An ultrahigh performance liquid chromatography-tandem mass spectrometry(UHPLC-MS/MS) with EtOH/K2HPO4·3H2O aqueous two-phase extraction was developed for the simultaneous determination of eight alkaloids in Cynoglossum amabile Stapf et Drumm.Effects of aqueous two phase extraction conditions on extraction efficiencies of eight alkaloids were optimized by single factor experiment.The optimum conditions were as follows:30%(mass ratio) of ethanol,20%(mass ratio) of K2HPO4·3H2O,6∶1 for an extractant to sample ratio,and 4.0 for pH value.The extract was analyzed by UHPLC-MS/MS with acetonitrile-0.1 mol/L ammonium acetate as mobile phases,and eight alkaloids in the extract were well separated.There were good linear relationships for the analytes in the certain concentration ranges with their correlation coefficients(r2) larger than 0.99.The relative standard deviations(RSDs) for precision,reproducibility and recovery were all less than 3.0%.The method is convenient，sensitive and accurate，and is suitable for the determination and confirmation of eight alkaloids in Cynoglossum amabile Stapf et Drumm.