Kinetic and analytical comparison of horseradish peroxidase on bare- and redox-modified single-walled carbon nanotubes

Single-walled carbon nanotubes (SWNTs) were coated first with methylene blue (MB) by noncovalent adsorption and then by horseradish peroxidase (HRP) by cross-linking with glutaraldehyde. The MB-SWNT/HRP composites formed stable films on glassy carbon electrodes. MB was probably present with a coverage of a monolayer or less. The MB voltammetry was consistent with fast electron transfer to a surface-confined species. The presence of HRP did not significantly affect the MB electrochemistry. MB could mediate electron transfer from HRP in the presence of H₂O₂. Cyclic voltammograms of this process were used to determine the rate constants for the reactions of the native ferriperoxidase with H₂O₂ and of the oxyferryl Compound II with the reduced form of MB. For comparison purposes, the rate constant for the direct electrode reduction of the HRP oxyferryl π-cation radical Compound I was determined in MB-free SWNTs. The results indicate a considerably faster regeneration rate for native ferriperoxidase by the mediated reaction than by direct electrochemistry. Using the MB-SWNT/HRP composites, H₂O₂ could be calibrated by amperometry at −0.3 V vs. SCE. The optimized response (at pH 7.0) had a sensitivity of 661.0 μA mM⁻¹ cm⁻² and a limit of detection (3 × S/N) of 0.1 μM.