The effect of lignin supplementation to a diet contaminated with zearalenone (ZEA) on antioxidant status was studied in female chickens of the ISA BROWN laying strain. From the day of hatching to 2 weeks of age, four groups of chickens were fed the same uncontaminated control diet. After 14 days, Group 1 (control) continued to receive the uncontaminated diet, while Group 2 was fed an identical diet enriched with 0.5% chemically modified lignin. Simultaneously, chickens of Group 3 were switched to a diet contaminated with 7.9 mg/kg ZEA and those of Group 4 to an identical contaminated diet supplemented with 0.5% lignin. At 6 weeks of age blood and tissue samples were collected. Feeding of a diet contaminated with a high level of ZEA resulted in elevated glutathione peroxidase (GPx) activity in the duodenal mucosa and kidney tissues, and an increased γ-glutamyltransferase (GGT) activity in the plasma, indicative of oxidative stress. In the liver tissue, no mycotoxin-induced response in GPx and thioredoxin reductase (TrxR) activities occurred, and the malondialdehyde (MDA) level was even reduced. Neither the plasma levels of retinol and α-tocopherol nor the activities of superoxide dismutase in erythrocytes and GPx in blood were affected in birds fed the contaminated diet. The only effect of lignin supplemented to the contaminated feed was that it prevented the increase of GPx activity in the duodenal mucosa as an indicator of oxidative stress.