| Agaricus blazei Murill,having medicial and edible function,is a kind of fungi native of Brazil and belongs to the Agaricomycetes,Hymenomycetes,Agaricales,Mushrooms.At present,the research mainly concentrated on the separation and purification of its polysaccharides,whereas few on structure and biological activities.In this paper,the Agaricus blazei Murill as raw materials,its water-soluble polysaccharide have been discussed on purification,structure characterization and some biological activities.It can provide the basis for the development of health products and further theoretical research.The main results are as follows:(1)The polysaccharides were extracted and purified from Agaricus blazei Murill at first,and then the physicochemical properties of purified polysaccharide ABD was analysed.Crude polysaccharides was gained through hot water extraction,deproteinization by Sevage,repeated alcohol precipitation.The yield was 18.69%,and the sugar content was only about 47.50%.The ABD polysaccharide was obtained after purified by DEAE-sepharose Fast Flow anion exchange chromatography column,and its sugar content was 93.09%.It was identified as a single component polysaccharide by gel permeation chromatography and freeze thaw analysis.The weight average molecular weight was 2058 kDa.The results are obtained that ABD contains no starch,a very small amount of protein and uronic acid,which through the analysis of the physical and chemical properties.ABD is a neutral polysaccharide.(2)The structure characteristics of polysaccharide ABD were analyzed by infrared spectroscopy,nuclear magnetic resonance,acid hydrolysis and methylation.Through the infrared analysis,ABD proved to be a polysaccharide,which has characteristic absorption peaks at 3389 cm-1,2935 cm-1 and in the range of 1300~1000 cm-1,and the alpha and beta sugar rings at 853 cm-1and 929 cm-1 respectively.The monosaccharide composition of ABD was evaluated by high performance liquid chromatography(HPLC)and was found to consist of glucose(83.59%),galactose(5.46%),mannose(0.59%)and fucose(1.04%).The main linkages type of ABD were comprised of α-1,3-D-Glc,α-1,4-D-Glc and β-1,3-D-Gal,as well as β-1,3-D-Glc and α-1,6-D-Glc as side lain,and terminated with α-L-Man and α-L-Fuc residues based on the periodate oxidation-Smith degradation,methylation and NMR analysis.The ABD was showed that have no the three-helix conformation but have more brands andside lains by I2/KI and Congo red test.Simultaneous Thermogravimetry-Differential Scanning Calorimetry(STA/TG-DSC)was applied to analyzied the stability of ABD.The results showed that ABD had good thermal stability and the glass transition temperature and thermally decomposed temperature of ABD was around 65.6 and 305.0 °C respectively.(3)The antioxidant properties of ABD were studied.The reduction ability of ABD was proved by reduction of Fe3+.The inhibitory effect of ABD on free radicals was determined by the DPPH· scavenging rate.The total antioxidant capacity of ABD polysaccharide on active oxygen was characterized through the ORAC value.The results show that ABD polysaccharide has certain antioxidant capacity and effect good at high concentration.(4)Three kinds of cancer cells(Hela,MCF-7 and HepG-2)were used to carry out the anti-cancer experiment of ABD polysaccharide.There were three doses groups of ABD acted them.The inhibition rate of ABD on three cancerous cells was similar at low and moderate doses,but the inhibitory effect on MCF-7 and HepG-2 was higher than Hela at high concentration.The results showed that each concentration of ABD had a centain anti-cancer effect,furthermore the effect of high concentration is obvious.(5)The immunological and anti-inflammatory effects of ABD polysaccharide were determined.A cell model was constructed by RAW264.7 to measured that the secretion of NO,TNF-α and IL-6.The results prove that ABD polysaccharide can not only enhance the phagocytosis of macrophage and secretion of proinflammatory cytokines,to enhance the role of immunity;but can reduce the secretion of proinflammatory cytokines in excessive inflammation response induced by LPS,inhibiting inflammation furture expansion,playing an anti-inflammatory role.ABD polysaccharide has the effect of bidirectional regulation of immune function,and can maintain the balance of the immune system.The new ABD is a high weight molecular neutral polysaccharide,mainly composed of glucose and galactose.It has good thermal stability.Although it does not contain the space conformation of the three-helix,it has certain antioxidant capacity and anti-cancer ability in vitro,and has bidirectional regulation of immune function that maintained the balance of the immune system. |
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