Synthesis of thermo-sensitive copolymer with affinity butyl ligand and its application in lipase purification

In this study, thermo-sensitive N-alkyl substituted polyacrylamide polymer PNNB was synthesized by using N-hydroxymethyl acrylamide(NHAM), N-isopropyl acrylamide (NIPA) and butyl acrylate (BA) as monomers, and its low critical solution temperature (LCST) was controlled to be 28 °C. The recovery of the thermo-sensitive polymer was over 98%. Butanol as a hydrophobic ligand was covalently attached onto polymer PNNB and butyl ligand density was 80 μmol g−1 polymer. The affinity polymer was used for purification of lipase from crude material. Optimized condition was pH 7.0, 35 °C adsorption temperature, 120 min adsorption time and 0.5 mg ml−1 initial concentration of lipase. The adsorption isotherm accords with a typical Langmuir isotherm. The maximum adsorption capacity (Qm) of the affinity polymer for lipase was 24.8 mg g−1polymer. The affinity copolymer could be recycled by temperature-inducing precipitation and there was only about 6% loss of adsorption capacity after five recyclings. Specific activity of lipase was improved from 14 IU mg−1 to 506 IU mg−1 protein, and its recovery achieved 82%. The affinity polymer is suitable for the purification of target proteins from the crude material with large volume and dilute solution.