Rational construction of heterostructures (especially p-n heterostructures) is an excellent strategy for efficient charge separation and desirable photocatalytic performance. Regretfully, the measurements of charge separation efficiency of p-n heterojunction are often monotonous. In this paper, a well-designed p-CuAl2O4/n-Bi2MoO6 heterojunction was successfully synthesized by assembling Bi2MoO6 nanosheets (NSs) on the electrospun CuAl2O4 hollow nanofibers (HNFs) through a solvothermal method. The Bi2MoO6 NSs tightly connect with CuAl2O4 HNFs via strong chemical bonding, and the loading amount can be easily controlled by the concentration of precursor. Significantly, for the first time the charge separation efficiency of the heterojunction is systematically investigated via its photo electrochemical responses under visible light irradiation with and without trapping agents, and the highly efficient charge separation and transfer of the heterojunction is tightly confirmed. Photocatalytic experiments show that the highly efficient charge separation and transfer of the heterojunction results in the enhanced photocatalytic performance for degrading all pollutant models (Rhodamine B, Methyl Orange, Cr(VI), and 4-nitrophenol), whose reaction rate is 1 order of magnitude higher than the reference samples. The work may be useful for rational constructing p-n heterojunctions and provide novel insights to investigate the photoelectrochemical and photocatalytic performance.