1. A method for the isolation and cultivation of porcine hepatocytes and porcine duodenal enterocytes for the investigation of drug oxidation reactions has been established. 2. Hepatocytes as well as enterocytes metabolized ethoxyresorufin (EROD) and ethoxycoumarin (ECOD) effectively, the rate being 31+/-17 pmol/h x dish (EROD) and 9530+/-4062 pmol/h x dish (ECOD) in the case of hepatocytes, and 9+/-4 pmol/h x dish (EROD) and 510+/-467 pmol/h x dish (ECOD) in the case of enterocytes. Diazepam, another CYP monooxygenase substrate, was also metabolized by porcine hepatocytes but not with porcine enterocytes, thus indicating differences in the metabolic competence of the liver and the gut. 3. The ability to induce enzymes responsible for the metabolism of ethoxyresorufin and ethoxycoumarin was investigated in vitro on treatment of the cell cultures with either 50 microM 3-methylcholanthrene (3-MC) or 50 microM beta-naphthoflavone (beta-NF). With enterocyte cultures, ECOD activity was inducible up to 20-fold, whereas EROD remained unchanged following treatment with either 3-MC or beta-NF. 4. Western blotting provided additional evidence for the expression of CYP1A1 and CYP3A4 at the protein level and treatment of cultured enterocytes with 30 microM Aroclor 1254 or 50 microM beta-NF resulted in enhanced expression of the CYP1A protein, and CYP3A4 protein expression was induced following treatment with 50 microM DEX, 2 mM PB, 30 microM Aroclor 1254 or 50 microM beta-NF. 5. The metabolism of diazepam was also investigated with baculovirus-expressed human CYP enzymes (2C8, 2C9-ARG, 2C9-CYS, 2C19, 3A4, 3A4+cytochrome b5 and 3A5) and evidence was obtained to suggest the formation of temazepam and oxazepam by enzymes of the CYP3A subfamily. Small amounts (32+/-12 ng/ml) of desmethyldiazepam were additionally recovered in microsomal preparations of all CYP-transfected cell lines. 6. In conclusion, porcine duodenal enterocytes can successfully be cultured for a short period and may be used as a tool for studying intestinal metabolism, whereas porcine hepatocytes can be cultured for prolonged periods (>10 days) reliably to investigate hepatic drug oxidation reactions.