Electrospray-Orbitrap Analysis of Nitrate and Sulfate Isotopologues in Natural Samples: Performance and Constraints.

Traditional isotope ratio mass spectrometers are not optimally designed to analyze nonvolatile molecular solutes such as oxyanions or organics. Necessary sample conversion steps, often demanding micromole-scale samples, break chemical bonds and limit oxyanion analyses to elemental isotope ratios. Recent studies enthusiastically promote soft electrospray ionization coupled to high-resolution Orbitrap mass spectrometry (ESI-Orbitrap-MS) for intact-molecule isotopologue analysis. This combination uses widely accessible instrumentation to provide simultaneous multi-isotopologue and clumped isotope analyses using tens of nanomoles of dissolved sample. However, its isotopic capabilities have been evaluated primarily through technical assessments rather than on natural samples, despite the need for matrix sensitivity assessments and robust isotopic calibrations. Here, we establish a framework for natural-sample ESI-Orbitrap-MS isotopologue analysis of nitrate and sulfate, addressing matrix effects, isotopologue calibration, and instrumental stability. We demonstrate strong sensitivity to contaminants at concentrations 1000 times below target analytes and strong temperature-correlated drift over >30 min, highlighting the enhanced need for rigorous sample purification to suppress matrix effects, sample-reference bracketing to overcome drift, and isotopologue calibration to reduce biases and robustly quantify uncertainties. We achieve 0.4-1.3‰ precision and accurate measurements of δ15N-NO3-, δ18O-NO3-, Δ17O-NO3-, δ34S-SO42-, and Δ17O-SO42- relative to traditional techniques, consuming only 20 nmol nitrate or 10 nmol sulfate. We resolve relative nitrate clumped isotope values in ice core samples (Δ15N18O-NO3-: ±2.3‰; Δ18O18O-NO3-: ±3.3‰) yet observe no correlations with bulk nitrate isotopes. These results establish practical guidelines for applying ESI-Orbitrap-MS to isotopologue and clumped isotope analysis of trace analytes in challenging sample matrices under real-world conditions.