Functionalized ionic liquids containing ethyoxyl groups were synthesized and immobilized on magnetic silica nanoparticles (MSNP) prepared by two steps, i.e., Fe3O4 synthesis and silica shell growth on the surface. This magnetic nanoparticle supported ionic liquid (MNP-IL) were applied in the immobilization of penicillin G acylase (PGA). The MSNPs and MNP-ILs were characterized by the means of Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), and vibrating sample magnetometer (VSM). The results showed that the average size of magnetic Fe3O4 nanoparticles and MSNPs were similar to 10 and similar to 90 nm, respectively. The saturation magnetizations of magnetic Fe3O4 nanoparticles and MNP-ILs were 63.7 and 26.9 A.m(2).kg(-1), respectively. The MNP-IL was successfully applied in the immobilization of PGA. The maximum amount of loaded enzyme was about 209 mg.g(-1) (based on carrier), and the highest enzyme activity of immobilized PGA (based on ImPGA) was 261 U.g(-1). Both the amount of loaded enzyme and the activity of ImPGA are at the same level of or higher than that in previous reports. After 10 consecutive operations, ImPGA still maintained 62% of its initial activity, indicating the good recovery property of ImPGA activity. The ionic liquid modified magnetic particles integrate the magnetic properties of Fe3O4 and the structure-tunable properties of ionic liquids, and have extensive potential uses in protein immobilization and magnetic bioseparation. This work may open up a novel strategy to immobilize proteins by ionic liquids.