Dual-mode colorimetric and fluorometric detection of D-penicillamine via inhibition of peroxidase-mimetic activity of bimetallic N-doped carbon dots.

D-penicillamine (D-PEN) is used to treat Wilson's disease, rheumatoid arthritis, and cystinuria but requires careful monitoring due to its narrow therapeutic window and risks of nephrotoxicity, hematological disorders, and autoimmune reactions. To enable reliable detection, bimetallic FeCu co-doped carbon dots (FeCu@CDs) were developed as multifunctional nanozymes for dual-mode sensing of D-PEN. These nanozymes integrate peroxidase-like catalytic activity, intrinsic fluorescence, and selective thiol-binding affinity, allowing simultaneous colorimetric and ratiometric fluorescence detection. Fe and Cu doping enhanced the peroxidase-like activity and fluorescence emission of the CDs by promoting H₂O₂ activation into HO• and 1O₂ and improving charge mobility. D-PEN inhibited this catalytic activity by binding metal centers and altered the inner filter effect (IFE) between FeCu@CDs and 2, 3-diaminophenazine (DAP), enabling selective signal modulation. The assay achieved detection limits of 0.13 μM (colorimetric) and 0.03 μM (fluorescence), with recoveries ranging from 95.6 % to 105.1 % in real samples, highlighting its importance in clinical monitoring and bioanalytical applications.