The production of hydroxyl radical in HRP-NADH-H2O/O2 systems and its application in chlorobenzene removal

The mechanism of radical generation in HRP-NADH-O2/H2O2 systems and state-change of horseradish peroxidase (HRP) was investigated by using ESR and UV measurements, and the novel enzyme-coenzymatic systems were performed to degrade chlorobenzene as a non-phenolic persistent organic pollutants. The UV results showed that compound III was produced from HRP oxidized by hydrogen peroxide with the catalysis of NADH, which would generate hydroxyl radical. The ESR results demonstrated the production of *OH and O2-. in enzyme-coenzymatic system in the presence of O2 or H2O2 with DMPO and POBN as spin-trappers, respectively. In HRP-NADH-H2O2 system, compound III was the main state of HRP in the initial 10 min, and then converted to HRP with generating hydroxyl radical; and after the addition of oxygen, the production of hydroxyl radical was promoted rapidly, as 4 times as that of the system in absence of oxygen. The addition of SOD(Zn-Cu) decreased the production of hydroxyl radical significantly, resulting from that SOD eliminated O2 reduction to O2-. by NADH and then inhibited *OH formation. The results showed that NADH could improve by about 20% enzyme activity of HRP for phenol removal. The removal of chlorobenzene with HRP-NADH-H2O2 and HRP-NADH-H2O2-O2 systems reached 24.6% and 48.2%, respectively, which was much higher than that with traditional enzymatic system (1.42%), showing a promising prospect in proposal