Isolation And Antimicrobial Activity Analysis Of Green Tea Extrac

The production of summer tea is high,but due to it’s poor taste and production cost,a large amount of tea green is often difficult to make full use of,which is a waste of resources.Therefore,it is of great significance and value to develop new tea products to broaden the tea market.It was found that summer tea is more bitter and astringent because it has higher contents of tea polyphenols and caffeine than any other seasons,and these components can effectively inhibit the growth of bacteria.Based on this,this experiment screened out the varieties with better antibacterial effect from Guizhou featured tea varieties,and studied the extraction process and application of antibacterial components,and the main findings are as follows.1.Identification of antibacterial active ingredientsThe total flavonoid and total phenolic contents of the eight extracts were detected by NaNO₂-Al（NO₃）₃-NaOH colorimetric method and forinol method,and the contents of catechin fraction and caffeine were determined by high performance liquid chromatography,and the correlation analysis was done between the results of the content determination of each component and the size of the inhibition circle obtained in 1.The liquid-phase results showed that there was little difference in the types of the eight extract components and the contents varied.Among the components measured,total phenols,total flavonoids,catechins,EGCG,ECG,EC and EGC were the highest in QM,while caffeine and C were the highest in 6M.2.Comparison of antibacterial effect of green tea extractsUsing the double-layer plate method,the antibacterial effect of eight different green tea extracts on the test bacteria was compared by measuring the size of the inhibition circle,and the extract with the best effect was selected for the determination of the minimum inhibitory concentration（MIC）and the minimum bactericidal concentration（MBC）and the detection of the growth curve.The results showed that the inhibition circles of green tea alcohol extracts（QM,FM,6M,4M）of the four varieties（Qiancha 1 tea variety,Fuding,Qianmei 601,Qianmei 419）were larger than those of their respective water extracts（QW,FW,6W,4W）,and the largest inhibition circle was QM,which was 1.12-1.28 times larger than the smallest 4W.the MICs of QM against S.aureus,P.aeruginosa and E.coli were 3.125 mg·m L^-1,3.125 mg·m L^-1,12.500 mg·m L^-1and MBC were 12.500 mg·m L^-1,25.000 mg·m L^-1,12.500 mg·m L^-1,respectively.The bacterial growth curves showed no growth in the number of bacteria at MIC concentrations,while lower concentrations exhibited prolonged retardation,slowed logarithmic phase and decreased number of stable phase.The results indicated that the antibacterial effect of green tea alcohol extract was better than that of water extract,and among the four tea tree varieties,Qiancha 1 tea variety had the best antibacterial effect.The results of correlation analysis showed that the contents of total phenols,total flavonoids,catechins,EGCG,ECG and EC were correlated with the size of the inhibition circle,so the above components were selected as the test indexes in the subsequent experiments.3.Study on the extraction process of antibacterial active ingredients of Qiancha 1tea varietyThe influence of four extraction factors,namely extraction time,extraction temperature,material-to-liquid ratio and methanol concentration,on the extraction amount of the six components was investigated by single-factor test,and the weights of each component were assigned by Factor Analysis.The results showed that the optimal extraction conditions were as follows The results showed that the optimum extraction conditions were:material-liquid ratio of 1:25（g:m L）,methanol concentration of 75%,extraction temperature of 70℃and extraction time of 12 min.The contents of total phenols,total flavonoids,catechins,EGCG,ECG and EC in the extract of Qiancha 1 tea variety were（260.68±3.30）,（63.54±1.60）,（233.99±1.72）（100.19±2.84）,（46.28±1.24）,（28.32±0.81）mg·g^-1,respectively,and the mass fractions of the index components were basically stable and consistent with the orthogonal test data,with the RSD values within5%,indicating that the preferred process conditions were feasible.The results can provide a theoretical basis for further research of Qiancha 1 tea variety,and also provide a reference basis for the development of its tea products.4.Study on the combined application of green tea extract and antibioticsThe MIC of erythromycin on the three test bacteria was determined by micro-broth dilution method,and the MIC of QM and erythromycin combination on the three test bacteria was determined by micro-checkerboard method,and the partial inhibition concentration index（FICI）was calculated to determine the combined effect and calculate the percentage reduction of MIC when the two were used together.The results showed that the MICs of erythromycin against S.aureus,P.aeruginosa and E.coli were62.5000μg·m L^-1,15.6250μg·m L^-1and 7.8125μg·m L^-1,respectively.the FICIs of QM and erythromycin combination against S.aureus,P.aeruginosa and E.coli were 0.250,0.625 and 0.500,respectively,showing synergistic,additive and synergistic effects,respectively.The combination of erythromycin reduced the MICs of S.aureus,P.aeruginosa and E.coli by 87.5%,50.0%and 75.0%,respectively,compared with those of S.aureus,P.aeruginosa and E.coli alone.The best ratio is erythromycin 7.8125μg·ml^-1,QM 3.1230 mg·ml^-1.The results suggest that the combination of the two can reduce the use of antibiotics,thus reducing the development of bacterial resistance,and has promising applications.