A novel hybrid scan approach enabling the ion-mobility separation and the alternate data-dependent and data-independent acquisitions (HDDIDDA): Its combination with off-line two-dimensional liquid chromatography for comprehensively characterizing the multicomponents from Compound Danshen Dripping Pill.

Data-dependent acquisition (DDA) and data-independent acquisition (DIA)-based MSn strategies are extensively applied in metabolites characterization. DDA gives accurate MSn information, but receives low coverage, while DIA covers the entire mass range, but the precursor-product ions matching often yields false positives. Currently available MS scan approaches rarely integrate DIA and DDA within a duty circle. Utilizing a Vion™ IM-QTOF (ion mobility-quadrupole time-of-flight) mass spectrometer, we report a novel hybrid scan approach, namely HDDIDDA, which involves three scan events: 1) IM-enabled full scan (MS1), 2) high-definition MSE (HDMSE) of all precursor ions (MS2); and 3) high-definition DDA (HDDDA) of top N precursors (MS2). As a proof-of-concept, the HDDIDDA approach combined with off-line two-dimensional liquid chromatography (2D-LC) was applied to characterize the multiple ingredients from a reputable Chinese patent medicine, Compound Danshen Dripping Pill (CDDP) used for treating the cardiovascular diseases. An off-line 2D-LC system by configuring an XBridge Amide column and an HSS T3 column showed a measurable orthogonality of 0.92 and enhanced the separation of co-eluting components. A fit-for-purpose HDDIDDA methodology was developed in the negative mode to characterize saponins and salvianolic acids, while tanshinones in the positive mode. Computational workflows to efficiently process the acquired HDMSE and HDDDA data were established, and the searching of an in-house CDDP library (recording 712 compounds) eventually characterized 403 components from CDDP, indicating approximate 12-fold improvement compared with the previous report. The HDDIDDA approach can measure collision cross section of each component, and merges the merits of DIA and DDA in MS2 data acquisition.

An ion mobility-enabled and high-efficiency hybrid scan approach in combination with ultra-high performance liquid chromatography enabling the comprehensive characterization of the multicomponents from Carthamus tinctorius.

Liquid chromatography/mass spectrometry (LC/MS) is extensively applied for the untargeted/targeted analyses of the herbal components, utilizing data-dependent acquisition (DDA) or data-independent acquisition (DIA) to record the fragmentation information useful for the structural elucidation. A new trend recently has emerged by integrating DDA and DIA to render the hybrid scan, which, unfortunately, has rarely been reported. Herein, by using the Vion™ ion-mobility quadrupole time-of-flight mass spectrometer, a hybrid scan strategy (HDMSE-HDDDA) was presented and validated by the untargeted characterization of the multicomponents from Carthamus tinctorius (safflower), in combination with reversed-phase ultra-high performance liquid chromatography (RP-UHPLC). Good chromatographic separation was achieved on an HSS T3 column within 26 min, while HDMSE-MS/MS was used to acquire the collision-induced dissociation MS2 data in the negative mode. Automatic workflows (e.g., data correction, precursors/product ions matching, and peak annotation) were well established on UNIFI™ (incorporating an in-house library recording 261 known compounds) to process the obtained MS2 data. Compared with single DDA or DIA, the hybrid approach of HDMSE-HDDDA better balanced between the coverage and reliability, led to high-definition MS spectra, offered useful collision cross section (CCS) information, and showed satisfactory identification performance comparable to MSE. A total of 141 components (involving 41 quinochalcones, 66 flavanols/flavones, 11 flavanones, 6 organic acids, 1 polyacetylene, and 16 others) were characterized from safflower. Moreover, CCS prediction could assist isomers characterization, to some extent. Conclusively, this hybrid scan approach enables a dimension-enhanced MS data acquisition strategy providing the complementary structural information, which more suits the chemical characterization of complex samples.

Highly selective monitoring of in-source fragmentation sapogenin product ions in positive mode enabling group-target ginsenosides profiling and simultaneous identification of seven Panax herbal medicines.

In-source fragmentation of ginsenosides in the positive ESI mode (pISF-G) frequently occurs, which results in little fragment information useful for the structural elucidation. We are aimed to unveil the genesic mechanism and explore its potential significance in quality control of Ginseng and the related compound formulae. By applying six high-resolution mass spectrometers from Agilent, Waters, and Thermo Fisher, we could primarily demonstrate the susceptibility of pISF-G. The ion clusters in the positive full-scan MS1 spectra were generated from the protonated sapogenins by successive elimination of H2O, and showed specificity for ginsenoside classification. Selective ion monitoring (SIM) of the sapogenin product ions could delineate group-target ginsenoside profiles from Ginseng. A high-selectivity characteristic chromatogram (CC) was elaborated for Ginseng, on the Vion™ IMS-QTOF mass spectrometer by IM (ion mobility) separation and quadrupole filtering of four sapogenin fragments (m/z 407.37/CCS 206.24 Å2; m/z 423.36/CCS 211.26 Å2; m/z 439.36/CCS 209.60 Å2; m/z 457.37/CCS 217.81 Å2). Chemometric analysis, based on the CC data of seven Ginseng drugs (P. ginseng, P. quinquefolius, P. notoginseng, Red ginseng, leaf of P. ginseng, P. japonicus, and P. japonicus var. major), disclosed 35 marker compounds. We could readily discriminate among P. ginseng, P. quinquefolius, and P. notoginseng, in 15 different compound formulae by identifying these marker compounds on both the Vion IMS-QTOF and QTrap 4500 mass spectrometers. Conclusively, SIM of the pISF-G sapogenin product ions renders a new concept of CC enabling the group-target profiling of ginsenosides and authentication of Ginseng and the related compound formulae.