Herein, we developed the dual-function template method to fabricate hollow magnetic nano-spheres (denoted as HMFe-Si-Cn, n=16, 18) with a mesoporous shell and hollow interior structure using alkyl chain trimethoxysilane templating. The shorter chain template directed formation of HMFe-Si-C16 with size of 119 nm, having disordered inkbottle type mesopores and saturation magnetization of 50.01 emu/g higher than that of HMFe-Si-C18 with cylindrical type mesopores. By contrast, Mitomycin C (MMC) loading efficiency of HMFe-Si-C16 was higher owing to the fact that the pore size, surface area, and pore volume of HMFe-Si-C16 were larger than those of HMFe-Si-C18. Besides, MMC loaded HMFe-Si-Cn hollow spheres showed a clear pH-dependent drug release behavior, having a higher release rate in acidic environments of pH 5.7. For the pH 5.7 and 7.4 release, the release kinetic for HMFe-Si-C16-MMC composites follows pseudo-first-order attributable to its special pore structure. For this reason the inner cavity of HMFe-Si-C16 could be labeled with radioisotope 99Tcm to study the magnetic targeting distribution of HMFe-Si-C16 in vivo, and its cytotoxicity against in vitro HeLa cells was also studied. These results indicate the potential of HMFe-Si-C16 in the magnetic targeted drug delivery system.