Bimetallic Fe/Cu-doped carbon-based nanozymes with peroxidase-like properties for multimodal determination of neurodegenerative related substance acetylcholinesterase

Acetylcholinesterase (AChE) is a crucial substance in mammalian nervous system, which plays a pivotal role in regulating the levels of neurotransmitters. In this study,a bimetallic iron/copper (Fe/Cu)-doped carbon-based peroxidase-like nanozyme (Fe/Cu N₈₀₀) with graphitic carbon nitride (g-C₃N₄) as nitrogen source was successfully synthesized through high-temperature pyrolysis. Fe/Cu N₈₀₀ exhibited higher peroxidase-like activity compared with Fe N₈₀₀ and Cu N₈₀₀, separately, indicating thatFe/Cu co-doping could enhance catalytic activity. With peroxidase-like properties, Fe/Cu N₈₀₀could catalyze hydrogen peroxide (H₂O₂) to generate reactive oxygen species, which oxidized o-phenylenediamine (OPD) to yellow 2,3-diaminophenazine underultraviolet lightabsorptionat 419 nm and fluorescence emission at 560 nm.Notably, AChE could catalyze the hydrolysis of acetylthiocholine (ATCh) to generate thiocholine (TCh), which hindered the oxidation process of OPD and thus weakened the absorbance and fluorescence emission.Further, this sensing system could use smartphones to achieve visual detection based on color change of the hydrogel device. Thus, by monitoring the fluorescenceof the sensing system at 560 nm and hue intensity of hydrogel device, a dual signal sensing platform formonitoring AChE wassuccessfully established. The linear calibration results of fluorescence and smartphone-based methods for AChE detection were in the range of 0.2–10 and 0.5–15 U·L⁻¹, separately, while the limits of detection were 0.07 and 0.13 U·L⁻¹, respectively. Finally, this developed method yielded satisfactory results for quantification of AChE in human serum and whole blood samples, providing a possibility for convenient and quick determination of neurodegenerative substances and for diagnosing AChE-related diseases.