Extraction,Separation And Purification Of Ginkgo Biloba L. Polysaccharide And Bacteriostatic Study

Ginkgo biloba,alias:Ginkgo biloba,was the most famous plant in the world.It can be used as a cosmetic and was a dual-use plant.China's Ginkgo biloba yielded accounts for about 70%of world production.Ginkgo biloba polysaccharide had many important biological activities and functions,and it was one of the hot spots of drug development and research.In recent years,there had been many studies on the extraction,purification,structure identification and biological activity of plant polysaccharides.However,there were few studies on Ginkgo biloba polysaccharides and even polysaccharides in Ginkgo biloba leaves.In this paper,the leaves of Ginkgo biloba leaves were taken as the research object,and the Ginkgo granules were extracted and purified,and the Ginkgo granules with higher purity were obtained.The antibacterial effect of Ginkgo biloba polysaccharides was studied and the antibacterial activity was deliminator explored.The mechanism was followed by compounding into a mixed polysaccharide with Zea mays,screening the best compounding ratio,and studying the stability of the antibacterial activity of the compound polysaccharide.Finally,the toxicological experiment was used to provide experimental basis for further application.The specific research contents and results are as follows:The dried Ginkgo biloba leaves were used as raw materials,and the extraction yield of crude polysaccharides was used as the experimental index.The optimal process conditions for extracting Ginkgo biloba leaves polysaccharides by cellulase method were optimized by response surface method:enzyme dosage 1.65%,enzymatic hydrolysis temperature 50?,enzyme The solution has a pH of 5.0 and a hydrolysis time of 65 min.The maximum extraction yield can reach 9.1733%±0.1348%.The polysaccharide content of the crude polysaccharide was determined to be 88.27%,and the property was identified as a non-starch polysaccharide.The effects of Sevage method,TCA method,isoelectric point method and enzymatic method on the removal of protein from Ginkgo biloba L.polysaccharide were compared with the results of protein removal rate and polysaccharide loss rate.The optimal protein removal method was selected to enzymatically remove protein.The enzymatic method combined with isoelectric point method was the best process for removing protein from Ginkgo biloba polysaccharide:the enzyme addition was 0.90%?calculated as substrate?,the enzymatic hydrolysis time was 1.1 h,the enzymatic hydrolysis temperature was 46?,and the enzymatic hydrolysis pH was 4.7.Under this condition,the protein loss rate was 92.73%,and the polysaccharide loss rate was 9.98%.The decolorization treatment of Ginkgo biloba leaves polysaccharides with AB-8 macroporous resin showed that the time should not exceed 7 h during static decolorization.When the loading amount and the loading concentration were certain,1.5 mL/min was the best elution flow rate for decolorization.When the loading amount and flow rate were constant,it was more appropriate to select the 40 mg/mL loading concentration for decolorization.The three polysaccharides were further isolated,named GBLP-60,GBLP-70 and GBLP-80,of which the highest content of GBLP-60,accounting for 88.14%of the total polysaccharide,and GBLP-70 and GBLP-80 accounted of the total polysaccharide,respectively1.23%and 0.51%,the sum of the three polysaccharides accounts for 89.88%of the total polysaccharide,which was silver The main components of apricot deciduous polysaccharides;physical and chemical identification results:the total sugar content of GBLP and alcohol precipitation components was between 92.07%⁹7.32%,and contains a small amount of protein and uronic acid,UV spectrum identification of Gingko deciduous polysaccharides in 190 to 200 There was obvious absorption peak at nm,and there was no other peak,which was consistent with the characteristics of polysaccharide.The heterogeneous polysaccharide formed by polymerization of seven kinds of monosaccharides was determined by high performance liquid chromatography.The molar ratio of composition was mannose:rhamnose:glucose:Galactose:Xylose:Arabinose:Fucose=1.28:3.35:1.42:3.58:1:2.66:2.75.The inhibition of the inhibition zone by the Oxford Cup method showed that GBLP-60 had a relatively strong antibacterial activity against Bacillus cereus in Escherichia coli and Gram-positive bacteria in Gram-negative bacteria.By measuring the length of hyphae,it was determined that GBLP-60 inhibited the growth of Aspergillus niger.After 1 day of culture,the growth effect of GBLP-60 inhibiting Aspergillus niger was beginning to be obvious,and the effect of high concentration of GBLP-60 on Aspergillus niger was observed.The best,after 4 days of culture,the effect of high and medium concentration of GBLP-60on the growth of Aspergillus niger mycelium continued to be significant,but the antibacterial effect of low concentration of GBLP-60 began to be insignificant.To investigate the effect of GBLP-60 on the growth and development of test bacteria,the results showed that GBLP-60 can inhibit the growth of Escherichia coli?G-?and Bacillus subtilis?G+?,and its growth curve changes,which was different from normal cells.Using Escherichia coli and Bacillus cereus as test bacteria,the inhibitory pathway of GBLP-60 against bacteria and the effects on protein,DNA synthesis and ATPase activity were studied.The results showed that GBLP-60 destroyed the cell membrane by making it.The nucleotides in the bacteria were dissolved to inhibit the growth of bacteria.GBLP-60 inhibited the synthesis of proteins,DNA and ATPase activity in the bacteria.The polysaccharides from Ginkgo biloba L.were combined with the polysaccharides from Porphyra yezoensis,and the optimal compounding ratio of Ginkgo biloba polysaccharides and P.yezoensis polysaccharides was determined by using the diameter of the inhibition zone as the index.The results showed that the optimal compounding ratio was 1:1,the diameter of the inhibition zone is up to 34.75 mm±0.893.Escherichia coli was used as the test bacteria,and the minimum inhibitory concentration?MIC?of the compound polysaccharide was investigated to be 1 mg/mL.Taking the absorbance of the bacterial liquid as an indicator,four factors affecting the stability of the antibacterial activity of the compound polysaccharide were discussed:pH,temperature???,ultraviolet irradiation time?h?and metal cation.The results showed that the antibacterial activity of the compounded polysaccharide solution was better under neutral or alkaline conditions,and the antibacterial activity was lower under acidic conditions.When the temperature of the compounded polysaccharide solution was lower than 80?,the antibacterial activity was not obvious.The effect was higher than 80?.The antibacterial activity of the compound polysaccharide solution under UV light was less than 6 h,and it has no obvious effect on the antibacterial activity.The antibacterial activity of the compound polysaccharide solution after 6 h treatment After 8 h,its properties tend to be stable;K⁺had no significant change on the antibacterial activity of the compounded polysaccharide solution,and Mg²⁺and Zn²⁺had certain effects on the antibacterial activity of the compounded polysaccharide solution,but the overall change was not obvious,Na⁺The antibacterial activity of the compound polysaccharide solution decreased first and then decreased,and Na⁺had the greatest influence on the antibacterial activity.Through acute and subacute toxicology experiments,no compound polysaccharides were found to produce meaningful pathological changes in mice.