【作者】 陈仁玉；

【导师】 刘朝贵；

【作者基本信息】 西南大学 ， 微生物学， 2012， 硕士

【摘要】 多糖是一类重要的生物活性物质,在人类及动植物医疗保健、动物养殖、病虫害防治、农作物栽培等领域有十分广阔的应用前景。现代医学研究发现,食用菌中能显著增强癌症患者抵抗力的生理活性物质即为食用菌多糖,并且其毒性很小,易与菌体分离。与植物多糖相比,食药用菌中的多糖生产周期短,产量及质量稳定,性价比较高,具有免疫调节、抗病毒、抗肿瘤、抗氧化、降低血脂、抗血栓等其它多糖不具备的特殊功效。花脸蘑作为一种还没有大规模人工栽培的食用菌,其子实体含有丰富的氨基酸和糖类等营养物质,而且矿质元素种类十分丰富。据报道,花脸蘑子实体中多糖含量为10.31%,海藻糖的含量为8.34%,其海藻糖含量高于常见的食用菌。本试验以花脸蘑为载体,研究了胞内多糖和海藻糖在花脸蘑中积累的规律及酶活性变化规律,为今后综合开发利用食用菌的多糖和海藻糖资源作了初步的探索,取得了有价值的结果。另外对花脸蘑胞内多糖的理化性质以及单糖组分进行了研究,这些都为工业化生产花脸蘑胞内多糖,以及开发花脸蘑多糖系列产品提供了一定的理论基础及帮助。试验结果如下：①花脸蘑液体深层发酵产胞内多糖的优水平组合为：玉米粉20g,酵母膏4g,pH=7,CuS04的浓度为1.5×10-4g/100mL,1000 mL水,菌龄为12d。其干生物量可达到1.13g/100ml,湿菌丝体中胞内多糖含量达24.72%,高于其子实体中多糖的含量。②当N含量高时,有利于花脸蘑菌丝的生长,菌丝体得率高；而随着C含量升高时,有利于胞内多糖的积累。但是,单糖并不利于花脸蘑菌丝生长,其菌丝利用玉米粉的效率较高。当玉米粉质量浓度增加到25g/L后,由于玉米粉溶解性差、粘度大,造成了培养基溶氧量的下降,使菌丝生长受到了阻碍,胞内多糖产量也开始下降。因此,碳源浓度是影响花脸蘑菌丝体胞内多糖产量的决定因子③同灵芝、灰树花、姬松茸等其它食用菌一样,花脸蘑菌丝对有机氮源的利用优于无机氮源。pH值与微量元素对花脸蘑菌丝的生长及胞内多糖的积累影响较小。菌龄以8d与12d较好。④花脸蘑菌丝体胞内多糖呈白色粉末状,无气味,溶于水,不溶于乙醚、丙酮、95%的乙醇等有机溶剂。且为非还原性多糖,不含有蛋白质成分。其单糖组分为葡萄糖、木糖、甘露糖,并且是以葡萄糖为主组成的多糖。⑤花脸蘑菌丝体海藻糖液体发酵的优水平组合为：蔗糖20g,酵母膏4g,pH=7,CuSO4为1.5×10-4g/100mL,1000 mL水,菌龄为12d。湿菌丝体中海藻糖的含量为19.02%,明显高于子实体体中海藻糖的含量。⑥单糖并不利于花脸蘑菌丝体海藻糖的液体发酵,以葡萄糖作为碳源进行液体发酵时,其海藻糖产量最低。菌丝体利用有机氮源中的酵母膏、蛋白胨来液体发酵时,其菌丝体内的海藻糖含量比利用无机氮源作为氮源时要高。微量元素对液体发酵产海藻糖的影响较小。影响花脸蘑菌丝体海藻糖产量的决定因子是菌龄。⑦花脸蘑菌丝体胞内多糖和海藻糖得率最高均在第12天,即在稳定期同时达到了最高峰。据此,可以在稳定期提取胞内多糖的同时也提取海藻糖,从而实现对花脸蘑菌丝体的综合利用。⑧不同碳源、氮源、菌龄对花脸蘑胞外酶活性的大小有一定的影响,但是并没有改变酶活性的变化规律。花脸蘑菌丝对有机碳源、有机氮源更容易吸收利用,相应的酶活性也会相应增加,这与菌丝体生物量及糖得率有一定的相关性。而pH值、微量元素对酶活性的影响比较小。更多还原

【Abstract】 Polysaccharide is an important class of biologically active substances which has very broad application prospects in the field of human, animal and plant health care, animal breeding, pest control, crop cultivation. Modern medical researchs have found that mushroom polysaccharide, the physiologically active substances in edible fungi and easily separated from mushroom, can significantly enhance the patients’ resistance to cancer and whose toxicity is very small. Compared with plant polysaccharides, the polysaccharides in edible and medicinal fungi has the features, such as short production cycle, the stable yield and quality, higher performance to price ratio and some special effects-immunomodulatory, antiviral, antitumor, antioxidant, reduce blood lipids, anti-thrombosis while other polysaccharides don’t have.Lepista sordida is not large-scale artificially cultivated yet, whose fruiting body is rich in nutrients such as amino acids and carbohydrates, and the species of mineral composition.According to reports, the contents of trehalose in Lepista sordida fruiting body is 8.34%, more than other common edible mushrooms apparently. This test studied the law of accumulation of Lepista sordida’s intracellular polysaccharide and trehalose and the change of enzyme activity, and made a preliminary exploration for the future comprehensive development and utilization of edible fungus polysaccharides and trehalose resources. In addition, this test studied the physicochemical properties and the monosaccharide composition of intracellular polysaccharide.These reseach provided a theoretical basis and help for industrial production Lepista sordida’s intracellular polysaccharide and development of products of Lepista sordida’s polysaccharide.The test result are as follows:①The intracellular polysaccharide’s deep fermentation liquid optimal level combination was Sucrose 20g, yeast extract 4g, pH=7, the concentration of CuSO4 1.5 ×10-4g/100mL, water 1000 mL, strain age 12d. The dry biomass can be achieved 1.13g/100ml and the content of intracellular polysaccharide in wet mycelium is 24.72%, wich is higher than the fruiting bodies.②When the N content is high, It is conducive to the growth of the mycelium. With the rise of the C content, It is beneficial to the accumulation of intracellular polysaccharide. Monosaccharide is not conducive to the growth of mycelium, and Mycelium can be a good use of corn flour. When the mass concentration of corn flour increased to 25g/L, The intracellular polysaccharide production began to decline. It is because of Corn flour’s low solubility and high viscosity resulting in a decline in dissolved oxygen in medium, mycelial growth has been hampered. The carbon source is the decisive factor of the intracellular polysaccharide production in Lepista sordida mycelium.③As with other edible fungi, such as Ganoderma lucidum, Grifola frondosa, Agaricus blazei Murill, its mycelium can make better use of organic nitrogen than inorganic nitrogen. Trace element and pH value on mycelial growth and the Intracellular polysaccharide Accumulations have a smaller impact. It was better that the strain age was 8d to 12d.④The pure Intracellular polysaccharide of Lepista sordida is White powdered, Odorless, Soluble in water but not in organic solvent, such as ether and acetone,95% ethanol etc. It’s a kind of non-reducing polysaccharide and does not contain a protein component. The monosaccharides group is divided into glucose, xylose and mannose. And the main Composition is glucose.⑤The Lepista sordida mycelium trehalose liquid fermentation excellent level combination was sucrose 20g, yeast extract 4g, pH=7, the final concentration of CUSO4 1.5×10-4g/100mL, water 1000mL, strain age 12d.Trehalose content in the wet mycelium was 19.02%, significantly higher than the content of trehalose in the body.⑥Monosaccharide is not conducive to the Lepista sordida mycelium trehalose liquid fermentation. Using glucose as the carbon source for liquid fermentation resulted in that trehalose content was the lowest. The trehalose content of the mycelial body was higher when The Mycelium using organic nitrogen sources-yeast extract, peptone-to liquid fermentation than using inorganic nitrogen as a nitrogen source. trace elements had Less impact on liquid fermentation of trehalose. The strain age is the decisive factor of the trehalose production in Lepista sordida mycelium.⑦The Maximum yield of intracellular trehalose and polysaccharide in Lepista sordida mycelia appeared in the 12d, that is to say in stationary phase, the yield be maximized. Accordingly, we can extract both trehalose and polysaccharide in stationary phase, so as to achieve the comprehensive utilization of Lepista sordida mycelia.⑧Different carbon sources, nitrogen source, fungus age have certain effect on extracellular enzyme activity, but do not change the change rule of enzyme activity. Hyphae of Lepista sordida would be easier to absorb and utilize organic carbon sources, organic nitrogen source, the corresponding enzyme activity also can increase. It can be explained by that mycelia biomass and sugar yield have a certain correlation. But pH value, trace elements have little influence on enzyme activity.更多还原