Ceftibuten undergoes H+-coupled uphill transport across rat small intestinal brush-border membrane vesicles. The effects of amino acids, peptides, folate, and beta-lactams on the uptake of ceftibuten were examined. Uptake of ceftibuten was competitively inhibited by dipeptides or tripeptides. A counter-transport effect on ceftibuten uptake was observed in the vesicle preloaded with these peptides, and the transport was temporarily against a concentration gradient (overshooting). On the other hand, ceftibuten uptake was not changed by amino acids and a tetrapeptide. Therefore, ceftibuten is predominantly transported via the oligopeptide transport system in the brush-border membranes. The relationship of ceftibuten transport to folate and other oral antibiotics was also investigated. Cyclacillin, cephradine, and cefadroxil exhibited both inhibitory and countertransport effects, but folate, cefaclor, and cephalexin showed only a slight inhibitory effect. As the transport of cefaclor showed no uphill uptake in the presence of a H+ gradient and its H+ stimulated uptake was small, a H+ gradient-independent carrier-mediated system seems to participate in its transport. These findings suggest that two different carrier-mediated transport systems, H+ gradient dependent and independent, may exist for oral cephems.