It is well known that cattle ingesting aflatoxin B1contaminated feed commodities excrete aflatoxin M1into their milk. As aflatoxin M1originates from hepatic metabolism, measures to prevent aflatoxin M1formation need to be directed to either the immobilization of aflatoxin B1in the gastrointestinal tract or the modification of hepatic metabolism of aflatoxin B1. Here we studied the influence of oltipraz and a second dithiolthione, (1,2) dithiolo (4,3‐c)‐1,2‐dithiole‐3,6 dithione (DDD) on bovine hepatic aflatoxin B1biotransformation. Oltipraz inhibited aflatoxin B1metabolism as no aflatoxin M1and no aflatoxin B1‐dihydrodiol, the second metabolite found in bovine hepatocytes, was formed. DDD did not significantly inhibit aflatoxin B1metabolism. It could be demonstrated that the inhibition of aflatoxin B1metabolism was due to the inhibition of several cytochrome P450 enzyme activities by oltipraz. In contrast, DDD inhibited only ethoxyresorufinO‐deethylation activity. These findings suggest a high efficacy of oltipraz in inhibiting aflatoxin M1contamination of milk from dairy cows exposed to aflatoxin B1contaminated feeds.