Direct introduction of amine groups into cellulosic paper for covalent immobilization of tyrosinase: support characterization and enzyme properties

Abstract

Tyrosinase is used to eliminate phenolic compounds from wastewater. Therefore, its immobilization is important to enhance catalytic efficiency. Papery materials are of particular interest for use as support for enzyme immobilization since the porous microstructure of fiber networks in papers can provide a suitable reaction environment, especially in flow-type catalytic reactions. However, immobilization of protein onto papery structure needs chemical modifications in severe conditions. To overcome this challenge, a cellulosic paper was directly amine-functionalized in moderate conditions and used for tyrosinase immobilization. The support was pretreated with HCl (0.5 N) solution and then sequentially immersed in ethylenediamine (EDA), glutaraldehyde solution (2% v/v) and the crude enzyme. In comparison with the untreated one, the immobilized enzyme on the EDA-treated support offered a 3.7-fold increase in activity. The FTIR spectra as well as EDX analysis proved the presence of amine groups in the cellulosic paper and also covalent immobilization of tyrosinase on the modified support. When considering the effect of pH on the activity at 25 °C, a maximum relative activity of 134% at pH 6 was revealed. Similarly, evaluating the effect of temperature on the activity at pH 7 displayed a maximum relative activity of 152% at 35 °C. The immobilized enzyme was suitable for use for more than four cycles to degrade a phenolic compound at severe pH and temperature conditions. Additionally, the immobilized enzyme was active after treatment of the surface at different pHs and temperatures for 105 min. The chemically modified cellulosic paper can be used as a support for enzyme immobilization.