Simple enzyme-free detection of uric acid by an in situ fluorescence and colorimetric method based on Co-PBA with high oxidase activity

Abstract

In this work, we prepared a simple and low-cost cobalt-doped Prussian blue analog (Co-PBA), which can directly oxidize 10-acetyl-3,7-dihydroxyphenoxazine and 3,3′,5,5′-tetramethylbenzidine (TMB) to produce resorufin (ox-AR) with high fluorescent quantum yield and ox-TMB with blue color, respectively, without the need for unstable H₂O₂. Using the Michaelis–Menten curve and Lineweaver–Burk equation, the Michaelis–Menten constant of Co-PBA and the substrate TMB was found to be 0.033 mM, which was much lower than horseradish peroxidase and other reported nanozymes, showing satisfactory substrate affinity. Uric acid (UA) can cause erosion of the Co-PBA structure, and it significantly reduces the catalytic activity of Co-PBA, resulting in the decrease of the fluorescence emission signal of ox-AR and the absorption signal of ox-TMB. Based on this, a simple, sensitive, and fast fluorescence/colorimetric dual-mode uric acid detection platform was established. The detection range for UA by fluorescence method is 0.625–40 μM, and the detection limit (LOD, S/N = 3) is as low as 0.389 μM. The detection system was applied to serum samples with good recovery and can be used for field detection of UA in biological samples under different environments to meet different needs.