OBJECTIVE: To investigate the relationship between the intestinal barrier function and pigment gallstone formation. METHODS: Ninety Guinea pigs were divided randomly into 3 groups: normal control (CON) group receiving normal forage, pigment gallstone (PS) group receiving pigment gallstone-forming forage, and intestinal mucosa protection group receiving pigment gallstone-forming forage with supplemental glutamine intestinal (GLN), a mucosa protector. The guinea pigs were observed for 8 weeks, the gallstone-forming rate, plasma diamine oxidase ( DAO), serum endotoxin, proportionality of urine lactulose/mannitol, and biliary beta-glucuronidase were detected. PCR was used to detect the bacteria in abdominal lymph node taking 16SrRNA as the target gene common in most bacteria. 32 gallstone patients, 16 with cholesterol gallstone and 16 with pigmental gallstone, and 27 patients with non-gastroenterological diseases, as controls, underwent detection of the plasma DAO and serum endotoxin. Another 109 gallstone patients, 31 with cholesterol gallstone and 78 with pigmental gallstone, and 21 patients with nongastroenterological diseases, as controls, underwent detection of urine technetium-labeled diethylenetriamine-pentaacetate (99mTc-DTPA). RESULTS: The gallstone-forming rate of the guinea pigs of the GLN group was 44.4% was, significantly lower than that of the PS group (73.9%, P < 0.05). The plasma DAO, serum endotoxin levels, proportionality of urine lactulose/mannitol, and activity of biliary beta-glucuronidase of the PS group were all significantly higher than those of the CON group (P < 0.05 or P < 0.01). The plasma endotoxin level of the pigmental GLN group was significantly lower than that of the PS group (P < 0.01). The positive rate of bacteria in abdominal lymph node of the PS group was 80%, significantly higher than those of the CON and GLN groups (30% and 45% respectively, P < 0.01 and P < 0.05). The level of plasma DAO and endotoxin of the pigmental gallstone patients were significantly higher than those of the controls (P < 0.05 and P < 0.01). The urine 99mTc-DTPA excretion rate of gallstone patients was 11.4%, significantly higher than that of the controls (4.7%, P < 0.01). CONCLUSION: Intestinal barrier function is correlated with pigment gallstone forming. Intestinal barrier function disorder may promote pigment gallstone formation through bacteria translocation, endotoxemia, and increase of biliary beta-glucuronidase.