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Key value for chemical safety assessment

Genetic toxicity in vitro

Description of key information

Experimental study on genetic toxicity in vitro was conducted by James L. Simset al.( Biochemistry, 1982) to determine the mutagenic nature of target substance pyrazine-2-carboxylic acid (98-97-5). DNA repair test was conducted for 2-Pyrazinecarboxylic acid (98 -97 - 5) by using human lymphocytes. It was damaged by treating DNA damaging agents such as UV irradiation or N-methyl-N’-nitro-N-nitrosoguanidine (MNNG). The lymphocytecells were exposed to the test substance pyrazine-2-carboxylic acid for 6 hr incubation with [3H]thymidine in the presence of 10 mM hydroxyurea.at 37C.Lymphocyte cells not treated with DNA damaging agents are used as a control. For the DNA repair assay, lymphocyte is prepared in the following manner- Normal human lymphocytes were isolated from peripheral blood and were suspended at 2 × 106cells/mL in complete medium composed of α- modified Eagles medium buffered with 25 mM Hepes, pH 7.2, and supplemented with 10% fetal calf serum, 50 units/mL penicillin, and 50 µg/mL streptomycin. The amount of [3H]dTMP incorporated by DNA-damaged cells incubated in the presence of hydroxyurea was taken as the 100% level for unscheduled DNA synthesis. Stimulation of unscheduled DNA synthesis in the presence of test substance was expressed relative to the 100% level. Results are presented as the means of assays performed in triplicates.96% of DNA synthesis was occurred by the test substance as compared to the control which shows 100% unscheduled DNA synthesis in DNA damaged lymphocytes. The test substance showed 0% inhibition of the enzyme and also was unable to stimulate unscheduled DNA synthesis. Thus, the test was considered to be negative. Therefore 2-Pyrazinecarboxylic acid (98 -97 -5) was not likely to be classified as gene mutant in vitro

Link to relevant study records
Reference
Endpoint:
in vitro DNA damage and/or repair study
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
Data is from publication.
Qualifier:
according to
Guideline:
other: As mention below
Principles of method if other than guideline:
To evaluate the mutagenic potential of 2- Pyrazinecarboxylic acid in Normal human lymphocytes by DNA Repair Synthesis.
GLP compliance:
not specified
Type of assay:
other: DNA Repair Synthesis.
Specific details on test material used for the study:
- Name of test material (as cited in study report): Pyrazine-2-carboxylic acid
- Molecular formula (if other than submission substance): C5H4N2O2
- Molecular weight (if other than submission substance): 124.098 g/mol
- Substance type: Organic
- Physical state: Solid
Target gene:
Thymidine
Species / strain / cell type:
lymphocytes: Normal human
Details on mammalian cell type (if applicable):
Not applicable
Additional strain / cell type characteristics:
not specified
Cytokinesis block (if used):
Not specified
Test concentrations with justification for top dose:
2mM
Vehicle / solvent:
Not specified
Untreated negative controls:
not specified
Negative solvent / vehicle controls:
not specified
True negative controls:
not specified
Positive controls:
not specified
Details on test system and experimental conditions:
Human lymphocyte cells not treated with DNA damaging agents are used as a control.
Rationale for test conditions:
Not specified
Evaluation criteria:
The amount of [3H]dTMP incorporated by DNA-damaged cells incubated in the presence of hydroxyurea was taken as the 100% level for unscheduled DNA synthesis.
Stimulation of unscheduled DNA synthesis in the presence of test substance was expressed relative to the 100% level.
Statistics:
Results are presented as the means of assays performed in triplicate.
Key result
Species / strain:
lymphocytes: Human
Metabolic activation:
not specified
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:
-Other- Pyrazine-2-carboxylic acid showed 0% inhibition of the enzyme, i.e; the compound did not inhibited the enzyme poly(ADP-ribose) polymerase and were unable to stimulate unscheduled DNA synthesis.
Conclusions:
The test substance 2-Pyrazinecarboxylic acid was unable to stimulate the unscheduled DNA synthesis in UV or MNNG treated human lymphocytes. Thus, the test result was found to be negative.
Executive summary:

DNA repair test was conducted for 2-Pyrazinecarboxylic acid (98 -97 - 5) by using human lymphocytes . It was damaged by treating DNA damaging agents such as UV irradiation or N-methyl-N’-nitro-N-nitrosoguanidine (MNNG). The lymphocytecells were exposed to the test substance pyrazine-2-carboxylic acid for 6 hr incubation with [3H]thymidine in the

presence of 10 mM hydroxyurea.at 37ᵒC.Lymphocyte cells not treated with DNA damaging agents are used as a control. For the DNA repair assay, lymphocyte is prepared in the following manner- Normal human lymphocytes were isolated from peripheral blood and were suspended at 2 × 106cells/mL in complete medium composed of α- modified Eagles medium buffered with 25 mM Hepes, pH 7.2, and supplemented with 10% fetal calf serum, 50 units/mL penicillin, and 50 µg/mL streptomycin.The amount of [3H]dTMP incorporated by DNA-damaged cells incubated in the presence of hydroxyurea was taken as the 100% level for unscheduled DNA synthesis. Stimulation of unscheduled DNA synthesis in the presence of test substance was expressed relative to the 100% level. Results are presented as the means of assays performed in triplicates.96% of DNA synthesis was occurred by the test substance as compared to the control which shows 100% unscheduled DNA synthesis in DNA damaged lymphocytes. The test substance showed 0% inhibition of the enzyme and also were unable to stimulate unscheduled DNA synthesis. Thus, the test was considered to be negative. Therefore 2-Pyrazinecarboxylic acid (98 -97 -5) was not likely to be classified as gene mutant in vitro.

Endpoint conclusion
Endpoint conclusion:
no adverse effect observed (negative)

Genetic toxicity in vivo

Endpoint conclusion
Endpoint conclusion:
no study available

Additional information

Genotoxicity In-vitro

Various publications were reviewed to determine the mutagenic nature of pyrazine-2-carboxylic acid (98-97-5). The studies are as mentioned below:

Experimental study on genetic toxicity in vitro was conducted by James L. Simset al.( Biochemistry, 1982) to determine the mutagenic nature of target substance pyrazine-2-carboxylic acid (98-97-5). DNA repair test was conducted for 2-Pyrazinecarboxylic acid (98 -97 - 5) by using human lymphocytes. It was damaged by treating DNA damaging agents such as UV irradiation or N-methyl-N’-nitro-N-nitrosoguanidine (MNNG). The lymphocytecells were exposed to the test substance pyrazine-2-carboxylic acid for 6 hr incubation with [3H]thymidine in the presence of 10 mM hydroxyurea.at 37C.Lymphocyte cells not treated with DNA damaging agents are used as a control. For the DNA repair assay, lymphocyte is prepared in the following manner- Normal human lymphocytes were isolated from peripheral blood and were suspended at 2 × 106cells/mL in complete medium composed of α- modified Eagles medium buffered with 25 mM Hepes, pH 7.2, and supplemented with 10% fetal calf serum, 50 units/mL penicillin, and 50 µg/mL streptomycin. The amount of [3H]dTMP incorporated by DNA-damaged cells incubated in the presence of hydroxyurea was taken as the 100% level for unscheduled DNA synthesis. Stimulation of unscheduled DNA synthesis in the presence of test substance was expressed relative to the 100% level. Results are presented as the means of assays performed in triplicates.96% of DNA synthesis was occurred by the test substance as compared to the control which shows 100% unscheduled DNA synthesis in DNA damaged lymphocytes. The test substance showed 0% inhibition of the enzyme and also was unable to stimulate unscheduled DNA synthesis. Thus, the test was considered to be negative. Therefore 2-Pyrazinecarboxylic acid (98 -97 -5) was not likely to be classified as gene mutant in vitro.

 Supporting gene mutation toxicity was predicted for Pyrazine-2-carboxylic acid (98-97-5) using the battery approach from Danish QSAR database (2017). The study assumed the use of Salmonella typhimurium bacteria in the Ames test. The end point for gene mutation has been modeled in the Danish QSAR using the three software systems Leadscope, CASE Ultra and SciQSAR. Based on predictions from these three systems, a fourth and overall battery prediction is made. The battery prediction is made using the so called Battery algorithm. With the battery approach it is in many cases possible to reduce “noise” from the individual model estimates and thereby improve accuracy and/or broaden the applicability domain.

In a Gene mutation toxicity study was performed by Errol Zeiger et al.( Environmental Mutagenesis,1987) to determine the mutagenic nature of Pyrazinamide (RA CAS no98-96-4; IUPAC name: pyrazine-2-carboxamide . The read across substances share high similarity in structure and log kow .Therefore, it is acceptable to derive information on mutation from the analogue substance. Mutagenicity effect of Pyrazinamide was evaluated in Salmonella typhimurium. Salmonella typhimurium strains TA100, TA1535, TA1537, TA98 were used in the study. Mutagenic assay was performed by preincubation method. The assay performed in presence and absence pf metabolic activator i.e. Aroclor 1254-induced rat liver S-9 and Aroclor 1254-induced hamster liver S-9.DMSO was used as solvent for test substance.The test chemical, Salmonella culture and S-9 mix or buffer incubated at 37 °C for 20 min without shaking. Agar was added on plate, content of tubes were mixed poured on surface of petri dish that contain Vogel Bonner medium. The mutant colonies on plates were counted after 2 days post incubation. Doses of test substance 0, 100, 333, 1000, 3333, 10000 ug/plate were tested in triplicate. Experiments were repeated 1 week following of initial trial. Sodium azide was used as positive control substance in absence of metabolic activation for strains TA100, TA1535. 9-aminocaridine and 4-nitro-o-phenylenediamine were positive control in absence of metabolic activation for strains TA1537 and TA98 respectively. The positive control with metabolic activation for all strains was 2-aminoanthracene. In results of study, there was no increase in dose related mean number of revertant colonies compared to solvent control across all strains.Therefore, Pyrazinamide was not mutagenic in bacteria Salmonella typhimurium. Hence Pyrazinamide not likely to be classified as gene mutation in vitro.

In a Gene mutation toxicity study was performed by T.B. Adams et al (Food and Chemical Toxicology, 2002) to determine the mutagenic nature of Pyrazinamide (98-96-4); IUPAC name: pyrazine-2-carboxamide. The read across substances share high similarity in structure and log kow .Therefore, it is acceptable to derive information on mutation from the analogue substance. Mutagenicity effect of Pyrazinamide was evaluated in Salmonella typhimurium. Salmonella typhimurium strains TA100, TA1535, TA1537, TA98 were used in the study. Mutagenic assay was performed by preincubationmethod. The assay performed in presence and absence pf metabolic activator i.e. Aroclor 1254-induced rat liver S-9 and Aroclor 1254-induced hamster liver S-9.DMSO was used as solvent for test substance.The test chemical, Salmonella culture and S-9 mix or buffer incubated at 37 °C for 20 min without shaking. Agar was added on plate, content of tubes were mixed poured on surface of petri dish that contain Vogel Bonner medium. The mutant colonies on plates were counted after 2 days post incubation. Doses of test substance 0, 100, 333, 1000, 3333, 10000 ug/plate were tested in triplicate. Experiments were repeated 1 week following of initial trial. Sodium azide was used as positive control substance in absence of metabolic activation for strains TA100, TA1535. 9-aminocaridine and 4-nitro-o-phenylenediamine were positive control in absence of metabolic activation for strains TA1537 and TA98 respectively. The positive control with metabolic activation for all strains was 2-aminoanthracene. In results of study, there was no increase in dose related mean number of revertant colonies compared to solvent control across all strains.Therefore, Pyrazinamide was not mutagenic in bacteria Salmonella typhimurium. Hence Pyrazinamide not likely to be classified as gene mutation in vitro.

Based on the data available for the target chemical and its read across substance, it is concluded that pyrazine-2-carboxylic acid (98-97-5) does not exhibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant in vitro.

Justification for classification or non-classification

Thus based on the data available and CLP criteria for the target chemical , it is concluded that pyrazine-2-carboxylic acid (98-97-5) does not exhibit gene mutation in vitro. Hence the test chemical is not likely to classify as a gene mutant in vitro.