Study On Substance Basis And Quality Control Of Kai-Xin-San Based On UPLC-Q-Orbitrap MS And GC-MS

In ancient books,the prescription named Kai-Xin-San(KXS)was first recorded in Beiji Qianjin Yaofang written by Sun Simiao in the Tang Dynasty.It consists of four herbs:Ginseng Radix Rhizoma,Polygalae Radix,Poria and Acori Tatarinowii Rhizoma.KXS used in this experiment is a powder made of Ginseng Radix Rhizoma,Polygalae Radix,Poria and Acori Tatarinowii Rhizoma in a ratio of 1:1:2:1 after optimization by our research group.KXS has a good clinical effect on depression,alzheimer’s disease and vascular dementia.The development of traditional Chinese medicine(TCM)in the world is greatly limited due to its complex chemical composition,large quality difference and unclear pharmacodynamic material basis.Therefore,it is of great importance to clarify the chemical composition and effective substance basis of TCM and establish reasonable quality control methods for the development of TCM and the internationalization of TCM.Wang xijun is put forward for the first time in the early 1990 s and has established the Chinese medicine serum drug chemical theory and research methods,he thinks that TCM after oral its effective substances must take blood as the medium to the target can play a role,the transitional composition in the blood is the direct effect substance of TCM,therefore,the blood migration components of KXS were detected in this study.The site of action of KXS is the brain,and drugs need to reach target organs through blood circulation to play a role.Many drugs cannot reach the brain due to the existence of blood-brain barrier.In this study,the components entering the brain were further detected to further clarify the pharmacodynamic substance basis of KXS in the treatment of neurodegenerative diseases.In this study,the analytical method of the chemical composition of KXS was established and comprehensively identified.The types of compounds in KXS are very complex,Ginseng Radix Rhizoma mainly contains ginsenoside.Tenuigenin,polyxanthone,and polygala sugar ester are mainly found in Polygalae Radix.Triterpenoid acid is the main component of Poria,and volatile oil is the main component of Acori Tatarinowii Rhizoma,the main components of this volatile oil are mainly phenylpropanoids,monoterpenoids and sesquiterpenoids.Therefore,GC-MS and UPLC-Q-Orbitrap MS chemical composition identification methods were established in this study,and the chemical composition of KXS was comprehensively characterized.A total of 211 compounds were identified by UPLC-Q-Orbitrap MS,among which 60 compounds were identified from Ginseng Radix Rhizoma,40 compounds were identified from Poria and 111 compounds were identified from Polygalae Radix.105 compounds were identified from KXS by GC-MS,mainly from Acori Tatarinowii Rhizoma.On the basis of the established qualitative analysis method of KXS,this project further established the analysis method of KXS into blood by optimizing the pretreatment method of plasma sample,mass spectrometry data collection mode and data processing process.UPLC-Q-Orbitrap MS was used for comparative analysis of blank plasma,drug-containing plasma and KXS extract.A total of 40 compounds and 112 metabolites were identified from drug-containing plasma.A total of 18 compounds were identified by HS-SPME-GC-MS for the comparative analysis of blank plasma,drug-containing plasma and KXS extract.After oral administration,effective substances of TCM must be transported to the target in the blood to play a role,and the transitional components in the blood are the direct substances of TCM.In clinical treatment of Alzheimer’s disease and depression,KXS acts on brain tissue.Due to the existence of blood-brain barrier,it is difficult for drugs to enter brain tissue through the blood-brain barrier to achieve therapeutic effect.Therefore,it is of great importance to determine the components entering brain and clarify the pharmacodynamic substance basis of KXS.On the basis of the established method for analyzing the components of KXS in blood,this study further optimized the sample pretreatment method of drug-containing brain tissue and established the method for analyzing the components of KXS in brain.UPLC-Q-Orbitrap MS was used for comparative analysis of blank brain tissue homogenate,drug-containing brain tissue homogenate and KXS extract.A total of 6compounds were identified from drug-containing brain tissue.The homogenates of blank brain tissue,drug-containing brain tissue and KXS extract were compared and analyzed by HS-SPME-GC-MS.Eighteen compounds were identified from drug-containing brain tissue.In this study,16 components were selected according to the research results of blood components and the requirements of quality control of single drug in KXS in The Chinese Pharmacopoeia in 2020,including sibiricose A5,sibiricose A6,sibiricaxanthone B,polygalaxanthone Ⅲ,3,6′-disinapoyl sucrose,tenuifoliside A,ginsenoside Re,ginsenoside Rf,ginsenoside Rg1,tenuifolin,ginsenoside Rb1,onjisaponin B,ginsenoside Rd,polyporenic acid C,poricoic acid A,pachymic acid were used as indicator components to establish a quantitative method for quality control of KXS.A adjusted parallel response monitoring model was established to determine the multi-component content of 16 blood prototype components in KXS,so as to establish a fast,accurate and comprehensive quality control method of KXS.In this project,the chemical components,blood and brain components of KXS in vitro were analyzed and identified based on UPl C-Q-Orbitrap MS and GC-MS,multiple data acquisition modes and Compound Discover 3.1 metabolic platform were used to determine the content of multiple index components in KXS.A fast and comprehensive quality control method for KXS was also provided.This study provides theoretical basis for the clinical application,pharmacological research and preparation production of KXS.