3-hydroxy-4-[(2-methoxy-5-nitrophenyl)azo]-N-(3-nitrophenyl)naphthalene-2-carboxamide

Endpoint summary

Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

In vitro gene mutation study in mammalian cells was conducted by NTP (NATIONAL TOXICOLOGY PROGRAM. Technical Report Series. No.411, 1992) to evaluate the mutagenic potential of target chemical 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4)  in mammalian cell line by chromosome aberration test. The chromosome aberration is a confirmatory test for genetic mutation.The genetic toxicity of C.I. Pigment Red 23 was assessed by testing the ability of the chemical to induce chromosome aberration in Chinese hamster ovary cells. In the Abs test without S9, cells were incubated in McCoy's 5A medium with 30, 50, 100 µg/ml concentration of C.1. Pigment Red 23 for 10-h Colcemid was added and incubation continued for 2 to 3 hours. The cells were then harvested by mitotic shake-off, fixed, and stained with Giemsa.For the Abs test with S9, cells were treated with 30, 50, 100 µg/ml concentration of C.1. Pigment Red 23 and S9 for 2 hours, after which the treatment medium was removed and the cells incubated for 11 hours in fresh medium, with Colcemid present for the final 2 hours. No induction of chromosome aberration was observed in Chinese hamster ovary cell in presence and absence of metabolic activation. Hence the substance can be classified as non mutagenic in vitro.

Link to relevant study records

Reference

Endpoint:

in vitro cytogenicity / chromosome aberration study in mammalian cells

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

secondary literature

Justification for type of information:

Data is from secondary source.

Qualifier:

according to guideline

Guideline:

other: As mention below

Principles of method if other than guideline:

The genetic toxicity of C.I. Pigment Red 23 was assessed by testing the ability of the chemical to induce chromosome aberration in Chinese hamster ovary cells.

GLP compliance:

not specified

Type of assay:

other: chromosome aberration test

Specific details on test material used for the study:

- Name of test material : 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide
- Common name : 3-hydroxy-4-[(2-methoxy-5-nitrophenyl)azo]-N-(3-nitrophenyl)naphthalene-2-carboxamide, C.I. Pigment Red 23
- Molecular formula : C24H17N5O7
- Molecular weight : 487.4263 g/mol
- Smiles notation : c12c(c(c(cc1cccc2)C(=O)Nc1cc(ccc1)[N+](=O)[O-])O)/N=N/c1c(ccc(c1)[N+](=O)[O-])OC
- InChl : 1S/C24H17N5O7/c1-36-21-10-9-17(29(34)35)13-20(21)26-27-22-18-8-3-2-5-14(18)11-19(23(22)30)24(31)25-15-6-4-7-16(12-15)28(32)33/h2-13,30H,1H3,(H,25,31)/b27-26+
- Substance type : Organic
- Physical state : Solid
-Purity->96 %

Species / strain / cell type:

Chinese hamster Ovary (CHO)

Details on mammalian cell type (if applicable):

Not specified

Additional strain / cell type characteristics:

not specified

Cytokinesis block (if used):

not specified

Metabolic activation:

with and without

Metabolic activation system:

Aroclor 1254-induced male Sprague-Dawley rats

Test concentrations with justification for top dose:

0,30, 50, 100 µg/ml

Vehicle / solvent:

Vehicle
- Vehicle(s)/solvent(s) used: DMSO

Untreated negative controls:

not specified

Negative solvent / vehicle controls:

yes

Remarks:

DMSO

True negative controls:

not specified

Positive controls:

yes

Positive control substance:

cyclophosphamide

mitomycin C

Details on test system and experimental conditions:

Details on test system and conditions
METHOD OF APPLICATION: in medium
DURATION
- Preincubation period: No data available
- Exposure duration:
Without metabolic activation- 15 hour
With metabolic activation- 2 hour

- Expression time (cells in growth medium): Without metabolic activation- 15 hour
With metabolic activation- 13 hour

SPINDLE INHIBITOR (cytogenetic assays): Colcemid

STAIN (for cytogenetic assays):stained with Giemsa


OTHER EXAMINATIONS:
- Determination of polyploidy: No data available
- Determination of endoreplication: No data available
- Other:

Evaluation criteria:

Classes of aberrations included simple (breaks and terminal deletions), complex (rearrangements and translocations), and other (pulverized cells, despiralized chromosomes, and cells containing 10 or more aberrations). were observed .

Statistics:

Not applicable.

Key result

Species / strain:

Chinese hamster Ovary (CHO)

Metabolic activation:

with and without

Genotoxicity:

negative

Cytotoxicity / choice of top concentrations:

not specified

Vehicle controls validity:

valid

Untreated negative controls validity:

not specified

Positive controls validity:

valid

Remarks on result:

other: No mutagenic effect were observed

Conclusions:

The pigment C.I. Pigment Red 23 was negative for the induction of chromosomal aberrations in Chinese hamster ovary cells both in the presence and absence of S9.

Executive summary:

The genetic toxicity of C.I. Pigment Red 23 was assessed by testing the ability of the chemical to induce chromosome aberration in Chinese hamster ovary cells.In the Abs test without S9, cells were incubated in McCoy's 5A medium with 30, 50, 100 µg/ml concentration of C.1. Pigment Red 23 for 10-h Colcemid was added and incubation continued for 2 to 3 hours. The cells were then harvested by mitotic shake-off, fixed, and stained with Giemsa.For the Abs test with S9, cells were treated with 30, 50, 100 µg/ml concentration of C.1. Pigment Red 23 and S9 for 2 hours, after which the treatment medium was removed and the cells incubated for 11 hours in fresh medium, with Colcemid present for the final 2 hours. No induction of chromosome aberration was observed in Chinese hamster ovary cell in presence and absence of metabolic activation.

Endpoint conclusion

Endpoint conclusion:

no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion

Endpoint conclusion:

no study available

Additional information

In different studies, 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4) has been investigated for potential of mutagenic response with and without metabolic activation. The studies are based on in vitro gene mutation study in mammalian cells and bacteria for target chemical C.I. Pigment Red 23. Different studies based on these experiments are concluded below

In vitro gene mutation study in mammalian cells was conducted by NTP (NATIONAL TOXICOLOGY PROGRAM. Technical Report Series. No.411, 1992) to evaluate the mutagenic potential of target chemical 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4)  in mammalian cell line by chromosome aberration test. The chromosome aberration is a confirmatory test for genetic mutation.The genetic toxicity of C.I. Pigment Red 23 was assessed by testing the ability of the chemical to induce chromosome aberration in Chinese hamster ovary cells. In the Abs test without S9, cells were incubated in McCoy's 5A medium with 30, 50, 100 µg/ml concentration of C.1. Pigment Red 23 for 10-h Colcemid was added and incubation continued for 2 to 3 hours. The cells were then harvested by mitotic shake-off, fixed, and stained with Giemsa.For the Abs test with S9, cells were treated with 30, 50, 100 µg/ml concentration of C.1. Pigment Red 23 and S9 for 2 hours, after which the treatment medium was removed and the cells incubated for 11 hours in fresh medium, with Colcemid present for the final 2 hours. No induction of chromosome aberration was observed in Chinese hamster ovary cell in presence and absence of metabolic activation. Hence the substance can be classified as non mutagenic in vitro.

Supported by In vitro gene mutation study in mammalian cells conducted by NTP (NATIONAL TOXICOLOGY PROGRAM. Technical Report Series. No.411, 1992) to evaluate the mutagenic potential of target chemical 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4)in mammalian cell line by sister chromatid exchange assay.C.I. Pigment Red 23were tested for mutagenicity with the Chinese hamster ovary cell with and without metabolic activation(Aroclor 1254-induced male Sprague-Dawley rats).In the SCE test without S9,CHO cells were incubated for 26 hours with C.1. Pigment Red 23 of 5, 10, 16 and 30 µg/ml concentration in McCoy's 5A medium supplemented with 10% fetal bovine serum, I-glutamine (2mM), and antibiotics.In the SCE test with S9, cells were incubated with C.I. Pigment Red 23 of 10, 16, 30 and 50 µg/ml concentration, serum-free medium, and S9 for 2 hours. In results,C.I. Pigment Red 23 induced SCE over a concentration range of 5 to50 µg/mL in the absence of S9 in an initial trial; the second trial performed without S9 also demonstrated an increase in SCEs, but only at a higher dose than was evaluated in the first trial. At higher dose a delayed harvest protocol was employed to offset the toxic effect of the pigment on cell cycle progression. There was no induction of SCEs was observed in CHO cells in the presence of liver S9 from Aroclor 1254-induced male Sprague-Dawley rats. Hence,the substance C.I. Pigment Red 23 is considered to be mutagenic in Chinese hamster ovary cell without metabolic activation and non-mutagenic with activation.

Further supported by in vitro gene mutation study in bacteria conducted by Kristien Mortelmans et.al (Environmental Mutagenesis,1986) to evaluate the mutagenic potential of target chemical 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4)in bacterial strain. The genetic toxicity of C.I. Pigment Red 23 was assessed by   screening test in Salmonella typhemurium strain TA1537, TA100 and TA98. The stains were exposed to 10, 33, 100, 333, 1000 and 3333µg/plate concentration of C.I. Pigment Red 23 in the presence and absence of metabolic activation (Hamster and rat liver). Reproducible dose-related increases in the number of revertant was observed. Under this condition the study gives positive mutagenic result. Therefore as per this study, the pigment C.I. Pigment Red 23 was considered to be mutagenic in Salmonella typhemurium strain TA1537, TA100 and TA98.

Bacterial reverse mutation assay is a screening test for mutagenic response, while mammalian cell gene mutation assay is a confirmatory test for mutagenic response. As per the above studies the target chemical 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4) is considered to be non mutagenic according to In vitro gene mutation study in mammalian cells. While according to in vitro gene mutation study in bacteria the target chemical is mutagenic. On the basis of the confirmatory tests, the test substance was observed to be non mutagenic with and without metabolic activator in mammalian cell by chromosome aberration test. Even in the sister chromatid exchange assay the test chemical was non mutagenic in the presence of S9 metabolic activation.This confirmatory test concludes that the test substance shows negative result for mutagenic potential. Thusbased on the available data for the target chemical,3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 does notexhibits gene mutation toxicity in vitro. Hence, the chemical isnot likely to classify as a gene mutant in vitro.

Justification for classification or non-classification

Thus based on the above annotation Bacterial reverse mutation assay is a screening test for mutagenic response, while mammalian cell gene mutation assay is a confirmatory test for mutagenic response. As per the above studies the target chemical 3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl) naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 ( 6471-49-4) is considered to be non mutagenic according to In vitro gene mutation study in mammalian cells. While according to in vitro gene mutation study in bacteria the target chemical is mutagenic. On the basis of the confirmatory tests, the test substance was observed to be non mutagenic with and without metabolic activator in mammalian cell by chromosome aberration test. Even in the sister chromatid exchange assay the test chemical was non mutagenic in the presence of S9 metabolic activation.This confirmatory test concludes that the test substance shows negative result for mutagenic potential. Thusbased on the available data for the target chemical,3-hydroxy-4-[(E)-2-(2-methoxy-5-nitrophenyl)diazen-1-yl]-N-(3-nitrophenyl)naphthalene-2-carboxamide; Other name:C.I. Pigment Red 23 does notexhibits gene mutation toxicity in vitro. Hence, the chemical isnot likely to classify as a gene mutant in vitro.