Stable isotope labeling for detection of ozonation byproducts in effluent organic matter with FT-ICR-MS

Despite effluent organic matter (EfOM) being a major consumer of ozone during wastewater treatment, little is known about ozonation byproducts (OBPs) produced from EfOM. To unambiguously identify OBPs, heavy ozone was used to ozonate EfOM, resulting in ¹⁸O labeled and unlabeled OBPs. Labeled OBPs mostly represent a single ¹⁸O transfer and were classified as either direct or indirect OBPs based on the ¹⁸O/¹⁶O intensity ratios of the isotopologues. Of the 929 labeled OBPs, 84 were unequivocally classified as direct OBPs. The remainder suggest a major contribution by indirect, hydroxyl radical induced formation of OBPs in EfOM. Overall, labelled OBPs possess a low degree of unsaturation and contributed most to OBP peak intensity - marking them as potential end products. A few direct and indirect OBPs with high peak intensity containing ¹⁸O and heteroatoms (N, S) were fragmented with CID FT-ICR-MS/MS and screened for indicative neutral losses carrying heavy oxygen. The neutral loss screening was used to detect the ¹⁸O location on the OBP and indicate the original functional group in EfOM based on known reaction mechanisms. We identified sulfoxide and sulfonic acid functional groups in selected OBPs - implying the presence of reduced sulfur in EfOM molecules - while no evidence for nitrogen containing functional groups reacting with ozone was found.