Discrimination and Quantification of Glutathione by Cu⁺-Based Nanozymes

Glutathione (GSH) is the most abundant low-molecular-weight biological thiol in vivo and has been linked to several diseases. The accurate quantification of GSH is therefore crucial for disease diagnosis and monitoring. In this study, we prepared self-assembled Cu(I)-Cys (cysteine) nanozymes through a two-step procedure. The Cu(I)-Cys nanoparticles exhibited peroxidase-mimicking activity. Upon the addition of H₂O₂, they were able to oxidize 3,3,5,5-tetramethylbenzidine (TMB) into oxTMB, resulting in a measurable increase in UV-Vis absorption at 655 nm. However, in the presence of GSH, oxTMB was reduced back to TMB, leading to a decrease in UV-Vis absorption at 655 nm. By utilizing these changes in the absorption intensity, we achieved the sensitive detection of GSH with a detection limit of 2.13 μM. Moreover, taking advantage of the different peroxidase-mimicking activities of Cu(I)-Cys nanoparticles at various pH values, a sensor array with Cu(I)-Cys nanoparticles at pH 4 and pH 5 was constructed. The discrimination of GSH among Cys and ascorbic acid was achieved and the practicability of the sensor array in human serum was validated. This novel approach holds significant promise for the precise discrimination and quantification of GSH and its potential applications in disease diagnosis and therapeutics.