【作者】 王俊永；

【导师】 梁生旺；

【作者基本信息】 广东药学院 ， 药物化学， 2008， 硕士

【摘要】 本文分析了近年来中药复方药效物质基础的研究现状和研究方法,探讨了中药复方的药效物质基础研究对中药创新发展的意义,归纳了研究对象脑得生丸中主要化学成分及其药理活性的研究概况。在中医药学理论和实践的指导下,设计研究路线,以追踪中药复方整体药效部位为核心,结合药效学试验筛选出脑得生丸的有效部位,借助大孔吸附树脂纯化技术对脑得生丸进行纯化分离得到脑得生洗脱部位,并采用适宜的分析手段建立药效部位的质量控制方法。研究以总皂苷、总黄酮、葛根素和羟基红花黄色素A(HSYA)等有效成分为评价指标,采用正交设计试验和大孔吸附树脂纯纯化技术优选提取纯化工艺为:将脑得生丸成方分为两部分,以三七、川芎和山楂为第一部分,粗粉混匀,用10倍量80%乙醇的溶剂回流提取2次,每次60min,合并提取液,减压回收溶剂,浸膏减压干燥,得脑得生丸第一部分提取样品Ⅰ;以葛根和红花为第二部分,用12倍量40%乙醇的溶剂回流提取2次,每次45min,,合并提取液,减压回收溶剂,浸膏减压干燥,得脑得生丸第二部分提取样品Ⅱ。脑得生的两部分提取物(样品Ⅰ、Ⅱ)均以A型吸附树脂为纯化填料,树脂柱径高比为H1,上样浓度分别为C1和Ca,上样量与树脂体积比均为N,以5B倍树脂体积的水清洗树脂柱后,分别用M%乙醇(样品Ⅰ)和N%乙醇(样品Ⅱ)洗脱树脂,洗脱剂用量分别为5B和6B量树脂体积。收集M%和N%乙醇洗脱液,合并,减压干燥至干,粉碎,混匀,制得脑得生纯化样品。建立大鼠全脑缺血为模型,以神经症状评分,脑组织含水量,脑梗死面积为评价指标初步筛选出脑得生丸的药效部位,并通过药效学试验验证所得纯化样品为脑得生丸的药效部位。利用HPLC法和UV法,建立了脑得生丸药效部位的质量控制方法。药效部位中的总皂苷含量为12.52%～12.71%,总黄酮含量为11.53%～11.75%,阿魏酸含量为0.226%～0.229%,异黄酮的含量总和为35.43%～36.07%,HSYA含量为2.14%～2.21%,脑得生有效部位中可测定的成分总含量大于60%。其中总皂苷、总黄酮、阿魏酸、异黄酮(葛根素、3‘-甲氧基葛根素、芹糖基葛根素和大豆苷之和)和HYSA的转移率分别为89.14%～90.81%,79.33%～81.13%,81.89%～84.25%,84.47%～85.72%,71.21%～73.38%,表明研究所建立的脑得生丸药效部位提取纯化工艺是合理的、可行的,各有效成分的含量和转移率均符合开发新药要求。本研究初步阐释了三七总皂苷、山楂总黄酮、阿魏酸、异黄酮(葛根素、3‘-甲氧基葛根素、芹糖基葛根素和大豆苷等)和HYSA为脑得生丸的药效物质,为脑得生丸现代创新药物的进一步研究开发奠定基础。更多还原

【Abstract】 In this paper, the present situation and development about the therapeutic basis of Traditional Chinese Medicine prescription and the methods which were used to study the constituents of Chinese medicine were summaried. The significance of the therapeutic basis of traditional Chinese medicine prescription in novel drug developments was mentioned. As the crucial chemical constituents and pharmacological activities, contained in Nao-Deshen pill, were discussed in recent researches. Under the direction of theroy and clinical practice of Traditional Chinese Medicine, this study scheme was designed to trace the therapeutic part as the core of Chinese medicine. Resorted to the extraction technology of macroporous adsorbing resin to purify Nao-Deshen pill, while the effective fraction was screened by the pharmacodynamic trial and its quality control method was established by suitable means.In Nao-Deshen prescription, there were some effective constituents, such as saponins, flavones, ferulaic acid, puerarin, isoflavones and hydroxysafflor yellow A (HSYA), which could be as evaluationt factors in the technology of extraction and purificaiton. Given the previous study, Nao-Deshen prescription was divided into two parts:: Part one contained Notoginseng, Szechwan Lovage Rhizome and Hawthorn Fruit; Part two included Kudzuvine Root and Safflower. The extractions were opimized by orthogonal design trial as follow: The extracting solvent of part one was 80% ethanol cunsumed ten times the amount of material in the volume, and two times for one hour each time, then the merged extracting liquid was dried by decompression as sampleⅠ; The extracting solvent of part two was 40% ethanol cunsumed twelve times the amount of material in the volume, and two times for three quarters forty five minutes each time, then the merged extracting liquid was dried by decompression as sampleⅡ. Then, A, a kind of Mqacroporous adsorbing resin, was selected as the packing material to purify two extracts (sampleⅠandⅡ) in the purification technology. For sampleⅠandⅡ, the same column in diameter/high was H1, and the ratio of loading capatity to resin volume was N, but the adsorbing concentrations, differently, were C1 and Ca respectively. After adsorbed, both sampleⅠand sampleⅠwere eluted by water with five times of the resin volume, and then eluted by certain concentration of ethanol respectively. As that, the column of sampleⅠwas eluted by M% ethanol with five times of the resin volume and t he column of sampleⅡwas eluted by N% ethanol with six times of the resin volume. Merging M% and N% elutiions, dried by decompression, pulverized and mixed well, and then the purified sample of Nao-Deshen pill was made. While a model of the rat global brain ischemia was made to screen and verify the effective fraction of Nao-Deshen by the nervous symptom, the oedema of brain , and the infarct size of brain.By analytical methods of High Performance Liquid Chromatography (HPLC) and Thin Layer Chromatography (TLC), the quality assessment method on the therapeutic part of Nao-Deshen pill was established. The contents of saponins, flavones, ferulaic acid, isoflavones and HSYA were 12.52%～12.71%, 11.53%～11.75%,0.226%～0.229%, 35.43%～36.07% and 2.14%-2.21% respectively. Therefore, the total content of five fractions was more than 60%. And the transferring rates of saponins, flavones, ferulaic acid, isoflavones and HSYA were 89.14%～90.81%,79.33%～81.13%,81.89%～84.25%,84.47%～85.72% and 71.21%～73.38% respectively. These met demands of the novel medicine development, showing that the technology on the study of Nao-Deshen were reasonable and feasible.This thesis preliminarily illuminated that the therapeutic substances of Nao-Deshen maybe be saponins from Notoginseng, flavones from Hawthorn Fruit, ferulaic acid, isoflavones and HYSA, which should provide a good foundation for further development of Nao-Deshen pill.更多还原