In Situ Synthesis of 3D Au@Pt Nanozyme Hydrogel SERS Substrates for Highly Sensitive and Quantitative Sensing

Quantitative detection of low-concentration molecules in complex samples remains a significant challenge in Surface-Enhanced Raman Spectroscopy (SERS) analysis. In this study, we synthesized a 3D Au@Pt nanozyme hydrogel SERS substrate (Au@Pt@Agar) with peroxidase and oxidase mimetic activities using an in situ one-step method. The integration of nanozymes with the hydrogel overcomes several limitations of SERS, including poor quantitative performance, matrix interference, poor reproducibility, and structural stability. The synthesized Au@Pt nanozyme hydrogel SERS substrate is homogeneous, stable, and exhibits efficient catalytic activity. Its three-dimensional network structure provides numerous active sites, facilitating the highly sensitive detection of small molecules. Au@Pt@Agar catalyzes the oxidation of colorless 3,3',5,5'-tetramethylbenzidine (TMB), resulting in a blue-colored oxide with a strong SERS signal. Utilizing diazotization and redox reactions, the nanozyme enabled the sensing of nitrite and glutathione (GSH) with detection limits as low as 2.27 × 10^-13 and 1.98 × 10^-10 M, respectively. The detection sensitivity is enhanced by the dual action of peroxidase and oxidase in the substrate, achieving at least 3 orders of magnitude higher sensitivity compared to previous methods reported in the literature. In addition, the method has been successfully applied to the SERS sensing of nitrite in food and glutathione in human serum without complicated sample pretreatment. The results demonstrated excellent accuracy and practicality, highlighting its broad application potential in food safety, healthcare, and related fields.