Determination of hydroxyl rate constants by a high-throughput fluorimetric assay: towards a unified reactivity scale for antioxidants.

We describe in this article the development of a new method for the determination of rate constants of reaction of the hydroxyl radical, generated by radiolysis of water, with almost any possible molecule. It has been designed to provide a fast and reliable screening of antioxidant banks using microplates. Our particular approach is based on the use of the coumarin molecule as a competitor against the tested molecules: after a fast pulse of low dose irradiation, the fluorescence of 7-hydroxycoumarin produced by the oxidation of coumarin is measured and is inversely proportional to the scavenging ability of the tested antioxidant.

High-throughput evaluation of antioxidant and pro-oxidant activities of polyphenols with thymidine protection assays.

A recently reported high-throughput screening strategy has been applied to the rapid selection of new water-soluble antioxidants that display strong protective activities. Based on a competitive immunoassay, a triple-screening procedure was used to evaluate the ability of different compounds to protect thymidine under different oxidative stresses. The pro-oxidant effect of norbadione A in the presence of iron was observed, while some pulvinic acid derivatives proved strongly protective during gamma radiolysis, UV irradiation, and Fenton-like oxidation.