Leaf rolling (LR), particularly in crops, is a dehydration avoidance mechanism that results from water loss in plants under drought conditions. Two maize cultivars (Zea mays L.) differing in their tolerance to drought (cultivar Side was selected as stress-tolerant and cultivar Karacay was selected as stress-sensitive) under PEG-induced drought stress were used in this study to understand the effects of polyamines (PAs) on LR response and the role of the antioxidant system in the delayed LR process. Aerial parts of the seedlings were excised prior to the experiment and then submerged in a Hoagland nutrient solution with or without 0.1 mM Putrescine and 0.1 mM Spermine for 17 h, after which they were submitted to osmotic stress treatments (5% PEG) for 24 h. LR was physically prevented in order to clarify the relationship between PAs and antioxidant systems in LR response. Plants were evaluated by measuring the accumulation of hydrogen peroxide (H2O2), lipid peroxidation, antioxidant enzyme activities, and nonenzymatic antioxidant levels. Exogenous applications of PAs prevented water loss and delayed LR in comparison with the control (seedlings treated only with PEG). PAs almost totally prevented increases in lipid peroxidation. Antioxidant enzyme activities in rolled leaves were induced in response to osmotic stress in both maize seedlings via PA application. Ascorbate, glutathione, and endogenous PA levels increased as a result of PA applications. H2O2 content was lower in PA-pretreated plants than in the control in both cultivars. The activity/effectiveness of antioxidant system components of the physical prevention of leaf rolling (PLR) group of plants were lower than those of the group of plants with rolled leaves, especially in sensitive Karacay, whereas more induced H2O2 content and lipid peroxidation were determined in the PLR group. In addition, exogenous applications of PAs further increased the components of Side's antioxidant system in comparison with Karacay. When considered together, the data suggest that exogenous PAs may have a H2O2-mediated regulatory role in the LR process through the induction of antioxidant machinery. A stimulated antioxidant system decreases oxidative stress damage from excess accumulation of H2O2, thus delaying LR. In summary, exogenous PAs could help plants invert the adverse effects of osmotic stress and might play a key role in providing tolerance in plants through modulating the antioxidant system and decreasing H2O2 levels and water loss in plants via LR, as an alternative drought-protection mechanism. In addition, maize cultivars with late LR by PA applications may be provided as an opportunity for improving yield potential in drought-prone areas.