Construction of a bimetallic Cu@Ce nanozyme cascade biosensor with multi-enzyme activities for detection of alendronate sodium

In this work, a bimetallic Cu@Ce nanozyme was fabricated via electrostatic self-assembly using positively-charged Arg-CeO₂ and negatively-charged Asp-Cu as building blocks, rather than through simple physical mixing of the two precursors. Benefiting from Cu@Ce interfacial electronic redistribution and abundant oxygen-vacancy defects, the resulting Cu@Ce exhibits well-balanced phosphatase-like, laccase-like and ascorbate oxidase-like triple enzyme-mimicking activities, which cannot be achieved by individual Asp-Cu or Arg-CeO₂ components. Leveraging the intrinsic multi-enzyme properties of Cu@Ce, a fluorescence-colorimetric dual-mode cascade sensing platform was constructed for the screening of bisphosphonate-class drugs without introducing additional natural or exogenous nanozymes. As a representative bisphosphonate, alendronate sodium (ALDS) can electrostatically adsorb onto the nanozyme surface and competitively suppress the phosphatase-like activity of Cu@Ce. Under the optimized conditions, the fluorescence channel shows a linear range of 0.5-80 μM with a limit of detection (LOD) of 0.14 μM, whereas the colorimetric channel delivers a broader detection window of 1-120 μM with an LOD of 0.68 μM. The two independent signal outputs enable mutual cross-validation, improving both the anti-interference capability and detection reliability of the system. The sensing platform has been successfully applied to the determination of ALDS in lake-water samples with satisfactory recoveries. Free from tedious chemical derivatization and bulky high-cost instruments, this platform acts as a highly promising complementary analytical tool for on-site rapid screening of bisphosphonate-class drugs.