Long-term total parenteral nutrition (TPN) is known to be associated with pigment gallstones. The objectives of this study were to identify the lithogenic effects of TPN and to determine whether glutamine (GLN)-enriched TPN can prevent such changes. Eighteen male Wistar rats were randomly divided into three groups of six animals. A central line was inserted via the jugular vein. The CHOW group received an infusion of saline and chow ad libitum; the TPN group received a standard TPN solution; the GLN group received TPN solution identical to the TPN group except for the addition of 2% glutamine. Diets were isonitrogenous and isocaloric and fed for 7 days. On Day 7, following laparotomy, the bile duct was cannulated, and bile flow was measured and collected for assay of total and unconjugated bilirubin (TB & UCB) and of total and ionized calcium (TCa & iCa). Caval blood was obtained for liver function studies. The results demonstrated that the hepatic bile flow was decreased in the TPN group compared with the CHOW and the GLN groups. The bilirubin concentration of bile, especially unconjugated bilirubin, in the TPN group was significantly higher when compared with the CHOW and GLN groups (P < 0.01). The ionized calcium concentration of bile in the TPN group was also increased significantly when compared with the CHOW and GLN groups (P < 0.05). From these data, we conclude that TPN induces lithogenic effects in hepatic bile, including increased bilirubin and calcium concentrations and decreased hepatic bile flow. Addition of glutamine to the TPN solution prevents these lithogenic changes.