Method of selecting antioxidants for use in topically applied compositions

An ex vivo method for screening compounds for antioxidant behavior in a composition to be topically applied to skin, the method comprises determining the compound's ability both to inhibit ultraviolet radiation-induced skin lipid hydroperoxide formation and to inhibit ultraviolet radiation-induced reactive oxygen species formation in the epidermis;

• wherein determining inhibition of ultraviolet radiation-induced reactive oxygen species formation in the epidermis comprises applying the composition containing said compound(s) to a surface of a skin tissue;
• treating the skin tissue with dihydrorhodamine, irradiating the treated skin tissue with a UV source;
• and imaging the skin tissue using two-photon fluorescence intensity imaging;
• wherein a base formula without said compound was used as a control;
• and wherein determining inhibition of UVR-induced skin lipid hydroperoxide formation comprises the steps of applying to distinct areas of skin of a subject an antioxidant-containing composition and a placebo composition to produce an antioxidant skin site containing antioxidant and skin lipids and a placebo skin site containing placebo and skin lipids;
• applying a strip to the antioxidant skin site and the placebo skin site to produce an antioxidant strip sample containing antioxidant and skin lipids and a placebo strip sample containing placebo and skin lipids;
• removing said strip samples from the skin and exposing said strip samples to UVR to form a UVR-induced antioxidant/lipid reaction product on the antioxidant strip sample and a UVR-induced placebo/lipid reaction product on the placebo strip sample;
• separately contacting the antioxidant strip sample and the placebo strip sample with solvent to prepare a first extract containing UVR-induced antioxidant/lipid reaction product and a second extract containing UVR-induced placebo/lipid reaction product;
• assaying said first and second extracts for lipid hydroperoxide content for each extract;
• and comparing the lipid hydroperoxide content of the first extract to the lipid hydroperoxide content of the second extract.