National TCM Key Lab of Serum Pharmacochemistry, Heilongjiang University of Chinese Medicine, Heping Road 24, Harbin 150040, China.
Analyst. 2013 Jun 7;138(11):3303-12. doi: 10.1039/c3an36902a.
To discover and screen the constituents or metabolites absorbed into blood after oral administration of herbal medicines tends to be more and more difficult. In this work, an integrative pattern recognition approach of principal component analysis (PCA) and orthogonal partial least squared discriminant analysis (OPLS-DA) was successfully applied for rapid discovery of natural compounds from herbal medicines. A rapid, sensitive, and reliable ultra performance liquid chromatography coupled with electrospray ionization/quadrupole-time-of-flight mass spectrometry (UPLC-ESI-Q-TOF-MS) method with Masslynx™ software was established to characterize the chemical constituents and rats metabolites of Phellodendri amurensis cortex (Guan Huangbai, GHB). The analysis was performed on a Waters UPLC HSS T3 column (2.1 × 100 mm, 1.8 μm) using gradient elution system. A hyphenated electrospray ionization and quadrupole-time-of-flight analyzer was used for the determination of accurate mass of the protonated or deprotonated molecule and fragment ion in both negative and positive modes. A total of 46 peaks were obtained, 41 of which were tentatively characterized from GHB. In the S-plot of OPLS-DA, 24 interested ions (17 ions in positive mode and 6 ions in negative mode) were extracted, among them, 12 absorbed prototype components of GHB and 12 metabolites were identified in vivo. Major metabolic reactions of GHB were demethylation, methylation and glucuronidation. This is the first report on systematic analysis of chemical constituents and in vivo metabolites of GHB. It is concluded that UPLC-MS coupled with pattern recognition approach for the identification of herbal constituents in biological samples has been successfully developed. The method can also be applied to rapid discovery and global characterization of the constituents in rat serum after oral administration of other herbal medicines.
用于发现和筛选草药口服给药后吸收到血液中的成分或代谢物变得越来越困难。在这项工作中，成功地应用了一种主成分分析（PCA）和正交偏最小二乘判别分析（OPLS-DA）的综合模式识别方法，用于快速发现草药中的天然化合物。建立了一种快速、灵敏、可靠的超高效液相色谱-电喷雾/四极杆飞行时间质谱联用（UPLC-ESI-Q-TOF-MS）方法，并用 Masslynx™软件对黄柏（关黄柏，GHB）的化学成分和大鼠代谢物进行了表征。分析在 Waters UPLC HSS T3 柱（2.1×100mm，1.8μm）上进行，采用梯度洗脱系统。串联电喷雾电离和四极杆飞行时间分析仪用于测定正负模式下质子化或去质子化分子和碎片离子的精确质量。共得到 46 个峰，其中 41 个峰被暂定表征为 GHB。在 OPLS-DA 的 S-plot 中，提取了 24 个感兴趣的离子（正模式下 17 个离子，负模式下 6 个离子），其中 12 个是 GHB 的吸收原型成分，12 个在体内被鉴定为代谢物。GHB 的主要代谢反应为去甲基化、甲基化和葡萄糖醛酸化。这是首次对 GHB 的化学成分和体内代谢物进行系统分析的报道。结论是，已经成功开发了将模式识别方法与 UPLC-MS 相结合用于鉴定生物样品中草药成分的方法。该方法还可应用于其他草药口服给药后大鼠血清中成分的快速发现和全面表征。
