Conference Agenda

Isotopocule analysis is a transformative tool for environmental research, offering insights into isotopic distributions across many molecules, even as simple as oxyanions such as nitrate and sulfate. Traditional isotopic methods using isotope ratio mass spectrometry (IRMS) often rely on labor-intensive sample conversions, which can obscure site-specific isotopic information. This presentation introduces the application of electrospray ionization-Orbitrap IRMS, a robust technique for direct isotopocule analysis, circumventing the limitations of classical approaches.

The methodology employs low-flow solution inlets via dual syringe pumps or HPLC autosamplers, ensuring high sensitivity (down to 1 nmol nitrate) and tolerance to matrix interferences. The workflow includes advanced data processing with the open-source R package isoorbi, facilitating isotope ratio calculations, delta value determination, and isotopocule-specific visualizations. Demonstrated applications include precise nitrate isotope analysis from environmental samples, highlighting potential proxies for ecological and agricultural processes.

This talk will showcase method validation using reference materials, comparison with established IRMS protocols, and the potential for unattended long-term sample throughput. The integration of computational tools with Orbitrap technology underlines its promise for advancing isotopic studies in atmospheric, aquatic, and terrestrial systems, paving the way for novel insights into environmental dynamics.

For decades, isotope geochemists have quantified elemental isotope ratios using mass spectrometry. Traditional hard-ionisation, low mass resolution methods such as IRMS or ICP-MS break apart sample molecules for analysis, often destroying the sample’s intramolecular isotopic detail.

In contrast, the recently repurposed, bioanalytical Electrospray-Orbitrap mass spectrometer employs soft ionisation and high-resolution mass analysis to directly measure isotopologue ratios on intact molecules. This innovation heralds a new era in stable isotope analysis, advancing beyond traditional elemental isotope ratios, to enable isotopologue-specific measurements, including isotope clumping quantifications.

We showcase the Electrospray-Orbitrap’s unique capability to simultaneously quantify an extended range of singly- and doubly-substituted nitrate and sulphate isotopologue ratios, using nanomole amounts of ice core and aerosol sample. This breakthrough enables the first-ever clumped isotopologue analysis of natural oxyanion samples– offering crucial insights into Earth’s past atmospheric oxidative capacity, as well as representing a potential new tool in the routine isotopic analysis of oxyanions.

Despite its potential, questions remain regarding the accuracy, precision and robustness of Electrospray-Orbitrap isotopic measurements compared to the established IRMS and ICP-MS methods. To validate its performance, we provide a comprehensive comparison of isotopic measurements obtained from the same alpine ice core and Antarctic aerosol samples using both Electrospray-Orbitrap and traditional techniques.

High precision natural abundance isotope ratio analysis of complex organic molecules has been an important tool in understanding environmental and planetary processes throughout the last decades. State of the art workflows often require the conversion of complex organic molecules to low molecular weight gases to perform a high precision isotope quantification using sector field MS. Recently, the high-resolution accurate-mass of Thermo Scientific^TM Orbitrap^TM MS has been shown to enable simultaneous, high precision analysis of intact isotopocules with isotopic substitutions from multiple elements, which bears new potential for gathering intramolecular isotopic information.

Here we highlight improvements in ESI-Orbitrap IRMS for the analysis of organic molecules using ¹³C, ²H, ¹⁸O and ¹⁵N substituted isotopocules of caffeine and vanillin as a model. Orbitrap results are compared and benchmarked against international reference materials and results obtained by sector field MS revealing accuracies and precisions down to <0.5 ‰ for carbon, 2 ‰ for oxygen, 10 ‰ for hydrogen and 1 ‰ for nitrogen isotopocule ratios. The presented data will show how challenges in Orbitrap-IRMS for complex organic molecules can be overcome by dedicated referencing and calibration procedures, highlighting the need for a wider availability of isotopically characterized reference materials. Performing efficient sample/standard comparison sets requirements on the sample introduction techniques. While many different techniques have been developed for reference introduction in the gas phase, liquid sample and reference introduction approaches still lack the efficiency and robustness of their gaseous counterparts. To this end, we have explored novel sample introduction techniques and online liquid chromatography for better automation and matrix tolerance specifically targeting high precision natural abundance isotopocule ratio quantification in complex organic mixtures. Among other application the developed methods aim to improve the analysis of isotope and isotopocule ratios of inorganic oxyanions and small organic molecules in water and soil samples.