Impact Of Effect Components In Formula Shenmai On Hepatic Uptake Of Ophiopogonin D Mediated By OATPs/Oatps And Its Compatibility Mechanism

Background:Shenmai Formula(Shenmai injection), derived from Shengmai San, is widely used in the treatment of diseases such as chronic heart failure, coronary atherosclerotic heart disease, viral myocarditis. Ophiopogonin D is one of the main active components of the injection. Previous studies found that ophiopogonin D was mediereted by OATPs/Oatps. The SD rats were respectively given shenmai injection and equivalent dose of ophiopogonin D through hale vene, result showed that the former?s blood concentrations of ophiopogonin D was significantly higher and the ratio of the blood concentration and hepto concentration was bigger, Which given us an idea that there are some mechanisms of interaction and mutual influence based on OATPs/Oatps of heptic between shenmai injection complex components.Consequently, it is necessary to study the influential components toward transpors of heptic of formula shenmai to ophiopogonin D by OATPs/Oatps, and further clarify and reveal the compatible application laws and internal mechanism of Traditional Chinese Medicine based on hepatic transporters, to provide new ideas and research method in the study of Traditional Chinese Medicine Modernization.Objectives:Through rat primary hepatocytes model, stable and high expression of OATP1B1/OATP1B3 of HEK293 T cell model,and animal model in rats in vivo to system study the effective-components of formula shenmai and separated components of shenmai injection toward ophiopogonin D uptake, explore the influence of material foundation of shenmai formula on ophiopogonin D liver transhipment,clarify the interactions and the internal compatibility mechanism of complex components of formula shenmai based on OATPs/Oatps-mediated.Methods:Firstly, establish a LC-MS way to measure ophiopogonin D in cell sample and rat plasma sample.Secondly, study the influence of effective-site and effective-composition of formula shenmai on ophiopogonin D transshipment in rat primary hepatpcytes and OATP1B1-HEK293T/OATP1B3-HEK293 T cells.Thirdly, using HPLC separation method gradually separate shenmai injection to study the effect of each elutes on ophiopogonin D transshipment in OATP1B1-HEK293T/OATP1B3-HEK293 T cells.Finally, SD rats are given ophiopogonin D alone, ophiopogonin D, Rb1 and Rd together by tail vein injection. The pharmacokinetic parameters of ophiopogonin D between two groups were compared and analyzed.Results:The establishment of LC-MS detection method for the OPD in cells and rat plasma with good specificity, high sensitivity, precision and accuracy are in line with relevant regulations.In rat primary hepatocytes and OATP1B1-HEK293T/OATP1B3-HEK293 T cells,we find that main effective position of famula shenmai is panaxadiol-type saponins that has a strong transit inhibition of ophiopogonin D in the three pieces of cell model with IC50 values of 0.94, 1.39 and 0.46?M, respectively. panaxatriol-type saponins has a weak transit inhibition of ophiopogonin D in the three species of cell model with IC50 values of 3.46, 5.37 and 2.44?M, respectively; its main effective component have strong transit inhibition of ophiopogonin D in the three species of cell model by Rb1 with IC50 values of 0.89 1.64 and 0.57?M, respectively, Rd with IC50 values of 0.44, 1.98 and 0.32?M, respectively. The weak transit inhibition of ophiopogonin D in the three pieces of cell model by Re with IC50 values of 4.80,35.89 and 10.71?M respectively, Rg1 with IC50 values of 8.83, 27.22 and 4.18?M,respectively.SMI-2(30-45min) has a weak transport inhibition of ophiopogonin D in the OATP1B1-HEK293 T and OATP1B3-HEK293 T cells with IC50 values of 19.25,3.39?M respectively, SMI-3(45-62min) has a strong transport inhibition of ophiopogonin D in the OATP1B1-HEK293 T and OATP1B3-HEK293 T cells with IC50 values of 1.48 and 0.50?M respectively, SMI-1(0-30min) and SMI-4(62-100min) have no obviously transport inhibition of ophiopogonin D.SMI-3-1(0-10min) and SMI-3-3(15-20min) have strong transport inhibition of ophiopogonin D in the OATP1B1-HEK293 T and OATP1B3-HEK293 T cells,SMI-3-1(0-10min) with IC50 values of 1.94 and 0.55?M respectively,SMI-3-3(15-20min) with IC50 values of 1.66 and 0.31?M respectively,SMI-3-2(11-15min) has no obviously transport inhibition of ophiopogonin D.Compared with contract intravenous injection of the Rb1, Rd and ophiopogonin D and equal doses of ophiopogonin D,the Cmax of former ophiopogonin D is 1.76 times of that for the latter, t1/2 is 2.04 times, the pharmacokinetic parameters of two groups such as AUC, CL, MRT there are significant differences(P < 0.05).Conclusions:Panaxadiol-type saponins could significantly inhibit OPD uptake, while panaxatiol-type saponins with weak inhibition, which infered that panaxadiol-type saponins may be the major effective-site of shenmai formula that effect the hepatic uptake of OPD mediate by organic anion-transporting PolypeptidesGinsenoside Rb1 and ginsenoside Rd could significantly inhibit OPD uptake, while ginsenoside Re and ginsenoside Rg1 with weak inhibition, which infered that ginsenoside Rb1 and ginsenoside Rd may be the major effective-components of shenmai formula that effect the hepatic uptake of Ophiopogonin D mediate by organic anion-transporting Polypeptides.Ginsenoside Rb1 and ginsenoside Rd were the the main material basis of complex components of shenmai formula that effect the hepatic uptake of Ophiopogonin D mediate by organic anion-transporting Polypeptides.Complex components of shenmai formula exists compatibility mechanism that based on hepatic transporters mediated.