A REAGENTLESS AMPEROMETRIC ELECTRODE BASED ON CARBON-PASTE, CHEMICALLY-MODIFIED WITH D-LACTATE DEHYDROGENASE, NAD(+), AND MEDIATOR CONTAINING POLYMER FOR D-LACTIC ACID ANALYSIS .1. CONSTRUCTION, COMPOSITION, AND CHARACTERIZATION

A reagentless carbon paste electrode was designed for D-lactic acid analysis in a flow injection system for the monitoring of the production of D-lactate in a batch fermentation. D-lactate dehydrogenase, nicotinamide adenine dinucleotide (NAD(+)), a synthetic redox polymer containing covalently attached toluidine blue O as mediator, graphite powder, and paraffin oil were used for the construction of the modified carbon paste electrode. D-Lactate selectivity was indicated by insignificant responses from a variety of possible interferences including L-lactate. The electrodes gave a linear response in the range between 0.05 and 5 mM D-lactate, with a detecting limit of 30 mu M, allowing a sample throughput of 20 h(-1). Preliminary investigations were made by covering the electrode surface with electropolymerized membranes. Satisfactory stability was observed, indicated by a reproducibility of 3.3% relative standard deviation (RSD, n = 31), with a non-membrane-covered electrode for the analysis of D-lactate in fermentation broth. A long-term stability (230 broth samples) was proven, suggesting the electrodes to have a good potential for use in on-line monitoring of fermentation processes. (C) 1995 John Wiley & Sons, Inc.