In an attempt to rationalize the search for new potential antimicrobial agents, a novel series of 1,4-disubstituted 1,2,3-triazole-imidazo[1,2-a]pyrazine conjugates have been synthesized via copper-catalyzed azide-alkyne cycloaddition. Eight novel compounds were characterized by( 1)H and C-13 NMR, IR, and mass spectra and elemental analyses. Their mesomorphic behavior was studied by polarizing optical microscopy (POM). All compounds are enantiotropically nematogenic without any smectogenic character, and phase transition temperatures and the textures were determined. The nematic textures are threaded or schlieren. The newly synthesized compounds have been assessed for their antimicrobial potential using eight microbial strains, including gram-positive and gram-negative bacteria and fungi (E. coli, P. aeruginosa, E. aerogenes, B. megaterium, S. aureus, B. subtilis, A. niger, and A. flavus). The compounds displayed superior activities to the standard drugs against various microbial strains.