Abstract: A sensitive method was developed for the simultaneous determination of mannitol and betaine in seaweed fertilizer by ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS). The real samples can be determined only by dissolution without further complicated pretreatment. The separation was performed on a waters BEH C18 column with 5 mM ammonium acetate and acetonitrile as the mobile phase at a flow rate of 0.3 mL／min. The temperature of column was 35 ℃. The electrospray source was operated in both positive and negative ion mode by using multiple reaction monitoring (MRM), and the results were calculated by external standard method．The research showed that the linear range of mannitol and betaine was 5-50 and 0.05-0.5 mg/L, and the detection limits were 0.18 and 0.083 mg/L, respectively. The average recoveries were within the range of 90％-110％.