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

Abstract

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.