N-ethyl-o(or p)-toluenesulphonamide

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:

2 September 1983 - 30 October 1983

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1984

Report date:

1984

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

Pre-GLP period

Principles of method if other than guideline:

Test procedures based on the work of Ames et al (1975), "Methods for detecting carcinogens and mutagens with the salmonella mammalian-microsome test"

GLP compliance:

no

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

N-ethyl-o(or p)-toluenesulphonamide

EC Number:

232-465-2

EC Name:

N-ethyl-o(or p)-toluenesulphonamide

Cas Number:

8047-99-2

Molecular formula:

C9H13NO2S

IUPAC Name:

N-ethyl-4-methylbenzene-1-sulfonamide

Test material form:

liquid: viscous

Remarks:

Colour: light yellow

Details on test material:

Ketjenflex 8 (NETSA)
Chemical name in report: N-ethyl-o/p-toluenesulfonamide (or N-substituted toluene sulphonamide)
Description: light yellow viscous liquid
Purity 100% (or 90%)

Test substance storage: at room temperature in the dark
Stability under storage conditions: stable

Specific details on test material used for the study:

- Test Material: Santicizer 8
- Purity: 89%
- Storage condition of test material: stored in the dark at ambient temperature
- Stability under test conditions: indicated not to be unstable to heat, light, and water by the sample submitter

Method

Target gene:

Histidine gene

Species / strain

Species / strain / cell type:

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

Metabolic activation:

with and without

Metabolic activation system:

Rat liver S9-mix ( as described by ames at el.(1975)

Test concentrations with justification for top dose:

TA 1535, TA 1537, TA 98, and TA 100 (without and with S9): 0.01, 0.04, 0.20, 1.00, 3.00 and 10.00 mg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: Solutions of the test material were prepared with ACS grade dimethylsulfoxide (DMSO)

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

2-acetylaminofluorene

4-nitroquinoline-N-oxide

9-aminoacridine

benzo(a)pyrene

other: NaNO2 and 2-aminoanthracene

Details on test system and experimental conditions:

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

METHOD OF APPLICATION:
- Method: plate incorporation method

DURATION
- Exposure duration: 72 hours at 37 degrees celsius

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

Evaluation criteria:

For the test substance to be considered mutagenic, there must be evidence of a dose-response relationship showing increasing numbers of mutant colonies with increasing dose levels. This was done using statistical methods to evaluate the test results ( see below).

Statistics:

Statistical analysis was performed on the plate incorporation assay results after transforming revertant/ plate values as log 10 (revertant/plate). Analysis included Bartlett’s test for homogeneity of variance and comparison of treatments with controls using within-levels pooled variance and a one-sided t-test. Grubb’s test was performed to determine if outliers were present. Dose response was evaluated with regression analysis for log 10 transformed doses and revertants/ plate. Significance of dose-response was evaluated by a t-tester.

Results and discussion

Test results

Test results 1

Key result

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Only the maximum concentration of 10 mg per plate was toxic in the test

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Key result

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Only the maximum concentration of 10 mg per plate was toxic in the test

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Key result

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Only the maximum concentration of 10 mg per plate was toxic in the test

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Key result

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

Only the maximum concentration of 10 mg per plate was toxic in the test

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS
- Precipitation: No precipitation was reported

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- In all strains toxicity was observed at the higher doses (at 10 mg /plate), both in the absence and/or presence of S9, with the exception of TA98 (with the absence of S9).

Applicant's summary and conclusion

Conclusions:

Under the conditions of this study, Santicizer 8 was determined to be not mutagenic and does not need to be classified for mutagenicity in the Salmonella typhimurium reverse mutation assay.

Executive summary:

The mutagenic activity of Santicizer 8 was conducted based on test procedures described by Ames et al (1975) according to OECDTG 471 . Salmonella typhimurium strains TA1535, TA1537, TA98 and TA100 were treated with the test item using the Ames plate incorporation method at up to six dose levels, in triplicate, both with and without the addition of S9-mix. Adequate negative and positive controls were included. Only four strains of bacteria were used, none of which would be able to detect certain oxidizing mutagens, cross- linking agents and hydrazines as detected by E.Coli WP2 strains or s.typhimurium TA102.

The test material, Santicizer 8, was not mutagenic towards salmonella typhimurium test strains TA98, TA100, TA1535 or TA1537 in the plate incorporation assays conducted with and without a rat microsomal activation system. A maximum of 10 mg per plate was used in the plate incorporation tests. Levels of 10 mg per plate were toxic in the plate incooperation test in the presence and absence of a rat microsomal activation system, whereas the lower concentrations tested were not. The plate incorporation test results indicated no significant mutagenic activity for Santicizer 8, and was therefore considered to be non-mutagenic under the conditions of this test.