Self-Assembled Integrated Nanozyme Cascade Biosensor with Dual Catalytic Activity for Portable Urease Analysis

In this work, a novel nanozyme (Cu@Zr) with all-in-one dual enzyme and fluorescence properties is designed by simple self-assembly. A nanozyme cascade sensor with disodium phenyl phosphate (PPDS) as substrate was first established by exploiting the dual enzymatic activities of phosphatase and laccase. Specifically, phosphatase cleaves the P-O bond of PPDS to produce colorless phenol, which is then oxidized by laccase and complexed with the chromogenic agent 4-aminoantipyrine (4-AP) to produce red quinoneimine (QI). Strikingly, the NH₃ produced by the urease hydrolysis of urea can interact with Cu@Zr, accelerating the electron transfer rate and ultimately leading to a significantly improved performance of the cascade reaction. Moreover, the fluorescence at 440 nm of Cu@Zr is further quenched by the inner filter effect (IFE) of QI. Thus, the colorimetric and fluorescence dual-mode strategy for sensitive urease analysis with LODs of 3.56 and 1.83 U/L was established by the proposed cascade sensor. Notably, a portable swab loaded with Cu@Zr was also prepared for in situ urease detection with the aid of a smartphone RGB readout. It also provides a potentially viable analytical avenue for environmental and biological analysis.