Studies on the application of a newly synthesized polymer for trypsin purification

A water soluble ligand bound polymer has been synthesized for the purpose of purification and stabilization of trypsin, an easily autodigestible enzyme. The affinity polymer was formed by copolymerizing N-acryloyl-m-aminobenzamidine and acrylamide in the absence of oxygen. The binding efficiency of trypsin to this polymer was dependent upon the ratio of acrylamide to the monomer N-acryloyl-m-aminobenzamidine used in the polymer synthesis. The amount of acrylamide present in the polymer solution also greatly affected the trypsin binding efficiency. The total binding capacity of trypsin molecules to ligand molecules approached the theoretical value of 120 : 1 which was considerably higher than that of insoluble gel matrices. Bound trypsin could be easily eluted by the trypsin inhibitor arginine in comparison to acid, salt and benzamidine. At low temperatures, the polymer solution was very stable and retained its high capacity for trypsin binding over a long period of time. The polymer could also be reused with trypsin binding close to 90% for three consecutive runs. This study provides a sufficient background for the development of a continuous and one-step process for trypsin purification.