Abstract: A method of LC-MS/MS was used for qualitative and quantitative analysis of flavonoids in Crotalaria sessiliflora L.. The sample was smashed and extracted with methanol,and then purified with D101 macroporous resin. UltimateTM C18 column was used as stationary phase and the mixture of methanol and 0.5% (φ) acetic acid was used as mobile phase for gradient elution. ESI was adopted in MS/MS. The main characteristic fragment ions were analyzed in the positive and negative ion scan mode. Vitexin,isovitexin,orientin,isoorientin and trematin were detected based on MS/MS data. Linear relationship were found between the peak area and the mass concentration of orientin,isoorientin,vitexin,isovitexin in a certain range,with detection limits (3S/N) of 50.2,40.5,52.1,42.1 μg·L-1,respectively. The method was applied for analysis of Crotalaria sessiliflora L. samples,giving recoveries determined by standard addition method in the range of 96.0%-103% with RSD (n=5) less than 3.0%.