Study On Pharmacodynamic Substance Foundation And Quality Control Of Hugan Qingzhi Granules

Nonalcoholic fatty liver disease(NAFLD)is a pathologic syndrome characterized by diffuse hepatocyte bullous adipose lesions,in the absence of any secondary causes of hepatic accumulation,such as alcohol consumption and other factors directed to liver injury.Hugan Qingzhi granules(HGQZ)is a traditional Chinese medicine formulated with Alismatis Rhizoma,Crataegi Fructus,Nelumbinis Folium,Typhae Pollen and Notoginseng Radix.HGQZ is effective for the treatment of hyperlipidemia and liver disorders.In recent years,many studies has focused on its mechanisms and pathways on NAFLD.However,the clinical application of HGQZ is limited by its insufficient information of chemical profiles,unkown pharmacodynamic substance foundation as well as poor quality control.In this study,the efficacy and chemical analysis were used to study the pharmacodynamic substance foundation of HGQZ.Firstly,a free fatty acid-induced LO2 cell model was established and the effect of two fraction on hepatic steatotic LO2 cell were compared.The results showed that ethanol extract is the active fraction of HGQZ and its lipid-lowering effect is superior than that of water extract.Meanwhile,the chemical components in two fraction was analyzed by ultra-high performance liquid chromatography coupled with triple quadrupole mass spectrometry(UHPLC-QqQ-MS/MS).The results demonstrated that the contents of components in ethanol extract were higher than water extract.This result was in accord with its bioactivity.Next,the active compounds in the King(Jun)—Alismatis Rhizoma was studied for the first time.Four alisols was detected in the process of preparation of HGQZ.It was found that the alisols are highly sensitive to changes in temperature and pH,and transform during heating.Alisol B 23-acetate can be transformed to alisol A and alisol A 24-acetate,and alisol B can be transformed to alisol A.Then,repetitive orthogonal test was used to optimize the ethanol concentration,solvent amount,extraction time and extraction times of extraction process.It was found that the optimized extraction condition is as follow:herbal medicine were refluxed three times with six times the volume of 70%ethanol for 1 h each time.Subsequently,the extract was condensed and dried as well as pelleted to obtain the granules.The chemical analysis showed that the transfer rate of the main components was higher than 65%,therefore the quality of the products was guaranteed.Finally,Thin Layer Chromatography(TLC)and High Performance Liquid Chromatography(HPLC)were adopted for the qualitative and quantification analysis,respectively.The results showed that the TLC spots of Alismatis Rhizoma,Crataegi Fructus,Nelumbinis Folium,Typhae Pollen and Notoginseng Radix were clear and well seperated without the interference of negative control.HPLC with variable wavelength method was proved to be well seperated and with good precision,repeatability,stability and accuracy.The results demonstrated that the contents determination of typhaneoside,hyperoside,nuciferine and alisol B 23-acetate were 1.1384±0.0021 mg/g,1.0894±0.0171 mg/g,0.1508±0.0022 mg/g and 0.6560±0.0151 mg/g.Our establish method can provide scientific base for the quality control of Hugan Qingzhi grancules.