The complementary DNAs of rat glutathione S-transferase (GST, EC 2.5.1.18) Ycl and of mouse Yc were expressed from a prokaryotic expression vector in E. coli. The purified proteins were analyzed for their activity toward aflatoxin B 1-8,9-epoxide (AFBO), the reactive intermediate of the fungal mycotoxin aflatoxin B 1 (AFB). The mouse Yc isozyme had about 50-fold higher conjugating activity toward AFBO than the rat Yc1 isozyme (144 nmol/mg/min versus 3.3 nmol/mg/min). The rat Yc1 isozyme had specific activities toward 1-chloro-2,4-dinitrobenzene, cumene hydroperoxide and ethacrynic acid of 10.7, 0.98 and 0.92 μmol/mg/min, respectively, whereas the mouse Yc isozyme had specific activities of 5.7, 2.1 and 0.1 μmol/mg/min for these substrates, respectively. These data provide further support for the hypothesis that the constitutive presence of the alpha class GST Yc isozyme in mouse liver protects mice from the hepatoearcinogenic effects of aflatoxin B 1.