Metabolome analysis of gram-positive bacteria such as Staphylococcus aureus by GC-MS and LC-MS.

The field of metabolomics has become increasingly important in the context of functional genomics. Together with other "omics" data, the investigation of the metabolome is an essential part of systems biology. Beside the analysis of human and animal biofluids, the investigation of the microbial physiology by methods of metabolomics has gained increased attention. For example, the analysis of metabolic processes during growth or virulence factor expression is crucially important to understand pathogenesis of bacteria. Common bioanalytical techniques for metabolome analysis include liquid and gas chromatographic methods coupled to mass spectrometry (LC-MS and GC-MS) and spectroscopic approaches such as NMR. In order to achieve metabolome data representing the physiological status of a microorganism, well-verified protocols for sampling and analysis are necessary. This chapter presents a detailed protocol for metabolome analysis of the Gram-positive bacterium Staphylococcus aureus. A detailed manual for cell sampling and metabolite extraction is given, followed by the description of the analytical procedures GC-MS and LC-MS. The advantages and limitations of each experimental setup are discussed. Here, a guideline specified for S. aureus metabolomics and information for important protocol steps are presented, to avoid common pitfalls in microbial metabolome analysis.

Synthesis and characterization of authentic standards for the analysis of ribofuranose-containing carbohydrates by the reductive-cleavage method.

Described herein is the synthesis of all positional isomers of partially methylated and acetylated or benzoylated 1,4-anhydro-D-ribitol. The benzoates are generated simultaneously from 1,4-anhydro-D-ribitol by sequential partial methylation and benzoylation or sequential partial benzoylation and methylation. The individual isomers are obtained in pure form by high-performance liquid chromatography. Debenzoylation and acetylation provided the corresponding acetates. Reported herein are the 1H NMR spectra of the benzoates and the electron-ionization mass spectra of the acetates and the tri-O-methyl derivative and also for the acetates and the tri-O-methyl derivative, their linear temperature programmed gas-liquid chromatography retention indices on three different capillary columns.