A Time-Efficient Cyclic Ion Mobility Spectrometry−Mass Spectrometry method for the Separation and Detection of Ergot Alkaloids

The diversity of ergot alkaloids (EAs) isomers poses significant challenges for traditional analytical methodologies. By adding an orthogonal rapid gas-phase separation step, Ion mobility mass spectrometry (IM-MS) enables faster workflows and higher throughput without sacrificing data quality. However, its application in this context remains limited. We hypothesize that high resolution IM–MS can shorten chromatographic analysis times for natural compounds while maintaining or even enhancing separation performance. In this study, we developed a high-throughput LC-cyclic IMS (cIMS) methodology for the analysis of 28 EAs including 9 isomeric pairs, which comprises epimers, conformational isomers and constitutional isomers, within 4 min by leveraging the additional dimension of separation provided by the cyclic ion mobility with high-resolution mass spectrometry. A 60% reduction in analysis time has been shown using LC-cIM-MS compared to LC-MS while keeping selectivity of the method. Ion mobility was particularly effective for the separation of small (<300 Da) and more polar conformational and constitutional isomers with peak-to-peak resolution, outperforming chromatographic separation. These results highlight the importance of complementary separation dimensions for the analysis of complex mixtures of natural compounds. The method developed in study allows to separate currently regulated EAs epimers within 4-minutes LC-cIM-MS run. • Cyclic ion mobility enables rapid separation of ergot alkaloids isomers • A 4-min LC–cIMS method achieves reduction of time without loss of selectivity • Eight of nine ergot alkaloid isomeric pairs are baseline-resolved by multipass cIMS • Ion mobility outperforms chromatography for small and polar ergot alkaloid isomers • The most extensive CCS database for ergot alkaloids (28 compounds) is reported