Background Intense physical exercise has been associated with an increase of free radical production. When the body's natural defense systems against free radicals are overwhelmed, oxidative stress increases. Aim of the study This study examined the effects of a vitamin antioxidant supplement, (composed of 600 mg α-tocopherol, 1000 mg ascorbic acid and 32 mg β-carotene) on oxidative, hormonal, and enzymatic exercise stress markers during habitual training activity over 35 days. Methods The plasma concentrations of ascorbic acid, α-tocopherol, β-carotene, testosterone, cortisol and lipid peroxides and the serum activities of lactate dehydrogenase and creatine kinase were measured at four time points: pre-supplementation (PS), pre-training (PT), after training (AT) and 24h after training (24h-AT) in 13 professional basketball players of the first Spanish Basketball League (ACB). Results Antioxidant supplementation led to a significant increase of α-tocopherol and β-carotene from PS to PT. Plasma lipid peroxides decreased about 27.7 % after 35 days of antioxidant treatment. A significant decrease of lactate dehydrogenase serum activity was observed during the 24h recuperation time. During this time the anabolic/catabolic balance increased about 29.8 % in the antioxidant supplemented group, although this increase did not reach statistical significance. Conclusions The results of the present study suggest that supplementation with α-tocopherol, β-carotene and ascorbic acid might partially account for the hormonal and enzymatic stress marker profile observed during habitual training activity of professional basketball players.