豆类丝核菌及其次级代谢产物有效成分的分离研究

发布时间：2018-06-14 13:44

  本文选题：豆类丝核菌 + 培养液　； 参考：《西北农林科技大学》2006年硕士论文

【摘要】： 目的:旨在探索豆类丝核菌菌丝体及其次级代谢产物的成分,分析其各成分的用途,为开拓豆类丝核菌的新用途奠定理论基础。 方法:1.定期用马铃薯葡萄糖琼脂培养基(PDA)移接豆类丝核菌,使之保持旺盛的生长力。2.将特定的豆类丝核菌菌株用改良的Czapek’s培养基进行培养,获得豆类丝核菌培养液和豆类丝核菌菌丝体。3.用不同的方法对菌丝体和次级代谢产物分别进行处理。①对豆类丝核菌菌丝体的处理:将自然干燥的豆类丝核菌菌丝体用0.5%的盐酸进行浸泡24h后应用40KHz超声波清洗器在70℃处理30min,收集提取液,如此重复三次,合并提取液。将提取液在60℃条件下减压浓缩,备用。用2mol/L盐酸调提取液为pH 2,加入新鲜配制的雷氏盐饱和水溶液,有沉淀生成,滤集生成的沉淀,并将沉淀溶于丙酮,滤除不溶物,向丙酮溶液中加入硫酸银饱和水溶液,使沉淀分解。过滤后向滤液中加入一定量氯化钡溶液,过滤,滤液用2mol/L氨水将之调pH 10,挥干得到褐色粘稠状带有特异性香味的物质。将其在90℃下减压升华得到白色晶体。②对豆类丝核菌培养液的处理:方法一,取1000ml预处理完备的豆类丝核菌培养液,在60℃减压浓缩,浓缩液调pH 2,用正丁醇萃取10次,得到有机相和酸水液。将酸水液调至pH 10,再用正丁醇萃取10次得到水相和有机相。有机相用0.2mol?L的稀硫酸萃取,得到水相和有机相。将水相用浓氨水碱化后浓缩至干,所得浸膏减压升华得到白色粉末状物质。方法二,取1000ml预处理完备的豆类丝核菌培养液,调pH 10,用H2O:CH2Cl2 (3∶1, v/v )萃取,弃去CH2Cl2层,将水层调pH 10后,再用H2O∶CH2Cl2(3∶1, v/v )萃取,弃去CH2Cl2层,将水层调pH 7,浓缩后上732型强酸性苯乙烯系阳离子交换树脂,先用去离子水冲洗,后用1mol/LNH4OH洗脱,收集氨洗液,调pH 7,浓缩后冻干,得到浅黄色粉末,将之减压升华得到白色粉末状物质。方法三,取1000ml预处理完备的豆类丝核菌培养液,调pH7,上大孔吸附树脂,反复富集5次,静置12h,用滴水冲洗,洗2个柱体积,收集水洗脱液,再用乙醇洗脱,收集约2个柱体积的醇水混合洗脱液,最后收集2个柱体积的醇洗脱液,分别减压浓缩各洗脱段的液体,经TLC检测,合并醇洗液,减压回收乙醇,余液冻干,得到褐色粉末。TLC则可检出,与标准品对照,均呈现淡紫色,但由于纯度较差难以升华。 结果:经过熔点测定仪和GC-MS检测,初步鉴定豆类丝核菌菌丝体中可能含有蒲公英甾醇、α-香树脂醇、大狼毒素以及两种未知的化合物。这几种物质均是首次从豆
[Abstract]:Objective: to explore the composition of the mycelium and its secondary metabolites of Rhizoctonia legume, and to analyze the uses of each component, so as to lay a theoretical foundation for the development of new uses of Rhizoctonia legume. Method 1: 1. The strain of Rhizoctonia legume was transfered on potato glucose Agar medium (PDAs) regularly to maintain its vigorous growth ability. 2. The specific strains of Rhizoctonia legume were cultured on the modified Czapeks medium to obtain the culture medium of Rhizoctonia legume and the mycelia of Rhizoctonia legume. Treatment of mycelium and secondary metabolites by different methods. 1 treatment of mycelium of Rhizoctonia legume: soaking naturally dried Rhizoctonia mycelium with 0.5% hydrochloric acid for 24 hours and then using 40kHz ultrasonic cleaning The scrubber was treated at 70 鈩，

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