RationaleThe biochemical mechanisms underlying lung function are incompletely understood.ObjectivesTo identify and validate the plasma metabolome of lung function using two independent adult cohorts: discovery—the European Prospective Investigation into Cancer–Norfolk (EPIC-Norfolk, n=10 460) and validation—the VA Normative Aging Study (NAS) metabolomic cohort (n=437).MethodsWe ran linear regression models for 693 metabolites to identify associations with forced expiratory volume in one second (FEV₁) and the ratio of FEV₁ to forced vital capacity (FEV₁/FVC), in EPIC-Norfolk then validated significant findings in NAS. Significance in EPIC-Norfolk was denoted using an effective number of tests threshold of 95%; a metabolite was considered validated in NAS if the direction of effect was consistent and p<0.05.Measurements and main resultsOf 156 metabolites that associated with FEV₁ in EPIC-Norfolk after adjustment for age, sex, body mass index, height, smoking and asthma status, 34 (21.8%) validated in NAS, including several metabolites involved in oxidative stress. When restricting the discovery sample to men only, a similar percentage, 18 of 79 significant metabolites (22.8%) were validated. A smaller number of metabolites were validated for FEV₁/FVC, 6 of 65 (9.2%) when including all EPIC-Norfolk as the discovery population, and 2 of 34 (5.9%) when restricting to men. These metabolites were characterised by involvement in respiratory track secretants. Interestingly, no metabolites were validated for both FEV₁ and FEV₁/FVC.ConclusionsThe validation of metabolites associated with respiratory function can help to better understand mechanisms of lung health and may assist the development of biomarkers.