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 (k_cat) 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 k_cat obtained for Fe₃O₄@SHS (GOx added externally during the assay), which is 14 times lower than that of GOx-Fe₃O₄@SHS.

Patturu Swathi

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