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Global profiling of the muscle metabolome: method optimization, validation and application to determine exercise-induced metabolic effects

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Abstract

Skeletal muscle represents a crucial metabolic organ in the body characterized by a tremendous metabolic plasticity and the ability to influence important metabolic events elsewhere in the body. In order to understand the metabolic implications of skeletal muscle, it is imperative to characterize the metabolites within the tissue itself. In this work we aimed at developing a suitable analytical pipeline to analyze the metabolome of muscle tissue. Methanol/chloroform/water at neutral pH was selected as the method of choice for metabolite extraction prior to analysis by chromatographic-mass spectrometry systems in five different platforms covering a relevant part of the muscle metabolome: organic acids, amines, nucleotides, coenzymes, acylcarnitines and oxylipins. This analytical pipeline was extensively validated and proved to be robust, precise, accurate and biologically sound. The capability of our analytical method to capture metabolic alterations upon challenges was finally tested using a small proof-of-concept study involving an exercise intervention. Mild but consistent metabolic patterns were observed, allowing the discrimination between non-exercised and exercised muscles. Despite the low numbers of subjects enrolled in this study (5), these results are indicative that our method is suitable to determine intervention effects in skeletal muscle tissue whenever applied to adequately powered and well characterized studies.

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Acknowledgments

This study was (co)financed by the Netherlands Metabolomics Centre (NMC) which is part of the Netherlands Genomics Initiative/Netherlands Organization for Scientific Research. We gratefully acknowledge Sietse-Jan Koopmans (Wageningen University) for providing pig muscle biopsies. Raymond Ramaker, Olga Willemsen, Theo van der Kaaij, Ayovi Almeida, Sabine Bos and Jan Steinz (NMC/Leiden University) for MS measurements and data preprocessing. Margriet Hendriks (NMC/Leiden University) for her input regarding mixed models and Boyd Rotteveld for the extraction efficiency experiments performed during his bachelor internship at the Analytical Biosciences Department (Leiden University).

Conflict of Interest

All authors declare that they have no conflict of interest.

Animal Rights Statement

All institutional and national guidelines for the care and use of laboratory animals were followed.

Informed Consent Statement

All procedures followed were in accordance with the ethical standards of the responsible committee on human experimentation (institutional and national) and with the Helsinki Declaration of 1975, as revised in 2000. Informed consent was obtained from all patients for being included in the study.

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Authors and Affiliations

  1. Netherlands Metabolomics Centre, Leiden University, Einsteinweg 55, 2300 RA, Leiden, The Netherlands

    Rodrigo D. A. M. Alves, Adrie D. Dane, Amy Harms, Katrin Strassburg, Ruud Berger, Thomas Hankemeier & Rob J. Vreeken

  2. Department of Analytical Biosciences, Leiden University, Einsteinweg 55, 2300 RA, Leiden, The Netherlands

    Rodrigo D. A. M. Alves, Adrie D. Dane, Amy Harms, Katrin Strassburg, Ruud Berger, Thomas Hankemeier & Rob J. Vreeken

  3. Department of Biotechnology, Kluyver Centre for Genomics of Industrial Fermentation, Delft University of Technology, Julianalaan 67, 2628 BC, Delft, The Netherlands

    Reza Maleki Seifar

  4. Department of Human Movement Sciences, NUTRIM School for Nutrition, Toxicology and Metabolism, Maastricht University Medical Centre, Maastricht, The Netherlands

    Lex B. Verdijk

  5. Division of Human Nutrition, Wageningen University, Dreijenlaan 2, 6703 HA, Wageningen, The Netherlands

    Sander Kersten

  6. Netherlands Nutrigenomics Centre, Top Institute Food and Nutrition, P.O. Box 557, 6700 AN, Wageningen, The Netherlands

    Sander Kersten

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  1. Rodrigo D. A. M. Alves
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  2. Adrie D. Dane
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  3. Amy Harms
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Correspondence to Rodrigo D. A. M. Alves.

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Alves, R.D.A.M., Dane, A.D., Harms, A. et al. Global profiling of the muscle metabolome: method optimization, validation and application to determine exercise-induced metabolic effects. Metabolomics 11, 271–285 (2015). https://doi.org/10.1007/s11306-014-0701-7

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