A MODEL SYSTEM FOR A FLUOROMETRIC BIOSENSOR USING PERMEABILIZED ZYMOMONAS-MOBILIS OR ENZYMES WITH PROTEIN CONFINED DINUCLEOTIDES

Using permeabilized Zymomonas mobilis or glucose-fructose oxidoreducta se isolated from this microorganism a model system for biosensors with a protein confined NADP(H) cofactor for the determination of glucose, fructose, gluconolactone, and sorbitol was developed. Either permeabi lized microorganisms containing the oxidoreductase or the pure enzyme were confined via membrane separation in a small measuring chamber, th at was integrated into a flow injection analysis system (FIA). The mea suring principle was the monitoring of the NAD(P)H fluorescence, excit ed at 360 nm and measured at 450 nm. NADP(H), which is confined in the protein complex, was oxidized or reduced during the enzymatic reactio ns and the changes in the fluorescence intensity were related to the s ubstrate concentration. The sensitivity of the system covered a range from 0.001 to 100 g/L of the analyte depending on substrate and operat ing conditions. The applicability of this model system for bioprocess monitoring was proved using samples from a Pseudomonas pseudoflava cul tivation.