2,4-bis[N'-(4-methylphenyl)ureido]toluene

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:

September 29, 1992 - November 2, 1992

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

The study has been performed according to OECD 471 (1983), EU Method B.14 (1984) and according to GLP principles. The study was not conducted with E.coli and/or with S. typhimurium TA102, which have an AT base pair at the primary reversion site, as currently required. This study is therefore considered a Salmonella typhimurium reverse mutation assay only. Limited information available to verify the composition of the used test substance.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1992

Report date:

1992

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Version / remarks:

(1983)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

other: EU Method B.14 (1984)

Deviations:

no

Principles of method if other than guideline:

As OECD 471 (1983) required to test Salmonella typhimurium TA1535, TA1537, TA98 and TA100 only, there is no deviation from the guideline.

GLP compliance:

yes (incl. QA statement)

Type of assay:

other: Salmonella typhimurium reverse mutation assay (Ames test)

Test material

Test material information

Constituent 1

Reference substance name:

2,4-bis[N'-(4-methylphenyl)ureido]toluene

EC Number:

411-790-5

EC Name:

2,4-bis[N'-(4-methylphenyl)ureido]toluene

Cas Number:

54390-87-3

Molecular formula:

C23H24N4O2, C24H34N4O2

IUPAC Name:

3-(2-methyl-3-{[(4-methylphenyl)carbamoyl]amino}phenyl)-1-(4-methylphenyl)urea; 3-(2-methyl-5-{[(4-methylphenyl)carbamoyl]amino}phenyl)-1-(4-methylphenyl)urea; 3-{2-methyl-3-[(octylcarbamoyl)amino]phenyl}-1-(4-methylphenyl)urea

Details on test material:

- Name of test material (as cited in study report): KY-MA
- Physical state: Light yellow solid
- Storage condition of test material: At room temperature

Method

Target gene:

Histidine

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 induced by Aroclor 1254 in olive oil

Test concentrations with justification for top dose:

Experiment 1
Preliminary test (without and with S9) TA98 and TA100: 3.3, 10, 33.3, 100, 333.3, 1000, 2500 and 5000 µg/plate
Main study: TA1535, TA1537:
Without and with S9-mix: 33.3, 100, 333.3, 1000, 2500 and 5000 µg/plate
Experiment 2:
Without and with S9-mix: 33.3, 100, 333.3, 1000, 2500 and 5000 µg/plate

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: DMSO
- Justification for choice of solvent/vehicle: because of its solubility properties

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

Positive controls:

yes

Positive control substance:

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

Details on test system and experimental conditions:

METHOD OF APPLICATION: in agar (plate incorporation)

DURATION
- Exposure duration: 48 hour

NUMBER OF REPLICATIONS:
- Doses of the test substance were tested in triplicate in each strain. Two independent experiments were conducted.

DETERMINATION OF CYTOTOXICITY
- Method: The reduction in the number of spontaneous revertants, a clearing of the bacterial background lawn or by degree of survival of treated cultures

OTHER EXAMINATIONS:
- The presence of precipitation of the test compound on the plates was determined.

Evaluation criteria:

- A test article is considered as positive if either a dose related increase in the number of revertants or a significant and reproducible increase for at least one test concentration is induced.
- A test article producing neither a dose related increase in the number of revertants nor a significant and reproducible positive response at any one of the test points is considered non-mutagenic in this system.
- A significant response is described as follows: A test article is considered as mutagenic if in strain TA100 the number of reversions is at least twice as high and in strains TA1535, TA1537 and TA98 it is at least 3 times higher as compared to the spontaneous reversion rate.
- Also, a dose-dependent increase in the number of revertants is regarded as an indication of possibly existing mutagenic potential of the test article regardless whether the highest dose induced the above described enhancement factors or not.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 3

Species / strain:

S. typhimurium TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no cytotoxicity

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

no 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 was observed up to and including the top dose of 5000 µg/plate

RANGE-FINDING/SCREENING STUDIES:
- No toxicity or mutagenicity was observed up to and including the top dose of 5000 µg/plate

COMPARISON WITH HISTORICAL CONTROL DATA: No data

ADDITIONAL INFORMATION ON CYTOTOXICITY:
- No toxicity or mutagenicity was observed up to and including the top dose of 5000 µg/plate

Applicant's summary and conclusion

Conclusions:

The substance is not mutagenic in the Salmonella typhimurium reverse mutation assay.

Executive summary:

The genetic toxicity of the substance was assessed at concentrations of ≤ 5000 µg/plate using S. typhimurium TA100, TA1535, TA98 and TA1537 strains, in accordance with OECD 471 (1983), EU Method B.14 (1984) and according to GLP principles.

All bacterial strains showed negative responses with and without metabolic activation. The study was not conducted with E.coli and/or with S. typhimurium TA102, which have an AT base pair at the primary reversion site, as currently required. Based on the results of this study, the substance is not mutagenic in the Salmonella typhimurium reverse mutation assay.