Abstract: A sensitive liquid chromatography-electrospray ionization-tandem mass spectrometry (LC-ESI-MS/MS) method for the determination of phytosterols in sargassum fusiforme has been developed and validated. Four kinds of phytosterols were derivatived by dansyl hydrazine and separated on a reversed-phase Agilent Poroshell 120 EC-C18 column(50 mm×3.0 mm, 2.7 μm) with a mobile phase of methanol-water (99:1, V:V) at a flow rate of 0.3 mL/min, and the injection volume was 10 μL. An electrospray ionization source was applied and operated in the positive ion mode, confirmed by the selected reaction monitoring (SRM) mode using an internal standard method.Phytosterols showed a wide linearity in 0.05~100 μg/mL, with correlation coefficients between 0.990 3 and 0.998 1.The limits of detection of phytosterols were between 0.2 and 20 ng/mL. This simple, selective and sensitive method can be successfully used for the determinations of different types of matrices, and has a 25~500 fold increase in sensitivity than that in high performance liquid chromatography.