Background Previous studies indicated that the prevalence of constipation in enuretic patients is higher than that in the general population. Several studies have revealed that successful treatment of constipation may be helpful in resolving enuresis. However, constipation affecting the efficacy of desmopressin in treating enuresis remains to be clarified. Objective This study aimed to determine whether the presence of constipation is associated with the efficacy of desmopressin in treating enuresis. Study design Patients diagnosed with nocturnal enuresis (NE) were studied prospectively. Treatment responses in different stratified groups of patients with NE were compared by Chi-squared tests or Wilcoxon rank sum test. A logistic regression model was performed to investigate the relationship between the possible factors and the effectiveness of desmopressin. Results In children with severe enuresis, patients with constipation had significantly lower complete response rate compared with patients without constipation. The presence of constipation was always related to the effectiveness of desmopressin whether in monosymptomatic NE or non-monosymptomatic patients with NE. With stratification for dose of desmopressin, non-constipated patients who received 0.2 mg of desmopressin had significantly higher complete response rate than patients with constipation. However, in subgroups of mild to moderate NE and 0.4 mg desmopressin, constipation was not associated with treatment response of enuresis. Logistic regression analysis revealed that constipation was significantly related to the effectiveness of desmopressin. Discussion This study confirmed the negative effects of constipation in response to desmopressin in patients with NE. To the best of authors knowledge, this work is the first study to evaluate the relationship of constipation in enuretic patients and the efficacy of desmopressin. Conclusions The presence of constipation negatively affects the response to desmopressin in patients with NE, especially in patients with severe enuresis and in patients prescribed with low dose of desmopressin.