Volatomic analysis identifies compounds that can stratify non-alcoholic fatty liver disease

Background: Analysis of volatile organic compounds (VOCs) in exhaled breath, ‘volatomics’, provides opportunities for non-invasive biomarker discovery and novel mechanistic insights into a variety of diseases. Aim: The purpose of this pilot study was to compare breath VOCs in an initial cohort of non-alcoholic fatty liver disease (NAFLD) patients and healthy controls. Methods: Breath samples were collected from 15 participants with Child-Pugh Class A NAFLD cirrhosis, 14 with non-cirrhotic NAFLD and 14 healthy volunteers. Exhaled breath samples were collected using an established methodology and VOC profiles were analysed by gas chromatography-mass spectrometry. The levels of 19 VOCs previously associated with cirrhosis were assessed. Peaks of the VOCs were confirmed and integrated using Xcalibur® software, normalized to an internal standard. Receiver Operating Characteristic (ROC) curves were used to determine the diagnostic accuracy of candidate VOCs. Results: Terpinene, dimethyl sulfide (DMS) and D-limonene provided the highest predictive accuracy to discriminate between study groups. Combining DMS with D-limonene led to even better discrimination of NAFLD cirrhosis from healthy volunteers (AUROC 0.98, 95% confidence interval (CI) 0.93 -1.00, p<0.001) and NAFLD cirrhosis from non-cirrhotic NAFLD (AUROC 0.91, 95% CI 0.82 – 1.00, p<0.001). Breath terpinene concentrations discriminated between non-cirrhotic NAFLD and healthy volunteers (AUROC 0.84, 95% CI 0.68 – 0.99, p=0.002). Conclusion: Breath terpinene, dimethyl sulfide and D-limonene are potentially useful volatomic markers for stratifying NAFLD; and a two-stage approach allows differentiation of non-cirrhotic and cirrhotic patients. These observations require validation in a larger NAFLD population. (ClinicalTrials.gov Identifier: NCT02950610).