This paper reports an alternative process to prepare hollow mesoporous silica spheres (HMS) using a single cationic surfactant with a tunable wall thickness and radially oriented pore structures. Using N,N-dimethylformide (DMF) as the intermediate solvent bridging the organic and aqueous phase, hollow mesoporous silica spheres were synthesized with interfacial hydrolysis reactions at the surface of liquid droplets. These spheres have an ordered pore structure aligned along the radial direction, and the wall thickness and sphere sizes can be tuned by adjusting the experimental conditions. Transmission electron microscopy and nitrogen absorption techniques were used to characterize HMS and its formation procedure. A hypothetic formation mechanism was proposed on the basis of a morphology transformation with the correct amount of DMF and a careful observation of the early hydrolysis stages. Au and magnetic Fe3O4 nanoparticles have been encapsulated in the HMS hollow core for potential applications.