AMPEROMETRIC NADH DETERMINATION VIA BOTH DIRECT AND MEDIATED ELECTRON-TRANSFER BY NADH OXIDASE FROM THERMUS-AQUATICUS YT-1

Using NADH oxidase extracted from Thermus aquaticus YT-1, the determin ation of NADH by the direct oxidation of labile FADH(2) and by mediate d electron transfer using the Ru(III/IV) redox couple is described. A scheme for enzyme amplification is outlined using alcohol dehydrogenas e (ADH) and NADH oxidase in solution, and enables 1-10 mu M NADH to be determined. Detection is performed at +150 mV using the RuCl62-/RuCl6 3- couple. Additionally, an electrode is modified by an ADH/NADH oxida se bilayer, enabling ethanol determination (20-200 mu M) at 0 mV via t he oxidation of ruthenium red.