The purpose of this study was to determine the interactions of erythromycin and various fluoroquinolones with P-glycoprotein (P-gp) and in turn assess their effects on transport kinetics across a model cell monolayer. MDCKII-MDRI cells were selected as a model monolayer to evaluate the effects of various fluoroquinolones, ie, norfloxacin, lomefloxacin, ofloxacin, enoxacin, grepafloxacin, levofloxacin, and sparfloxacin on the P-gp-mediated efflux of H-cyclosporine (CsA) and C-erythromycin. IC50 values associated with grepafloxacin-, levofloxacin-, and sparfloxacin-mediated inhibition of P-gp were calculated across Caco-2 cells with erythromycin as the model P-gp substrate. Transport of erythromycin was then studied with P-gp saturable concentrations of fluoroquinolones. Western blot analysis was performed on Caco-2 cells to confirm P-gp expression. Only grepafloxacin elevated the uptake of H-CsA across the MDCKII-MDRI cell monolayer, whereas norfloxacin, lomefloxacin, ofloxacin, and enoxacin did not exert any effect on H-CsA uptake. Inhibition studies indicate that grepafloxacin, levofloxacin, and sparfloxacin are potent inhibitors of P-gp-mediated efflux of C-erythromycin in the MDCKII-MDRI cell monolayer. Similar studies were conducted across Caco-2 cells and IC50 values were calculated. Inhibitory potency of sparfloxacin (IC50 = 607.6 microM) exceeded that of levofloxacin (IC50 = 1644 microM) and grepafloxacin (IC50 = 2266 microM). Permeability ratio (BL-AP/AP-BL) of C-erythromycin was found to be 8.67, which was reduced to 1.18, 1.83, and 1.39 in the presence of grepafloxacin (1 mmol/L), levofloxacin (5 mmol/L), and sparfloxacin (1 mmol/L), respectively. Log partition coefficient of grepafloxacin (1.58), levofloxacin (0.553), and sparfloxacin (0.45) were correlated with the inhibition of P-gp. Western blot analysis indicated the expression of P-gp in Caco-2 cells. Fluoroquinolones like grepafloxacin, levofloxacin, and especially sparfloxacin significantly inhibit the efflux of erythromycin, which can modulate oral absorption and disposition of macrolide drugs when administered concomitantly.