Orbitrap Exploris Isotope Solutions: Using multiple microscans to enhance precision and accuracy for the ratios of minor isotopologues

Significance of Topic

High precision and accuracy in isotope ratio mass spectrometry (IR-MS) play a critical role in environmental studies, geochemistry, biochemistry and industrial quality control. The ability to detect minor isotopologues at low natural abundance extends our insight into metabolic pathways, provenance studies and forensic tracing of materials. Enhancing the reliability of isotope ratio measurements directly supports decision making in research and regulated industries.

Objectives and Study Overview

This technical note investigates the impact of summing multiple Orbitrap transients (microscans) prior to enhanced Fourier transform (eFT) processing on precision and accuracy of low-abundance isotope ratios. Sulfate (HSO4–) is used as a model compound to illustrate how increasing microscan counts influences signal-to-noise ratio, peak resolution and isotope ratio accuracy for 33S and 34S18O isotopologues.

Instrumentation

The workflow employs Thermo Scientific Orbitrap Exploris 480 MS equipped with:

• Electrospray ionization source (ESI)
• Advanced Quadrupole Technology (AQT) for mass filtering
• Ion-routing multipole (IRM) for storage and optional fragmentation
• Orbitrap analyzer with bell-shaped outer electrodes and central spindle electrode

Sample introduction uses either a dual syringe inlet or an in-flow injection via Vanquish Neo UHPLC.

Methodology

Each Orbitrap cycle comprises ion accumulation (injection time, IT) and transient acquisition (analysis time, AT). Summing transients from multiple microscans before eFT enhances signal intensity proportionally to the number of microscans, while noise increases only by its square root, thus boosting S/N. Isotope ratios are calculated in IsoX software and refined via R scripts, using bracketing against a reference solution to compute δ-values.

Main Results and Discussion

Experiments comparing 1–10 microscans show:

• A linear increase in peak intensity; S/N improves by √n
• Significant resolution gains for low-abundance peaks (34S18O) when microscans exceed four
• Marked improvement in accuracy of δ34S18O/M0 ratios approaching values obtained with 10 microscans
• Minimal impact on high-abundance M0 peak resolution, which remains constant

Data highlight a bimodal distribution of peak resolution for low-abundance isotopologues at low microscope counts, shifting to absorption mode with higher counts.

Benefits and Practical Applications

• Enhanced detection limits for trace isotopologues
• Improved precision of δ-value measurements in environmental and forensic applications
• Reduced data file size by combining transients into single scans
• Streamlined data processing and storage efficiency

Future Trends and Potential Applications

Continued development in Orbitrap hardware and eFT algorithms may enable real-time transient summing and adaptive microscan selection. Integration with machine learning for automated optimization could further enhance throughput. Applications may expand to site-specific isotopic analysis and coupling with separation techniques for complex mixtures.

Conclusion

Summing multiple microscans prior to eFT processing in Orbitrap Exploris mass spectrometers substantially improves S/N, resolution and isotope ratio accuracy for low-abundance species. This approach offers a robust method for high-precision IR-MS applicable across research and industrial laboratories.

References

• Cai R, Huang W, Meder M, Bourgain F, Aizikov K, Riva M, Bianchi F, Ehn M. Anal. Chem. 2022, 94, 15746–15753.
• Lange O, Damoc E, Wieghaus A, Makarov A. Enhanced FT for Orbitrap Mass Spectrometry.
• Jørgensen BB, Findlay AJ, Pellerin A. Front. Microbiol. 2019, 10, 849.
• Canfield DE et al. Science 2010, 330(6009), 1375–1378.
• Eiler J et al. Int. J. Mass Spectrom. 2017, 422, 126–142.
• Makarov A, Denisov E. J. Am. Soc. Mass Spectrom. 2009, 20, 1486–1495.
• Mueller EP, Sessions AL, Sauer PE, Weiss GM, Eiler JM. Anal. Chem. 2022, 94(2), 1092–1100.