Overcoming limitations in exhaled breath aerosol analysis - Untargeted breathomics for the identification of endogenous breath biomarkers using LC-HRMS/MS.

BACKGROUND: Human exhaled breath (EB) aerosol is a suitable matrix for the non-invasive bioanalysis of both volatile but also non-volatile compounds. One of the key challenges in drug monitoring studies is the lack of standardization in EB sample collection. The aim of the present study was to use untargeted breathomics for identification of potential endogenous biomarkers in EB that could address these challenges.

RESULTS: The Breath Explor impaction device was used to collect microparticles in EB aerosol from six healthy volunteers and six different sampling time points over a 1-h period (total number of samples = 36). Sample preparation consisted of an extraction with methanol and analysis was performed using reversed-phase and hydrophilic interaction-phase liquid chromatography coupled to high-resolution mass spectrometry. Results revealed the presence of a feature that could be reliably detected across all samples, thus representing a potential biomarker for standardizing EB aerosol sample collection. Although an annotation of this feature was not possible, it showed robustness and reproducibility in EB samples from healthy volunteers and could also be successfully detected in patient samples.

SIGNIFICANCE AND NOVELTY: This study represents a step forward in overcoming the current limitation of EB analysis, the lack of standardization in sampling procedure. Nevertheless, further investigation including correlation studies and more diverse populations will be essential for the confirmation of the clinical relevance and application of this biomarker.