Dual-function colorimetric platform for Hg2+ detection and Cys/Hcy discrimination based on the peroxidase-like activity of CuAu nanozymes

Abstract

This study presents a colorimetric method for the detection of Hg²⁺ and discrimination of cysteine (Cys) and homocysteine (Hcy) based on the peroxidase (POD)-like activity of copper-gold nanoparticles (CuAuNPs). CuAuNPs with an average size of approximately 15 nm were synthesized via a one-step reduction method and exhibited intrinsic POD-like activity, catalyzing the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) by H₂O₂ to produce a blue-colored product with a characteristic absorbance at 652 nm the addition of Hg²⁺ specifically enhances the catalytic activity of CuAuNPs, resulting in a distinct color change of the TMB–H₂O₂ solution from light blue to dark blue. Conversely, introduction of Cys or Hcy leads to competitive binding between their thiol (–SH) groups and Hg²⁺, forming stable S–Hg complexes, resulting in a concentration-dependent fading of the blue color. The distinct suppression extents between Cys and Hcy enable their discrimination. Under optimized conditions, the method enables quantitative detection of Hg²⁺ in the range of 100–900 nM (R² = 0.990) with a detection limit of 10 nM, and achieves satisfactory recoveries of 90.4–110.2% in spiked emollient water samples. For Cys and Hcy, linear responses are obtained over 5–40 µM (R² = 0.991; LOD = 2.5 µM) and 6–50 µM (R² = 0.992; LOD = 2.3 µM), respectively. The assay was successfully applied to rat serum samples, yielding recoveries of 99.4–108.1% (RSD < 6.2%) for Cys and 95.0–104.1% (RSD < 6.4%) for Hcy. The proposed method is simple, rapid, and selective, offering a promising nanozyme-based platform for environmental monitoring and biomedical diagnostics.