【作者】 钟慧敏；

【导师】 王俊儒；

【作者基本信息】 西北农林科技大学 ， 药用植物学， 2010， 硕士

【摘要】 猪苓[Polyporus umbellatus (Pers.)Fries],在分类学上属于无隔担子菌亚纲、无褶菌目、多孔菌科(Polyporaceae)、多孔菌属(Polyporus)的药用真菌,它的干燥菌核即为猪苓,属于常用中药。猪苓具有利尿、抗癌、抗菌等药理活性,利水渗湿是其主要药理活性。猪苓甾体含量仅次于猪苓多糖,是猪苓的另一主要化学成分,具有诸多生物活性,且大多数无毒副作用,在医药工业上具有广泛的用途和良好的发展前景。此外,猪苓甾体可以作为猪苓药材质量评价指标。因此,猪苓甾体是一种很有开发价值的药用资源,同时猪苓甾体的成功开发也为猪苓资源的充分利用作出贡献甾醇是甾体生物合成途径中的一个重要环节,甾醇类化合物是甾体合成的中间体,在猪苓发酵液中添加甾醇,促进猪苓细胞内甾体合成相关的酶的产生,从而促进甾体的合成,增加猪苓甾体的产量。本课题选用β-谷甾醇,豆甾醇和胆固醇三种不同甾醇及其混合物作为诱导物,在植物甾醇最佳添加浓度范围内(0.05‰~0.20‰),选择三个浓度梯度,添加在猪苓液体发酵培养基中,并通过加入β-环糊精和超声增加甾醇诱导物的溶解度;于23℃悬浮振荡暗培养10d后,检测干燥猪苓菌丝体和发酵液中甾体积累量的变化,从而筛选出最佳诱导物,及其最佳添加浓度。通过实验,获得结果如下:(1)单一诱导物中,β-谷甾醇和胆固醇对猪苓菌丝体的生长都具有促进作用,当β-谷甾醇的浓度为0.020%时,菌丝体的生长状况最好,菌丝体总数目最多,浓度为0.015%时,甾体的含量最高;胆固醇的最佳浓度为0.020%,菌丝体的干重最多,甾体净增长量也是最高的。豆甾醇培养基无论是在菌丝体干重还是甾体的净增长量上,就都不及对照组。三者相比,胆固醇效果最好,菌丝体干重达到最大的浓度和甾体净增长量最高的浓度是一致的。胆固醇浓度为0.020%时,菌丝体甾体含量最大(53.224mg),是三者中诱导效果最好。(2)混合诱导物对猪苓甾体含量和猪苓菌丝体干重的影响混合诱导物对猪苓菌丝体的生长具有促进作用,菌丝体的总量的已增加,甾体的量都得到不同程度的提高。在菌丝体生长状况方面,E组(β-谷甾醇+胆固醇)在当甾醇浓度达到0.005%时,菌丝体干重最高。含有胆固醇的组(E组、F组G组)猪苓甾醇的净增长量明显高于不含胆固醇的组(D组);除了浓度为0.020%时,甾体净增长量低于其他三组,E组(β-谷甾醇+胆固醇)在其他两个浓度都高于其他组,且当浓度为0.005%时,甾体净增量达到最高(97.338),因此可以说β-谷甾醇与胆固醇混合,是混合诱导物中诱导效果最佳的。通过研究表明,混合诱导物对猪苓甾体的诱导效果强于单一诱导物对猪苓甾体诱导,胆固醇的诱导效果强于β-谷甾醇和胆固醇,β-谷甾醇与胆固醇的混合物诱导效果最好,当浓度为0.005%,猪苓甾体的净增长量最大,达到97.338mg。猪苓甾体对研究猪苓药用价值,充分利用猪苓资源,甾体药物的合成都具有很重要的意义,所以提高猪苓甾体的含量是很有必要的。更多还原

【Abstract】 Polyporus [Polyporus umbellatus (Pers.) Fries] is a medicinal Fungi ,which belongs to holobasidi- omycetidae,Aphyllophorales, Polyporaceae, Polyporus in the taxonomy.it shall be Polyporus dry sclerotia, are commonly used in Chinese medicine.Modern medicine study indicated that Polyporus umbellatus Fries have many pharmacological activities,for example, diuresis, anti-imflammation; anti-tumor, anti-radiation and enhancing immune function and so on,which has a good prosperity in apply and exploit in fulture. Therefore, polyporus steroid is a kind of valuable medicinal resources and beneficial to make full use of polyporus resource.steroids is biosynthetic intermediate in steroid synthetic. It can promote enzyme related to steroid synthetic product in cell of Polyporus.This topic selectedβ-sitosterol, stigmasterol and cholesterol and their mixtures as inducer, add inducers in the range of (0.05‰~ 0.20‰) to liquid fermented medium, by addβ-CD and ultrasound for improving inducer solubility.23℃suspended oscillating culture.Then testing steroid accumulation of Polyporus mycelium and broth. furthemore, three kinds of mixtures of sterol and its optimal concentration, which also compared the induction of sterol best.The results as follows:(1) single inducerβ-sitosterol and cholesterol promote the growth of mycelium of Polyporus umbellatus when theβ-sitosterol concentration is 0.020%, the best growth conditions of mycelium, concentration is 0.015%, the highest content of steroids; when cholesterol concentration is 0.020% dry weight of mycelium is biggest, the net increase amount of steroid is the highest. Stigmasterol can not promote Polyporus steroid accumulation.In a word,when cholesterol concentration is 0.020%, the largest concentration of steroids mycelium (53.224mg), so it is the best effect among the three.(2) Mixed inducerTests show that the hybrid inducer promote on the growth of mycelium of Polyporus .The amount of steroids were increased to varying degrees. Status in the growth of mycelium,when concentration of inducer as 0.005%, E group (β-sitosterol + cholesterol) dry weight of mycelium is the highest. Containing cholesterol group (E group, F Group G Group) Polyporus sterol was significantly higher than the net increase in non-cholesterol group (D group); addition to the concentration of 0.020%, the net increase of steroids was less than the other three groups,E group(β-sitosterol+cholesterol) in the other two levels are higher than the other group,and a concentration of 0.005%,the highest net increase steroid(97.338),it can be said thatβ–sitosterol and cholesterol mixed is the best inducer.The resule show that the mixed inducer was better than a single inducer .The induction effect of cholesterol is stronger thanβ-sitosterol and cholesterol,β-sitosterol and cholesterol mixture induction the best, when the concentration of 0.005%, Polyporus steroidal largest net increase, reaching 97.338mg. Polyporus steroidal medicinal value of research, make full use of Polyporus resources, the synthesis of steroidal drugs have very important meaning, so to improve the content of Polyporus steroid is necessary.更多还原