Discovery and validation of food intake biomarkers using untargeted metabolomics in human intervention and cross-sectional studies

Background: Banana is one of the most widely consumed fruits in the world. However, scarce information is available regarding its health effects. Biomarkers of banana intake would allow a more accurate assessment of its consumption in nutrition studies.Objective: Using an untargeted metabolomics approach, we aimed to identify the bananaderived metabolites present in urine after consumption, and among them new candidate biomarkers of banana intake.Methods: A randomized controlled study with a crossover design was performed on 12 healthy subjects, with two dietary interventions: 1) 250ml control drink (Fresubin) and 2) 240g banana+150ml control drink. Urine samplescollected over 24h were analyzed with UPLC-QTOFMS and GC×GC-MS. The discovered biomarkers were confirmedin a cross-sectional study (KarMeN) where 72 subjects were selected reflecting high, low, and non-consumers ofbanana. The confirmed biomarkers were examined singly or in combinations, for established criteria of validationfor biomarkers of food intake.Results: We identified 33 potentially bioactive banana metabolites, some of them being newly described. A panelof five metabolites, methoxyeugenol-glucuronide, dopamine-sulfate, salsolinolsulfate, xanthurenic-acid and 6-hydroxy-1-methyl-1,2,3,4-tetrahydro-b-carboline-sulfate, were confirmed as candidate intake biomarkers. We demonstrated that the combination of methoxyeugenolglucuronide and dopamine-sulfate performs best in predicting banana intake in high (AUCtest=0.92) and low (AUCtest=0.87) consumers. The new biomarkers met key criteria establishing their current applicability in nutrition and health research for assessing occurrence of banana intake.Conclusions: Our metabolomics study revealed new putative bioactive metabolites of banana and a combined biomarker of intake. Those findings will help to better decipher the health effects of banana in future focused studies