ENZYME STABILIZATION USING PECTIN AS A NOVEL ENTRAPMENT MATRIX IN BIOSENSORS

Abstract

ABSTRACT The development of biosensors is often based on the use of enzymes that commonly display an inherent lack of suitable stability. We report the use of pectin as a novel material for enzyme entrapment and stabilisation. This work relates to the development of an electrochemical enzyme-sensor array for the monitoring of fruit quality. The enzymes investigated are glucose oxidase, mutarotase, invertase and ascorbate oxidase. Sensors were constructed by screen-printing, with a metallised (rhodium) carbon working electrode, silver/silver chloride printed reference electrode and carbon counter electrode. Enzymes were dispensed in a pectin-based gel over the working electrode and retained by a cellulose acetate membrane. Enzyme stability as a function of dry storage time was assessed by the attained steady-state response of the single-use sensors upon removal from storage. Except for ascorbate oxidase, all enzyme sensors functioned via the detection of enzymatically generated hydrogen peroxide. Ascorbate oxidase stability was assessed by the efficacy of enzymatic elimination of ascorbic acid. All the measurements were carried out at +350 mV vs. Ag/AgCl. Pectin was compared to the established polymer hydroxyethyl cellulose (HEC) as an immobilisation matrix. The use of a pectin immobilisation matrix demonstrated either similar stability (glucose oxidase) or increased stability (mutarotase, invertase, ascorbate oxidase) with respect to HEC during a twenty-five week storage test period. Pectin immobilised enzymes demonstrated a greater stabilising effect and reduced response variability while larger signal currents were often observed for HEC immobilised enzymes.