Many recent studies and attempts have focused on developing new chiral nano-selector for simple and efficient chiral separations. In this study, a new class of PAMAM-magnetic polymeric materials was prepared by a microwave-assisted method and used for the immobilization of BSA to form chiral magnetic nanoparticles Fe₃O₄@SiO₂@PAMAM-BSA (BSA-PMNPs) (Fig.1). The binding capacity of BSA to PMNPs increases with each generation of growth, which can enhance the chiral discrimination of magnetic nanoparticles prepared in this way. UV-Vis spectrophotometry and confocal laser scanning microscopy were used to evaluate the binding capacity of BSA. The activity of the BSA on the PMNPs was tested using an immunoaffinity approach, with the eluent being assayed by sodium dodecylsulfate polyacrylamide gel electrophoresis. The functional magnetic nanoparticles were then used for the direct separation of D, L-tryptophan, and could be readily separated from the reaction mixtures with a magnet. These results were further confirmed by HPLC analysis of D, L-tryptophan. The reduction in the peak area of (–)-tryptophan was more dramatic than that of (+)-tryptophan and the BSA immobilized onto the G3PAMAM-MNPs showed a much stronger chiral recognition ability than that of the G0PAMAM-MNPs, suggesting that the BSA-PMNPs could be used to improve the chiral separation efficiency because larger amounts of BSA had been immobilized onto the PMNPs. The separation results indicated that the BSA-PMNPs showed much greater affinities for the (–)-enantiomers than the (+)-enantiomers, and effectively demonstrated the potential utility of this material for chiral separations.