Pulsed light (PL) was used to remove Ochratoxin A (OTA) from clear grape juice in this study. Firstly, the factors of OTA degraded by PL were explored, and the OTA degradation rate was 95.29% after optimization by response surface methodology. Dose-response curves were most suitable for kinetic fitting of OTA degradation by PL treatment (R2 = 0.9937). Besides, results showed that pH value, soluble solids content and total organic acid content did not change significantly, little changes in color difference (ΔE < 3), while the content of monomer phenol decreased (12.81%–30.95%) with increased PL application time. The contents of alcohols, esters and aldehydes significantly decreased in aroma components, while the contents of ketones and acids significantly increased, alkanes were not affected. Results of animal experiments showed that PL treatment greatly alleviated the toxic effects of high dose OTA on mice. LC-MS/MS also showed that OTA produced non-toxic OTα and phenylalanine when treated with PL.