Studies On The Transport Of Taxifolin And Astilbin In Vitro And Pharmacokinetics In Rats

Flavonoids are polyphenolic compounds present in many fruits,vegetables,and beverages.They have been strongly implicated as protectors in coronary heart disease and stroke as well as cancer.Astilbin and its aglycone,taxifolin,were isolated from many Chinese herbs,such as the rhizome of Smilax glabra,a Liliaceae plant, Dimorphandra mollis and Hypericum perforatum,and also have been found in many citrus fruits,especially grapefruit and orange.It has been reported that taxifolin and astilbin have many activities in anti-bacteria,anti-oxidative,anti-inflammation,as well as hepato-protection.Although astilbin and taxifolin provide many benefits to human health,the knowledge about the absorption,transport,metabolism,the interactions with drug-transporters in vivo and in vitro,and other pharmacokinetics profiles are still limited.The aims of this present study were to understand the drug metabolism and pharmacokinetic profiles of astilbin and taxifolin using in vivo and in vitro models.1.The transport of taxifolin and astilbin across the Caco-2 cell monolayers and the effects on the P-gp expressed in Caco-2 cellsThe aim of this study was to explore the potential absorption and transport mechanism of flavanonols astilbin and taxifolin across the Caco-2 cell monolayers,a model of human intestinal epithelial membrane.In the Caco-2 cells,these two flavanonols showed a low absorption.The apparent permeability coefficient(P_app) values of apical to basolateral direction for astilbin and taxifolin were less than 1×10^-6 cm/s.The efflux ratios of both compounds were much higher than 2.0.The efflux of taxifolin can be abolished in the presence of MK-571,the inhibitor of the multidrug resistance proterin 2(MRP2),and the efflux of astilbin can be abolished in the presence of P-glycoprotein(P-gp) inhibitors verapamil and GF 120918.Those findings indicated that astilbin and taxifolin were transported by P-gp and MRP2,respectively.Astilbin and taxifolin may have low bioavailability after oral administration because the two efflux transporters participating in the absorption of the tow flavanonols.The drug-drug interactions should be considered when astilbin or taxifolin is co-administrated with the inhibitors of P-gp or MRP2 in the clinic.Using Rhodamine 123(R123) as the substrate probe of P-gp,we studied the effects of astilbin and taxifolin on the P-gp,the most important transporter in the intestine. Astilbin and taxifolin had no significant inhibition effects on the P-gp-mediated transport of R123 across the Caco-2 cell monolayers.Caco-2 cells exposed to astilbin or taxiolin for 36 h exhibited higher P-gp activity through up-regulating P-gp expression at protein and mRNA levels.These results indicated that astilbin and taxifolin may have effects on the regulation the expression of P-gp.The increased expression of P-gp in Caco-2 cells may serve as an adaptation and defense mechanism in limiting the entry of xenobiotics into the body.2.The metabolism of taxifolin in Caco-2 cellsThe aim of this study was to find out the possible metabolism of taxifolin in human intestine using Caco-2 cells in vitro.After adding the taxifolin into the culture medium and incubating with Caco-2 cells for 48 h,we found four new compounds in the culture medium and Ml was the main metabolite.Using HPLC,MS and NMR,we prepared the metabolites and identified the structure of Mlas 3'-O-methyltaxifolin. 3.The pharmacokinetic studies of taxifolin in ratsIn this study,a sensitive ultra performance liquid chromatography-mass spectrometry (UPLC-MS) method has been developed and validated for the quantification of taxifolin and 3'-O-methyltaxifolin in rat plasma and urine.The present methods were successfully applied to the estimation of the pharmacokinetic parameters of taxifolin following intravenous and oral administration to rats.The absolute bioavailability of taxifolin was 0.17%.And the possible metabolites in rat plasma and urine were detected, especially 3'-O-methyltaxifolin,the main metabolite in rat plasma.Using DAS 2.0 software,it was found that the best fit pharmacokinetic model was single compartment model with weight of 1/C² for oral administration,and three compartments model with weight of 1/C for intravenous administration.The area under the curve(AUC) and C_max were non-proportional to the dose of oral administration ranged from 10 mg/kg to 100 mg/kg.T_max and t_1/2 were depended on the doses of oral administration.Those results indicated the absorption of taxifolin in rat by oral administration was a typical non-linear process.The biotransformation of taxifolin in rat was so fast that the metabolites can be detected at the first sampling time for both oral administration and intravenous administration.The results were helpful for studying the kinetics of taxifolin,and allowed us to make better understanding of the metabolism mechanism of taxifolin in human.4.The pharmacokinetic studies of astilbin in ratsIn this study,a sensitive UPLC-MS method has been developed and validated for the quantification of astilbin in rat plasma and was successfully applied to the estimation of the pharmacokinetic parameters of astilbin following intravenous and oral administration to rats.Using DAS 2.0 software,it was found that the best fit pharmacokinetic model was two compartments model with weight of 1/C² for both oral administration and intravenous administration.The absolute bioavailability of astilbin was 0.066%indicating that the oral absorption of astilbin in vivo may be poor.The AUC of astilbin in the rats with co-administration with verapamil was increased 2.57 times indicating that the P-gp plays an important role in the pharmacokinetics of astilbin in rats.