Selection of solvent and positive control concentration for enhanced Ames test conditions for N-nitrosamine compounds

• Published in: Regulatory toxicology and pharmacology Vol. 154; p. 105711
• Main Authors: Kajavadara, Chetan K., Patel, Satyam N., Shukla, Rushikesh M., Valani, Darshan T., Patel, Rajesh J., Bhatt, Laxit K., Sundar, Rajesh, Jain, Mukul R.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier Inc 01.12.2024
• Subjects: 1-Cyclopentyl-4-nitrosopiperazine; Enhanced Ames test conditions; Metabolic activation system; N-nitrosamine impurities; N-nitrosodimethylamine; Solvents; Solvents; N-nitrosamine impurities; Enhanced Ames test conditions; N-nitrosodimethylamine; Metabolic activation system; 1-Cyclopentyl-4-nitrosopiperazine; 1-cyclopentyl-4-nitrosopiperazine
• ISSN: 0273-2300; 1096-0295; 1096-0295
• DOI: 10.1016/j.yrtph.2024.105711

The Ames test is a widely used bacterial mutagenicity assay to evaluate the potential of chemical compounds to induce mutations. In recent years, there has been growing concern regarding the presence of N-nitrosamines in pharmaceuticals, food, and other consumer products. N-Nitrosamines are probable mutagens and carcinogens. To address the reduced sensitivity of the standard Ames test for N-nitrosamines, particularly N-nitrosodimethylamine, the European Medicines Agency (EMA) and United States Food and Drug Administration (FDA) have recently published recommendations for enhanced Ames test (EAT) conditions. However, there is a lack of clear guidance on the selection of N-nitrosamine positive control concentrations, particularly for 1-cyclopentyl-4-nitrosopiperazine, and the amount of solvent to be used in the EAT. This study aims to address the current gap in concentration and volume specifications by providing a comprehensive guide to set up enhanced Ames test conditions specifically for N-nitrosamine compounds using appropriate amounts of solvent, new solvents, and strain-specific positive control concentrations. •TA1535 is the most sensitive strain for both NDMA and CPNP.•CPNP ≥50 μg/plate yielded positive results in the enhanced Ames test with hamster liver S9 in base-pair excision mutant strains.•CPNP 5000 μg/plate yielded positive results in the enhanced Ames test with rat liver S9 in base-pair excision mutant strains.•10 μg dose volume is appropriate organic solvent amount for enhanced Ames test.•Water, DMSO, methanol, acetonitrile, DMF, and EGME were found to be compatible solvents in the enhanced Ames test.