Incubation of human plasma with the tear gas 2-chloroacetophenone (CN) led to the formation of acetophenone (AcPhen)-adducts with the small plasma thiols glutathione, homocysteine, and cysteine persulfide. The AcPhen-adducts were either bound to an organic thiol-group or to an organic persulfide. The adducts were detected with a mass spectrometry-based method working in the parallel reaction monitoring (PRM) mode.

ABSTRACT

As the tear gas of the highest toxicity, 2-chloroacetophenone (CN) poses a potential threat for exposed individuals. Several fatality cases following exposure to CN are documented, but an unambiguous identification of CN exposure is still missing. Thus, we herein present the identification of in vitro reaction products between CN and endogenous molecules useful as biomarkers. After incubation of human plasma with CN, diverse acetophenone (AcPhen)-adducts were formed with the small molecule thiols homocysteine (HCys), glutathione (GSH), and cystine (Cys-Cys). All adducts were detected by microbore liquid chromatography-electrospray ionization high-resolution tandem mass spectrometry (μLC-ESI MS/HRMS) working in the parallel reaction monitoring (PRM) mode and were characterized as potential biomarkers of CN exposure. Time- and concentration-dependent adduct formations were investigated, and the stability of AcPhen-adducts in plasma during 4 freeze-and-thaw cycles and in the autosampler was tested. The limit of identification (LOI) of identified AcPhen-adducts in vitro was found at about 6 μM (concentration of CN in plasma) but showed quite limited in vitro stability. The AcPhen-adducts herein presented might be beneficial for future studies addressing CN exposure in vivo.