Antioxidants were characterized in needles from mature spruce trees [Picea abies (L.) Karst.] grown in a forest on the Wank mountain in the Bavarian Calcareous Alps at three altitudes (870 m, 1270 m, and 1700 m). The effect of elevation, season and needle age on the activities of superoxide dismutase, glutathione reductase, ascorbate peroxidase, and the content of ascorbate and glutathione was analyzed. Ascorbate peroxidase and superoxide dismutase activity were little affected by stress from higher altitudes. Superoxide dismutase activity decreased with increasing needle age at all three altitudes. Maximum levels of glutathione and glutathione reductase were found in needles from the highest altitude, but the increase was not linear with increasing elevation. At the middle altitude, levels of glutatione reductase and glutatione were as low as, or even lower than at the lowest site. The mean ascorbate content increased with increasing altitude by about 2 μmol g−1 fresh weight per site. The seasonal change of the ascorbate content in the needles was twofold higher at the upper than at the lower sampling site. These findings suggest that superoxide dismutase and ascorbate peroxidase activities were sufficient to cope with a higher production of toxic oxygen species which probably occurs at high altitudes, and that adaptation was necessary for the antioxidant substrates and their regeneration system.