Protein-Protected Gold Nanocluster-Based Biosensor for Determining the Prooxidant Activity of Natural Antioxidant Compounds

In this work, chicken egg white protein (CEW)-protected gold nanoclusters (CEW-AuNCs) were prepared from CEW and HAuCl4 to measure the Cu(II)-induced prooxidant activity of antioxidant compounds such as epicatechin, epigallocatechin gallate, catechin, rosmarinic acid, resveratrol, ascorbic acid, and glutathione. These compounds reduced Cu(II) to Cu(I), and the latter was mainly bound to thiol groups in the CEW-AuNC structure. As the protein-bound Cu(I) may act as a catalytic center for generating reactive oxygen species, the Cu(II) reducing ability of antioxidants is an indirect measure of their prooxidant potency. The bound Cu(I) may be released with the cuprous-selective ligand neocuproine (Nc), forming the basis of a spectrophotometric method measuring absorbance at 450 nm wavelength of the Cu(I)Nc chelate. The developed method involved a one-pot synthesis and determination without preseparation and was applied to binary synthetic mixtures of studied antioxidant compounds and to certain herbal plant (green tea, linden, echinacea, and artichoke leaf) extracts to determine the total prooxidant activities. The obtained results were statistically compared with those of the literature Cu(II)-Nc assay using a calcium proteinate-based solid biosensor. The developed biosensor was durable, reliable, easily applicable, and of low cost and wide linear range and could determine the prooxidant activities of natural antioxidant samples with high reproducibility.