Some biomarkers play important roles in the endothelial dysfunction of patients with pulmonary arterial hypertension (PAH), including nitric oxide (NO), endothelin‐1 (ET‐1), asymmetric dimethylarginine (ADMA), galectin‐3 (Gal‐3), B‐type natriuretic peptide (BNP), and uric acid (UA). However, studies on these biomarkers in pulmonary artery blood in congenital heart disease–PAH (CHD–PAH) and the effect of iloprost on the regulation of biomarkers are lacking. This study investigated potential CHD–PAH biomarkers and their association with the severity of disease. The effect of iloprost on the regulation of these biomarkers was also studied. A total of 31 patients with CHD–PAH were enrolled. 7 with positive effects of iloprost (the average reduction in mPAP 11.13 ± 1.73 mm Hg) and 19 with negative effects of iloprost [the average reduction in mPAP 4.21 ± 4.87 mm Hg, iloprost positive group (IPG) vs iloprost negative group (ING), P < 0.01]—and 5 age‐matched controls were studied. The pulmonary artery blood sample was collected before and after inhaling iloprost, and the plasma concentrations of Gal‐3, ADMA, ET‐1, and NO were measured. A significant positive linear relationship was observed between mPAP and plasma ET‐1, BNP, ADMA, and UA levels in all patients with CHD–PAH. ET‐1, ADMA, BNP, and UA levels had a significant linear relationship with mean pulmonary arterial pressure, which could be used to predict the severity of CHD–PAH. ET‐1 might be a potential biomarker to pre‐evaluate the effect of iloprost on CHD–PAH. Iloprost could affect the expression of Gal‐3 and, therefore, the process of fibrosis could be influenced by iloprost. This article is protected by copyright. All rights reserved.