Biology subjects

Bird, S. S.

Publications and source records attributed to Bird, S. S..

2 recordsLinked to original sources

bioRxiv2026

Full Scan enhanced Dynamic Range MS improves metabolite coverage and cancer cell-line discrimination in untargeted metabolomics

Metabolite detection with mass spectrometry (MS) in untargeted metabolomics is limited by the wide concentration range of metabolites, where high-abundance signals dominate MS1 scans and suppress detection of low-abundance features. This reduces metabolite coverage and obscures biologically relevant signals, particularly in complex cellular systems. Full Scan enhanced Dynamic Range (eDR) MS addresses these limitations by partitioning the MS1 mass range into multiple subscans and mass windows, reducing saturation effects from dominant ions. Here, we systematically evaluate different eDR acquisition strategies for untargeted metabolomics. Across four hepatocellular carcinoma cell lines, Full Scan eDR MS increased detectable features up to [~]3.5-fold compared to Full Scan MS. Among equidistant window configurations, 12 windows yielded the highest feature count and broadest dynamic range, while custom window distributions further improved detection in ion-dense regions. In particular, allocating smaller window sizes to the low m/z region selectively increased detection of low-mass features while preserving performance for higher mass ions. Full Scan eDR MS also improved data quality, reducing variation and increasing signal-to-noise ratios, especially for low-abundance metabolites. MS2 coverage and metabolite identifications increased substantially, resulting in unique detection of cancer-relevant metabolites. Importantly, the increased depth of metabolite detection enabled improved discrimination between cancer cell lines, supporting deeper interrogation of metabolic heterogeneity. Overall, these results establish Full Scan eDR MS as a flexible strategy to improve sensitivity and metabolome coverage in untargeted metabolomics. Customization of window size and distribution enable targeted expansion of dynamic range within predefined mass regions, allowing MS acquisition to be tailored to sample complexity and metabolites of interest.

biochemistry↗

bioRxiv2026

Nanoflow ion-pairing LC-MS for ultra-low-input polar metabolomics and isotope tracing

Low-input and single-cell metabolomics remain constrained by the poor retention of polar metabolites in conventional reversed-phase nanoflow LC-MS workflows. Here, we establish nanoflow tributylamine (TBA) ion-pairing LC-MS as a platform for ultra-low-input polar metabolomics and stable isotope tracing. By adapting an analytical-flow TBA ion-pairing method to the nanoflow scale, this workflow extends the sensitivity and inline concentration advantages of nanoflow chromatography to charged metabolites involved in central carbon metabolism. Using mouse liver metabolite extracts, we show that the nanoflow method preserves chromatographic retention and separation of chemically diverse metabolite classes, including adenine nucleotides, nucleotide cofactors, TCA cycle intermediates, acyl-CoAs, and bile acid isomers. Despite loading 20-fold less tissue-equivalent material on column, nanoflow LC-MS produced higher signal intensity than the analytical-flow method for many metabolites. Across representative compounds, the nanoflow workflow reduced the biomass required for detection by approximately 20- to >600-fold, with pronounced gains for low-abundance metabolites such as NADPH and acetyl-CoA. TBA ion-pairing also enabled trap-and-elute nanoflow analysis of retained polar metabolites from single-cell-equivalent inputs. ATP was detected from one cell equivalent using both full-scan and targeted parallel reaction monitoring acquisition, with targeted acquisition further increasing signal over blank. Finally, we applied the workflow to stable isotope tracing in uniformly labeled 13C-glucose-treated cells. 13C-labeled ATP isotopologues were detectable from single-cell-equivalent input, and targeted acquisition improved isotopologue measurement near the detection limit. Together, these results demonstrate that nanoflow TBA ion-pairing LC-MS enables retained, high-sensitivity analysis of polar metabolites from ultra-low inputs and provides a foundation for extending central carbon metabolite analysis and isotope tracing toward single-cell-scale applications.

biochemistry↗