General Preparation of Heme Protein Functional Fe₃O₄@Au-Nps Magnetic Nanocomposite for Sensitive Detection of Hydrogen Peroxide

Stable magnetic nanocomposite of gold nanoparticles (Au-NPs) decorating Fe₃O₄ core was successfully synthesized by the linker of Boc-L-cysteine. Transmission electron microscope (TEM), energy dispersive X-ray spectroscopy (EDX) and cyclic voltammograms (CV) were performed to characterize the as-prepared Fe₃O₄@Au-Nps. The results indicated that Au-Nps dispersed homogeneously around Fe₃O₄ with the ratio of Au to Fe₃O₄ nanoparticles as 5–10/1 and the apparent electrochemical area as 0.121 cm². After self-assembly of hemoglobin (Hb) on Fe₃O₄@Au-Nps by electrostatic interaction, a hydrogen peroxide biosensor was developed. The Fe₃O₄@Au-Nps/Hb modified GCE exhibited fast direct electron transfer between heme center and electrode surface with the heterogeneous electron transfer rate constant (Ks) of 3.35 s⁻¹. Importantly, it showed excellent electrocatalytic activity towards hydrogen peroxide reduction with low detection limit of 0.133 μM (S/D=3) and high sensitivity of 0.163 μA μM⁻¹, respectively. At the concentration evaluated, the interfering species of glucose, dopamine, uric acid and ascorbic acid did not affect the determination of hydrogen peroxide. These results demonstrated that the introduction of Au-Nps on Fe₃O₄ not only stabilized the immobilized enzyme but also provided large surface area, fast electron transfer and excellent biocompatibility. This facile nanoassembly protocol can be extended to immobilize various enzymes, proteins and biomolecules to develop robust biosensors.