Validation of volatile metabolites of pulmonary oxidative injury: a bench to bedside study

Changes in exhaled volatile organic compounds (VOCs) can be used to discriminate between respiratory diseases, and increased concentrations of hydrocarbons are commonly linked to oxidative stress. However, the VOCs identified are inconsistent between studies, and translational studies are lacking.In this bench-to-bedside study, we captured VOCs in the headspace of A549 epithelial cells after exposure to hydrogen peroxide (H2O2), to induce oxidative stress, using high-capacity polydimethylsiloxane sorbent fibres. Exposed and unexposed cells were compared using targeted and untargeted analysis. Breath samples of invasively ventilated ICU patients (n=489) were collected on sorbent tubes and associated with the inspired oxygen fraction (FiO2) to reflect pulmonary oxidative stress. Headspace samples and breath samples were analysed using gas-chromatography and mass-spectrometry.In the cell, headspace octane concentration was decreased after oxidative stress (p=0.0013), while the other VOCs were not affected. 2-ethyl-1-hexanol, showed an increased concentration in the headspace of cells undergoing oxidative stress in untargeted analysis (p=0.00014). None of the VOCs that were linked to oxidative stress showed a significant correlation with FiO2 (Rsrange: −0.015 to −0.065) or discriminated between patients with FiO2≥0.6 or below (AUC range: 0.48 to 0.55).Despite a comprehensive translational approach, validation of known and novel volatile biomarkers of oxidative stress was not possible in patients at risk of pulmonary oxidative injury. The inconsistencies observed highlight the difficulties faced in VOC biomarker validation and that caution is warranted in the interpretation of the pathophysiological origin of discovered exhaled breath biomarkers.