Simultaneous use of dehydrogenases and hexacyanoferrate(III) ion in electrochemical biosensors for l-lactate, d-lactate and l-glutamate ions

Abstract

The l-lactate, d-lactate and l-glutamate selective amperometric electrochemical biosensors presented were designed so that the last electron-transfer step is hexacyanoferrate(II) oxidation on a platinum electrode. A single enzyme sensor is described for l-lactate assay, where a lactate dehydrogenase extracted from yeast, immobilized on a membrane, will accept potassium hexacyanoferrate(III) as an electronic relay. It is possible to determine l- and d-lactate using bienzymatic sensors with NAD +-dependent dehydrogenases immobilized or in solution. In such a case, a second enzymatic reaction [a diaphorase-catalysed NADH oxidation by hexacyanoferrate(III)] enabled the detection limit to be lowered. For the l-glutamate-specific sensor, the two preceding enzymes were associated with a third one that catalyses a substrate product transformation, making it possible to exploit the enzyme amplification phenomenon. In each instance, the required presence of hexacyanoferrate(III) in the samples to be assayed makes it possible to suggest a simple apparatus with two slightly polarized electrodes. The advantages of enzyme fixation in increasing sensor stability and lowering the detection limit are also highlighted.