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Development of a novel method for quantification of sterols and oxysterols by UPLC-ESI-HRMS: application to a neuroinflammation rat model

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Abstract

Cholesterol and oxysterols are involved as key compounds in the development of severe neurodegenerative diseases and in neuroinflammation processes. Monitoring their concentration changes under pathological conditions is of interest to get insights into the role of lipids in diseases. For numerous years, liquid chromatography coupled to mass spectrometry has been the method of choice for metabolites identification and quantification in biological samples. However, sterols and oxysterols are relatively apolar molecules poorly adapted to electrospray ionization (ESI). To circumvent this drawback, we developed a novel and robust analytical method involving derivatization of these analytes in cholesteryl N-4-(N,N-dimethylamino)phenyl carbamates under alkaline conditions followed by ultra-performance liquid chromatography–high resolution mass spectrometry analysis (UPLC-HRMS). Optimized UPLC conditions led to the separation of a mixture of 11 derivatized sterols and oxysterols especially side chain substituted derivatives after 6 min of chromatographic run. High sensitivity time-of-flight mass analysis allowed analytes to be detected in the nanomolar range. The method was validated for the analysis of oxysterols and sterols in mice brain in respect of linearity, limits of quantification, accuracy, precision, analyte stability, and recovery according to the Food and Drug Administration (FDA) guidelines. The developed method was successfully applied to investigate the impact of lipopolysaccharide (LPS) treatment on the rat cerebral steroidome.

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Acknowledgment

The post-doctoral position of S.A. was funded by ANR Chol AD (French-Canadian Cooperation-2010-MALZ-10303) and the PhD position of M.G. by Technologie Servier (Orléans, France). O.L. is indebted to Fondation pour la Recherche Médicale, Région-Île-de-France and Centre National de la Recherche Scientifique for their financial support. We thank Fathia Djelti (INSERM U745) for providing the C57BL/6 mice brain tissue.

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Author notes

  1. Mathieu Gaudin

    Present address: Biomolecular Medicine, Department of Surgery and Cancer, Faculty of Medicine, Imperial College London, SW7 2AZ, London, UK

Authors and Affiliations

  1. Chimie-Toxicologie Analytique et Cellulaire, EA 4463, Université Paris Descartes, Sorbonne Paris Cité, Faculté des Sciences Pharmaceutiques et Biologiques, 75006, Paris, France

    Sophie Ayciriex, Anne Regazzetti, Mathieu Gaudin, Delphine Dargère, France Massicot, Nicolas Auzeil & Olivier Laprévote

  2. Division métabolisme, Technologie Servier, 45000, Orléans, France

    Mathieu Gaudin

  3. Centre de Recherche de Gif, Institut de Chimie des Substances Naturelles, CNRS, Avenue de la Terrasse, 91198, Gif-sur-Yvette Cedex, France

    Mathieu Gaudin

  4. UMR 8638 CNRS, Université Paris Descartes, Sorbonne Paris Cité, Faculté des Sciences Pharmaceutiques et Biologiques, 75006, Paris, France

    Elise Prost

  5. Service de Toxicologie Biologique, Hôpital Lariboisière, 4 rue Ambroise Paré, 75475, Paris cedex 10, France

    Olivier Laprévote

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  1. Sophie Ayciriex
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Correspondence to Nicolas Auzeil.

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Sophie Ayciriex and Anne Regazzetti contributed equally to this work.

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Ayciriex, S., Regazzetti, A., Gaudin, M. et al. Development of a novel method for quantification of sterols and oxysterols by UPLC-ESI-HRMS: application to a neuroinflammation rat model. Anal Bioanal Chem 404, 3049–3059 (2012). https://doi.org/10.1007/s00216-012-6396-6

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  • DOI: https://doi.org/10.1007/s00216-012-6396-6

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