The Screen Of A Polysaccharid-producing Marine Bacteria And The Study Of Its Polysaccharide Fermentation

In recent years, with the development and application of marine resources, marinepolysaccharides, especially marine bacterial polysaccharides has been drawn increasingattention by scientists.In this paper, marine bacteria502A (Aerococcus urinaeequi HZ)has been Systematic researched in the sample collection, separation and purification ofthe strains, the strain of producing polysaccharide to select, species identification andphylogenetic relationship to build, optimization of fermentation conditions,polysaccharide separation purification and structure identification ect.. Results are asfollows：1. In this experiment, active strain228has been selected from the cultureenvironment of abalone, the strain23of producing polysaccharide has been detected byusing the phenol-sulfuric acid method, which polysaccharide production of8strains ismore than0.5g/L, after repeated experiments, a strain named502A which growthcycle short, yield stability, the strain502A which long rod-shaped has beendetermined as Gram-negative bacteria by staining, in Zobell2216E solidmedium,colonies (diameter2mm) are larger, white, round, uplift, round edges, opaque,non-migratory. After16SrDNA sequence right and the retrieval of NCBI, its species isdetermined as the Aerococcus urinaeequi and its phylogenetic relationship has beenbuilt..2.On the basis of existing single-factor experiment, the medium components of thestrain502A which products polysaccharide by fermentation is done by orthogonaloptimization, the optimal conditions is3%sucrose,0.25%beef extract and3%salt;Through Plackett-Burman design, from a number of fermentation factors to filter out themore significant factors: incubation time, temperature and rotation speed, theexperiment has been further designed by center combination,with the correspondingsurface analysis,the optimum fermentation conditions: the rotation speed is230r/min,temperature is26°C, incubation time is35hours, under optimal conditions,thehighest production of polysaccharide is2.34g/L.3. Crude polysaccharide has been extracted from fermentation broth by the way ofwater extraction and alcohol precipitation, and through Sevage reagents, the protein hasbeen removed from the crude polysaccharides, by UV scanning detection,there is noresidual nucleic acids, proteins;polysaccharide samples have been further purified by DEAE-52column and Sephadex G100column;by high performance gel permeationchromatography (HPGPC), the relative purity of polysaccharide is uniform;and itsweight average molecular weight: Mn=2.84×10~5, number average molecular weight:Mw=2.22×10~5, the distribution coefficient is1.28.4. By the analysis of GC-MS, IR, NMR can get that the EPS-1is mainly composedwith xylose, glucose, mannose, and contains a small amount of glucoside,galactose,and glyceraldehyde; and they are all β comfiguration.Marine bacteria lives in the low temperature environment, temperature conditionwhich polysaccharide synthesis is easy to meet, once applied in the industrialproduction, can effectively reduce energy consumption, and conforms to the trend ofindustrialization of low-carbon production; at home and abroad,the study of bacterialpolysaccharide, especially in the synthesis of polysaccharide of marine bacteria lacks ofresearch, there is empty in industrialization;at present there is no report that Aerococcusurinaeequi products polysaccharide at home and abroad, the results in the area havegood research value and application prospect.