<p>Drought is a major abiotic stress that adversely impacts on crop productivity and global food security. Proteomics greatly facilitates the study of stress response mechanism in crop plants by identifying new stress-responsive proteins. Currently, most proteomic studies on osmotic stress response in cereal crops, such as maize, have mainly focused on medium to long-term scales, but few studies investigated the early osmotic stress response. In this study, maize roots were subjected to 30&#xa0;min 10% polyethylene glycol (PEG)-6000 induced osmotic stress, the physiological, biochemical and proteomic changes were analyzed. The results showed that short-term osmotic stress led to the significant increase of reactive oxygen species (ROS, especially O₂⁻) level, the content of thiobarbituric acid reactive substances (TBARS) and proline, and the activity of catalase (CAT) and superoxide dismutase (SOD) in maize root tips. Proteomics analysis identified a set of 20 stress-responsive proteins, which were mainly involved in antioxidative stress and energy metabolism (e.g., ATP synthase), and most of them were more abundant under osmotic stress. Obviously, the antioxidant system and energy metabolism pathway play critical roles in the early root response to short-term osmotic stress. Our work provides insights into the mechanisms underlying the early response of plants to osmotic stress. </p>