基于Maxent模型-DNA条形码-HPLC的西藏沙棘叶质量分析研究

发布时间：2018-03-08 04:06

  本文选题：西藏沙棘叶　切入点：生态适宜性　出处：《成都中医药大学》2016年硕士论文　论文类型：学位论文

【摘要】：藏医药历史悠久,理论体系完整,有其独特的疗效作用,使用人群众多,是具有代表性的传统民族医药之一。沙棘为典型多基原藏药,据《晶珠本草》记载藏药沙棘分为大、中、小3种,小沙棘即为西藏沙棘Hippophae thibetana Schlechtend(藏文名“萨达尔”)。西藏沙棘是青藏高原上分布较广的沙棘种类,且其根系较为发达,在高原地区有着极为重要的保持水土作用,具有重要的生态价值。西藏沙棘果已被收录于《四川省藏药材标准》(2014年版),是沙棘膏主要来源之一,也可单独应用,具有“滋补肝阴的功效”。已有研究发现沙棘叶中也富含黄酮类及其他营养成分,且沙棘叶资源丰富便于采集,是药品、保健食品、化妆品及饲料等产品的原料,具有重要的经济价值。《四川省藏药材标准》(2014版)收载了沙棘叶(中国沙棘叶、云南沙棘叶)的质量标准。但目前,对西藏沙棘叶的相关研究较少,为充分利用西藏沙棘资源,本文考察了西藏沙棘资源分布及生态适宜性特点；对西藏沙棘叶进行品种鉴定；对西藏沙棘叶中黄酮类成分进行研究；并建立西藏沙棘叶的质量评价方法,保证了其临床用药的有效性和药用资源的可持续性,为西藏沙棘资源开发利用提供依据。目的：1.调查西藏沙棘资源分布和生态适宜性特点,为西藏沙棘资源利用提供依据。2.鉴别西藏沙棘的真伪。3.对西藏沙棘叶中黄酮类成分进行分离纯化,建立西藏沙棘叶TLC鉴别、HPLC含量测定方法,初步建立西藏沙棘叶质量标准。方法：1.基于Maxent生态位模型及GIS研究西藏沙棘H. thibetana Schlechtend,在我国的地理分布,以及影响西藏沙棘地理分布的主要生态因子特征；对西藏沙棘潜在分布区域进行了生长适宜性等级划分。2.采用DNA条形码技术,对西藏沙棘进行基原鉴定。3.采用大孔吸附树脂、Sephadex LH-20.聚酰胺等柱色谱法对西藏沙棘叶乙醇提取物分离纯化,结合1H-NMR、13C-NMR、MS等波谱技术确定化合物的结构。以槲皮素、山奈素、异鼠李素为对照品,建立西藏沙棘叶的TLC鉴别和HPLC含量测定方法,同时对西藏沙棘叶药材的性状、检查项、浸出物等进行研究。结果：1.采用人工(Manual)分级方法分析,划分出各生态因子图层相应的西藏沙棘适宜性分布区域,结果表明西藏沙棘分布在青藏高原的高寒草原区域,主要受7月、8月、12月降水量、2月气温等影响。在我国适宜区面积47.778万平方千米,较适宜区面积为47.55万平方千米。2.ITS2序列能成功鉴别西藏沙棘与混伪品。3.从西藏沙棘叶乙醇提取物中分离得到11个黄酮类化合物,分别为：槲皮素-3-O-β-D-葡萄糖-7-O-α-L-鼠李糖苷(1)；异鼠李素-3-O-β-D-半乳糖-7-O-α-L-鼠李糖苷(2)；异鼠李素-3-O-β-D-葡萄糖-7-O-α-L-鼠李糖苷(3)：异鼠李素-3-O-α-L-阿拉伯吡喃糖-7-O-α-L-鼠李糖苷(4)；水仙苷(5);槲皮苷(6)；异鼠李-3-O-β-D-葡萄糖苷(7)；异鼠李素-7-O-a-L-鼠李糖苷(8)：异鼠李素(9)；山奈素(10)；槲皮素(11)。4.确定了西藏沙棘叶性状、TLC鉴别方法；规定了西藏沙棘叶检查项的限度,即水分不得超过11.0%,总灰分不得超过5.0%,酸不溶性灰分不得超过0.4%,醇溶性的浸出物不得少于14.0%；测定了西藏沙棘叶中黄酮类含量,槲皮素含量在0.092%～0.282%之间,平均值为0.183%；异鼠李素含量在0.225%～0.666%之间,平均值为0.388%；山奈素含量在0.080～0.324%之间,平均值为0.195%。以干燥品计算,槲皮素不得少于0.146%、异鼠李素不得少于0.310%、山奈素不得少于0.156%。结论：1.西藏沙棘的资源分布主要在青藏高原的高寒草原区域,生长密度较大。生态因子中海拔及温度季节性变化标准差对西藏沙棘的生长适宜度影响最大。2.ITS2序列能准确快速鉴别西藏沙棘叶,为西藏沙棘叶的鉴别提供新方法。3.研究了西藏沙棘叶中黄酮类化学成分。4.初步建立西藏沙棘叶的质量控制方法。
[Abstract]:Tibetan medicine has a long history, a complete theoretical system, has its unique curative effect, the use of the masses more than the traditional ethnic medicine is one of the representative. As a typical multi Seabuckthorn original Tibetan medicine, according to the < jingzhubencao > record of seabuckthorn, divided into 3 kinds, small, small sea buckthorn is Hippophae thibetana of seabuckthorn in Tibet Schlechtend (Tibetan name "Sardar"). Tibet is widely distributed species of sea buckthorn Hippophae on Qinghai Tibet Plateau, and its roots are more developed, has a very important role in soil and water conservation in the plateau region, has important ecological value. Tibet has been included in the sea backthern fruit in Sichuan Province, Tibetan medicine standard < > (2014 version), is one of the main sources of sea buckthorn cream, can also be used alone, with the efficacy of nourishing liver yin ". Studies have found that sea buckthorn leaves are rich in flavonoids and other nutrients, and sea buckthorn leaves rich resources to collect, medicine, health care Food, cosmetics and feed products such as raw materials, has an important economic value in Sichuan province. The Tibetan medicine standard "(2014 Edition) containing the seabuckthorn leaves (Chinese seabuckthorn leaf, Yunnan seabuckthorn leaf) quality standards. But at present, the relevant research on Tibet sea buckthorn leaves less, to make full use of Tibet seabuckthorn resources. This paper examines the Tibet sea buckthorn resources distribution and ecological suitability characteristics; species identification of Tibet seabuckthorn leaf; Study on Flavonoids of seabuckthorn leaves in Tibet; and establish the quality evaluation method of Tibet seabuckthorn leaves, to ensure the sustainability and effectiveness of the clinical use of medicinal resources, provide the basis for the development and utilization of seabuckthorn resources Tibet. Objective: 1. to investigate the Tibet characteristics of seabuckthorn resources distribution and ecological suitability, provide the basis for the identification of.2..3. on the authenticity of Tibet Seabuckthorn flavonoids from Tibet seabuckthorn leaves for Tibet seabuckthorn resources utilization Separation and purification, the establishment of Tibet sea buckthorn leaves TLC identification, HPLC method for the determination of content, the initial establishment of Tibet seabuckthorn leaf quality standard. Methods: 1. based on the Maxent model and the niche of seabuckthorn in Tibet H. thibetana Schlechtend GIS, geographic distribution in China, and the main ecological factors influencing the characteristics of geographical distribution of seabuckthorn in Tibet; the Tibet sea buckthorn potential the regional distribution of growth suitability grades of.2. using DNA barcode technology based on Tibet sea buckthorn identification by.3. macroporous resin, polyamide and Sephadex LH-20. column chromatography of ethanol extracts from leaves of seabuckthorn in Tibet purification, combined with 1H-NMR, 13C-NMR, MS spectroscopy to determine the structure of compounds. With quercetin, Yama Nasu. Isorhamnetin as reference, to establish a method for determination of TLC identification and HPLC content of Tibet seabuckthorn leaves, while the Tibet sea buckthorn leaf medicinal properties. Check and study extract. Results: 1. using artificial classification method (Manual) analysis, divides the regional distribution of the ecological factors of seabuckthorn in Tibet proper layer corresponding, the results show that the Tibet sea buckthorn distribution in alpine steppe regions of the Tibetan Plateau, mainly by July, August, December, precipitation, influence of temperature in February. In China the suitable area of 477 thousand and 780 square kilometers, the suitable area is 475 thousand and 500 square kilometers.2.ITS2 sequence can get 11 flavonoids isolated from ethanol extracts from leaves of seabuckthorn in Tibet in the successful identification of Tibet Seabuckthorn and adulterants.3. respectively: Quercetin -3-O- beta -D- glucose -7-O- alpha -L- rhamnoside (1); isorhamnetin -3-O- beta -D- galactose -7-O- alpha -L- rhamnoside (2); isorhamnetin -3-O- beta -D- glucose -7-O- alpha -L- rhamnoside (3): Isorhmnetin -3-O- -L- alpha -7-O- alpha -L- in Arabia pyranoses Li Tanggan (4); water Xian glycosides (5); quercitrin (6); isorhamnetin -3-O- beta -D- glucoside (7); isorhamnetin -7-O-a-L- rhamnoside (8): Isorhmnetin (9); kaempferol quercetin (10); (11).4. to determine the Tibet sea buckthorn leaf traits, the identification method of TLC; the provisions of the Tibet sea buckthorn leaves check limit, namely water shall not exceed 11% of the total ash not more than 5%, acid insoluble ash shall not exceed 0.4%, alcohol soluble extract of not less than 14%; the content of flavonoid in leaves of seabuckthorn in Tibet were determined, the content of quercetin in 0.092% ~ 0.282%, the average value of 0.183%; isorhamnetin the content in 0.225% ~ 0.666%, the average value is 0.388%; kaempferol content in 0.080 ~ 0.324% between the average value of 0.195%. calculated on dry goods, quercetin Isorhmnetin shall not be less than 0.146%, not less than 0.310%, not less than 0.156%. of kaempferol: 1. Tibet sea buckthorn resources distribution 鍦ㄩ潚钘忛珮鍘熺殑楂樺瘨鑽夊師鍖哄煙,鐢熼暱瀵嗗害杈冨ぇ.鐢熸，

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