Identification of flavonoid glycosides in Rosa chinensis flowers by liquid chromatography–tandem mass spectrometry in combination with 13C nuclear magnetic resonance

Abstract

Flowers of Rosa chinensis are widely used in traditional Chinese medicine as well as in food industry. Flavonoid glycosides are believed to be the major components in R. chinensis that are responsible for its antioxidant activities. In this work, a liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method was developed for analysis of flavonoid glycosides presented in ethyl acetate extract of dried R. chinensis flowers. Twelve flavonoid glycosides were separated and detected. By comparing the retention times, UV spectra, and tandem MS fragments with those of respective authentic compounds, eight flavonoid glycosides were unequivocally identified. Although the other four were also identified as flavonoid glycosides, the glycosylation positions could not be determined due to lack of authentic compounds. Fortunately, the glycosylation effects were clearly observed in the 13C NMR spectrum of the extract. The detailed structural information was, therefore, obtained to identify the four flavonoid glycosides as quercetin-3-O-d-glucoside, quercetin-3-O-d-xyloside, kaempferol-3-O-d-xyloside and quercetin-3-O-d-(6″-coumaroyl)-galactoside. These flavonoid glycosides were detected and identified for the first time in this botanic material. This work reports on the first use of 13C NMR of a mixture to enhance a rapid HPLC–MS/MS analysis. The proposed analytical protocol was validated with a mixture of authentic flavonoid glycosides.