Abstract:Abelmoschus manihot（L.）Medic flowers were used as the raw materials for extracting flavonoids via ultrasound-assisted ethanol/ammonium sulfate aqueous two-phase extraction.Based on single-factor experiments，the extraction process was optimized using response surface methodology. The flavonoid components were analyzed by ultra-high performance liquid chromatography-orbit ion trap mass spectrometry（UPLC-Orbitrap-MS）. The antioxidant activities of the flavonoids extracted by this method were compared with those extracted by traditional methods，and the correlation between the main flavonoid components and antioxidant activity was analyzed. The results showed that the optimal extraction conditions for flavonoids from A. manihot flowers were as follows：ethanol mass fraction of 31%，ammonium sulfate mass fraction of 20%，ultrasonic power of 210 W，and A.manihot flower powder amount of 0.5 g.Under these conditions，the two-phase volume ratio was 1.16± 0.03，the partition coefficient was 50.13±2.50，and the flavonoid extraction rate was（98.31±0.13）%.A total of 63 flavonoid compounds were identified by UPLC-Orbitrap-MS，including 13 previously unreported compounds. Compared to aqueous two-phase extraction，ultrasound extraction，and alcohol extraction，ultrasound-assisted aqueous two-phase extraction exhibited stronger antioxidant activity，with EC50 values for DPPH and ABTS+ radicals of 6.37 μg/mL and 17.71 μg/mL，respectively. The ultrasound-assisted aqueous two-phase method efficiently extracted flavonoids from A.manihot flowers，which possessed high antioxidant activity.