Preparation of insulin-imprinted poly (HEMA-MAH) cryogels

Insulin is one of the most important hormone and it has profound effects on metabolism by controlling of glucose homeostasis. When insulin production or the use of insulin ability decrease these disorders cause diabetic illness. Separation and purification of insulin or purified proteins are very important because of to obtain as a purified raw materials of diabetics drugs. Protein separation techniques gain more importance at this point. Protein purification via adsorption is of significant importance in biomedical and biotechnological purposes.In present study, affinity adsorption technique is studied for insulin adsorption. For this aim insulin-imprinted supermacroporous cryogel was prepared for the insulin adsorption. N- methacryloyl- L- histidin methylester (MAH) was choosen as the monomer. MAH was synthesized using methacryloyl chloride and L- histidinemethyl ester by Hacettepe University Faculty of Science Department of Biochemistry Prof. Dr. Adil Denizli?s group.In the first step, insulin was complexed with MAH and insulin-imprinted p(HEMA-MAH) cryogel was prepared by free radical polymerization by polyhydroxymethacrylate p(HEMA), N,N,N?,N?? tetrametiletilendiamin (TEMED) and ammmonium persulphate (APS) in an ice bath. Then, insulin was removed from cryogel by using 0.1 M glycine-HCl buffer (pH: 3.5). Cryogel was characterized by using scanning electron microscopy (SEM), Fourier transfer infrared spectroscopy and swelling test. The equilibrium swelling ratios of the cryogels were 8.56 g H2O/g polymer for p(HEMA) and 7.20 g H2O/g polymer for insulin-imprinted p(HEMA-MAH). Insulin adsorption experiments were carried out under different conditions in continuous system (i.e., flow rate, medium pH, insulin concentration, temperature and ionic strength). Insulin adsorption capacity of p(HEMA-MAH) cryogel from aqueous solution was found to be 8.13 mg/g polymer. It was observed that insulin could be repeatedly adsorbed and desorbed with p(HEMA-MAH) cryogel without any significant loss in the adsorption capacity.