETERMINATION OF CYTOTOXICITY
- Method: plates were assessed for numbers of revertant colonies and examined for effects on the growth of the bacterial background lawn.

Evaluation criteria:

Acceptance Criteria:

The reverse mutation assay may be considered valid if the following criteria are met:
All tester strain cultures exhibit a characteristic number of spontaneous revertants per plate in the vehicle and untreated controls.
The appropriate characteristics for each tester strain have been confirmed, eg rfa cell-wall mutation and pKM101 plasmid R-factor etc.
All tester strain cultures should be in the approximate range of 1 to 9.9 x 109 bacteria per ml.

Evaluation criteria:

The mean number of revertants per plate and the standard deviation was calculated for each concentration and strain. A test result was considered
positive if for any strain, a significant increase over the negative control in the number of revertants per plate was observed which was concentration-dependent. A significant increase was a two-fold increase when the background was 50 revertants/plate or greater; a three-fold increase when the background was between 10 and 49 revertants/plate; and a four-fold increase when the background was less than 10 revertants/plate.

Statistics:

Standard deviation

Results and discussion

Test results

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:

no cytotoxicity nor precipitates, but tested up to recommended limit concentrations

Remarks:

Tested up to maximum recommended dose of 5000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Solubility: The test sample was dissolved in tetrahydrofuran (THF) at 100 mg/ml. All dilutions were made fresh in THF. A fresh stock solution was
made for eachassay.
- Precipitation: No test material precipitate was observed on the plates at any of the doses tested in either the presence or absence of S9-mix.

RANGE-FINDING/SCREENING STUDIES:
The test sample was non-toxic to the strains of bacteria used. The test sample formulation and S9-mix were both shown to be sterile.

COMPARISON WITH HISTORICAL CONTROL DATA:
Prior to use, the master strains were checked for characteristics, viability and spontaneous reversion rate (all were found to be satisfactory).

Results for the negative controls (spontaneous mutation rates) were considered to be acceptable.

All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies thus confirming the activity of the S9-mix and the sensitivity of the bacterial strains.

ADDITIONAL INFORMATION ON CYTOTOXICITY: None

Remarks on result:

other: all strains/cell types tested

Remarks:

Migrated from field 'Test system'.

Any other information on results incl. tables

RESULTS

Prior to use, the master strains were checked for characteristics, viability and spontaneous reversion rate (all were found to be satisfactory). 

Results for the negative controls (spontaneous mutation rates) are were considered to be acceptable. 

The mean number of revertant colonies and the standard deviations for the test sample, vehicle and positive controls both with and without metabolic activation, are presented as Appendix 1 in 'Attached documents'.

The test sample caused no visible reduction in the growth of the bacterial background lawn at any dose level and was tested up to the maximum recommended dose level of 5000 µg/plate. No test sample 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 of the bacterial strains, at any dose level either with or without metabolic activation.

All of the positive control chemicals used in the test induced marked increases in the frequency of revertant colonies thus confirming the activity of the S9-mix and the sensitivity of the bacterial strains.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
negative

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

Executive summary:

Introduction.

The purpose of this study was to investigate the potential of the test substance, OS 68022C, to induce mutations in Salmonella typhimurium using an in vitro mutagenesis assay.

Methods.

Salmonella typhimurium strains TA1535, TA1537, TA98 and TA100 were treated with the test sample using the Ames plate incorporation method at six dose levels, in triplicate, both with and without the addition of a rat liver homogenate metabolising system (10% liver S9 in standard co-factors) except for TA1537 (absence of S9 -mix only).

Results.

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

The test sample caused no visible reduction in the growth of the bacterial background lawn at any dose level and was tested up to the maximum recommended dose level of 5000 µg/plate. No test sample 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 of the bacterial strains, with any dose of the test material, either with or without metabolic activation.

Conclusion.

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