Drought and heat stress often co-occur in nature, and their combined effects are a major driver of crop losses, causing more severe damage to plant metabolism than when they occur individually. This study investigates the responses of three edamame cultivars (AGS429, UVE14, and UVE17) to combined drought and heat (DH) stress, with emphasis on the reactive oxygen species (ROS), antioxidative mechanisms and cell wall modifications. Malondialdehyde (MDA), electrolyte leakage (EL), and hydrogen peroxide (HO) were used to measure oxidative stress and membrane damage. The non-enzymatic (ascorbic acid, AsA) and enzymatic (superoxide dismutase, ascorbate peroxidase (APX), guaiacol peroxidase, and glutathione reductase) antioxidant responses were determined spectrophotometrically. Cell wall biomass composition (cellulose, hemicellulose, lignin, and phenols) was determined using Fourier transform Infrared Spectroscopy and spectrophotometry. Ascorbate peroxidase activity and AsA content in DH-stressed AGS429 at flowering strongly correlated to reduced lipid peroxidation (r = -0.97 and - 0.98). Cultivar UVE14 accumulated high AsA under DH stress at both growth stages, which, in turn, was positively associated with total phenolic content (r = 0.97), APX activity, and holocellulose, suggesting enhanced ROS-dependent oxidative polymerisation. On the contrary, poor ROS quenching in UVE17 led to MDA accumulation (p ≤ 0.05), leading to high EL and poor cellulose synthesis at pod-filling (r = -0.88). Therefore, at the physio-biochemical level, AGS429 and UVE14 showed DH stress tolerance through enhanced antioxidative responses and cell wall modifications, while UVE17 was susceptible. Identifying the key biochemical traits linked to DH stress tolerance in edamame offers novel insights for breeding more resilient edamame cultivars.