Within the realm of advancing photocatalysis, the investigation of material defects has perennially commanded paramount attention. However, a nuanced comprehension of the intricate interplay between these defects and the catalytic milieu necessitates deeper exploration. In the present study, oxygen vacancies (Vo) were adeptly harnessed to induce precise carrier localization, thereby effectuating a heightened augmentation of photocatalytic activity. This augmentation was deftly achieved through the deliberate modulation of external illumination parameters, resulting in the extension of carrier lifetimes. Specifically, the drastically enhanced carrier effective mass (m*) and the density of unoccupied state are directly uncovered by the density functional theory (DFT) calculation and Raman spectra. Through the utilization of wavelength dispersive in-situ fluorescence spectroscopy (WDIFS) experiments, it was observed that Vo-rich ZnO nanoparticles (NPs) displayed noteworthy photocatalytic activity around the central frequency of Vo (5.17 × 1014 Hz, 580 nm), owing to the highly efficient utilization of localized carriers. Moreover, based on Einstein rate equations, the charge carrier lifetime was found to be further prolonged with increased external illumination intensity and excitation wavelength. This work provides a methodology for optimizing the functionality of point defect on photocatalysis, where the photo-induced charge carrier localization is expected to be extensively used in further photocatalysis research.