Previous studies have implicated protective effects of vitamin D on insulin secretion and pancreas beta cell function. The goal of the present study is to determine if a combination therapy of 25-hydroxyvitamin D3 and insulin had any advantage over insulin therapy alone on lipid peroxidation and antioxidant activity in the streptozotocin (STZ)-induced diabetic rat. The lipid peroxidation product, thiobarbituric acid-reacting substances (TBARS), was measured to assess free radical activity in the heart, kidney and liver tissues. The enzymatic activities of glutathione peroxidase (GSH-Px), superoxide dismutase (SOD) and catalase (CAT) were measured as indicators of antioxidation in these tissues. Sprague-Dawley rats were made diabetic with a single injection of STZ (75 mg/kg i.p.). Rats were separated into three groups, each containing 10 animals: Group 1, non-diabetic and no drug treatment was given; Group 2, diabetic rats were treated with 3 IU/day subcutaneous (s.c.) insulin; and Group 3, diabetic rats were treated with 3 IU/day (s.c.) insulin plus 1 mg/kg/day per oral (p.o.) 25-hydroxyvitamin D3 for a period of 4 weeks. At the end of the study, TBARS contents of the liver, kidney and heart tissues in Groups 2 and 3 were found to be significantly increased as compared to Group 1 (P < 0.05) and kidney MDA levels in Group 3 were also significantly increased as compared to Group 2 (P < 0.05). The SOD and CAT contents of the heart in Group 2 were significantly increased as compared to Groups 1 and 3 (P < 0.05). GSH-Px activity was unaltered in all groups (P > 0.05). We suggest that a combination of insulin with 25-hydroxyvitamin D3 treatment would not be more beneficial than the use of insulin alone in antioxidant defence of diabetic liver and kidney tissues.