Study On Extraction, Purification, And The Primary Efficacy Activity Of The Polysaccharide Isolated From Ganoderma Sinense

The study was designed to determine the efficacy of polysaccharide isolated from Ganoderma sinense. By using it as raw materials, we have adopted three different methods of conventional water bath extraction, ultrasonic assisted water extraction and supercritical carbon dioxide extraction cooperated with water to extract polysaccharide from Ganoderma sinense(GSP). Then do the experiment in vitro antioxidation and antitumorthe with polysaccharide and select active strong coarse polysaccharide to separate and purify according to the characterization the primary structure. The main research are as follows:1Experimental content(1) Using different methods to extrac polysaccharide from Ganoderma sinense.Using three different methods of water bath extraction, ultrasonic assisted water extraction and supercritical CO2extraction assisted water to extract polysaccharide from Ganoderma sinense.(2) The best extraction technology of polysaccharide from Ganoderma sinenseThis paper investigates the effects on·OH, the effects on·DPPH and the restrain on EC-109of polysaccharide from Ganoderma sinense and according to the test results, we have chosen the methods for extracting polysaccharide. The best extraction technology is selected by taking the extraction ratio of polysaccharide from Ganoderma sinense as index.(3) Decolorization and deproteinization technology of polysaccharide Five kinds of resin-AB-8, D-101, ADS-7, and DM-301and717are evaluated based on decoloration rate, protein removal rate, total sugar retention rate and composition weighted score as indicators. Then, one kind of resin is confirmed to carry out for single-factor experiments. The orthogonal tests L9(33) are carried out based on single factor test to choose the optimal condition of decolorization and deproteinization of polysaccharide.(4) Isolation, purification and molecular weight determination of polysaccharide from Ganoderma sinenseA DEAE-52celluloseion exchange column and a Sephadex G-150gels filter column are successively used to further separate crude polysaccharides, and then determine the product purity and molecular weight with high performance gels filtration chromagraphy (HPGFC).(5) Biological activity research on polysaccharide from Ganoderma sinenseThis paper investigates the effects on-OH, the effects on DPPH and the restrain on EC-109of three purification polysaccharide from Ganoderma sinense, Through the experiment the auther compared polysaccharide component oxidation and antitumor ability.2The results of the experiment(1) The colours of polysaccharide from Ganoderma sinense, which is extracted with the water bath and ultrasonic assisted water are brown powder. The colour which is extracted with supercritical CO2assed water is pale brown. They had no significant difference in extraction yield. The experimental results showed that:the effect of polysaccharide with water bath extraction is the best in withdrawing on·OH and·DPPH and the polysaccharide with water bath extraction is the highest in restraining on EC-109, too. The clearances rate of·OH and·DPPH are87.76%and75.14%when its concentration is2.5mg/mL, and the rate of inhibition of EC-109function of24h is35.31%when its concentration is80mg/L.The results also show that polysaccharide from Ganoderma sinense with water bath extraction has a strong activity.(2) The optimal reflux extraction conditions are determined with single-factor method and orthogonal method. The results indicated the following:the ratio of solution to material20, time2h, temperature90℃, and the rate of extracting is4.23%.(3) ADS-7resin can be used as a high effective one to remove pigment and protein of polysaccharide from Ganoderma sinense. The optimal technological parameters are as follows:the ratio of resin and crude polysaccharides solution (the concentration of polysaccharides solution was2mg/mL) is0.03g/mL, temperature40℃, time1.5h. The discoloration rate is98.36%, the protein removal rate is90.20%, and the total sugar retention rate is63.07%.(4) Three fractions of GSP,, GSP2, GSP3are obtained after fractionating DEAE-52cellulose and Sephadex G-150column. They are all single polysaccharide by HPGPC identification. GSP1, with a relative molecular weight of54200Da; GSP2, with a relative molecular weight of2511.8Da, GSP3, with a relative molecular weight of248.9Da.(5) In the removal of-OH and DPPH, GSP1, GSP2and GSP3have the ability to scavenge. The sequence is:GSP1>GSP2>GSP3. In the test range, there is a certain dose-effect relationship in scavenging capacity of polysaccharide solution with the concentration, when the concentration of GSP1reached2.5mg/mL, the removal rate-OH and DPPH are94.76%and87.69%. As to the rate of inhibition of EC-109function of24h, GSP1and GSP2show their strong activity when their concentration is80mg/L, the maximum inhibition rates of GSP1, GSP2are48.31%and25.33%, and no obvious inhibitory effect of GSP3on EC-109is found.