Pyrazinamide-derived 1,2,3-triazoles: Antimicrobial evaluation, selective antimycobacterial activity and mechanism of action insights

A series of pyrazinamide-derived 1,2,3-triazoles featuring systematic chlorination of the pyrazine ring and diverse aryl substituents was synthesized and evaluated for antimycobacterial activity. Biological activity screening revealed broad-spectrum antimycobacterial activity and good selectivity toward mycobacteria over other pathogens, with 11 of the prepared compounds showing activity against Mycobacterium tuberculosis (Mtb) H37Ra and/or Mtb H37Rv (MIC ≤ 62.5 μg/mL). Structure–activity relationship analysis showed that 5-Cl substitution on the pyrazine ring was associated with improved antimycobacterial activity, with the best MIC values observed for compound 7 against Mtb H37Ra (MIC = 1.98 μg/mL) and compound 37 against Mtb H37Rv (MIC = 1.56 μg/mL). The tested compounds retained activity against drug-resistant Mtb isolates and partially against naturally resistant Mycobacterium abscessus , while showing low in vitro cytotoxicity in the HepG2 cell line and favorable selectivity indices. During advanced cytotoxicity testing, both compounds 7 and 37 displayed substantially lower hemolytic activity than bedaquiline, indicating a favorable erythrocyte safety profile within the tested concentration ranges. In vivo toxicity testing on Galleria mellonella showed low acute toxicity for both compounds. Mechanistic studies on compounds 7 , 27 , 31 , and 37 revealed reduced biosynthesis of fatty acids and derived lipids, a phenotype consistent with interference with the Fatty Acid Synthase I (FAS I) system.