Application of GC Orbitrap mass spectrometry for untargeted metabolomics of pathogenic microorganisms

Significance of the Topic

Infections by pathogenic microorganisms such as Staphylococcus aureus and Candida albicans pose severe clinical challenges. Understanding the small molecule interactions within mono and polymicrobial biofilms is critical for developing new diagnostic and therapeutic approaches.

Study Objectives and Overview

This work aimed to apply untargeted gas chromatography high resolution mass spectrometry to profile metabolic changes in individual and co cultured C albicans and S aureus cells and their growth media. The goal was to identify key metabolites underlying the synergistic virulence observed in mixed biofilms.

Methodology and Used Instrumentation

Sample preparation involved chemical derivatization by methoximation and silylation of intracellular and extracellular extracts. Internal standards included 13C6 glucose, D27 myristic acid and scyllo inositol. The analytical platform consisted of:

• Thermo Fisher Scientific Q Exactive GC Orbitrap mass spectrometer
• Thermo Fisher Scientific TRACE 1310 gas chromatograph
• Thermo Fisher Scientific TraceGOLD TG 5Sil MS column
• Thermo Fisher Scientific TriPlus RSH robotic sample introduction

Main Results and Discussion

Data processing utilized Compound Discoverer for alignment, peak detection and multivariate statistics and TraceFinder for deconvolution and putative compound annotation. Key findings included:

• Distinct PCA clustering separating media only, individual cultures and co culture samples
• Altered amino acid and sugar profiles in spent media, with glycine depletion in S aureus media
• Elevated sedoheptulose 7 phosphate levels in co culture indicating potential antifungal control metabolites
• Confirmation of metabolite identities via library matching and comparison with pure standards

Benefits and Practical Applications

This workflow offers rapid untargeted metabolite discovery with sub ppm mass accuracy and robust dynamic range. It supports high confidence compound identification and quantitation for microbial interaction studies and biomarker discovery.

Future Trends and Potential Applications

High resolution GC MS is poised to advance integrative metabolomics and multi omics studies in microbiology. Future developments may include automated library expansion, retention time prediction algorithms and real time metabolic monitoring in clinical diagnostics and drug development.

Conclusion

The Q Exactive GC Orbitrap system provides a powerful platform for untargeted metabolomics of pathogenic microorganisms. Its full scan sensitivity, high mass resolution and seamless data processing enable detailed metabolic insights into microbial interactions.

Reference

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