Two human hepatoma cell lines, KYN-2 and Mz-Hep-1 were characterized in terms of glucuronidation capacity and inducibility of cytochrome P4501A1/1A2 and several UDP-glucuronosyltransferases (UGTs). Cytochrome P4501A1/1A2 activity was measured using 7-ethoxyresorufin and that of UGTs with 16 different substrates. The effects of dimethyl sulfoxide (DMSO), 13-naphthoflavone, ~-naphthoflavone, and rifampicin on these drug-metabolizing enzyme activities were studied. DMSO treatment increased in a dose-dependent manner the ethoxyresorufin O-deethylase (EROD) activity in KYN-2 cells, while an opposite effect was observed in Mz-Hep-1 cells. In KYN-2 cells, EROD was more responsive toward 13-naphthoflavone treatment in combination with DMSO. This activity was enhanced in Mz-Hep-1 cells more than 83 times by 13-naphthoflavone. The enhancement of EROD activity by DMSO and 13-naphthoflavone treatments of KYN-2 cells was abolished by ~-naphthoflavone treatment. In Mz-Hep-1, only the inducing effect of ~naphthoflavone was abolished by ct-naphthoflavone treatment. Rifampicin treatment of KYN-2 cells reversed both the DMSO and 13-naphthoflavone effects on the EROD activity. Glucuronidation of steroids, bile acids, fatty acids and drugs was effective in KYN-2 and Mz-Hep-1 cells. Both 1-naphthol glucuronidation and the level of UGTI*6 protein detected by immunoblot and supporting this activity were lowered by DMSO treatment and increased by 13-naphthoflavone treatment in KYN-2 cells. In Mz-Hep-1 cells, DMSO and 13-naphthoflavone had no effect on 1-naphthol glucuronidation activity. DMSO, ~-naphthoflavone and rifampicin also affected the glucuronidation of various substrates supported by different UGT isoforms. These results indicate that KYN-2 and Mz-Hep-1 cells can be used as new in vitro models for the studies of drug metabolism and the regulation of the corresponding enzymes.