AIM--To determine whether pancreatic enzymes are present in hepatic bile and in intrahepatic bile duct epithelium. METHODS--The activity and proteins of pancreatic enzymes (pancreatic alpha-amylase, lipase, trypsin/trypsinogen) in hepatic bile were investigated using biochemical and western blot analyses in 25 patients with extrahepatic bile duct obstruction. Immunolocalization of enzyme proteins was evaluated by immunohistochemistry in 20 necropsy livers with extrahepatic bile duct obstruction. RESULTS--Western blot analysis showed proteins of pancreatic alpha-amylase, lipase, and trypsin in 19 of 25 (76%), 10 of 25 (40%), and 14 of 25 (56%) patients, respectively. Pancreatic alpha-amylase and lipase activities was present in every bile specimen. Radioimmunoassay showed that trypsin was present in every bile sample. Immunohistochemically, the immunoreactivity of the three enzymes was present in epithelia and in the lumina of intrahepatic large bile ducts, septal bile ducts, and peribiliary glands in all cases. CONCLUSIONS--These results strongly suggest that biliary epithelia of larger intrahepatic ducts produce pancreatic alpha-amylase, lipase, and trypsin, and that these enzymes are secreted into the lumina of intrahepatic bile ducts.