QCM-D as a method for monitoring enzyme immobilization in mesoporous silica particles

Enzyme immobilization in mesoporous materials is a field of great interest, with applications in biocatalysis and biosensing. However, the actual immobilization process is not well understood and has mainly been studied by indirect measurements. The present work demonstrates a direct method for real time study of enzyme immobilization in mesoporous silica particles using quartz crystal microbalance with dissipation monitoring (QCM-D). Silica-coated sensors were grafted with amine groups followed by adsorption of small (40 nm), spherical mesoporous silica particles, after which the enzyme immobilization into the mesoporous particles could be studied in real time. The influence of pH on the immobilization efficiency was studied using two different enzymes; lipase from Rhizopus oryzae and feruloyl esterase from Fusarium oxysporum. The results showed that the silica particles adsorbed readily to the amine-grafted surface. The QCM-D measurements indicated that considerably more enzyme was immobilized into mesoporous silica particles than to non-porous silica particles and to a flat silica surface. The viscoelastic effect of the immobilized enzymes was visualized by plotting the frequency shift against the corresponding dissipation. It was observed that the immobilization into the porous particles can be divided into two regimes where the first regime is suggested to represent adsorption to the outer surface and pore openings and the second regime represents further adsorption inside the pores. In summary, we demonstrated QCM-D as a novel method for understanding enzyme immobilization in mesoporous particles in real time and the approach may be of general use for studies of entrapment of molecules into porous particles.