PurposeThe detection of hydrolysis products of Novichok agents in biological samples from victims is important for confirming exposure to these agents. However, Novichok agents are new class of nerve agent and there have been only few reports on analyses of Novichok agent degradation products. Here, we developed hydrophilic interaction liquid chromatography (HILIC)–tandem mass spectrometry (MS/MS) methods to detect Novichok agent degradation products in human urine with simple pretreatment and high sensitivity.MethodsA Poroshell 120 HILIC-Z column was used to analyze six Novichok agent degradation products. For urine samples, we used a simple pretreatment method, which consisted of deproteinization with acetonitrile and microfiltration. We calculated the pKavalues of the OH groups, the log P values, and the molecular weights to investigate the difference in chromatographic behaviors of the Novichok agent degradation products and the degradation products of conventional nerve agents.ResultsSix Novichok agent degradation products, includingN-(bis-(diethylamino)methylidene)-methylphosphonamidic acid (MPGA), which could not be detected by our previous method, could be analyzed with sufficient peak shape and mutual separation. The detection limits of six Novichok agent degradation products were sufficiently low (1–50 ng/mL) and the calibration curves showed sufficient linearity. The physicochemical parameters of Novichok agent degradation products were different from those of conventional nerve agent degradation products, and this explains the difference in chromatographic behaviors.ConclusionSix Novichok agent degradation products were successfully analyzed by HILIC–MS/MS. Due to the absence of a derivatization step, throughput performance was higher than our previous derivatization-liquid chromatography–MS/MS method.Supplementary InformationThe online version contains supplementary material available at 10.1007/s11419-022-00656-4.