Mixed Monolayers of Ferrocenylalkanethiol and Encapsulated Horseradish Peroxidase for Sensitive and Durable Electrochemical Detection of Hydrogen Peroxide

This paper describes the construction of a mixed monolayer of ferrocenylalkanethiol and encapsulated horseradish peroxidase (HRP) at a gold electrode for amperometric detection of H2O2 at trace levels. By tuning the alkanethiol chain lengths that tether the HRP enzyme and the ferrocenylalkanethiol (FcC(11)SH) mediator, facile electron transfer between FcC(11)SH and HRP can be achieved. Unlike most HRP-based electrochemical sensors, which rely on HRP-facilitated H2O2 reduction (to H2O), the electrocatalytic current is resulted from an HRP-catalyzed oxidation reaction of H2O2 (to O-2). Upon optimizing other experimental conditions (surface coverage ratio, pH, and flow rate), the electrocatalytic reaction proceeding at the electrode was used to attain a low amperometric detection level (0.64 nM) and a dynamic range spanning over 3 orders of magnitude. Not only does the thin hydrophilic porous HRP capsule allow facile electron transfer, it also enables H2O2 to permeate. More significantly, the enzymatic activity of the encapsulated HRP is retained for a considerably longer period (>3 weeks) than naked HRP molecules attached to an electrode or those wired to a redox polymer thin film. By comparing to electrodes modified with denatured HRP that are subsequently encapsulated or embedded in a poly-L-lysine matrix, it is concluded that the encapsulation has significantly preserved the native structure of HRP and therefore its enzymatic activity. The electrode covered with FcC(11)SH and encapsulated HRP is shown to be capable of rapidly and reproducibly detecting H2O2 present in complex sample media.