Trypsin activity in imidazolium based ionic liquids: evaluation of free and immobilized enzyme

详细信息	查看全文 | 推荐本文 |

• 作者：Paula C.A.G. Pinto ; Susana P.F. Costa ; Andreia D.F. Costa ; Marieta L. Passos ; José ; L.F.C. Lima ; M. Lú ; cia M.F.S. Saraiva ; ^{lsaraiva@ff.up.pt}
• 关键词：Trypsin ; Enzymatic peptide synthesis ; Immobilization ; Ionic liquids
• 刊名：Journal of Molecular Liquids
• 出版年：2012
• 期刊代码：61_01677322
• 类别：ch
• 出版时间：July, 2012
• 卷：171
• 期：Complete
• 页码：16-22
• 文件大小：613 K

摘要

This work aimed the evaluation of ionic liquids as an alternative to conventional organic solvents in the enzymatic synthesis of peptides. For this, the effect of imidazolium ionic liquids on the activity and stability of immobilized trypsin was studied. Enzyme activity was studied by means of a model reaction, the hydrolysis of N-伪-benzoyl-DL-arginine-p-nitroanilide (BAPNA), optimized by means of batch and flow methodologies. The activity of the enzyme was studied in strictly aqueous media and in the presence of increasing concentrations of emim [BF₄], emim [Ms], emim [OTf] and bmim [BF₄]. It was observed that bmim [BF₄] seemed to be the less inhibiting IL, leading to a maximum enzyme inhibition of 69.3%when presented in a concentration of 85%, in the assays with free enzyme. The results were similar on both batch and flow assays. Moreover, an enhanced catalytic efficiency of immobilized trypsin in IL media was observed when compared with that of free enzyme. These observations evidence the compatibility between ILs and the utilization of enzymes immobilized in glass beads. The results gathered from this work can be of most importance for the large-scale industrial implementation of the trypsin catalysed peptide synthesis since enzyme stabilization through immobilization showed to be adequate to high yield synthetic biotransformations in biphasic media (IL-water). There were no evidences of enzyme leakage from the glass beads during its continued use in ionic liquid media.

The SIA methodologies showed to be interesting tools for the evaluation of trypsin's activity in ionic liquids as they proved to be robust and exhibited good repeatability in all the assay conditions leading also to a reduction of the consumption of solvents as well as of effluent production.