A protocol for lab-scale production of 13C yeast extract as internal standard for metabolomics and quantification of intracellular metabolites

Cammaert, M.; Wouters, R. I.; van Ede, J. M.; de Hulster, E. A. F.; Mooiman, C. M.; van Dam, P. T. N.; Pabst, M.; van Gulik, W. M.; Daran-Lapujade, P.

doi:10.64898/2026.06.12.731807

bioRxiv · 10.64898/2026.06.12.731807

A protocol for lab-scale production of 13C yeast extract as internal standard for metabolomics and quantification of intracellular metabolites

Cammaert, M.·Wouters, R. I.·van Ede, J. M.·de Hulster, E. A. F.·Mooiman, C. M.·van Dam, P. T. N.·Pabst, M.·van Gulik, W. M.·Daran-Lapujade, P.

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Abstract

Metabolomics enables the profiling of small-molecule metabolites and thereby captures the biochemical state of a living organism at a given moment and enables to monitor its cellular responses to stimuli. This technique has become a powerful tool in pharmaceutical research, the food industry, and microbial research. Metabolomics aims to obtain an unbiased metabolic profile; however, this is complicated by compound instability, complex and often extensive sample processing, and nonlinear responses in mass spectrometry. Therefore, correcting for metabolite loss and mass spectrometry-related artifacts is essential, typically achieved through relative quantification against an isotopically labelled internal standard for each metabolite of interest. This article describes how to produce 13C-labelled yeast extract and its use as internal standard for metabolomics. More specifically, it provides step-by-step protocols for the fed-batch fermentation, quenching, metabolite extraction, and LC-MS and GC-MS characterization of the internal standard. It also includes a protocol explaining how to use the internal standard for the quantification of metabolites in yeast samples.

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Cammaert, M., Wouters, R. I., van Ede, J. M., de Hulster, E. A. F., Mooiman, C. M., van Dam, P. T. N., Pabst, M., van Gulik, W. M., Daran-Lapujade, P.. 2026-06-16. A protocol for lab-scale production of 13C yeast extract as internal standard for metabolomics and quantification of intracellular metabolites. https://doi.org/10.64898/2026.06.12.731807

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