OBJECTIVE To investigate the tolerance time limits from warm ischemia to cold preservation of liver grafts. METHODS Orthotopic liver transplantations (OLTs) were performed on Bama miniature swine. Morphological and functional changes of the liver grafts and biliary tracts after 10 minutes of warm ischemia followed by different durations of cold preservation and its reversibility were investigated. RESULTS When the grafts were subjected to 10 minutes of warm ischemia followed by less than 16 hours of cold preservation, all animals could survive 1 week and there was no animal death from biliary necrosis. However, when the cold preservation time exceeded 16 hours, the incidence of biliary necrosis was significantly increased (P<0.05), and recipient death from bile leaks occurred. With further prolongation of the cold preservation time, primary graft nonfunction and intraoperative or early postoperative deaths occurred and the living animals all developed biliary necrosis. When compared with the less than 16 hours cold preservation group, the morphological scores and apoptosis index of the epithelial cells of bile ducts in grafts after reperfusion were significantly elevated in the more than 16 hours cold preservation group (P<0.05) and the activity of Na+-K+-ATPase and Ca2+-ATPase of bile ducts in grafts were also significantly reduced (P<0.05). Liver function tests showed that the recoveries of AST, AST, GGT and ALP were quicker in the 16 hours cold preservation group then those over 16 hour preservation ones. Correlation analysis revealed that the incidence of biliary necrosis was significantly correlated with the morphological score (r = 0.972) and with the apoptosis index of the epithelial cells of bile ducts in grafts after reperfusion (r = 0.931) and also correlated negatively (P<0.01) with the activity of Na+-K+-ATPase (r = -0.973) and Ca2+-ATPase (r = -0.973). CONCLUSIONS It is concluded that with 10 minutes of warm ischemia, cold preservation of the grafts should not be longer than 16 hours in order to avoid early biliary necrosis, and the corresponding tolerance time limit of the livers to the cold preservation was less than 20 hours.