The invention discloses a preparation method of a cubic iron trioxide monocrystal. The iron trioxide monocrystal with high crystallinity is synthesized by mainly using ferric trichloride hexahydrate and natrium carbonicum calcinatum by adopting a hydrothermal process without adding a dispersing agent and a surfactant, wherein the substances such as a template and the like are not added in the preparation process so that the aftertreatment is convenient. The preparation method has the advantages of low reaction temperature, short time and simple process, low cost, environment friendliness and no use of any toxic iron source and organic solvent.