P-gp by ginsenoside Rh2 epimers in vivo, the pharmacokinetics of Rh2 epimers in vivo, the possible metabolism, and evaluation of P-gp regulatory effects in vitro were all included. Moreover, the differential P-gp regulations of Rh2 epimers were further confirmed by applying Rh2 epimers as P-gp regulators in reversal of P-gp mediated multi-drug resistance. Our study provides a new case describing the chiral characteristics of P-gp. It is also a meaningful trial to elucidate the stereoselective P-gp regulation mechanisms of ginsenoside Rh2 epimers in vivo from a pharmacokinetic view. at 5 mg/kg prior to i.g. administration of digoxin, the AUC and Cmax of digoxin were significantly enhanced. But, when the dosage of 20-Rh2 was elevated to 50 mg/kg, the absorption of digoxin was not changed significantly compared with control group. The dose-effect trends of 20-Rh2 and 20Rh2 on the oral pharmacokinetics of digoxin were just opposite. ” Stereoselective LC-MS quantification of ginsenoside Rh2 epimers and the deglycosylation metabolites ginsenoside Ppd epimers The chromatograms shown in Fig. 3 demonstrated that the present LC-MS conditions applied for analysis of Rh2 and Ppd epimers provided appropriate separation with the retention time of 6.9, 7.9, 14.2, 14.7 and 6.7 min for 20-Rh2, 20-Rh2, 20Ppd, 20-Ppd and digitoxin respectively. The specificity of the method was evaluated by screening blank biological matrix in selected ion monitoring mode, and no interference had been observed. The method showed good linearity in a range of 1 1000 nM with a correlation coefficient R2 exceeding 0.995 for the analytes. Stereoselective oral pharmacokinetics of ginsenoside Rh2 epimers in rats As seen in Fig. 4, there was significant difference in oral pharmacokinetics of ginsenoside Rh2 epimers in rats. With the same dosage for oral administration, the Cmax and AUC of 20Rh2 were 15-fold and 10-fold higher than those of 20-Rh2 respectively: the Cmax of 20-Rh2 was nearly 1000 nM while the Cmax of 20-Rh2 was no higher than 50 nM, which suggested better oral absorption of 20-Rh2 than 20-Rh2. Furthermore, chiral inversions between ginsenoside Rh2 epimers were observed. When 20-Rh2 was orally administered, 20Rh2 was also detected in plasma, with Cmax only one eighth of 20-Rh2 and AUC only one tenth of 20-Rh2. Similarly, when 20-Rh2 was orally administered, 20-Rh2 was also detected in plasma, and the concentrations of 20-Rh2 were much lower than those of 20-Rh2. Otherwise, the deglycosylation metabolite of 20-Rh2 was also monitored in plasma when 20-Rh2 was orally administered, and the configuration of Ppd was confirmed by the standard substance of 20-Ppd. But, no Ppd was found in plasma after oral administration of 20-Rh2. Results Effects of 20-Rh2 and 20-Rh2 on oral pharmacokinetics of digoxin in rats Digoxin has been proved as a classic P-gp substrate, and its intestinal absorption is mainly restricted by P-gp. When 20-Rh2 was i.g. administered to rats prior to i.g. administration of digoxin, the oral absorption of digoxin was enhanced with increasing concentrations of 20-Rh2. The AUC and Cmax of digoxin were elevated by 1.8-fold and 1.6-fold respectively by 50 mg/kg 20-Rh2. The AUCs were calculated and listed in Effects of 20-Rh2, 20-Rh2, 20-Ppd and ” 20-Ppd on P-gp functions in AZ-6102 site Caco-2 cells Caco-2 cell model is a classic approach in the research of P-gp. As shown in Fig. 6A, 20-Rh2 decreased the efflux ratio of digoxin crossing Caco-2 cell monolayers in a concentr