The invention discloses a preparation method for a one-step synthetic multilayer nanometer box-shaped CuO-CeO2 catalyst for CO preferential oxidation reaction in hydrogen-rich gas. The catalyst is composed of a plurality of layers of homocentric-square-shaped nanometer box structures which are composed of nanometer CuO and CeO2, triblock copolymer serves as a template, cupric nitrate and cerium nitrate serve as raw materials, and a simple one-step method is adopted to synthesize the catalyst. The multiple layers of cavity structures of the prepared catalyst expose more CuO contact interfaces and CeO2 contact interfaces, and therefore more active sites are provided for the CO preferential oxidation reaction. The catalyst with the special features has good performance that CO complete conversion is achieved at the low temperature, and wide temperature windows with CO being completely converted exist.