A new sorbent was prepared by loading rhodamine B on Amberlite IR-120. Various physico-chemical parameters such as effects of adsorbate concentration, contact time, pH, and temperature on the sorption of the dye have been studied. Thermodynamic parameters (ΔH° and ΔS°) were also evaluated for the sorption of dye. Kinetic studies revealed that the sorption of the dye was best fit for pseudo-second-order kinetic. The metal ion uptake in different solvent systems has been explored through column studies. On the basis of distribution coefficient (Kd), some heavy metal ions of analytical interest from binary mixtures have been separated. The limit of detection (LOD) for the Ni2+ and Fe3+ metal ions was 0.81 and 0.60 µg L−1, and the limit of quantification (LOQ) was found to be 2.72 and 2.0 µg L−1. This sorbent has also been successfully applied in the analysis of multivitamin formulation. The applicability of the modified resin in the separation of heavy metals constituting real and synthetic samples has been explored.