Elsevier

Analytical Biochemistry

Volume 370, Issue 1, 1 November 2007, Pages 17-25
Analytical Biochemistry

Microbial metabolomics: Toward a platform with full metabolome coverage

https://doi.org/10.1016/j.ab.2007.07.022Get rights and content

Abstract

Achieving metabolome data with satisfactory coverage is a formidable challenge in metabolomics because metabolites are a chemically highly diverse group of compounds. Here we present a strategy for the development of an advanced analytical platform that allows the comprehensive analysis of microbial metabolomes. Our approach started with in silico metabolome information from three microorganisms—Escherichia coli, Bacillus subtilis, and Saccharomyces cerevisiae—and resulted in a list of 905 different metabolites. Subsequently, these metabolites were classified based on their physicochemical properties, followed by the development of complementary gas chromatography–mass spectrometry and liquid chromatography–mass spectrometry methods, each of which analyzes different metabolite classes. This metabolomics platform, consisting of six different analytical methods, was applied for the analysis of the metabolites for which commercial standards could be purchased (399 compounds). Of these 399 metabolites, 380 could be analyzed with the platform. To demonstrate the potential of this metabolomics platform, we report on its application to the analysis of the metabolome composition of mid-logarithmic E. coli cells grown on a mineral salts medium using glucose as the carbon source. Of the 431 peaks detected, 235 (=176 unique metabolites) could be identified. These include 61 metabolites that were not previously identified or annotated in existing E. coli databases.

Section snippets

In silico metabolomes

The complete lists of metabolites present in the in silico metabolomes of E. coli and Bacillus subtilis were deduced from the metabolite databases for these microorganisms as downloaded in April 2005 from http://www.ecocyc.org (version 9.0 [15]) and http://biocyc.org/BSUB/organism-summary?object=BSUB and the metabolite database for yeast as downloaded in August 2006 from http://systemsbiology.ucsd.edu/organisms/yeast.html[16]. These metabolite lists were curated manually; compounds with

Metabolite composition of microbial metabolomes

For the development of an analytical platform that allows the detection of all metabolites present in microbial metabolomes, it is essential to have access to information about the type and number of metabolites potentially present in microorganisms. To this end, we established the in silico metabolomes of three phylogenetically different and commonly applied microorganisms: E. coli, B. subtilis, and Saccharomyces cerevisiae. Metabolite lists were downloaded from the compounds sub-databases in

Discussion

We have presented a strategy for the development of a metabolomics platform toward the analysis of full microbial metabolomes. Notwithstanding the large chemical diversity of metabolites, using a combination of six different GC–MS and LC–MS methods only, of the 399 metabolites for which commercial standards could be purchased, 380 could be analyzed with this metabolomics platform. Notably, two of the methods of our metabolomics platform, the OS–GC–MS and IP–LC–MS methods, were very powerful and

Acknowledgments

The authors thank Nicole van Luijk and Anna Conesa for downloading the Bacillus subtilis metabolite database; Roelie Bijl for cultivating Escherichia coli and extracting the samples; Maud Koek, Leo van Stee, and Richard Bas for GC–MS and LC–MS analysis; Roche Vitamins (currently DSM DNP, Basel, Switzerland) for financial support; and Markus Wyss and Werner Bretzel (currently DSM DNP) for fruitful discussions.

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    Current address: Division of Analytical Biosciences, Leiden/Amsterdam Center for Drug Research, 2300 RA Leiden, The Netherlands.

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