We report that endotoxin treatment results in decreased amounts of peroxisomes as well as changes in structure and function of peroxisomal membranes. Peroxisomes isolated from the liver of control and treated animals showed a marked decrease in total protein, but no significant alteration in the sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) protein profile. However, the Western blot study of the peroxisomal beta-oxidation enzymes and catalase showed an increase in those enzymes in the peroxisomal peak of normal density in endotoxin-treated rats. Disintegration of peroxisomal membranes by carbonate treatment from endotoxin-treated liver and change in the fluidity of peroxisomal membranes suggests alterations in peroxisomal membrane structure. No such alterations were found in mitochondrial or microsomal membranes of endotoxin-treated livers. The lipid analysis of these organelles showed that the only organelle affected was the peroxisome, with a significant decrease in the phospholipid and cholesterol concentrations. To understand the mechanism of endotoxin-mediated alterations in peroxisomes, we studied the possible role of Kupffer cell secreted soluble factors (tumor necrosis factor alpha [TNF-alpha]) on the peroxisomal structure/function. Inactivation/elimination of Kupffer cells by gadolinium chloride before endotoxin treatment did not normalize the overall peroxisomal protein amount and the lipid composition of isolated peroxisomes. However, the levels of individual protein amount in remaining peroxisomes were normalized. Endotoxin also decreased peroxisomal beta-oxidation, and this was partially restored with gadolinium treatment. These results clearly show that peroxisomes are severely affected by endotoxin treatment and suggest that the damage to this organelle may contribute, at least in part, to endotoxin-induced hepatic cytotoxicity.