Electrochemically Reduced Graphene Oxide on Electrochemically Roughened Gold as a Support for Horseradish Peroxidase

Graphene oxide (GO) is composed of isolated graphene-like sheets containing various oxygen functionalities. The electrochemical reduction of GO enables the direct deposition of the graphene layer on electrode surfaces. Furthermore, a partial reduction of GO can be performed, enabling easy modification of the surface with oxygen-containing groups. In this contribution, the electrochemical reduction of GO is carried out on a gold electrode roughened by electrochemical oxidation–reduction cycles. Such a gold electrode contains nanostructures ranging in size from tens to 300 nm. The electrochemically reduced GO layer is subsequently used for horseradish peroxidase (HRP) immobilization. Two types of electrochemically reduced GOs, partially reduced graphene oxide (ERGOP) and fully reduced graphene oxide (ERGOF), are used in this study. Although HRP immobilized on both types of surfaces directly catalyzes electroreduction of the hydrogen peroxide, the enzyme immobilized on the ERGOP layer exhibits slightly higher current values compared with those of ERGOF. In contrast, HRP adsorbed directly on the roughened gold reveals negligible activity. To evaluate the effect of the roughened gold, we studied HRP immobilized on an ERGOF layer on glassy carbon. Additionally, infrared and Raman spectroscopy were used to investigate the composition of GO, ERGOP, and ERGOF as well as the HRP structure at the studied surfaces. The infrared results indicate a random orientation of the enzyme molecules on the bare roughened gold and ordered HRP layers on the surfaces covered with either ERGOP or ERGOF.