Higher uptake and translocation of copper (Cu) into plant tissues can cause serious physiological and biochemical alterations in root and leaf tissues of plants. The present study investigates the ameliorative role of salicylic acid (SA) and ascorbic acid (AsA) against Cu-induced toxicity changes in cotton genotypes (two parental lines (J208, Z905) and their hybrid line (ZD14)). To study the tolerance potential against Cu (100 μM) stress, 2-week-old cotton seedlings were pretreated with 100 μM either SA or AsA for three days. Elevated Cu concentration in nutrient media increased Cu accumulation in roots and shoots of all the three cotton genotypes studied. Roots were the main Cu storage site, followed by leaves and stems. Increased cellular Cu concentration significantly inhibited the root and shoot development, although leaf growth was more sensitive toward Cu toxicity. Cu-induced oxidative stress to cotton leaves was evident from significantly increased hydrogen peroxide (H2O2) contents and lipid membrane damage. Increasing Cu translocation toward cotton leaves strongly influenced the malondialdehyde (MDA) contents, which, in turn, inhibited biomass production. SA and AsA pretreated cotton seedlings showed better growth under Cu stress. Despite increase in overall Cu uptake, the SA-pretreated seedlings could defy Cu toxicity through inhibited Cu translocation and modification in the activities of antioxidative enzymes. Whereas, tolerance to Cu-induced toxicity in AsA pretreated plants was associated with Cu exclusion from tissues and reduction of the overall Cu uptake. The present study revealed that the alleviatory role of AsA was significantly higher than SA regarding Cu stress in our experimental cotton genotypes. Furthermore, the hybrid cotton genotype (ZD14) performed well followed by J208 and Z905 in the present experimental setup.