In-depth profiling, characterization, and comparison of the ginsenosides among three different parts (the root, stem leaf, and flower bud) of Panax quinquefolius L. by ultra-high performance liquid chromatography/quadrupole-Orbitrap mass spectrometry

Despite Panax quinquefolius L. serving as a crucial source for food additives, healthcare products, and herbal medicines, unawareness of the metabolome differences among three parts (root, PQR; stem leaf, PQL; flower bud, PQF) seriously restricts its quality control. Ultra-high performance liquid chromatography/quadrupole-Orbitrap mass spectrometry (UHPLC/Q-Orbitrap-MS) was fully utilized to comprehensively identify and compare the ginsenoside compositions among PQR, PQL, and PQF. Metabolite profiling and characterization were performed by coupling reversed-phase UHPLC (a CSH C18 column) and improved untargeted data-dependent MS2 acquisition in the negative mode. A novel vehicle, "Ginsenoside Sieve," was proposed by developing fixed tolerance (± 10 mDa), discrete mass defect filtering (MDF) based on the m/z features of 499 known ginsenosides, which assisted in the screening of 71 (from 3453 ions), 89 (from 6842), and 84 (from 7369) target precursor masses for PQR, PQL, and PQF, respectively. The newly established data-dependent acquisition (DDA) approach exhibited 14% improvement in characterization of targeted components (using a PQL sample), and comparable performance in identifying the unknown, compared with conventional DDA. We could characterize 347 saponins (147 from PQR, 173 from PQL, and 195 from PQF), and 157 thereof not ever-isolated from the Panax genus. These potentially new saponins have 63 unknown masses. Subsequent untargeted metabolomics analysis unveiled 20 marker saponins, among which m-Rb1, Rb1, Ro, m-Rb2, and m-Rb1 isomer are the most important diagnostic for differentiating the three parts. Conclusively, the established improved DDA represents a potent ginsenoside characterization strategy, and the results obtained in this work would benefit better quality control of P. quinquefolius. Graphical abstract.