Isolation And Purification, Structural Investigation And Bioactivity Of Polysaccharide From Craterellus Aureus Berk. Et Curt.

Craterellus aureus Berk. et Curt. is an edibable and medicinal fungi, which is widelydistributed, delicious and nutritious. At present, the research of Craterellus aureus Berk. etCurt. focuses on the nutrients, antibacterial activity and so on. But there are few studies onCraterellus aureus Berk. et Curt. polysaccharide. This paper mainly studies the extraction,separation, purification, primary structure and biological activity of Craterellus aureus Berk.et Curt. polysaccharide. The main results are as follows:The content of crude protein, total polysaccharide, crude fat, crude fiber, ash and waterin the dry sample were:22.82%,49.58%,4.36%,4.58%,9.18%and6.98%. Craterellusaureus Berk. et Curt. was rich in minerals and essential amino-acid.The optimized water extracting technology of Craterellus aureus Berk. et Curt.polysaccharide was investigated. The single factor experiment and orthogonal experimentshowed that the influence factors of polysaccharide yield were: extraction times> extractiontime> extraction temperature> liquid-solid ratio. The optimum water conditions were asfollows: extraction temperature90℃, liquid-solid ratio30:1（mL/g）, extraction time3h,extraction times2times, the yield could come to10.9%in these conditions.The separation and purification of Craterellus aureus Berk. et Curt.polysaccharide（CAB）: CAB was obtained after polysaccharide extracted in the optimumconditions, with decolored, ethanol subsided, deproteinized and ethanol subsided. CAB wasseparated and purified by DEAE-52ion-exchange column chromatography, Sepharose CL-6Bgel column chromatography and100KD ultrafiltration centrifugal tube. Then three purifiedpolysaccharides were obtained which is named CAB â… , CABâ…¡-2, CAB â…¢. The weightaverage relative molecular mass of CABâ… , CABâ…¡-2and CAB â…¢ were16271Da,19513Daand56004183Da which were detected by GPC.The structure of CAB â… w as analysed. The infrared spectra showed CABâ…  contained thestretching vibration of O-H at3412cm^-1, the stretching vibration of C-H at2925cm^-1, thestretching vibration of a group of carboxylic at1417cm^-1. CAB â…  contained pyranoid ring andmannose because of the absorption peak at1151¹043cm^-1and867cm^-1,800cm^-1.Monosaccharide composition analysis of CABâ… by GC demonstrated CAB â… was consistedof fucose, mannose, glucose and galactoside with the mole ratios was1:2.11:3.13:6.45.Periodate oxidation and Smith degradation showed CABâ…  contained1â†'6,1â†',1â†'2and1â†'3type glycosidic bond. Fucose and mannose couldn’t be oxidized and should be1â†'3typeglycosidic bond. Large amounts of glucose could be oxidized, the glycosidic bond mainly was 1â†'6,1â†'or1â†'2. All galactoside could be oxidized, the glycosidic bond was1â†'6,1â†'or1â†'2. NMR result analysis proved that CABâ… contained α, β-configuration, and partsubstitutional bond was C-2, C-3and C-6, non substitutional bond was C-4.The study results of biological activity of CABâ… , CABâ…¡and CABâ…¢ showedCraterellus aureus Berk. et Curt. polysaccharides had ability of scavenging DPPH· freeradicals and good ability of reducing power. Both of them indicated they had antioxidantactivity. Anticomplement research chose guinea pig blood serum as the complement, andresult showed Craterellus aureus Berk. et Curt. polysaccharides owned good anticomplementactivity. Antitumor activity of Craterellus aureus Berk. et Curt. polysaccharides wasdetermined on two kinds of cancer cells, human hepatoma cells Hep G2and human breastcancer cells MCF-7by MTT assay. The results showed that CABâ… , CABâ…¡and CABâ…¢ havecertain inhibition to this two kinds of tumor.In the three biological activities Craterellusaureus Berk. et Curt. polysaccharides was dose-dependent on the whole and the purifiedpolysaccharide showed good biological activity.