Studies On Liquid Fermentation Of Pleurotus Nebrodensis And Bioactivities Of Exopolysaccharides

Pleurotus nebrodensis is a kind of edible and medical fungus. Polysaccharides from Pleurotus nebrodensis possess a wide variety of physiological activities, such as antitumour, anti-aging, immunomodulating, etc. In this paper, a study was carried out on the submerged fermentation process, active substances of mycelia of Pleurotus nebrodensis, extraction and purification of exopolysaccharides, and physical and chemical properties of exopolysaccharides from Pleurotus nebrodensis. The biological activities of exopolysaccharides were also studied.By orthogonal test, the optimum medium composition had been obtained. It was as follows: glucose 1.0%, corn powder 3.0%, yeast extract 0.1%, soybean powder 2.0%, KH2PO4 0.3%, MgSO4·7H2O 0.1% , CaSO40.1%.Also the effects of pH, medium capacity, inoculation quantity, shaking frequency on the mycelia growth were studied. The optimum shaking culture condition was as follows: initial medium pH 6.5, 10% inoculation quantity, 160r/min shaking frequency, 100ml medium in 250ml shaking flask, 6 days incubation. The ferment condition of 5L fermentor was as follows: 60% inoculation quantity, 25±1℃,100r/min spurring speed,4 days incubation. Productivity of 2.9g/100ml was obtained, under the condition as above. Analyses of active substances of the mycelia of shaking culture and the fruit bodies of Pleurotus nebrodensis were compared. The ultraviolet spectrum showed that the shapes of peaks of the sample were approximately the same, and the mycelia had different nucleic acid substances from the fruit bodies. The composition of amino acids in the mycelia of Pleurotus nebrodensis was the same as that of fruit bodies.The suitable procedure for extraction exopolysaccharides from Pleurotus nebrodensis was determined as follows: pH of ferment liquid was changed to 6.0, concentrated to 1/2 volume at 60℃and then precipitated at a final 75% ethanol at 4℃overnight .The pellets were harvested by centrifuge, washed several times with 100% ethanol, removed protein by Sevag method, removed low molecular substances by dialyze and dried by vacuum refrigerator.The raw exopolysaccharides precipitate was further purified by DEAE-DE 52 and Sephadex-G 100 column chromatogram to obtain the homogeneous component, which was proved by polyacylamide gel electrophoresis and Sephacryl S-300 column. It contained 90.5% polysaccharides. The component of exopolysaccharides was determined to be composed of D-Gal, D-Glu, D-Man, D-Ara and D-Xyl in a molar ratio of 2.448 : 1.466 : 0.684 : 0.700 : 0.586 by gas chromatography. The infrared spectrum indicated that the glucosidal linkage in exopolysaccharides wasβ-pyranose.The Anti-aging Effect of the exopolysaccharides from Pleurotus nebrodensis was detected in vivo and vitro. The results showed that comparing with contrast, the exopolysaccharides of Pleurotus nebrodensis have notable ability of defending against free radical inhibit the generation of MDA and decreasing the swelling of mitochondria by·OH at the low dose. The exopolysaccharides could obviously strengthen the activities of SOD, GSH-PX and CAT in blood and liver tissue, reduce the content of MDA. These results suggested that exopolysaccharides from Pleurotus nebrodensis liquid fermentation have bioactivities.