Emulsion-Templated Porous Polymers as Support Materials for Covalent Enzyme Immobilization

Abstract

It has been observed that poly(High Internal Phase Emulsion) (polyHIPE) materials can be used as a biocatalysts, via the covalent immobilization of Candida Antarctica Lipase B (CAL-B). Recently, it has been shown that polyHIPEs can be prepared with epoxy functionality, which show potential for the covalent immobilization of enzymes.
The aims of our work were, firstly, to produce an open-void glycidyl methacrylate (GMA)-based polyHIPE material. Secondly, these materials were then to be developed for use within a continuous flow set-up. Thirdly, the post-polymerisation of these materials was to be investigated. Finally, these materials were to be used as a support for the covalent immobilization of enzymes.
Highly porous, open-void GMA-based polyHIPE materials were accomplished via the photo- initiation, rather than thermal initiation of the continuous phase of the emulsion. The rapid cure of the emulsion effectively ‘locks’ the emulsion morphology, prior to emulsion destabilisation, that is more prominent in the slower thermally initiated HIPEs. Photopolymerised GMA-based polyHIPE materials were further developed for use within a continuous flow-set up. GMA-based polyHIPE materials were functionalized post-polymerisation with tris(2-aminoethyl)amine, morpholine and O,O’-bis(3-aminopropyl)polyethylene glycol. The functionalization of these GMA-based materials was observed via a number of analysis techniques, such as FT-IR spectroscopy, XPS spectroscopy, elemental analysis, Fmoc number determination, 1H HR-MAS NMR spectroscopy, and the covalent attachment of ninhydrin and FITC. Elemental analysis of the morpholine and tris(2-aminoethyl)amine polyHIPE showed that a near quantitative conversion, of 72 and 82 % respectively, was accomplished via the reaction being conducted at reflux for 24 hours. The enzymes, Lipase from Candida Antarctica and Proteinase K from Tritirachium album were immobilized either directly onto the polyHIPE material or via a hydrophilic spacer group, O,O’-bis(3-aminopropyl)polyethylene glycol. CAL was immobilized with a loading of between 5.4 and 7.5 wt. % per g of polyHIPE material.