A simple and fast method for the simultaneous quantification of six flavonoids in Fructus aurantii by UPLC–PDA and confirmation by UPLC/ESI-Q-TOF-MS

Abstract

Fructus aurantii, the dried, immature fruit of Citrus aurantium L. and its cultivated varieties (Family Rutaceae), is one of the commonly used Chinese medicinal herbs in which flavonoids are considered as the major bioactive constituent. In this study, reverse phase ultra-performance liquid chromatography coupled with photo diode array detector (UPLC–PDA) method was developed for the simultaneous quantification of six flavonoids, namely narirutin, naringin, hesperidin, neohesperidin, nobiletin, and tangeretin in Fructus aurantii. The analysis was performed on a Waters Acquity UPLC BEH C18 column (50 mm × 2.1 mm, 1.7 μm) with 1% aqueous acetic acid-acetonitrile gradient elution. The runtime was as short as 8 min. The method was validated in terms of linearity, sensitivity, precision, stability, and accuracy. It was found that all calibration curves showed good linearity (r² > 0.9991) within the test ranges. The limits of detection (LOD) and limits of quantification (LOQ) were all lower than 35 and 107 ng mL⁻¹. The overall relative standard deviations (RSDs) for intra- and inter-day repeatability were less than 2.94% and 2.74%, respectively. The sample was stable for at least 48 h. The recoveries were 95.33–103.78%. The newly proposed method was found to be simple, rapid, and reproducible for quantitative analysis of six flavonoids in ten samples from different origins. In addition, the samples were also analyzed on an ultra-performance liquid chromatography-electrospray ionization-time-of-flight mass spectrometry (UPLC/ESI-Q-TOF-MS) system to confirm the identification results.