Studies On Isolation,Purification,Structure And Properties Of Acidic Polysaccharide From Aureobasidium Pullulans G16

Microbial extracellular polysaccharides possess excellent physical properties and biological activities,as well as short production period.It can realize industrial production through submerged fermentation on a large scale.Therefore,it is of great significance to develop a new type of microbial extracellular polysaccharide with good properties.Pullulan secreted by Aureobasidium pullulans has been widely used because of its excellent properties.Besides,acidic polysaccharide can also be secreted by A.pullulans.A.pullulans G16 preserved in our laboratory can secrete pullulan and acidic polysaccharide simultaneously.In this paper,structure and properties of acidic polysaccharide which isolated from fermentation broth of A.pullulans G16,were investigated after purification.And effects of three kinds of nitrogen source on production of extracellular polysaccharides of A.pullulans G16 were also studied.Acidic polysaccharide from A.pullulans G16 was obtained by ethanol precipitation,protein removal with enzyme-Sevag treat and CTAB precipitation.And then it was purified by fractional precipitation with ethanol and DEAE-52 cellulose column chromatography.It was proved to be homogeneous by Sephadex G-200 analysis and Ultraviolet whole wavelength scanning.The result of high performance gel filtration chromatography showed that the average molecular weight of the acidic polysaccharide was 495450.By means of chemical analysis,gas chromatography,infrared spectroscopy,periodate oxidation,Smith degradation,partial acid hydrolysis,enzymatic hydrolysis and liquid chromatography-mass spectrometry,this polysaccharide was proved to be a kind of heteropolysaccharide containing 12.76% of mannuronic acid.And it was composed of mannose,glucose and galactose in a molar ratio of 2.53:27.21:0.75.What's more,the backbone of the acidic polysaccharide was composed of glucose,mannose and galactose in a molar ratio of 2.48:15.92:0.72.The molar ratio of ?-(1?3)linked mannose to galactose was 1.56:12.80 in backbone,and the rest of mannose,glucose as well as galactose were linked by ?-(1?4)glucosidic bonds.The branch of acidic polysaccharide was composed of ?-(1?6)linked glucose.The solubility of acidic polysaccharide from A.pullulans G16 was poor.It was slightly soluble in deionized water,NaOH and HCl solution,and insoluble in DMSO,ether and acetone.Viscosity of the acidic polysaccharide solution was high and increased with increasing solution concentration.In the range of 30°C to 80°C and pH 4.0 to 10.0,the viscosity of the acidic polysaccharide solution was stable.K+ and Ca2+ had little effect on viscosity of the acidic polysaccharide solution and it behaved salt-resistant stability.When the concentration was in the range of 0 to 1 mg/m L,reduction power,scavenging activities of hydroxyl radical,superoxide radical and DPPH radical of acidic polysaccharide solution increased with increasing solution concentration.And IC50 of scavenging activities of hydroxyl radical,superoxide radical and DPPH radical were 1.45 mg/mL,3.62 mg/m L and 1.11 mg/mL respectively.Effects of three kinds of nitrogen source on production of extracellular polysaccharides of A.pullulans G16 were studied.The results showed that(NH4)2SO4 and higher concentration(?1.8 g/L)of yeast extract were better nitrogen source for pullulan production of A.pullulans G16;NaNO3 and low concentration(<1.8 g/L)of yeast extract were better nitrogen source for acidic polysaccharide production of A.pullulans G16.