A hydrophilic interaction liquid chromatography tandem mass spectrometry-based method for the simultaneous determination of maleic hydrazide (MH) and its two glucosides in tobacco leaves was developed. The method was based on the synthesis of MH-O-ß-D-glucoside and the identification of MH-O-ß-D-glucoside and MH-N-ß-D-glucoside in tobacco leaves. The developed method was compared with the reference methods (including the tobacco industry standard YCT405.5-2011) and the sample preparation of this method was found to be simpler, more efficient, safer, and with higher throughput than those of the reference methods. This method was also found to have shorter instrument analysis time and higher sensitivity than that of the reference methods. The distribution of MH and its glucosides in tobacco leaves can also be determined with this method. Method validation was performed and correlation coefficients (r²) of MH and MH-O-ß-D-glucoside were in the range of 0.9971-0.9972. Recoveries were in the range of 83-112 %, with intra-day repeatability 2.7-3.8 %, inter-day repeatability 7.1-8.3 %, limits of quantitation 0.8~1.0 mg/kg, and limits of detection 0.3~0.5 mg/kg. The developed method was used for the study of the metabolism of MH in tobacco leaves. At day 28 after MH spraying, content of MH in tobacco leaves decreased by 80.8 %, of which only 7.6 % was transformed to MH-glucosides. The transformation of the remainder of the MH needs to be studied.