二维分离纯化技术在高山物种研究中的应用

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	二维分离纯化技术在高山物种研究中的应用
	党军
学位类型	博士
导师	陶燕铎 ; 王启兰
	2014-05
学位授予单位	中国科学院研究生院
学位授予地点	北京
学位专业	植物学
关键词	高山物种二维分离纯化技术样品前处理制备结构鉴定
摘要	高山环境主要具有低温、缺氧、日照时数长、紫外辐射强、昼夜温差大、物种生长季节短等特点。高山物种由于其生境的特殊性，以其独特的生理生化机制来应答极端环境，在与极端环境的长期对抗中产生出独特的初级、次级代谢产物，这些独特的代谢产物是天然药物的潜在来源之一。近年来，二维液相色谱分离纯化技术是天然产物活性成分分离纯化技术中极具潜力的新技术之一，具有高选择性、普适性以及节约能源等显著的优越性。本研究将从物质基础到目标制备系统阐述二维分离纯化技术在高山物种化学或活性化学成分研究中的应用。研究中选取唐古特虎耳草、麻花艽、枸杞叶以及黄绿蜜环菌子实体四种典型高山物种为研究对象，开展了以下方面的研究： ⑴系统阐述唐古特虎耳草中活性成分的二维分离纯化方法，包括样品前处理、二维分离纯化方法学的建立以及活性化合物的反相/反相二维分离纯化。 ⑵响应面法优化麻花艽中龙胆苦苷的提取工艺以及建立麻花艽中龙胆苦苷二维分离纯化工艺，通过反相/亲水二维分离纯化，实现了龙胆苦苷的克级制备。 ⑶正交实验法优化枸杞叶中甜菜碱的提取工艺以及建立枸杞叶中甜菜碱二维分离纯化工艺。通过缓冲盐反相/反相二维分离纯化体系，建立了枸杞叶中甜菜碱的分离纯化工艺。 ⑷通过正相/亲水二维分离纯化结合实时无标记动态细胞分析技术体外抗肝癌活性筛选跟踪，从黄绿蜜环菌子实体丙酮提取物中分离纯化出三个具有体外抑制肝癌细胞增殖活性的甾醇类化合物。研究结果表明，二维分离纯化技术极其适用于高山物种化学成分以及目标化合物的分离纯化。通过发展合适的样品前处理方法，建立具有良好正交性的二维分离纯化体系，可以实现高效的对高山物种物质基础研究以及对特定物种中目标化合物的规模化制备。
其他摘要	Alpine environment are characterized by low temperature, anoxia, long sunshine hours, intense UV radiation, large temperature difference between day and night, and short growing periods of species. As the special living environment, high mountain species possess unique physiological and biochemical mechanism for adapting to extreme environments. Alpine species produce unique primary and secondary metabolites during the long confrontation with extreme environments, and these unique metabolites were one of the potential sources of natural medicines. Two-dimensional liquid chromatography separation and purification technique was one of the promising new technologies in the isolation and purification of natural active constituents in recent years, which possesses the superiority of high selectivity, universality and saving energy. This research will systematically elaborate the application of two-dimensional separation and purification technique in alpine species active constituent isolation and purification from material basis research to targeted preparation research. Saxifraga tangutica Engl., Gentiana Straminea Maxim., Lycium barbarum leaves and fruit body of Armillaria luteo-virens were selected as the objects of study, and conducted the parts of research as following: (1) Systematically expounding the two-dimensional separation and purification method of the active ingredients in Saxifraga tangutica Engl., including sample pretreatment, establishing two-dimensional separation and purification methodology and preparing the active compounds by the reversed phase × reversed phase two-dimensional separation and purification technology. (2) Optimizing the extraction process of gentiopicroside from Gentiana Straminea Maxim. by response surface methodology and establishing two-dimensional separation and purification technology of gentiopicroside from Gentiana Straminea Maxim., finally, achieving gram-scale preparation through reversed phase × hydrophilic two-dimensional separation and purification technology. (3) Optimizing the extraction process of betaine from Lycium barbarum leaves the orthogonal experimental method and establishing two-dimensional separation and purification technology of betaine from Lycium barbarum leaves. By adding buffered salt system reversed phase × reversed phase two-dimensional separation and purification technology, establishing the isolation and purification process of betaine from Lycium barbarum leaves. (4) Normal phase × hydrophilic two-dimensional separation and purification combined with real time cellular analysis liver cancer activity screening in vitro, successfully preparing three active sterols from the acetone extract from fruit body of Armillaria luteo-virens. The research result showed that two-dimensional separation and purification technology was extremely suitable to isolation and purification research of alpine species chemical composition and targeted compounds. By developing appropriate sample pretreatment method combined with establishing good orthogonal two-dimensional separation and purification system, which can realize efficient to study the chemical composition of alpine species and prepare targeted compound large-scale from given species.
文献类型	学位论文
条目标识符	http://210.75.249.4/handle/363003/4008
专题	中国科学院西北高原生物研究所
推荐引用方式 GB/T 7714	党军. 二维分离纯化技术在高山物种研究中的应用[D]. 北京. 中国科学院研究生院,2014.

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