Electrochemical biosensors utilizing the electron transfer of hemoglobin immobilized on cobalt-substituted ferrite nanoparticles–chitosan film

Cobalt ferrite nanoparticles (Co_xFe_3−xO₄) and chitosan (CS) film were used to immobilize/adsorb hemoglobin (Hb) to create a protein electrode to study the direct electron transfer between the redox centers of the proteins and the electrode. X-ray diffraction (XRD) and transmission electron microscopy (TEM) revealed that the Co_xFe_3−xO₄ particles were nanoscale in size and formed an ordered layered structure. The native structure of the immobilized Hb was preserved as indicated by Fourier-transform infrared (FTIR) and UV–visible (UV–vis) spectroscopy. The Hb-Co_xFe_3−xO₄–CS modified electrode showed a pair of well-defined and quasi-reversible cyclic voltammetric peaks at −0.373 V (vs. SCE) and exhibited appreciable electrocatalytic activity for the reduction of H₂O₂. The catalysis currents increased linearly with H₂O₂ concentration in a wide range of 5.0 × 10⁻⁸ to 1.0 × 10⁻³ mol L⁻¹ with a detection limit of 1.0 × 10⁻⁸ mol L⁻¹ (S/N = 3) and had long-term stability. Finally, the proposed method was applied to investigate the coexistence of hydrogen peroxide with the interfering substances. Experimental results showed that the ascorbic acid, glucose, l-cysteine, uric acid, and dopamine at corresponding concentrations did not influence the detection of H₂O₂.