Interaction And Transport Of The Main Effective Substances Of Shengmai Formula Mediated By Drug Transporters

Background:Shengmai Formula（SMF）is a famous Chinese herbal prescription in ancient China,which is composed of ginseng,ophiopogon japonicus and schisandra chinensis.There are Shengmai Powder,Shengmai Yin and Shengmai Injection of SMF preparation in clinic practice.It is indicated for deficiency of both Qi and Yin,and has the effect of Supplementing Qi and Nourishing Yin.We previously selected 12 known effective components in SMF and found that ginsenoside Re and ginsenoside Rd were potential inhibitors of OAT1;ginsenoside Re was a potential substrate of NTCP;schizandrol B involved in the BCRP mediated interactions at both cellular and vesicle levels.These studies all suggested that the above transporters mediated the transport process of the main effective components of SMF.Organic cation transporter 2（OCT2）is a kind of uptake transporter with high expression in the kidney,which plays a crucial role in the renal elimination of drugs.However,the interaction and transport of the above components mediated by OCT2 remains unclear,which is worthy of study.In addition,the clinical efficacy of traditional Chinese medicine（TCM）is the result of the combined action of the effective substances groups.Based on the overall concept of TCM,we should pay more attention to the interaction and transport of the TCM compound preparations when multiple components coexist from the perspective of the effective substances groups,comprehensively reveal the compatibility mechanism of complex components of traditional Chinese herbal mediated by hepatic and renal transporters.Objectives:The cell models with high expression of OCT2 were used to explore the interaction and the transport process of the 15 main effective components of SMF which mediated by OCT2.The two main effective substances groups of total ginsenosides and schisandra total lignans were selected to study the transport process of the components in SMF mediated by NTCP,BCRP,OCT2,OAT1 and the excretion,the pharmacokinetic changes of relevant effective components.All of this researches were aimed to interpret the interaction and compatibility mechanism of the main effective substances groups in SMF,which were mediated by hepatic and renal transporters.The conclusion from this studies provide theoretical and experimental basis for clinical safe and rational use of SMF.Methods:1.A stable,sensitive and reliable LC-MS analytical method was established for the detection of the concentrations of 15 main effective components in matrices such as cells,rat plasma,bile and urine of SMF.2.The MDCK-OCT2 and MDCK-MOCK cell models were used to study the interaction and the transport process of the 15 effective components of SMF,which were mediated by OCT2.3.The cell models,overexpressing NTCP,OCT2,OAT1 and BCRP respectively,were used to study the interaction and the transport process of the main effective substances groups of SMF mediated by the above transporters.4.In order to study the interaction and compatibility mechanism of the main effective groups of SMF mediated by hepatic and renal transporters in vivo,the excretion of target components through bile and urine and their pharmacokinetic changes were investigated using an in vivo rat model.Results:1.The quantitative LC-MS method developed in our laboratory for the detection of the main effective components of SMF in different matrices met the relevant requirements for the analysis of biological samples in terms of specificity,precision and accuracy.2.Compared to MDCK-MOCK cells,the uptake of ginsenoside Rb1 and methylophiopogonanone A were significantly higher than those in MDCK-OCT2cells among the 15 main effective components of SMF.Besides,adding the inhibitors of OCT2,the uptake of them were observed significantly decrease in MDCK-OCT2cells.With the OCT2 classical substrate ASP⁺inhibition assay,it was found that ginsenoside Rd,Re and schzandrin B inhibited OCT2 transport activity by 47%,56%and 21%at the concentration of 10μM.In interaction studies,it was found that ginsenoside Rd,Re can significantly inhibite the uptake of ginsenoside Rb1 mediated by OCT2;while ginsenoside Re and schizandrin B had no significantly effect on the uptake methylophiopogonanone A mediated by OCT2.3.In the cell model highly expressing NTCP,BCRP,OCT2 and OAT1 cell lines,with the transport experiment of the main effective substances groups（total ginsenoside and schisandra total lignans）,it was found that NTCP mediated the transport process of ginsenoside Re,OCT2 mediated the transport process of ginsenoside Rb1,Rd and BCRP mediated the transport process of schisandrin A and schisandrin B.Schisandra total lignans inhibited the uptake of ginsenoside Re mediated by NTCP and the uptake of ginsenoside Rb1,Rd mediated by OCT2.4.In rat model,the plasma AUC_0-∞of ginsenoside Rb1,Rd,Re increased significantly,after the combination administration of the main effective substances groups,and the CL declined significantly.The cumulative excretion of schizandrin B in bile within 4 h and the cumulative excretion of ginsenoside Rb1,Rd in urine within8 h decreased significantly.Conclusions:1.Based on the studies of the main effective componenets of SMF mediated by OCT2,it suggest that ginsenoside Rb1 and methylophiopogonanone A are potential substrates for OCT2.The ginsenoside Re,ginsenoside Rd,schisandrin B have a significant inhibitory effect on OCT2-mediated ASP⁺uptake and may be the potential inhibitors of OCT2.There are interaction and compatibility mechanism of the main effective components in SMF mediated by OCT2.2.The studies of the main effective substances groups suggest that NTCP,BCRP and OCT2 mediate the transport process of the ginsenoside Rb1,Rd,schisandrin A and schisandrin B.When total ginsenoside and schisandra total lignans were administered together,the effective components such as ginsenoside Rb1,Rd and schisandrin B take place the transporter-mediated competitive inhibition by affecting their excretion process and pharmacokinetics.The interaction and compatibility mechanism mediated by the hepatic and renal transporter network exist among the main effective substance groups of SMF.