Peroxidase-Catalyzed Co-oxidation of 3,3´,5,5´-Tetramethylbenzidine with 2-Amino-4-nitrophenol, 4,4´-Dihydroxydiphenylsulfone, and Their Polydisulfides in Aqueous and Micellar Media

E. I. Karasyova¹, Yu. P. Losev², and D. I. Metelitza^1*

¹Institute of Bioorganic Chemistry, National Academy of Sciences of Belarus, ul. Kuprevicha 5/2, Minsk, 220141 Belarus; fax: (375)-(172) 63-7274; E-mail: enzyme@ns.iboch.ac.by

²School of Chemistry, Belorussian State University, Leningradskaya ul. 14, Minsk, 220050 Belarus; fax: (375)-(172) 26-5942; E-mail: organic@chem.bsu.unibel.by

^* To whom correspondence should be addressed.

Received April 17, 2000; Revision received January 23, 2001

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The effects of different concentrations of 2-amino-4-nitrophenol (ANP) and of its polydisulfide (poly(ADSNP)) on peroxidase-catalyzed oxidation of 3,3´5,5´-tetramethylbenzidine (TMB) were studied at 20°C in reversed micelles of AOT (0.2 M) in heptane and in mixed reversed micelles of AOT (0.1 M)-Triton X-100 (0.1 M) in isooctane supplemented with 15% hexanol. The oxidation of TMB was activated nearly twofold in the presence of ANP and nearly fourfold in the presence of poly(ADSNP) in reversed micelles of AOT, whereas in the mixed micelles oxidation of the TMB-ANP pair was associated with inhibition of TMB conversion and poly(ADSNP) activated oxidation of TMB. The co-oxidation of TMB with 4,4´-dihydroxydiphenylsulfone (DDS) and with its polydisulfide (poly(DSDDS)) at different concentrations of phenol components was accompanied by activation of TMB conversion in 0.01 M phosphate buffer (pH 6.4) supplemented with 5% DMF and in reversed micelles of AOT in heptane. The effect of pH of the aqueous solution on the initial oxidation rate of the TMB-DDS and TMB-poly(DSDDS) pairs and also the effect of hydration degree of reversed micelles of AOT on conversion of the same pairs by peroxidase were studied. A scheme of peroxidase-dependent co-oxidation of “aromatic amine-phenol” pairs is proposed and discussed. A significant part of this scheme is a nonenzymatic exchange of phenoxyl radicals with amines and of aminyl radicals with phenols.

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KEY WORDS: horseradish peroxidase, tetramethylbenzidine, 2-amino-4-nitrophenol, 4,4´-dihydroxydiphenylsulfone, phenol polydisulfides, co-oxidation, inhibition of peroxidase reactions

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