基于核磁定量与核磁共振代谢组学技术的秦艽品质评价与祛风湿作用研究

In this study, verifying Qinjiao as an example, with content determination, spices discrimination, dispelling wind-dampness activity investigation as starting point, quantitative NMR (qNMR) and NMR based metabolomics (NMRM) were systematically applied to quality and activity investigation of traditional Chinese medicine (TCM). The characteristics of qNMR and NMRM were fully embodied in this work, including quantification not relying on authentic reference and reflecting qualitative information, identification based on whole chemical information and directly reflexing inner quality, activity evaluation from a holistic-metabolism perspective and fitting well with TCM theory, et.al. Specific contents were as follows: establishing a qNMR method for determinations of gentiopicroside, loganic acid, and swertiamarin in Qinjiao; developing a NMRM method for spices discrimination of G. macrophμlla Pall., G. straminea Maxim., G. crassicaulis Duthie ex Burk. and G. dahurica Fisch., and exploring chemical difference among spices; employing a NMRM method to investigate the dispelling wind-dampness activity of Qinjiao on collagen-induced arthritis model, and revealing the mechanism underlying. This study shows great significance for quality evaluation and activity study of Qinjiao, prompts application of qNMR and NMRM to quality and activity investigation of TCM, and develops a new mode for quality evaluation and activity investigation of TCM.

本研究以秦艽为例，以含量测定、种属鉴别、祛风湿作用研究为切入点，将核磁定量与核磁共振代谢组学技术系统应用于中药的品质评价与活性研究中，充分体现了定量不依赖专属对照品、定量同时反映定性信息、鉴别基于全部化学信息、鉴别直接反映药材质量、活性研究从机体整体出发、活性研究与中医整体性思想相吻合等优点。具体工作如下：建立核磁定量法测定秦艽中龙胆苦苷、马钱苷酸与獐牙菜苦苷等成分含量；建立核磁共振代谢组学方法进行秦艽、麻花艽、粗茎秦艽、小秦艽等4种秦艽种间鉴别，并探讨种间化学差异；建立核磁共振代谢组学方法评价秦艽对胶原诱导性关节炎模型的祛风湿作用，并深入探讨祛风湿作用机制。本研究对秦艽的品质评价与祛风湿作用研究具重要意义，促进了核磁定量与核磁共振代谢组学技术在中药品质与活性研究中应用，开启了中药品质评价与活性研究的新方法与新模式。

核磁共振技术以无响应歧视、样本制备简单等优势，在药物分析以及代谢组学领域有着广泛的应用，然而其在中药质量评价与作用机制领域研究较少。秦艽为我国重要的传统药材，《中国药典》收载为龙胆科植物秦艽、麻花秦艽、粗茎秦艽或小秦艽的干燥根，具祛风湿、止痹痛之功效。秦艽中的环烯醚萜成分为其药效作用物质基础。秦艽多基源、多成分特征增加了其质量评价与机制研究的难度。本项目以秦艽为研究对象，采用靶向核磁定量技术对秦艽中化学成分进行定量测定；采用核磁共振技术，结合中药基因条码技术、超高分辨质谱技术等建立多基源药材质量评价方法；并采用核磁共振代谢组学技术对秦艽发挥保肝作用的机制进行初步探讨。本项目建立：（1）靶向轮廓分析法测定秦艽中龙胆苦苷与马钱苷酸含量测定方法；（2）整合ITS2、1H-NMR、TOF-MS组学技术的秦艽质量评价方法；（3）代谢组学技术揭示秦艽保肝作用代谢调控机制。本项目创造性建立靶向轮廓核磁定量法与基因条码-代谢组学融合技术进行秦艽的质量评价，对其他多基源药材质量评价具有借鉴意义；本研究促进了核磁定量与核磁共振代谢组学技术在中药品质与活性研究中应用，开启了中药品质评价与活性研究的新方法与新模式。