Direct electrochemistry of horseradish peroxidase immobilized on electrografted 4-ethynylphenyl film via click chemistry

In this paper, a simple two-step approach for redox protein immobilization was introduced. Firstly, alkynyl-terminated film was formed on electrode surface by electrochemical reduction of 4-ethylnylphenyl (4-EP) diazonium compound. Then, horseradish peroxidase (HRP) modified with azido group was covalently immobilized onto the electrografted film via click reaction. Reflection absorption infrared (RAIR) spectroscopy and electrochemical methods were used to characterize the modification process. The results indicate that HRP retains its native structure and shows fast direct electron transfer. Moreover, the immobilized HRP shows excellent electrocatalytic reduction activity toward H2O2 with a linear range of 5.0 × 10−6 to 9.3 × 10−4 mol L−1.

Graphical abstractHydrogen peroxide biosensor was developed based on electrochemically assisted aryldiazonium salt chemistry and click chemistry.Figure optionsDownload full-size imageDownload as PowerPoint slideHighlights► A simple, versatile two-step approach, which is based on electrochemically assisted aryldiazonium salt chemistry and Cu(I)-catalyzed azide alkyne cycloaddition (CuAAC) reaction has been developed for covalent redox proteins immobilization and biosensing for the first time. In this work, azido group modified HRP was covalently grafted on 4-ethylnylphenyl diazonium compound via CuAAC reaction and a novel electrochemical hydrogen peroxide biosensor was successfully fabricated.