Rapid quantitation and identification of the chemical constituents in Danhong Injection by liquid chromatography coupled with orbitrap mass spectrometry

ABSTRACT Danhong Injection (DHI) is a Chinese medicine patent drug to treat cardiovascular diseases. It is derived from the herbal medicines Dan-shen and Hong-hua. The bioactive compounds of DHI are polar phenolic acids and flavonoid glycosides. Thus far, the contents of major compounds in DHI are not well understood, and the identification of minor compounds lacks rapid methods. In this work, quantitative and qualitative analyses of DHI compounds were performed using ultra-high performance liquid chromatography coupled with orbitrap mass spectrometry (UHPLC/orbitrap-MS). DHI was separated on an Acquity HSS T3 column (1.8 µm, 100 mm × 2.1 mm) and eluted with acetonitrile-water (containing 0.1% formic acid) to determine the contents of 12 compounds within 6 minutes. The method was fully validated according to the ICH guidance. To identify the minor compounds, an ion statistics-based strategy was used to dig for 4 filtering ions and 6 diagnostic ions from 22 reference standards. A total of 117 compounds, including 76 phenolic acids, 20 flavonoids, and 21 other compounds were tentatively identified. The poor stability of salvianolic acid A upon storage was also discussed.