The optimisation and application of a metabolite profiling procedure for the metabolic phenotyping of Bacillus species

The rapid advances in sequencing technologies over the last decade have enabled routine sequencing of microbial genomes. Despite notable achievements, metabolomics/metabolite profiling has not progressed with the same rapidity, which in part is due to the intrinsic complex chemical nature of the metabolome. However, well characterised metabolomes are essential if a comprehensive understanding of biological function and biotechnological applications are to be revealed and implemented. In the present study a hyphenated MS metabolite profiling procedure has been developed, predominantly for Bacillus species. The approach has been systematic in its development, delivering optimised procedures for the quenching of bacterial metabolism, extraction of metabolites, the separation and detection of components as well as data analysis, integration and visualisation workflows. Collectively, the procedure has enabled the detection of 27 % of the predicted Bacillus subtilis metabolome in the industrial HU36 strain. The analytical platform developed has been used to assess the chemotype of commercially used probiotic Bacillus strains, including a novel pigmented Bacillus strain HU36 that has potential either as a probiotic or source of antioxidants. The results are discussed in a biochemical context, revealing: (i), specific metabolic networks associated with pigment biosynthesis in HU36 and (ii), biotechnological applications through the demonstration of substantial equivalence.