Oxidative stress is induced under diabetic conditions and possibly causes various forms of tissue damage in patients with diabetes. Recently, it has become aware that susceptibility of pancreatic β-cells to oxidative stress contributes to the progressive deterioration of β-cell function in type 2 diabetes. A hypoglycemic sulfonylurea, gliclazide, is known to be a general free radical scavenger and its beneficial effects on diabetic complications have been documented. In the present study, we investigated whether gliclazide could protect pancreatic β-cells from oxidative damage. One hundred and fifty μM hydrogen peroxide reduced viability of mouse MIN6 β-cells to 29.3%. Addition of 2μM gliclazide protected MIN6 cells from the cell death induced by H₂O₂ to 55.9%. Glibenclamide, another widely used sulfonylurea, had no significant effects even at 10μM. Nuclear chromatin staining analysis revealed that the preserved viability by gliclazide was due to inhibition of apoptosis. Hydrogen peroxide-induced expression of an anti-oxidative gene heme oxygenase-1 and stress genes A20 and p21^CIP1/WAF1, whose induction was suppressed by gliclazide. These results suggest that gliclazide reduces oxidative stress of β-cells by H₂O₂ probably due to its radical scavenging activity. Gliclazide may be effective in preventing β-cells from the toxic action of reactive oxygen species in diabetes.