【作者】 张万全；

【导师】 陈庆富； 杨庆雄；

【作者基本信息】 贵州师范大学 ， 植物学， 2009， 硕士

【摘要】 以原产贵州、四川等地的淫羊藿属（Epimedium L.）13个物种43个不同地方居群植物为材料,对淫羊藿属植物的总黄酮含量、黄酮组分含量、黄酮化学指纹、黄酮提取工艺、黄酮单体分离及其与牛血清白蛋白相互作用活性等进行了研究,目的是为淫羊藿属植物中药现代化研究提供指导。所得主要结果如下:（1）以紫外分光光度法测定了生长在相似条件下的淫羊藿属11种植物32群247个植株的总黄酮含量。11种淫羊藿的平均总黄酮含量是6.994%,种间变异大,变异系数为44.581%。平均总黄酮含量超过10%的淫羊藿仅1种,含量在5-10%的有6种,含量低于5%的有4种。研究还发现,粗毛淫羊藿和柔毛淫羊藿不同季节之间总黄酮含量有显著差异,且表现出相同的变化规律:9月＞7月＞3月＞5月。（2）考虑提取次数、乙醇含量、溶剂用量（料液比）、pH值四个因素,研究了粗毛淫羊藿的冷浸提取工艺。结果表明,影响提取效果的首要因素是提取次数。以总黄酮的获得量为标准,最佳工艺条件是15倍量80%乙醇不加碱提取2次。以淫羊藿苷获得量为标准,最佳工艺条件是:20倍量60%乙醇不加碱提取2次。（3）通过硅胶柱层析和葡聚糖凝胶柱层析,从粗毛淫羊藿提取物中分离出6个黄酮成分,经ESI-MS,¹H NMR和¹³C NMR谱鉴定,它们是:淫羊藿苷,淫羊藿次苷Ⅱ,2″-鼠李糖基淫羊藿次苷Ⅱ,大花淫羊藿苷B,脱水淫羊藿素,4″-鼠李糖基淫羊藿次苷Ⅱ。（4）用RP-HPLC法测定了生长在相似条件下的淫羊藿属13种植物40个植株的淫羊藿苷、淫羊藿次苷Ⅱ、2″-鼠李糖基淫羊藿次苷Ⅱ、大花淫羊藿苷B、脱水淫羊藿素、4″-鼠李糖基淫羊藿次苷Ⅱ等6种黄酮成分的含量。淫羊藿苷的含量普遍较高,其它5种黄酮成分的含量普遍很低。以各种植物的6种黄酮成分的平均含量为标准用最短距离法进行聚类分析,可把13种淫羊藿分为5类。（5）对10批次不同来源的黔产粗毛淫羊藿的HPLC图谱进行分析,得到共有峰10个。5号峰为淫羊藿苷,7号峰为4″-鼠李糖基淫羊藿次苷Ⅱ,8号峰为淫羊藿次苷Ⅱ。2、3、4号峰,不能直接指认,根据文献分析,极有可能是朝藿定A、朝藿定B、朝藿定C。1、6、9、10号峰为未知峰。（6）在室温下,用淫羊藿苷、大花淫羊藿苷B、4″-鼠李糖基淫羊藿次苷Ⅱ和淫羊藿次苷Ⅱ等四种黄酮成分分别对牛血清白蛋白溶液进行荧光滴定实验。结果发现这4种黄酮成分都能牛血清白蛋白猝灭的荧光。更多还原

【Abstract】 The contents of total flavone and flavonoid monomers in 43 accessions of 13 barrenwort species of genus Epimedium L.,and their chemical fingerprinting,extracting technics and interactive effects with bovine serum albumin were studied,in order to provide some new clues for Chinese medicine modernization.The main results are as follows:（1） The ultraviolet spectrophotometric analysis showed that the average flavone content of 247 plants of 32 accessions in 11 barrenwort species was 6.994%,with the variation coefficient of 44.581%.Among them,E.ecalcaratum has the highest average flavone content（13.166%）.E.pubescense,E.pauciflorum,E.wushanense,E. acuminatum,E.chlorandrum and E.myrianthum have average flavone content of 5-10%, and E.pudingense,E.leptorrhizum,E.luodianense,and E.lishihchenii the low average flavone content（less than 5%）.This study also showed that there are significant differences of flavone content of E.pubescense and E.acuminatum among different seasons:September＞July＞March＞May.（2） The extraction technics of flavones in E.acuminatum leaves under room temperature has been studied according to the orthogonal design involving in four factors （extracting times,the ethyl alcohol content,the solvent amount,and pH value）.The results showed that the extracting times is a key factor to flavone extraction effect. According to the extracting effect of total flavone,the optimum condition is;80%ethyl alcohol solution with 15 time weight more than the raw material,extracting 2 times under non-alkaline condition;As for extracting effect of total icariin,the optimum condition is: 60%ethyl alcohol solution with 20 time weight more than the raw material amount, extracting 2 times under non-alkaline condition.（3）Six flavonoid monomers were isolated from the extract of E.acuminatum by silica gel column chromatography and Sephadex column chromatography.They are identified as Icariin,Icariside-Ⅱ,and 2"-o- rhamnosyl icariside-Ⅱ,4"-o- rhamnosyl icariside-Ⅱ,Anhydroicartin and IkarisosideB by means of ESI MS,¹H NMR,and ¹³C NMR spectra.The others need further studying.（4） The contents of six flavonoid monomers（Icariin,Icariside-Ⅱ,2"-o- rhamnosyl icariside-Ⅱ,4"-o- rhamnosyl icariside-Ⅱ,Ikarisoside B,and Anhydroicartin ） in 40 plants of 13 barrenwort species growing under the same conditions were tested by RP-HPLC. The results indicated that icariin content is relatively high and the others very low.（5）10 common peaks were found in 10 accessions of E.acuminatum from different places of Guizhou Pronvince by HPLC.Among them,the fifth peak is icariin,the seventh peak the is 4"-o- rhamnosyl icariside-Ⅱthe eighth peak is icariside-Ⅱ:the second,the third,and the fourth peaks need further identification.Perhaps they are epimedin A, epimedin B and epimedin C according to the previous reports:the first,the sixth,and the tenth peaks are unknown.（6）In room temperature,2.0 ml solution containing appropriate concentration of BSA was titrated in the Tris-HCl buffer solution（pH7.40） by successive addition of icariin,icariside-Ⅱ,Anhydroicartin and 4"-o- rhamnosyl icariside-Ⅱsolution,respectively. The results indicated that the fluorescence intensities of bovine serum albumin were decreased regularly by the four flavone monomers.更多还原