Novel carbonylomics with stable isotope-coded derivatization for non-targeted analysis of reactive carbonyl species...

Reactive carbonyl species (RCS), both carcinogenic and widespread in the environment, disrupt cell function through biomolecular modifications. However, to date, the study of RCS has largely been via targeted analysis. Herein, we introduce a novel carbonylomics workflow integrating liquid chromatography-high-resolution mass spectrometry (LC-HRMS) with stable isotope-coded derivatization (SICD) using d₀- and d₃-2,4-dinitrophenylhydrazine (DNPH) to perform the non-targeted analysis of RCS in cooking oils. Our method enables comprehensive detection of RCS, entirely relying on examining four characteristic features of RCS-DNPH derivatives with well-defined instrument settings, while SICD enhances specificity by reducing false positives. Applying this workflow to soybean oil (SBO) and palm oil (PO) before and after heating identified numerous known and unknown RCS, with SBO exhibiting a greater variety in RCS (increasing from 23 to 129 ions vs. 18-75 ions for PO) and up to ∼11-fold greater peak intensities for shared RCS, indicating higher susceptibility to thermal oxidation. Among them, trans,trans-2,4-undecadienal and 2,3-octanedione were, for the first time, exclusively identified in oxidized SBO. Additionally, the approach was successfully applied to human urine, demonstrating broader applicability to biological matrices. Given the widespread presence of RCS, originating from environmental and endogenous sources, these findings highlight the utility of carbonylomics in elucidating RCS formation and identifying unknown toxicants, providing a critical tool for investigating food safety and other public health concerns related to RCS exposure.