Endoscopic retrograde cholangiopancreatography (ERCP) is associated with risk of complications. Methods to facilitate ductal cannulation may reduce complications. Intraduodenal acid infusion is a physiological approach to stimulate secretin release in human body and may facilitate cannulation. The objective of this study was to investigate the effect of intraduodenal acid infusion on pancreatic duct cannulation during ERCP. It was a single center, double-blind, randomized controlled trial. Consecutive patients undergoing first ERCP for pancreatic diseases were randomized toreceive 50 mL acetic acid intraduodenal infusion at ERCP (acetic acid group) or 50 mL saline (control group). The primary outcome, difficult cannulation rate (cannulation time >5 min), in the two groups was compared. Two hundred ten patients were included in the final analysis (105 in each group). The difficult cannulation rate were 39.1% in the control group and 20.9% in the acetic acid group, and the difference was statistically significant (P = 0.004). The overall successful deep cannulation rate was 89.5% and 85.7%, respectively (P = 0.402). The cannulation time was remarkably shortened (182 vs 286 s, P = 0.018), and the cannulation attempts were significantly decreased (3.4 vs 4.4, P = 0.008). The fluoroscopy time was also significantly reduced (60 vs 86 s, P = 0.028). The incidence of post-ERCP pancreatitis and hyperamylasemia was comparable (7.6% vs 10.5% and 10.5% vs 19.1%, P > 0.05). Intraduodenal acetic acid infusion can significantly decrease difficult pancreatic cannulation rate, facilitate pancreatic duct cannulation, and reduce radiation exposure (ClinicalTrials.gov number, NCT02800772). © 2018 Journal of Gastroenterology and Hepatology Foundation and John Wiley & Sons Australia, Ltd.