Ritonavir dramatically increases the bioavailability of a variety of concurrently administered drugs by inhibition of metabolic enzymes and drug transporters. The purpose of this study was to investigate the extent to which ritonavir's inhibition of drug transporters and/or CYP3A contributes to the increased oral bioavailability in mice. The area under the plasma concentration-time curves (AUC) for orally administered saquinavir after coadministration with 50 mg/kg ritonavir dramatically increased (325-fold). As a result, the bioavailability, Fa·Fg and Fh increased 75-, 38- and twofold, respectively. In addition, the increase in the AUC predicted from the in vitro Ki value was ninefold, which was derived from the inhibition of metabolic enzymes by ritonavir in the liver. The remaining 36-fold increase in the AUC was considered to be derived from the inhibition in the small intestine. The AUCinf for probe substrate midazolam, fexofenadine, and pravastatin increased after the oral administration of ritonavir by only five-, 13-, and sevenfold, respectively. Moreover, the AUC0-12 for saquinavir was affected negligibly by itraconazole. These results indicate ritonavir mainly affects the first-pass effect of saquinavir in the small intestine, increasing the bioavailability of orally administered saquinavir. Furthermore, cyp isoforms other than CYP3A, which contribute to the metabolism of saquinavir in humans, are involved in the metabolism of saquinavir in mice.