Purification of lysozyme from egg white by Reactive Blue 4 and Reactive Red 120 dye-ligands immobilised composite membranes

Abstract

A composite membrane was synthesized from 2-hydroxyethylmethacryl ate (HEMA) and chitosan (pHEMA/chitosan) via UV initiated photo-polymerisation. Reactive Blue 4 (Blue-4) and Reactive Red 120 (Red-120) were immobilised onto pHEMA/chitosan membranes. In the first part of this study, the binding characteristics of lysozyme to different dye-ligand immobilized membranes have been studied from aqueous solution using the plain membrane as a control system. The polarity of the investigated membranes was determined by contact angle measurement. The adsorption capacities of both dye-ligand immobilised membranes increased with increasing temperature but decreased with increasing NaCl concentration. The adsorption isotherm fitted both the Langmuir and the Freundlich models. A theoretical analysis has been conducted to estimate the thermodynamic contributions (changes in enthalpy, entropy and Gibbs free energy) for the adsorption of lysozyme to different dye-ligand immobilised composite membranes. In the second part, their purification efficacy of lysozyme from egg white was investigated. The purity of the eluted lysozyme was analysed by HPLC. The purity of lysozyme extracted from egg white was 84% with Red-120 immobilised membrane. This was 21% for Blue-4 immobilised membrane. The recovery yields were 72% and 16% for Red-120 and Blue-4 immobilised membranes, respectively. The Red 120 immobilised membrane provided an efficient method to purify lysozyme from egg white. showing high adsorption capacity and high selectivity for the lysozyme. On the other hand, the Blue-4 immobilised composite membrane had a lower adsorption capacity and selectivity than that of the Red-120 immobilised one. Purification was monitored by determination of lysozyme activity using Micrococcus lysodeiktictis as substrate. The dye-ligand immobilised composite membranes are stable when subjected to sanitization with sodium hydroxide after repeated separation-elution cycles. (C) 2004 Elsevier Ltd. All rights reserved.