4-chloro-2-nitroaniline

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:

1983

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: well performed GLP and OECD guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1983

Report date:

1983

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

OECD Guideline 472 (Genetic Toxicology: Escherichia coli, Reverse Mutation Assay)

Deviations:

no

GLP compliance:

yes

Type of assay:

bacterial reverse mutation assay

Test material

Test material information

Constituent 1

Reference substance name:

4-chloro-2-nitroaniline

EC Number:

201-925-4

Cas Number:

89-63-4

Molecular formula:

C6H5ClN2O2

IUPAC Name:

4-chloro-2-nitroaniline

Method

Species / strain

Species / strain 1

Species / strain / cell type:

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

Species / strain 2

Species / strain / cell type:

S. typhimurium TA 1538

Species / strain 3

Species / strain / cell type:

E. coli WP2 uvr A

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254(R) induced rat liver S9-mix

Test concentrations with justification for top dose:

Preliminary toxicity tests were performed with all tester strains at doses of 4 to 10 000 µg/plate and proved to be toxic to most of the strains at a dose of 2 500 µg/plate. Therefore for mutagenicity testing 5 000 µg/plate was chosen as the highest dose.

Toxicity experiment and dose range finding: 0, 4, 20, 100, 500, 2 500, 10 000 µg/plate (with and without metabolic activation)
Mutagenicity experiment: 0, 4, 20, 100, 500, 2 500, 5 000 µg/plate without metabolic activation
0, 4, 20, 100, 500, 1 000, 2 500, 5 000 µg/plate with metabolic activation

With strains TA 98 and TA 1538 a second mutagenicity expermiment was performed

Vehicle / solvent:

- Vehicle(s)/solvent(s) used: dest. water

Controls

Untreated negative controls:

no

Negative solvent / vehicle controls:

yes

True negative controls:

no

Positive controls:

yes

Positive control substance:

other: Na-azide, 9-Aminoacridine, 2-Nitrofluorene, N-Methyl-N-nitro-N-nitrosoguanidine, Benzo[a]pyrene, 2-Aminoanthracene

Details on test system and experimental conditions:

METHOD OF APPLICATION:
in agar (plate incorporation)

DURATION
- Exposure duration: incubation for at least 48 to 72 hours

NUMBER OF REPLICATIONS: 3 plates per strain and dose level, including controls, two independent experiments

Evaluation criteria:

negative result: no significant and no dose dependant increase in the number of revertant colonies

Statistics:

Arithmetic means of the counted colonies were calculated.

Results and discussion

Test results

Test results 1

Species / strain:

S. typhimurium TA 98

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

10 000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 2

Species / strain:

S. typhimurium TA 98

Metabolic activation:

with

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

10 000 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

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:

cytotoxicity

Remarks:

2 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 4

Species / strain:

S. typhimurium TA 1535

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

2 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 5

Species / strain:

S. typhimurium TA 1537

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

2 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 6

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

2 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 7

Species / strain:

S. typhimurium TA 1538

Metabolic activation:

with

Genotoxicity:

positive

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

2 500 µg/plate

Vehicle controls validity:

valid

Untreated negative controls validity:

not examined

Positive controls validity:

valid

Test results 8

Species / strain:

E. coli WP2 uvr A

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

at 2 500 µg/plate

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'.

Any other information on results incl. tables

In the absence of the metabolic activation system the test compound did not show a dose dependent increase in the number of revertants in any of the bacterial strains. In the presence of metabolic activation, treatment of the cells with the test item resulted in relevant increases in the number of revertant colonies with the S. typhimurium strains TA 98 and TA 1538.

The test compound proved to be toxic to most of the bacterial strains at 2 500 µg/plate.

Table 1: Results Experiment I TA 98 and TA 1538 with metabolic activation

Dose µg/plate	TA 98	TA 1538
	Number of colonies (mean)
0	28	11
4	26	14
20	26	17
100	27	14
500	33	32
1 000	48	42
2 500	48	29

Table 1: Results Experiment II TA 98 and TA 1538 with metabolic activation

Dose µg/plate	TA 98	TA 1538
	Number of colonies (mean)
0	31	16
4	23	19
20	33	16
100	35	24
500	63	53
1 000	68	59
2 500	68	56

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):
positive with metabolic activation

The test item, a mixture containing 50 % 4-chloro-2-nitroaniline, is mutagenic in S. typhymurium strains TA 98 and 1538 in the presence of exogenous metabolic activation.

Executive summary:

Mutagenic activity of the test item was investigated in Salmonella typhymurium strains 1535, 1537, 98, 100 and 1538 as well as Escherichia coli WP2 uvrA with and without metabolic activation (induced rat liver S9-mix) at concentrations of 0, 4, 20, 100, 500, 1 000, 2 500 and 5 000 µg/plate. Under the conditions tested the test item, a mixture containing 50 % 4 -chloro-2 -nitroaniliine, caused an increase in the number of revertant colonies in S. typhymurium TA 98 and TA 1538 in the presence of metabolic activation.