Benchmarking Polymeric Cryogels for Immobilized Metal Affinity Chromatography

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Hacettepe Journal of Biology and Chemistry, vol.51, no.1, pp.125-132, 2023 (Peer-Reviewed Journal) identifier


Cryogels are polymers prepared in frozen milieu, and garnered significant attention in the field as new separation mat - rices. They have denoted significant benefits including supermacroporosity, short diffusion path, low pressure, and resistance for both adsorption and elution. Macro- and connected pores give polymeric cryogels a unique spongy structure. Immobilized metal affinity chromatography (IMAC) is a standard analytical separation method for the purification of bio - molecules. Several transition ions generate stable complexes with electron-rich compounds. IMAC sorbent is obtained by complexing first-order transition metal ions over chelating agents. On the other hand, lysozyme is an enzyme found in vari - ous vertebrate cells and secretions. Common applications include its use as a cell disrupting agent, as an anti-bacterial agent, as a meal additive, and as a medicine against infections and ulcers. In this study, cryogel-based polymeric material was prepared by free-radical polymerization method with hydroxyethyl methacrylate/glycidyl methacrylate monomer pair that were covalently interacted with iminodiacetic acid metal chelating agent. The regions showing affinity for lysozyme enzyme were formed by binding with Ni(II) metal ions. The polymeric cryogel was first characterized using Fourier transform inf - rared spectrophotometer, scanning electron microscopy, thermal gravimetric analysis, X-ray photoelectron spectroscopy and swelling degree test. Then, the effects of pH, concentration, temperature, salt concentration and flow rate on enzyme adsorption capacity were evaluated, and optimum conditions were found. According to the optimization experiments, the maximum adsorption capacity of polymeric cryogel was reported as 11.82 mg/g at pH 7.4 and 25°C with a 0.5 mL/min flow rate and without ionic strength.