Measurement of intracellular metabolites of primary metabolism and adenine nucleotides in chemostat cultivated Penicillium chrysogenum

Biotechnology and Bioengineering
Uly NasutionJ J Heijnen

Abstract

An experimental platform has been developed for rapid sampling and quenching of chemostat cultivated Penicillium chrysogenum broth for metabolome analysis in highly dynamic experiments, aimed at the elucidation of the in vivo kinetic properties of metabolism. The sampling and quenching protocol available from Saccharomyces cerevisiae had to be modified for Penicillium chrysogenum mainly because of its filamentous character. Intracellular metabolites of glycolysis, TCA cycle, and adenine nucleotides were measured with isotope dilution mass spectrometry (IDMS) using a U-(13)C-labeled metabolite mix produced from yeast cells as internal standard. By addition of the U-(13)C internal standard mix prior to the metabolite extraction procedure, partial degradation of metabolites as well as non-linearity and drift of the LC-MS/MS could be successfully compensated for. It was found that there is a serious matrix effect on metabolite extraction between different organisms, which is however completely corrected for by the IDMS approach. Intracellular metabolites could be analyzed with standard deviations of around 5%. A comparison of the metabolite levels between Saccharomyces cerevisiae and Penicillium chrysogenum showed both significant ...Continue Reading

References

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Citations

Aug 1, 2007·Applied Microbiology and Biotechnology·Marco OldigesChristiane Wiendahl
Jun 22, 2011·Applied Microbiology and Biotechnology·Friederike SchädelEzequiel Franco-Lara
Jul 8, 2008·Biotechnology Letters·Christoph J Bolten, Christoph Wittmann
Sep 8, 2012·Applied Biochemistry and Biotechnology·Ming-Zhu DingYing-Jin Yuan
Oct 27, 2012·Microbial Cell Factories·Zheng ZhaoS Aljoscha Wahl
Nov 9, 2006·Biotechnology Letters·Mlawule R MashegoJoseph J Heijnen
Sep 15, 2015·Metabolomics : Official Journal of the Metabolomic Society·Francisca LameirasWalter M van Gulik
Jul 14, 2011·Biotechnology Journal·Lodewijk P de JongeWalter M van Gulik
Oct 23, 2012·Journal of Separation Science·Stefan NeubauerGunda Koellensperger
Jul 27, 2012·Metabolomics : Official Journal of the Metabolomic Society·Lodewijk P de JongeWalter M van Gulik
Dec 10, 2019·Biotechnology and Bioengineering·Guan WangSiliang Zhang
Jun 19, 2013·Biotechnology and Bioengineering·Katja SchmitzStephan Noack
Mar 6, 2021·Biotechnology and Bioengineering·Peng LiuHenk Noorman
Apr 7, 2007·Analytical Chemistry·Christoph J BoltenChristoph Wittmann

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