The ability of plants to survive oxygen deficiency is associated with significant changes in metabolism. Metabolic profiling of wheat seedlings under anoxia and subsequent reoxygenation conditions was performed using GC-MS. A total of 374 and 298 compounds were detected in root and shoot metabolomes, respectively. All intermediates of central metabolism were identified. Early anoxic responses of root and shoot metabolomes showed similarity, leading to the accumulation of amino acids (Ala, GABA and Tyr), carboxylates (lactate and succinate), nucleotides and amines, together with a decrease in sugars. The metabolic response to long-term anoxia varied significantly in the roots and shoots of wheat seedlings and was related to the redistribution of carbon flux from glycolysis predominantly to lipids in the roots, while it was directed to carboxylates and GABA in the shoots. Imposition of 24 h of reaeration after short-term anoxia (6 h) switched the metabolome toward a normoxic profile, predominantly in roots. Anaerobically down-regulated metabolites were accumulated, while anaerobic intermediates were depleted post-anoxia. The effects of more prolonged anoxia on wheat seedling metabolomes were less reversible, particularly in shoots. Interestingly, several metabolites with not fully understood roles (e.g., hydroxyl carboxylates, α,ω-dicarboxylic acids, polyols) were detected under anoxic conditions in wheat seedlings, which could potentially serve as markers of plant sensitivity to oxygen deficiency. © 2025 by the authors.