IMMOBILIZED METAL AFFINITY CHROMATOGRAPHIC MEMBRANE FOR TRYPSIN SEPARATION

Abstract

As a new technology, Immobilized Metal Affinity Chromatographic Membrane (IMAC) has proven its efficiency for protein purification. It is also a separation technique that use covalently bound chelating compounds on the membrane supports to immobilize metal ions, which serve as affinity ligands for various proteins. This study aims to prepare and characterize highly specific IMAC for trypsin separation. Chitosan and polyethylene glycol were used as modification solutions to impart the membrane porosity and flux recovery ratio (FRR) of IMAC matrix. The modified PSf with chitosan improved its FRR up to 82.11% which indicated that PSf/Chitosan was a suitable matrix for the affinity membrane. Glutaraldehyde and Cu served as crosslinker agents and affinity ligands respectively. Maximum immobilization capacity of Cu occurred at 120 ppm within 60 minutes’ incubation time. The optimum capacity of trypsin adsorption (12.67 mg/cm) onto IMAC membrane occurred when 0.3 M ionic strength of trypsin solution was used. Desorption of the enzyme by displacing salt of potassium chloride showed the highest trypsin recovery at 72.3%.