Among microelectronic materials, silicon (Si) has the most mature and low-cost technology; hence, several research groups are approaching Si-compatible technology as an innovative platform for biosensors. Porous silicon has been intensively investigated for a variety of applications such as chemical and biological sensors, medical diagnostics, optical band pass filters, microchemical reactors, and microfuel cells. Moreover, Si-based matrixes have been proved to be a very useful support for the immobilization of enzymes thanks to their capability of retaining biological activity. Silicon (Si) received a lot of attention due to its specific semiconductor properties and furthermore because it allows the development of a broad range of micropatterning processes in order to achieve functional features for future integration in complex systems.
Furthermore, the Si-H and Si-OH groups on porous silicon surface can be easily modified by many reactive reagents and derivatives with receptors, thus enabling the identification of ligands. Microreactors are miniaturized reaction systems fabricated by microtechnology and precision engineering. The microreactors work with micro and nanoliter volumes of reaction media and ensure high efficiency and reproducibility of biocatalytic processes. Enzymatic microreactors have been already proposed as integrated components termed lab-on-a-chip for analytical applications in micrototal analysis systems (MTAS). There, the immobilization strategies to graft different chemical substances on the surface of a microreactor, a support, are used for a design of necessary conditions within the microreactor spaces. Surface modification by silanization is a very common method for particle functionalization. High density of free amino groups (-NH2) lying outwards the particle surface provides an excellent media for further chemical surface modification such as enzyme cross-linking with glutaraldehyde. The immobilization of enzymes in microreactors is mostly carried out in a covalent way. The main advantage of covalent immobilization is the retention of the enzyme during the whole biocatalytic process. Actually, immobilization is a well-established approach in a wide range of industrial applications. Both synthetic and natural inorganic materials such as clay, glass beads, silice-based materials, and celite have been used to immobilize enzymes, the natural catalysts for many biological processes. Among them, mesoporous silicates are the most interesting due to their attractive properties, availability, and simple preparation. Peroxidase immobilization on inorganic mesoporous silicates has proven to be an interesting alternative to improve enzyme functionality. The large regular repeating structures of photonic porous silicon structure offer the possibility of adsorbing or entrapping large biomolecules within their pores, providing a suitable microenvironment to stabilize the enzyme.
Peroxidases (EC 220.127.116.11, etc.) belong to a large family of enzymes that participate in a large number of natural processes developed in living organisms. They are ubiquitous in fungi, plants, and vertebrates. Their principal active sites contain a heme prosthetic group or, alternately, residues of redox-active cysteine or seleno-cysteine groups that are able to oxidize a large number of organic compounds initiated by one electron oxidation step. For all peroxidases, the natural substrate is hydrogen peroxide, but the oxidative process can be performed with many other organic hydro-peroxides such as lipid peroxides. In the oxidation of phenols or aromatic amines, peroxidases produce free radicals that may dimerise or polymerize and thus, in general, form products that are much less soluble in water. This property might be used in removing carcinogenic aromatic amines and phenols from industrial aqueous effluents. Enzymes are also involved in degradation of aromatic compounds and other xenobiotics, including pesticides, polycyclic aromatic hydrocarbons, and dioxins, and thus can be used for removal of aromatic pollutants[12, 13] as antioxidant, as indicators for food processing, in bioelectrodes and in the synthesis of conducting materials. Peroxidases could also be used in the synthesis of fine chemicals and optically and biologically active oxide. Despite the obviously practical value of peroxidases, at present, their commercial uses are limited, primarily due to its low stability in the presence of hydrogen peroxide, their natural substrate. All heme-proteins, including peroxidases, are inactivated in the presence of some concentrations of hydrogen peroxide. This process, described as a suicide inactivation, is especially important in the absence of reducing substrates, but its mechanism has not been yet fully elucidated. Although the interest to peroxidase started several decades ago, their application as biocatalysts in industrial processes is still negligible due to its inherent instability under operational conditions, mainly caused by the inactivation in the presence of hydrogen peroxide. The development of techniques for enzyme stabilizing can improve a number of biocatalytic industrial processes. In this work, peroxidase enzyme has been immobilized onto porous silicon (PS) supports for the possible prevention from its self-inactivation and its stability under different operational conditions has been analyzed.