Provided is a method for preparing a compound semiconductor substrate. The method includes coating a plurality of spherical balls on a substrate, growing a compound semiconductor epitaxial layer on the substrate coated with the spherical balls while allowing voids to be formed under the spherical balls, and cooling the substrate on which the compound semiconductor epitaxial layer is grown so that the substrate and the compound semiconductor epitaxial layer are self-separated along the voids. The spherical ball treatment can reduce dislocation generations. In addition, because the substrate and the compound semiconductor epitaxial layer are separated through the self-separation, there is no need for laser lift-off process.