Immobilization of enzymes in mesostructured materials via the nonsurfactant-templated sol-gel chemistry

We have synthesized and characterized a variety of mesoporous enzymecontaining sol-gel materials that combine the functionality of biomolecules with stability of inorganic oxides via the nonsurfactant template pathway. The enzyme molecules become immobilized in situ in mesostructured composites during the sol-gel reactions of silicon alkoxides in the presence of the nonsurfactant pore forming agent or template, such as D-glucose, D-fructose, sucrose or glycerol. Removing the nonsurfactant templates from dried as-synthesized organic-inorganic nanocomposites by water extraction yields mesoporous biogels with high remaining enzymatic activities. The catalytic activity of encapsulated enzymes, such as alkaline phosphatase, acid phosphatase or horseradish peroxidase, has been assayed and correlated with the microstructures of the host silica or organically modified silica materials derived from tetramethyl orthosilicate, tetraethyl orthosilicate or phenyltrimethoxysilane under varied synthesis conditions.; Nitrogen adsorption and transmission electronic microscopy characterizations show that the sol-gel matrices obtained by removing the templates possess a three-dimensional network with interconnected mesopores. The pore structure parameters of the sol-gel matrices and the apparent catalytic activity and stability of the entrapped enzymes are closely related to and fine-tunable by the content of templates in the synthesis. The enzymes encapsulated in the nonsurfactant-templated mesoporous sol-gel materials exhibit remarkably higher apparent catalytic activity, from a few-fold to three orders of magnitude greater, than those in the nontemplated conventional microporous hosts synthesized in the absence of the templates under otherwise identical conditions. The activity improvement of enzymes in mesoporous materials over that in microporous sol-gels is believed to arise from relatively larger pore sizes, resulting in less internal diffusion resistance and steric hindrance and greater accessibility of the entrapped enzymes to the substrates. Because of confinement in the host cages, the thermal and operational stabilities of enzymes are greatly enhanced. We also have explored the use of nonionic poly(ethylene oxide) copolymer surfactants as templates for in situ immobilization of enzymes in mesostructured sol-gel materials. This study demonstrates that the novel sol-gel immobilization methods are versatile in terms of enzymes, available templates and matrix chemical compositions, leaving much room for further modifications and optimizations. The optically transparent, biologically doped sol-gel materials have potential applications in biocatalysis, biosensor devices, etc.