Simultaneous determination of 16 flavonoids and ferulic acid in honey by solid phase extraction and high performance liquid chromatography-tandem mass spectrometry

Abstract

Flavonoids are polyphenol compounds that are widely distributed in honey; they play an important role in its antioxidant activity and aid in distinguishing various species of honey. An analytical method based on solid phase extraction and high performance liquid chromatography with tandem mass spectrometry (SPE-HPLC-MS/MS) for the simultaneous determination of rutin, myricetin, morin, quercetin, naringenin, hesperetin, luteolin, genistein, kaempferol, isorhamnetin, apigenin, pinocembrin, wogonin, chrysin, galangin, genkwanin, and ferulic acid in honey was developed and validated. The honey samples were diluted with HCl solution (pH 2) and cleaned up on a C18 column by solid phase extraction; the methanol eluent was filtered through a nylon membrane and analyzed by LC-MS/MS on an ODS-3 column (150 mm×4.6 mm, 3 μm). Quantitative detection was performed by LC-MS/MS in the multiple reaction monitoring (MRM) mode under negative electrospray ionization (ESI-) conditions. The external standard method was used for quantitation. The method validation data were obtained. The calibration standard curve solutions were prepared by honey matrix-matching; the linear range was 0-200 μg/kg, with correlation coefficients of the calibration standard curves greater than 0.997. The limits of quantification (LOQs) of this method were 20 μg/kg. The recoveries were in the range of 64.5%-113%, and the relative standard deviations were 1.4%-14.5% at three spiked levels (20, 40, and 100 μg/kg). The developed method is convenient, rapid, and effective, and it is suitable for the determination and confirmation of flavonoids in honey samples.