Serum C-peptide exhibits various biological activities. The relationship between C-peptide and atrial cardiomyopathy remains unknown. We aimed to investigate the association between C-peptide level and atrial cardiomyopathy in nondiabetic adults. Our study enrolled 4578 participants without diagnosed diabetes from the Third National Health and Nutrition Examination Survey (NHANES III). Atrial cardiomyopathy was defined as a deep terminal negative P wave in V1 below − 100 µV (more negative), according to the electrocardiogram. The participants were categorized into low C-peptide (≤ 1.46 nmol/L) and high C-peptide (> 1.46 nmol/L) groups, according to the receiver operating characteristic analysis. Odds ratio (OR) and 95% confidence interval (CI) for the association between C-peptide level and atrial cardiomyopathy were generated using multivariate logistic regression analysis. The prevalence of atrial cardiomyopathy was higher in the high C-peptide group than in the low C-peptide group (5.62% vs. 2.31%, P < 0.001, respectively). Multivariate logistic regression analysis showed that participants in the high C-peptide group had a 3.60-fold (95% CI 1.81–6.99) higher risk of atrial cardiomyopathy than those in the low C-peptide group. Per standard deviation increase in C-peptide was linked to a 1.20-fold (95% CI 1.00–1.41) higher risk in atrial cardiomyopathy. High C-peptide level might be an independent risk factor for atrial cardiomyopathy in nondiabetic adults.