Reactive oxygen species generation by copper(II) oxide nanoparticles determined by DNA damage assays and EPR spectroscopy

• Published in: Nanotoxicology Vol. 11; no. 2; pp. 278 - 288
• Main Authors: Angelé-Martínez, Carlos, Nguyen, Khanh Van T., Ameer, Fathima S., Anker, Jeffrey N., Brumaghim, Julia L.
• Format: Journal Article
• Language: English
• Published: England Taylor & Francis 01-03-2017; Taylor & Francis Ltd
• Subjects: Ascorbic acid; Biological Assay; Copper; Copper - chemistry; Copper - toxicity; Copper compounds; Cytotoxicity; Deoxyribonucleic acid; DNA; DNA Damage; Electron Spin Resonance Spectroscopy; Electrophoresis; Escherichia coli - drug effects; Escherichia coli - genetics; Free radicals; Hydrogen; Hydrogen peroxide; Hydroxyl radicals; Industrial applications; nano-surfaces; Nanoparticles; Nanoparticles - chemistry; Nanoparticles - toxicity; nanotoxicology; Plasmids; Reactive oxygen species; Reactive Oxygen Species - metabolism; Solubility; Spectroscopy; Superoxide; Surface Properties; Time dependence; Time Factors; Nanoparticles; nano-surfaces; DNA damage; nanotoxicology
• ISSN: 1743-5390; 1743-5404
• DOI: 10.1080/17435390.2017.1293750

Copper(II) oxide nanoparticles (
NP
CuO) have many industrial applications, but are highly cytotoxic because they generate reactive oxygen species (ROS). It is unknown whether the damaging ROS are generated primarily from copper leached from the nanoparticles, or whether the nanoparticle surface plays a significant role. To address this question, we separated nanoparticles from the supernatant containing dissolved copper, and measured their ability to damage plasmid DNA with addition of hydrogen peroxide, ascorbate, or both. While DNA damage from the supernatant (measured using an electrophoresis assay) can be explained solely by dissolved copper ions, damage by the nanoparticles in the presence of ascorbate is an order of magnitude higher than can be explained by dissolved copper and must, therefore, depend primarily upon the nanoparticle surface. DNA damage is time-dependent, with shorter incubation times resulting in higher EC
50
values. Hydroxyl radical (
*
OH) is the main ROS generated by
NP
CuO/hydrogen peroxide as determined by EPR measurements;
NP
CuO/hydrogen peroxide/ascorbate conditions generate ascorbyl, hydroxyl, and superoxide radicals. Thus,
NP
CuO generate ROS through several mechanisms, likely including Fenton-like and Haber-Weiss reactions from the surface or dissolved copper ions. The same radical species were observed when
NP
CuO suspensions were replaced with the supernatant containing leached copper, washed
NP
CuO, or dissolved copper solutions. Overall,
NP
CuO generate significantly more ROS and DNA damage in the presence of ascorbate than can be explained simply from dissolved copper, and the
NP
CuO surface must play a large role.