The development of nanomaterials that mimic oxidase-like activities has recently attracted an increasing amount of attention. Obtaining highly active and cost-effective oxidase mimics has posed a significant challenge in this area of research. In this study, we successfully synthesized nickel-doped ferrous disulfide nanocubes (Ni–FeS2) via a facile one-step method. Characterization by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) revealed that Ni was predominantly distributed within the surface layer of the Ni–FeS2 nanocubes. The incorporation of nickel in density functional theory (DFT) calculations effectively reduced the d-band center of Fe, resulting in weakened adsorption to intermediates and thereby enhancing its catalytic efficiency. Moreover, we developed a novel approach based on Ni–FeS2 (the Ni–FeS2 method) for detecting reducing substances, which exhibited good sensitivity toward ascorbic acid (AA), glutathione (GSH), and cysteine (Cys). Remarkably, the established Ni–FeS2 method was successfully employed for in vitro assessment of total antioxidant capacity (TAC) in cellular and organ samples, thereby enabling discrimination between normal, senescent, and malignant cells as well as distinguishing among healthy liver tissue, cancerous liver tissue, and metastatic organs.
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