| Objective:The aim of this study was to investigate the absorption mechanism of2,3,5,4'-tetrahydroxy diphenylethylene-2-O-?-D-glucoside?TSG?by in vitro and in vivo experiments,to explore the effects of P-gp and MRP2 on the absorption of TSG,and to investigate the interaction between TSG and P-gp,MRP2 by western blot.Methods:The gastrointestinal perfusion model of rats was established to explore the absorption mechanism of TSG.The concentration of TSG in gastrointestinal perfusion was determined by HPLC.Phenol red labeling was used to correct the volume of intestinal circulating fluid.The percentage of absorption?PA?was calculated to investigate the absorption of TSG in gastrointestinal.A Caco-2 cell monolayer model was established,which was administered on the AP side and the BL side,respectively,to simulate the intestinal absorption of TSG.The Papp?AP?BL?and Papp?BL?AP?were calculated to further explore the intestinal absorption of TSG.In addition,the subject also investigated the role of P-gp and MRP2 in the intestinal absorption of TSG,and combined with western blotting to explore the effect of TSG on the expression of P-gp and MRP2.At last,pharmacokinetic experiments were performed to verify the effects of P-gp and MRP2 on TSG.Results:The PA of TSG in the stomach is higher than that in the intestine.The PA of TSG in the stomach of high,medium and low concentrations are?28.65±9.75?%??17.04±3.98?%??13.75±5.94?%,and?3.36±0.40?%??3.36±0.79?%??4.74±1.05?%in intestine,respectively.After adjusting the pH of intestinal perfusion to 6.4 and 5.4,the PA of TSG in the small intestine was?6.91±1.75?%??5.34±1.15?%,which was greater than the control group.Caco-2 cell experiment show that the Papp?AP?BL?of TSG are?5.70±0.35???5.52±1.02???7.28±0.53?×10-6cm·s-1,and Papp?BL?AP?were?4.57±0.37???4.70±0.41???8.03±0.44?×10-6cm·s-1,respectively.The Papp of the bilateral transport of TSG was significantly increased in the environment with a pH of 6.4 and 5.4 compared to the control group with pH of 7.4,which consistent with the results of intestine perfusion experiments.The PA of TSG in the intestine and the Papp?AP?BL?on the Caco-2 cell monolayer were increased significantly after adding verapamil hydrochloride,quinidine and probenecid.Treating with TSG,the expression of P-gp in Caco-2 cells increased and the expression of MRP2 decreased.Pharmacokinetic experiments showed that the Tmaxax was?33.33±9.43?min after administrating alone with TSG.After intravenous injection of verapamil hydrochloride,quinidine and probenecid,the Tmaxax of TSG was shortened obviously,which were?16.00±4.90?,?14.00±4.90?,?13.33±4.71?min,respectively.Moreover,the C?max?and AUC was increased,and the HLLambdaz were prolonged,but no significant difference.Conclusions:TSG is mainly absorbed in the stomach,less absorption in the intestine.The absorption mechanism in the stomach is passive transport,and the absorption increases with increasing concentration.While there are passive transport and P-gp,MRP2-mediated primary active transport mechanisms exist in the intestinal absorption,and TSG is better absorbed in a weakly acidic environment.TSG is affected by P-gp and MRP2 during the transport process,and also counteracts the protein,which induces the expression of P-gp and decreases the expression of MRP2.P-gp and MRP2 not only affect the absorption of TSG,but also also prolong the residence time and reduce the elimination rate of TSG. |
PDF Full Text Request