Objective To investigate the cause of elevation of bile amylase level in patients with normal pancreaticobiliary junction and the effect of endoscopic treatment. Methods The clinical data of 96 patients with normal pancreaticobi-liary junction who had laparoscopic cholecystectomy from January 2016 to December 2019 were retrospectively analyzed. Gallbladder bile was gotten before gallbladder resection and the amylase in bile was measured. Duodenoscopy, endoscopic retrograde cholangiopancreatography (ERCP), endoscopic pancreaticobiliary separation (EPBS) and bile duct amylase (BDA) was done in some cases. Results Gallbladder bile amylase (GBA) in all 96 cases was (484.2±200.0) U/L. Sixty-two cases had duodenoscopy. Diagnoses was as follows: long nipple: 7 cases; papillary atrophy: 5 cases; parapapillary diverticulum: 4 cases; papillary tumor: 3 cases; papillitis: 2 cases; parapapillary diverticulum and papillitis: 26 cases; parapapillary diverticulum and long nipple: 8 cases; long nipple and papillitis: 4 cases; parapapillary diverticulum and papillary atrophy: 3 cases. Fifty-nine cases had ERCP among whom 45 cases had elevated level of BDA[(386.9±137.3) U/L] higher than normal (110 U/L), but lower than GBA [(466.3±189.4) U/L] with significant difference (t=2.276, P=0.025). EPBS was performed in all 45 cases and ERCP was performed again one month later. BDA returned to normal [(65.9±13.6) U/L] with significant difference before EPBS and after (P<0.001). Conclusions Pancreaticobiliary junction disease might be an important cause of elevated bile amylase in patients with normal pancreaticobiliary junction. EPBS is an effective treatment.