A method for identification of glucosinolate in three kinds of cruciferous vegetable leaves was developed by using HPLC-Q-TOF-MS. Firstly, the mass spectrometry fragmentation pathways of two standard glucosinolates was studied. The bond of glucose, sulfated thiohydroximate and R-group was easily fractured and the rearrangement reaction was observed for the forming the fragment ions. Then, the structure of 8 glucosinolates including 3-(methylsulfinyl) propyl (1), 4-(methylsulfinyl) butyl (2), 3-butenyl (3), 4-hydroxybenzyl (4), 1-hydroxyindol-3-ylmethyl (5), Indol-3-ylmethyl (6), 1-methoxyindol-3-ylmethyl (7), 6-(methylsulfonyl) hexyl glucosinolate (8) were identified based on the fragmentation pathways of two standard glucosinolates, There were 4 kinds of glucosinolate (1, 2, 4, 5) in Baota Broccoli, 6 kinds of glucosinolate (2, 3, 5, 6, 7, 8) in Qinghuacai and 7 kinds of glucosinolate(1, 2, 4, 5, 6, 7) in Zise Broccoli.