|
Phellinus igniarius is a fungus of high medicinal value and promising utilization perspective. The polysaccharide, which is one of many bioactive components, plays an important treating role. This study was focusing on submerged culture, extraction and purification of polysaccharide for Phellinus igniarius. To enhance the yield of mycelia biomass, the experiments were done, including the improvement of submerged culture conditions and the extraction and purification of the polysaccharide.Carbon source tests showed that the corn flour was the best carbon for producing mycelia biomass in all the tested carbon sources. Nitrogen source tests showed that the powder of bean is superior to the other nitrogen sources. Inorganic salts and growth factors tests showed KH2PO4, MgSO4 can promote the yield of mycelia biomass. The optimal medium was determined was corn flour 4%, powder of bean 1.5%, KH2PO4 0.1% MgSO4 0.1% through the orthogonal test. Response-plane of Carbon source and Nitrogen source was: z=1.7950x+1.1967y-0.1967x2-0.1267y2-0.185xy-2.9722 z: mycelia biomass, x: Carbon source,y: Nitrogen sourceThe culture condition factors including temperature, initial pH value, loading capacity of culture medium, rotating speed, inoculating amount and culture duration, the results showed that the optimal culture conditions were culture temperature 26℃,initial pH 6.5, rotating speed 130 rpm, the culture duration 7 days, the inoculating amount 15%, containing 100 ml culture medium in the 250 ml Erlenmeyer flasks.The extraction condition of polysaccharide was combination with time, temperature, times, and solvent proportion. The optimal extraction condition was determined that the best of extraction condition were 2.5 hours, 60℃,3 times, and solvent proportion 14 times. The orthogonal test which polysaccharide isolates protein by Sevag was made. The result showed that the best conditions were what volume ratio of chloroform to butanol in Sevag solution is 10:3, volume ratio of sample to Sevag solution is 1:1, and surge time is 10 min. Response-plane of removing protein rate was:z=121.5167x+32.4499y-50.0999x2-4.4299y2-2.5600xy-46.7733z: removing protein rate, x: volume ratio of sample to Sevag solution （V/V）,y: volume ratio of chloroform to butanol（V/V）In this study, the firstly polysaccharide of Phellinus igniarius was purified with macroporous resin and DEAE-cellulose, and then it was purified with Sephadex G-100. After the primary separation, the two polysaccharide substances, H-A and H-B, were obtained. For the further separation, basing on the amount of molecular, H-A and H-B were puritied by Sephadex G-100, obtained H-A1 was 95.6%, H-B1 was 97.21%. So as to we know the polysaccharide of Phellinus igniarius was purified and classification well by Sephadex G-100.
|