Oxidation of lactose to lactobionic acid by a Microdochium nivale carbohydrate oxidase: Kinetics and operational stability

Abstract

Oxidation of lactose to lactobionic acid by a Microdochium nivale carbohydrate oxidase was studied. The K_m‐value for lactose, obtained by a traditional enzymatic assay, was 0.066 mM at pH 6.4 and 38°C. The effect of oxygen on the enzymatic rate of reaction as well as the operational stability of the enzyme was studied by performing reactions at constant pH and temperature in a stirred tank reactor. Catalase was included in all reactions to avoid inhibition and deactivation of the oxidase by hydrogen peroxide. At pH 6.4 and 38°C, K_m for oxygen was 0.97 mM, while the catalytical rate constant, k_cat, was 94 s⁻¹. Furthermore, we found that the operational stability of the oxidase was dependent on the type of base used for neutralization of the acid produced. Thus, when 2 M NaOH was used for neutralization of a reaction medium containing 50 mM phosphate buffer, significant deactivation of the oxidase was observed. Also, we found that the oxidase was protected against deactivation by base at high lactose concentrations. A simple model is proposed to explain the obtained results. Biotechnol. Bioeng. 2007;97: 694–707.