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    Nanostructured modified ultrananocrystalline diamond surfaces as immobilization support for lipases

    Year: 2018

    Journal: Diam. Relat. Mat., Volume 90, NOV, page 32–39

    Authors: Merker, Daniel; Kesper, Magdalena; Kailing, Lyn Lisette; Herberg, Friedrich; Reithmaier, Johann Peter; Pavlidis, Ioannis V.; Popov, Cyril

    Organizations: University of Kassel; ZFF-PROJEKT 1963

    Keywords: Ultrananocrystalline diamond films; Surface modification and functionalization; Lipase; Covalent and non-covalent immobilization

    Ultrananocrystalline diamond (UNCD) is a promising and - so far - relatively unexploited material for biocatalytic applications. In this study, UNCD films, deposited on silicon wafers by microwave plasma-assisted chemical vapor deposition, were investigated as supports for immobilization of the enzyme lipase B from Candida antarctica (CalB). The motivation for this work was the potential of utilization of the resulting bio-materials for the fabrication of flow bioreactors, harnessing both the benefits of the UNCD and the biocatalysts. Prior to immobilization, the UNCD surfaces were structured by reactive ion etching with a randomly distributed gold nanodroplet mask that was applied on top of the UNCD films, in order to increase the area for interaction with the enzyme. Furthermore, structured UNCD films were subjected to modification by UV/ozone treatment or trifluoromethane plasma which rendered the surfaces oxygen-terminated and hydrophilic or fluorine-terminated and hydrophobic, respectively. Two established coupling strategies, based on carbodiimide and oxalyl chloride chemistry, were investigated for covalent immobilization of CaIB on structured oxygen-terminated surfaces. However, non-covalent immobilization on fluorine- and as-grown hydrogen-terminated surfaces, mediated by physical adsorption of the enzyme, appeared to be more efficient. Moreover, the data strongly suggested that the chemical nature of the termination and the hydrophobicity of the surface influence the substrate selectivity of the enzyme. Non-covalently immobilized CaIB on strongly hydrophobic fluorine-terminated surfaces showed a similar to 5.5-fold higher preference for the short-chain substrate p-nitrophenyl butyrate over the medium-chain ester p-nitrophenyl laurate, whereas on hydrophilic O-terminated surfaces this selectivity was only similar to 1.9-fold. Moderately hydrophobic H-terminated surfaces conferred a selectivity ratio of similar to 3.5, which is in between the two former values and therefore in line with the hypothesized dependence of the substrate selectivity on the hydrophobicity of the surface material. These primary results can help on the optimization of the design of immobilized biocatalysts on UNCD surfaces to produce columns for flow bioreactors. Prime novelty: Nanostructured modified ultrananocrystalline diamond (UNCD) surfaces were studied as immobilization support for lipases. The physical adsorption of lipase CaIB was more efficient on hydrophobic F- or H-terminated surfaces than the two covalent coupling approaches on hydrophilic O-terminated UNCD. Furthermore, the chemical nature of the surface termination had an impact on the substrate selectivity by catalyzed hydrolysis.
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