Oranges are a rich source of bioactive compounds with recognized benefits for human health. To guarantee high quality and production levels, citrus farms usually employ the combination of selected cultivars with well adapted rootstocks. This study analyzes the impact of four different citrus rootstocks (Forner-Alcaide no.5, ‘Cleopatra mandarin’, Citrus volkameriana and Carrizo citrange) on the bioactive compounds and antioxidant activity of two orange cultivars (‘Salustiana’ and ‘Sanguinelli’) widely used in the orange juice industry. For the hydrophilic fraction, the phenolic compound, anthocyanin, and organic acid profiles were determined by HPLC-DAD-HRMS, and the antioxidant activity by ABTS, DPPH, and ORAC assays. Besides, the total carotenoids and ABTS concentrations were calculated for the hydrophobic fraction. A set of three flavanones, one flavone, and eight anthocyanins were tentatively identified and quantified in the orange cultivars tested. The predominant phenolic compounds obtained in both orange cultivars were hesperidin and narirutin, while cyanidin-3-O-(6″-malonyl) glucoside followed by cyanidin-3-O-rutinoside and cyanidin-3-O-glucoside were the main anthocyanins found in the ‘Sanguinelli’ cultivar. Citric acid, followed by malic, oxalic, and ascorbic acids were the main organic acids. The higher amount of antioxidant compounds was found in fruit from the Forner-Alcaide no.5 rootstock. These results indicate that Forner-Alcaide n.5 affects positively the phenolic and organic acid composition and the antioxidant capacity of ‘Sanguinelli’ and ‘Salustiana’ cultivars, and is therefore a good option for the sector based on the healthy promoting properties.