In the current industrial scenario, chromium (Cr) as a metal is of great importance but poses a major threat to the ecosystem. In the present study, the effect of different levels of Cr, i.e., 0 (no Cr), 50, and 100 µM in the soil on growth, photosynthetic pigments, gas exchange characteristics, oxidative stress biomarkers, antioxidants machinery (enzymatic and non-enzymatic antioxidants), ions uptake, organic acids exudation, and Cr uptake in different parts of plant were investigated with and without the exogenous application of choline chloride i.e., 0 (no choline chloride), 2–5 mM in Cr-stressed spinach (Spinacia oleracea L.). Our results depicted that Cr addition to the soil significantly (P < 0.05) decreased plant growth and biomass, gas exchange attributes, and minerals uptake by S. oleracea as compared to the plants grown without addition of Cr. However, Cr toxicity boosted the production of reactive oxygen species (ROS) by increasing the contents of malondialdehyde (MDA), which is the indication of oxidative stress in S. oleracea and was also manifested by hydrogen peroxide (H₂O₂) contents and electrolyte leakage to the membrane-bounded organelles. Although activities of various antioxidative enzymes like superoxidase dismutase (SOD), peroxidase (POD), catalase (CAT), and ascorbate peroxidase (APX) and non-enzymatic antioxidants like phenolic, flavonoid, and ascorbic acid, anthocyanin contents initially increased up to a Cr level of 50 µM but decreased gradually with the further increased in the Cr level of 100 µM in the medium, compared to those plants which were grown in the control treatment. Results also revealed that the soluble sugar, reducing sugar, and non-reducing sugar were decreased in plants grown under elevating Cr levels but increased the Cr accumulation in the roots and shoots of S. oleracea. Although results also illustrated that the application of choline chloride also decreased Cr toxicity in S. oleracea seedlings by increasing antioxidant capacity and, thus, improved the plant growth and biomass, photosynthetic pigments, gas exchange characteristics, and decrease oxidative stress in the roots and shoots of S. oleracea seedlings, compared to those plants which were not artificially supplied by choline chloride. Research findings, therefore, suggested that the choline chloride application can ameliorate Cr toxicity in S. oleracea seedlings and resulted in improved plant growth and composition under metal stress as depicted by balanced exudation of organic acids.