Identifying mechanisms involved in cold tolerance can play an important role in maintaining yield. In this study, physiological and biochemical responses, including malondialdehyde (MDA), cell membrane stability index (MSI), and activities of antioxidant enzymes in five autumn bread wheat cultivars and one spring tolerant cultivar in four treatments: under normal conditions (23°C), acclimation (4°C) and cold stress (-5°C) were studied in acclimated and non-acclimated wheat plants. Analysis of variance showed significant differences between the effects of cultivars and environment temperatures on the membrane stability index, while in other indices, significant differences derived between interactive effects. Results showed that lower temperatures increased MDA, while decreased MSI in different cultivars. As plants exposed to cold stress, their total protein content as well as activities of superoxide dismutase, catalase, ascorbate peroxidase and guaiacol peroxidase increased only in cold-acclimated plants, while decreased significantly in non-acclimated strands. Results also showed lower decrease of membrane stability index and MDA in Norstar and Baran cultivars under stress conditions. It seems that in non-acclimated plants under cold stress, decrease in activity of antioxidant enzymes has impaired plant's ability to withstand stress. Therefore, a decrease in cell damages as well as an increase in capacity of cell defense systems, coming down to acclimation process, can be regarded as important factors contributing to tolerance to cold stress.