2 FIBEROPTIC SENSORS WITH CONFINED ENZYMES AND COENZYMES - DEVELOPMENT AND APPLICATION

A fibre-optic biosensor based on the fluorimetric detection of NAD(P)H consumed or generated during enzymatic reactions is described. The design combines a miniaturized fluorimeter with a low-volume membrane reactor and utilizes flow-injection analysis principles. Two types of membrane reactor were developed and tested. In the first version, enzymes and a macromolecular nicotinamide adenine dinucleotide coenzyme derivative of polyethyleneglycol, PEG-N6-(2-aminoethyl)-NAD(P)(H), are retained in the direct vicinity of the sensor tip. In the second version, a solid-supported liquid membrane was used to retain native coenzyme, while substrates and products were transported via selective carriers through an organic membrane barrier. The automated determination of phenylpyruvate, pyruvate, lactate, formate, ethanol and mannitol concentrations in model media was possible in the range of 50-1500 mu-mol l-1. The usefulness of the biosensor concept reported for real process monitoring in biotechnology or for medical purposes was demonstrated on different examples including the determination of ethanol for Saccharomyces cerevisiae cultivation.