Multi reflecting TOF MS approaching resolution of 1,000,000 in a wide mass range

Abstract

Time-of-flight mass spectrometry (TOFMS) is a cornerstone of analytical chemistry, renowned for its exceptional combination of speed, sensitivity, resolution, and mass accuracy. Multi-reflecting TOFMS (MRT) instruments significantly enhance TOFMS resolution by repeatedly folding and extending the ion trajectory. Historically, MRT instruments have achieved resolutions of up to 200,000 across a wide mass range. While higher resolutions (reaching 1 million) have been demonstrated in trajectory looping configurations, these are associated with a narrower mass range that shrinks inversely with the number of loops. This research presents an advanced MRT instrument that overcomes these limitations by achieving both a resolving power of approximately 1 million (R ≈ 1,000,000) and a wide mass range, not limited by the analyzer design. This was achieved through numerous instrumental enhancements, and primarily by extending the flight path to approximately 100 m, corresponding to a flight time of 2.55 ms for m/z = 1000 Th ions. Prolonged flight times inevitably reduce the duty cycle of the orthogonal accelerator. In one practical acquisition method, encoded frequent pulsing (EFP) at an average pulsing rate of 50 kHz, recovered the OA duty cycle to 10 %. This enabled the instrument to record high-resolution MS/MS spectra at a rate of 10 Hz and identifying peptides in a concentration range of 10⁻⁸ to 10⁻⁴ M, enabling high-throughput MS/MS analysis. At higher sample concentrations, the instrument is sensitive to space charge effects within the analyzer, which start affecting resolution as early as 20 ions per packet. At 50 kHz EFP method, this limit corresponds to 10⁶ ion/peak/s and allows ion fluxes up to 10⁸ ion/s in the case of complex spectra containing numerous peaks. The major part of this publication is focused on characterizing the ultimate performance of the prototype MRT. To minimize spectral artifacts, most characterization experiments were conducted using a rare pulsing method (push and wait) at a pulsing rate of 500 Hz. Extended spectral acquisition times allowed for the accumulation of sufficient ion statistics, enabling the exploration of fine details within MS/MS spectra of peptides. The achieved standard deviation of mass accuracy was approximately 100 ppb over a dynamic range of 10⁵. This research comprehensively characterizes the high-resolution MRT instrument, focusing on its capabilities and limitations. While analytical applications are not discussed in this paper, the presented data provides a solid foundation for understanding the instrument's potential.