Abstract
Hydroxyl radicals (•OH) can be generated via Fenton chemistry catalyzed by transition metals. An in vitro Fenton system was developed to test both the inhibition and stimulation of •OH formation, by monitoring salicylate aromatic hydroxylation derivatives as markers of •OH production. The reaction was optimized with either iron or copper, and target analytes were determined by means of an original HPLC method coupled to coulometric detection. The method granted good sensitivity and precision, while method applicability was tested on antioxidant compounds with and without chelating properties in different substance to metal ratios. This analytical approach shows how Fenton’s reaction can be monitored by HPLC coupled to coulometric detection, as a powerful tool for studying molecules′ redox behavior.
Highlights
Oxidative stress is defined as the lack of balance between the presence of chemically reactive oxidative species, reactive oxygen species (ROS) and reactive nitrogen species (RNS), and the ability of the organism to counteract their action through its antioxidant protective systems, including both antioxidant enzymes and endogenous antioxidants
An optimized salicylate hydroxylation assay coupled to an original high-performance liquid chromatography with electrochemical detection (HPLC-ED) method was used to monitor OH production within the Fenton’s reaction
The method was applied to monitoring the Fenton’s reaction for the study of known or prospective antioxidant substances of natural and synthetic origin and iron/copper chelators. This method confirms that the redox behavior of these bioactive molecules can be at least in part related to their capacity to block or increase OH production via Fenton chemistry
Summary
Oxidative stress is defined as the lack of balance between the presence of chemically reactive oxidative species, reactive oxygen species (ROS) and reactive nitrogen species (RNS), and the ability of the organism to counteract their action through its antioxidant protective systems, including both antioxidant enzymes and endogenous antioxidants. This redox balance can be broken by the presence of prooxidant factors (drugs and other xenobiotics, radiation, inflammation) [1] and/or through a decline in systems of protection from oxidation, leading to a significant decrease in the body capacity to contrast oxidative attacks directed toward biomolecular targets [2]. One of the main biological iron features is its ability to donate or receive an electron, i.e., to convert between its ferrous (Fe2+ , Fe(II)) and ferric (Fe3+ , Fe(III)) forms
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