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The Use of an Enzyme Electrode in the Analysis of Indole-3-acetic Acid Oxidase Activity in Avena

¹ Department of Biology, The College of Wooster, Wooster, Ohio 44691

A flexible analytical system which allows for the continuous potentiometric monitoring of the disappearance of an electrochemical species, ferrocyanide, by the peroxidase enzyme is described. The ability of peroxidase to mediate the oxidation of indole-3-acetic acid is followed by observing the competition of indole-3-acetic acid with ferrocyanide for the peroxidase enzyme. This is accomplished by examining potentiometrically the decrease in the rate of ferrocyanide oxidation with increasing indole-3-acetic acid concentration. Homogenates of Avena sativa coleoptiles are investigates for both peroxidase and indole-3-acetic acid oxidase activity. Observations are made with respect to H₂O₂ and ferrocyanide in the presence and absence of indole-3-acetic acid and naphthalene acetic acid and several interpretations of the reaction kinetics are postulated. Solutions previously assayed for indole-3-acetic acid oxidase activity, when dialyzed and reassayed for peroxidase activity, demonstrated an unimpaired ability to oxidize ferrocyanide peroxidatively, suggesting interpretations of the bisubstrate situation which differ slightly from interpretations given in the literature.