Oxygen radicals have been implicated in the pathogenesis of renal injury induced by extrahepatic cholestasis. We conduct this study to investigate whether melatonin can have a protective effect against such injury. Either normal saline or gentamicin with or without melatonin was injected into rats that received either a bile duct ligation or a sham operation. The serum levels of malondialdehyde and total antioxidative activity were measured. The kidney was fixed for histologic scoring of renal injury. The serum malondialdehyde level was highest in the rats that received both bile duct ligation and gentamicin injection. Treatment with melatonin significantly increased the serum total antioxidative activity and reduced the serum malondialdehyde concentration. The mean score of renal injury, highest in the rats receiving bile duct ligation and gentamicin injection, was significantly reduced with melatonin treatment. By reducing the systemic oxygen radicals, supplementation with antioxidants exerts a protective effect on the renal injury induced by extrahepatic cholestasis.