Supermacroporous hydrophobic affinity cryogels for protein chromatography |
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| Affiliation: | 1. Mersin University, Faculty of Pharmacy, Department of Toxicology, Mersin, Turkey;2. Near East University, Faculty of Pharmacy, Department of Toxicology, Nicosia, Cyprus;3. Hacettepe University, Faculty of Science, Department of Chemistry, Ankara, Turkey;4. Mersin University, Vocational School of Technical Sciences, Mersin, Turkey;1. Hacettepe University, Department of Chemistry, Ankara, Turkey;2. Sinop University, Department of Bioengineering, Sinop, Turkey;3. Anadolu University, Department of Chemistry, Eskişehir, Turkey;1. Hacettepe University, Ankara, Turkey;2. Aksaray University, Aksaray, Turkey;1. Department of Analytical Chemistry, Faculty of Chemistry, University College of Science, University of Tehran, P.O. Box 14155-6455, Tehran, Iran;2. Department of Applied Chemistry, Faculty of Science, University of Mohaghegh Ardabili, Ardabil, Iran |
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| Abstract: | N-Methacryloyl-l-tryptophan (MATrp) containing poly(2-hydroxyethyl methacrylate) based supermacroporous cryogel [PHEMATrp] was prepared for lysozyme purification form chicken egg white. MATrp was synthesized by reacting methacryloyl chloride with l-tryptophan methyl ester and provided hydrophobic functionality to the cryogel. PHEMATrp cryogel with 60–100 μm pore size was obtained by free radical polymerization of HEMA and MATrp having a specific surface area of 50 m2/g. PHEMATrp cryogel was characterized by swelling studies, FTIR and SEM. The equilibrium swelling ratios of the cryogels were 7.18 g H2O/g for PHEMA and 6.99 g H2O/g for PHEMATrp. Lysozyme adsorption experiments were investigated under different conditions in continuous system (i.e., medium pH, flow-rate, protein concentration, temperature, salt type). Lysozyme adsorption capacity of PHEMA and PHEMATrp cryogels from aqueous solutions was estimated as 2.9 and 46.8 mg/g (0.49 and 7.85 mg/mL), respectively. Lysozyme molecules were desorbed with 0.5 M ethylene glycol solution with 91% recovery. It was observed that PHEMATrp cryogel can be used without significant decrease in lysozyme adsorption capacity after five adsorption–desorption cycles. PHEMATrp cryogel was used for the purification of lysozyme from chicken egg white. Purity of lysozyme was estimated by SDS-PAGE. Possible denaturation of purified lysozyme was checked with fluorimetric measurements. Specific activity of the purified lysozyme was found as 43,140 U/mg using Micrococcus lysodeikticus as substrate. |
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