The invention discloses a precolumn derivatization-UPLC(ultra performance liquid chromatography)-ESI(electronic spray ion)+-MS / MS (mass spectrometry) detection method of glyphosate and glufosinate-ammonium pesticide residue in tea, sodium borate is used as a buffer salt, a relatively pure target derived product can be obtained by derivatization of glyphosate and glufosinate-ammonium and FMOC-Cl and then ultra performance liquid chromatography (UPLC) washing out and separation, and response signals can be detected by tandem mass spectrometry (MS-MS). According to the method, competition between the two pesticide residue derivatization can be eliminated, the detection effect caused by matrix effect can be effectively weakened, and the derivatization time is shortened. By experimental verification, the detection method of glyphosate and glufosinate-ammonium pesticide (herbicide) residue in tea has the characteristics of being efficient, fast and accurate, good in sensitivity and the like, is applicable in detection of the glyphosate and glufosinate-ammonium pesticide residue in tea samples of sundried green crude tea, fermented tea and substitutional tea (green food substitutional tea of agricultural industry standards of NY / T2140-2012 of the people's Republic of China) and finished tea (brick tea, Tuo tea, melon-shaped tea, cake tea, scattered tea and bagged tea and the like).