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Microbial Metabolomics pp 225–239Cite as

Determining the Mode of Action of Antimalarial Drugs Using Time-Resolved LC-MS-Based Metabolite Profiling

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1859))

Abstract

Methods for assessing the mode of action of new antimalarial compounds identified in high throughput phenotypic screens are needed to triage and facilitate lead compound development and to anticipate potential resistance mechanisms that might emerge. Here we describe a mass spectrometry-based approach for detecting metabolic changes in asexual erythrocytic stages of Plasmodium falciparum induced by antimalarial compounds. Time-resolved or concentration-resolved measurements are used to discriminate between putative targets of the compound and nonspecific and/or downstream secondary metabolic effects. These protocols can also be coupled with 13C-stable-isotope tracing experiments under nonequilibrative (or nonstationary) conditions to measure metabolic dynamics following drug exposure. Time-resolved 13C-labeling studies greatly increase confidence in target assignment and provide a more comprehensive understanding of the metabolic perturbations induced by small molecule inhibitors. The protocol provides details on the experimental design, Plasmodium falciparum culture, sample preparation, analytical approaches, and data analysis used in either targeted (pathway focused) or untargeted (all detected metabolites) analysis of drug-induced metabolic perturbations.

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Acknowledgments

This work was supported by grants from the Australian National Health and Medical Research Council (NHMRC). M.J.M. is a NHMRC Principal Research Fellow, and S.A.C. is a University of Melbourne Early Career Research Fellow.

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Authors and Affiliations

  1. Department of Biochemistry and Molecular Biology, Bio21 Institute of Molecular Science and Biotechnology, The University of Melbourne, Parkville, 3010, Victoria, Australia

    Simon A. Cobbold & Malcolm J. McConville

Authors
  1. Simon A. Cobbold
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  2. Malcolm J. McConville
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Corresponding author

Correspondence to Malcolm J. McConville .

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Editors and Affiliations

  1. Joint BioEnergy Institute, Lawrence Berkeley National Laboratory, Emeryville, CA, USA

    Edward E.K. Baidoo

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Cobbold, S.A., McConville, M.J. (2019). Determining the Mode of Action of Antimalarial Drugs Using Time-Resolved LC-MS-Based Metabolite Profiling. In: Baidoo, E. (eds) Microbial Metabolomics. Methods in Molecular Biology, vol 1859. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8757-3_12

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  • DOI: https://doi.org/10.1007/978-1-4939-8757-3_12

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-8756-6

  • Online ISBN: 978-1-4939-8757-3

  • eBook Packages: Springer Protocols

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