The antiallergic agent bepotastine besilate is a nonsedating, second-generation H1-antagonist with high oral absorption and negligible distribution into brain. To clarify the role of P-glycoprotein (P-gp) in the pharmacokinetics of bepotastine, intestinal absorption and brain penetration studies were performed. [(14)C]Bepotastine transport in P-gp-overexpressed LLC-PK1 cells indicated that bepotastine was a substrate of P-gp. The affinity of bepotastine to P-gp estimated by ATPase activity assay was low, with a K(m) value of 1.25 mM. After i.v. administration, the brain/plasma free concentration ratio in mdr1-knockout mice was 3 times higher than that in wild-type mice. The in situ intestinal absorption studies of [(14)C]bepotastine in rats showed a clear regional difference, showing highest permeability at the upper part of small intestine with a decreasing permeability in the descending part of small intestine. The apparent absorption rate constant (ka) of [(14)C]bepotastine in the small intestine was greatly increased by cyclosporin A and verapamil, especially in the distal portion, and the site-specific absorption of [(14)C]bepotastine disappeared. The concentration dependence of ka of [(14)C]bepotastine was observed with a higher ka at higher concentration (20 mM) compared with that at lower concentration (1 microM). In conclusion, bepotastine is a substrate for P-gp, and P-gp clearly limited the brain distribution of bepotastine, whereas the effect of P-gp on intestinal absorption of bepotastine was minimal, presumably because of high membrane permeability at the upper region of small intestine where P-gp is less expressed. Such intestinal absorption property of bepotastine is distinctly different from the low membrane-permeable P-gp substrate fexofenadine.