Follow-up studies on knock-out mutants confirm this complex picture [8,9]. As this filamentous bacterium grows in heterogeneous pellets, it has been difficult to perform reproducible cultivations in shake flasks. Recently, we introduced a dedicated optimized batch fermentation strategy as part of a technical platform for generating reproducible expression data of Streptomyces coelicolor . The central processes inside the cell during transition phase are turning off biosynthesis of cell building blocks while preserving synthesis of antibiotic precursor
metabolites and energy production for maintenance purposes. The comprehensive Inhibitors,research,lifescience,medical analysis of intra- and extracellular metabolite pools is called metabolomics, and mass spectrometry (MS) and nuclear magnetic Inhibitors,research,lifescience,medical resonance (NMR) have become the two most popular technologies in this field . The advantages of MS are the high sensitivity and excellent resolution when combined with gas and/or liquid chromatographic (GC/LC) separation of complex biological extracts. GC/LC-MS analysis is either Inhibitors,research,lifescience,medical performed with a non-targeted or a targeted strategy. The former approach aims at identifying
metabolites (e.g., biomarkers) of the biological system under study, while the latter aims at quantifying known compounds. Targeted analysis is of particular interest in systems biology since quantified metabolite pool data can be used directly in the integrated data analysis . As the metabolome comprises compounds with a wide diversity in physico-chemical properties, ranging from highly charged to selleck chem hydrophobic metabolite species, it is necessary to use several GC-MS and LC-MS methods for a comprehensive coverage of the most abundant metabolites [13,14]. In particular, Inhibitors,research,lifescience,medical LC-MS has become an invaluable
tool in profiling families of secondary metabolites [15,16,17], and an LC-MS based method has only recently been applied in a metabolite analysis study of a synthetic metabolic Inhibitors,research,lifescience,medical switch in S. coelicolor . In the present study, we have used both GC-MS and LC-MS/MS to monitor changes in the intracellular pools of important metabolite groups like amino and non-amino organic acids, phosphometabolites and nucleotides during the entire lifetime of S. coelicolor batch fermentations including in particular the period of metabolic AV-951 selleck products switching to antibiotic production phase. The term ‘metabolic switching’ is nowadays often used in the sense of describing the molecular events during the transition from primary to secondary metabolism, and in the present study, we have chosen to use the term in this broader molecular sense. To our knowledge, this is the first report of a high resolution time-course metabolic profiling study in Streptomyces spp. to understand how the primary intracellular metabolite pools change in response to metabolic switching from growth to secondary metabolite production phase. 2. Results and Discussion 2.1.