基于体内过程与失眠蛋白网络关联研究酸枣仁的有效成分群

Semen Ziziphi Spinosae(SZR), originating from a medicinal plant Ziziphus jujuba var. spinosa (Bunge) Hu ex H. F. Chow, has been widely used as a Chinese herbal medicine (CHM) for the treatment of insomnia in clinical practice. Under the mode of isolation-to-bioassay or bioassay guided isolation, the previous efforts in isolating and screening single active compounds from SZR are less satisfactory as expected. However, the mode removes the importance of interrelationships of the multi-components., and the standard of quality control can not stand for the whole therapeutic efficacies of SZR. In this project, the combination of liquid-mass techenologies were developed for the analysis of.phytochemical profile in SZR. Using the method of traditional Chinese medicine serum pharmacochemistry and in vivo metabolism, the absorbed components and metabolites in vivo in insomnia rat were identified by using “in vitro to in vivo translation” approach. Meanwile, the bioactive compounds were clarified. Based on the interaction of components in plasma and the significant correlation protein in the network of isonomia, the candicate effective componets screening of SZR were selected. Finally, the real-time components trapping and combining system for sample preparation were applied. The effect of these active components in treatment of insomnia was proved in body. The action features of SZR through multi-component, multi-channel and multi-target were elucidated. The results mentioned above revealed the chemical base of SZR in treatment of insomnia,provided a good option for quality evaluation in SZR, and served as a prerequisite for the multi-component pharmacokinetics and interactions.. The proposed method processed the acedamic value for inspiring an herb medicine into a clear component, quality controlled and effective new drug based discovery of effective components screening.

酸枣仁来源于药用植物酸枣，是治疗失眠的常用中药。在以活性为导向的传统分离模式下，已阐明了酸枣仁治疗失眠的一些有效成分。但该模式难以从整体上揭示其有效成分的全貌，无法建立体现中药特点的质量评价标准。本项目整合多种液质联用技术，全面表征酸枣仁整体化学成分群。通过血清药物化学和体内代谢分析结合，在病理动物模型下，开展酸枣仁体内复杂成分群与体外化学成分群的关联分析研究，明确酸枣仁体内化学成分群。将入血成分与失眠蛋白网络靶标进行映射，遴选潜在有效成分群；最终，利用目标化学成分剔除技术，结合整体动物模型，确证酸枣仁治疗失眠的有效成分群，阐明其多成分、多靶点、多途径整合作用效应。研究结果不仅可以揭示酸枣仁治疗失眠的化学基础，为其资源质量评价提供科学依据，为后期多组分协同作用及药动学研究提供实验基础，对以酸枣仁有效成分群开发创新组分中药具有理论价值。

酸枣仁的本底化学成分基本清晰，且在以活性为导向的传统分离模式下，阐明了酸枣仁催眠作用的部分有效成分。然而，这种追踪酸枣仁中一种或几种有效成分的研究模式，难以从整体上揭示其有效成分的全貌以中医药整体观为指导。鉴于此，本项目拟以酸枣仁的体内过程和失眠蛋白网络关联，并辅以药效学实验验证，从吸收、代谢和网络多重角度共同锁定酸枣仁的有效成分群。.建立了大鼠脑组织中5-HT，GABA，Glu，NE和DA含量的方法，并用于失眠模型的评价；基于1HNMR代谢组学技术探讨了酸枣仁改善睡眠作用机制；建立了集样品前处理、分析和数据采集模式一体化的高分辨质谱体内分析新策略；首次全面表征了酸枣仁口服给予大鼠后尿液中56种成分（包括30种原型和26种代谢产物）；血清中43种成分（包括25个原型和18个代谢产物）；粪便中63种（包括34种原型成分，29种代谢产物）；胆汁中27种成分（包括16种原型成分，11种代谢产物）；黄酮C糖苷部分在肠道发生水解反应，继而发生肝脏I相和II相代谢反应，通过尿液排出；生物碱类成分通过II相代谢反应经尿液排泄；皂苷类成分部分以原型的形式排出。皂苷元及次级苷应该是通过粪便排泄。建立了酸枣仁体内成分群的成分-失眠蛋白网络，初步筛选出乌药碱、酸李碱、芹菜素、斯皮诺素和酸枣仁皂苷A 5个入血原型成分与GABA、E类、Ach类、5-HT类、Glu类和DA类受体等失眠靶点具有直接相关性。建立了酸枣仁水提取物质量控制方法，并应用于酸枣仁提取物的质量评价；从整体动物水平，确证了酸枣仁总黄酮和总皂苷均为改善睡眠的潜在有效成分部位，为酸枣仁质量标志物的确定奠定了坚实的基础。.该项目以第一作者发表研究性论文3篇（SCI 2篇），综述性文章2篇；以通讯作者共发表研究性论文6篇（SCI 1篇）。研究期间获得专利授权1项，另有3项公开。培养硕士研究生4名，其中2名已毕业。