The cytochrome P450 (P450) 3A family is considered to be the most important and abundantly expressed P450 subfamily in mammals. The mRNA expression levels of four P450 3A enzymes in porcine liver and small intestine were investigated using real-time reverse transcriptase-polymerase chain reaction (RT-PCR). The expression of P450 3A mRNAs (P450 3A pool) was higher in the liver than that in the small intestine. In the small intestine, the P450 3A mRNAs were gradually decreased from the duodenum to the ileum. P450 3A29 and P450 3A22 were predominantly expressed both in liver and small intestine tissues with larger ratios in the P450 3A pool than the other P450 3A enzymes. These results demonstrate that P450 3A29 and P450 3A22 probably serve as the major P450 3A contributors for both the hepatic and intestinal P450 3A pool. This work provides a deeper comprehension of the contribution of P450 3A enzymes to xenobiotic metabolism in pigs.