Enzyme Immobilization in Polymerized Ionic Liquids-based Hydrogels for Active and Reusable Biocatalysts

Abstract

Enzyme immobilization in polymerized ionic liquids (PILs)promises to be a versatile tool for simple recovery and reuse of catalysts. In this study, a raw extract of CalB was encapsulated in poly (VEImBr) and assessed with respect to solvent, temperature, amount of enzyme, leaching behavior, and reusability on the example of the kinetic resolution of rac-1-phenylethanol with vinyl acetate. This immobilization method increased the enzyme activity of the CalB raw extract in comparison to the non-immobilized enzyme. The desired product was synthesized with high enantiomeric excess (ee) and no leaching of active enzyme was observed in the experiments. The immobilization method was compared to Novozyme 435 and Lipozyme RM IM, as commercially available immobilisates. Nonpolar solvents, including n-heptane and n-dodecane, proved to be the best reaction solvents, showing nearly full conversion and high catalytic activities. The encapsulated lipase was easily recovered from the reaction mixture and reused for ten cycles. Key words immobilization, enzyme, lipase, ionic liquid, recyclingThe majority of biocatalytic industrial processes operate with heterogeneous catalysts, using immobilized enzymes that are easily removed from the process stream. 1 Immobilization is regarded as the key enabling technology for enhancing handling and stability of the enzymes. Other advantages include an easier reactor operation, simple product separation, application in continuous processes, and a wider reactor choice. In terms of 'green' and 'sustainable' chemistry, complete recovery of the enzyme and a pure product uncontaminated with protein reduce possible allergenicity. [2][3][4] These reasons account for the increasing importance of immobilization, for which several methods are already known, including immobilizing biocatalysts on solid supports or in synthetic or natural polymers by covalent attachment, adsorption, and physical entrapment. 5,6 One carrier-free immobilization technique involves crosslinking of enzymes, which, with the help of their superficial amino groups, are connected to each other by a bifunctional reagent, commonly glutaraldehyde. 6 Generally, two procedures for this process are known. First, to form insoluble crosslinked enzyme crystals (CLECs), the enzyme must crystallize from an aqueous buffer before the crosslinker is added. 4 The second method is much easier because it replaces crystallization with precipitation. Salts (e.g., ammonium sulfate) or water-miscible organic solvents are added to an aqueous enzyme solution. After the crosslinker is added, the crosslinked enzyme aggregates (CLEAs) precipitate. Thereafter, no purification is necessary, and the enzyme can be employed immediately. 7 However, immobilization via crosslinking has several drawbacks, including low activity, poor reproducibility, and low mechanical stability, 8 the latter of which can be improved by entrapping enzymes into a solid support. Apart from the traditional carriers, which include polysaccharides, synthetic resins, and zeol...

Poly(ionic liquid) composites

“…Moreover, the enzyme catalyst was easily recyclable due to the high mechanical and chemical stability of the PILs. 166 In addition, PIL-biosubstance composites can be used in other biofields. DNA-polycation complexes are known as one of the most promising systems in gene therapy due to their salient features, such as aqueous solubility, charge neutralization, pH sensitivity, and tunable physicochemical properties.…”

Advances in Recombinant Lipases: Production, Engineering, Immobilization and Application in the Pharmaceutical Industry

“…In physical adsorption, enzyme leaching is a critical problem that limits the use of the immobilized enzyme in different reaction conditions [138]. To solve this problem, entrapment is employed to restrict the enzyme in the polymer frameworks [139]. In the entrapment method, the enzyme is locked inside the lattice of a polymer matrix or membrane so that it is retained while allowing the penetration of the substrate.…”

Recent Developments in Carriers and Non-Aqueous Solvents for Enzyme Immobilization

“…CalB was encapsulated in poly(VEIm-Br) and assessed in different solvents. The results suggested that activity was enhanced and there was no leaching of enzyme activity [90]. Enzymatic biodiesel production has been hindered by reusability, but the enzyme-IL system helps reuse of enzymes over more cycles [91].…”