【作者】 李绪全；

【导师】 杨晓彤；

【作者基本信息】 上海师范大学 ， 微生物学， 2005， 硕士

【摘要】 本研究对灵芝属内五个种(Ganoderma lucidum, GL; Ganoderma oerstedii, GO;Ganoderma resinaceum, GR; Ganoderma subamboinense, GS; Ganoderma tsugae,GT)的25 个菌株进行了深层液体发酵,比较了不同菌株的生物量。菌丝体进一步用热水提取、乙醇沉淀后用苯酚-硫酸法和Lowry 法测定了提取物中胞内多糖及蛋白质的产量。结果表明,25 个菌株的总体平均生物量为(14.02±3.49)g/L,以GR-1 菌株的最高(20.52 g/L),GT-3 最低(7.12 g/L),有15 个受试菌株超过平均值;胞内多糖产量从0.10～0.55g/L 不等,平均为(0.28±0.13)g/L,其中GT-4 的多糖产量(0.55g/L)最高,GT-2 的多糖产量(0.10g/L)最低,有11 个受试菌株超过平均水平。蛋白质产量则从0.05～0.68g/L 不等,平均为(0.28±0.18)g/L,其中GL-6 的蛋白质产量(0.68g/L)最高,GT-1 的蛋白质产量(0.05g/L)最低,也有11 个受试菌株超过平均水平。Ganoderma lucidum 种的GL-1、GL-4、GL-6、GL-7 和GL-8 这5 个菌株在生物量、胞内多糖和蛋白质产量三项指标上均表现良好。胞内多糖用三氟醋酸水解后,单糖用1-苯基-3-甲基-5-吡唑啉酮(1-phenyl-3--methyl-5-pyrazolone,PMP)进行衍生,然后用自行建立的反相高效液相色谱条件对三种不同类型(酸性、中性、碱性)的9 种单糖进行鉴定和含量测定。结果表明, 25 个菌株胞内多糖均含有Glc、Gal、Man、GlcN 和Fuc 等五种单糖,均以Glc 为主要单糖组分;GL-3、GL-5、GL-6、GL-7、GO-3、GR-3、GR-4、GR-5、GT-1、GT-2、GT-3、GT-5、GS-2 等13 个菌株还含有Xyl,GL-5、GL-6、GL-7、GL-8、GO-3、GR-3、GR-4、GT-2、GT-3、GT-4、GT-5 等11 个菌株还含有Rha,GL-1、GL-3、GL-8、GR-3、GS-1、GT-1、GT-2、GT-3 、GT-4 、GT-5、GT-6、GT-7 等12 个菌株含有痕量GalUA,而所有25 个菌株均不含有GlcUA。HPGPC 凝胶色谱分析表明25 个菌株的多糖重均分子量介于2.03×104～1.63×105Da 之间,其中重均分子量最大的菌株为GR-1,最小的菌株为GS-1。25 种胞内多糖均为结合蛋白多糖,含有3 或4 个不同的多糖组分,它们的主峰保留时间在16.22～17.47min 之间,以分子量小于5×104Da 的部分为主,处于该区段的多糖部分占全部多糖比例的80.26～98.84%。更多还原

【Abstract】 Twenty-five Ganoderma strains belonging to five Ganoderma species(Ganodermalucidum, GL; Ganoderma oerstedii, GO; Ganoderma resinaceum, GR; Ganodermasubamboinense, GS; Ganoderma tsugae, GT)were cultivated by using submergedliquid fermentation. Their mycelial biomasses were compared and the intra-mycelialpolysaccharides were further extracted using hot water and ethanol precipitation. Thepolysaccharide and protein contents were measured by using phenol-sulfuric acidmethod and Lowry method, respectively. The result showed that the total averagebiomass of 25 Ganoderma strains was 14.02±3.49 g/L, GR-1 strain produced thehighest biomass (20.52 g/L) while GT-3 strain produced the lowest (7.12g/L).Intra-mycelial polysaccharide yields of 25 strains were in a range of 0.10～0.55g/L infermentation broth. The highest was produced by strain GT-4 (0.55g/L) and the lowest(0.10 g/L) was by GS-2. Protein yields were from 0.05g/L to 0.68g/L and the averagewas at (0.28±0.18) g/L. Protein yield of GL-6 was the highest (0.68g/L) while that ofGT-1 was the lowest (0.10g/L). Strains (GL-1、GL-4、GL-6、GL-7 and GL-8) belongedto Ganoderma lucidum showed a favorable balance among biomass, polysaccharide andprotein yield.The monosaccharide compositions of intra-mycelial polysaccharides wereanalyzed by reverse phase HPLC with a condition capable to simultaneously detect 9monosaccharides. Samples were hydrolyzed by trifluoroacetic acid andmonosaccharides in hydrolyzates was derived with 1-phenyl-3-methyl-5-pyrazolone(PMP). The derivates were eluted with a phosphate buffered acetonitrile gradient on aC18 column and quantified. The results showed that all 25 intra-mycelialpolysaccharides contained glucose, galactose, mannose, glucosamine and fucose, andglucose was the major monosaccharide. Xylose and rhamaose were also detected inpolysaccharides from strain GL-3、GL-5、GL-6、GL-7、GO-3 、GR-3、GR-4、GR-5、GT-1、GT-2、GT-3、GT-5、GS-2 and strain GL-5、GL-6、GL-7、GL-8、GO-3、GR-3、GR-4、GT-2、GT-3、GT-4、GT-5, respectively. Trace amount of galacturonic acidwas detected in strain GL-1、GL-3、GL-8、GR-3、GS-1、GT-1、GT-2、GT-3 、GT-4 、GT-5、GT-6、GT-7 polysaccharides,while no glucuronic acid was found.High performance gel permeation chromatography (HPGPC) analysis showed thatthe weight-mean molecular weights (MW) of all strains polysaccharides ranged from2.03×104 Da to 1.63×105 Da. MW of GR-1 polysaccharide was the biggest and that ofGS-1 was the lowest. Proteins were found to be associated with polysaccharides, and allthe samples tested possessed 3 to 4 peaks. The MW of majority peak was below5×104Da which accounted for 80.26% to 98.84% in total substances.更多还原