Electrode size-dependent reaction-diffusion fields revealed by in-situ electrochemical-fluorescence lifetime imaging

Electrode size governs mass-transport geometry and thereby modulates reaction-diffusion balances at electrochemical interfaces. Fluorescence imaging provides a spatially resolved means to interrogate these dynamics. In electrochemistry-fluorescence systems where redox transformations and photophysical processes proceed concurrently, quantitative interpretation requires integration of electrochemical signals with both fluorescence intensity and lifetime. Here, we examine the resazurin (RZ)-resorufin (RF)-dihydroresorufin (DH) redox-fluorogenic network using an in-situ electrochemical-fluorescence lifetime imaging (EC-FLIM) platform. A 12.5 μm Au ultramicroelectrode (UME) is directly compared with a 1.6 mm Au disk macroelectrode. Cyclic voltammetry reveals a strongly attenuated return-sweep oxidation current at the UME. This behavior is consistent with hemispherical diffusion, which sustains RZ supply while limiting local DH accumulation. Under these conditions, comproportionation (DH + RZ ⇌ 2RF) is favored, reducing the pool of species available for reoxidation. During linear potential sweeps, fluorescence at the UME remains above baseline even at potentials where RF is further reduced to DH. In contrast, the macroelectrode exhibits rapid signal decay under planar diffusion and pronounced reactant depletion. Depth-resolved FLIM (xy - and xz -plane) identifies, at the UME, a near-electrode off-surface zone of maximal fluorescence activity. This feature is attributed to RF regeneration within the RZ-DH coexistence region. Simulated RF concentration profiles qualitatively reproduce these spatial trends and further indicate that, at the macroelectrode, a regeneration maximum may occur beyond the experimental imaging window. Finally, RF lifetimes decrease systematically with increasing RZ fraction. A lifetime-based Stern-Volmer relationship (τ₀/τ vs . [Q]) displays strong linearity, establishing RZ as a dynamic quencher of RF. Collectively, these results connect electrode-size-dependent diffusion geometry to interfacial redox pathways, fluorescence persistence and regeneration, and lifetime modulation.