A dual-signal sensing platform based on single atomic iron-nitrogen-carbon nanozyme for the detection of neurodegenerative disease-related bioactive molecules acetylcholinesterase and dopamine

Abstract

In this work, a novel iron-nitrogen-carbon single-atom nanozyme (Fe-N800 SAzyme) with peroxidase-like activity was synthesized by doping Fe in zeolitic imidazolate frameworks (ZiF-8) and utilizing graphitic C3N4 (g-C3N4) as a nitrogen source in the pyrolysis process. Acetylcholine (ACh) was continuously oxidized by acetylcholinesterase (AChE) and choline oxidase (CHO) to generate H2O2 through an enzymatic cascade reaction. The interaction between Fe-N800 SAzyme and H2O2 led to the generation of reactive oxygen species (ROS) that could then oxidize 3,3’,5,5’-tetramethylbenzidine (TMB) to oxidized TMB (oxTMB), which exhibited an absorption peak at 650 nm. Lysozyme (LZ)-functionalized 5-methyl-2-thiouracil (MTU) gold/silver nanoclusters (Au/Ag/LZ-MTU), which exhibited a fluorescence peak at 610 nm, could be quenched by oxTMB through the inner filter effect (IFE). Dopamine (DA) was able to consume ROS and hinder the oxidation of TMB. Therefore, by monitoring the change of fluorescence at 610 nm and absorption at 650 nm, a dual signal fluorometric and colorimetric sensing platform for AChE and DA was established. The AChE activity was monitored through the variation of fluorescence (I610) or colorimetric (A650) intensity, which had a low limit of detection (LOD) of 0.057 U/L and 0.11 U/L, respectively. The fluorescence and colorimetric methods were also able to sensitively monitor the DA activity with a LOD of 0.83 μM and 2.7 μM, respectively. Finally, the developed method could satisfactorily quantify AChE and DA in human serum, whole blood, and DA hydrochloride injection samples.