Background Atrial fibrillation (AF) is the most common cardiac arrhythmia, but the pathogenesis is not completely understood. The application of metabolomics could help in discovering new metabolic pathways involved in the development of the disease. Methods and Results We measured 112 baseline fasting metabolites of 3770 participants in the Malmö Diet and Cancer Study; these participants were free of prevalent AF. Incident cases of AF were ascertained through previously validated registers. The associations between baseline levels of metabolites and incident AF were investigated using Cox proportional hazard models. During 23.1 years of follow-up, 650 cases of AF were identified (incidence rate: 8.6 per 1000 person-years). In Cox regression models adjusted for AF risk factors, 7 medium- and long-chain acylcarnitines were associated with higher risk of incident AF (hazard ratio [HR] ranging from 1.09; 95% CI, 1.00-1.18 to 1.14, 95% CI, 1.05-1.24 per 1 SD increment of acylcarnitines). Furthermore, caffeine and acisoga were also associated with an increased risk (HR, 1.17; 95% CI, 1.06-1.28 and 1.08; 95% CI, 1.00-1.18, respectively), while beta carotene was associated with a lower risk (HR, 0.90; 95% CI, 0.82-0.99). Conclusions For the first time, we show associations between altered acylcarnitine metabolism and incident AF independent of traditional AF risk factors in a general population. These findings highlight metabolic alterations that precede AF diagnosis by many years and could provide insight into the pathogenesis of AF. Future studies are needed to replicate our finding in an external cohort as well as to test whether the relationship between acylcarnitines and AF is causal.