The precise identification of predominant toxic disinfection byproducts (DBPs) from disinfected water is a longstanding challenge. We propose a new acellular analytical strategy, the 'Thiol Reactome', to identify thiol-reactive DBPs by employing a thiol probe and nontargeted mass spectrometry (MS) analysis. Disinfected/oxidized water samples had reduced cellular oxidative stress responses of 46 ± 23% in Nrf2 reporter cells when preincubated with glutathione (GSH). This supports thiol-reactive DBPs as the predominant drivers of oxidative stress. This method was benchmarked using seven classes of DBPs including haloacetonitriles, which preferentially reacted with GSH via substitution or addition depending on the number of halogens present. The method was then applied to chemically disinfected/oxidized waters, and 181 tentative DBP-GSH reaction products were detected. The formulas of 24 high abundance DBP-GSH adducts were predicted, among which nitrogenous-DBPs (11) and unsaturated carbonyls (4) were the predominant compound classes. Two major unsaturated carbonyl-GSH adducts, GSH-acrolein and GSH-acrylic acid, were confirmed by their authentic standards. These two adducts were unexpectedly formed from larger native DBPs when reacting with GSH. This study demonstrated the "Thiol Reactome" as an effective acellular assay to precisely identify and broadly capture toxic DBPs from water mixtures.
Keywords: Nrf2; disinfection byproducts; high-resolution mass spectrometry; nontargeted analysis; α,β-unsaturated carbonyls.