The mutagenicity of human bile was examined in the Ames Salmonella/microsome assay. Bile samples were obtained from the gallbladders resected from patients with cholelithiasis, choledocholithiasis, gallbladder cancer, extrahepatic bile duct cancer and other diseases. For extraction of mutagenic components, the bile samples were treated with blue rayon and the adsorbed materials were assayed with Salmonella typhimurium TA98 in the presence of S9 mix. Twenty-four bile samples were tested and positive mutagenic activity was found in 14 samples. A 200-microliter bile equivalent material gave 6.3 times as many revertant colonies as the solvent control. With several samples that had undergone two cycles of blue rayon extraction, clear dose-response relationships in mutagenicity were demonstrated.