Abstract: In order to establish a rapid and effective method for the quantitative determination of microcystins (MC-LR, MC-YR and MC-RR) in bivalves such as mussels, by taking the mussels as the research object and optimizing the factors such as the extracting solvent, the concentration of extracting solvent, and dilution ratio, etc., a method for the determination of three microcystins (MC-LR, MC-YR and MC-RR) in mussels was established by using the direct dilution combined with ultra-high performance liquid chromatography-tandem mass spectrometry. The results showed that: the determination method of three toxins in mussels was as follows: after being extracted with 90% methanol aqueous solution, the extract fluid was diluted with 0.5% acetic acid methanol solution, and the diluent was directly determined by ultra-high performance liquid chromatography-tandem mass spectrometry. The limit of detection (LOD) was 5 μg·kg^-1 and the limit of quantification (LOQ) was 15 μg·kg^-1. The mixed standard substance of 15, 30, and 150 μg·kg^-1 microcystins was added to mussels for the spiked recovery test, and the recovery rates of three microcystins (MC-LR, MC-RR and MC-YR) were 89.4%-91.8%, 77.1%-106.3%, and 75.8%-102.3%, respectively. The relative standard deviations were 3.5%-13.5%, 5.6%-9.2%, and 4.5%-8.1% (n=6), respectively. The determination method was simple and fast, with low cost and strong operability, which could meet the technical requirements of the current batch quantitative detection of three microcystins in mussels.