Construction of hyaluronan-silver nanoparticle–hemoglobin multilayer composite film and investigations on its electrocatalytic properties

In this work, hyaluronan-silver nanoparticles (HSNPs) were prepared by UV-initiated photoreduction, and protein hemoglobin (Hb) was then alternately assembled with the prepared negatively charged HSNPs into layer-by-layer (LBL) films on solid surface. The electrochemical behavior and electrocatalytic activities toward oxygen and hydrogen peroxide of the resulting films were studied. It was found that the HSNPs greatly enhanced the electron transfer reactivity of Hb as a bridge. The assembly films showed a pair of nearly reversible redox peaks with a formal potential of −0.32 V (vs. Ag/AgCl) for the heme Fe(III)/Fe(II) redox couple. The immobilized Hb in the films maintained its biological activity, showing a surface-controlled process with a heterogeneous electron transfer rate constant (ks) of 1.0 s−1 and displaying the same features of a peroxidase in the electrocatalytic reduction of oxygen and hydrogen peroxide. This work provides a novel model to fabricate LBL films with protein, polysaccharide and nanoparticles, which may establish a foundation for fabricating new type of biosensors based on the direct electron transfer of redox proteins immobilized in nanocomposite multilayer films with underlying electrodes.