Systematic Revealing of Plant-Derived Nerve Agent Exposure Markers by In-Source Fragmentation–Enabled HRMS Nontargeted Screening

Identifying diverse, uncharacterized plant-derived nerve agent adducts is a formidable challenge for environmental exposure confirmation and forensic attribution. We developed a fragmentation pattern–driven nontargeted screening strategy integrating in-source fragmentation with Full MS, all-ion fragmentation (AIF), and neutral loss (NL)–triggered ddMS 2 acquisition. By utilizing agent-specific neutral losses and diagnostic fragment ions, 256 tentative adduct-related features were identified across four nerve agents and four plant species, establishing an expansive repository of previously unreported candidate markers. Structural characteristics and preferential adduction sites were elucidated through integrated interpretation of high-resolution MS data, fragmentation pathways, and density functional theory calculations. Notably, 9 features exhibited conservation across plant species, while 11 features remained detectable at low-level exposure in Arabidopsis thaliana, highlighting their potential as viable markers. This strategy expands the detectable chemical space for plant-derived nerve agent adducts and provides a transferable framework for environmental exposure assessment and emerging contaminants research.