Designing a strategy for encasing enzymes and nanozymes in microreactors with spatial confinement in a way to improve the selectivity and activity of nanozymes is an exciting goal. In the present work, we report a facile route to encapsulate glucose oxidase (GOx) and poly(ethylenimine) (PEI)-conjugated magnetite nanoparticles (Fe3O4-PEI) in the hollow interior of hybrid microreactors. The microreactors are prepared by polyallylamine hydrochloride (PAH)-mediated silica (SiO2) nanoparticle assembly on calcium carbonate (CaCO3) particles as a removable core. By tuning both shape and phase (vaterite/calcite and pure calcite) of CaCO3, it allows generation of GOx and Fe3O4-PEI-encapsulated silica hollow microspheres (GOx-Fe3O4@SHS) and microcubes (GOx-Fe3O4@SHC). As observed, in a biomimetic cascade catalysis, the confined GOx in the microreactors is able to catalyze oxidation of glucose to gluconic acid and hydrogen peroxide (H2O2), followed by the activation of H2O2 by Fe3O4-PEI for the oxidation of the chromogenic substrate o-phenylenediamine (oPD) to 2,3-diaminophenazine. Comparison of the peroxidase-like activity of the encapsulated Fe3O4-PEI shows that the hollow microspheres (GOx-Fe3O4@SHS) result in activity 14 times higher than that of the hollow microcubes (GOx-Fe3O4@SHC), which in turn is corroborated to the differential loading capacity of GOx in microspheres and microcubes. The evaluation of kinetic parameters indicates a fivefold increase in the catalytic constant (kcat) of Fe3O4-PEI confined in hollow microspheres (GOx-Fe3O4@SHS) compared to the mixture comprising free GOx and Fe3O4-PEI in solution. It suggests that the confined space in the microreactors allows the tandem reactions of GOx and Fe3O4-PEI to take place in close proximity, leading to an improved overall activity. This indeed is seen in the kcat obtained for Fe3O4@SHS (GOx added externally during the assay), which is 14 times lower than that of GOx-Fe3O4@SHS.
Keywords: glucose oxidase; microreactors; peroxidase; self-assembly; silica.