Oxidation kinetics of 1,5-dihydroflavin by oxygen in non-aqueous solvent.

Abstract

The oxidation of reduced flavin anion by oxygen in aprotic dimethylformamide starts with the rapid preequilibrium formation of an anionic intermediate (ɛ375∼ 8.0 mM−1· cm−1). The preferred irreversible decay mode of this complex is the heterolytic rupture yielding oxidized flavin and hydrogen peroxide anion. The oxidation kinetics of neutral reduced flavin in unbuffered dimethylformamide are strictly dependent on the flavin concentration. In dilute solution, the reaction kinetics are identical to those observed with the anionic form, which requires proton eviction on formation of the flavin-oxygen complex. The whole preequilibrium step is extremely fast. Increasing the amount of neutral reduced flavin results in secondary kinetic effects corresponding to neutralization (pKa in dimethylformamide, pK*= 11.1) of the intermediate. The rate constant for the heterolytic splitting of the neutral complex is 2300-fold lower than that of the anionic form. Two sorts of pH-dependent coupled reactions between the peroxydihydroflavin adduct and uncomplexed reduced flavin appear in buffered dimethylformamide. A dismutation-type reaction yielding hydrogen peroxide and 2 mol of the flavin radical, forms the first mode. Its rate is maximum around the pK* of reduced flavin (10.15). Dismutation of the flavin radical results in autocatalytic kinetics. The second concerted mode (pK*= 10.4) allows simultaneous release of two oxidized flavin and water molecules; it has been characterized as a two-step process. The flavin-oxygen complex also forms in aqueous media, on the acidic side, with an estimated pK near neutrality. On the other hand, direct electron transfer without intermediate formation is the preferred path under strongly alkaline aqueous conditions, yielding flavin and hyperoxide radicals as the primary products.