Simultaneous determination of hypoxanthine, inosine and inosine 5′-monophosphate with serially connected three enzyme reactors

A biosensor system for simultaneous determination of hypoxanthine (H-x), inosine (HxR) and inosine 5'-monophosphate (TMP) was developed to measure fish freshness indices such as K-i and H value in a convenient way. The biosensor which is composed of three enzyme reactors packed with enzyme-bonded chitosan porous beads (Chitopearl) was operated in a flow injection analysis mode with a serial circuit system. The optimum combination of column length for the H-x-HxR-IMP reactor which permitted a complete reaction between substrate and immobilized enzyme at the respective enzyme reactor was 15, 12 and 7 cm, respectively. The concentrations of H-x, HxR and IMP were determined simultaneously based on the sensor response from each enzyme reactor. The accuracy of this system was proved by a model study, and the K-i- and H-values for various fish samples were determined. The analytical results by the biosensor system did correlate well with those obtained by liquid chromatography. (C) 1999 Elsevier Science B.V. All rights reserved.