REACTIONS BETWEEN DIAPHORASE AND QUINONE COMPOUNDS IN BIOELECTROCATALYTIC REDOX REACTIONS OF NADH AND NAD(+)

A thermostable diaphorase (EC 1.6.99.-) is used to oxidize NADH electrocatalytically, during which a variety of quinone compounds and several kinds of flavins are studied as mediators. The enzyme-catalysed electrolytic oxidation, i.e. bioelectrocatalytic oxidation, of NADH proceeds very rapidly with the quinone compounds and flavins as mediators; analysis of the bioelectrocatalytic current by the theory of steady state catalytic currents reveals that the bimolecular reaction rates between the enzyme and the quinones whose redox potentials are more positive than -0.28 V vs. Ag\AgCl\KCl(sat.) at pH 8.5 are as high as 10(8) M(-1) s(-1), suggesting the reactions to be diffusion controlled. The redox potential of the enzyme is determined to be more positive by 42 mV than that of NAD(+)\NADH at pH 8.5 using a spectroscopic method. From the thermodynamic point of view, based on the above results, the bioelectrocatalytic reduction of NAD(+) is predicted to be favourable under alkaline conditions. The reaction is realized effectively at pH 10.0 with 7-deoxyadriamycinon as a mediator.