Novel bio-conjugate materials: soybean peroxidase immobilized on bioactive glasses containing Au nanoparticles

Abstract

In the field of implantation, the delivery and/or immobilization of biomolecules developing a specific action on bone mineralization has attracted great attention in the last few years. In fact, a wide spectrum of enzymes and proteins have been grafted with different methods onto/within implanted materials. Bioactive glasses and glass-ceramics, due to their tailorable properties in terms of chemical composition, reactivity, and easiness of manufacturing, represent good scaffolds for enzyme immobilization. These biomaterials are well known for their peculiar surface reactivity promoting, when contacted with real or simulated body fluids, the formation of an hydroxy-carbonate apatite layer. The aim of the present contribution has been to immobilize, via a covalent linkage, an enzyme on the glass surface through the formation of self-assembled monolayers (SAMs), in order to obtain a stable bio-conjugate useful as a material bio-implantable into the human body. The innovation of this study resides in the use of a new method of protein immobilization on the glass surface. Unlike other works, in which a preliminary silanization process has often been used, the introduction of gold nanoparticles (AuNPs) in the glass composition allowed us to exploit the easy SAMs formation process on the AuNPs dispersed in the bioactive glass matrix and, consequently, to immobilize an enzyme (soybean peroxidase, SBP, in the present case) on the SAMs. A thorough characterization of the materials, at different steps of the functionalization process, has been also reported, together with in vitro activity tests of immobilized SBP, compared with merely adsorbed SBP, and cytotoxicity tests using human osteoblast (MG-63) cells. Overall, a new bio-conjugate material, able to maintain its activity over time and to decrease the oxidative stress when in contact with MG-63 cells, has been obtained.