The nature of compounds present in mixtures of oxidized and reduced flavin mononucleotides

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Abstract
The composition of the equilibrium mixtures obtained by mixing known amounts of FMN and FMNH2 have been studied by spectrophotometry, fluorimetry and electron-spin-resonance spectrometry. The major component of the mixtures containing more than 2 mM total flavin is a species absorbing at 900 m[mu] which we regard as a charge-transfer complex between FMNH2 and FMN. Comparison of the spectrophotometric and electron-spin-resonance data show a close correlation between the extinction at 570 m[mu] and the concentration of free radical. When the radical concentration is plotted against total percentage of reduced flavin at constant flavin concentration, a strongly asymmetric curve is obtained with a maximum at about 70% reduction. Evidence is presented that this asymmetry is due to the presence of another radical species, a complex between the semiquinone and FMNH2. The formation of the complex between semiquinone and FMNH2 is inhibited in the presence of solutes such as caffeine and urea and in various organic solvents. In aqueous solution (0[center dot]12M-phosphate, pH6[center dot]3) our data are suitably fitted by the following equilibria: [image] The effect of changing the solvent on the relation between the extinction at 570 m[mu] and that at 900 m[mu] leads us to the conclusion that the charge-transfer complex (900 m[mu] band) is a precursor of the free semiquinone.
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