Amines, C10-14-branched and linear alkyl, bis[2-[(4,5-dihydro-3-methyl-5-oxo-1-phenyl-1H-pyrazol-4-yl)azo]benzoato(2-)]chromate(1-)

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Based on the available information, the test item is not mutagenic in the Ames test under the selected experimental conditions.

Link to relevant study records

Reference

Endpoint:

in vitro gene mutation study in bacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

01 Mar 2000 to 19 Dec. 2000

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 471 (Bacterial Reverse Mutation Assay)

GLP compliance:

yes (incl. QA statement)

Type of assay:

bacterial reverse mutation assay

Specific details on test material used for the study:

- Name of the test substance used in the study report: Neozapon Gelb R Stuecke ber. 100%
- Purity: Ca. 96.6%

performed under old CAS 73297-13-9
new CAS 85029-58-9, same structure

Target gene:

S. typhimurium: his
E. coli: trp

Species / strain / cell type:

S. typhimurium TA 1535, TA 1537, TA 98, TA 100 and E. coli WP2

Metabolic activation:

with and without

Metabolic activation system:

Aroclor-induced rat liver S-9 mix resp. uninduced hamster liver S-9 mix

Test concentrations with justification for top dose:

20 µg - 5000 µg/plate (Standard Plate Test)
4 µg - 2,500 µg/plate (Prival Preincubation Test)

Vehicle / solvent:

- Vehicle: DMSO
- Justification for choice of solvent/vehicle: Due to the limited solubility of the test substance in water, DMSO was selected as the vehicle, which had been demonstrated to be suitable in bacterial reverse mutation tests and for which historical control data are available

Untreated negative controls:

yes

Negative solvent / vehicle controls:

yes

True negative controls:

yes

Positive controls:

yes

Positive control substance:

4-nitroquinoline-N-oxide

9-aminoacridine

congo red

other: 2-aminoanthracene (2-AA), benzidine, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), 4-nitro-o-phenylendiamine (NOPD)

Details on test system and experimental conditions:

Ames Standard Plate Test and Prival Preincubation Test

Evaluation criteria:

The test chemical is considered positive in this assay if the following criteria are met :
A dose-related and reproducible increase in the number of revertant colonies, i .e. about doubling of the spontaneous mutation rate in at least one tester strain either without S-9 mix or after adding a metabolizing system .
A test substance is generally considered nonmutagenic in this test if:
The number of revertants for all tester strains were within the historical negative control range under all experimental conditions in two experiments carried out
independently of each other.

Statistics:

no statistics, standard deviation calculated

Key result

Species / strain:

S. typhimurium, other: TA 1535, TA 1537, TA 98, TA 100

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

A bacteriotoxic effect was observed under all test conditions

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Key result

Species / strain:

E. coli WP2

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

cytotoxicity

Remarks:

A bacteriotoxic effect was observed under all test conditions

Vehicle controls validity:

valid

Untreated negative controls validity:

valid

Positive controls validity:

valid

Additional information on results:

An increase in the number of his+ or trp+ revertants was not observed either in the Ames standard plate test or in the Prival preincubation test without S-9 mix or after the addition of a metabolizing system.

TOXICITY
A weak bacteriotoxic effect (slight decrease in the number of revertants, slight reduction in the titer) was observed in the Ames standard plate test using the Salmonella strains depending on the strain and test conditions from about 2,500 pg/plate onward . In the Prival preincubation assay bacteriotoxicity (reduced his- or trp- background growth, decrease in the number of revertants, reduction in the titer) was observed depending on the strain and test conditions from about 100 pg - 500 pg/plate onward .
SOLUBILITY
Test substance precipitation was found from about 500 pg/plate onward .

Table: Standard Test (average result of 3 plates, respectively)

Concentration [µg/plate]	TA 1535		TA 1537		TA 98		TA 100		E. coli WP2 uvrA
	-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9
Solvent control (DMSO)	18	19	10	11	31	38	114	128	36	36
20	20	17	9	10	21	30	96	116	25	45
100	18	18	8	12	20	43	91	113	29	43
500	17	15	5	11	20	25	98	86	25	34
2500	11	11	4	5	13	16	58	24	25	37
5000	11	7	3	1	12	5	62	16	25	35
Positive control	647	165	601	103	1081	580	619	722	550	225

Table: Preincubation test (average result of 3 plates, respectively)

Concentration [µg/plate]	TA 1535		TA 1537		TA 98		TA 100		E. coli WP2 uvrA
	-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9	-S9	+S9
Solvent control (DMSO)	17	17	9	13	25	35	108	109	36	29
Test item
4	19	17	8	12	21	35	105	105	37	25
20	16	16	5	11	20	30	60	89	33	23
100	13	16	4	10	15	25	56	81	27	27
500	12	11	1	5	12	20	31	59	26	25
2500	8	6	2	1	10	9	49	14	24	19
Positive control
MNNG	1093						1190
2-AA		178		104		561		622		220
AAC			632
NOPD					990
CONGOR.						370
BENZID						579
4-NQO									519

MNNG: N-methyl-N'-nitro-N-nitrosoguanidine

2 -AA: 2-aminoanthracene

AAC: 9-aminoacridine

NOPD: 4 -nitro-o-phenylendiamine

4 -NQO: 4 -Nitroquinoline-N-oxide

BENZID: Benzidine

CONGOR.: Congo red

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

Genetic toxicity in vitro:

In a reverse gene mutation assay in bacteria (Ames-test), strains TA 1535, TA 100, TA 1537 and TA 98 of Salmonella typhimurium and strain WP2 uvrA of E. coli were exposed to the test substance at concentrations of 20, 100, 500, 2500 and 5000 µg/plate (BASF, 2000). Standard plate test and preincubation test both were conducted with and without metabolic activation (Aroclor induced rat liver S-9 mix). The positive controls induced the appropriate responses in the corresponding strains. There was no evidence of induced mutant colonies over background. Based on the available information, the test substance is not mutagenic in the Ames test under the selected experimental conditions.

Justification for classification or non-classification

Based on the available information classification for genetic toxicity is not warranted in accordance with EU Classification, Labeling and Packaging of Substances and Mixtures (CLP) Regulation No. 1272/2008.