Abstract: A potentiometric biosensor for the analysis of isocitrate was developed by using a CO32--selective electrode and enzyme immobilization in flow injection analysis (FIA). The biosensor consisted of a peristaltic pump, injector, enzyme reactor, reference electrode, working electrode, pH/mV meter and record. The experimental parameters that influenced the sensitivity and selectivity of the biosensors were optimized in the FIA system. A linear correlation between the potential difference and logarithmic isocitrate concentration was obtained in the range of 10^-3-10-1 M isocitrate under optimal conditions. The interference effect of major sugars and organic acids on the sensor system was less than 5%. Isocitrate concentrations of some fruit juices analyzed by the isocitrate sensor system were compared with those analyzed by gas chromatography (GC). There was no significant difference between the two analytical methods in any of the fruit juices. This suggests that the isocitrate sensor system is reliable in determining the isocitrate concentrations of foods.

Abstract: The oxidation of NADH using NADH oxidase from Bacillus licheniformis or diaphorase from Clostridium kluyveri, was found to produce superoxide anion (O-2(-)). The generated O-2(-) reduced water-soluble tetrazolium salt WST-1 to WST-1 formazan. The formation of WST-1 formazan was monitored as a change in the absorbance at 438 nm. The formation of WST-1 formazan was found to be proportional to the NADH concentration. As a result, a linear curve was obtained within the range of 0.5 muM-50 muM NADH concentration. The concentration of NADH was determined by chemiluminescence using lucigenin specific for O-2(-) instead of WST-1. Application of NADH oxidase from Bacillus licheniformis was found to be 400 times stronger chemiluminescence than that of diaphorase with a lower detection limit of 15 nM. The present method was applied to determine ethanol combined with yeast alcohol dehydrogenase. The ethanol concentrations of various kinds of alcoholic beverages obtained by the present method were compared with those obtained using F-kit method. A good linear correlation was observed between them (r=0.9997) and the slope (1.005) was very close to unity, suggesting that the present method could be applied for the determination of ethanol in practical samples. (C) 2001 Elsevier Science Ltd. All rights reserved.

Abstract: A biosensor for the determination of fructose was constructed by the immobilization of the enzyme fructose dehydrogenase (FDH) behind a thin non-conducting electropolymer film of 1,3 phenylene diamine-resorcinol. Two different types of electrochemical mediators, soluble hexacyanoferrate (III) (FeCN) and the water insoluble tetracyanoquinodimethane (TCNQ) were applied as redox mediators for the amperometric measurement of fructose. Optimization of the biosensor was done with respect to operating conditions by varying the operating parameters such as pH, temperature, mediator concentration, and enzyme loading. The optimized biosensor was found to have a minimum detection limit of 10 µM with a linear range upto 1.0 mM. The performance of the two mediators was also compared with respect to stability. A simple, lumped parameter mathematical model was proposed to describe the steady state response of the biosensor and experimentally verified with the constructed biosensors.

Abstract: A two-line flow injection analysis biosensor was developed which can simultaneously detect bacterial spoilage and the progress of aging. This FIA biosensor was composed of a putrescine oxidase immobilized electrode and a xanthine oxidase immobilized electrode as detectors. The putrescine electrode measures putrescine and cadaverine which are produced by bacteria, and the xanthine electrode measures hypoxanthine and xanthine which accumulate in meat with aging. The analytical conditions for this system were set as follows; flow rate, 1 ml/min; water bath temperature, 30°C; flow buffer, 0.1 M phosphate buffer (pH 7.0); injection volume for putrescine electrode, 200 µl; injection volume for xanthine electrode, 40 µl; and measurement cycle, 2 min. The linear relationship for standard solution was between 20 and 800 nmol/ml in the putrescine electrode and between 0.1 and 3.0 µmol/ml in the xanthine electrode. The coefficients of variation in standard solutions were 2.14% with the putrescine electrode and 2.83% with the xanthine electrode. The coefficients of variation values in the specimen solution were 3.22% and 3.76%, respectively. This two-line FIA biosensor was applied to the vacuum-packed beef stored at 0, 5 and 10°C. The progress of aging could be monitored at all temperatures, and the bacterial spoilage could be detected before the appearance of putrid odor at 5 and 10°C. However, at 0°C the putrid odor did not appear during storage, and neither putrescine nor cadaverine was detected. Thus, this FIA biosensor was confirmed to be useful for the quality control of beef aging at 5 and 10°C, but not at 0°C. A two-line flow injection analysis biosensor was developed which can simultaneously detect bacterial spoilage and the progress of aging. This FIA biosensor was composed of a putrescine oxidase immobilized electrode and a xanthine oxidase immobilized electrode as detectors. The putrescine electrode measures putrescine and cadaverine which are produced by bacteria, and the xanthine electrode measures hypoxanthine and xanthine which accumulate in meat with aging. (22 References)