南方菟丝子盐炙增强补肾固精的药效物质基础及炮制原理研究

Semen Cuscutae (Tu-Si-Zi), is a well-known traditional Chinese medicine used to nourish liver and kidney. Processing Tu-Si-Zi has a long history. The crude and processed products such as yan zhi pin (YZP) are widely used clinically. YZP means that the crude Tu-Si-Zi was processed by stir-heating with salt solution. Based on the traditional Chinese medicinal theory, YZP was used to improve kidney tropism and tonify the kidney to arrest spontaneous emission. Unfortunately, few studies on the comparing between the crude Tu-Si-Zi and YZP have been reported so far. Cuscuta australis R. Br. has been listed as genuine cuscutae semen in Chinese Pharmacopoeia (edition 2010), for Cuscuta chinensis Lam. is rarely used. After the preliminary study on the Cuscuta australis R.Br., We found that the group of the YZP's total polysaccharides and flavonids can increase the weights of sexual organs in immature male mouses, and also found that the total contents of polysaccharides increased significantly, the contents of hyperoside and quercetin increased significantly while that of astragalin and kaempferol reduced significantly after processed by stir-heating with salt solution. So a research hypothesis is proposed that the changes of the contents and constituents of polysaccharides and flavonoids lead to tonify the kidney to arrest spontaneous emission after processed by stir-heating with salt solution. On the basis of the existing study, a research proposal has be made . First, the effective constiuents to tonify the kidney to arrest spontaneous emission after processed by stir-heating with salt solution will be made clear. The pharmacodynamical study on the kidney-yang deficiency rats induced by hydrocortisone and discussion the mechanism on reproductive system in rats will be developed between the polysaccharides and flavonoids of the crude and those of YZP. Second, the contents and constituents of polysaccharides and flavonoids between the crude and YZP will be compared. Many morden techniques for analysis, isolation and identification, such as HPLC-MSn, high performance gel permeation chromatography (HPGPC) coupled with chemometrics, NMR, etc. will be used. Third, the study on how the two main factors, salt and heating, influence the effection on tonifying the kidney to arrest spontaneous emission and the contents and constituents of polysaccharides and flavonoids. Fourth, the Yan Zhi processing technology wil be optimized. Finally, methods with specificity constituents, such as total polysaccharides, total flavonoids, hyperoside, quercetin, kaempferol, astragalus as the markers, will be established, and the contents of above components will be determined in the crude and YZP. The study will have the importance on explanation on the processing theory of tonifying the kidney to arrest spontaneous emission after processed by stir-heating with salt solution, quality control on the processing products and deeply developing on Cuscuta australis R. Br.

菟丝子滋补肝肾、固精缩尿，盐炙可增强补肾固精作用，但其炮制原理和药效物质如何变化尚不清楚。南方菟丝子作为商品主要来源，被药典列为正品。前期研究发现：盐炙品总多糖和总黄酮能显著增加幼年小鼠性器官重量；盐炙后多糖含量明显增加，总黄酮中金丝桃苷和槲皮素明显增加，山奈酚和紫云英苷明显降低；由此提出 "菟丝子盐炙后黄酮和多糖的变化导致补肾固精作用增强" 的假说。本课题拟对其生品、盐炙品的黄酮、多糖进行补肾固精药效学对比及作用机制研究，阐明药效物质基础；采用HPLC-MS、凝胶色谱、核磁共振等分析、分离、鉴定技术并结合化学计量学方法研究比较盐炙前后其黄酮、多糖组分结构和含量的变化，找出主要影响成分；研究食盐和加热两个主要因素与补肾固精作用及黄酮、多糖组分变化的相关性；依据药效专属成分优化炮制工艺，建立生品和盐炙品质量评价体系，以上对于阐明菟丝子盐炙理论、控制饮片质量，充分发挥其临床疗效具有重要意义。

菟丝子滋补肝肾、固精缩尿，盐炙可增强补肾固精作用，南方菟丝子作为商品主要来源，被药典列为正品。古今临床和药理均证实盐炙菟丝子增强补肾固精的功效，但其盐炙后药效物质基础如何变化，食盐和加热对药效和成分有何影响，炮制增效原理是什么，尚不清楚，制约了菟丝子的进一步开发利用。本课题研究，对于科学阐述中药菟丝子“盐炙增强补肾固精”的传统炮制理论、有效控制菟丝子饮片质量，确保临床疗效具有重要意义，对于深入开发利用南方菟丝子具有参考价值。. 本课题主要研究了：（1）采用药效专属成分优化盐炙工艺，优化了南方菟丝子的总黄酮和总多糖的提取工艺，建立了南方菟丝子黄酮和多糖含量的测定方法，完善生品、盐炙品质量评价体系。. （2）通过对南方菟丝子生品、盐炙品中的总黄酮、总多糖对肾阳虚大鼠生殖和免疫系统影响的对比研究及机制探讨，阐明了菟丝子盐炙增强补肾固精的药效物质基础：南方菟丝子总黄酮和总多糖对肾阳虚大鼠均有保护作用，能改善肾阳虚大鼠免疫力低下的问题，同时能够对氢化可的松对大鼠造成的肾损伤起到保护作用，但是南方菟丝子总黄酮和总多糖的保护途径不同，其中总黄酮通过降低尿素氮的含量对其产生保护作用，而总多糖则是通过降低肌酐的含量起到保护作用。. （3）采用HPLC-MSn、凝胶色谱等多种分析、分离、结构鉴定方法研究了盐炙前后黄酮和多糖组分的变化：实验得知盐炙后没有新的化学成分产生；利用PCA对南方菟丝子生品和盐炙品黄酮进行了分析，其生品和盐炙品具有显著性差异，其中绿原酸、槲皮素-3-O-β-D半乳糖-（2→1）-β-D芹糖苷、金丝桃苷、槲皮素盐炙后上升，异鼠李素糖苷、山奈酚、异鼠李素盐炙后下降。对菟丝子总多糖精制前后单糖组成进行分析，结果表明菟丝子总多糖精制前后基本没有明显变化，以甘露糖为主，约占多糖总量的50%以上，对菟丝子生品和盐炙品中的总多糖含量进行了测定，以甘露糖为对照品，测得菟丝子生品中的总多糖含量为6.71%，盐炙后总多糖含量升高，为14.71%。. （4）研究了食盐、加热两个关键因素对补肾固精药效和黄酮、多糖成分的影响并探讨了谱——效、量——效关系：南方菟丝子盐炙后没有新的化学成分产生，结合化学成分的变化及补肾固精药效的比较可知，加热和加盐的方式是菟丝子盐炙增强补肾固精药效的主要因素。. 以上对于阐明菟丝子盐炙理论、控制饮片质