【作者】 郭立佳；

【导师】 陶文沂；

【作者基本信息】 江南大学 ， 微生物与生化药学， 2006， 硕士

【摘要】 本文以吸附层析为主,探索研究了适合于从红豆杉粗提物及内生真菌发酵液中分离纯化紫杉醇的技术。传统的液液萃取由于溶剂用量大、溶液易乳化等缺点限制了其在真菌发酵液中分离紫杉醇的应用。本文以树脂层析取代传统的液液萃取步骤,解决了上述不足。从六种树脂中筛选出了吸附性能最好的HZ818大孔吸附树脂,通过单因素及正交试验对工艺进行了优化。通过一步树脂层析可使原料中的紫杉醇同时达到脱色和纯化的双重目的。此步平均回收率在98%以上。以正相层析作为整体工艺的第二步。研究了硅胶及碱性氧化铝两种介质柱的层析效果。通过考察洗脱液组成、洗脱液强度、洗脱速率、高径比等因素对两种柱进行了对比,其中样品经氧化铝层析后,紫杉醇含量有所增加,这主要是由于氧化铝的催化作用,使其它某些紫杉烷类物质转化成了紫杉醇。树脂处理后的样品再经氧化铝层析后纯度达到30%,平均回收率在125%左右。最后一步采用C18反相层析对前两步纯化得到的产品进行精制,通过优化层析洗脱剂组成、洗脱剂强度、洗脱方式以及进料量等操作条件,采用双柱串联的方法,得到纯度在93-94%的紫杉醇,再继以结晶重结晶步骤,除去少量杂质得到纯度在98%左右的紫杉醇晶体。经本工艺分离纯化后的紫杉醇总回收率约为88%。采用波长扫描、高压液相及质谱对制得产品进行鉴定,结果与文献上报道得紫杉醇性质一致。更多还原

【Abstract】 Separation and purification of taxol from Taxus Yunnanensis extract and fermentation liquor of yew endophyte were mainly carried out by adsorption chromatography.Because of large usage of organic solvent and emulsification , traditional liquor-liquor extraction was limited on some degree,especially on separation from fermentation liquor of yew endophyte. This thesis used resin chromatography insteadly as the first step,and HZ818 macroreticular resin was screened from six kind resins,it showed excellent effects of adsorption and separation. Meanwhile, single-factor and orthogonal experiments were carried out to optimize the technological parameters. Through this chromatography process,decoloration and purification were achieved,the even recovery of taxol was more than 98%.Normal-phase chromatography process was established as the second step. Silica gel and Al2O3 were chosen to study. Elutent composition、strength and rate of elution、rate of height and diameter were optimized. Results showed that the content of taxol was greatly increased after Al2O3 chromatography. It might be that some taxol-like taxanes converted to taxol under the catalyzing of Al2O3. Through resin and Al2O3 chromatography,the purity of taxol reached 30%, and the recovery reached about 125%.C18 reversed-phase chromatography was selected as the final step to further purify taxol. By optimization of eluent composition, strength of elution, elution mode and capacity of loading, with two column in series, the purity of taxol can be reached nearly 93-94%, followed with crystallization and recrystallization, the purity can increase to about 98%. Through this whole process, the total recovery of taxol was about 88%.The structure of taxol was identified with IR、HPLC and MS, which was identical with the results that reported on documents.更多还原