Identification and Semi-quantification of 36 Compounds from Violae herba (Zihuadiding) via UHPLC-Q-Orbitrap-MS/MS as Well as Proposal of Anti-counterfeiting Quality-Marker for Pharmacopeia

Violae herba (Zihuadiding) is a common Chinese herbal medicine. The current Chinese Pharmacopeia (ChP) defines Viola yedoensis Makino as the sole plant origin and esculetin as its sole quality-marker (Q-marker). Esculetin, however, occurs in some counterfeits as well. This implies that current ChP criterion actually cannot recognize the counterfeits of V. yedoensis. The study thus created a specialized MS library using ultra-high-performance liquid chromatography with quadrupole-orbitrap tandem mass spectrometry (UHPLC-Q-Orbitrap-MS/MS) analysis. Through library-comparison, 36 compounds were putatively identified from V. yedoensis; especially, four isomers were successfully distinguished, that is, vitexin vs isovitexin and schaftoside vs isoschaftoside. The subsequent UHPLC-Q-Orbitrap-MS/MS semi-quantification suggested that the chemical contents of 36 compounds varied from 0.001 to 1.958% and the old Q-marker esculetin had high content (0.484 ± 0.028%). According to the relevant principles, flavonoid luteolin was proposed as the new and additional Q-marker. The proposal offers a preliminary evidence to recognize seven adulterations (or counterfeits) of V. yedoensis.