Development and application of immobilized lipase assembled with ionic liquid on poly(acrylic acid) beads

다국어 초록 (Multilingual Abstract)

Immobilized enzymes have attracted much attention due to the characteristics of operational stability, enhanced activity, and reusability as well as simple separation. Most of commercial lipases are immobilized form and are used to convert fatty acids to fatty acid alkyl esters (FEs) via an environment-friendly process. Immobilization of enzymes on resin beads has been utilized as a common tool in biological studies as well as bio-industrial field.
However, factors controlling the stability of enzyme on resin beads have not been fully explored. So, esterification reactions are designed to evaluate how the stability of Candida antarctica lipase B (CALB) depends on the hydrophilic/hydrophobic properties with or without ionic liquids (ILs) on poly(acrylic acid) (PA) beads, because the ILs can provide a tunable microenvironment to the lipase. The catalytic activities of the immobilized lipases were compared under several factors such as bead properties, immobilization methods, loading levels of ILs, chain lengths of alkyl group on ILs, and solvents.
From these results, we found that the IL-grafted PA beads which contained more hydrophilic properties (octyl group) showed better catalytic performance than hydrophobic ones (dodecyl group). The use of tert-butanol as a solvent was a pivotal factor to maintain the activity of the immobilized lipase. Surprisingly, the single type immobilized lipase, PA-C8Im-Lipase, (C8Im loading level, 1.2 mmol/g) showed the best catalytic performance (95% yield) at 50 oC even though the network type immobilized lipase was expected to show the best performance. The single type immobilized lipase can be reused with slightly loss of activity.
In conclusion, octyl group provided positive influence to the lipase activity on IL-grafted PA beads without blocking the hydrophobic active site of lipase.

목차 (Table of Contents)

• Contents
• Abstract ...............................................i
• List of Abbreviations .................................vi
• List of Figures .....................................viii
• List of Schemes .......................................ix
• List of Tables .........................................x
• Chapter 1. Introduction ................................1
• 1.1 General Introduction of Lipase .....................1
• 1.1.1 Progress of Enzyme Catalysts .....................1
• 1.1.2 Properties of Free Lipase ........................2
• 1.1.3 Industrially Valuable Ester Compounds.............3
• 1.2 Enzyme Immobilization...............................4
• 1.2.1 Strategies for Enzyme Immobilization..............4
• 1.2.2 Role of Polymer Supports in Lipases Immobilization6
• 1.2.3 Polymer Grafting for Enzyme Immobilization Assembled with Ionic Liquids ...........................7
• 1.3 Research objectives.................................8
• Chapter 2. Experiments .................................9
• 2.1 General ............................................9
• 2.1.1 Materials ........................................9
• 2.1.2 Instruments .....................................11
• 2.2 Modification of Poly(acrylic acid) Beads ..........12
• 2.2.1 Preparation of Ionic Liquid-grafted Poly(acrylic acid) Beads ...........................................12
• 2.2.2 Immobilization of Candida Antarctica Lipase B on Ionic Liquid-grafted Poly(acrylic acid) Beads .........16
• 2.3 Enzyme Activity Test...............................18
• 2.3.1 Hydrolysis of p-Nitrophenyl Butyrate.............18
• 2.3.2 Esterification of Fatty Acids ...................19
• Chapter 3. Results and Discussion .....................20
• 3.1 Preparation and Characterization of Immobilized Lipase.................................................20
• 3.2 Catalytic Activities of the IL-grafted PA Beads....25
• 3.2.1 Hydrolysis of p-Nitrophenyl Butyrate ............25
• 3.2.2 Esterification Activity of Immobilized Lipases ..27
• 3.2.3 Reusability of Immobilized Lipase ...............33
• Conclusion ............................................34
• References ............................................35
• Abstract in Korean ....................................38
• Appendix ..............................................40