Monodispersed spherical TiO2 particles were prepared by hydrothermal crystallization and/or calcination of spherical amorphous particles, synthesized by thermal hydrolysis of TiCl4. The crystallized spherical particles were secondary agglomerates of primary nanocrystallites. Different crystallization routes and conditions provided the spherical TiO2 particles with wide particle characteristics, such as the fraction of crystallization, the size and shape of the primary nanocrystallites, and the specific surface area. The photocatalytic activity showed complex dependence on the crystallization routes and conditions. The complex dependence behavior could be explained by combining the effects of the fraction of crystallization, the specific surface area, and the adsorption ability for hydroxyl ions. Especially, in the present study, the hydrothermally crystallized TiO2 particles with large primary nanocrystallites showed the highest photocatalytic activity. The high photocatalytic activity mainly resulted from the high surface adsorption ability for hydroxyl ions, which was closely related to the well-developed (flat and faceted) morphology of primary nanocrystallite.