Preliminary Studies On The Biological Characters And Bioactive Compounds Of Xylaria Nigripes

The biological characters and some bioactive compounds of Xylaria nigripes , which originally collected from Northern Zhejiang province were studied by means of tissue isolation, liquid fermentation, PCR, column chromatography, TLC, HPLC and spectrum technology . The main contents included the optimization of the nutritious factors and the environmental conditions for its growth, screening of its liquid fermentation condition;determination of the growth curve of the spawn in the process of fermentation;analysis of the ITS sequence of rhizomorph and mycelium;isolution, purification and structural analysis of polysaccharide extracted from the mycelia of Xylaria nigripes;extraction and determination of the water-soluble polysaccharide ,mannitol and nucleotides . The result showed as below:1. The optimum nutritious sources of solid medium and environmental condition for Xylaria nigripes were as follows: the glucose and the fructose were the optimum carbon source for it' s growth;the Soybean powder, peanut powder and silkworm net powder were the optimum nitrogen source;the citric acid can promoted the growth of mycelium. Xylaria nigripes was a middle-temperture fungus which can grow at the circumstance of 22℃-30℃, but the optimum temperture is 25℃-26℃and the optimum pH for growth is about 6.0.2. The optimum medium constitutes for the liquid fermentation of Xylaria nigripes were: potato juice 20%, peptone 5%, silkworm net powder10%, glucose 20%, magnesium sulfate 1%, Zinc sulfate 1%, potassium dihydrogen phosphate 0. 5%,dipotassium hydrogen phosphate l%03. Four typical growth periods in the liquid fermentation were described: the hesitation growth period, the rapid growth period, the steady growth period and the old period. The best fermentation time is about 66 hourso4. The internal transcribed spacer (including ITS,, ITS^ and 5. 8SrDNA gene) of nuclear ribosomal DNA from the mycelium and rhizomorph of Xylaria nigripes were amplified by means of PCR technologyo The PCR products electrophoretic patterns and nucleotide sequence were complete accordant, The result indicated that the fermentable myceliunK cultured mycelium which grew on the solid medium and the rhizomorph are the same material which origined from a single strain, meanwhile, the ITS sequence similarity between mycelium and sclerotia ■> Xylaria kedahae Lioyd (Xiangyin, Hunan) > Xylaria hypoxylon (L.)Grew. ( Guiyang , China) were 87. 91% > 77. 90% > 71.18% respectively05. The extraction optimination of water-soluble polysaccharide from mycelium was determined by orthogonal experiment in four factors and three levels, the temperature was 95°C, marinated in 3 multiple volume solution for 5 hourso A maximum extraction of 20.45 mg/g has been obtainedo6.Crude polysaccharide was extracted from the cultured mycelium of Xylaria. nigripes by means of extraction with hot water and precipitation with alcohol, The proteins in it was removed by Sevage method, constituent XN-1 and XN~2 were isolated and purified by DEAE-52 chromatography. Their homogeneity were verified by UV scan and Sephadex G-100 chromatography, TLC shows that XN-1 was partly composed by glucoseo The reaction of periodic acid suggested that the sugarresidue partly be interlinked by 1—6 bondso7.There was plenty of mannitol in the Xylaria nigripes , the sum of mannitol was 57. 5 mg/g, 53.5mg/g, 65mg/g in the mycelium, rhizomorph and sclerotia respectivelyo8. Composition of nucleosides in fermental mycelia ^ rhizomorph and sclerotia were determined by means of high performance liquid chromatography(HPLC), The results showed that the content of uridine in the fermentsl mycelia ^ sclerotia and rhizomorph was 8. 79mg/g^ 2. 20mg/g> 0. 29mg/g respectively;the adenosine was 0. 32mg/g^ 2. 07mg/g^ 0. 24mg/g respectively, besides, other three nucleosides whose content were comparatively high were discovered but do not know what they are, and need further researcho...