[8-[(p-aminophenyl)azo]-7-hydroxy-2-naphthyl]trimethylammonium chloride

Genetic toxicity: in vitro

Administrative data

Endpoint:

in vitro gene mutation study in mammalian cells

Remarks:

Type of genotoxicity: gene mutation

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

4 (not assignable)

Data source

Reference

Reference Type:

review article or handbook

Title:

SCIENTIFIC COMMITTEE ON CONSUMER PRODUCTS SCCP Opinion on Basic Brown 17.

Author:

European Commission, Directorate – General for Health & Consumers.

Year:

2008

Bibliographic source:

Scientific Committee on Consumer Safety (SCCS) - report no. SCCP/1173/08

Materials and methods

Test guideline

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 476 (In Vitro Mammalian Cell Gene Mutation Test)

Principles of method if other than guideline:

In vitro Mouse Lymphoma assay (tk locus) was carried out using L5178Y Mouse lymphoma cells.

GLP compliance:

not specified

Type of assay:

other: In vitro Mouse Lymphoma assay (tk locus)

Test material

Test material information

Constituent 1

Reference substance name:

[8-[(4-amino-3-nitrophenyl)azo]-7-hydroxy-2-naphthyl]trimethylammonium chloride

EC Number:

269-944-0

EC Name:

[8-[(4-amino-3-nitrophenyl)azo]-7-hydroxy-2-naphthyl]trimethylammonium chloride

Cas Number:

68391-32-2

IUPAC Name:

8-[(4-amino-3-nitrophenyl)diazenyl]-7-hydroxy-N,N,N-trimethylnaphthalen-2-aminium chloride

Constituent 2

Reference substance name:

(8-((4-Amino-3-nitrophenyl)azo)-7-hydroxy-2-naphthyl)trimethylammonium chloride

IUPAC Name:

(8-((4-Amino-3-nitrophenyl)azo)-7-hydroxy-2-naphthyl)trimethylammonium chloride

Test material form:

other: Dark brown powder

Details on test material:

- Name of test material (as cited in study report): (8-((4-Amino-3-nitrophenyl)azo)-7-hydroxy-2-naphthyl)trimethylammonium chloride- Molecular formula (if other than submission substance): C19H20N5O3.Cl- Molecular weight (if other than submission substance): 401.852 g/mol- Smiles notation (if other than submission substance):[N+](c1cc(\N=N\c2c3cc([N+](C)(C)C)ccc3ccc2O)ccc1N)([O])=O.[ClH]- InChl (if other than submission substance): 1S/C19H19N5O3.ClH/c1-24(2,3)14-7-4-12-5-9-18(25)19(15(12)11-14)22-21-13-6-8-16(20)17(10-13)23(26)27;/h4-11H,1-3H3,(H2-,20,21,22,25);1H- Structural formula attached as image file (if other than submission substance): No data available- Substance type: Organic- Physical state: Solid- Analytical purity: No data available- Impurities (identity and concentrations): No data available- Composition of test material, percentage of components: No data available- Isomers composition: No data available- Purity test date: No data available- Lot/batch No.: No data available- Expiration date of the lot/batch: No data available- Radiochemical purity (if radiolabelling): No data available- Specific activity (if radiolabelling): No data available- Locations of the label (if radiolabelling): No data available- Expiration date of radiochemical substance (if radiolabelling): No data available- Stability under test conditions: No data available- Storage condition of test material: No data available - Other: No data available

Method

Target gene:

tk locus

Species / strain

Species / strain / cell type:

mouse lymphoma L5178Y cells

Test concentrations with justification for top dose:

Experiment I: 8.1, 16.3, 32.5, 65.0 and 97.5 μg/ml without S9-mix16.3, 32.5, 65.0, 130.0 and 195.0 μg/ml with S9-mixExperiment II: 8.0, 16.0, 32.9, 64.0, 128.0 and 192.0 μg/ml without

Vehicle / solvent:

Deionised water was used as a solvent.

Controls

Untreated negative controls:

yes

Negative solvent / vehicle controls:

not specified

True negative controls:

not specified

Positive controls:

yes

Details on test system and experimental conditions:

METHOD OF APPLICATION: No data availableDURATION- Preincubation period: No data available- Exposure duration:Experiment I: 4 h treatment without and with S9-mix Experiment II: 24 h treatment without S9-mix- Expression time (cells in growth medium): Experiment I: expression period 72 h Experiment II: expression period 48 h- Selection time (if incubation with a selection agent):Experiment I: selection period of 10-15 daysExperiment II: selection period of 10-15 days- Fixation time (start of exposure up to fixation or harvest of cells): No data availableSELECTION AGENT (mutation assays): No data availableSPINDLE INHIBITOR (cytogenetic assays): No data availableSTAIN (for cytogenetic assays): No data availableNUMBER OF REPLICATIONS: Two parallel cultures in 2 independent experimentsNUMBER OF CELLS EVALUATED: No data availableDETERMINATION OF CYTOTOXICITY- Method: mitotic index; cloning efficiency; relative total growth; other: No data availableOTHER EXAMINATIONS:- Determination of polyploidy: No data available- Determination of endoreplication: No data available- Other: No data availableOTHER: No data available

Evaluation criteria:

Toxicity was measured in the main experiments as percentage relative total growth of the treated cultures relative to the total growth of the solvent control cultures.

Statistics:

No data available

Results and discussion

Test results

Species / strain:

mouse lymphoma L5178Y cells

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

not specified

Untreated negative controls validity:

not specified

Positive controls validity:

not specified

Additional information on results:

TEST-SPECIFIC CONFOUNDING FACTORS- Effects of pH: No data available- Effects of osmolality: No data available- Evaporation from medium: No data available- Water solubility: No data available- Precipitation: In experiment I, precipitation was noted at 97.5 and 130.0 μg/ml without S9-mix and at 195.0 and 260.0 μg/ml with S9-mix; in experiment II precipitation occurred at 128.0 and 192.0 μg/ml.- Other confounding effects: The appropriate level of toxicity (10-20% survival after the highest dose) was not reached in experiment with S9-mix pointing to insufficient exposure of the cells.Both in experiment I and II no biological relevant and dose dependent increase in the number mutant colonies was observed independent of the presence or absence of S9-mix.RANGE-FINDING/SCREENING STUDIES: Test concentrations were based on the results of a pre-test on toxicity measuring relative suspension growth.COMPARISON WITH HISTORICAL CONTROL DATA: No data availableADDITIONAL INFORMATION ON CYTOTOXICITY: No data available

Remarks on result:

other: other: L5178Y Mouse lymphoma cells

Remarks:

Migrated from field 'Test system'.

Applicant's summary and conclusion

Conclusions:

Interpretation of results (migrated information):other: Negative (with and without)The test substance (8-((4-Amino-3-nitrophenyl)azo)-7-hydroxy-2-naphthyl)trimethylammonium chloride did not induce the mutation in the L5178Y Mouse lymphoma cells (with and without metabolic activation system). Thus, the test was considered to be negative.

Executive summary:

In vitro Mouse Lymphoma assay (tk locus) was carried outfor gene mutations at thetklocus of mouse lymphoma cells both in the absence and presence of S9 metabolic activation.

 

Test concentrations were based on the results of a pre-test on toxicity measuring relative suspension growth. Test chemical conc. used for the two experiment was given below-

 

Experiment I:

8.1, 16.3, 32.5, 65.0 and 97.5 μg/ml without S9-mix

16.3, 32.5, 65.0, 130.0 and 195.0 μg/ml with S9-mix

 

Experiment II:

8.0, 16.0, 32.9, 64.0, 128.0 and 192.0 μg/ml without

 

Deionised water was used as a solvent.

 

In the main test, cells were treated for 4 h or 24 h (without S9 in experiment II only) followed by an expression period of 72 or 48 h to fix the DNA damage into a stabletkmutation. Liver S9 fraction from phenobarbital/β-naphthoflavone-induced rats was used as exogenous metabolic activation system. Toxicity was measured in the main experiments as percentage relative total growth of the treated cultures relative to the total growth of the solvent control cultures. Negative and positive controls were in accordance with the OECD guideline.

 

In experiment I, precipitation was noted at 97.5 and 130.0 μg/ml without S9-mix and at 195.0 and 260.0 μg/ml with S9-mix; in experiment II precipitation occurred at 128.0 and 192.0 μg/ml.

 

The appropriate level of toxicity (10-20% survival after the highest dose) was not reached in experiment with S9-mix pointing to insufficient exposure of the cells. Both in experiment I and II no biological relevant and dose dependent increase in the number mutant colonies was observed independent of the presence or absence of S9-mix.

 

The test substance(8-((4-Amino-3-nitrophenyl)azo)-7-hydroxy-2-naphthyl)trimethylammonium chloridedid not induce the mutation in theL5178Y Mouse lymphoma cells(with and without metabolic activation system). Thus, the test was considered to be negative.Thus, it can be concluded that the test substance has negative genetic toxicity effects and based on the CLP criteria for classification, it cannot be classified as genotoxic.