Author
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Jacks, Thomas |
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Submitted to: Analytical Letters
Publication Type: Peer Reviewed Journal Publication Acceptance Date: 2/1/2004 Publication Date: 4/1/2004 Citation: Jacks, T.J. 2004. Spectrofluorometric determination of heme and nonheme haloperoxidase activity. Analytical Letters. 37:1177-1184. Interpretive Summary: It is important to be able to measure enzymes present in crop plants because they can effect the quality and quantity of agricultural products. This paper describes a new method for measuring enzymes that cause discoloration of plant products and also protect plants against infection. The method is based on the reaction of the enzyme with a chemical to yield a product that fluoresces when illuminated with ultraviolet light. The intensity of the fluorescence is directly proportional to both the amount and the strength of the enzyme and is easily measured. In this manner, scientists, who are the principal users of this technological advance, are able to readily and reliably measure the amounts and strengths of the enzymes. Technical Abstract: A spectrofluorometric procedure for determining catalytic activities of heme and nonheme haloperoxidases is described. Haloperoxidases (E.C. 1.11.1.10) are oxidoreductases that catalyze the oxidation of halide ions to hypohalites by hydrogen peroxide. Reaction products of the enzymes were found to react with kojic acid, generating intense fluorescence with excitation and relaxation (emission) wavelenghts at 395 and 495 nm, respectively. Continuous monitoring of the enzymically catalyzed reactions was accomplished spectrofluorometically. Nonheme haloperoxidase, which differs from other haloperoxidases not only by the absence of a heme prosthetic group but also by catalyzing the peroxidation of alkyl acids, was assayed by coupling peroxyacid generation to bromide ion oxidation. Utility of the assay procedure was shown by determining activities of plant peroxidase, fungal chloroperoxidase, mammalian myeloperoxidase and bacterial nonheme haloperoxidase. |
