Mass Spectrometric Identification and Isomer Differentiation of Novichok‐Related Ethanimidamides

Chemical warfare agent (CWA) analysis faces a significant challenge when degradation or precursor compounds lack reference spectra in databases. This study focuses on three structural isomers, N-methyl-N-propylethanimidamide, N-methyl-N-isopropylethanimidamide, and N,N-diethylethanimidamide, which are relevant hydrolysis by-products/precursors of Novichok nerve agents (Schedule 1.A.13/1.A.14) yet are absent from standard spectral libraries. All three isomers were synthesized and then comprehensively characterized by gas chromatography-electron ionization mass spectrometry (GC-EI-MS), gas chromatography-chemical ionization mass spectrometry (GC-CI-MS), and liquid chromatography-high-resolution mass spectrometry (LC-HRMS). Distinctive fragmentation signatures were identified for each isomer. Trimethylsilyl derivatization of the ethanimidamides enhanced GC performance and revealed additional diagnostic ions. LC-HRMS results confirmed the fragment formulas and allowed unambiguous differentiation based on key ion intensity ratios. This work provides the first systematic mass spectral data for these Novichok-related ethanimidamides. The value of these reference spectra was demonstrated in the 57^th OPCW Proficiency Test (PT-57): an unknown spiking chemical was successfully identified as N-methyl-N-propylethanimidamide by comparison to the generated spectra. The study thus delivers new analytical capabilities for CWA forensics-enabling confident identification of challenging isomeric degradation products through a combination of tailored derivatization, multimode mass spectrometry, and expert interpretation of fragmentation pathways.