Objective To establish a novel bioartificial liver (BAL) consisting of spheroids of porcine hepatocytes in a hollow-fiber bioreactor, and to perform an in vitro study on its metabolic effects on the serum from severe hepatitis B patients.Methods Hepatocytes were isolated from pup pigs and cultured as aggregate spheroids through rotation and vibration. Phase-contrast microscopy, transmission electron microscopy, and scanning electron microscopy were used for morphological detection of hepatocyte spheroids. The hepatocyte spheroids were then transferred into the shell of a polysulfone hollow-fiber bioreactor, creating a novel BAL. Diluted serum samples of severe hepatitis B patients were circulated for 3 hours each into the bioreactor, by using an extracorporeal circulatory system. Every half hour, including both before and after perfusion, serum samples were collected to assay total bilirubin (TBIL), total protein (TP), albumin (ALB), and globulin (GLB) concentrations in order to judge the metabolic effects of this novel BAL. Results Most hepatocytes had formed spheroids with high viability after 24 hours in culture. After 3 hours of perfusion, when compared with the control group, the serum concentration of TBIL in the treatment group decreased significantly (P<0.01), but the serum concentrations of TP and ALB increased significantly (P<0.05). Conclusions Hepatocytes can be conveniently cultured as aggregate spheroids through a rotation and vibration method. The novel BAL is efficient in removing bilirubin from the serum of severe hepatitis B patients, and in supplying the serum with ALB. Thus, the BAL might provide effective therapy for patients with severe hepatitis B.The extracorporeal bioartificial liver support system (EBLSS) has been established to compensate for hepatic function in patients with severe hepatopathy, and to provide supportive therapy for these patients. EBLSS has an important status in the research of artificial organs. At present, the study of artificial livers has progressed to the stage of the hybrid bioartificial liver (BAL) consisting of mammalian hepatocytes in artificial synthetic materials.1 This type of artificial liver can not only provide the biological functions of living hepatocytes, but can also effectively prevent rejection from occurring. In recent years, noticeable progress has been achieved in hybrid BAL research. Fundamental research and animal experiments involving hybrid BAL are carried out continuously. Meanwhile, clinical studies showed that patients with fulminant hepatic failure (FHF) have been successfully treated using BAL.2,3Patients with severe hepatitis have a high morbidity in China. Since causal treatment and traditional supportive therapy cannot compensate for the function of hepatocytes, patients with severe hepatitis also have a high mortality rate. BAL can theoretically substitute for liver function during the necrosis stage of hepatocytes and provide favorable conditions for the rehabilitation of liver function.4 Thus the condition and the cure rate of patients with severe hepatitis can be improved. Few studies, however, reported the use of BAL. In this study, hepatocytes were cultured as aggregate spheroids and transferred into the shell of a hollow-fiber bioreactor in order to set up a novel BAL. The diluted serum of patients with severe hepatitis B was circulated regularly in an extracorporeal system including the BAL. By this way, we sought to find an effective new method to treat severe hepatitis B.