Abstract: Unsubstituted, halogenated and/or alkylated pyrazine derivatives connected via -CONH- bridge with substituted anilines were synthesized and tested against Mycobacterium tuberculosis strain H37Rv. Condensation of chlorides of substituted pyrazine-2-carboxylic acids with ring-substituted amines yielded a series of amides of pyrazine-2-carboxylic acid, 6-chloropyrazine-2-carboxylic acid, 5-tert-butylpyrazine-2-carboxylic acid or 5-tert-butyl-6-chloropyrazine-2-carboxylic acid, respectively. The synthetic approach, analytical, spectroscopic, lipophilicity and biological data of twelve newly synthesized compounds are presented. Structure-activity relationships among the chemical structure, the antimycobacterial, antifungal, photosynthesis-inhibiting and antialgal activity of the evaluated compounds are discussed. Pyrazine-2-carboxylic acid (2-trifluormethyphenyl)amide (2) has shown the highest activity against Mycobacterium tuberculosis H37Rv (99% inhibition). The highest antifungal effect against Trichophyton mentagrophytes, the most susceptible fungal strain tested, was found for 5-tert-butylpyrazine-2-carboxylic acid (3-trifluormethyphenyl)amide (8, MIC = 62.5 µmol mL-1). The highest reduction of chlorophyll content in Chlorella vulgaris was found for 6-chloropyrazine-2-carboxylic acid (3-trifluormethyphenyl)amide (5, IC50 = 12.1 µmol L-1).