Short communication Immobilized glucose oxidase on different supports for biotransformation removal of glucose from oligosaccharide mixtures D. Mislovic ˇova ´ a, * , E. Micha ´lkova ´ b , A. Vikartovska ´ a a Institute of Chemistry, Slovak Academy of Sciences, Du ´bravska ´ cesta 9, SK-845 38 Bratislava, Slovakia b Faculty of Ecology and Environmental Science, Technical University Zvolen, T.G. Masaryka 24, SK-965 03, Zvolen, Slovakia Received 17 August 2006; received in revised form 27 October 2006; accepted 3 November 2006 Abstract Four immobilized forms of glucose oxidase (GOD) were used for biotransformation removal of glucose from its mixture with dextran oligosaccharides. GOD was biospecifically bound to Concanavalin A-bead cellulose (GOD-ConA-TBC) and covalently to triazine-bead cellulose (GOD-TBC). Eupergit C and Eupergit CM were used for preparation of other two forms of immobilized GOD: GOD-EupC and GOD-EupCM. GOD-ConA-TBC and GOD-EupC exhibited the best operational and storage stabilities. pH and temperature optima of these two immobilized enzyme forms were broadened and shifted to higher values (pH 7 and 35 8C) in comparison with those of free GOD. The decrease of V max values after immobilization was observed, from 256.8  7.0 mmol min 1 mg GOD 1 for free enzyme to 63.8  4.2 mmol min 1 mg GOD 1 for GOD- ConA-TBC and 45  2.7 mmol min 1 mg GOD 1 for GOD-EupC, respectively. Depending on the immobilization mode, the immobilized GODs were able to decrease the glucose content in solution to 3.8–15.6% of its initial amount The best glucose conversion, was achieved by an action of GOD-EupCM on a mixture of 100 g dextran with 9 g of glucose (i.e. 98.7% removal of glucose). # 2006 Elsevier Ltd. All rights reserved. Keywords: Glucose oxidase; Immobilization; Stability; Glucose biotransformation; Bead cellulose; Eupergit 1. Introduction Glucose oxidase (GOD) is a flavoprotein which catalyses the oxidation of b-D-glucose using molecular oxygen to hydrogen peroxide and d-gluconolactone, which spontaneously hydro- lyses non-enzymically to gluconic acid [1]. This enzyme is of considerable commercial importance. GOD is widely used in clinical and chemical analyses as well as in food industry [2–5] for the removal of glucose [6] or oxygen from food products and for production of gluconic acid [7,8]. In order to enhance the enzyme properties such as reusability, operational stability, recovery and shelf life, GOD has been immobilized on different supports involving various immobilization methods [9–11]. The choice of the support as well as of the technique depends on the nature of the enzyme, nature of the substrate and the support application. GOD has been immobilized on various particular supports with the purpose of its economic reuse, for the preparation of gluconic acid [12], for removal of oxygen from a beer [13], and for removal of glucose from eggs [14–16]. GOD was also used for the removal of glucose from pentosan of wheat flour [17]. Determination of pentosan content of wheat flour is routinely based on hydrolysis of the polysaccharides present in the sample followed by determination of released pentoses by HPLC. The interference of relatively high concentration of starch-derived glucose with pentosan monosaccharides was eliminated by enzymatic conversion of the glucose into gluconic acid catalysed with GOD [17]. The aim of this study was to obtain an immobilized GOD suitable for the removal of glucose from pharmaceutical and food products. The presence of glucose in these products causes their browning during dehydration and long-term storage, which is mainly due to the Maillard reaction. This is a major obstacle in the manufacture of dehydrated egg powder [15], and also is a serious problem at hydrolysis of high molecular weight dextrans to oligosaccharides utilized for a preparation of pharmaceutical products. The resulting colored compounds are not compatible with pharmaceutical and food rules for formulation acceptibility. In the present work we have prepared and characterized four samples of Aspergillus niger GOD immobilized on bead cellulose and on Eupergit, either by covalent or biospecific www.elsevier.com/locate/procbio Process Biochemistry 42 (2007) 704–709 * Corresponding author. Tel.: +421 2 59410263; fax: +421 2 59410222. E-mail addresses: chemmisl@savba.sk, danica.mislovicova@savba.sk (D. Mislovic ˇova ´). 1359-5113/$ – see front matter # 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.procbio.2006.11.001