Study On Antibacterial Active Constituents From Traditional Chinese Medicine Euphorbia Helioscopia Linn.

The purpose of this thesis is to study the in vitro antimicrobial active ingredients and their mode of action from Zeqi(Euphorbia helioscopia Linn.(Euphorbiaceae)).In the thesis,based on the preliminary screening of the antibacterial activity of the extracts from traditional Chinese medicine,the target medicinal plant material for further study was determined.Through the activity oriented extraction,separation of the plant chemical components,the antimicrobial compounds from the plant were characterized.The antimicrobial effects of the compounds used alone,and their effects in combination with conventional antibiotics were systematically evaluated.This thesis is composed of four parts.In the first part,21 common Chinese medicinal materials are screened in vitro for antibacterial activity.The screening results are that 13 kinds of Chinese medicinal materials such as Celosia cristata Linn,Euphorbia helioscopia Linn.,etc.showed better antibacterial activity in vitro with broad-spectrum antibacterial activity.the second part is to track and separate the antibacterial compounds from the Euphorbia helioscopia Linn,trace the chemical composition of the active part of the Euphorbia helioscopia Linn.and identify its structure;the first part of the third part passes the separated compounds and 11 compounds in the Euphorbia helioscopia Linn(commercially available).Initial screening of antibacterial activity in vitro was conducted to select monomer compounds with better antibacterial activity in vitro.In the second section,the compounds screened in the first section with better antibacterial activity in vitro and the six commonly used antibiotics(penicillin,Imipenem/cilastatin,amikacin,ceftriaxone,ampicillin,Piperacillin/Tazobactam)through the checkboard microdilution method and time-killing curve method to determine the combined action mode,and the combination of synergistic antibacterial and synergistic sterilization mode was selected to provide a certain reference for clinical application;In the fourth part,after the standard bacteria of Staphylococcus aureus(CMCC(B)26003)and methicillin-resistant Staphylococcus aureus(MRSA166)were treated with different concentrations of Licochalcone A,the images were collected under the scanning electron microscope to observe the passing compound The surface structure of bacteria changes after treatment.The cytotoxicity of Licochalcone A on human lung adenocarcinoma drug-resistant cell line A549 and human gastric cancer cell line NCI-N87 was measured by CCK8 method,and the cytotoxicity of Licochalcone A was observed.By using the method of biological activity-directed separation,the in vitro antibacterial activity test of the drug is combined with the separation and extraction,and the active components with better antibacterial activity are purposefully tracked and separated to avoid blind separation.The activity of the extraction layer and the fractions were measured by agar punching method and broth microdilution method,and the active extraction layer and the fractions were separated through silica gel column and combined with the Pop data(1H NMR,13 C NMR)and literature Contrast to determine the structure of monomer compounds.A total of 6 known compounds were isolated and identified from the Euphorbiaceae plant Euphorbia helioscopia Linn,which were Compound 1(ethyl stearate),Compound 2(?-sitosterol),Compound 3(euphornin),Compound 4(palmitic acid),compound 5(gallic acid)and compound 6(quercetin).Among them,compound 3(euphornin)is a characteristic chemical component in lacquer,which belongs to jatrophane type diterpene.The in vitro antibacterial activity test results of the tested monomer compounds showed that: 11 compounds out of 6 compounds and 11 commercially available compounds had a certain antibacterial effect on each standard bacteria and their resistant bacteria strains.Among them,the myricetin and Licochalcone A monomer compounds have better activity on MRSA.Checkerboard microdilution method showed that when Licochalcone A and six antibiotics including penicillin used in combination with 10 MRSA strains,most of them showed synergistic or additive effects,and a few show irrelevant or antagonistic effects.Time-killing curve method showed that when Licochalcone A combined with antibiotics,two of them showed synergistic bactericidal effect,most of the rest were additive,and a small part showed synergistic and antagonistic effects.Scanning electron microscope and cytotoxicity results showed that at 8MIC and4 MIC concentrations,the number of bacteria is small and no obvious aggregation occurs,and the number of bacteria begins to increase from 1MIC to 1/4MIC.After 8MIC to 2MIC,the bacterial morphology did not change significantly.Licochalcone A showed a significant dose-effect relationship on the inhibition of proliferation of the two tumor cells.The IC50 of A549 cells and NCI-N87 cells were 86.15?g/ml and 46.72?g/ml,respectively.The results in this thesis preliminarily clarified the antimicrobial active substancebasis of Chinese medicine Zeqi,which provided a scientific basis for the further development of the resources of Zeqi medicinal materials and the exploration of the antimicrobial composition of Zeqi and its mode of action,so as to overcome the problem of clinical drug-resistant bacteria infection.