Identification of alkaloid constituents from Fangchi species using pH control liquid‐liquid extraction and liquid chromatography coupled to quadrupole time‐of‐flight mass spectrometry

Rationale Fangchi ( F .) species from four different origins have been widely used to treat or prevent diseases, and their main constitutes are several types of alkaloids. Identification of alkaloids in F . species is a necessary step to understand the therapeutic properties of each different origin, but this has not yet been fully performed. Methods Several types of alkaloids were extracted from F . species using ultrasonication with 70% CH 3 OH and the extract was partitioned at pH 2 and 12 to enrich alkaloid constituents and to remove interferences. The separation of alkaloids in the Fangchi extract was performed on a C18 column using gradient elution and their tandem mass spectra were obtained by quadrupole time‐of‐flight tandem mass spectrometry (QTOF‐MS/MS) to perform accurate mass measurements of fragment ions for the alkaloid constituents. Results Several types of alkaloids were successfully separated and identified by LC/ESI‐MS/MS. The structural assignment of individual alkaloids was performed based on convergence of MS/MS spectral data, pH partitioning behavior, LC retention behavior, and accurate mass measurements. The pH partition of the extract provided structural information about unknown alkaloids extracted from Fangchi species. A total of 28 compounds were identified and tentatively characterized, and of these 10 alkaloids were reported for the first time in the investigated F . species. Conclusions The chemical profiling of alkaloids in F . species with different origins was performed for the first time and provided diagnostic ions for diverse alkaloids in F . species. Marker compounds were suggested based on the 28 characterized compounds for quality evaluation and the differentiation of Fangchi species with four different origins. Copyright © 2015 John Wiley & Sons, Ltd.