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Real-time 2D separation by LC × differential ion mobility hyphenated to mass spectrometry

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Abstract

The liquid chromatography–mass spectrometry (LC-MS) analysis of complex samples such as biological fluid extracts is widespread when searching for new biomarkers as in metabolomics. The success of this hyphenation resides in the orthogonality of both separation techniques. However, there are frequent cases where compounds are co-eluting and the resolving power of mass spectrometry (MS) is not sufficient (e.g., isobaric compounds and interfering isotopic clusters). Different strategies are discussed to solve these cases and a mixture of eight compounds (i.e., bromazepam, chlorprothixene, clonapzepam, fendiline, flusilazol, oxfendazole, oxycodone, and pamaquine) with identical nominal mass (i.e., m/z 316) is taken to illustrate them. Among the different approaches, high-resolution mass spectrometry or liquid chromatography (i.e., UHPLC) can easily separate these compounds. Another technique, mostly used with low resolving power MS analyzers, is differential ion mobility spectrometry (DMS), where analytes are gas-phase separated according to their size-to-charge ratio. Detailed investigations of the addition of different polar modifiers (i.e., methanol, ethanol, and isopropanol) into the transport gas (nitrogen) to enhance the peak capacity of the technique were carried out. Finally, a complex urine sample fortified with 36 compounds of various chemical properties was analyzed by real-time 2D separation LC×DMS-MS(/MS). The addition of this orthogonal gas-phase separation technique in the LC-MS(/MS) hyphenation greatly improved data quality by resolving composite MS/MS spectra, which is mandatory in metabolomics when performing database generation and search.

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Acknowledgments

EV and GH wish to thank M. Himmelsbach (visiting scientist in the LSMS group) for performing the FT-ICR-MS experiments as well as T. Bruderer (Ph.D. student in the LSMS group). GH also acknowledges R. Falchetto (Novartis) for his support with the FT-ICR-MS instrument. EV and GH would like to acknowledge Dionex (F. Steiner, M. Martin, and F. Sabini) for providing the UltiMate 3000 Dual RSLC system. Finally, the authors would also thank B.B. Schneider (AB Sciex) for his support with the DMS device. D. Simmons and L. Burton/R. Bonner (AB Sciex) for the MS resolution calculator and PeakView software support, respectively.

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Authors and Affiliations

  1. School of Pharmaceutical Sciences, University of Geneva, University of Lausanne, Life Sciences Mass Spectrometry, Quai Ernest-Ansermet 30, 1211, Geneva 4, Switzerland

    Emmanuel Varesio & Gérard Hopfgartner

  2. AB Sciex, 71 Four Valley Drive, Concord, ON, L4K 4V8, Canada

    J. C. Yves Le Blanc

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  1. Emmanuel Varesio
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  2. J. C. Yves Le Blanc
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  3. Gérard Hopfgartner
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Correspondence to Emmanuel Varesio.

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Published in the special issue Analytical Sciences in Switzerland with guest editors P. Dittrich, D. Günther, G. Hopfgartner, and R. Zenobi.

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Varesio, E., Le Blanc, J.C.Y. & Hopfgartner, G. Real-time 2D separation by LC × differential ion mobility hyphenated to mass spectrometry. Anal Bioanal Chem 402, 2555–2564 (2012). https://doi.org/10.1007/s00216-011-5444-y

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  • DOI: https://doi.org/10.1007/s00216-011-5444-y

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