Quantitation of γ-Glutamylcysteine-Formaldehyde Conjugate in Formaldehyde- and Oxidative Stress-Exposed Cells by Liquid Chromatography-Tandem Mass Spectrometry.

Humans are constantly exposed to formaldehyde (FA) of both exogenous and endogenous sources, and FA exposure is associated with the development of many human diseases, including cancers. Marker molecules that can provide information on exposure history and amounts will assist disease risk assessment and early interventions. To develop marker signatures of FA exposure, we explored in this study the conjugation reaction of FA with γ-glutamylcysteine (GGC), one of the precursors to glutathione biosynthesis, under physiologically relevant conditions. The results showed that the reaction produced a stable metabolite of FA, (S)-1-((((R)-2-amino-2-carboxyethyl)thio)methyl)-5-oxopyrrolidine-2-carboxylic acid (COCA). Using liquid chromatography-tandem mass spectrometry coupled to a stable isotope-dilution method, we then quantitated for the first time the formation of this novel metabolite in FA- and Fe2+-EDTA-exposed human cells. The results revealed the exposure time- and concentration-dependent formation of COCA in FA- or Fe2+-EDTA-exposed cells, suggesting that COCA may serve as a biomarker of FA and oxidative stress exposure. Furthermore, the study sheds light on a previously unknown protective role of GGC against FA and oxidative stress.