Uniting targeted (Zeno MRM-HR) and untargeted (SWATH) LC-MS in a single run for sensitive high-resolution exposomics.

Traditionally, chemical exposure has been assessed by low-resolution mass spectrometry via targeted ap-proaches due to the typically extremely low concentration of such compounds in biological samples. Never-theless, untargeted approaches are now becoming a promising tool for a broader investigation of the expo-some, covering additional compounds, their biotransformation products and possible metabolic alterations (metabolomics). However, despite broad compound coverage, untargeted metabolomics still underperforms in ultra-trace biomonitoring analysis. To overcome these analytical limitations, we present the development of the first combined targeted/untargeted LC-MS method, merging MRM-HR and SWATH experiments in one analytical run, making use of the Zeno technology for improved sensitivity. MRM transitions were optimized for 135 highly diverse toxicants including mycotoxins, plasticizers, PFAS, personal care products ingredients and industrial side products as well as potentially beneficial xenobiotics such as phytohormones. As a proof of concept, standard reference materials of human plasma (SRM 1950) and serum (SRM 1958) were ana-lyzed with both, Zeno MRM-HR + SWATH and SWATH-only methodologies. Results demonstrated a signifi-cant increase in sensitivity represented by the detection of lower concentration levels in spiked SRM materi-als (mean value: 2.2x and 3x more sensitive for SRMs 1950 and 1958, respectively). Overall, detection fre-quency was increased by 45% (from 22 to 40 positive detections) in MRM-HR+SWATH mode compared to the SWATH-only. This work presents a promising avenue for addressing the outstanding challenge in the small-molecule omics field: finding balance between high sensitivity and broad chemical coverage. It was demonstrated for exposomic applications but might be transferred to lipidomics and metabolomics work-flows.