In order to study the DNA protection and possible mechanisms of vitexin, it was measured by using three spectrophotometric assays, including DNA protection, Fe²⁺-complexing, and Cu²⁺ reducing. In addition, its Fe²⁺-complexing reaction was also analyzed using UV-Vis spectra. The results showed that the IC₅₀ values of vitexin were (919.6±23.6), (9.3±1.2) and (421.7±29.2) μmol/L, respectively, for DNA protection assay, Fe²⁺-complexing assay, and Cu²⁺ reducing assay. When mixed with Fe²⁺, vitexin gave a UV-Vis peak at 554 nm \[molar absorptivity ε=2 680.74 L/(mol·cm)\]. In conclusion, vitexin can effectively protect DNA from ·OH induced damage. The protective mechanism may include electron-transfer (ET) and Fe²⁺-complexing, and its Fe²⁺-complexing site is possibly between 4-carbonly and 5-hydroxyl groups.