Department of Pharmaceutical Analysis, Hospital of Integrated Traditional Chinese and Western Medicine Affiliated to Nanjing University of Chinese Medicine, Nanjing, Jiangsu, People's Republic of China.
Department of Metabolomics, Jiangsu Province Academy of Traditional Chinese Medicine and Jiangsu Branch of China Academy of Chinese Medical Sciences, Nanjing, Jiangsu, People's Republic of China; State Key Laboratory of Natural Medicines, Department of Chinese Medicines Analysis, China Pharmaceutical University, Nanjing 210009, PR China.
J Chromatogr A. 2019 Dec 20;1608:460418. doi: 10.1016/j.chroma.2019.460418. Epub 2019 Aug 8.
High performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (HPLC-QTOF-MS) is widely used to qualitatively characterize the chemical profiles of herbal medicines, in which the generated adducts and fragments are crucial for confirming molecular ion (deprotonated/protonated ion) and deducing structure of detected components. However, how chromatographic and mass spectrometric (LC-MS) conditions/parameters affect the quantity and intensity of adducts and fragments of detected components is scarcely concerned. In present study, three types of triterpene saponins from the root of Ilex asprella (RIA) were selected as a case study to systematically investigate the effects of LC/MS conditions/parameters on their ionization and fragmentation, so as to obtain higher intensity (higher detection sensitivity) and quantity (rich information) of adducts and fragments for the characterization of components in RIA. It was found that for LC conditions, methanol as organic phase was more benefit for generating more adducts with higher intensity; formic acid as a modifier suppressed the formation of [M-2H], thus promoted the generation of other types of adducts at lower concentration but inhibited the generation when the concentration exceeded 0.1%. MS parameters affect scarcely the quantity but mainly intensity of adducts, cone voltage, source temperature and desolvation gas flow have relatively higher impacts when compared with other parameters. Collision energy affected both quantity and intensity of fragments. MS parameters at the medium value largely increased the quantity and intensity of adducts and fragments. Three-types of triterpene saponins presented structurally specific ionization and fragmentation due to their amounts of acidic substitutes. A total of 55 components were detected and definitely or tentatively identified in RIA under the optimized LC-MS conditions, among which 35 triterpene saponins were firstly discovered. This is the first report that proposes and validates a systematic approach for assessing the effects of LC/MS conditions/parameters on the ionization and fragmentation of analytes, which could be helpful for the optimization of LC-MS conditions for effective chemical profiling analysis of herbal medicines.
高效液相色谱-四极杆飞行时间质谱联用（HPLC-QTOF-MS）广泛用于定性分析中草药的化学成分，其中生成的加合物和碎片对于确认分子离子（去质子化/质子化离子）和推断检测成分的结构至关重要。然而，色谱和质谱（LC-MS）条件/参数如何影响检测成分的加合物和碎片的数量和强度却很少受到关注。在本研究中，选择了三种来自冬青科苦丁茶的三萜皂苷作为研究对象，系统研究了 LC/MS 条件/参数对它们的电离和碎裂的影响，以获得更高强度（更高检测灵敏度）和数量（丰富的信息）的加合物和碎片，从而对苦丁茶中的成分进行表征。结果表明，对于 LC 条件，甲醇作为有机相更有利于生成更高强度的加合物；甲酸作为修饰剂抑制了[M-2H]的形成，从而在低浓度下促进了其他类型的加合物的生成，但在浓度超过 0.1%时抑制了其生成。MS 参数对加合物的数量影响较小，但主要影响加合物的强度，与其他参数相比，锥电压、源温度和脱溶剂气流的影响相对较高。碰撞能量影响碎片的数量和强度。在中等值的 MS 参数下，加合物和碎片的数量和强度都有较大幅度的增加。三种三萜皂苷由于其酸性取代基的数量不同，表现出特定的电离和碎裂结构。在优化的 LC-MS 条件下，共检测并确定或推测出苦丁茶中的 55 种成分，其中 35 种三萜皂苷为首次发现。这是首次提出并验证一种系统的方法来评估 LC/MS 条件/参数对分析物电离和碎裂的影响，这有助于优化 LC-MS 条件，以有效地对中草药进行化学特征分析。
