基于质谱响应探针和纸基微流控的中药质量评价方法研究

Quality assessment plays a critical role in modernization and internationalization of traditional Chinese medicines (TCMs). Chemical testing is currently the prevailing approach used for TCM quality assessment. However, TCMs contain often a myriad of diverse phytochemicals, and quantification of one or several specific constituents of interest cannot fully account for the efficacy. Therefore, there has been increasing interest in developing biological assays that correlate with the drug’s intended mechanism of action or clinical effect to complement the chemical tests. However, of the bioassays that have been designed, none meet the assay requirements which include multiplexed detection (simultaneous measurement of multiple bioactivities) and integrated detection (integration of chemical and biological tests on the same analytical platform). Inspired by the chemical derivatization method in proteome research, we design novel mass spectrometry-responsive probes using tag molecules with high ionization efficiency, which facilitate simultaneous detection of multiple bioactivities of TCMs. By integrating mass spectrometry-responsive probes with ambient mass spectrometry and paper-based microfluidics, we develop a rapid, sensitive and interference-resistant method for quality assessment of TCMs, where chemical and biological tests are integrated. The method is exemplified on QiShenYiQi Pills to provide an overall evaluation of the drug quality. This study is expected to constitute a valuable addition to the arsenal of TCM quality assessment.

推进中药质量评价技术的创新发展对于实现中药现代化和国际化具有重要意义。中药生物活性检测已成为中药质量评价技术的研究前沿并取得一定进展，但依然面临着诸多挑战，其中的瓶颈性难题包括多重检测（同时检测多个生物指标）以及融合检测（在一个分析平台同步完成生物活性检测和化学检测）。本研究以芪参益气滴丸为研究对象，从检测原理层面进行创新，提出一种中药生物活性检测的新方法——质谱响应探针。通过在可特异性识别生物靶标的肽段上修饰易于离子化的标签分子合成质谱响应探针，实现多靶标高灵敏同步检测。在此基础上，本研究以纸基微流控为检测载体，以原位电离质谱为检测手段，将中药生物/化学检测相融合，建立快速、灵敏、耐干扰的中药质量评价方法。该研究有助于合理、全面地评价芪参益气滴丸的质量，对于中药质量评价技术的创新发展具有重大科学意义和学术价值。

为探索创建中药生物活性多重检测（同时检测多个生物指标）以及化学-生物融合检测（在一个分析平台同步完成生物活性检测和化学检测）方法，本研究从检测原理层面进行创新设计，提出一种基于质谱响应探针的中药质量评价方法。以凝血酶和血管紧张素转化酶（ACE）为靶标，通过在能够特异性识别生物靶标的肽段上修饰易于离子化的标签分子1-（2-嘧啶基）哌嗪，合成得到两个新型质谱响应探针，用于多靶标高灵敏同步检测。以丹参注射液为研究对象，将质谱指纹图谱和双酶检测整合在DART-MS平台上，通过优化实验流程和仪器参数，在两分钟内即可完成化学成分一致性和生物活性的评价。针对色谱指纹图谱耐用性差等问题，提出一种衍生多反应监测（MRM）的指纹图谱分析方法，通过衍生MRM将四极杆飞行时间质谱（QTOF-MS）的MS2质谱参数转化为三重四级杆质谱（QqQ-MS）的MRM质谱参数，无需对照品即可获取MRM，从而建立中药“高密度”指纹图谱，具有耐用性好、效率高、专属性强、灵敏度高等优点。本项目对于中药质量评价技术的创新发展具有重大科学意义和学术价值，有助于更合理、全面地评价中药及其产品的质量。