Protective Effect Of Panax Quinquefolium Saponins On Myocardial Cell Injury

Objective:Collect 16 batches of Panax quinquefolium from different origins and different ages to establish fingerprints,and determine the content of saponins in them,to compare the differences in the saponins in them.The total saponin components in American ginseng were extracted,separated,and analyzed qualitatively and quantitatively using traditional Chinese medicine chemical technology.Through the application of biological cell technology,the protective effect of total saponins of American ginseng on myocardial cell injury was studied,providing experimental evidence for the clinical mechanism of American ginseng in protecting the heart and product development.Methods:1.16 batches of different specifications of American ginseng chromatograms were obtained by HPLC,and evaluated using the similarity evaluation software for chromatographic fingerprints of traditional Chinese medicine（2012 version）.2.The content of 11 saponins was determined using external standard method,and cluster analysis and principal component analysis were used to distinguish and compare them.3.A single factor study was conducted on the extraction rate of total saponins from Panax quinquefolium by investigating the type of solvent,solvent fold amount,extraction temperature,and extraction time.An orthogonal design scheme was designed for the extraction process.The total saponins extracted from Panax quinquefolium fixed on D101 macroporous adsorption resin were separated and purified using water-60%ethanol by investigating the sample volume and elution fold amount.4.The content of the total saponins of Panax quinquefolium was determined by UV spectrophotometry,and the content of common monomer saponins was determined by HPLC.The saponins and their structures in Panax quinquefolium were identified by UHPLC-Q Active Orbitrap/MS in combination with databases and relevant literature.5.Adopt 600μMol L^-1of H₂O₂was used for damage modeling of H9c2 at a concentration of 2.5μg/mL、5μg/mL、10μg/mL of total saponins from Panax quinquefolium were administered and their survival rate was measured using the CCK8 method.6.Cell flow cytometry was used to detect the impact of total saponins of Panax quinquefolium on ROS,ELISA was used to detect the impact of SOD,MDA,and myocardial enzyme levels,JC-1 was used to detect its impact on mitochondrial membrane potential,and Western blot was used to detect the expression of Bax,Bcl-2,and other proteins in cells.Results:1.The fingerprint similarity of 16 batches of Panax quinquefolium was greater than 0.90.The contents of 11 ginsenosides（Rg₁,Re,Rg₂,Ro,Rb₁,Rc,Rb₂,Rb₃,Rd,Rg₃,Rh₂）were determined,but no ginsenoside Rf was detected.The higher the growth years of American ginseng,the higher the saponin content.The quality of American ginseng in Northeast China is higher than that in Shandong Province.The results of cluster analysis and principal component analysis for 16 batches of American ginseng were similar.2.The extraction solvent used for extracting the total saponins of Panax quinquefolium is 80%ethanol,the solvent content is 40 times,and the extraction time is 2 hours at 100℃.The sample loading amount is 125 mL of crude drug extraction solution with a concentration of0.139 g/mL,and the elution volume is 60%of the volume of three columns of ethanol solution.3.The purity of the total saponins obtained from American ginseng is about 80%,and the content of 11 common ginsenosides accounted for 65.93%of the total saponins.Through the mass charge ratio of the compound mother nucleus and its ionic fragments,another 21 saponin components were identified,and a total of 32 saponin compounds were identified from the total saponins of American ginseng.4.Use 600μmol/L of H₂O₂to damage H9c2,the cell survival rate was 67%.The total saponins of American ginseng can improve the survival rate of cardiac muscle cells by about 10%.5.The total saponins of Panax quinquefolium can reduce the levels of MDA,ROS,and myocardial enzymes,and increase the level of SOD;Meanwhile,total saponins of Panax quinquefolium can increase mitochondrial membrane potential.The expression level of Bcl-2 protein was increased,while the expression level of Bax protein was significantly decreased compared to the model group.Conclusion:The fingerprint and content determination method established in this study are accurate and reliable,providing a reference for the quality control of American ginseng.The total saponins of American ginseng were extracted and separated,and qualitative and quantitative analysis were conducted.Total saponins of Panax quinquefolium play a protective role in H₂O₂induced myocardial injury by influencing oxidative stress and myocardial enzyme levels,thereby reducing cell apoptosis.