Mapping and Modeling Interfacial pH Dynamics at Enzymatic Microelectrode
Abstract
Redox enzymes are promising biocatalysts for energy conversion and sensing, particularly when immobilized on electrodes to facilitate charge transfer and enhance stability. While they offer the advantages of operating under mild conditions and near-neutral pH, high reaction rates can profoundly alter the immobilized enzyme microenvironment. This requires understanding the interplay between localized concentration gradients and enzyme activity at the electrocatalytic interface. Here, we elucidate this dynamic coupling between reaction kinetics and local pH at an enzyme-modified electrode through a combined modeling and experimental approach. We present a detailed 2D-axisymmetric finite element model of a graphite microelectrode modified with bilirubin oxidase from Myrothecium verrucaria catalyzing the oxygen reduction reaction via direct electron transfer. The model explicitly accounts for buffer equilibria through ion activities and incorporates experimentally determined pH-dependent kinetic parameters, including apparent turnover and Michaelis constants. The simulations were validated experimentally using in situ and operando fluorescence confocal laser scanning microscopy (FCLSM) with the pH-sensitive dye fluorescein in weakly buffered electrolytes. Both numerical and experimental results reveal substantial interfacial alkalinization, with the local pH increasing by more than two units under the weakest buffering conditions. The model successfully reproduces cyclic voltammetry and quasi-steady-state chronoamperometry profiles using a single enzyme-coverage-dependent parameter. Furthermore, spatial fluorescence profiling closely corroborates the simulated pH gradients, while revealing evidence of natural convection occurring over extended time scales. Our study provides a quantitative framework crucial for designing stable, high-performance bioelectrochemical interfaces, which can be readily extended to other key redox enzymes, particularly those involved in CO₂ and H₂ conversion.
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BibTeX
@article{liu2026mapping,
title = {{Mapping and Modeling Interfacial pH Dynamics at Enzymatic Microelectrode}},
author = {Ruoyi Liu and Sayaka Nishida and Ami Kobayashi and Keisei Sowa and Elisabeth Lojou and Anne de Poulpiquet and Ievgen Mazurenko},
journal = {Journal of the American Chemical Society},
year = {2026},
month = oct,
doi = {10.1021/jacs.6c15657},
url = {https://hal.science/hal-05786271v1},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Mapping and Modeling Interfacial pH Dynamics at Enzymatic Microelectrode
AU - Ruoyi Liu
AU - Sayaka Nishida
AU - Ami Kobayashi
AU - Keisei Sowa
AU - Elisabeth Lojou
AU - Anne de Poulpiquet
AU - Ievgen Mazurenko
PY - 2026
DA - 2026/10/08
JO - Journal of the American Chemical Society
DO - 10.1021/jacs.6c15657
UR - https://hal.science/hal-05786271v1
AB - Redox enzymes are promising biocatalysts for energy conversion and sensing, particularly when immobilized on electrodes to facilitate charge transfer and enhance stability. While they offer the advantages of operating under mild conditions and near-neutral pH, high reaction rates can profoundly alter the immobilized enzyme microenvironment. This requires understanding the interplay between localized concentration gradients and enzyme activity at the electrocatalytic interface. Here, we elucidate this dynamic coupling between reaction kinetics and local pH at an enzyme-modified electrode through a combined modeling and experimental approach. We present a detailed 2D-axisymmetric finite element model of a graphite microelectrode modified with bilirubin oxidase from Myrothecium verrucaria catalyzing the oxygen reduction reaction via direct electron transfer. The model explicitly accounts for buffer equilibria through ion activities and incorporates experimentally determined pH-dependent kinetic parameters, including apparent turnover and Michaelis constants. The simulations were validated experimentally using in situ and operando fluorescence confocal laser scanning microscopy (FCLSM) with the pH-sensitive dye fluorescein in weakly buffered electrolytes. Both numerical and experimental results reveal substantial interfacial alkalinization, with the local pH increasing by more than two units under the weakest buffering conditions. The model successfully reproduces cyclic voltammetry and quasi-steady-state chronoamperometry profiles using a single enzyme-coverage-dependent parameter. Furthermore, spatial fluorescence profiling closely corroborates the simulated pH gradients, while revealing evidence of natural convection occurring over extended time scales. Our study provides a quantitative framework crucial for designing stable, high-performance bioelectrochemical interfaces, which can be readily extended to other key redox enzymes, particularly those involved in CO₂ and H₂ conversion.
SN - 0002-7863
SN - 1520-5126
ER -