Determination of maleic hydrazide and its glucosides in tobacco leaves using hydrophilic interaction liquid chromatography-tandem mass spectrometry

A hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method was established for the simultaneous determination of maleic hydrazide (MH) and its two glucosides in tobacco leaves. Ultrasonic assisted extraction of MH and its glucosides was performed using acetonitrile-methyl tert-butyl ether-water (7:10:13, volume ratio). The extraction solution was then centrifuged, and the subnatant was transferred for solvent replacement using acetonitrile. The extract in acetonitrile was then analyzed using HILIC-MS/MS. Method validation was performed, and the linear ranges for MH and MH-O-beta-D-glucoside were 5-150 mg/kg with correlation coefficients (r(2)) greater than 0.9971 for matrix-matched calibration curves. Limits of detection for MH and MH-O-beta-D-glucoside were 0.5 mg/kg and 0.3 mg/kg, and limits of quantification were 1.0 mg/kg and 0.8 mg/kg, respectively. The recoveries were in the range of 83.1%-112.3% at three spiked levels (10, 40, 80 mg/kg), with intra-day repeatability of 2.7% and 3.8%, inter-day repeatability of 8.3% and 7.1% at 40 mg/kg. The established method was used for the study of MH metabolism in tobacco leaves. By the 28th day after MH spraying, the content of MH in tobacco leaves had decreased by 80.8%, of which only 7.6% transformed to MH-glucosides. The disposition of the remainder needs to be studied.