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Combined infections from  Candida albicans  and  Staphylococcus aureus  are a leading cause of death in the developed world. Evidence suggests that  Candida  enhances the virulence of  Staphylococcus —hyphae penetrate through tissue barriers, while  S. aureus  tightly associates with the hyphae to obtain entry to the host organism. Indeed, in a biofilm state,  C. albicans  enhances the antimicrobial resistance characteristics of  S. aureus . The association of these microorganisms is also associated with significantly increased morbidity and mortality. Due to this tight association we hypothesised that metabolic effects were also in evidence.    Objectives  To explore the interaction, we used a novel GC-Orbitrap-based mass spectrometer, the Q Exactive GC, which combines the high peak capacity and chromatographic resolution of gas chromatography with the sub-ppm mass accuracy of an Orbitrap system. This allows the capability to leverage the widely available electron ionisation libraries for untargeted applications, along with expanding accurate mass libraries and targeted matches based around authentic standards.    Methods  Optimised  C. albicans  and  S. aureus  mono- and co-cultured biofilms were analysed using the new instrument in addition to the fresh and spent bacterial growth media.    Results  The targeted analysis experiment was based around 36 sugars and sugar phosphates, 22 amino acids and five organic acids. Untargeted analysis resulted in the detection of 465 features from fresh and spent medium and 405 from biofilm samples. Three significantly changing compounds that matched to high scoring library fragment patterns were chosen for validation.    Conclusion  Evaluation of the results demonstrates that the Q Exactive GC is suitable for metabolomics analysis using a targeted/untargeted methodology. Many of the results were as expected: e.g. rapid consumption of glucose and fructose from the medium regardless of the cell type. Modulation of sugar-phosphate levels also suggest that the pentose phosphate pathway could be enhanced in the cells from co-cultured biofilms. Untargeted metabolomics results suggested significant production of cell-wall biosynthesis components and the consumption of non-proteinaceous amino-acids.    Electronic supplementary material  The online version of this article (doi:10.1007/s11306-016-1134-2) contains supplementary material, which is available to authorized users.

A novel targeted/untargeted GC-Orbitrap metabolomics methodology applied to Candida albicans and Staphylococcus aureus biofilms