Natriuretic peptides are associated with malnutrition and volume overload. Over-hydration cannot simply be explained by excess extracellular water in patients undergoing hemodialysis. We assessed the relationship between the extracellular and intracellular water (ECW/ICW) ratio, N-terminal pro-B-type natriuretic peptide (NT-proBNP), human atrial natriuretic peptide (hANP), and echocardiographic findings. Body composition was examined by segmental multi-frequency bioelectrical impedance analysis in 368 patients undergoing maintenance dialysis (261 men and 107 women; mean age, 65 ± 12 years). Patients with higher ECW/ICW ratio quartiles tended to be older, were on dialysis longer, and had higher post-dialysis blood pressure and lower body mass index, ultrafiltration volume, serum albumin, blood urea nitrogen, and creatinine levels (p < 0.05). The ECW/ICW ratio significantly increased with decreasing ICW, but not with ECW. Patients with a higher ECW/ICW ratio and lower percent fat had significantly higher natriuretic peptide levels. After adjusting for covariates, the ECW/ICW ratio remained an independent associated factor for natriuretic peptides (β = 0.34, p < 0.001 for NT-proBNP and β = 0.40, p < 0.001 for hANP) and the left ventricular mass index (β = 0.20, p = 0.002). The ICW-ECW volume imbalance regulated by decreased cell mass may explain the reserve capacity for fluid accumulation in patients undergoing hemodialysis.