基于HPLC-MSn-SPE-CryoNMR联用技术的中药道地性物质基础研究新方法及应用——以川产道地药材附子为例

中药道地性物质基础是道地中药材质优效佳的根本原因，也一直是中药研究的热点。目前相关研究存在难以体现道地差异全面性和药材功效，以及定量指标与工作量难以平衡的问题。申请人以川产道地药材附子为例，经前期研究发现附子多种组分均存在一定道地差异，并创新发现附子不同组分核磁定量的关键信号。因此，本项目基于HPLC-MSn-SPE-CryoNMR联用技术提出中药道地性物质基础研究的新模式。本项目将开展如下研究：以大鼠、小鼠整体动物模型评价不同产地附子毒效差异；应用HPLC-MSn-SPE-CryoNMR联用技术，快速发现及分离鉴定附子道地差异成分；运用该联用技术定量各差异成分和组分，并建立不同产地指纹图谱，从而构建单体-组分-谱图的多层次道地性关系；利用斑马鱼和H9c2细胞模型进一步阐明附子毒效-道地性关系。通过上述研究，期望阐明附子道地性物质基础的科学内涵，并建立中药道地性物质基础研究新模式。

The material basis of genuineness of traditional Chinese medicine(GTCM) is the fundamental reason for the superior quality and effectiveness of genuine medicinal materials, and it has always been a hot spot in Chinese medicine research. The current related research has problems that it is difficult to reflect the comprehensiveness of genuine differences, the efficacy of medicinal materials, and it is difficult to balance quantitative indicators and workload. The applicant took Fuzi(Aconitum carmichaelii Debx.), a genuine medicinal herb in Sichuan, as an example. In the previous research, it was found that there are some differences in some components of Fuzi from different producing areas. Besides, the key signals of different components of Fuzi in quantitative nuclear magnetic resonance have been innovatively discovered. Therefore, this project proposes a new model based on HPLC-MSn-SPE-CryoNMR to carry out the material basis research of GTCM. Firstly, the differences in toxicity and efficacy of Fuzi from different producing areas are evaluated by rat and mouse animal models. Then, the rapid discovery, isolation and identification of different components of Fuzi are researched by HPLC-MSn-SPE-CryoNMR. Next, the different compounds and components are accurately quantified based on the combined technology, and fingerprints of Fuzi from different origins were established, so as to construct a multi-level genuine relationship of compound-component-fingerprint. Finally, the relationship between toxicity-effect and genuineness of Fuzi is further clarified by zebrafish and H9c2 cell models. Through the above research, it is hoped to clarify the scientific connotation of the genuine material basis of Fuzi, and establish a new mode of research on the material basis of GTCM.