Dual-modal colorimetric and fluorescent sensing of antineoplastic drug fludarabine via bimetallic-doped nitrogen carbon dots nanozyme

Accurate quantification of fludarabine (FLD) is of paramount clinical importance due to its narrow therapeutic index, where insufficient dosing compromises anticancer efficacy while overdosing leads to severe immunosuppression and toxicity. Reliable monitoring is therefore essential for personalized therapy and improved patient outcomes. To address this challenge, we designed bimetallic copper–nickel co-doped nitrogen-rich carbon dots (CuNi@NCDs) as a multifunctional nanozyme for dual-mode FLD detection. The synergistic Cu/Ni co-doping endowed the NCDs with robust peroxidase-like activity and enhanced photoluminescence by facilitating H₂O₂ activation into highly reactive hydroxyl radicals (HO•) and accelerating charge transfer across the carbon framework. This dual functionality enabled simultaneous colorimetric and ratiometric fluorescence sensing, providing cross-validated outputs that improve analytical reliability. Mechanistically, FLD selectively inhibited the catalytic activity through strong chelation with Cu/Ni centers while modulating the inner filter effect (IFE) of the oxidation product 2, 3-diaminophenazine (DAP), thereby generating distinct and complementary signal responses. The proposed assay achieved ultralow detection limits (16.0 nM by absorbance and 2.6 nM by fluorescence) and excellent recoveries (96.7–105.0%) in pharmaceutical and biological matrices, outperforming many existing analytical platforms. These findings demonstrate the potential of CuNi@NCD-based nanozymes as robust and versatile tools for clinical monitoring, therapeutic drug management, and broader bioanalytical applications.