Cross-Linked Bentonite-Acrylamide-histidine-Based Metal-Chelate Affinity Microcomposites for Lysozyme Separation From Egg White

Abstract

Among different adsorbents, a new support for immobilized metal affinity chromatography was used for lysozyme separation from egg white. For this purpose, acrylamide, bisacrylamide, bentonite, and L-histidine in the presence of an initiator (azobisisobutyronitrile, AIBN) were mixed and bentonite-acrylamide-histidine microcomposites were prepared by bulk polymerization. Characterization studies were performed by SEM, FT-IR, XRD, and BET. Specific surface area of the bentonite and BABH microcomposite was found to be 33.4 and 1.42 m2/g, respectively. Cu2+ ions (20 mu mol/g) were chelated on these bentonite-acrylamide-histidine (50 mu mol/g L-histidine incorporation) microcomposites. The lysozyme adsorption capacity of the Cu2+ ions chelated microcomposites increased the lysozyme adsorption up to 100 mg/g while the lysozyme adsorption capacity of the bentonite-acrylamide-histidine was 24 mg/g. More than 89% of the adsorbed lysozyme was desorbed in 1 h in the desorption medium containing 1.0 M NaCl. Bentonite-acrylamide-histidine-Cu(II) was used for the purification of lysozyme from chicken egg white. The purity of lysozyme was estimated by SDS-PAGE. Specific activity of the purified lysozyme was found as 50,000 U/mg using Micrococcus lysodeikticus as substrate. The new metal-chelate affinity microcomposites were suitable for repeated use for five cycles without a noticeable loss of capacity.