Oxidation of Biomolecules by Emerging Inorganic Nanoparticles

Oxidation of Biomolecules by Emerging Inorganic Nanoparticles

Antonia Luna-Velasco, Reyes Sierra-Alvarez, Jim A. Field

Department of Chemical and Environmental Engineering

University of Arizona, P.O. Box 210011, Tucson, Arizona 85721, USA

In recent years, inorganic nanoparticles (NPs) are finding increasing application in manufacturing and consumer products. The unusual properties of these emerging pollutants (e.g., very small size, high surface area and reactivity) has lead to growing public and regulatory concern about their potential toxicity. An important mechanism suggested for nanotoxicity is oxidative stress caused by reactive oxygen species (ROS). Testing the chemical generation of ROS by the interaction of NPs with biomolecules could be a helpful tool for nanotoxicity screening. In this study, a rapid screening assay was developed to determine the chemical generation of ROS by NPs through their interaction with biomolecules susceptible to oxidation such as, L-3,4-dihydroxyphemylalanine (L-dopa) and 1,2-dihydroxybenzene (catechol). The assay utilized the dye dichlorofluorescin (DCFH), which is oxidized to its highly fluorescent product dichlorofluorescein (DCF) by ROS. Additionally, oxidation of proteins by NPs was evaluated using an ELISA assay (based on detecting carbonyl groups in amino acid side chains).

The interaction of nanosized CeO₂ with L-dopa or catechol significantly enhanced the oxidation of the ROS indicator dye. Nanoscale CeO₂, Fe₂O₃ and Fe⁰ enhanced ROS production during the autooxidation of L-dopa by more than four-fold in reactions that were dependent on O₂.  Mn₂O₃ NPs oxidized DCFH in the absence of O₂ or L-dopa, suggesting a direct oxidation of the dye. Mn₂O₃ NPs also significantly oxidized the protein bovine serum albumin indicating their potential reactivity with cell components. ROS generation by NPs (CeO₂, Mn₂O₃ and Fe⁰) in aerobic aqueous suspensions was confirmed by electron paramagnetic resonance (EPR) signature for hydroxyl radicals with spin traps. The results taken as a whole indicate that NPs can generate ROS via chemical reactions with medium components and biomolecules susceptible to oxidation, such as L-dopa. The rapid assay with L-dopa and DCFH is a method proposed to screen for chemical ROS production by NPs.