Comprehensive chemical profiling of the flowers of Citrus aurantium L. var. amara Engl. and uncovering the active ingredients of lipid lowering

The flowers of Citrus aurantium L. var. amara Engl. (FCAVA) is popularly consumed as an edible tea for anti-hyperlipidemia. But the active ingredients are not fully clear. In this study, ultra-high performance liquid chromatography coupled to quadrupole time of flight tandem mass spectrometry (UHPLC-QTOF-MS/MS) with diagnostic product ions and neutral loss filtering strategy were successfully used for comprehensive characterization of chemical components in FCAVA. A total of 228 constituents, including 46 organic acids, 12 coumarins and 170 flavonoids, were tentatively characterized (30 confirmed with reference standards). Among them, nineteen flavonoids in 70 batches of FCAVA from different geographical origins were quantified by UHPLC tandem triple quadrupole mass spectrometry (QQQ-MS), which displayed satisfactory linearity, sensitivity, precision, accuracy, and stability. According to analytical results, the distribution of nineteen flavonoids in different geographical origins of FCAVA was clarified. In addition, the effect on LDL uptake of twenty-five flavonoids was investigated in HepG2 cell. It was found that the acacetin, diosmetin and rutin dose-dependently enhanced LDL uptake in HepG2 cells comparing to control. Furthermore, in a hyperlipidemia C57BL/6J mice model, administration of acacetin, diosmetin and rutin (30 mg/kg/d, intragastric, for three weeks) significantly decreased the levels of total cholesterol (TC), triglyceride (TG) and low density lipoprotein cholesterol (LDL-C) in plasma, respectively. Overall, these findings indicated the potential of FCAVA in the development of functional food or medicine for the prevention and treatment of hyperlipidemia, which could be considered for the improvement of quality standardization of FCAVA.