Carbonic anhydrase (CA) from erythrocytes of rainbow trout Oncorhynchus mykiss was purified by affinity chromatography through a process comprising hemolysate preparation, sepharose-4B-L-tyrosine-sulfanylamide affinity get chromatography, and dialysis. Then the effects of three antibiotics-sodium ampicillin, streptomycin, and trimethoprim plus sulfamethoxazole-on this enzyme were investigated in vitro and in vivo. Inhibition effects were determined in vitro by means of the Lineweaver-Burk and activity-antibiotic regression graphs. The dissociation constant of the enzyme inhibitor complex (K-i) and 50% inhibitory values were 2.16 +/- 0.40 and 1.87 mM for sodium ampicillin, 7.40 +/- 0.310 and 5.21 mM for streptomycin, and 18.50 +/- 0.35 and 9.74 mM for trimethoprim-sulfamethoxazole. The in vivo experiment demonstrated that CA was significantly inhibited by trimethoprim-sulfamethoxazole but not by sodium ampicillin and streptomycin.