Using alkyl trimethyl ammonium chloride (CTAC) as template, tetraethyl orthosilicate (TEOS) as silica source, methanol as co-solvent, sodium hydroxide as catalyst, single scattered and highly ordered mesoporous silica microspheres with different size were synthesized by hydrothermal method, and the synthesized microspheres were characterized by SEM. The effects of the CTAC concentration, the TEOS concentration, the methanol ratio and the temperature on particle size and dispersibility of the prepared silica spheres were studied, respectively. At the same time, the synthesized samples were characterized by XRD, BET, HRTEM and other methods. The results showed that the synthesis of monodispersed spherical silica had ordered hexagonal mesoporous structure and high specific surface area.