【作者】 柏建山；

【导师】 夏立秋；

【作者基本信息】 湖南师范大学 ， 微生物学， 2004， 硕士

【摘要】 蕌头(Allium chinense G.Don)，百合科葱属植物，在传统医学中已经认识到它的消炎和保健的重要作用，但在使用过程中缺乏科学依据，而且目前对蕌头中有效药用成分的研究缺乏定性、定量的分析。随着科学技术的高速发展和人类对健康水平要求的提高，对蕌头等中草药有效成分的分离提纯也必须要达到国际市场的标准和规范，并使之能够进行产业化生产，这样才能增强我国在国际医药市场上的竞争力。 1 藠头抗菌抑癌成分的分离纯化和结构鉴定 藠头经过搅碎、酶解后，其中的有效成分用有机溶剂二氯甲烷萃取，然后浓缩。浓缩后的样品通过Sephadex G-25凝胶柱分离提纯其中的有效成分，以白色念球菌(Candida albican)进行生物活性测定，并用气相色谱-质谱(GC-MS)法对其化学成分进行分离鉴定，GC法测定各种化合物的相对百分含量，共鉴定了9种成分，约占总提取物的82.12％，其中，主要物质是甲基丙烯基二硫化物。 2 藠头抗菌抑癌成分的理化性质 藠头上清液对痢疾志贺菌(Shigella dysenteriae)等细菌具有很好的抗菌活性，对白色念球菌等真菌也有很好的抗菌效果。经过Sephadex G-25凝胶柱分离提纯其中的有效成分，紫外/可见光分光光度计扫描发现有效成分的特征峰出现在237nm附近，以此可以通过吸光值A来进行定量的分析。在抑制肿瘤细胞生长的研究中发现，藠头洗脱液中的有效成分对肝癌细胞(HepG-2)和海拉细胞(Hela)都有强烈的抑制活性，但是藠头洗脱液对HepG-2细胞的抑制效果要好于对Hela细胞的抑制效果，藠头上清液的抑制肿瘤细胞的作用远不如藠头洗脱液。温度对藠头洗脱液中的有效成分具有一定的影响，在50℃具有最大吸收峰，随着温度的升高，其吸收峰逐渐下降，生物活性也逐渐消失。 3 藠头抗菌抑癌成分的生物功能分析 莫头洗脱液处理的HepG一2细胞和Hela细胞经流式细胞仪分析 (FCM)发现，这两种肿瘤细胞并没有发生细胞调亡，处于各个细胞周期的细胞数目有所改变，对其总DNA的电泳分析未发现梯形条带。在体外条件下，测定莫头洗脱液对木瓜蛋白酶，纤维素酶，蛋白酶K的酶活力影响，研究发现低浓度莫头洗脱液使木瓜蛋白酶活力增高，高浓度的慕头洗脱液使木瓜蛋白酶活力降低，但是相同浓度的蓖头洗脱液对纤维素酶，蛋白酶K的酶活力几乎没有什么影响。这说明嘉头洗脱液可能和一些琉基酶起作用，从而具有抗菌抑癌的生物活性。更多还原

【Abstract】 Allium chinense is a kind of plants of Allium , which has important role in inflammation and health care in the traditional medicine. But it is lack of scientific proof, furthermore at present it is lack of analysis of quality and quantity to the effective medical components of Allium chinense. The isolation and purification of effective components of Chinese herb, for example Allium chinense, must accord with the standard of criterion of international markets as the development of science and technique and the improvement of people health level, we hope we can industrialize the product of Allium chinense, so we can increase the competition in the international medicine market.1. The isolation and identification of antibacterial and anticancer components of Allium chinenseThe effective components of Allium chinenses were extracted by Dichloromethane after they were crushed and enzymed. The sample was isolated by sephadex G-25 gel filtration column after concentration. Then bioactivity was assayed by Candida albicans. The chemical components were isolated and identified by GC-MS. GC determined the relative percentage content of the extract, nine kinds of components were separated, identified and accounted for about 82.12%, the major compound was propenyl methyl disulfide.2. The physical and chemical character of antibacterial and anticancer components of Allium chinenseAllium chinenses had antibacterial property to Shigella dysenteriae and Candida albicans. We utilized Uv/vis spectrophotometer to scan the sample after the effective componets were purified by sephadex G-25 gel filtration column, we found that the specific peak was near 237nm. So we could determine the quantity of the sample by absorbance A. Theeffective componets from elution solution of A Ilium chinenses had strong inhibition to HepG-2 cells and Hela cells, but the elution solution had better inhibition to HepG-2 cells than Hela cells. The anticancer effect of supernate was much lower than the elution solution. The temperature had influence on the effective components of elution solution, we found the maximal absorbance peak at 50℃, then the absorbance peak droped with the rise of temperature, the bioactivity disappeared at the same time. 3. Analyzing of biological function of antibacterial and anticancer components of Allium chinenseFCM analyzed the HepG-2 cells and Hela cells which were treated by the elution solution of Allium chinenses. The two kinds of cancer cells did not induce apoptosis after they were treated by elution solution, but the cells changed their cell cycle distribution. We did not find the DNA ladder in the electrophoresis of total DNA. Then we determined the influence on the enzyme ativity of papain, cellulase and proteinase K treated by elution solution of Allium chinenses. We found the enzyme activity of papain was improved by low concentration and inhibited by high concentration of elution solution, but the same concentration had no influence on the enzyme activity of cellulase and proteinase K. All the things proved that the elution solution of Allium chinenses maybe regulated enzymatic activity of SH containing enzymes by thiol-disulfide exchange reactions.更多还原