Seasonal changes in levels of reactive oxygen species (ROS), low-molecular weight antioxidants and activities of antioxidant enzymes were analyzed in relation to the freezing tolerance of 1-year-old needles from four populations of Norway spruce. Throughout the study period (from January until May), no significant changes were observed in the superoxide anion radical (O2-) or hydrogen peroxide (H2O2) levels in the needles. By contrast, a marked reduction was observed in concentrations of lowmolecular weight antioxidants, including flavonoids (FL), ascorbic acid (AsA) and slight glutathione (GSH), during deacclimation. The activities of superoxide dismutase (SOD) (EC. 126.96.36.199.) and guaiacol peroxidase (PO) (EC. 188.8.131.52.) also decreased significantly. The activity of catalase (CAT) (EC. 184.108.40.206.) did not change significantly. Levels of low-molecular weight antioxidants (AsA, FL and GSH) and SOD activity were correlated significantly with freezing tolerance in the studied populations. The reactions were similar in all populations. This suggests that the response of the antioxidant system depends more strongly on climatic conditions than on population origin. The ability of spruce trees to cope with active oxygen species is discussed as an aspect of defense and a factor associated with freezing tolerance.