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To elucidate the response of oxidative metabolism, triggered by elevated ammonium (NH4+) concentrations, on root growth of wheat seedlings, Yumai 49 (NH4+-tolerant) and Lumai 15 (NH4+-sensitive) cultivars were supplied with either 5.0 mmol L–1 NH4+-N (EAC) or 5.0 mmol L–1 NO3–-N (CON) under hydroponic conditions. Root growth in both cultivars was significantly reduced under EAC, and the negative effect was greater in Lumai 15. EAC enhanced the activities of monodehydroascorbate reductase and dehydroascorbate reductase in the roots of both cultivars, while it decreased ascorbic acid (ASA) content and GDP-mannose pyrophosphorylase (GMPase) activity at the 12th day after treatment in Lumai 15 by 62.0 and 71.4％; and in Yumai 49 by 38.8 and 62.2％, respectively, indicating that the regeneration of ASA was increased, but the biosynthesis of ASA was reduced under EAC treatment. Moreover, EAC increased DHA/ASA, reactive oxygen species (ROS), and malondialdehyde contents, as well as antioxidant enzyme activities in the roots of both cultivars. Relatively greater increases in ROS and soluble sugar, and lower antioxidant enzyme activities in Lumai 15 indicate severe disruption of oxidative metabolism when compared to Yumai 49. Results reveal that the reduction of ASA biosynthesis via decreased GMPase activity under the EAC condition probably acts as a trigger for accumulated ROS and imbalanced redox status, resulting in root growth inhibition during wheat seedling growth stage. Yumai 49, being an NH4+ -tolerant cultivar, had the stronger capacity to protect itself from oxidative stress, which allowed it to retain a lower DHA to ASA ratio by maintaining a better redox homeostasis than could be maintained in the NH4+ -sensitive cultivar Lumai 15.