Study On The Quality Differences Of Tetrastigma Hemsleyanum And Its Biologically Active Material Basis

Tetrastigma hemsleyanum Diels et Gilg is a rare Chinese herbal medicine in the folk,which belonging to the grape family of Vitaceae.T.hemsleyanum is widely distributed in different producing areas such as Zhejiang,Fujian,Guizhou,Guangxi,etc.The clinical application of T.hemsleyanum has great potential.It has shown outstanding medicinal value in anti-coronavirus disease 2019(COVID-19)and now it has a broad market.In order to improve the quality of T.hemsleyanum medicinal materials and discover the basis of its efficacy,this article firstly evaluates the quality of the samples from 5 origins and the underground parts of T.hemsleyanum from Zhejiang,and further studies the material basis of its biological activity under the guidance of biology.Specifically,it is divided into the study of the difference of antibacterial ability and antioxidant capacity of different extract parts of T.hemsleyanum,preliminary discussion on the antibacterial mechanism of the biologically active extract parts and the analysis and identification of effective parts.The main research conclusions are as follows:1.Comparison of the differences in the contents of effective components in different parts of T.hemsleyanum underground of Zhejiang.Routine testing and determination of active ingredient contents of the tuber of T.hemsleyanum from different origins.Through the determination of moisture and alcohol-soluble extract content of the roots of T.hemsleyanum from 5 producing areas,the results showed that the moisture content of T.hemsleyanum from different producing areas ranged from 5.97 to 11.01%.The extract content range is 9.24～18.82%,and the average extract content is 15.04%,which all meet the requirements of "Zhejiang Traditional Chinese Medicine Processing Regulations"(2015 Edition)(≥ 9%);The highest total flavonoid and the phenol contents are in Zhejiang Wenling(ZJ-WL)and Fujian Shaxian;the highest content of polydatin and resveratrol is in Fujian Shaxian.Astragalin is the highest in Guizhou Tianzhu.The highest content of quercetin and kaempferol is in ZJ-WL.T.hemsleyanum from the Zhejiang province have higher effective component biomass than others.In addition,the analysis in this article found that there is a correlation between the different indicators of T.hemsleyanum.2.The content of the effective components in the roots(large tubers,small tubers and fibrous roots)of T.hemsleyanum in ZJ-WL and the fibrous roots of T.hemsleyanum in ZJ-HZ were determined.The results showed that the total flavonoids and phenols in the fibrous roots of T.hemsleyanum were significantly higher than those in the tubers.Polygonin,resveratrol and kaempferol were the highest in the fibrous roots.The content of astragalin is the highest in large tubers.Overall,the effective components of the fibrous roots in the underground parts of T.hemsleyanum are significantly higher than tubers.3.Study on the antibacterial and antioxidant activity of different extracts of T.hemsleyanum.According to the method of DPPH,ABTS+ combined with FRAP,the antioxidant activity of different extraction parts of T.hemsleyanum was compared,and the results are all the same that BAE of T.hemsleyanum had the strongest antioxidant capacity.And according to the antibacterial activity of different extraction parts of T.hemsleyanum,the study found that PEE,EAE and BAE have the best antibacterial effect on E.coli.E.faecalis and P.aeruginosa have the best antibacterial effect on BAE.BAE has the strongest effect on S.aureus.PEE,EAE and BAE all show antibacterial activity against B.subtilis.However,the four extraction parts of T.hemsleyanum on C.albicans all have or have very weak antibacterial activities.It can be seen that BAE of T.hemsleyanum is preliminarily judged to be the material basis for the antibacterial and antioxidant effects of T.hemsleyanum.4.Study on the antibacterial mechanism of BAE of T.hemsleyanum.Through AKP content determination and scanning electron microscope observation of the effect on the cell wall,it can be seen that BAE of T.hemsleyanum can rupture the cell membrane and cell wall by destroying the cell structure.In this way,BAE causes bacteria to die and achieves antibacterial effect.Moreover,the boundary between the bacteria is not obvious and the phenomenon of adhesion,causing the contents to ooze.Therefore,E.coli is more sensitive to the treatment of BAE,and the higher the concentration,the better the antibacterial effect,which is dose-dependent.5.Based on UHPLC-QE-MS technology to quickly analyze the bioactive substance basis of T.hemsleyanum.58 compounds were identified in BAE of T.hemsleyanum,mainly including flavonoids,alkaloids,organic acids and fatty acids,and a small amount of other chemical components such as polyphenols,benzophenones and amino acid derivatives.18 flavonoids are identified including 4 flavonoids,6 flavonols,2 flavanols,and 2 dihydroflavonols(Silicristin and(2R)-6,8-Diglucopyranosyl-4’,5,7-trihydroxyflavanone),1 dihydroflavonoid(Ampelopsin D),1 procyanidin(Procyanidin C1)and 1 isoflavone(Genistein).Nobiletin,Genistein,Silicristin,Mulberrin,(2R)-6,8-Diglucopyranosyl-4’,5,7trihydroxyflavanone are the first discovered in flavonoids.8 alkaloid compounds were identified,including 4 indole alkaloids(Psilocin,1-Methoxyindole-3-carbaldehyde,Lenticin and Trigonelline),and 2 β-carbolin alkaloids(HarMaline and Harmanine),1 quinoline alkaloid(5,6,7,8-Tetrahydro-2,4-dimethylquinoline)and 1 pyrrolidine alkaloid(5’-Hydroxycotinine).These 8 kinds constituents of alkaloids are all found in the tuber roots of T.hemsleyanum for the first time.6 organic acids,8 fatty acids,and 17 other chemical components were also detected.This result provides a research idea of the material basis of the efficacy of T.hemsleyanum.