The aim of this study was to examine the role of PEPT2, a proton-coupled oligopeptide transporter of the SLC15 family, on the disposition of the antibiotic cefadroxil in the body, particularly the kidney and brain. Pharmacokinetic, tissue distribution, and renal clearance studies were performed in wild-type and PEPT2 null mice after intravenous bolus administration of [(3)H]cefadroxil at 1, 12.5, 50, and 100 nmol/g body weight. Studies were also performed in the absence and presence of probenecid and quinine. Cefadroxil disposition kinetics was clearly nonlinear over the dose range studied (1-100 nmol/g), which was attributed to both saturable renal tubular secretion and reabsorption of the antibiotic. After an intravenous bolus dose of 1 nmol/g cefadroxil, PEPT2 null mice exhibited a 3-fold greater total clearance and 3-fold lower systemic concentrations of drug compared with wild-type animals. Renal clearance studies further demonstrated that the renal reabsorption of cefadroxil was almost completely abolished in PEPT2 null versus wild-type mice (3% versus 70%, p < 0.001). Of the 70% of cefadroxil reabsorbed in wild-type mice, PEPT2 accounted for 95% and PEPT1 accounted for 5% of reabsorbed substrate. Tissue distribution studies indicated that PEPT2 had a dramatic effect on cefadroxil tissue exposure, especially in brain where the cerebrospinal fluid (CSF)-to-blood concentration ratio of cefadroxil was 6-fold greater in PEPT2 null mice compared with wild-type animals. These findings demonstrate that renal PEPT2 is almost entirely responsible for the reabsorption of cefadroxil in kidney and that choroid plexus PEPT2 limits the exposure of cefadroxil (and perhaps other aminocephalosporins) in CSF.