SUBSTITUTION OF PEROXIDASE IN TRINDER REAGENT WITH IRON(II) FOR THE DETERMINATION OF HYDROGEN-PEROXIDE IN ENZYMATIC-REACTIONS BY APPLYING FLOW-INJECTION

The kinetics and mechanism of the reactions between 4-aminoantipyrine and/or 3,5-dichloro-2-hydroxybenzenesulfonic acid with Fe2+ in the pre sence of H2O2 have been investigated. A differential spectrophotometri c flow injection (FI) method for the determination of H2O2 has been de veloped, by substituting horseradish peroxidase (PO) in Trinder's reag ent with Fe2+. This method has been applied to the determination of gl ucose in human serum, with good results, by using an enzymic reactor w ith immobilized glucose oxidase and an automated Fl system, after opti mizing its parameters. The agreement between the results of this metho d with those of the routine method used by a hospital clinical laborat ory for the determination of glucose in human serum was very good. The correlation line of the two methods shows a slope of 1.03 and a corre lation coefficient (r) of 0.9997. The method was also applied to the d etermination of H2O2. The working curve, with Fe2+ as promoter, was li near for 10-400 mumol 1-1 H2O2 compared with 5-170 mumol l-1 H2O2 when PO was used. The slopes of the curves for Fe2+ and PO are 1.69 and 4. 35 A l mmol-1, respectively, showing a reduced sensitivity for the pro posed method compared with the PO method.