Review of sample preparation strategies for MS-based metabolomic studies in industrial biotechnology.
Fermentation and cell culture biotechnology in the form of so-called "cell factories" now play an increasingly significant role in production of both large (e.g. proteins, biopharmaceuticals) and small organic molecules for a wide variety of applications. However, associated metabolic engineering optimisation processes relying on genetic modification of organisms used in cell factories, or alteration of production conditions remain a challenging undertaking for improving the final yield and quality of cell factory products. In addition to genomic, transcriptomic and proteomic workflows, analytical metabolomics continues to play a critical role in studying detailed aspects of critical pathways (e.g. via targeted quantification of metabolites), identification of biosynthetic intermediates, and also for phenotype differentiation and the elucidation of previously unknown pathways (e.g. via non-targeted strategies). However, the diversity of primary and secondary metabolites and the broad concentration ranges encompassed during typical biotechnological processes means that simultaneous extraction and robust analytical determination of all parts of interest of the metabolome is effectively impossible. As the integration of metabolome data with transcriptome and proteome data is an essential goal of both targeted and non-targeted methods addressing production optimisation goals, additional sample preparation steps beyond necessary sampling, quenching and extraction protocols including clean-up, analyte enrichment, and derivatisation are important considerations for some classes of metabolites, especially those present in low concentrations or exhibiting poor stability. This contribution critically assesses the potential of current sample preparation strategies applied in metabolomic studies of industrially-relevant cell factory organisms using mass spectrometry-based platforms primarily coupled to liquid-phase sample introduction (i.e. flow injection, liquid chromatography, or capillary electrophoresis). Particular focus is placed on the selectivity and degree of enrichment attainable, as well as demands of speed, absolute quantification, robustness and, ultimately, consideration of fully-integrated bioanalytical solutions to optimise sample handling and throughput.
Division of Analytical Chemistry, Department of Chemistry, University of Natural Resources and Life Sciences (BOKU Vienna), Muthgasse 18, Vienna, 1190, Austria; Austrian Centre of Industrial Biotechnology (acib), Petersgasse 14, Graz, 8010, Austria. Electronic address: tim.causon@boku.ac.at.
Tim J
TJ
Causon
Division of Analytical Chemistry, Department of Chemistry, University of Natural Resources and Life Sciences (BOKU Vienna), Muthgasse 18, Vienna, 1190, Austria; Austrian Centre of Industrial Biotechnology (acib), Petersgasse 14, Graz, 8010, Austria.
Stephan
S
Hann
D001709:Biotechnology; D002851:Chromatography, High Pressure Liquid; D007221:Industry; D055432:Metabolomics; D013048:Specimen Handling; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*
Clean-up; Derivatisation; Enrichment; High-performance liquid chromatography; Industrial biotechnology; Metabolic engineering