A new Prussian blue analog (PBA) that contains three metal elements and has peroxidase-like activity was synthesized by a simple method. Then, AgNO₃ solution was added slowly to the PBA solution under continuous stirring. We found that this synthesis method could be used to prepare other PBAs, and that the anchoring of Ag⁺ on the surface of PBA could enhance the peroxidase-like activity of the material, suggesting potential applications for the Ag⁺-decorated Prussian blue analog (Ag-PBA) in traditional Chinese medicine. Ag-PBA is a new type of multi-metal cubic nano-enzyme that exhibits good stability and excellent peroxidase-like activity; as such, it could catalyze the oxidation of 3,3',5,5'-tetramethylbenzidine (TMB) in the presence of H₂O₂ and Ag-PBA. We then developed a new method to measure the content of antioxidant substances in Chinese herbs by using the excellent peroxidase-like activity of Ag-PBA. Using the Chinese herb Lycium ruthenicum Murr. as a model compound, we measured the content of the antioxidant substances in Lycium ruthenicum Murr. by this new method. After optimization of reaction temperature, concentrations of TMB and H₂O₂, and reaction time, the content of the antioxidant substances was measured and calculated in comparison with anthocyanidin standards. The results of the Ag-PBA method and the classical DPPH method were compared by a paired t-test, with no statistically significant difference found between the methods. Hence, these two methods can be used interchangeably, although the Ag-PBA method had the advantages of simplicity, rapidness, and good stability. Moreover, the Ag-PBA method has a low limit of quantification and a shorter reaction time, which are improvements on the DPPH method, and it is not necessary to avoid light. Therefore, we anticipate that the Ag-PBA method may be used widely for the measurement of the content of antioxidant substances in Chinese herbs.

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