Multifunctional magnetic microspheres have enormous potential in diverse fields. In this work, surface chiral-modified magnetic microspheres as chiral selectors, are prepared by polymerizing maleic anhydride-β-cyclodextrin (MAH-β-CD) for the enantioselective absorption of four enantiomers. The successful grafting of MAH-β-CD onto the surface of magnetic microspheres is confirmed by transmission electron microscopy, X-ray diffraction, Fourier transform infrared spectroscopy and thermogravimetric analysis. The prepared functional microspheres have a three-ply structure with an average particle size of 550 nm and a high saturation magnetization of 60 emu g⁻¹. The amount of MAH-β-CD modified on the P(MBAAm)@Fe₃O₄ was about 149.1 mg g⁻¹. The analysis results of specific rotation and capillary electrophoresis reveal that the MAH-β-CD-modified Fe₃O₄ microspheres show stronger complexation of (−)-enantiomers than (+)-enantiomers. In addition, the MAH-β-CD-modified Fe₃O₄ microspheres have stronger enantioselective absorption for double-ring chiral compounds. These chiral-functionalized magnetic microspheres are therefore expected to be an efficient and economical chiral separation method for use in further research.