Stopped-flow spectrophotometric determination of hydrogen peroxide with hemoglobin as catalyst

A rapid and sensitive method was proposed for the determination of hydrogen peroxide based on the catalytic effect of hemoglobin using o-phenylenediamine as the substrate. Stopped-flow spectrophotometric method was used to study the kinetic behavior of the oxidation reaction. The catalytic effectiveness of hemoglobin was compared with other four kinds of catalysts. The initial rate of the formation of the reaction product 2,3-diaminophenazine at the wavelength of 425 nm was monitored, permitting a detection limit of 9.2×10−9 mol/l H2O2. A linear calibration graph was obtained over the H2O2 concentration range 5.0×10−8–3.5×10−6 mol/l, and the relative standard deviation at a H2O2 concentration of 5.0×10−7 mol/l was 2.08%. Satisfied results were obtained in the determination of H2O2 in real samples by this method.