Study Onabsorption And Metabolism Of Quercetin And Its Glycosides In Caco-2 Cells

A model for the simulation of human intestine epithelia was established from Caco-2 cells based on the procedures reported in literature. The absorption and metabolism of quercetin and its glycosides were studied on this model, in order to provide information on absorption and metabolism of quercetin and its glycosides in vivo and related mechanisms.Six parts were included in this study. The major results were summarized as followings.1. The model of Caco-2 cells was established and culture conditions were optimized. The specific inhibitors for P-gp and MRP2 were applied to validate the model. The results showed that the model was reliable.2. The adequate optimal incubation conditions and LC/MS technique were developed. The incubation time should be less than 2.5h and the concentration of tested chemicals lower than 100ug/ml. The quercetin, quercetin glycosides and methylated metabolites were analyzed using the LC/MS procedure developed in this experiment. The results indicated this LC/MS procedure was sensitive, linear in certain range of concentration.3. The absorption of quercetin and quercetin glycosides was investigated in Caco-2 cell monolayer. The quercetin was transported across the monolayer bilaterally, in concentration and time dependent manners. The transport from basolateral to apical was more than from apical to basolateral. The Papp(A→B) and Papp(B→A) on the loading side were 24 and 8 times higher than the receiver side, respectively, indicating that quercetin underwent extensive transformation after absorption. Quercitrin and isoquercitrin were also transported across the monolayer bilaterally, in concentration and time dependent manners. Possible metabolism also occurred to them during the transportation across the Caco-2 cell monolayer.4. The roles of SGLT1 and GLUT2 in the absorption of quercetin and quercetin glycosides were investigated on Caco-2 cell monolayer. The glucose inhibited the transportation of quercetin across the monolayer, which was in a manner independent of the concentration of glucose. It was, therefore, suggested that quercetin and glucose competed for the same transporters on the monolayer during the absorption processes. Phloridzin and phloretin exhibited an effect similar to glucose on quercetin absorption, indicating that both SGLT1 and GLUT2 were involved in the processes of quercetin absorption. Phloridzin and phloretin acted differently on the absorption of quercitrin and isoquercitrin, which implied that the difference in sugar moiety affected the process of absorption on Caco-2 cell monolayer.5. The roles of P-gp and MRP2 in the absorption of quercetin and quercetin glycosides were also investigated on Caco-2 cell monolayer. The addition of specific inhibitors for P-gp and MRP2 decreased the transportation of quercetin across Caco-2 cell monolayer. However, no effect was observed for quercitrin and isoquercitrin. It was demonstrated that quercetin was the substrate for P-gp and MRP2, whereas quercitrin and isoquercitrin were not.6. The metabolism of quercetin and quercetin glycosides were analyzed on Caco-2 cell monolayer. Two methylated metabolites, isorhamnetin and tamarixetin were detected in the loading and receiver sides, which accounted for 10% of quercetin loaded, suggesting the existence of other metabolic pathways for quercetin. More metabolites were further identified, such as quercetin mono-sulphate, quercetin tri-sulphate, quercetin glucuronide, quercetin glucuronide sulphate, methylquercetin sulphate and methylquercetin glucuronide.In conclusion, quercetin and quercetin glycosides were transported intact across the Caco-2 cell monolayer and experienced an extensive metabolism. The quercetin glycosides appeared relatively slower than quercetin in the process of absorption and metabolism. The methylated, sulphated and glucuronidated metabolites were identified, respectively. The SGLT1 participated in the process of absorption of quercetin and quercetin glycosides, while the GLUT2 was involved in the process of absorption of quercitrin. Quercetin was the substrate for P-gp and MRP2. However, the absorption of quercitrin and isoquercitrin was not affected by P-gp and MRP2.