Minipigs have become a popular alternative to the traditional non-rodent species although little information is available on their P450 system. The total P450, the enzyme activity and immunochemical levels of some of the most important drug metabolizing isoenzymes CYP1A2, CYP2A6, CYP2C19, CYP2D6, CYP2E1 and CYP3A4 were measured in liver microsomes from 8 minipigs and 12 conventional pigs of both sexes and castrate conventional pigs. The mRNA expression was analyzed for 3 isoenzymes: CYP1A2, CYP2A6 and CYP2E1. The total P450 activity was slightly higher in minipigs compared to conventional pigs but no sex differences were detected. CYP1A2 activity was 4 times higher in female than in male minipigs. The activity of the male minipigs possessed the same activity as were identical to the conventional females, males and castrates. The activity of CYP2E1 was 4 times higher in female than in male minipigs and 2 times higher in female than in male pigs. No activity of CYP2D6 or CYP2C19 could be detected. The CYP3A4 activity detected in minipigs was higher than the activity in conventional pigs. A slight sex difference was seen in both strains. Correlations between enzyme activity and immunochemical levels were found for CYP1A2, CYP2A6 and CYP3A4 but not for CYP2E1. The mRNA concentration of CYP1A2, CYP2A6 and CYP2E1 was determined because the activity of these enzymes showed marked sex differences. A Spearman ranking correlation analysis between mRNA expression and enzyme activity showed a weak correlation for CYP2A6, but not for CYP1A2 and CYP2E1. These results seem to indicate that CYP2A6 could be transcriptionally regulated, whereas CYP1A2 might be post-transcriptionally regulated.