Abstract

In this study combined cross-linked aggregates of catalase from bovine liver and glucose-oxidase from Aspergillus niger were prepared, and the effects of the precipitant and crosslinking agents, as well as the use of bovine serum albumin (BSA) as a feeder protein, on enzyme immobilization yield and thermal stability of both enzymes, were evaluated. Combi- crosslinking of enzyme aggregates (CLEAs) prepared using dimethoxyethane as precipitant, 25 mM glutaraldehyde and BSA/enzymes mass ratio of 5.45 (w/w), exhibited the highest enzyme activities and stabilities at 40 °C, pH 6.0, and 250 rpm for 5 h. The stability of both immobilized enzymes was fairly similar, eliminating one of the problems of enzyme coimmobilization. Combi-CLEAs were used in gluconic acid (GA) production in a bubble column reactor operated at 40 °C, pH 6.0 and 10 vvm of aeration, using 26 g L−1 glucose as the substrate. Results showed conversion of around 96% and a reaction course very similar to the same process using free enzymes. The operational half-life was 34 h, determined from kinetic profiles and the first order inactivation model. Combi-CLEAs of glucose-oxidase and catalase were shown to be a robust biocatalyst for applications in the production of gluconic acid from glucose.

Highlights

  • Gluconic acid (GA) is a multifunctional carboxylic acid with a low corrosive capacity which presents good complexation with metal ions

  • Combi-crosslinking of enzyme aggregates (CLEAs) evaluation was based on the expressed activity of the individual activities of the glucose-oxidase (GOD) and catalase (CAT) enzymes

  • The aggregates were centrifuged, the supernatants were removed and the precipitates were re-dissolved in 100 Michaelis-Menten constant (mM) of sodium phosphate buffer at pH

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Summary

Introduction

Gluconic acid (GA) is a multifunctional carboxylic acid with a low corrosive capacity which presents good complexation with metal ions. This allows GA and its salts (calcium, sodium or potassium gluconates) to be widely used in the food, pharmaceutical and textile industries [1,2,3]. Glucose oxidase (GOD; β-d-glucose: oxygen-1-oxidoreductase, EC 1.1.3.4) is able to catalyze GA production [7,8,9,10] through a Bi-Bi Ping-Pong kinetic bisubstrate mechanism [11]. The decomposition of hydrogen peroxide by enzymes is quite convenient, avoiding any side-reaction [12]. The decomposition of hydrogen peroxide to water and oxygen can be catalyzed by catalases (CAT; E.C.1.11.1.6) [4,7,13,14]

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