Study On Component Analysis And Effects On Anti-atherosclerosis Of Traditional Chinese Medicine “Sini Decoction”

Sini decoction(SND),as one of the classical Chinese medicine formulas,is known for the efficacy of "warming the middle and dispelling cold,returning yang to save the rebellion".At the same time,it has been widely recognized for its excellent efficacy in anti-atherosclerosis(AS),anti-cardiovascular disease and anti-tumor.The complex chemical composition of SND is the material basis for its manifestation of various pharmacological activities.In this regard,previous studies were generally less rapid and comprehensive,and all concentrated on the part of supernatant while ignoring the precipitates spontaneously generated during the decoction process.The naturally-occurring precipitation have shown similar or even better pharmacological effects compared to the supernatant,the elucidation of the main components and formation mechanisms of naturally-occurring enable to better understand SND and its therapeutic relevance from a holistic level.In addition,the anti-AS efficacy of SND has been confirmed,but the active substances and mechanisms have not been clarified.Therefore,taking composition analysis as the core point,this study aims to comprehensively reveal the chemical composition of both the supernatant and naturally-occurring precipitation in SND,explore the possible formation pathways of precipitation,at the same time,network pharmacology molecular docking and cell experiments were combined to analyze the pharmacological components and key mechanisms of SND for the treatment of AS.This study first reports on a comprehensive analysis of the chemical compositions of both the supernatant and naturally-occurring precipitate in SND using ultra high-performance liquid chromatography hyphenated with quadrupole-Orbitrap high-resolution mass spectrometry(UHPLC-Q-Orbitrap HRMS),a total of 99 components were tentatively identified including 40 flavonoids,34 alkaloids,7 terpenoids and 18 other compounds.Among them,70 compounds were present in both supernatant and precipitate,indicating a high degree of similarity between the two parts,which confirmed the potential pharmacological value of naturally-occurring precipitate.The presence of 10 pairs of acid-base complexes in SND was observed by a cold spray ionization(CSI)device combined with high-resolution mass spectrometry at the condition of low temperature,the results of isothermal titration calorimetry and computational chemistry demonstrated that acid-base compounds can react spontaneously to form macromolecular complexes,the non-covalent interactions such as van der Waals forces and hydrogen bonds may be the main driving force for the complex formation,which provides a scientific basis and mechanism reference for the formation of naturally-occurring precipitation in SND and other traditional Chinese medicines.Based on qualitative experiments,we developed a strategy combining network pharmacology,molecular docking and cellular experiments to reveal the active components and mechanism of SND in the treatment of AS.The results showed that SND against AS is a multicomponent,multi-target and multi-pathway process: fisetin,tetrahydrocurcumin and boldine are considered to be the main pharmacological components;IL6,ALB,AKT1,TNF,VEGFA,MAPK3,MAPK1,and EGFR are the possible critical targets;related pathways especially oxidative stress and inflammation,are key pathways.The results of the cellular experiments have strongly confirmed the significant antioxidant damage effect of SND and three active ingredients on HUVECs cells,which is expected to be further studied and applied.In conclusion,we finally characterized 99 chemical components in supernatant and naturally-occurring precipitation of SND,there are compositional similarities between them,which proves naturally-occurring precipitation could be put into clinical use.;10 possible acidbase complexes in naturally-occurring precipitation were characterized,the main formation mechanisms may be van der Waals forces,hydrogen bonds and other non-covalent interactions;The active components of SND anti-AS may be fisetin,tetrahydrocurcumin and boldine,antioxidative damage is the key mechanism of action,which provides an experimental basis for further research and application.