Journal of Chromatography B 2010-05-01

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

Libo Zhou, Junfen Wan, Xuejun Cao, Libo Zhou, Junfen Wan, Xuejun Cao, Libo Zhou, Junfen Wan, Xuejun Cao

Index: J. Chromatogr. B. Analyt. Technol. Biomed. Life Sci. 878(15-16) , 1025-30, (2010)

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Abstract

In this study, thermo-sensitive N-alkyl substituted polyacrylamide polymer P(NNB) 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 degrees C. The recovery of the thermo-sensitive polymer was over 98%. Butanol as a hydrophobic ligand was covalently attached onto polymer P(NNB) and butyl ligand density was 80 micromol g(-1) polymer. The affinity polymer was used for purification of lipase from crude material. Optimized condition was pH 7.0, 35 degrees 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 (Q(m)) of the affinity polymer for lipase was 24.8 mg g(-1)polymer. 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 14IU 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.2010 Elsevier B.V. All rights reserved.